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3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

BOOK OF ABSTRACTS<br />

2 ND WORLD CONFERENCE OF STRESS<br />

3 RD CELL STRESS SOCIETY INTERNATIONAL CONGRESS ON<br />

STRESS RESPONSES IN BIOLOGY AND MEDICINE AND<br />

2 ND WORLD CONFERENCE OF STRESS<br />

CELEBRATION OF THE CENTENNIAL BIRTH ANNIVERSARY<br />

OF HANS SELYE<br />

23-26 AUGUST 2007, BUDAPEST, HUNGARY<br />

1


2<br />

23-26 August 2007,<br />

Budapest, Hungary


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

TABLE OF CONTENTS<br />

PLENARIES 5<br />

CORE MODULE 1; Stress proteins - molecular stress 15<br />

CORE MODULE 2; Stress and cellular functi<strong>on</strong>s 73<br />

MODULE 3; System level interdisciplinary approaches 169<br />

MODULE 4; Plant stress 185<br />

MODULE 5; Stress at level <str<strong>on</strong>g>of</str<strong>on</strong>g> the organism 233<br />

MODULE 6; Stress in medicine 305<br />

MODULE 7; Psychosocial stress 389<br />

MODULE 8; Others 477<br />

AUTHORS’ INDEX 483<br />

3


4<br />

23-26 August 2007,<br />

Budapest, Hungary


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

PLENARIES<br />

5


6<br />

23-26 August 2007,<br />

Budapest, Hungary


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

P1<br />

PROTEIN MISASSEMBLY: CHLOROPLASTS, CHAPERONES AND CROWDING<br />

R. John Ellis<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biological Sciences, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Warwick, UK<br />

e-mail: R.J.Ellis@warwick.ac.uk<br />

The majority <str<strong>on</strong>g>of</str<strong>on</strong>g> denatured proteins will refold correctly under appropriate c<strong>on</strong>diti<strong>on</strong>s<br />

so why are molecular chaper<strong>on</strong>es required inside the cell A possible answer is that all<br />

cells experience problems due to protein misassembly because all proteins are made <strong>on</strong><br />

polysomes in a crowded envir<strong>on</strong>ment. Synthesis <strong>on</strong> polysomes brings partly folded<br />

identical chains within touching distance while crowding increases associati<strong>on</strong><br />

c<strong>on</strong>stants by two to three orders <str<strong>on</strong>g>of</str<strong>on</strong>g> magnitude. Protein misassembly is defined as the<br />

associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> two or more polypeptide chains to produce n<strong>on</strong>functi<strong>on</strong>al, <str<strong>on</strong>g>of</str<strong>on</strong>g>ten<br />

neurotoxic, products, and is preferred to the more comm<strong>on</strong>ly used term `aggregati<strong>on</strong>`<br />

since some aggregates are functi<strong>on</strong>al. Misassembly is highly specific, requiring the<br />

chains to be identical or very similar in sequence. Protein misfolding is usually defined<br />

in structural terms, as the appearance <str<strong>on</strong>g>of</str<strong>on</strong>g> n<strong>on</strong>-native c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>s during the folding process, but if such<br />

c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>s have no biological c<strong>on</strong>sequences, this definiti<strong>on</strong> lacks meaning. A better definiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

misfolding is the <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> that is unable to reach the native functi<strong>on</strong>al c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <strong>on</strong> a<br />

biologically relevant time scale. On these definiti<strong>on</strong>s all misassemblies are misfolded, but there are <strong>on</strong>ly a few<br />

reports <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins that misfold in vitro but remain m<strong>on</strong>omeric. The c<strong>on</strong>clusi<strong>on</strong> is that most proteins are able<br />

to fold correctly; their problem is to avoid misassembly, a problem aggravated by the crowded nature <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

polysomal envir<strong>on</strong>ment. The universality <str<strong>on</strong>g>of</str<strong>on</strong>g> this misassembly problem could explain the ubiquity <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

molecular chaper<strong>on</strong>es. The observati<strong>on</strong> that some, but not all, chaper<strong>on</strong>es are also stress resp<strong>on</strong>se proteins,<br />

reflects the increased need to combat misassembly under stress c<strong>on</strong>diti<strong>on</strong>s. These ideas will be discussed in<br />

the c<strong>on</strong>text <str<strong>on</strong>g>of</str<strong>on</strong>g> the history <str<strong>on</strong>g>of</str<strong>on</strong>g> the molecular chaper<strong>on</strong>e c<strong>on</strong>cept and some <str<strong>on</strong>g>of</str<strong>on</strong>g> the misc<strong>on</strong>cepti<strong>on</strong>s that pervade<br />

this field.<br />

7


23-26 August 2007,<br />

Budapest, Hungary<br />

P2<br />

GLUCOCORTICOIDS AS SCULPTORS OF ADAPTATION<br />

Mary F. Dallman, Norman C. Pecoraro, Susan F. Akana, James P. Warne, Abigail B. Ginsberg,<br />

Michelle T. Foster<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Physiology, UCSF, SanFrancisco CA USA<br />

e-mail: mary.dallman@ucsf.edu<br />

Hans Selye coined the terms glucocorticoids (GC), stressor and stress to describe,<br />

respectively, the characteristics <str<strong>on</strong>g>of</str<strong>on</strong>g> adrenal steroid horm<strong>on</strong>es, stimuli and resp<strong>on</strong>ses that<br />

occur when organisms are challenged. In the periphery, stressors, through the acti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

GC, mobilize energy stores from muscle and fat and increase gluc<strong>on</strong>eogenetic capacity<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the liver, and also increase circulating insulin, thus providing adequate glucose <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

use by brain and red blood cells, while freeing free fatty acids <str<strong>on</strong>g>for</str<strong>on</strong>g> use by muscle.<br />

Provisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> adequate energy sources is <strong>on</strong>ly <strong>on</strong>e part <str<strong>on</strong>g>of</str<strong>on</strong>g> the role <str<strong>on</strong>g>of</str<strong>on</strong>g> the GC, however.<br />

Once provided, the energy must be used through changes in behavior that are also<br />

mediated by stressor-induced secreti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these protean horm<strong>on</strong>es. GC feed <str<strong>on</strong>g>for</str<strong>on</strong>g>ward <strong>on</strong><br />

the brain to affect not <strong>on</strong>ly learning and memory, but also to enhance the intensity <str<strong>on</strong>g>of</str<strong>on</strong>g> behaviors that are<br />

determined by the immediate c<strong>on</strong>text. Thus, if male rats are in a social dominance-subordinance situati<strong>on</strong>, the<br />

steroids increase the level <str<strong>on</strong>g>of</str<strong>on</strong>g> aggressi<strong>on</strong> observed; if wheels are available, rats run more with high GC, and, if<br />

palatable food is available, they eat more. These resp<strong>on</strong>ses are in all likelihood a c<strong>on</strong>sequence <str<strong>on</strong>g>of</str<strong>on</strong>g> the direct<br />

acti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> the GC <strong>on</strong> m<strong>on</strong>oaminergic cell groups in brain as well as their pr<strong>on</strong>ounced augmentati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

corticotropin-releasing factor (CRF) in the emoti<strong>on</strong>al, limbic brain. Chr<strong>on</strong>ic stressors activate the limbic CRF<br />

network, which then biases behavioral, aut<strong>on</strong>omic and neuroendocrine outputs, thus changing resp<strong>on</strong>se<br />

characteristics. These changes occur with the alarm resp<strong>on</strong>se and characterize adaptati<strong>on</strong> to the stress<br />

resp<strong>on</strong>se. Together, the peripheral and central acti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> GC give us the fitness to survive and even flourish<br />

in times <str<strong>on</strong>g>of</str<strong>on</strong>g> adversity. (Support: DK28172, DA16944)<br />

8


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

P3<br />

THE BIOLOGY OF HAPPINESS AND POSITIVE WELL-BEING<br />

Andrew Steptoe<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Epidemiology and Public Health, University College L<strong>on</strong>d<strong>on</strong>, L<strong>on</strong>d<strong>on</strong>, UK<br />

e-mail: a.steptoe@ucl.ac.uk<br />

Happiness and positive well-being appear to have favourable effects <strong>on</strong> l<strong>on</strong>gevity<br />

and the development <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic illness. In a recent meta-analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> 29 studies<br />

involving several thousand healthy individuals, we found that positive affect was<br />

associated with an average 19% reducti<strong>on</strong> in the hazard ratio <str<strong>on</strong>g>for</str<strong>on</strong>g> mortality (p < .001).<br />

The processes resp<strong>on</strong>sible are not well understood, but both lifestyle factors (healthy<br />

habits) and direct biological pathways may be involved. This presentati<strong>on</strong> will<br />

describe studies indicating that happiness and health might be linked through<br />

psychobiological processes – namely differential central nervous system activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the neuroendocrine, immune, inflammatory and cardiovascular resp<strong>on</strong>ses that<br />

c<strong>on</strong>tribute to the development <str<strong>on</strong>g>of</str<strong>on</strong>g> physical disorders. We have carried naturalistic, clinical and experimental<br />

studies to assess the associati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> positive psychological states with cortisol output, markers <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

inflammati<strong>on</strong> (C-reactive protein, interleukin-6 and fribrinogen), cardiac aut<strong>on</strong>omic c<strong>on</strong>trol (heart rate<br />

variability) and blood pressure. Data will be presented which indicate that favourable patterns <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

neuroendocrine, inflammatory and cardiovascular resp<strong>on</strong>ses are plausible mediators <str<strong>on</strong>g>of</str<strong>on</strong>g> associati<strong>on</strong>s between<br />

happiness and health. Importantly, these associati<strong>on</strong>s are independent <str<strong>on</strong>g>of</str<strong>on</strong>g> depressi<strong>on</strong> and other measures <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

negative affect, suggesting that positive states have a distinctive relati<strong>on</strong>ship with biological resp<strong>on</strong>ses, and<br />

may c<strong>on</strong>tribute to prol<strong>on</strong>ged healthy adult life.<br />

9


23-26 August 2007,<br />

Budapest, Hungary<br />

P4<br />

NOVEL CHLOROPLAST-TARGETED PROTEINS REGULATING THE<br />

SUSCEPTIBILITY OF PHOTOSYNTHESIS TO ENVIRONMENTAL STRESSES<br />

Eva-Mari Aro<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Turku, FI-20014 Turku, Finland<br />

e-mail: evaaro@utu.fi<br />

Chloroplasts are key organelles in regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> plant growth via photosynthesis and<br />

producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> different metabolites and redox active compounds that functi<strong>on</strong> as<br />

signalling molecules. Accordingly, a proper functi<strong>on</strong> and development <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

chloroplasts in any envir<strong>on</strong>mental c<strong>on</strong>diti<strong>on</strong> is a prerequisite <str<strong>on</strong>g>for</str<strong>on</strong>g> stress tolerance. Due<br />

to this, chloroplasts are equipped with a battery <str<strong>on</strong>g>of</str<strong>on</strong>g> chaper<strong>on</strong>es and assisting proteins,<br />

as well as protein post-translati<strong>on</strong>al modificati<strong>on</strong>s, to guarantee the dynamic<br />

functi<strong>on</strong>, turnover and repair <str<strong>on</strong>g>of</str<strong>on</strong>g> the photosynthetic light reacti<strong>on</strong>s, particularly the<br />

light-sensitive Photosystem II, in a diversity <str<strong>on</strong>g>of</str<strong>on</strong>g> envir<strong>on</strong>mental stress c<strong>on</strong>diti<strong>on</strong>s. Our<br />

knowledge <strong>on</strong> the multitude and functi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> these auxiliary proteins and protein<br />

modificati<strong>on</strong>s is <strong>on</strong>ly emerging.<br />

We have taken several approaches, ranging from transcript pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iling and proteomics studies under various<br />

envir<strong>on</strong>mental stress c<strong>on</strong>diti<strong>on</strong>s to reverse genetics <str<strong>on</strong>g>of</str<strong>on</strong>g> several candidate genes, in order to identify novel<br />

proteins and protein post-translati<strong>on</strong>al modificati<strong>on</strong>s that assist the acclimati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the photosynthetic<br />

apparatus to adverse envir<strong>on</strong>mental c<strong>on</strong>diti<strong>on</strong>s.<br />

It is dem<strong>on</strong>strated that state transiti<strong>on</strong>s and Lhcb protein phosphorylati<strong>on</strong>, regulated by chloroplast STN7<br />

kinase according to redox c<strong>on</strong>diti<strong>on</strong>s, protect plants from high light stress and allow maximal photosynthesis<br />

under fluctuating light and elevated CO2, the combinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> envir<strong>on</strong>mental c<strong>on</strong>diti<strong>on</strong>s that str<strong>on</strong>gly<br />

challenge the capacity <str<strong>on</strong>g>of</str<strong>on</strong>g> photosynthetic light reacti<strong>on</strong>s. Another nuclear gene resp<strong>on</strong>ding to chloroplast<br />

signals is a 40 kDa thylakoid-associated phosphoprotein CAS (earlier identified as an Extracellular Ca 2+<br />

Sensing receptor in the plasma membrane: Han Sh. et al. 2003 Nature, 425, 196-200). We localize CAS to the<br />

thylakoid membrane, particularly to the stroma-exposed thylakoid regi<strong>on</strong>s. Phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> CAS in<br />

chloroplasts is catalysed by the STN8 kinase <strong>on</strong> the C-terminus <str<strong>on</strong>g>of</str<strong>on</strong>g> the protein. Similar to the transcript levels,<br />

also the c<strong>on</strong>tent <str<strong>on</strong>g>of</str<strong>on</strong>g> the CAS protein in the thylakoid membrane is dependent <strong>on</strong> growth irradiance, and both<br />

the light intensity and the availability <str<strong>on</strong>g>of</str<strong>on</strong>g> water str<strong>on</strong>gly influence the phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> CAS. CAS and its<br />

phosphorylati<strong>on</strong> seem to play a role in accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> lumenal proteins and thereby the functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

photosynthetic machinery.<br />

The expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> another chloroplast-targeted protein, DnaJ (J8), is rapidly suppressed in moderate light.<br />

The lower is the light intensity the higher are the DnaJ transcript and protein c<strong>on</strong>tents with maximum levels<br />

occurring in darkness. The physiological role <str<strong>on</strong>g>of</str<strong>on</strong>g> DnaJ in darkness (or the lack <str<strong>on</strong>g>of</str<strong>on</strong>g> the protein in the light) is<br />

discussed.<br />

Yet another set <str<strong>on</strong>g>of</str<strong>on</strong>g> chloroplast targeted auxiliary proteins was isolated from the thylakoid fracti<strong>on</strong> where the<br />

degradati<strong>on</strong> and synthesis <str<strong>on</strong>g>of</str<strong>on</strong>g> the photodamaged D1 protein <str<strong>on</strong>g>of</str<strong>on</strong>g> Photosystem II takes place. These novel<br />

proteins were shown crucial <str<strong>on</strong>g>for</str<strong>on</strong>g> efficient repair <str<strong>on</strong>g>of</str<strong>on</strong>g> photodamaged PSII centers and their specific functi<strong>on</strong>s<br />

will be discussed.<br />

10


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

P5<br />

GENES AND GENE NETWORKS CONTROLLING TOLERANCE TO HIGH<br />

TEMPERATURE<br />

Ung Lee, Jane Larkindale Eman Basha, Heather O’Neill, Elizabeth Vierling<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry & Molecular Biophysics, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Ariz<strong>on</strong>a, Tucs<strong>on</strong>, AZ 85721<br />

e-mail: vierling@u.ariz<strong>on</strong>a.edu<br />

The l<strong>on</strong>g term goals <str<strong>on</strong>g>of</str<strong>on</strong>g> our research are to understand the genetic pathways and<br />

biochemical comp<strong>on</strong>ents that c<strong>on</strong>fer tolerance to high temperature in plants. We are<br />

using <str<strong>on</strong>g>for</str<strong>on</strong>g>ward and reverse genetics, gene microarray methods and biochemistry to<br />

define comp<strong>on</strong>ents involved in the resp<strong>on</strong>se to high temperatures in the model plant<br />

Arabidopsis thaliana. Using <str<strong>on</strong>g>for</str<strong>on</strong>g>ward genetics, we isolated mutants unable to acclimate<br />

to heat stress (hot mutants). The first mutant uncovered was <str<strong>on</strong>g>of</str<strong>on</strong>g> the molecular<br />

chaper<strong>on</strong>e Hsp101 (hot1). We are c<strong>on</strong>tinuing genetics and biochemistry to define the<br />

mechanism and targets <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp101 and <str<strong>on</strong>g>of</str<strong>on</strong>g> related proteins in chloroplasts. Major<br />

recent ef<str<strong>on</strong>g>for</str<strong>on</strong>g>ts c<strong>on</strong>cern the hot5 locus, which encodes S-nitrosoglutathi<strong>on</strong>e reductase<br />

(GSNOR), a highly c<strong>on</strong>served type III alcohol dehydrogenase. GSNOR metabolizes GSNO, which is a nitric<br />

oxide (NO) adduct <str<strong>on</strong>g>of</str<strong>on</strong>g> glutathi<strong>on</strong>e. hot5 null mutants accumulate excess NO and an excess <str<strong>on</strong>g>of</str<strong>on</strong>g> nitrosated<br />

proteins. Scavenging <str<strong>on</strong>g>of</str<strong>on</strong>g> NO rescues the hot5 thermotolerance defect, c<strong>on</strong>sistent with the importance <str<strong>on</strong>g>of</str<strong>on</strong>g> NO<br />

regulati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> thermotolerance. Preliminary data suggest reversible oxidati<strong>on</strong> may regulate GSNOR enzyme<br />

activity. To identify heat stress signaling comp<strong>on</strong>ents we have screened both EMS and T-DNA mutagenized<br />

transgenic plants expressing an Hsp101promoter::Luciferase transgene. Reverse genetics and biochemistry are<br />

being used to investigate the family <str<strong>on</strong>g>of</str<strong>on</strong>g> small HSPs, including proteins localized to the cytosol (<str<strong>on</strong>g>for</str<strong>on</strong>g> which we<br />

have a crystal structure), the chloroplast, mitoch<strong>on</strong>dri<strong>on</strong> and peroxisome. We have also used whole-genome<br />

microarrays to identify new genes that may be involved in the acquisiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> thermotolerance.<br />

11


23-26 August 2007,<br />

Budapest, Hungary<br />

P6<br />

CHAPERONE-MEDIATED FOLDING IN THE MAMMALIAN ENDOPLASMIC<br />

RETICULUM<br />

Yuki Okuda-Shimizu, Yuichiro Shimizu, Ying Shen, Yi Jin, Linda M. Hendershot<br />

St. Jude Children’s <str<strong>on</strong>g>Research</str<strong>on</strong>g> Hospital, 332 N. Lauderdale, Memphis, TN 38105 USA<br />

e-mail: linda.hendershot@stjude.org<br />

The endoplasmic reticulum is the entry into the secretory pathway <str<strong>on</strong>g>of</str<strong>on</strong>g> eukaryotic cells<br />

and represents a major and particularly crowded site <str<strong>on</strong>g>of</str<strong>on</strong>g> protein biosynthesis. In additi<strong>on</strong><br />

to the complexity <str<strong>on</strong>g>of</str<strong>on</strong>g> protein folding in any organelle, the ER envir<strong>on</strong>ment poses further<br />

dangers and c<strong>on</strong>straints to the process. A quality c<strong>on</strong>trol mechanism exists to m<strong>on</strong>itor<br />

the maturati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins in the ER. Nascent polypeptide chains are specifically<br />

prevented from traveling further al<strong>on</strong>g the secretory pathway until they have completed<br />

their folding or assembly. Both the folding and m<strong>on</strong>itoring <str<strong>on</strong>g>of</str<strong>on</strong>g> proper maturati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

nascent proteins is c<strong>on</strong>trolled by molecular chaper<strong>on</strong>es. Proteins that cannot achieve a<br />

proper c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> are recognized and removed from the ER <str<strong>on</strong>g>for</str<strong>on</strong>g> degradati<strong>on</strong> by the<br />

26S proteasome. Recently progress has been made in understanding how chr<strong>on</strong>ically<br />

unfolded proteins are identified and a number <str<strong>on</strong>g>of</str<strong>on</strong>g> comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g> the cellular machinery<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> retrotranslocating these proteins to the cytosol have been identified. Finally, the homeostasis <str<strong>on</strong>g>of</str<strong>on</strong>g> the ER is<br />

vigilantly m<strong>on</strong>itored and changes that impinge up<strong>on</strong> the proper maturati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins in this organelle lead<br />

to the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a signal transducti<strong>on</strong> cascade that serves to restore balance to the ER. Recent studies<br />

suggest that some <str<strong>on</strong>g>of</str<strong>on</strong>g> these diverse functi<strong>on</strong>s may be achieve due to the organizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the ER into functi<strong>on</strong>al<br />

and perhaps even physical sub-domains.<br />

12


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

P7<br />

CHAPERONES AND MISFOLDED PROTEINS IN CELL STRESS AND DISEASE<br />

Tjakko van Ham 1 , Karen Thijssen 1 , R<strong>on</strong>ald Plasterk 2 , Ellen Nollen 1<br />

1 UMCG, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Genetics, Gr<strong>on</strong>ingen, The Netherlands,<br />

2Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Functi<strong>on</strong>al Genomics, Hubrecht Laboratories, Utrecht, The Netherlands<br />

e-mail: e.a.a.nollen@medgen.umcg.nl<br />

Protecti<strong>on</strong> against protein damage is vital to our cells. Accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> misfolded<br />

proteins can lead to cellular dysfuncti<strong>on</strong> and cell death. Protein misfolding occurs<br />

under stress c<strong>on</strong>diti<strong>on</strong>s and in various age-related neurodegenerative diseases, which<br />

includes Parkins<strong>on</strong>’s and Huntingt<strong>on</strong>’s disease. To identify processes involved in<br />

cellular protecti<strong>on</strong>, we search <str<strong>on</strong>g>for</str<strong>on</strong>g> genes that modify oligomerizati<strong>on</strong> and aggregati<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> misfolded proteins. We have developed a C. elegans model <str<strong>on</strong>g>for</str<strong>on</strong>g> Parkins<strong>on</strong>’s disease<br />

which captures several features <str<strong>on</strong>g>of</str<strong>on</strong>g> the disease. These include the age-dependent<br />

accumulati<strong>on</strong> and immobilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> alpha-synuclein in microscopically visible foci<br />

and recogniti<strong>on</strong> by an antibody to a disease specific phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> alphasynuclein<br />

at serine 129. We have per<str<strong>on</strong>g>for</str<strong>on</strong>g>med a genome-wide RNAi screen <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

modifiers <str<strong>on</strong>g>of</str<strong>on</strong>g> alpha-synuclein foci-<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> and identified 109 genes that enhanced foci-<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> after<br />

knock down by RNAi, most <str<strong>on</strong>g>of</str<strong>on</strong>g> which have human orthologues. Previously, we have identified genes that,<br />

when knocked down, cause premature protein aggregati<strong>on</strong> in a C.elegans model <str<strong>on</strong>g>for</str<strong>on</strong>g> polyglutamine aggregati<strong>on</strong>.<br />

Within this set are genes involved in protein synthesis, folding, degradati<strong>on</strong> and RNA synthesis and<br />

processing. Interestingly, although some <str<strong>on</strong>g>of</str<strong>on</strong>g> the major functi<strong>on</strong>al classes that have been previously picked up<br />

as suppressors <str<strong>on</strong>g>of</str<strong>on</strong>g> polyglutamine aggregati<strong>on</strong> modify foci <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> alpha-synclein as well, n<strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

genes identified overlap. These results suggest that cells employ different strategies to protect themselves<br />

against different misfolded proteins. In all, our data provide an understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular protecti<strong>on</strong> against<br />

misfolded proteins and may yield targets <str<strong>on</strong>g>for</str<strong>on</strong>g> therapy against misfolded protein diseases.<br />

13


14<br />

23-26 August 2007,<br />

Budapest, Hungary


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

CORE MODULE 1<br />

STRESS PROTEINS -<br />

MOLECULAR STRESS<br />

15


23-26 August 2007,<br />

Budapest, Hungary<br />

SYMPOSIA<br />

• 1A. Small Hsps (August 26 th Sunday morning)<br />

(Organizers and chairs: Andre-Patrick Arrigo, and Robert M. Tanguay)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> A1, Building A groundfloor<br />

• 1B. The Hsp70 family (August 25 th Saturday afterno<strong>on</strong>)<br />

(Organizers and chairs: Costa Georgopoulos and Kevin A. Morano)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> A1, Building A groundfloor<br />

• 1C. HSF activati<strong>on</strong> (August 25 th Saturday morning)<br />

(Organizer and chair: Lea Sist<strong>on</strong>en)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> A1, Building A groundfloor<br />

• 1D. Extracellular chaper<strong>on</strong>es (August 26 th Sunday morning)<br />

(Organizer and chair: Stuart Calderwood)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> A1, Building A groundfloor<br />

POSTER SYMPOSIA<br />

• 1E. Other chaper<strong>on</strong>e relates topics (Hsp60, Hsp90, Cochaper<strong>on</strong>es, Hsp crystallography<br />

and structures, Stress <str<strong>on</strong>g>of</str<strong>on</strong>g> single molecules)<br />

August 25 th Saturday, Poster secti<strong>on</strong> A1, Building A groundfloor<br />

• 1F. Pharmacological modulators <str<strong>on</strong>g>of</str<strong>on</strong>g> stress protein expressi<strong>on</strong><br />

August 25 th Saturday, Poster secti<strong>on</strong> A1, Building A groundfloor<br />

16


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

1A. SMALL HSPS<br />

(ANDRE-PATRICK ARRIGO, ROBERT M. TANGUAY)<br />

1A_01_S<br />

HSP27 (HSPB1) AS A THERAPEUTIC TARGET<br />

André-Patrick Arrigo<br />

Stress, Chaper<strong>on</strong>s and Cell Death Laboratory, CGMC, CNRS UMR 5534, Claude Bernard University LYON<br />

1, 16 Rue Dubois, Bat. Gregor Mendel, 69622 Villeurbanne, France<br />

e-mail: arrigo@univ-ly<strong>on</strong>1.fr<br />

Human Hsp27(HspB1) is a molecular chaper<strong>on</strong>e which is c<strong>on</strong>stitutively expressed in several mammalian cells,<br />

particularly in pathological c<strong>on</strong>diti<strong>on</strong>s. This protein has functi<strong>on</strong>s as diverse as protecti<strong>on</strong> against toxicity<br />

mediated by aberrantly folded proteins or oxidative-inflammati<strong>on</strong> c<strong>on</strong>diti<strong>on</strong>s. In additi<strong>on</strong>, it has antiapoptotic<br />

properties and is tumorigenic when expressed in cancer cells. Hsp27(HspB1) has implicati<strong>on</strong>s,<br />

either positive or deleterious, in pathologies such as neurodegenerative diseases, asthma, and cancers.<br />

Moreover, mutati<strong>on</strong>s in hsp27 gene have been detected which are resp<strong>on</strong>sive <str<strong>on</strong>g>of</str<strong>on</strong>g> hereditary motor<br />

neuropathies. Hsp27(HspB1) is there<str<strong>on</strong>g>for</str<strong>on</strong>g>e an active determinant in health and disease and not just a passive<br />

storage device. Approaches as well as preliminary results towards therapeutic strategies aimed at modulating<br />

the expressi<strong>on</strong> and/or the activities <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp27(HspB1) will be presented.<br />

1A_02_S<br />

HSPB8 FORMS WITH BAG3 A CHAPERONE COMPLEX STIMULATING<br />

MACROAUTOPHAGY: POSSIBLE INVOLVEMENT OF SHSP IN PROTEIN<br />

QUALITY CONTROL<br />

Jacques Landry<br />

<str<strong>on</strong>g>Centre</str<strong>on</strong>g> de recherche en cancérologie de l’Université Laval, L’Hôtel-Dieu de Québec, Québec, Canada, G1R 2J6,<br />

e-mail: jacques.landry@med.ulaval.ca<br />

Intracellular protein aggregati<strong>on</strong> can occur up<strong>on</strong> proteotoxic stress or genetic mutati<strong>on</strong>s and represents a<br />

major threat in the crowded envir<strong>on</strong>ment <str<strong>on</strong>g>of</str<strong>on</strong>g> the cell. The Hsp70/Hsp90 proteins and their associated cochaper<strong>on</strong>es<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g>m a protein quality c<strong>on</strong>trol system which can recognize such proteins and either assist them in<br />

renaturati<strong>on</strong> or target them <str<strong>on</strong>g>for</str<strong>on</strong>g> degradati<strong>on</strong>. Proteins from the small Hsp family, and in particular HspB1<br />

(Hsp27) and HspB8, may similarly act as molecular chaper<strong>on</strong>es as evidenced by their associati<strong>on</strong> with human<br />

diseases featuring protein c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> defects. Diverse types <str<strong>on</strong>g>of</str<strong>on</strong>g> activities are associated with each <str<strong>on</strong>g>of</str<strong>on</strong>g> these<br />

proteins. On <strong>on</strong>e hand, HspB1 (Hsp27) but not HspB8 potentiates the renaturati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> damaged proteins in<br />

the cells. In this activity, HspB1 recognizes the substrate and then recruits a renaturating machinery likely<br />

involving Hsp70 family members. On the other hand, HspB8, but not HspB1, <str<strong>on</strong>g>for</str<strong>on</strong>g>ms in cells a stable protein<br />

complex with Bag3, a Bag family member previously identified as a Hsp70 co-chaper<strong>on</strong>e. In associati<strong>on</strong> with<br />

Bag3 but independently Bag3 binding to Hsp70, HspB8 efficiently limits the accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> aggregati<strong>on</strong>pr<strong>on</strong>e<br />

protein substrates by targeting them to destructi<strong>on</strong> by macroautophagy, a lysosome-based protein<br />

degradati<strong>on</strong> system capable <str<strong>on</strong>g>of</str<strong>on</strong>g> degrading large structures and insoluble protein aggregates. In this complex<br />

HspB8 likely acts as a molecular chaper<strong>on</strong>e resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> the recogniti<strong>on</strong> the damaged substrates whereas<br />

Bag3 is resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> recruiting and stimulating the macroautophagy machinery. Accordingly, HspB1<br />

engineered to interact with Bag3 can also target unstable proteins to degradati<strong>on</strong>. Hence HspB proteins can,<br />

17


23-26 August 2007,<br />

Budapest, Hungary<br />

like Hsp70/Hsp90, functi<strong>on</strong> in protein quality c<strong>on</strong>trol by recognizing protein substrates, the fate <str<strong>on</strong>g>of</str<strong>on</strong>g> which<br />

being determined by associated co-chaper<strong>on</strong>es that are specific to individual chaper<strong>on</strong>es.<br />

1A_03_S<br />

RECOVERY OF MACROMOLECULAR SYNTHESIS AFTER STRESS: THE ROLE OF<br />

SMALL HEAT SHOCK PROTEIN<br />

Nicolette H. Lubsen<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biomolecular Chemistry, Radboud University Nijmegen<br />

e-mail: N.Lubsen@science.ru.nl<br />

Cells stressed by heat downregulate protein synthesis. During recovery from heat stress protein synthesis<br />

slowly recovers. In the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> either αB-crystallin or Hsp27, both small heat shock proteins, the rate <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

recovery is enhanced. Using fluorescence recovery after photobleaching we showed that Hsp27, but not αBcrystallin,<br />

increased the pool <str<strong>on</strong>g>of</str<strong>on</strong>g> mobile stress granule-associated EGFP-eIF4E in heat shocked cells. Hsp27<br />

also partially prevented the sharp decrease in the pool <str<strong>on</strong>g>of</str<strong>on</strong>g> mobile cytoplasmic EGFP-eIF4G, supporting other<br />

evidence <str<strong>on</strong>g>for</str<strong>on</strong>g> a direct interacti<strong>on</strong> between eIF4G and Hsp27. sHsps did not prevent the phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

eIF2α by a heat shock, but promoted dephosphorylati<strong>on</strong> during recovery. Blocking the endogenous heat<br />

shock resp<strong>on</strong>se by expressing a dominant negative HSF1 mutant during recovery from a heat shock slows the<br />

rate <str<strong>on</strong>g>of</str<strong>on</strong>g> recovery <str<strong>on</strong>g>of</str<strong>on</strong>g> protein synthesis and blocks the restorative effect <str<strong>on</strong>g>of</str<strong>on</strong>g> sHsps, showing that sHsps need to<br />

cooperate with other Hsps <str<strong>on</strong>g>of</str<strong>on</strong>g> which the synthesis is induced by heat shock. Translati<strong>on</strong>al recovery 24 hours<br />

after heat shock does not differ between cells expressing dnHSF1 and c<strong>on</strong>trol cells. However, if cells express<br />

dnHSF1 as well as the C-terminal fragment <str<strong>on</strong>g>of</str<strong>on</strong>g> GADD34, which causes c<strong>on</strong>stitutive dephosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

eIF2α, translati<strong>on</strong> does not recover. These data show that two partially redundant pathways are involved in<br />

translati<strong>on</strong>al recovery from a heat shock.<br />

1A_04_S<br />

REGULATION OF SMALL HEAT SHOCK PROTEINS IN AGEING AND<br />

RESISTANCE TO STRESS<br />

Robert M. Tanguay, Geneviève Morrow, Hyun-Ju Kim, Sébastien Michaud<br />

Lab Cellular Developmental Genetics, CREFSIP, Dept Medecine, Pav. Marchand, Université Laval, Québec,<br />

Canada G1K 7P4<br />

e-mail: Robert.tanguay@rsvs.ulaval<br />

Small HSP are involved in the refolding and/or disposal <str<strong>on</strong>g>of</str<strong>on</strong>g> protein aggregates, a feature <str<strong>on</strong>g>of</str<strong>on</strong>g> many ageassociated<br />

diseases. In Drosophila melanogaster, there are 4 main small Hsps each residing in a different<br />

intracellular compartment. Targeting the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the mitoch<strong>on</strong>drial Hsp22, to different cell types can<br />

increase lifespan by more than 30%. L<strong>on</strong>g-lived flies expressing Hsp22 in motorneur<strong>on</strong>s have an increased<br />

resistance to oxidative stress and maintain their locomotor activity l<strong>on</strong>ger. Over expressing the cytosolic<br />

Hsp23 in a pan-neur<strong>on</strong>al fashi<strong>on</strong> (transgenic GAL4/UAS system) also increases l<strong>on</strong>gevity by 15%.<br />

C<strong>on</strong>versely, a strain carrying an inserti<strong>on</strong> in the promoter <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp23 which downregulates its expressi<strong>on</strong> in<br />

specific cells <str<strong>on</strong>g>of</str<strong>on</strong>g> the embry<strong>on</strong>ic CNS and in adults has a decreased lifespan. The acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these chaper<strong>on</strong>es <strong>on</strong><br />

lifespan likely involves different pathways as suggested by the l<strong>on</strong>gevity curves, and their respective site and<br />

developmental pattern <str<strong>on</strong>g>of</str<strong>on</strong>g> expressi<strong>on</strong>. Microarrays analysis was used to unveil the mechanisms involved in<br />

18


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

lifespan. The transcripti<strong>on</strong>al changes brought by Hsp22 overexpressi<strong>on</strong> occur early in adulthood and are<br />

associated with genes involved in energy metabolism, protein biosynthesis, and protein folding. The relati<strong>on</strong><br />

between the insulin/IGF signaling pathway and the heat shock resp<strong>on</strong>se has also been examined using flies<br />

with mutati<strong>on</strong>s in the heat shock factor HSF and dFOXO. Altogether, these results c<strong>on</strong>firm a beneficial role<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> small chaper<strong>on</strong>es and corroborate the pivotal role <str<strong>on</strong>g>of</str<strong>on</strong>g> the nervous system and the<br />

insulin/IGF pathway in the ageing process. Supported by CIHR (Canada) and the EU 6 th Framework<br />

Programme MiMage.<br />

1A_05_S<br />

(poster secti<strong>on</strong> A1, poster board #1, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

INHIBITION OF APOPTOTIC CELL DEATH BY A NOVEL 16.2 KD HEAT SHOCK<br />

PROTEIN VIA HSP90 MEDIATED LIPID RAFTS STABILIZATION AND AKT<br />

ACTIVATION PATHWAY<br />

Balazs Sumegi 1 , Szabolcs Bellyei 1,2 , Eva Pozsgai 1 , Andras Szigeti 1,2 , Arpad Bor<strong>on</strong>kai 1,2 ,<br />

Ferenc Gallyas Jr. 1<br />

Departments <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry and Medical Chemistry 1 , Oncotherapy 2 , University <str<strong>on</strong>g>of</str<strong>on</strong>g> Pécs, Pécs, Hungary<br />

Corresp<strong>on</strong>dence to: Balazs Sumegi, PhD, DSci, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry and Medical Chemistry, University <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Pécs, 12 Szigeti Street, Pécs H-7624, Hungary. Tel: +36-72-536-276 Fax36-72-536-277<br />

e-mail: balazs.sumegi@aok.pte.hu<br />

AlphaB-crystallin homology, heat stress inducti<strong>on</strong> and chaper<strong>on</strong>e activity suggested that a previously<br />

encl<strong>on</strong>ed gene product is a novel small heat shock protein (Hsp16.2). Suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp16.2 by siRNA<br />

sensitized cells to hydrogen peroxide or taxol induced cell-death. While over-expressing <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp16.2 protected<br />

cells against stress stimuli by inhibiting cytochrome c release from the mitoch<strong>on</strong>dria, nuclear translocati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

AIF and end<strong>on</strong>uclease G, and caspase 3 activati<strong>on</strong>. Recombinant Hsp16.2 protected mitoch<strong>on</strong>drial<br />

membrane potential against calcium induced collapse in vitro indicating that Hsp16.2 stabilizes mitoch<strong>on</strong>drial<br />

membrane systems. Hsp16.2 <str<strong>on</strong>g>for</str<strong>on</strong>g>med self-aggregates and bound to Hsp90. Inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90 by<br />

geldanamycin diminished the cytoprotective effect <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp16.2 indicating that this effect was Hsp90-mediated.<br />

Hsp16.2 over-expressi<strong>on</strong> increased lipid rafts <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> as dem<strong>on</strong>strated by increased cell surface labeling<br />

with fluorescent cholera toxin B, and increased Akt phosphorylati<strong>on</strong>. The inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> PI-3-kinase-Akt<br />

pathway by LY-294002 or wortmannin significantly decreased the protective effect <str<strong>on</strong>g>of</str<strong>on</strong>g> the Hsp16.2. These<br />

data indicate that the over-expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp16.2 inhibits cell death via the stabilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>drial<br />

membrane system, activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90, stabilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> lipid rafts and by the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> PI-3-kinase - Akt<br />

cytoprotective pathway.<br />

19


23-26 August 2007,<br />

Budapest, Hungary<br />

1A_06_S<br />

(poster secti<strong>on</strong> A1, poster board #2, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

MODULATION OF THE CHAPERONE-LIKE FUNCTION OF SMALL HEAT SHOCK<br />

PROTEINS BY METHYLGLYOXAL<br />

Ram H. Nagaraj, Ashis Biswas, Manjunatha Bhat<br />

Case Western Reserve, University and Cleveland Clinic Foundati<strong>on</strong>, Cleveland, OH, USA<br />

Alpha-crystallin and Hsp27 bel<strong>on</strong>g to the family <str<strong>on</strong>g>of</str<strong>on</strong>g> small heat shock proteins. They are stress proteins and<br />

play an important role in preventing protein aggregati<strong>on</strong> and cell death by external stress. Methylglyoxal is an<br />

ubiquitous alpha-dicarb<strong>on</strong>yl compound produced from the triose phosphate intermediates <str<strong>on</strong>g>of</str<strong>on</strong>g> glycolysis. It<br />

reacts rapidly with arginine, cysteine and lysine residues in proteins and chemically modifies them to <str<strong>on</strong>g>for</str<strong>on</strong>g>m<br />

stable adducts. We have studied the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> methylglyoxal modificati<strong>on</strong> <strong>on</strong> the chaper<strong>on</strong>e-like functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

alpha-crystallin and Hsp27. Methylglyoxal modificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these two stress proteins increased their chaper<strong>on</strong>e<br />

functi<strong>on</strong> <strong>on</strong> a c<strong>on</strong>centrati<strong>on</strong>-dependent manner. Specific arginine modificati<strong>on</strong> to argpyrimidine was found to<br />

be resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> the enhanced chaper<strong>on</strong>e functi<strong>on</strong>. Site-directed mutagenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> methylglyoxal-modifiable<br />

arginine residues to alanine mirrored the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> methylglyoxal. Introducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> additi<strong>on</strong>al guanidino<br />

groups abolished the chaper<strong>on</strong>e-like functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> alphaA-crystallin but subsequent modificati<strong>on</strong> by<br />

methylglyoxal not <strong>on</strong>ly revived the chaper<strong>on</strong>e-like functi<strong>on</strong> but also made it better than the unmodified<br />

protein. Modificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> both substrate proteins and alphaA-crystallin by methylglyoxal further enhanced<br />

resistance to protein aggregati<strong>on</strong> by thermal and chemical stress. These results suggest that physiological<br />

dicarb<strong>on</strong>yl methylglyoxal promotes the chaper<strong>on</strong>e functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> alphacrystallin and Hsp27 and prevents<br />

aggregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins and thus may play a vital role in cell resp<strong>on</strong>se to stress in health and disease.<br />

1A_07_S<br />

(poster secti<strong>on</strong> A1, poster board #3, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

TWO SMALL HEAT SHOCK PROTEINS OF A FISSION YEAST FUNCTION IN<br />

DIFFERENT MANNER TO COPE WITH WIDE RANGE OF TEMPERATURES AND<br />

VARIOUS DENATURED PROTEINS<br />

Masafumi Yohda 1* , Chika Sugino 1 , Maya Hirose 1 , Ryo Iizuka 1 , Masafumi Shimizu 2 ,<br />

Shun-ichi Kidokoro 3 , Noriyuki Ishii 4<br />

1Tokyo University <str<strong>on</strong>g>of</str<strong>on</strong>g> Agriculture and Technology, Koganei, Tokyo;<br />

3Tokyo University <str<strong>on</strong>g>of</str<strong>on</strong>g> Technology, Hachioji, Tokyo;<br />

4Nagaoka University <str<strong>on</strong>g>of</str<strong>on</strong>g> Technology, Nagaoka-shi, Niigata;<br />

5Nati<strong>on</strong>al Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Advanced Industrial Science and Technology, Tsukuba-shi, Ibaraki, Japan<br />

*e-mail: yohda@cc.tuat.ac.jp<br />

There exist two small heat shock proteins (SpHsp15.8 and SpHsp16.0) in the fissi<strong>on</strong> yeast,<br />

Schizosaccharomyces pombe (S. pombe). At the elevated temperatures, both SpHsp15.8 and SpHsp16.0<br />

dissociate into small oligomers and then interact with denatured substrate proteins. SpHsp16.0 exhibited clear<br />

enthalpy change <str<strong>on</strong>g>for</str<strong>on</strong>g> denaturati<strong>on</strong> at over 60 ºC in differential scanning calorimetry. In additi<strong>on</strong>, there was<br />

another small enthalpy change at about 50 ºC, which is likely to corresp<strong>on</strong>d to oligomer dissociati<strong>on</strong>. The<br />

oligomer dissociati<strong>on</strong> and interacti<strong>on</strong> with denatured protein <str<strong>on</strong>g>of</str<strong>on</strong>g> SpHsp15.8 and SpHsp16.0 were analyzed by<br />

fluorescence polarizati<strong>on</strong> analysis (FPA). Both sHsps exhibited temperature dependent decrease <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

20


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

fluorescence polarizati<strong>on</strong>, which correlates with the dissociati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> large oligomers to small oligomers. The<br />

dissociati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> SpHsp15.8 oligomer started to occur at about 35 ºC and proceeds gradually. On the c<strong>on</strong>trary,<br />

SpHsp16.0 oligomer was stable up to about 45 ºC, but dissociate to small oligomers abruptly at the<br />

temperature. Interacti<strong>on</strong> between sHsps and denatured CS at the elevated temperature was also examined by<br />

FPA. Interestingly, SpHsp16.0 is likely to interact with denatured CS in the dissociated state, but SpHsp15.8<br />

in the large complex in c<strong>on</strong>trast. The results suggest that S. pombe, utilizes two sHsps to cope with wide<br />

range <str<strong>on</strong>g>of</str<strong>on</strong>g> temperature and various denatured proteins.<br />

21


23-26 August 2007,<br />

Budapest, Hungary<br />

22<br />

1A_01_P<br />

(poster secti<strong>on</strong> A1, poster board #4, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HSPB8 AND BAG3: A NEW CHAPERONE COMPLEX STIMULATING MUTATED<br />

HUNTINGTIN DEGRADATION BY MACROAUTOPHAGY<br />

S. Carra* 1 , S. Seguin 1 , J. Vinet 2 , H. Lambert 1 , A. Sik 2 , Jacques Landry 1<br />

1<str<strong>on</strong>g>Centre</str<strong>on</strong>g> de recherche L'Hôtel-Dieu de Québec, Université Laval, 9 rue McMah<strong>on</strong>, Québec, Canada;<br />

e-mail: jacques.landry@med.ulaval.ca<br />

2<str<strong>on</strong>g>Centre</str<strong>on</strong>g> de recherche Robert-Giffard, 2601 Chemin de la Canardière, Québec, Canada<br />

HspB8 is a member <str<strong>on</strong>g>of</str<strong>on</strong>g> the small heat shock proteins or HspB family <str<strong>on</strong>g>of</str<strong>on</strong>g> molecular chaper<strong>on</strong>es, which<br />

comprises ten members in mammals (HspB1-10). We previously dem<strong>on</strong>strated that, in cultured cells,<br />

overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HspB8, but not <str<strong>on</strong>g>of</str<strong>on</strong>g> HspB1 or HspB5, totally blocked the insolubilizati<strong>on</strong> and accumulati<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> a pathogenic aggregati<strong>on</strong>-pr<strong>on</strong>e <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> Huntingtin (Htt43Q). Here we report that HspB8 stably and<br />

sotichiometrically interacts with the co-chaper<strong>on</strong>e Bag3. HspB8 associati<strong>on</strong> with Bag3 is essential <str<strong>on</strong>g>for</str<strong>on</strong>g> both its<br />

structural stability and chaper<strong>on</strong>e functi<strong>on</strong>, as dem<strong>on</strong>strated by knocking down Bag3 expressi<strong>on</strong> by siRNA<br />

technique. We next investigated the chaper<strong>on</strong>e functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the HspB8/Bag3 complex. We found that Bag3<br />

overexpressi<strong>on</strong>, like HspB8, facilitated Htt43Q degradati<strong>on</strong> in cultured cells. Treatment with specific<br />

macroautophagy inhibitors dramatically decreased the HspB8/Bag3 chaper<strong>on</strong>e activity and lead to the<br />

accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> aggregated Htt43Q. Moreover, HspB8 and Bag3 both increased the number <str<strong>on</strong>g>of</str<strong>on</strong>g> cells<br />

c<strong>on</strong>taining LC3 positive-vacuoles and stimulated the lipidati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> LC3, a step which is necessary <str<strong>on</strong>g>for</str<strong>on</strong>g> LC3<br />

incorporati<strong>on</strong> into the autophagosomes. These results str<strong>on</strong>gly suggest the implicati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

macroautophagy process in the HspB8/Bag3 complex mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> acti<strong>on</strong>. By joining the ability <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

recognizing endogenous misfolded proteins and <str<strong>on</strong>g>of</str<strong>on</strong>g> stimulating the macroautophagic vacuole <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>, the<br />

new HspB8/Bag3 chaper<strong>on</strong>e complex may play a crucial role in the protein quality c<strong>on</strong>trol system<br />

resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> eliminating potentially harmful aggregating proteins.<br />

1A_02_P<br />

(poster secti<strong>on</strong> A1, poster board #5, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PHOSPHORYLATION OF HUMAN SMALL HEAT SHOCK PROTEIN HSP22 BY<br />

CAMP-DEPENDENT PROTEIN KINASE<br />

Ant<strong>on</strong> A. Shemetov, Ivan S. Chernik, Alim S. Seit-Nebi, Nikolai B. Gusev<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, School <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, Moscow State University, Moscow 119992, Russia<br />

The data <str<strong>on</strong>g>of</str<strong>on</strong>g> literature (Benndorf et al., J. Biol. Chem.276, 26753, 2001) indicate that small heat shock protein<br />

with molecular mass 22 kDa (HSP22, HspB8 or H11 kinase) can be phosphorylated by protein kinase C and<br />

p44 MAP kinase. Ser24 and Ser57 <str<strong>on</strong>g>of</str<strong>on</strong>g> human HSP22 are located in c<strong>on</strong>sensus sequence RXS recognized by<br />

cAMP-dependent protein kinase and we supposed that this enzyme could also be involved in HSP22<br />

phosphorylati<strong>on</strong>. To check this suggesti<strong>on</strong> we obtained three mutants (S24D, S57D, S24,57D) <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP22 and<br />

analyzed phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the wild type protein and these mutants by cAMP-dependent protein kinase.<br />

Wild type HSP22 and its S24D mutants were rapidly phosphorylated up to 1 mole <str<strong>on</strong>g>of</str<strong>on</strong>g> phosphate per mole <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

protein, whereas S57D and S24,57D mutants were not phosphorylated by cAMP-dependent protein kinase.<br />

These data indicate that S57D is the primary site phosphorylated by cAMP-dependent protein kinase.<br />

Phosphorylati<strong>on</strong> (or mutati<strong>on</strong>s mimicking phosphorylati<strong>on</strong>) does not affect the oligomeric structure <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

HSP22 analyzed by size-exclusi<strong>on</strong> chromatography and has no effect <strong>on</strong> chemical crosslinking <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP22. At


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

the same time mutati<strong>on</strong>s mimicking phosphorylati<strong>on</strong> affect accessibility <str<strong>on</strong>g>of</str<strong>on</strong>g> Trp residues to solvent and<br />

increase susceptibility <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP22 to chymotrypsinolysis. Mutati<strong>on</strong>s (or phosphorylati<strong>on</strong>) decrease chaper<strong>on</strong>elike<br />

activity <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP22 if insulin or rhodanase were used as model protein substrates. It is c<strong>on</strong>cluded that<br />

HSP22 can be phosphorylated by cAMP-dependent protein kinase and phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Ser residues<br />

located in the N-terminal domain might affect its structure and chaper<strong>on</strong>e-like activity.<br />

1A_03_P<br />

(poster secti<strong>on</strong> A1, poster board #6, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

NEW IDEAS ON MECHANISM OF PROTEIN AGGREGATION AND MECHANISM<br />

OF PROTECTIVE ACTION OF α-CRYSTALLIN<br />

K. A. Markossian, B. I. Kurganov<br />

Bach Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, Russian Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences, 119071 Moscow, Leninsky pr. 33<br />

e-mail: markossian@inbi.ras.ru, boris@kurganov.com<br />

The kinetics <str<strong>on</strong>g>of</str<strong>on</strong>g> thermal aggregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins (β-crystallin from calf eye lens, glyceraldehyde-3-phosphate<br />

dehydrogenase from rabbit skeletal muscle and mitoch<strong>on</strong>drial aspartate aminotransferase from porcine heart)<br />

were studied by dynamic light scattering. On the basis <str<strong>on</strong>g>of</str<strong>on</strong>g> the study <str<strong>on</strong>g>of</str<strong>on</strong>g> aggregati<strong>on</strong> kinetics a new mechanism<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> protein aggregati<strong>on</strong> was developed. The first stage <str<strong>on</strong>g>of</str<strong>on</strong>g> protein aggregati<strong>on</strong> is the stage <str<strong>on</strong>g>of</str<strong>on</strong>g> the <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the start aggregates. The size <str<strong>on</strong>g>of</str<strong>on</strong>g> the start aggregates was estimated by c<strong>on</strong>structi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the light scattering<br />

intensity versus hydrodynamic radius plots. The hydrodynamic radius <str<strong>on</strong>g>of</str<strong>on</strong>g> the start aggregates remains c<strong>on</strong>stant<br />

at variati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> temperature and the protein c<strong>on</strong>centrati<strong>on</strong>. The sec<strong>on</strong>d stage <str<strong>on</strong>g>of</str<strong>on</strong>g> the aggregati<strong>on</strong> process is<br />

sticking together <str<strong>on</strong>g>of</str<strong>on</strong>g> the start aggregates. This stage proceeds in the kinetic regime wherein the rate <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

aggregati<strong>on</strong> is limited by diffusi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the particles (the regime <str<strong>on</strong>g>of</str<strong>on</strong>g> “diffusi<strong>on</strong>-limited cluster-cluster<br />

aggregati<strong>on</strong>”). Under this regime each collisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the interacting particles results in their sticking together.<br />

Suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> thermal aggregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> α-crystallin is due to diminishing <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

size <str<strong>on</strong>g>of</str<strong>on</strong>g> the start aggregates, increase in the durati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the latent period over which the start aggregates are<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g>med, and transiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the aggregati<strong>on</strong> process into the kinetic regime wherein the sticking probability <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

the colliding particles becomes less than unity (the regime <str<strong>on</strong>g>of</str<strong>on</strong>g> “reacti<strong>on</strong>-limited cluster-cluster aggregati<strong>on</strong>”).<br />

1A_04_P<br />

(poster secti<strong>on</strong> A1, poster board #7, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

TRANSDUCED TAT-HSP40 PROTECTS CELLS AGAINST OXIDATIVE STRESS<br />

Jung-Ho<strong>on</strong> Yo<strong>on</strong> 1 , Soo-A Kim 2 , Sang-Gun Ahn 1,*<br />

1Oral Biology <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute, BK21 projects, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Pathology, Chosun University College <str<strong>on</strong>g>of</str<strong>on</strong>g> Dentistry,<br />

Gwangju, KOREA, 2 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, D<strong>on</strong>gguk University College <str<strong>on</strong>g>of</str<strong>on</strong>g> Oriental Medicine, Gye<strong>on</strong>gju,<br />

Korea, * corresp<strong>on</strong>ding author<br />

Heat shock protein (Hsp) 40 acts as a co-chaper<strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 by facilitating the ATPase activity <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 as<br />

well as promoting the protein folding and renaturati<strong>on</strong> by Hsp70. In the present study, Hsp40 gene was fused<br />

with a gene fragment encoding the HIV-1 TAT protein transducti<strong>on</strong> domain (YGRKKRRQRRR) in a<br />

bacterial expressi<strong>on</strong> vector pTAT-HA to produce the TAT-fused Hsp40 (TAT-Hsp40). Purified TAT-Hsp40<br />

was effectively transduced into the HEK 293 cells in a time- and dose-dependent manner. To examine the<br />

effect <str<strong>on</strong>g>of</str<strong>on</strong>g> TAT-Hsp40 up<strong>on</strong> oxidative stress, HEK293 cells were exposed to H 2 O 2 . Oxidative stress induced<br />

23


23-26 August 2007,<br />

Budapest, Hungary<br />

the rapid increase <str<strong>on</strong>g>of</str<strong>on</strong>g> proteasome activity followed by cell death in HEK 293 cells. However, HEK 293 cells<br />

transduced by TAT-Hsp40 showed resistance against oxidative stress-induced cytotoxicity. TAT-Hsp40<br />

transduced cells showed decreased proteasome activity and inhibited Hsp70 degradati<strong>on</strong>. These results<br />

suggest that Hsp40 might protect cell death from oxidative stress by preserving the cellular level <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70.<br />

24<br />

1A_05_P<br />

(poster secti<strong>on</strong> A1, poster board #8, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECT OF ALPHA-CRYSTALLIN ON THERMAL DENATURATION AND<br />

AGGREGATION OF MITOCHONDRIAL ASPARTATE AMINOTRANSFERASE<br />

N. V. Golub, K. A. Markossian<br />

Bach Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry RAS, 33 Leninsky av., Moscow, 119071, Russia<br />

e-mail: ngolub@inbi.ras.ru<br />

Thermal denaturati<strong>on</strong> and aggregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>drial aspartate aminotransferase have been investigated by<br />

differential scanning calorimetry (DSC) and dynamic light scattering. The dependence <str<strong>on</strong>g>of</str<strong>on</strong>g> excess heat capacity<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> AAT <strong>on</strong> temperature passes through a maximum at 72.4 degrees C and can be described by a scheme in<br />

which the denaturati<strong>on</strong> process is c<strong>on</strong>sidered as an irreversible first-order reacti<strong>on</strong>. Inactivati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> AAT<br />

follows the exp<strong>on</strong>ential law. The parameters <str<strong>on</strong>g>of</str<strong>on</strong>g> Arrhenius equati<strong>on</strong> have been determined from the DSC data<br />

and temperature dependence <str<strong>on</strong>g>of</str<strong>on</strong>g> the inactivati<strong>on</strong> rate c<strong>on</strong>stant. Calculati<strong>on</strong>s show that at 57.5 degrees C the<br />

inactivati<strong>on</strong> c<strong>on</strong>stant (k in ) is 28.9 times higher than the denaturati<strong>on</strong> rate c<strong>on</strong>stant (k den ) determined from the<br />

DSC data. The k in /k den ratio decreases with temperature and becomes equal to 1.3 at 77 degrees С. It has been<br />

shown that alpha-crystallin, the protein possessing a chaper<strong>on</strong>e-like activity, reduces thermostability <str<strong>on</strong>g>of</str<strong>on</strong>g> ААТ<br />

and accelerates thermoinactivati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the enzyme. Suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> thermal aggregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ААТ in the<br />

presence <str<strong>on</strong>g>of</str<strong>on</strong>g> alpha-crystallin has been dem<strong>on</strong>strated. The protective effect <str<strong>on</strong>g>of</str<strong>on</strong>g> alpha-crystallin is due to the<br />

decrease in the size <str<strong>on</strong>g>of</str<strong>on</strong>g> the start aggregates and transiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the aggregati<strong>on</strong> process from the regime <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

diffusi<strong>on</strong>-limited cluster-cluster aggregati<strong>on</strong> (the sticking probability <str<strong>on</strong>g>for</str<strong>on</strong>g> the colliding particles is equal to<br />

unity) to the regime <str<strong>on</strong>g>of</str<strong>on</strong>g> reacti<strong>on</strong>-limited cluster-cluster aggregati<strong>on</strong> (the sticking probability <str<strong>on</strong>g>for</str<strong>on</strong>g> the colliding<br />

particles is less than unity). The study was supported by the Russian Foundati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> Basic <str<strong>on</strong>g>Research</str<strong>on</strong>g> (grants<br />

05-04-48691-a and 06-04-39008), Program “Molecular and Cell Biology” <str<strong>on</strong>g>of</str<strong>on</strong>g> the Presidium <str<strong>on</strong>g>of</str<strong>on</strong>g> the Russian<br />

Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences and INTAS (grant 03-51-4813).<br />

1A_06_P<br />

(poster secti<strong>on</strong> A1, poster board #9, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HSP26 FROM SACCHAROMYCES CEREVISIAE IS A POLYDISPERSE SMALL HEAT-<br />

SHOCK PROTEIN<br />

J. A. Aquilina, J. L. P. Benesch, R. A. Lindner, A. Rekas, E. Vierling, C. V. Robins<strong>on</strong>, J. A. Carver<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Woll<strong>on</strong>g<strong>on</strong>g, Northfields Avenue, Woll<strong>on</strong>g<strong>on</strong>g, NSW, Australia 2522<br />

e-mail: aquilina@uow.edu.au<br />

Here we report investigati<strong>on</strong>s into the oligomeric organizati<strong>on</strong> and dynamics <str<strong>on</strong>g>of</str<strong>on</strong>g> the Saccharomyces cerevisiae<br />

small heat-shock protein ScHSP26. Using both multi-angle light scattering and mass spectrometry (MS)<br />

approaches we have shown that at ambient temperature this protein is exists as a polydisperse assembly,<br />

comprising numerous oligomeric states. Relative quantificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these oligomeric states using a tandem MS


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

technique dem<strong>on</strong>strated the 24mer to be anomalously abundant, c<strong>on</strong>sistent with previous reports which have<br />

culminated in a 3D structure <str<strong>on</strong>g>for</str<strong>on</strong>g> this oligomeric state. Significantly, we dem<strong>on</strong>strate <str<strong>on</strong>g>for</str<strong>on</strong>g> the first time, that<br />

ScHSP26 also <str<strong>on</strong>g>for</str<strong>on</strong>g>ms larger oligomers, up to 40 subunits in size at room temperature. The oligomers observed<br />

were exclusively composed <str<strong>on</strong>g>of</str<strong>on</strong>g> an even number <str<strong>on</strong>g>of</str<strong>on</strong>g> subunits. This str<strong>on</strong>gly suggests there to be a basic dimeric<br />

‘building block’, an observati<strong>on</strong> c<strong>on</strong>sistent with the 3D structure <str<strong>on</strong>g>of</str<strong>on</strong>g> the ScHSP26 24mer, as well as studies<br />

per<str<strong>on</strong>g>for</str<strong>on</strong>g>med <strong>on</strong> sHSPs from bacteria, plants and mammals. Using an <strong>on</strong>line thermo-c<strong>on</strong>trolled device we<br />

investigated the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> heat stress <strong>on</strong> the oligomeric structure <str<strong>on</strong>g>of</str<strong>on</strong>g> ScHSP26. We found that as the<br />

temperature was increased, dissociati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the oligomers into dimers and m<strong>on</strong>omers was observed. Such<br />

dissociati<strong>on</strong> has previously been observed <str<strong>on</strong>g>for</str<strong>on</strong>g> ScHSP26 and other sHSPs. The remaining ScHSP26 existed in<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g>ms 32 subunits or larger, with the majority having assembled into a 40mer. There is a possibility that these<br />

larger <str<strong>on</strong>g>for</str<strong>on</strong>g>ms are unusually resistant to thermal stress, or ScHSP26 preferentially associates into these <str<strong>on</strong>g>for</str<strong>on</strong>g>ms at<br />

elevated temperatures.<br />

1A_07_P<br />

(poster secti<strong>on</strong> A1, poster board #10, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SMALL HSPS FORM INTRACELLULAR GRANULES WITH AKT/PKB AND P38<br />

DISTINCT FROM TIA/TIAR-CONTAİNİNG STRESS GRANULES<br />

Anastassiia Vertii, Alexey Kotlyarov, Matthias Gaestel<br />

From the Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, Medical School Hannover, Hannover 30625, Germany<br />

Small heat shock proteins are rapidly phosphorylated in resp<strong>on</strong>se to cellular stress up<strong>on</strong> activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> p38<br />

kinase pathway. Phosphorylati<strong>on</strong> occurs at Ser 15 and Ser 86 in mouse Hsp25 and at Ser 15 , Ser 78 and Ser 82 in<br />

human Hsp27 resulting in changes in their oligomeric state. Stress causes accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp25/27 in an<br />

insoluble fracti<strong>on</strong> and <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> intracellular granular structures. Although stress-induced Hsp27 granules<br />

have been known <str<strong>on</strong>g>for</str<strong>on</strong>g> a l<strong>on</strong>g time, the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> other proteins in these granules remains enigmatic. Certain<br />

stress granules (SGs) c<strong>on</strong>tain mRNA-binding proteins, such as TIA/TIAR, and sequested processing bodies<br />

with mRNA and translati<strong>on</strong> initiati<strong>on</strong> factors which play a role in translati<strong>on</strong>al inhibiti<strong>on</strong>. Here we report<br />

stress-independent interacti<strong>on</strong> between Hsp25 and Akt/PKB and characterised the regi<strong>on</strong>s in Hsp25 and Akt<br />

resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> this interacti<strong>on</strong>. We further analysed stress-induced insolubilisati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> small Hsps and<br />

dem<strong>on</strong>strate stress-induced insolubilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Akt together with Hsp25. By immun<str<strong>on</strong>g>of</str<strong>on</strong>g>luorescence we show<br />

that Hsp25/27-c<strong>on</strong>tainig SGs, which are <str<strong>on</strong>g>for</str<strong>on</strong>g>med in resp<strong>on</strong>se to arsenite treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> HeLa cells, include Akt<br />

and p38. Although it is known that MK2 interacts with p38 we were not able to detect overexpressed MK2 in<br />

these SGs. Furthermore, these SGs are distinct from cytoplasmic granules that c<strong>on</strong>tain TIA/TIAR. siRNA<br />

silencing and nuclear/cytoplasmic fracti<strong>on</strong>ati<strong>on</strong> was used to further describe the role <str<strong>on</strong>g>of</str<strong>on</strong>g> these novel SGs <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

the functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Akt.<br />

25


23-26 August 2007,<br />

Budapest, Hungary<br />

26<br />

1A_08_P<br />

(poster secti<strong>on</strong> A1, poster board #11, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SMALL HEAT SHOCK PROTEINS EFFECTIVELY PROTECT MYOSIN S1 AND F-<br />

ACTIN FROM THERMALLY INDUCED AGGREGATION<br />

A. V. Pivovarova 1,3 , D. I. Markov 1,2 , N. B. Gusev 2 , D. I. Levitsky 1<br />

1A.N. Bach Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, Russian Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences, Moscow; 2 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, School<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, and 3 School <str<strong>on</strong>g>of</str<strong>on</strong>g> Bioengeneering and Bioin<str<strong>on</strong>g>for</str<strong>on</strong>g>matics, Moscow State University, Moscow, Russia<br />

We have found that small heat shock proteins (sHsp) do not affect thermal denaturati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> actin filaments<br />

(F-actin) measured by differential scanning calorimetry (DSC), but effectively prevent their heat induced<br />

aggregati<strong>on</strong>. The presence <str<strong>on</strong>g>of</str<strong>on</strong>g> sHsp significantly increases the temperature <str<strong>on</strong>g>of</str<strong>on</strong>g> F-actin aggregati<strong>on</strong>. It has been<br />

shown by co-sedimentati<strong>on</strong> experiments that sHsp do not bind to native F-actin. However, after heating to<br />

75 o C they <str<strong>on</strong>g>for</str<strong>on</strong>g>m soluble complexes with thermally denatured actin, which do not precipitate at high-speed<br />

centrifugati<strong>on</strong>, unlike F-actin in the absence <str<strong>on</strong>g>of</str<strong>on</strong>g> sHsp that fully precipitates under the same c<strong>on</strong>diti<strong>on</strong>s.<br />

Sedimentati<strong>on</strong> coefficients <str<strong>on</strong>g>of</str<strong>on</strong>g> these complexes, as estimated by analytical centrifugati<strong>on</strong>, and their size (as<br />

determined by dynamic light scattering experiments) were much less than these parameters <str<strong>on</strong>g>for</str<strong>on</strong>g> native F-actin<br />

and thermally denatured actin. The gel filtrati<strong>on</strong> was used to estimate a stoichiometry sHsp/actin in these<br />

complexes. We also studied the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> sHsp <strong>on</strong> thermal denaturati<strong>on</strong> and aggregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> isolated myosin<br />

head (myosin subfragment 1, S1). It has been shown that under heat shock c<strong>on</strong>diti<strong>on</strong>s (up<strong>on</strong> incubati<strong>on</strong> at<br />

43 o C) sHsp effectively prevent aggregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> S1 induced by its thermal denaturati<strong>on</strong>, but these proteins have<br />

no appreciable influence <strong>on</strong> the thermal unfolding <str<strong>on</strong>g>of</str<strong>on</strong>g> S1, as studied by DSC, and <strong>on</strong> thermally induced<br />

inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> myosin ATPase. Supported by grants from RFBR and the Program “Molecular and Cellular<br />

Biology” <str<strong>on</strong>g>of</str<strong>on</strong>g> Russian Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences.<br />

1A_09_P<br />

(poster secti<strong>on</strong> A1, poster board #12, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DYNAMICAL PROPERTIES OF SHSPS<br />

Justin L. P. Benesch 1 , Alexander J. Painter 1 , J. Andrew Aquilina 2 ,<br />

John Carver 3 , Elizabeth Vierling 4 , Carol V. Robins<strong>on</strong> 1<br />

1 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Chemistry, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Cambridge, Cambridge, CB2 1EW, UK<br />

2 School <str<strong>on</strong>g>of</str<strong>on</strong>g> Biological Sciences, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Woll<strong>on</strong>g<strong>on</strong>g, Woll<strong>on</strong>g<strong>on</strong>g, NSW 2522, Australia<br />

3 School <str<strong>on</strong>g>of</str<strong>on</strong>g> Chemistry & Physics, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Adelaide, Adelaide, SA 5005, Australia<br />

4 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry & Molecular Biophysics, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Ariz<strong>on</strong>a, Tucs<strong>on</strong>, AZ, USA<br />

The functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a protein is determined by the combinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> its structural and dynamical characteristics.<br />

Am<strong>on</strong>gst molecular chaper<strong>on</strong>es the sHSPs are the least well understood, with high-resoluti<strong>on</strong> structures<br />

existing <str<strong>on</strong>g>for</str<strong>on</strong>g> <strong>on</strong>ly a small subset <str<strong>on</strong>g>of</str<strong>on</strong>g> the family. One observed characteristic <str<strong>on</strong>g>of</str<strong>on</strong>g> these proteins appears to be an<br />

inherently dynamic nature. Under stress c<strong>on</strong>diti<strong>on</strong>s these proteins have been purported to undergo structural<br />

rearrangement, thereby exposing binding surfaces necessary <str<strong>on</strong>g>for</str<strong>on</strong>g> their chaper<strong>on</strong>e functi<strong>on</strong>. Moreover, these<br />

proteins have also been shown to undergo rapid exchange <str<strong>on</strong>g>of</str<strong>on</strong>g> subunits, presumably reflecting their flexible<br />

functi<strong>on</strong> within the stress resp<strong>on</strong>se. Here we will present data obtained by applying a mass spectrometry<br />

approach to several different sHSP systems. The stoichiometries <str<strong>on</strong>g>of</str<strong>on</strong>g> sHSPs from Arabidposis thaliana<br />

(HSP17.6 and 18.1), Saccharomyces cerevisiae (HSP26), and Pisum sativum (HSP18.1) are all elucidated in<br />

this way, and the changes incurred under thermal stress are investigated. Furthermore we present an


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

automated approach towards determining the kinetics <str<strong>on</strong>g>of</str<strong>on</strong>g> subunit exchange <str<strong>on</strong>g>of</str<strong>on</strong>g> these proteins. From these<br />

studies we obtain kinetic parameters <str<strong>on</strong>g>for</str<strong>on</strong>g> the fast exchange <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP17.6 and 18.1, and HSP26. By per<str<strong>on</strong>g>for</str<strong>on</strong>g>ming<br />

these reacti<strong>on</strong>s at a range <str<strong>on</strong>g>of</str<strong>on</strong>g> temperatures we shown how the mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> exchange varies at different<br />

temperatures and how this reflects the different structural states populated by these proteins. We c<strong>on</strong>clude<br />

that these proteins are very dynamic, and are capable <str<strong>on</strong>g>of</str<strong>on</strong>g> freely exchanging subunits <strong>on</strong> a rapid time-scale.<br />

C<strong>on</strong>sidering that this process must surely occur in vivo allows us to speculate about the importance <str<strong>on</strong>g>of</str<strong>on</strong>g> this as<br />

a general characteristic regarding the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> sHSP chaper<strong>on</strong>e acti<strong>on</strong>.<br />

27


23-26 August 2007,<br />

Budapest, Hungary<br />

28<br />

1B. THE HSP70 FAMILY<br />

(COSTA GEORGOPOULOS, KEVIN A. MORANO)<br />

1B_01_S<br />

REGULATION OF SMALL HEAT SHOCK PROTEINS IN AGEING AND<br />

RESISTANCE TO STRESS<br />

R<strong>on</strong>ald S. Ullers 1 , Debbie Ang 1 , Françoise Schwager 1 , Pierre Genevaux 2 , Costa Georgopoulos 1<br />

1Département de Microbiologie et Médecine Moléculaire, <str<strong>on</strong>g>Centre</str<strong>on</strong>g> Médical Universitaire,<br />

1, Rue Michel-Servet, CH-1211 Geneva, Switzerland,<br />

2Laboratoire de Microbiologie et Génétique Moléculaires, Institut de Biologie Cellulaire et de Génétique, <str<strong>on</strong>g>Centre</str<strong>on</strong>g><br />

Nati<strong>on</strong>al de la Recherche Scientifique, Université Paul-Sabatier, 118 Route de Narb<strong>on</strong>ne,<br />

31062 Toulouse Cedex 09, France<br />

Polypeptides emerging from the ribosome are assisted by a pool <str<strong>on</strong>g>of</str<strong>on</strong>g> molecular chaper<strong>on</strong>es and targeting<br />

factors, which enable them to efficiently partiti<strong>on</strong> as cytoplasmic, integral membrane, or exported proteins.<br />

In Escherichia coli, the chaper<strong>on</strong>es SecB, Trigger Factor (TF), and DnaK are key players in this process.<br />

Here, we report that, as with dnaK or dnaJ mutants, a secB null strain exhibits a str<strong>on</strong>g cold-sensitive (Cs)<br />

phenotype. Through suppressor analyses, we found that inactivating mutati<strong>on</strong>s in the tig gene encoding TF<br />

fully relieve both the Cs phenotype and protein aggregati<strong>on</strong> observed in the absence <str<strong>on</strong>g>of</str<strong>on</strong>g> SecB. This<br />

antag<strong>on</strong>istic effect <str<strong>on</strong>g>of</str<strong>on</strong>g> TF depends <strong>on</strong> its ribosome-binding and chaper<strong>on</strong>e activities but unrelated to its<br />

peptidyl-prolyl cis/trans isomerase’s (PPIase) activity. Furthermore, in c<strong>on</strong>trast to the previously known<br />

synergistic acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> TF and the DnaK/DnaJ chaper<strong>on</strong>e machine above 30°C, a tig null mutati<strong>on</strong> partially<br />

suppresses the Cs phenotype exhibited by a compromised DnaK/DnaJ chaper<strong>on</strong>e machine. The<br />

antag<strong>on</strong>istic role <str<strong>on</strong>g>of</str<strong>on</strong>g> TF is further exemplified by the fact that the secB dnaJ double mutant is viable <strong>on</strong>ly in<br />

the absence <str<strong>on</strong>g>of</str<strong>on</strong>g> TF. Finally, we show that, in the absence <str<strong>on</strong>g>of</str<strong>on</strong>g> TF, more SecA and ribosomes are associated<br />

with the inner membrane, suggesting that the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> TF directly or indirectly somehow interferes with<br />

the process <str<strong>on</strong>g>of</str<strong>on</strong>g> cotranslati<strong>on</strong>al protein targeting to the Sec transloc<strong>on</strong>. Various models to explain our results<br />

(occasi<strong>on</strong>ally seemingly c<strong>on</strong>tradictory) will be <str<strong>on</strong>g>of</str<strong>on</strong>g>fered.<br />

1B_02_S<br />

ROLE OF HSP110 IN THE FUNCTIONAL NETWORK OF HSP70 CHAPERONES<br />

Jocelyne Fiaux, Claes Andréass<strong>on</strong>, Heike Rampelt, Heather Sadlish,<br />

Matthias Mayer, Bernd Bukau<br />

ZMBH, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Heidelberg, Im Neuenheimer Feld 282, D-69120 Heidelberg, Germany,<br />

e-mail: bukau@zmbh.uni-heidelberg.de<br />

Eukaryotic cells express Hsp110 proteins that c<strong>on</strong>stitute a diverged branch <str<strong>on</strong>g>of</str<strong>on</strong>g> the Hsp70 molecular<br />

chaper<strong>on</strong>e superfamily. Recently, both the yeast Hsp110 member Sse1p and the mammalian orthologue<br />

Hsp105 were found to act as potent nucleotide exchange factors <str<strong>on</strong>g>for</str<strong>on</strong>g> cytosolic Hsp70 chaper<strong>on</strong>es. We set out<br />

to characterize the steps <str<strong>on</strong>g>of</str<strong>on</strong>g> the nucleotide exchange cycle <str<strong>on</strong>g>of</str<strong>on</strong>g> the yeast Hsp70, Ssa1p, catalyzed by Sse1p using<br />

H/D exchange and mass spectrometry. The mechanism was found to involve <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a stable but<br />

nucleotide-sensitive complex. Furthermore, Sse1p itself displays nucleotide binding and hydrolysis and adopts<br />

c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>s dependent <strong>on</strong> the nucleotide binding status. Interestingly, nucleotide binding by Sse1p appears<br />

to regulate its activity as a nucleotide exchange factor. We also used H/D exchange and mass spectrometry to


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

map the interacti<strong>on</strong> surface <str<strong>on</strong>g>of</str<strong>on</strong>g> the Sse1p-Ssa1p complex. Based <strong>on</strong> these results, we can now propose a<br />

detailed model <str<strong>on</strong>g>for</str<strong>on</strong>g> the Sse1p-catalyzed nucleotide exchange cycle.<br />

1B_03_S<br />

NETWORKS OF HSP70 AND J-PROTEIN MOLECULAR CHAPERONES<br />

Elizabeth A. Craig, Alis<strong>on</strong> Meyer, Chandan Sahi<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Wisc<strong>on</strong>sin, Madis<strong>on</strong> WI, 53706, USA<br />

e-mail: ecraig@wisc.edu<br />

Highly c<strong>on</strong>served molecular chaper<strong>on</strong>es functi<strong>on</strong> in a wide variety <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular processes, including protein<br />

folding, translocati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins across membranes and remodeling <str<strong>on</strong>g>of</str<strong>on</strong>g> protein complexes. J-proteins are<br />

obligate partners <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70s that act via their J-domains to stimulate Hsp70’s ATPase activity, stabilizing their<br />

interacti<strong>on</strong>s with client proteins. In additi<strong>on</strong> many J-proteins c<strong>on</strong>tain additi<strong>on</strong>al domains, some <str<strong>on</strong>g>of</str<strong>on</strong>g> which are<br />

capable <str<strong>on</strong>g>of</str<strong>on</strong>g> binding client proteins and allowing J-proteins to “deliver” them to their partner Hsp70. Both<br />

Hsp70s and J-proteins are encoded by large multigene families. Our results indicate that certain Hsp70s and<br />

J-proteins have evolved to functi<strong>on</strong> in specialized cellular processes and/or have “n<strong>on</strong>traditi<strong>on</strong>al” functi<strong>on</strong>s.<br />

For example, Ssz1 <str<strong>on</strong>g>for</str<strong>on</strong>g>ms a stable heterodimer with the ribosome-associated J-protein Zuo1 and is required<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> Zuo1’s efficient stimulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the ATPase activity <str<strong>on</strong>g>of</str<strong>on</strong>g> its partner Hsp70 Ssb. Jjj1, is an example <str<strong>on</strong>g>of</str<strong>on</strong>g> a<br />

specialized J-protein. It plays an important role in the biogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> 60S ribosomal subunits, likely facilitating<br />

the dissociati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> factors from preribosomes to generate subunits competent <str<strong>on</strong>g>for</str<strong>on</strong>g> translati<strong>on</strong>. In c<strong>on</strong>trast,<br />

other J-proteins are multifuncti<strong>on</strong>al. The most abundant J-protein <str<strong>on</strong>g>of</str<strong>on</strong>g> the yeast cytosol, Ydj1, functi<strong>on</strong>s in<br />

multiple cellular processes. Surprisingly, however, expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> <strong>on</strong>ly a J-domain at normal levels is capable <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

rescuing the severe growth defect caused by the absence <str<strong>on</strong>g>of</str<strong>on</strong>g> Ydj1. Thus many functi<strong>on</strong>s carried out by this<br />

highly c<strong>on</strong>served and complex J-protein require <strong>on</strong>ly the capacity to stimulate Hsp70s ATPase activity, not<br />

the “delivery” <str<strong>on</strong>g>of</str<strong>on</strong>g> client proteins.<br />

1B_04_S<br />

HSP110 PROTEIN CHAPERONE FUNCTION IN YEAST<br />

Kevin Morano<br />

Université Laval, Departement <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Ave Medecine, G1K 7P4, Quebec, Canada<br />

SSE1 and SSE2 encode the essential yeast members <str<strong>on</strong>g>of</str<strong>on</strong>g> the Hsp70-related Hsp110 molecular chaper<strong>on</strong>e<br />

family. Both mammalian Hsp110 and the Sse proteins functi<strong>on</strong>ally interact with cognate cytosolic Hsp70s as<br />

nucleotide exchange factors, and in yeast Sse1 is required <str<strong>on</strong>g>for</str<strong>on</strong>g> Hsp90 chaper<strong>on</strong>ing. We dem<strong>on</strong>strate that Sse1<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g>ms high affinity heterodimeric complexes with both yeast Ssa and mammalian Hsp70 chaper<strong>on</strong>es, and that<br />

ATP binding to Sse1 is required <str<strong>on</strong>g>for</str<strong>on</strong>g> binding to Hsp70s. The nucleotide binding domains (NBD) <str<strong>on</strong>g>of</str<strong>on</strong>g> both<br />

Sse1/2 and the Hsp70s dictate interacti<strong>on</strong> specificity, and are sufficient to mediate heterodimerizati<strong>on</strong> with<br />

no discernable c<strong>on</strong>tributi<strong>on</strong> from the peptide binding domains (PBD). However, the PBD is required <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

NEF activity. To better understand the roles <str<strong>on</strong>g>of</str<strong>on</strong>g> the Hsp110 chaper<strong>on</strong>es, we are investigating the participati<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> Sse1 in cellular processes requiring Hsp70. To that end, we have generated a novel temperature sensitive<br />

allele <str<strong>on</strong>g>of</str<strong>on</strong>g> SSE1 that should shed light <strong>on</strong> the essential functi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> this intriguing chaper<strong>on</strong>e family.<br />

29


23-26 August 2007,<br />

Budapest, Hungary<br />

30<br />

1B_05_S<br />

(poster secti<strong>on</strong> A1, poster board #13, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EVOLUTION AND DIVERSITY OF HUMAN HSP70: IMPLICATIONS FOR HEALTH<br />

AND DISEASE<br />

Luciano Brocchieri 1 , Everly C<strong>on</strong>way de Macario 2 , Alberto J. L. Macario 2<br />

1University <str<strong>on</strong>g>of</str<strong>on</strong>g> Florida, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular Genetics and Microbiology and UF Genetics Institute,<br />

Gainesville, FL, USA<br />

2Center <str<strong>on</strong>g>of</str<strong>on</strong>g> Marine Biotechnology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Maryland Biotechnology Institute, Baltimore, MD, USA<br />

Defective chaper<strong>on</strong>es are implicated in disease, the chaper<strong>on</strong>opathies, and senescence. Defective chaper<strong>on</strong>e<br />

identificati<strong>on</strong> and elucidati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> pathogenetic role in disease and ageing requires full gene identificati<strong>on</strong> in the<br />

genome; characterizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> genes and proteins in silico, in vitro and in vivo; and clinicopathological studies.<br />

We applied a series <str<strong>on</strong>g>of</str<strong>on</strong>g> complementary bioin<str<strong>on</strong>g>for</str<strong>on</strong>g>matics and evoluti<strong>on</strong>ary methods (chaper<strong>on</strong>omics) to the study<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> human Hsp70, and identified 47 loci encoding Hsp70-like sequences: 16 were functi<strong>on</strong>al genes, encoding<br />

proteins c<strong>on</strong>served at the N-terminal but not at the C-terminal domain, <str<strong>on</strong>g>of</str<strong>on</strong>g> which eight encoded typical<br />

Hsp70s (65-80kDa) with nucleotide-binding and substrate-binding domains, two encoded products lacking<br />

most or all <str<strong>on</strong>g>of</str<strong>on</strong>g> the C-terminal domain, and six encoded heavier proteins (> 81 kDa) with atypical C-terminal<br />

domains. Also, 31 hsp70-related pseudogenes were identified. Evoluti<strong>on</strong>ary analyses showed that ER-residing<br />

Hsps evolved by duplicati<strong>on</strong> while the six typical genes whose products reside in the cytosol/nucleus, and the<br />

majority <str<strong>on</strong>g>of</str<strong>on</strong>g> the pseudogenes, originated from retrotranspositi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> <strong>on</strong>e gene, HSPA8. These evoluti<strong>on</strong>ary<br />

processes generated a group <str<strong>on</strong>g>of</str<strong>on</strong>g> genes with wide diversity further amplified by the occurrence <str<strong>on</strong>g>of</str<strong>on</strong>g> multiple<br />

mRNA variants and protein is<str<strong>on</strong>g>of</str<strong>on</strong>g>orms. The variety <str<strong>on</strong>g>of</str<strong>on</strong>g> the Hsp70 proteins is reflected in the diversity <str<strong>on</strong>g>of</str<strong>on</strong>g> their<br />

patterns <str<strong>on</strong>g>of</str<strong>on</strong>g> expressi<strong>on</strong> and localizati<strong>on</strong> in an assortment <str<strong>on</strong>g>of</str<strong>on</strong>g> tissues, cell types and sub-cellular compartments,<br />

through the stages <str<strong>on</strong>g>of</str<strong>on</strong>g> development and ageing. C<strong>on</strong>sequently, the pathogenetic impact <str<strong>on</strong>g>of</str<strong>on</strong>g> defective Hsp70s<br />

should be widespread.<br />

1B_06_S<br />

(poster secti<strong>on</strong> A1, poster board #14, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HEAT SHOCK COGNATE PROTEIN HSC70 AS A REGULATOR OF THE<br />

ACTIVIN/NODAL/TGF-ß-SMAD2 SIGNALLING PATHWAY DURING ZEBRAFISH<br />

DEVELOPMENT<br />

Michiaki Yamashita, Misako Hojo, Takeshi Yabu<br />

Nati<strong>on</strong>al <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Fisheries Science, Fukuura, Yokohama 236-8648, Japan<br />

e-mail: mic@affrc.go.jp<br />

The cytosolic heat shock protein 70 (HSP70) and heat shock cognate protein 70 (HSC70) are thought to<br />

combine to functi<strong>on</strong> as a molecular chaper<strong>on</strong>e. The protein binds transiently to nascent polypeptides and<br />

unfolded proteins, and prevents intramolecular and intermolecular interacti<strong>on</strong>s, which can result in<br />

misfolding or aggregati<strong>on</strong>. Precise c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> Nodal proteins, which are members <str<strong>on</strong>g>of</str<strong>on</strong>g> the trans<str<strong>on</strong>g>for</str<strong>on</strong>g>ming growth<br />

factor-ß (TGF-ß) superfamily, has been identified as a key endogenous mesoderm inducer in vertebrates. In<br />

the present study, we report that the HSC70 molecular chaper<strong>on</strong>e promotes the mesoderm inducti<strong>on</strong><br />

activities <str<strong>on</strong>g>of</str<strong>on</strong>g> Nodal signalling. To study the biological functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSC70, the endogenous expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

HSC70 was knocked down by the injecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a specific antisense morpholino olig<strong>on</strong>ucleotide (HSC-MO).<br />

The HSC-MO-injected embryos showed reduced phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Smad2 by Nodal signalling, and this


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

phenotype was rescued by co-injecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the HSC70 protein. However, the HSP70 mutant that lacked the<br />

C-terminal tetrapeptide (EEVD) motif showed no activity up<strong>on</strong> inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Nodal signalling. The Activin<br />

type IIB receptor (ActRIIB) was immunoprecipitated with HSC70, and its autophosphorylati<strong>on</strong> was<br />

enhanced in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> HSC70, which suggests that HSC70 binds directly to the receptor and modulates<br />

its activity. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, HSC70 plays essential roles in the <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> and activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the type IIB receptor<br />

up<strong>on</strong> Nodal signalling.<br />

31


23-26 August 2007,<br />

Budapest, Hungary<br />

32<br />

1B_01_P<br />

(poster secti<strong>on</strong> A1, poster board #15, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE PAM18/TIM14-PAM16/TIM16 COMPLEX OF THE MITOCHONDRIAL<br />

TRANSLOCATION MOTOR: THE FORMATION OF A STABLE COMPLEX FROM<br />

MARGINALLY STABLE PROTEINS<br />

Ohad Iosefs<strong>on</strong><br />

Tel Aviv University, Biochemistry, 69978, Tel Aviv, POB 39040, Israel<br />

The majority <str<strong>on</strong>g>of</str<strong>on</strong>g> the mitoch<strong>on</strong>drial proteins encoded in the nucleus, synthesized in the cytosol and<br />

transported into the mitoch<strong>on</strong>dria. The presequence translocase associated import motor is required in order<br />

to import matrix-targeted proteins across the mitoch<strong>on</strong>drial inner membrane. This transport machinery is<br />

composed <str<strong>on</strong>g>of</str<strong>on</strong>g> the following c<strong>on</strong>stituents: mitoch<strong>on</strong>drial Hsp70, the nucleotide exchange c<str<strong>on</strong>g>of</str<strong>on</strong>g>actor- GrpE,<br />

Tim44, Tim14 and Tim16. All <str<strong>on</strong>g>of</str<strong>on</strong>g> these proteins are essential to the eukaryotic cell viability. However, in order<br />

to shed light <strong>on</strong> the accurate cooperati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> this intriguing motor, in-vitro analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> the purified<br />

comp<strong>on</strong>ents needs to be d<strong>on</strong>e. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, our work is focused <strong>on</strong> in-vitro characterizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the structurefuncti<strong>on</strong><br />

relati<strong>on</strong>ship <str<strong>on</strong>g>of</str<strong>on</strong>g> the two motor c<strong>on</strong>stituents: Tim14 and Tim16. We characterized the interacti<strong>on</strong><br />

between these proteins using two different methods: fluorescence polarizati<strong>on</strong> and heat thermal denaturati<strong>on</strong>.<br />

In order to examine the stability <str<strong>on</strong>g>of</str<strong>on</strong>g> the Tim14-Tim16 complex by plotting their heat thermal denaturati<strong>on</strong><br />

curve Circular dichroism spectrophotometer was used. In additi<strong>on</strong>, by using fluorescence polarizati<strong>on</strong><br />

approach, we measured the affinity <str<strong>on</strong>g>of</str<strong>on</strong>g> these two proteins and determined the dissociati<strong>on</strong> c<strong>on</strong>stant (K d ) <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

their complex.<br />

1B_02_P<br />

(poster secti<strong>on</strong> A1, poster board #16, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HSP70 IS DIFFERENTIALLY EXPRESSED IN CATTLE AND SHEEP LEUKOCYTES<br />

Linda L. Agnew, Ian G. Colditz<br />

CSIRO Livestock Industries, Armidale, NSW, Australia<br />

Flow cytometry is a rapid and quantitative method <str<strong>on</strong>g>of</str<strong>on</strong>g> determining intracellular heat shock protein (hsp) 70<br />

expressi<strong>on</strong> in individual cells from a heterogeneous populati<strong>on</strong> such as peripheral blood m<strong>on</strong><strong>on</strong>uclear cells.<br />

Hsp70 expressi<strong>on</strong> is induced in vivo in a range <str<strong>on</strong>g>of</str<strong>on</strong>g> leukocyte types in resp<strong>on</strong>se to a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> stressors. The<br />

applicati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a mild, transient, n<strong>on</strong>-lethal, in vitro heat shock is a technique that allows the researcher to<br />

examine the cellular stress resp<strong>on</strong>se which is a highly c<strong>on</strong>served defence mechanism. Thus c<strong>on</strong>stitutive and in<br />

vitro induced leukocyte hsp70 expressi<strong>on</strong> may be a useful indicator <str<strong>on</strong>g>of</str<strong>on</strong>g> an organism’s adaptati<strong>on</strong> to<br />

envir<strong>on</strong>mental/physiological stress. In the current studies, flow cytometric methods were used to<br />

dem<strong>on</strong>strate that hsp70 is c<strong>on</strong>stitutively expressed in ovine and bovine leukocytes but that the level <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

expressi<strong>on</strong> varies c<strong>on</strong>siderably between different leukocyte types and between species. The optimum<br />

temperature <str<strong>on</strong>g>for</str<strong>on</strong>g> heat shock <str<strong>on</strong>g>of</str<strong>on</strong>g> leukocytes from sheep and cattle without a loss <str<strong>on</strong>g>of</str<strong>on</strong>g> cell viability is 43.5°C. In<br />

sheep, the magnitude <str<strong>on</strong>g>of</str<strong>on</strong>g> upregulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp70 expressi<strong>on</strong> by in vitro heat shock was CD14+ cells > γδ T<br />

cells > neutrophils > B cells > CD4 = CD8. A similar ranking was observed in cattle. Best results were<br />

obtained from fresh samples; after storage at room temperature <str<strong>on</strong>g>for</str<strong>on</strong>g> 24 hours upregulati<strong>on</strong> was highly variable<br />

between animals and less than in fresh samples. These studies dem<strong>on</strong>strate that evaluati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> leukocyte hep70<br />

expressi<strong>on</strong> by flow cytometry is a robust, reproducible method <str<strong>on</strong>g>for</str<strong>on</strong>g> use in the evaluati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> stress resp<strong>on</strong>ses in<br />

livestock species.


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

1B_03_P<br />

(poster secti<strong>on</strong> A1, poster board #17, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

LEPTIN AND CERULENIN DIFFERENTLY REGULATE HEAT-SHOCK PROTEIN-<br />

70 (HSP-70) GENE EXPRESSION IN CHICKEN TISSUES<br />

Denise Figueiredo, Arieh Gertler, Eddy Decuypere, Johan Buyse, Gérard Cabello, Sami Dridi *<br />

Katholieke Universiteit Leuven, MeBioS, 3001 Kasteelpark Arenberg B-30, Heverlee, Belgium<br />

Lines <str<strong>on</strong>g>of</str<strong>on</strong>g> evidence suggested that systems involved in the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress resp<strong>on</strong>ses and <str<strong>on</strong>g>of</str<strong>on</strong>g> energy<br />

homeostasis are highly integrated. Since leptin and cerulenin (the natural fatty acid synthesis inhibitor) have<br />

been shown to affect food intake and energy homeostasis, and since the stress biormarker Hsp-70 gene was<br />

found to interact directly with fatty acids, we hypothesized that leptin or cerulenin may regulate Hsp-70 gene<br />

expressi<strong>on</strong>. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, the present study was undertaken to examine this issue. Leptin and Cerulenin<br />

significantly (P


23-26 August 2007,<br />

Budapest, Hungary<br />

inducible Hsp70 were chosen <strong>on</strong> the base <str<strong>on</strong>g>of</str<strong>on</strong>g> in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> structure analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> the protein sequence [1].<br />

Influence <str<strong>on</strong>g>of</str<strong>on</strong>g> recombinant Hsp70 and the peptides <strong>on</strong> IFN-γ producti<strong>on</strong> and marker surface expressi<strong>on</strong> was<br />

estimated up<strong>on</strong> 24 h incubati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the cells with IL-2. Hsp70 as well as peptides 399-408 and 411-424 costimulated<br />

IFN-γ producti<strong>on</strong> in IL-2-activated NK cells suggesting that these peptides may c<strong>on</strong>tain Hsp70<br />

sites resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> the interacti<strong>on</strong> with NK cells. The same peptides modulated NK cell surface antigen<br />

expressi<strong>on</strong> in Hsp70-like manner. CD3 - CD16 - CD56 + cell populati<strong>on</strong> was more susceptible to Hsp70 acti<strong>on</strong><br />

comparing with CD3 - CD16 + CD56 + populati<strong>on</strong>. The Hsp70 effects in the experimental system did not<br />

depend <strong>on</strong> LPS. In sum the results testify that exogenous Hsp70 affects directly human NK cells, leading to<br />

the strengthening <str<strong>on</strong>g>of</str<strong>on</strong>g> IFN-γ producti<strong>on</strong> and modulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> antigen surface expressi<strong>on</strong> in c<strong>on</strong>diti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> cell<br />

stimulati<strong>on</strong> by activating cytokines. 1. Nekrasov A. N., Entropy <str<strong>on</strong>g>of</str<strong>on</strong>g> protein sequences: an integral approach, J<br />

Biomol Struct Dyn 20: 87-92, 2002.<br />

1B_05_P<br />

(poster secti<strong>on</strong> A1, poster board #19, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PRIMARY CULTURES OF BOVINE INTERVERTEBRAL DISC CELLS HAVE A<br />

REDUCED RESPONSE TO THERMAL STRESS<br />

S. J. Owen, S. Roberts, C. A. Sharp<br />

RJAH Orthopaedic Hospital, Gobowen, UK, e-mail: shar<strong>on</strong>.owen@rjah.nhs.uk<br />

Intervertebral discs (IVD) are avascular tissues that separate the b<strong>on</strong>y vertebral bodies and act as shock<br />

absorbers <str<strong>on</strong>g>for</str<strong>on</strong>g> the spinal column. At the centre <str<strong>on</strong>g>of</str<strong>on</strong>g> each disc is the nucleus pulposus (NP), a regi<strong>on</strong> rich in<br />

proteoglycan and type II collagen produced and maintained by ch<strong>on</strong>drocyte-like NP cells. In vivo, NP cells<br />

have adapted to a low oxygen envir<strong>on</strong>ment and c<strong>on</strong>stant mechanical loading. We have studied the resp<strong>on</strong>se<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> NP cells to thermal stress. NP cells were isolated from bovine coccygeal IVDs. Primary cells were cultured<br />

up to passage 4 (P4). Type I and II collagen (Col I/II), aggrecan and GAPDH gene expressi<strong>on</strong> were<br />

determined by PCR at each passage. Cells were cultured <str<strong>on</strong>g>for</str<strong>on</strong>g> 3 days following isolati<strong>on</strong> or passage (P0, 2 and<br />

4) then heat shocked at 45 o C <str<strong>on</strong>g>for</str<strong>on</strong>g> 1h and allowed to recover at 37 o C <str<strong>on</strong>g>for</str<strong>on</strong>g> 6, 24 and 48h. Total cell number at<br />

each time point was determined using propidium iodide uptake after fixati<strong>on</strong>. At P2 Col I expressi<strong>on</strong> was upregulated<br />

and by P3 Col II expressi<strong>on</strong> was down-regulated whereas aggrecan expressi<strong>on</strong> remained unchanged.<br />

Primary NP cell HSP70 producti<strong>on</strong> was maximal (0.5ng/1000 cells) after 24h recovery whereas those at P2<br />

and P4 were maximally elevated (2.8 and 1.7ng/1000 cells respectively) after 6h. Total cell number at P0, 2<br />

and 4 were <strong>on</strong>ly marginally affected by heat shock (110, 94 and 102% respectively) compared with c<strong>on</strong>trols.<br />

Viability was unaffected. Freshly isolated bovine NP cells have an attenuated resp<strong>on</strong>se to thermal stress and<br />

dedifferentiate in culture losing their ch<strong>on</strong>drocyte-like phenotype. NP cell dedifferentiati<strong>on</strong> is accompanied<br />

by increased HSP70 producti<strong>on</strong> after heat shock. In vivo, NP cell phenotype reflects their tissue envir<strong>on</strong>ment<br />

which is modified by cell culture.<br />

34


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

1B_06_P<br />

(poster secti<strong>on</strong> A1, poster board #20, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HEAT SHOCK PROTEINS IN PATHOLOGICAL HUMAN INTERVERTEBRAL DISCS<br />

C. A. Sharp, H. Evans, S. Roberts, S. J. Owen<br />

RJAH Orthopaedic Hospital, Gobowen, UK, e-mail: chris.sharp@rjah.nhs.uk<br />

Back pain is associated with intervertebral disc (IVD) degenerati<strong>on</strong>, resulting from the failure <str<strong>on</strong>g>of</str<strong>on</strong>g> disc cells to<br />

maintain a functi<strong>on</strong>al tissue matrix. IVDs are avascular and exchange <str<strong>on</strong>g>of</str<strong>on</strong>g> nutrients & waste product is slow.<br />

This envir<strong>on</strong>ment may be detrimental to disc cell functi<strong>on</strong> & survival. Disc degenerati<strong>on</strong> begins with loss <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

proteoglycans & fluid, resulting in loss <str<strong>on</strong>g>of</str<strong>on</strong>g> disc height & functi<strong>on</strong>al impairment. Disc herniati<strong>on</strong> begins with<br />

the protrusi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the nucleus pulposus (NP) into the surrounding annulus fibrosus (AF). If the AF ruptures<br />

the NP can extrude into the spinal canal & become separated from the disc. Scoliosis is a spinal de<str<strong>on</strong>g>for</str<strong>on</strong>g>mity<br />

with lateral curvature <str<strong>on</strong>g>of</str<strong>on</strong>g> the spine. Hsp27&72 have been identified in “normal” post-mortem (PM) human<br />

IVDs & Hsp72 appeared to increase with degenerati<strong>on</strong>. Little is known <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsps in discs from patients with<br />

disc pathologies. We have studied HSF1, Hsp27&72 in IVDs from 33 patients with herniated (n=19, mean<br />

age 43y) & degenerate (14, 45y) discs, 5 with scoliosis (15y) removed at surgery & 5 c<strong>on</strong>trol discs (55y)<br />

removed at PM. IHC was per<str<strong>on</strong>g>for</str<strong>on</strong>g>med using specific mAbs to Hsp27&72 & a pAb to HSF1 & labeled with<br />

peroxidase and DAB. On each secti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> disc 200 cells were counted. Data are presented as the mean % <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

+vely stained cells. More cells were +ve <str<strong>on</strong>g>for</str<strong>on</strong>g> Hsp27 & 72 in herniated discs (49% & 38%) than in degenerate<br />

(37%&26%), scoliotic (26%&21%) or c<strong>on</strong>trols (33%&30%). Hsp27+ve cells were greater in herniated than<br />

scoliotic discs (p=0.02). HSF1 staining was greater in c<strong>on</strong>trols (24%) than in herniated (16%), degenerate<br />

(6%) or scoliotic (6%) discs. There were no trends with age <str<strong>on</strong>g>for</str<strong>on</strong>g> any <str<strong>on</strong>g>of</str<strong>on</strong>g> the antigens. HSF1, Hsp27&72 are<br />

detectable in IVD tissue. Overall Hsp levels were greatest in herniated and lowest in scoliotic discs that may<br />

imply less capacity to mount a stress resp<strong>on</strong>se.<br />

1B_07_P<br />

(poster secti<strong>on</strong> A1, poster board #21, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

POLYAMINE COMPOUND DEOXYSPERGUALIN INHIBITS HEAT SHOCK<br />

PROTEIN-INDUCED ACTIVATION OF IMMATURE DENDRITIC CELLS<br />

Atsushi Sugawara<br />

Sapporo Medical University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Pathology, 060-8556 South 1 West 17 Chuo-ku, Sapporo, Japan<br />

(OBJECTIVES) Deoxyspergualin (DSG),a spermidinyl, - hydrocyglycyl, 7-guanidinoheptanoyl<br />

peptidomimetic, is a potent immunosuppressive agent. <str<strong>on</strong>g>for</str<strong>on</strong>g> rejecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> organ transplant., whose mechanism<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> acti<strong>on</strong> remains unknown. Dendritic cells (DC) are c<strong>on</strong>sidered the most potent antigen-presenting cells<br />

(APC) and they can stimulate CD4and CD8T cells. HSP 70 bind to DC and induce DC maturati<strong>on</strong> and<br />

activated DC to elicit immune resp<strong>on</strong>ce. It has been reported that DSG and DSG analogs bind to Hsp70 and<br />

Hsp90. We report here DSG inhibit Hsp70 binding to DC by flow cytometry analysis and prevent Hsp70<br />

induced DC activati<strong>on</strong>.by TNF-alpha releasing assey.These data indicate that interacti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> DSG and Hsp70<br />

involve in immunosupresive ability <str<strong>on</strong>g>of</str<strong>on</strong>g> DSG. (MARERIALS and METHODS) B<strong>on</strong>e marrow-derived<br />

immature DCs were generated from the femurs and tibia <str<strong>on</strong>g>of</str<strong>on</strong>g> C57BL/6 mice,which were incubated in<br />

complete RPMI-1640 with 10%heatinactivated FCS and 20 ng/ml GM-CSF . Flow cytometry analysis was<br />

per<str<strong>on</strong>g>for</str<strong>on</strong>g>med to comfirm Hsp70 binding to DCs. Immature DCs were incubated with Alexa 488-labeled Hsp70<br />

at 4°C <str<strong>on</strong>g>for</str<strong>on</strong>g> 15 minutes with DSG, DSGanalog(which does not have immunosuppressive activity). To evaluate<br />

35


23-26 August 2007,<br />

Budapest, Hungary<br />

the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> immunosuppresi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> DSG, tumor necrosis factor α (TNF-α) release assay was d<strong>on</strong>e.Immature<br />

DCs <str<strong>on</strong>g>of</str<strong>on</strong>g> mice were incubated <str<strong>on</strong>g>for</str<strong>on</strong>g> 12hours with Hsp70 al<strong>on</strong>e, with DSG or DSG analog. (Result) Binding <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Hsp70 to DCs decreased under DSG presence.Hsp70 induced TNF-α release <str<strong>on</strong>g>of</str<strong>on</strong>g> DCs was suppresed by<br />

DSG. (C<strong>on</strong>clusi<strong>on</strong>) These results supposed that Hsp70 and Hsp70 family was involved in DSG<br />

immunosuppresive functi<strong>on</strong>.<br />

1B_08_P<br />

(poster secti<strong>on</strong> A1, poster board #22, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

RECOMBINANT EXPRESSION, PURIFICATION AND CHARACTERISATION OF<br />

THE COMPLEX BETWEEN HSP70 PROTEINS AND THEIR HSP40 CO-<br />

CHAPERONE PARTNERS<br />

A. Vydyanath, S. Smerd<strong>on</strong>, M. Odell<br />

School <str<strong>on</strong>g>of</str<strong>on</strong>g> Biosciences, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Westminster, L<strong>on</strong>d<strong>on</strong>, W1W 6UW<br />

e-mail: anu.keshav@gmail.com<br />

Nascent polypeptides emerging from the ribosome are exposed to a complex medium in the cytoplasm<br />

crowded with macromolecules such as cellular proteins and RNA. The <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> unwanted interacti<strong>on</strong>s<br />

can pose a threat to the proper folding <str<strong>on</strong>g>of</str<strong>on</strong>g> the nascent chain and can lead to aggregati<strong>on</strong>. Assistance to<br />

prevent this process comes in the <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> molecular chaper<strong>on</strong>es, Heat shock proteins. The 70kDa members<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the heat shock protein family (eg. Hsp70) functi<strong>on</strong> as molecular chaper<strong>on</strong>es by binding to exposed<br />

hydrophobic patches <strong>on</strong> nascent polypeptides <str<strong>on</strong>g>for</str<strong>on</strong>g>ming n<strong>on</strong>-covalent interacti<strong>on</strong>s, thereby preventing their<br />

aggregati<strong>on</strong> and facilitating their proper folding. The folding reacti<strong>on</strong> comprises <str<strong>on</strong>g>of</str<strong>on</strong>g> cyclic binding and release<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the unfolded substrate powered by ATP hydrolysis. Hsp70 requires the assistance <str<strong>on</strong>g>of</str<strong>on</strong>g> a co-chaper<strong>on</strong>e,<br />

generally provided by the Hsp40 group <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins, <str<strong>on</strong>g>for</str<strong>on</strong>g> the cycle <str<strong>on</strong>g>of</str<strong>on</strong>g> protein folding. Biochemical analyses<br />

have mapped the possible sites <str<strong>on</strong>g>of</str<strong>on</strong>g> interacti<strong>on</strong> between the Hsp70 and Hsp40 proteins and predicted a<br />

bipartite mode <str<strong>on</strong>g>of</str<strong>on</strong>g> interacti<strong>on</strong> am<strong>on</strong>gst the comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g> the chaper<strong>on</strong>e cycle, Hsp70, Hsp40 and the<br />

substrate. However, structural investigati<strong>on</strong>s into the mechanistic features <str<strong>on</strong>g>of</str<strong>on</strong>g> the folding cycle have been<br />

hampered by the transient nature <str<strong>on</strong>g>of</str<strong>on</strong>g> interacti<strong>on</strong>. We have devised a cl<strong>on</strong>ing strategy <str<strong>on</strong>g>for</str<strong>on</strong>g> the expressi<strong>on</strong> and<br />

purificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a recombinant human Hsp70/Hsp40 complex from E.coli .The complex was purified using<br />

Ni-NTA affinity chromatography and gel filtrati<strong>on</strong> column. The gel filtrati<strong>on</strong> eluti<strong>on</strong> pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ile matched neither<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the individual Hsp70 or Hsp40 comp<strong>on</strong>ents. We are analysing the stability and behaviour <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

recombinant complex with both nucleotides and substrates. The recombinant complex will be used to<br />

understand the mechanistic differences in the interacti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> the Hsp70 with Hsp40 and Hsp70 with<br />

substrate during the stages <str<strong>on</strong>g>of</str<strong>on</strong>g> the folding cycle. We will present our human Hsp70/Hsp40 complex cl<strong>on</strong>ing,<br />

expressi<strong>on</strong> and purificati<strong>on</strong> strategy and its preliminary biochemical characterisati<strong>on</strong>.<br />

36


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

1B_09_P<br />

(poster secti<strong>on</strong> A1, poster board #23, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SURFACE HSP70 EXPRESSION AND APOPTOSIS IN NORMAL AND MALIGNANT<br />

LEUCOCYTE POPULATIONS<br />

Francesca Le<strong>on</strong>i, Nina C. Dempsey, Claire, Hunter-Lavin, Christine Hoyle, John H. H. Williams<br />

Chester <str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g> Stress <str<strong>on</strong>g>Research</str<strong>on</strong>g>, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biological Science, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Chester<br />

Parkgate Road, Chester, United Kingdom, CH1 4BJ, e-mail: f.le<strong>on</strong>i@chester.ac.uk<br />

Hsp70 is normally an intracellular protein, however there is an increasing amount <str<strong>on</strong>g>of</str<strong>on</strong>g> data dem<strong>on</strong>strating their<br />

release from cells and interacti<strong>on</strong> with membrane lipids. Surface Hsp70 expressi<strong>on</strong> has been shown to be<br />

restricted to tumour cells and has an established role as a target <str<strong>on</strong>g>for</str<strong>on</strong>g> tumour immunity. We have initially<br />

examined surface expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 in leukemic patients finding an increased expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> surface Hsp70<br />

in malignant cells as expected. Jurkat cells do not express large levels <str<strong>on</strong>g>of</str<strong>on</strong>g> surface Hsp70 at normal growing<br />

temperatures. However, we have shown an increase <str<strong>on</strong>g>of</str<strong>on</strong>g> surface Hsp70 expressi<strong>on</strong> at 42°C but not at 45°C.<br />

Pro caspase-2 and the effector caspase-3 were both activated after heat shock, with a peak <str<strong>on</strong>g>of</str<strong>on</strong>g> caspase-3 after<br />

2.5 hours and a decrease in viability and necrosis after l<strong>on</strong>ger time periods and higher temperatures. The time<br />

course <str<strong>on</strong>g>of</str<strong>on</strong>g> the appearance <str<strong>on</strong>g>of</str<strong>on</strong>g> surface Hsp70 correlated with phosphatidilserine (PS) externalisati<strong>on</strong> (detected<br />

using annexin V) after 42°C heat shock. This associati<strong>on</strong> is in accordance with previous studies showing the<br />

localisati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 <strong>on</strong> the cell surface within the lipid raft structures which are partly comprised <str<strong>on</strong>g>of</str<strong>on</strong>g> PS. We<br />

have also found surface expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 in neutrophils and m<strong>on</strong>ocytes from normal lysed whole blood,<br />

and observed an increase in Hsp70 surface expressi<strong>on</strong> in these populati<strong>on</strong>s after heat shock at 42°C but not<br />

at 45°C. Again these data were associated with the increased externalisati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> phosphatidilserine (PS) <strong>on</strong> the<br />

outside <str<strong>on</strong>g>of</str<strong>on</strong>g> the cells, detected with Annexin-V. We will present further data <strong>on</strong> the relati<strong>on</strong>ship between<br />

inserti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 into the plasma membrane and the secreti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 from cells.<br />

1B_10_P<br />

(poster secti<strong>on</strong> A1, poster board #24, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SERUM INDUCIBLE HSP70 LEVELS REFLECT DISEASE SEVERITY AND DEGREE<br />

OF TISSUE DAMAGE IN SUBJECTS WITH PATHOLOGICAL PREGNANCIES AND<br />

CHRONIC HEART FAILURE<br />

Z. Prohászka 1 , T. Gombos 1 , K. Madách 2 , J. Rigó Jr. 3 , A. Molvarec 4<br />

1: III rd Dept. Int. Med. and Szentágothai Knowledge Center, Semmelweis University<br />

2: Dept. Anesth. and Intensive Ther., Semmelweis University<br />

3: Dept. Obstet. and Gynecol., Kútvölgyi Clinical Center, Semmelweis University<br />

4: I st Dept. Obstet. and Gynecol, Semmelweis University, Budapest, Hungary<br />

Background: Increasing body <str<strong>on</strong>g>of</str<strong>on</strong>g> evidence suggests that Hsp70 may be released from viable cells within<br />

exosomes, or from necrotized cells. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> our cross-secti<strong>on</strong>al studies was to identify the clinical<br />

associati<strong>on</strong>s and biological correlates <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 serum levels in healthy subjects, in females with normal and<br />

pathological pregnancies and in patients with chr<strong>on</strong>ic heart failure. The influence <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70-2 genotypes <strong>on</strong><br />

Hsp70 levels and <strong>on</strong> its associati<strong>on</strong>s to clinical data was also investigated. Methods and Results: Serum<br />

c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the inducible Hsp72 was measured with a sandwich enzyme-linked immunosorbent assay<br />

and the Hsp70-2 A(1267)G SNP was genotyped using PCR-RFLP. In healthy pregnant women Hsp72 levels<br />

were significantly lower as compared to n<strong>on</strong>-pregnant women, and there was a negative correlati<strong>on</strong> between<br />

37


23-26 August 2007,<br />

Budapest, Hungary<br />

maternal age and serum Hsp72 c<strong>on</strong>centrati<strong>on</strong> (Spearman R=-0.35; p


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

1C. HSF ACTIVATION<br />

(LEA SISTONEN)<br />

1C_01_S<br />

MULTI-SITE POST-TRANSLATIONAL MODIFICATIONS AND FUNCTIONAL<br />

INTERPLAY OF HSF1 AND HSF2<br />

Lea Sist<strong>on</strong>en<br />

Turku <str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g> Biotechnology and Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, Åbo Akademi University, BioCity, Tykistökatu 6,<br />

20520 Turku, Finland, e-mail: lea.sist<strong>on</strong>en@btk.fi<br />

Heat shock factors, HSFs, are specific transcripti<strong>on</strong>al regulators <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock genes encoding heat shock<br />

proteins, Hsps, that functi<strong>on</strong> as molecular chaper<strong>on</strong>es in protecting cells against proteotoxic stress. The<br />

activity <str<strong>on</strong>g>of</str<strong>on</strong>g> HSFs is under stringent, mainly post-translati<strong>on</strong>al c<strong>on</strong>trol, e.g. acetylati<strong>on</strong>, phosphorylati<strong>on</strong>,<br />

sumoylati<strong>on</strong>, and ubiquitylati<strong>on</strong>. Am<strong>on</strong>g the functi<strong>on</strong>al domains <str<strong>on</strong>g>of</str<strong>on</strong>g> HSFs, the amino-terminal helix-turn-helix<br />

DNA-binding domain is the most c<strong>on</strong>served and it also designates membership to the HSF family. The<br />

activati<strong>on</strong>-induced trimeric assembly <str<strong>on</strong>g>of</str<strong>on</strong>g> HSFs is mediated by hydrophobic heptad repeats and is unusual, as<br />

proteins c<strong>on</strong>taining leucine zippers <str<strong>on</strong>g>of</str<strong>on</strong>g>ten <str<strong>on</strong>g>for</str<strong>on</strong>g>m dimers. Four HSFs have been identified in vertebrates, HSF1<br />

and HSF2 being ubiquitously expressed and well c<strong>on</strong>served throughout evoluti<strong>on</strong>, whereas HSF3 has been<br />

found <strong>on</strong>ly in avian species and HSF4 <strong>on</strong>ly in mammals. The different members <str<strong>on</strong>g>of</str<strong>on</strong>g> the mammalian HSF<br />

family have been c<strong>on</strong>sidered to be functi<strong>on</strong>ally distinct; HSF1 is essential <str<strong>on</strong>g>for</str<strong>on</strong>g> the heat shock resp<strong>on</strong>se,<br />

whereas HSF2 and HSF4 are refractory to stress stimuli but are important <str<strong>on</strong>g>for</str<strong>on</strong>g> differentiati<strong>on</strong> and<br />

development, including corticogenesis, spermatogenesis, and maintenance <str<strong>on</strong>g>of</str<strong>on</strong>g> sensory organs, e.g. lens and<br />

olfactory epithelium. We have, however, evidence <str<strong>on</strong>g>for</str<strong>on</strong>g> a functi<strong>on</strong>al interplay between HSF1 and HSF2. These<br />

factors can interact through their trimerizati<strong>on</strong> domains, and in resp<strong>on</strong>se to stress, they can bind to hsp<br />

promoters as well as to satellite III repeats at locus 9q12. In both cases, an intact HSF1 is required, suggesting<br />

that HSF1 influences the DNA-binding activity <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF2. At the transcripti<strong>on</strong>al level, HSF2 is able to<br />

modulate HSF1-mediated expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the target genes in a gene-specific manner.<br />

1C_02_S<br />

HSF1 ACTIVATION AND THE CONTROL OF APOPTOSIS IN CHEMORESISTANT<br />

CANCERS<br />

G. Belardo, A. Rossi, S. Ciafre’, A. Ciucci, P. Gianferretti, S. Roberts, M. G. Santoro<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Rome Tor Vergata and Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Neurobiology and Molecular Medicine,<br />

CNR, Rome, Italy; Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Chemistry, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Manchester, Manchester, UK<br />

Activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the heat shock resp<strong>on</strong>se (HSR) via HSF1 c<strong>on</strong>tributes to preserve cellular functi<strong>on</strong> and<br />

homeostasis under stress c<strong>on</strong>diti<strong>on</strong>s, and to establish a cytoprotective state in several human diseases. In<br />

cancer, however, HSR activati<strong>on</strong> has been associated with both anti- and pro-apoptotic resp<strong>on</strong>ses. Heatinduced<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cytoprotective and antiapoptotic heat shock proteins (HSP) is a known complicati<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> hyperthermia, resulting in cancer cell thermotolerance and chemoresistance. In some instances, however,<br />

HSF1 activati<strong>on</strong> may result in apoptosis inducti<strong>on</strong>. We have developed a library <str<strong>on</strong>g>of</str<strong>on</strong>g> novel potent inducers <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

HSF1 which are characterized by pro-apoptotic activity in several types <str<strong>on</strong>g>of</str<strong>on</strong>g> chemoresistant cancers. We have<br />

previously shown that HSF1 inducti<strong>on</strong> prevents TNFα - and mitogen-induced activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> NF-κB, a nuclear<br />

factor playing an important role in promoting inflammati<strong>on</strong>, as well as cell proliferati<strong>on</strong> and survival. NF-κB<br />

39


23-26 August 2007,<br />

Budapest, Hungary<br />

has been found to be c<strong>on</strong>stitutively activated in several types <str<strong>on</strong>g>of</str<strong>on</strong>g> chemoresistant cancers where it suppresses<br />

cell death pathways by switching <strong>on</strong> genes that dampen pro-apoptotic signals. We now show that activati<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> HSF1 by diverse chemical inducers, as well as by hyperthermia itself, results in inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> c<strong>on</strong>stitutive<br />

NF-κB activity and rapid down-regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> NF-κB-dependent survival genes, triggering<br />

apoptosis in aggressive cancers presenting aberrant NF-κB regulati<strong>on</strong>. The results suggest that the block <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

anti-apoptotic signaling pathways utilizing the IκB kinase IKK may play an important role in modulating<br />

HSR pro-apoptotic effects in chemoresistant cancers.<br />

40<br />

1C_03_S<br />

ROLES OF HSF1 IN INFLAMMATORY AND IMMUNE RESPONSE<br />

Akira Nakai<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry and Molecular Biology, Yamaguchi University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Ube, Japan,<br />

e-mail: anakai@yamaguchi-u.ac.jp<br />

Inflammatory cytokines such as IL-1, IL-6, and TNF-α are induced in resp<strong>on</strong>se to bacterial infecti<strong>on</strong> and<br />

diseases. These cytokines elicit the febrile resp<strong>on</strong>se that is a complex physiological reacti<strong>on</strong> to disease<br />

including cytokine-mediated rise in body temperature and activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> inflammatory systems. Fever plays<br />

beneficial roles <strong>on</strong> disease prognosis clinically. Experimentally, pretreatment <str<strong>on</strong>g>of</str<strong>on</strong>g> animals with heat shock also<br />

increases survival in a LPS-injected endotoxic model. These beneficial roles <str<strong>on</strong>g>of</str<strong>on</strong>g> fever are mediated partly by<br />

suppressing pyrogenic and inflammatory cytokines as expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> TNF-α, IL-1β, and IL-6 reduces in heatshocked<br />

cells and in whole body exposed to high temperature. It was shown that HSF1 inhibits expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

cytokines by binding directly to TNF-α promoter, or by physically interacting with NF-IL6, an activator <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

IL-1β. However, molecular mechanisms underlining fever-mediated suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cytokine gene expressi<strong>on</strong><br />

are uncovered yet. Here we show that heat shock suppresses LPS-induced inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> IL-6 by activating<br />

HSF1 that induces ATF3, a negative regulator <str<strong>on</strong>g>of</str<strong>on</strong>g> IL-6. In vivo analysis using HSF1-null and ATF3-null mice<br />

reveals that HSF1 as well as ATF3 acts as a negative regulator <str<strong>on</strong>g>of</str<strong>on</strong>g> IL-6 expressi<strong>on</strong> in whole body and is<br />

required to inhibit fever. Unexpectedly, overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ATF3 into cells has no effect <strong>on</strong> IL-6 expressi<strong>on</strong> in<br />

the absence <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF1. HSF1 also binds directly to IL-6 promoter, and is required not <strong>on</strong>ly <str<strong>on</strong>g>for</str<strong>on</strong>g> a repressor<br />

ATF3, but also an activator NF-kB to bind to IL-6 promoter by partially opening chromatin structure. Taken<br />

together with the effects <strong>on</strong> expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> TNF-α and IL-1β, these results indicate that HSF1 plays a major<br />

role in feedback regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the febrile resp<strong>on</strong>se.<br />

1C_04_S<br />

HSF2 INFLUENCES THE DECISION BETWEEN PROLIFERATION AND<br />

MIGRATION FOR NEURAL CORTICAL PROGENITORS<br />

Diane Trouillet, Anne Le Mouël, Rachid El Fatimy, Laurence Denis, Valérie Mezger<br />

CNRS UMR8541, Ecole Normale Supérieure, 46 rue d’Ulm 75005 Paris<br />

Heat Shock Factors (HSFs) are not <strong>on</strong>ly resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> resp<strong>on</strong>se to envir<strong>on</strong>mental stress, but are involved in<br />

developmental processes. We showed in collaborati<strong>on</strong> with Lea Sist<strong>on</strong>en’s lab that HSF2 is involved in<br />

meiosis in both genders and in brain development (Kallio et al., 2002 ; Chang et al., 2006). We will describe<br />

how HSF2 influences the radial migrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> young postmitotic neur<strong>on</strong>s during cortical development, but<br />

also the proliferati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> their neural progenitors (NPCs), using loss and gain <str<strong>on</strong>g>of</str<strong>on</strong>g> functi<strong>on</strong> models : Hsf2 -/- mice


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

and the overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF2 in the chick neural tube, by in ovo electroporati<strong>on</strong>. The identificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> new<br />

HSF2 target genes suggests us that HSF2 directly regulates genes that are involved in the c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

microtubule dynamics, either in mitosis or during migrati<strong>on</strong>. The questi<strong>on</strong> is how HSF2, which is active in<br />

NPCs as well as in migrating postmitotic neur<strong>on</strong>s, might differentially regulate distinct pools <str<strong>on</strong>g>of</str<strong>on</strong>g> target genes<br />

in these cell populati<strong>on</strong>s. The ability <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF2 to differentially recruit transcripti<strong>on</strong> factors and chromatin<br />

modifiers as well as its distinct biochemical properties in NPCs versus postmitotic neur<strong>on</strong>s has been<br />

investigated by biochemical and chromatin immunoprecipitati<strong>on</strong> (ChIP) analyses and will be discussed. Such<br />

a combinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> events and properties, coupled with the geography <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF2 binding sites and <str<strong>on</strong>g>of</str<strong>on</strong>g> other<br />

transcripti<strong>on</strong> factors sites –characteristic <str<strong>on</strong>g>of</str<strong>on</strong>g> a given target gene – should allow HSF2 to induce or repress<br />

transcripti<strong>on</strong> and there<str<strong>on</strong>g>for</str<strong>on</strong>g>e to allow the decisi<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> a NPC to c<strong>on</strong>tinue to divide or to exit the cell cycle and<br />

start to migrate. Grant supports : ARC (Associati<strong>on</strong> pour la Recherche c<strong>on</strong>tre le Cancer) and ANR<br />

Neurosciences (Agence nati<strong>on</strong>ale pour la Recherche).<br />

1C_05_S<br />

(poster secti<strong>on</strong> A1, poster board #26, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DEVELOPMENTAL ACTIVITY OF HSF1 REVEALED BY LOSS OF FUNCTION<br />

EXPERIMENTS<br />

Elizabeth Chritians<br />

Université Paul Sabatier - Toulouse III, <str<strong>on</strong>g>Centre</str<strong>on</strong>g> de Biologie du Développement - UMR 5547, 118,<br />

Route de Narb<strong>on</strong>ne-4R3B3, 31062 Toulouse, France<br />

Traditi<strong>on</strong>al descripti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF1 activity is based <strong>on</strong> stress inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> molecular modificati<strong>on</strong>s which enable<br />

DNA binding and transcripti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> target genes. This view seems to imply that HSF1 is mainly inactive in<br />

normal, physiological situati<strong>on</strong>. Previously published data and the work presented here str<strong>on</strong>gly suggest that<br />

this descripti<strong>on</strong> needs to be revised. Loss <str<strong>on</strong>g>of</str<strong>on</strong>g> functi<strong>on</strong> by gene targeting <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsf1 in mice revealed a complex<br />

phenotype including HSF1 maternal requirement in the oocyte. Hsf1-/- females produce oocyte but they<br />

appear to be unable to properly accomplish expected developmental steps such as meiotic maturati<strong>on</strong>,<br />

fertilizati<strong>on</strong> and activati<strong>on</strong>, which are mandatory <str<strong>on</strong>g>for</str<strong>on</strong>g> embry<strong>on</strong>ic development. In order to better understand<br />

the link between HSF1 and those developmental steps, we searched <str<strong>on</strong>g>for</str<strong>on</strong>g> known and unknown targets <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

HSF1 in oocytes. Using RT combined with real time PCR, we show that Hsp genes are differentially<br />

expressed and regulated by HSF1 in oocytes under normal, physiological c<strong>on</strong>diti<strong>on</strong>s. Am<strong>on</strong>g HSF1 known<br />

targets, Hsp86 (Hsp90alpha), which is described as the inducible <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90, is significantly reduced in<br />

immature oocyte be<str<strong>on</strong>g>for</str<strong>on</strong>g>e meiotic maturati<strong>on</strong>. Additi<strong>on</strong>al experiments will be discussed to show how Hsp90<br />

alpha can be <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the key link between HSF1 and early developmental steps undertaken by the oocytes.<br />

1C_06_S<br />

(poster secti<strong>on</strong> A1, poster board #27, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DUAL ROLES OF HEAT SHOCK FACTOR 1 (HSF1) IN CARDIOPROTECTION,<br />

PATHOLOGIC HYPERTROPHY AND HEART FAILURE IN TRANSGENIC MICE<br />

András Orosz, Ivor Benjamin<br />

Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Cardiology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Utah, Salt Lake City, USA, 84112<br />

e-mail: andras.orosz@hsc.utah.edu<br />

Heat shock factor 1 (HSF1), the major stress-inducible transactivator binds to heat shock elements (HSE)<br />

embedded in the promoter <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock proteins (HSPs) under stressful c<strong>on</strong>diti<strong>on</strong>s and up-regulates the<br />

41


23-26 August 2007,<br />

Budapest, Hungary<br />

synthesis <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP genes. In principle, HSF1 activati<strong>on</strong> in the heart provides protecti<strong>on</strong> against ischemic insults<br />

and other stressful episodes through the increased synthesis and chaper<strong>on</strong>e activities <str<strong>on</strong>g>of</str<strong>on</strong>g> HSPs. HSF1 level is<br />

also up-regulated in physiological cardiac hypertrophy following strenuous exercise regimens. To study HSF1<br />

functi<strong>on</strong>s in the heart, we generated a transgenic mouse model expressing cardiac specific, inducible, but<br />

c<strong>on</strong>stitutively active HSF1 whose transcripti<strong>on</strong>al competency does not require stressors. Two transgenic<br />

mouse lines were characterized, termed mHSF1(+) Low Tg and High Tg, with transgene expressi<strong>on</strong> ranging<br />

between equivalent and ~5-10 fold <str<strong>on</strong>g>of</str<strong>on</strong>g> the endogenous HSF1 level, respectively. In mHSF1(+) Low Tg mice,<br />

7 days expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> mHSF1(+) induced ~3-fold upregulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> each major HSP groups, which c<strong>on</strong>ferred<br />

significantly increased ischemic protecti<strong>on</strong> in an ex vivo heart injury model. In c<strong>on</strong>trast, doxycyclin feeding <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

mHSF1(+) High Tg line triggered ~5-10-fold increase in mHSF1(+) and target HSPs levels and ~30%<br />

cardiac hypertrophy in 7 days. Interestingly, 3 weeks mHSF1(+) expressi<strong>on</strong> resulted in decompensated<br />

cardiac hypertrophy, massive cell death and fibrosis al<strong>on</strong>g with greatly decreased cardiac functi<strong>on</strong> and overt<br />

heart failure in transgenic mice. Mechanistically, we hypothesize that upregulated class I and class II hist<strong>on</strong><br />

deacetylase family members, direct or indirect targets <str<strong>on</strong>g>of</str<strong>on</strong>g> mHSF1(+), may potentiate cardiomyocyte<br />

hypertrophy, whereas decreased expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> PGC-1α a transcripti<strong>on</strong>al co-activator <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>drial<br />

biogenesis accelerates the metabolic demise <str<strong>on</strong>g>of</str<strong>on</strong>g> mHSF1(+) High Tg hearts. Our studies have clearly<br />

established an intriguing dichotomy, based <strong>on</strong> low and high levels <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF1 transactivati<strong>on</strong> in both protective<br />

and pathological cardiac gene expressi<strong>on</strong> programs and, perhaps, human pathologies.<br />

42


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

1C_01_P<br />

(poster secti<strong>on</strong> A1, poster board #28, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CRYSTAL STRUCTURE OF PYROCOCCUS FURIOSUS HEAT SHOCK REGULATOR,<br />

A MOLECULAR CHIMERA REPRESENTING EUKARYAL AND BACTERIAL<br />

FEATURES<br />

Wei Liu 1 , Gudrun Vierke 2 , Ann-Kathrin Wenke 2 , Michael Thomm 2 , Rudolf Ladenstein 1<br />

1 Karolinska Institutet, Novum, Center <str<strong>on</strong>g>for</str<strong>on</strong>g> Structural Biochemistry, 141 57 Huddinge, Sweden<br />

2 Archaea Center, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Regensburg, 93053 Regensburg, Germany<br />

e-mail: wei.liu@biosci.ki.se<br />

All living organisms share a comm<strong>on</strong> molecular stress resp<strong>on</strong>se up<strong>on</strong> rapidly up-shifted envir<strong>on</strong>mental<br />

temperature. The heat shock resp<strong>on</strong>se is characterized by a dramatic change in gene expressi<strong>on</strong> patterns and<br />

elevated syntheses <str<strong>on</strong>g>of</str<strong>on</strong>g> a family <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock protein, most <str<strong>on</strong>g>of</str<strong>on</strong>g> which functi<strong>on</strong> as molecular chaper<strong>on</strong>es in<br />

preventing the aggregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> denatured proteins and/or helping protein refolding. The expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> most<br />

heat shock genes is strictly repressed under normal c<strong>on</strong>diti<strong>on</strong>s, but activated <strong>on</strong>ce stress resp<strong>on</strong>se is triggered.<br />

The mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock regulati<strong>on</strong> differs am<strong>on</strong>g the three kingdoms. Compared to bacteria and<br />

eukaryotes little is known <strong>on</strong> heat shock regulati<strong>on</strong> in archaea. The first transcripti<strong>on</strong>al regulator selectively<br />

inhibiting cell-free transcripti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> archaeal heat shock promoters has been recently identified from the<br />

hyperthermophilic archae<strong>on</strong> P. furiosus (1). The 24 kDa protein named as Phr <str<strong>on</strong>g>for</str<strong>on</strong>g>ms a homodimer and<br />

specifically inhibits the transcripti<strong>on</strong> in vitro and in vivo by binding to a 29-bp DNA sequence overlapping<br />

the transcripti<strong>on</strong> start site in heat shock promoters. Phr established a novel protein family with n<strong>on</strong>homologous<br />

amino acid sequence with eukaryotic HSFs. The regulator specifically represses the expressi<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock genes at physiological temperature in vitro and in vivo but is released from the promoters up<strong>on</strong><br />

heat shock resp<strong>on</strong>se. We report here the crystal structure <str<strong>on</strong>g>of</str<strong>on</strong>g> Phr which represents the first characterized heat<br />

shock transcripti<strong>on</strong> factor in archaea (2). Structure analysis revealed a stable homodimer, each subunit<br />

c<strong>on</strong>sisting <str<strong>on</strong>g>of</str<strong>on</strong>g> a N-terminal winged helix DNA-binding domain (wH-DBD) and a C-terminal antiparallel<br />

coiled coil helical domain. The overall structure shows as a molecular chimera with significant folding<br />

similarity <str<strong>on</strong>g>of</str<strong>on</strong>g> its DBD to the bacterial SmtB/ArsR family, while its C-terminal part was found to be a remote<br />

homologue <str<strong>on</strong>g>of</str<strong>on</strong>g> the eukaryotic BAG domain. The dimeric protein recognizes a palindromic DNA sequence.<br />

Molecular docking and mutati<strong>on</strong>al analyses suggested a novel binding mode in which the major specific<br />

c<strong>on</strong>tacts occur at the minor groove interacting with the str<strong>on</strong>gly basic wing c<strong>on</strong>taining a cluster <str<strong>on</strong>g>of</str<strong>on</strong>g> 3 arginine<br />

residues. These results argue <str<strong>on</strong>g>for</str<strong>on</strong>g> an unprecedented DNA binding mode.<br />

43


23-26 August 2007,<br />

Budapest, Hungary<br />

1C_02_P<br />

(poster secti<strong>on</strong> A1, poster board #29, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

GLOBAL CHIP-CHIP PROMOTER ANALYSIS IN MOUSE TESTIS REVEALS HSF2<br />

BINDING TO THE Y CHROMOSOME<br />

Malin Åkerfelt 1,2 *, Eva Henrikss<strong>on</strong> 1,2 *, Asta Laiho 1 , Lea Sist<strong>on</strong>en 1,2<br />

1 Turku <str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g> Biotechnology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Turku and Åbo Akademi University, Turku, Finland<br />

2 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, Åbo Akademi University, Turku, Finland. * Equal c<strong>on</strong>tributi<strong>on</strong><br />

e-mail: malin.akerfelt@btk.fi, eva.henrikss<strong>on</strong>@btk.fi<br />

Heat shock factor 2, HSF2, is a member <str<strong>on</strong>g>of</str<strong>on</strong>g> an evoluti<strong>on</strong>ary c<strong>on</strong>served HSF transcripti<strong>on</strong> factor family, which<br />

in mammals c<strong>on</strong>sists <str<strong>on</strong>g>of</str<strong>on</strong>g> three genes, Hsf1, Hsf2 and Hsf4. The HSFs were originally found as regulators <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the heat shock resp<strong>on</strong>se, but mutant analyses revealed HSFs also to be important developmental factors. The<br />

Hsf2-/- mice show defects in the development <str<strong>on</strong>g>of</str<strong>on</strong>g> the CNS and in the functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the reproducti<strong>on</strong> systems<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> both genders. In males, increased apoptosis in testis as well as reduced sperm count are detected. Further,<br />

the structure <str<strong>on</strong>g>of</str<strong>on</strong>g> the seminiferous tubules is altered, displaying less differentiating spermatocytes and extensive<br />

vacuolizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the tubules. The molecular mechanisms causing these defects have remained obscure. Thus,<br />

we per<str<strong>on</strong>g>for</str<strong>on</strong>g>med a chromatin immunoprecipitati<strong>on</strong> <strong>on</strong> promoter microarray (ChIP-chip) screen <strong>on</strong> wild type<br />

mouse testis to find HSF2 target genes. We have now identified a large set <str<strong>on</strong>g>of</str<strong>on</strong>g> novel target promoters <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF2<br />

in mouse testis. In additi<strong>on</strong> to DNA binding, we have verified that HSF2 is transcripti<strong>on</strong>ally active as the<br />

corresp<strong>on</strong>ding mRNAs are altered in Hsf2-/- testis. Many <str<strong>on</strong>g>of</str<strong>on</strong>g> the novel target genes <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF2 are indeed<br />

involved in spermatogenesis. The ChIP-chip screen surprisingly revealed a significant accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF2<br />

<strong>on</strong> genes located <strong>on</strong> the Y chromosome and important in sperm differentiati<strong>on</strong>. Taken together, our results<br />

provide str<strong>on</strong>g evidence <str<strong>on</strong>g>for</str<strong>on</strong>g> HSF2 playing significant role in spermatogenetic processes.<br />

1C_03_P<br />

(poster secti<strong>on</strong> A1, poster board #30, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

COREST REPRESSES HEAT SHOCK RESPONSE MEDIATED BY HSF1<br />

A. V. Gómez 1 , E. Battaglioli 2 , J. C. Mass 3 , G. Mandel 4 , M. Kukuljan 3 , M. E. Andrés 1<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Cellular and Molecular Biology, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Biological Sciences,<br />

P<strong>on</strong>tificia Universidad Catolica de Chile, Santiago, Chile<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology and Genetics <str<strong>on</strong>g>for</str<strong>on</strong>g> Medical Sciences, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Milan, Milan, Italy<br />

3Centro de Neurociencias Integradas, Iniciativa Científica Milenio y Programa de Fisiología y Bi<str<strong>on</strong>g>of</str<strong>on</strong>g>ísica, ICBM,<br />

Facultad de Medicina, Universidad de Chile, Santiago, Chile<br />

4Howard Hughes Medical Institute, Vollum Institute at the Oreg<strong>on</strong> Health & Science University, Portland, Oreg<strong>on</strong>,<br />

USA<br />

The transcripti<strong>on</strong>al corepressor CoREST is a main comp<strong>on</strong>ent <str<strong>on</strong>g>of</str<strong>on</strong>g> a chromatin modifying complex that<br />

recruits the hist<strong>on</strong>e deacetylases, HDACs1/2, and the hist<strong>on</strong>e lysine specific demethylase, LSD1. First<br />

characterized as a corepressor <str<strong>on</strong>g>of</str<strong>on</strong>g> the RE1 Silencing Factor/Neural restrictive silencing factor (REST/NRSF),<br />

its transcripti<strong>on</strong>al regulator role has been already dem<strong>on</strong>strated in the c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>al<br />

genes. The high evoluti<strong>on</strong>ary c<strong>on</strong>servati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> CoREST complex, allows suspecting that it might c<strong>on</strong>trol the<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> another subset <str<strong>on</strong>g>of</str<strong>on</strong>g> genes. By a yeast two hybrid screening assay, using CoREST as bait, we<br />

identified the molecular chaper<strong>on</strong>e heat shock protein 70 (Hsp70) as a CoREST interacting protein. We<br />

44


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

corroborated the associati<strong>on</strong> between both proteins, and delimited the interacting regi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> CoREST to its<br />

both SANT domains. C<strong>on</strong>sidering that Hsp70 is involved in the heat shock resp<strong>on</strong>se as a transcripti<strong>on</strong>al<br />

corepressor <str<strong>on</strong>g>of</str<strong>on</strong>g> Heat Shock Factor 1 (HSF1), we evaluated whether CoREST regulate the heat shock<br />

resp<strong>on</strong>se. Our results show that CoREST overexpressi<strong>on</strong> represses the heat shock inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a reporter<br />

gene driven by the human Hsp70 promoter. Moreover, CoREST represses HSF1 transcripti<strong>on</strong>al activity in<br />

both c<strong>on</strong>trol and heat shock treatment. Next, we analyzed if Hsp70 repressor effect depend <strong>on</strong> CoREST<br />

expressi<strong>on</strong>. Reducing CoREST expressi<strong>on</strong> by using a short hairpin RNA induces a significant increase over<br />

the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the Hsp70 promoter mediated by HSF1. Moreover, overexpressing Hsp70 is not able to<br />

repress the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF1 in cells with reduced CoREST expressi<strong>on</strong>. In c<strong>on</strong>clusi<strong>on</strong>, we dem<strong>on</strong>strated that<br />

CoREST regulates the heat shock resp<strong>on</strong>se at transcripti<strong>on</strong>al level repressing HSF1 activity.<br />

1C_05_P<br />

(poster secti<strong>on</strong> A1, poster board #31, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HEAT SHOCK FACTOR (HSF) 4A INHIBITS HEMIN-INDUCED ACTIVATION OF<br />

HSF2<br />

Sang-Gun Ahn 1 , Jung-Ho<strong>on</strong> Yo<strong>on</strong> 1 , Soo-A Kim 2, *<br />

1Oral Biology <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute, BK21 projects, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Pathology, Chosun University College <str<strong>on</strong>g>of</str<strong>on</strong>g> Dentistry,<br />

Gwangju, KOREA, 2 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, D<strong>on</strong>gguk University College <str<strong>on</strong>g>of</str<strong>on</strong>g> Oriental Medicine, Gye<strong>on</strong>gju,<br />

KOREA, * corresp<strong>on</strong>ding author<br />

The inducible regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock gene transcripti<strong>on</strong> is mediated by a family <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock factors<br />

(HSFs) that resp<strong>on</strong>d to diverse <str<strong>on</strong>g>for</str<strong>on</strong>g>ms <str<strong>on</strong>g>of</str<strong>on</strong>g> physiological and envir<strong>on</strong>mental stress including temperature, heavy<br />

metals, oxidative stress. Although HSFs have been extensively studied with respect to their regulati<strong>on</strong> by<br />

molecular chaper<strong>on</strong>es, regulatory mechanism between HSF family members are poorly understood. In this<br />

study, we identified the interacti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF2 with HSF4A but not with HSF4B under n<strong>on</strong>-stressed c<strong>on</strong>diti<strong>on</strong>s.<br />

Using immunoprecipitati<strong>on</strong> assay, we found that leucine zipper domain 1-3 <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF4A was directly interacts<br />

with HSF2 and inhibits transcripti<strong>on</strong>al activity <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF2. Interestingly, HSF4A was accumulated to the<br />

nucleus up<strong>on</strong> hemin treatment and inhibited trimerizati<strong>on</strong> and following target gene transcripti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF2.<br />

These observati<strong>on</strong>s suggest HSF4A as an active repressor <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF2-mediated transcripti<strong>on</strong>.<br />

1C_06_P<br />

(poster secti<strong>on</strong> A1, poster board #32, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

GENETIC EVIDENCE FOR A PROTECTIVE ROLE OF HEAT SHOCK FACTOR 1<br />

AGAINST GASTRIC ULCER AND COLITIS RELATED TO HUMAN<br />

INFLAMMATORY BOWEL DISEASE<br />

Ken-ichiro Tanaka, Tohru Mizushima<br />

Graduate School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical and Pharmaceutical Sciences, Kumamoto University<br />

Gastric lesi<strong>on</strong>s result from an imbalance between aggressive and defensive factors. Indirect lines <str<strong>on</strong>g>of</str<strong>on</strong>g> evidence<br />

suggest that heat shock factor 1 (HSF1)-dependent inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock proteins (HSPs) by various<br />

aggressive factors provide a major protective mechanism. On the other hand, inflammatory bowel disease<br />

(IBD) involves infiltrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> leukocytes into intestinal tissue and both pro-inflammatory cytokines (such as<br />

TNF-α) and cell adhesi<strong>on</strong> molecules (CAMs) play an important role in this step. However, the role <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF1<br />

45


23-26 August 2007,<br />

Budapest, Hungary<br />

in development <str<strong>on</strong>g>of</str<strong>on</strong>g> IBD has remained unknown. In this study, we examined the producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> gastric lesi<strong>on</strong>s<br />

and DSS-induced colitis, animal model <str<strong>on</strong>g>of</str<strong>on</strong>g> IBD, using HSF1-null mice lacking HSF1. The producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

gastric lesi<strong>on</strong>s by ethanol or hydrochloric acid was stimulated in HSF1-null mice. Ethanol administrati<strong>on</strong> upregulated<br />

gastric mucosal HSPs, in particular HSP70, in an HSF1-dependent manner, and more apoptotic<br />

cells were observed in the gastric mucosa <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF1-null mice than in wild-type mice. Geranylgeranylacet<strong>on</strong>e<br />

(GGA), a clinically used anti-ulcer drug with HSP-inducing activity, suppressed ethanol-induced gastric<br />

lesi<strong>on</strong>s in wild-type mice but not in HSF1-null mice. The results suggest that the producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> irritantinduced<br />

gastric lesi<strong>on</strong>s in HSF1-null mice is due to their inability to up-regulate HSPs, leading to apoptosis.<br />

It is also suggested that the HSP-inducing activity <str<strong>on</strong>g>of</str<strong>on</strong>g> GGA c<strong>on</strong>tributes to the drug’s anti-ulcer activity. On<br />

the other hand, the DSS-induced colitis was worsened in HSF1-null mice. DSS-administrati<strong>on</strong> up-regulated<br />

HSP70 at col<strong>on</strong>ic tissues in an HSF1-dependent manner. Comparing to wild-type mice, levels <str<strong>on</strong>g>of</str<strong>on</strong>g> proinflammatory<br />

cytokines (such as TNF-α) and CAMs at col<strong>on</strong>ic tissues with DSS-administrati<strong>on</strong> were<br />

increased in HSF1-null mice. Macrophages prepared from HSF1-null mice showed higher activity <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

lipopolysaccharide-stimulated generati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> TNF-α. Suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> hsf1 expressi<strong>on</strong> stimulated<br />

lipopolysaccharide-induced up-regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> CAMs. These results suggest that HSF1 play a protective role<br />

against DSS-induced colitis. Furthermore, this protective role seems to involve suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

TNF-α and CAMs. This study provides the first direct genetic evidence that HSF1 c<strong>on</strong>fer protecti<strong>on</strong> against<br />

the development <str<strong>on</strong>g>of</str<strong>on</strong>g> gastric lesi<strong>on</strong>s and colitis related to human IBD.<br />

1C_07_P<br />

(poster secti<strong>on</strong> A1, poster board #33, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CHARACTERIZATION OF THE DNA-BINDING SITES OF HSF4 THAT<br />

CONSTITUTIVELY FORMS A TIMER<br />

Mitsuaki Fujimoto, Eiichi Takaki, Naoki, Hayashida, Sachiye Inouye, Akira Nakai<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry and Molecular Biology, Yamaguchi University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Minami-Kogushi,<br />

Ube, Japan, e-mail: yoji530@yamaguchi-u.ac.jp<br />

Heat shock transcripti<strong>on</strong> factors (HSFs) regulates expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock genes and genes related to<br />

development. HSFs bind to heat shock element (HSE), which is composed <str<strong>on</strong>g>of</str<strong>on</strong>g> at least three inverted repeats<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> c<strong>on</strong>sensus sequence nGAAn. Under normal c<strong>on</strong>diti<strong>on</strong>, HSF1 and HSF2 exist as a m<strong>on</strong>omers and a<br />

dimmer, respectively. HSF1 is c<strong>on</strong>verted to a trimer up<strong>on</strong> heat shock. In c<strong>on</strong>trast, HSF4 c<strong>on</strong>stitutively stays a<br />

trimer that binds to the HSE under normal c<strong>on</strong>diti<strong>on</strong>. Recent observati<strong>on</strong>s showed that HSFs plays roles in<br />

physiology including developmental processes and complements or competes with each other. However, we<br />

do not understand how HSFs do so. To examine their cooperativity, here we examined specificity <str<strong>on</strong>g>of</str<strong>on</strong>g> DNAbinding<br />

by HSFs. We firstly determined the HSE-binding specificity <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF4 by a random olig<strong>on</strong>ucleotide<br />

selecti<strong>on</strong> method. We found that the G nucleotide in 5’-nGAAn-3’ in all <str<strong>on</strong>g>of</str<strong>on</strong>g> the selected sequences is<br />

c<strong>on</strong>served, but there is little preferences <str<strong>on</strong>g>for</str<strong>on</strong>g> the nucleotides in As. Mutati<strong>on</strong>s in the A nucleotides in the<br />

nGAAn caused decreased binding <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF1 and HSF2 whereas those affected HSF4 binding <strong>on</strong>ly a little.<br />

There results indicate that the HSF4-binding sequence is an inverted repeats <str<strong>on</strong>g>of</str<strong>on</strong>g> nGnnn. We next per<str<strong>on</strong>g>for</str<strong>on</strong>g>med<br />

in vivo chromatin immunoprecipitati<strong>on</strong> (CHIP) assay, and 71 candidate target genes were identified.<br />

C<strong>on</strong>sensus sequence <str<strong>on</strong>g>of</str<strong>on</strong>g> the putative HSF4-binding sites was the same as that identified by a random<br />

olig<strong>on</strong>ucleotide selecti<strong>on</strong> method. These results indicate that HSF4 possesses str<strong>on</strong>ger affinity <str<strong>on</strong>g>for</str<strong>on</strong>g> a set <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

DNA-binding sites, whose sequences weakly matched with a can<strong>on</strong>ical HSE sequence, than HSF1 and HSF2.<br />

46


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

1C_08_P<br />

(poster secti<strong>on</strong> A1, poster board #34, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HSFS, A MOLECULAR BASIS FOR FETAL ALCOHOL SYNDROME (FAS)<br />

Rachid El Fatimy 1 , Anne Le Mouel 1 , Leslie Schwenndimann 2 , Pierre Gressens, Valérie Mezger 1<br />

1CNRS UMR8541, École Normale Supérieure, 46 rue d’Ulm, 75005 PARIS, France<br />

2INSERM 676 et service de Neuropédiatrie, Hôpital Robert Debré, 48 bd Sérurier, 75019 PARIS, France<br />

The transiti<strong>on</strong> from normal growth c<strong>on</strong>diti<strong>on</strong>s to stress c<strong>on</strong>diti<strong>on</strong>s induces the cell protective heat shock<br />

proteins (HSPs). Hsp genes are activated by Heat Shock Factors (HSFs). HSF1, the major stress-resp<strong>on</strong>sive<br />

transcripti<strong>on</strong> factor in mammals, is activated through a multi-step pathway including posttranslati<strong>on</strong>al<br />

modificati<strong>on</strong>s and binding to Heat shock Elements (HSE) upstream <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp genes. Although reported to be<br />

desactivated by heat shock (HS), HSF2 c<strong>on</strong>tributes to Hsp inducti<strong>on</strong> through interplay with HSF1. HSFs are<br />

also involved in development. Cerebral cortex c<strong>on</strong>sists in 6 layers <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>s arising from migrati<strong>on</strong>. We<br />

showed that Hsf2 -/- cerebral cortices display mispositi<strong>on</strong>ing <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>s due to disturbances in the expressi<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> target genes which c<strong>on</strong>trol neur<strong>on</strong>al migrati<strong>on</strong> signaling pathways (Chang Y, 2006 Genes & Dev). We are<br />

interested by the intriguing role <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF1 and HSF2 as coordinators between stress and development.<br />

Chr<strong>on</strong>ic alcoholic intoxicati<strong>on</strong> (CAI) causes neur<strong>on</strong>al migrati<strong>on</strong> defects in brain development, which are part<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the fetal alcohol syndrome (FAS), but whose molecular basis remain elusive. Since alcohol induces HSPs in<br />

cultured cells, we hypothesized that CAI disturbs HSF1/HSF2 activities in fetal cortices. Such alterati<strong>on</strong>s<br />

could disturb the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> target genes that c<strong>on</strong>tain HSE and are regulated by HSF2 <strong>on</strong>ly during normal<br />

neur<strong>on</strong>al migrati<strong>on</strong>. We showed that HSF1 and HSF2 activities are disturbed both in ex vivo alcoholic<br />

exposure <str<strong>on</strong>g>of</str<strong>on</strong>g> cultured cells and in vivo CAI <str<strong>on</strong>g>of</str<strong>on</strong>g> fetal cortices. Differential HSF1 posttranslati<strong>on</strong>al modificati<strong>on</strong>s<br />

were induced by alcohol versus HS. Modificati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF activities correlated to disturbances in the<br />

expressi<strong>on</strong> levels <str<strong>on</strong>g>of</str<strong>on</strong>g> various HSF2 target genes involved in neur<strong>on</strong>al migrati<strong>on</strong>. By ChIP (chromatin<br />

immunoprecipitati<strong>on</strong>), we characterized the differential in vivo occupancies by HSF1 versus HSF2 <strong>on</strong> these<br />

genes. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e HSFs provide a molecular explanati<strong>on</strong> to the FAS aspect <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>al migrati<strong>on</strong> defects.<br />

1C_09_P<br />

(poster secti<strong>on</strong> A1, poster board #35, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

A NEW MATHEMATICAL MODEL FOR THE EUKARYOTIC HEAT SHOCK<br />

RESPONSE<br />

I. Petre*, C. Hyder*, A. Mizera*, D. Preoteasa # , A. Mikhailov*, J. E. Erikss<strong>on</strong>*, L. Sist<strong>on</strong>en*,<br />

R.-J. Back*<br />

*Abo Akademi University, 20520 Turku, # University <str<strong>on</strong>g>of</str<strong>on</strong>g> Helsinki, 00014 Helsinki, Finland,<br />

e-mail: ipetre@abo.fi<br />

We present a math model <str<strong>on</strong>g>of</str<strong>on</strong>g> the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the heat shock protein hsp70 in eukaryotic cells. The hsp70<br />

proteins interact with misfolded proteins, helping them to refold. Experimental results show that under heat<br />

shock, there is <strong>on</strong>ly a transient period <str<strong>on</strong>g>of</str<strong>on</strong>g> sustained transactivity <str<strong>on</strong>g>of</str<strong>on</strong>g> the hsp70-encoding genes, even under<br />

c<strong>on</strong>tinued heat shock. The transactivity <str<strong>on</strong>g>of</str<strong>on</strong>g> the hsp70-encoding genes is regulated by the heat shock factor<br />

(HSF) proteins that trimerize under heat shock and bind to the heat shock elements (HSE). Once the level <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

hsp70 is sufficient, its further synthesis is blocked: hsp70 helps unbinding the HSF trimer from HSE.<br />

Moreover, hsp70 blocks subsequent <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF trimers by binding to HSF m<strong>on</strong>omers. Once a new<br />

heat shock is encountered, hsp70 resumes its chaper<strong>on</strong>e activity, releasing the HSF proteins, which then<br />

47


23-26 August 2007,<br />

Budapest, Hungary<br />

trimerize and promote the transactivity. We c<strong>on</strong>sider in our model the detailed kinetics <str<strong>on</strong>g>of</str<strong>on</strong>g> this network. Some<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the model parameters were deduced from literature, while others were estimated. Our current model<br />

predicts correctly the transient activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF trimers under heat shock at 42 o C, while showing low level<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> activity at 37 o C. Although the misfolding rate is about 10 times higher at 42 o C than at 37 o C, the predicted<br />

steady state at 42 o C is identical with that at 37 o C, except <str<strong>on</strong>g>for</str<strong>on</strong>g> the level <str<strong>on</strong>g>of</str<strong>on</strong>g> misfolded proteins. This suggests<br />

that if cells survive the initial stage <str<strong>on</strong>g>of</str<strong>on</strong>g> sustained cellular activity, they manage to return to the same levels as at<br />

37 o C. Sensitivity analysis shows, however, that the system is more sensitive at 42 o C than at 37 o C: small<br />

changes in the kinetics <str<strong>on</strong>g>of</str<strong>on</strong>g> certain reacti<strong>on</strong>s will yield greater change in the resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> the network at 42 o C<br />

than at 37 o C.<br />

48


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

1D. EXTRACELLULAR CHAPERONES<br />

(STUART CALDERWOOD)<br />

1D_01_S<br />

HSP RELEASE: PASSIVE VS ACTIVE RELEASE MECHANISMS<br />

Alexzander Asea<br />

Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Investigative Pathology, Scott & White Clinic and Texas A&M University System Health Science<br />

Center College <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, 2401 South 31 st Street, Temple, TX 76508 United States<br />

e-mail: asea@medicine.tamhsc.edu<br />

Thus far, two mechanisms are recognized by which heat shock proteins (HSP) are released from cells into the<br />

extracellular milieu and subsequently into systemic circulati<strong>on</strong>; a passive release mechanism as a result <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

necrotic cell death, severe blunt trauma, surgery and following infecti<strong>on</strong> with lytic viruses, and an active<br />

release mechanism which involves the n<strong>on</strong> classical protein release pathway by which HSP70 is released as<br />

free HSP72 and within highly immunologically potent exosomes. This presentati<strong>on</strong> covers the most recent<br />

findings <strong>on</strong> the mechanisms by which stress induces the release <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP72 into the systemic circulati<strong>on</strong> and<br />

addresses the biological significance <str<strong>on</strong>g>of</str<strong>on</strong>g> circulating HSP72 to host defense against disease.<br />

1D_02_S<br />

HSP70 SECRETION AND BINDING: ITS PLACE IN THE IMMUNE RESPONSE<br />

Stuart K. Calderwood, Salamatu S Mambula, Jimmy Theriault, Jianlin G<strong>on</strong>g<br />

Beth Israel Deac<strong>on</strong>ess Medical center, Harvard Medical School and Center <str<strong>on</strong>g>for</str<strong>on</strong>g> Molecular Stress Resp<strong>on</strong>se,<br />

Bost<strong>on</strong> University Medical School.<br />

Hsp70 plays a significant extracellular role within the immune resp<strong>on</strong>se and can carry out both proinflammatory<br />

/ pro-immune functi<strong>on</strong>s and anti-inflammatory effects depending <strong>on</strong> cellular and tissue<br />

c<strong>on</strong>text. However, understanding its place in the immune resp<strong>on</strong>se requires deciphering the mechanisms by<br />

which it is released and the cell surface structures which it binds <strong>on</strong> target cells in the immune system.<br />

We have examined mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 release from tumor cells and macrophages and have found<br />

evidence <str<strong>on</strong>g>for</str<strong>on</strong>g> an active secreti<strong>on</strong> mechanism. Our experiments indicate that hsp70 secreti<strong>on</strong> employs a similar<br />

pathway to that used by other “leaderless” proteins such as interleukin 1βIL-1β) involving the entry <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70<br />

into secretory lysosomes and ATP dependent release. Our evidence suggest however, that the trigger <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

hsp70 release is different from that which releases IL-1β: <str<strong>on</strong>g>for</str<strong>on</strong>g> instance, in macrophages exposed to E. coli, the<br />

trigger <str<strong>on</strong>g>for</str<strong>on</strong>g> IL-1β release is lipopolysaccharide, while Hsp70 release is triggered by a different signal. In<br />

additi<strong>on</strong>, IL-1β release is independent <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 release and Hsp70 blocking does not alter release IL-1β.<br />

Hsp70 released from cells binds to adjacent cells. Our experiments indicate that such receptors may include a<br />

diverse group <str<strong>on</strong>g>of</str<strong>on</strong>g> structures. Hsp70 signaling can be mediated by Toll Like receptors (TLR), CD40 and<br />

LRP/CD91. However, our recent studies indicate that Hsp70 internalizati<strong>on</strong> is mediated by scavenger<br />

receptors (SR) found <strong>on</strong> antigen presenting cells. We have found that at least 3 members <str<strong>on</strong>g>of</str<strong>on</strong>g> the SR including<br />

LOX-1, SREC-1 and FEEL-1/CLEVER-1 can bind and internalize Hsp70. As the SR play a pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ound role in<br />

the “cross presentati<strong>on</strong>” <str<strong>on</strong>g>of</str<strong>on</strong>g> extracellular proteins to CD8+ T lymphocytes, these experiments suggest a<br />

pathway al<strong>on</strong>g which secreted Hsp70 with chaper<strong>on</strong>ed peptide antigens can be taken up by APC and interact<br />

with immune cells.<br />

49


23-26 August 2007,<br />

Budapest, Hungary<br />

1D_03_S<br />

EXTRACELLULAR HEAT SHOCK PROTEINS: SEARCHING FOR A ROLE<br />

John H. H. Williams<br />

Chester <str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g> Stress <str<strong>on</strong>g>Research</str<strong>on</strong>g>, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biological Sciences, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Chester, Parkgate Road, Chester,<br />

CH1 4BJ, UK,<br />

e-mail: John.Williams@chester.ac.uk<br />

Hsp70 can be detected in serum, despite being an intra-cellular protein. Hsp70 must there<str<strong>on</strong>g>for</str<strong>on</strong>g>e be released by<br />

damaged cells, or secreted by healthy cells. We have previously dem<strong>on</strong>strated that Hsp70 and Hsp60 are<br />

released from cells. An increasing number <str<strong>on</strong>g>of</str<strong>on</strong>g> cell types, including peripheral blood m<strong>on</strong><strong>on</strong>uclear cells<br />

(PBMCs), have been dem<strong>on</strong>strated to release Hsps, including Hsp70, Hsp60 and Hsp90. This release <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp<br />

is stimulated, in vivo and in vitro, by a wide variety <str<strong>on</strong>g>of</str<strong>on</strong>g> stressors. Several cell types resp<strong>on</strong>d to extracellular<br />

Hsp exposure by releasing cytokines, such as TNFα. The type and source (human or bacterial) <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp alter<br />

the cytokine resp<strong>on</strong>se. Extracellular Hsp70 protects cells from heat shock without needing to enter the cells.<br />

It is unclear whether the interacti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these extracellular hsps with the membrane is directly with lipid or<br />

protein. The aims <str<strong>on</strong>g>of</str<strong>on</strong>g> this talk are to discuss: (1) routes <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp release, and through this challenge the<br />

paradigm that <strong>on</strong>ly necrotic cells release Hsps. (2) how extracellular Hsps interact with cells, and through this<br />

investigate their potential roles: as danger signals and as cell protectors.<br />

1D_04_S<br />

QUALITY CONTROL OF EXTRACELLULAR PROTEIN FOLDING:<br />

AN EMERGING FIELD<br />

Justin J. Yerbury, Amy R. Wyatt, Stephen Po<strong>on</strong>, Mark R. Wils<strong>on</strong><br />

School <str<strong>on</strong>g>of</str<strong>on</strong>g> Biological Sciences, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Woll<strong>on</strong>g<strong>on</strong>g, Northfields Avenue, Woll<strong>on</strong>g<strong>on</strong>g. NSW. 2522. Australia<br />

Many serious human diseases are thought to arise from the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> intra- or extracellular aggregates <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

proteins with n<strong>on</strong>-native c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>s. Inside cells, chaper<strong>on</strong>e and protease systems regulate protein<br />

folding; however, little is currently known about any corresp<strong>on</strong>ding mechanisms that operate extracellularly.<br />

Knowledge <str<strong>on</strong>g>of</str<strong>on</strong>g> these mechanisms is important because it may lead to the development <str<strong>on</strong>g>of</str<strong>on</strong>g> new disease<br />

therapies. We have identified a small family <str<strong>on</strong>g>of</str<strong>on</strong>g> abundant human plasma proteins which have a potent small<br />

heat shock protein-like chaper<strong>on</strong>e acti<strong>on</strong>. These proteins occur in plasma at levels <str<strong>on</strong>g>of</str<strong>on</strong>g> 0.1-2.0 mg/ml and thus<br />

in this locale are orders <str<strong>on</strong>g>of</str<strong>on</strong>g> magnitude more abundant than normally intracellular chaper<strong>on</strong>es (e.g. Hsp70)<br />

found extracellularly1. The abundant extracellular chaper<strong>on</strong>es (ECs) <str<strong>on</strong>g>for</str<strong>on</strong>g>m stable soluble complexes with<br />

misfolded and partially unfolded proteins to inhibit amorphous protein aggregati<strong>on</strong> induced by physical or<br />

chemical stresses. The ECs also exert potent effects <strong>on</strong> amyloid <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> and toxicity2. Recent data suggests<br />

that complexes <str<strong>on</strong>g>for</str<strong>on</strong>g>med between ECs and "damaged" (misfolded/unfolded) proteins are rapidly bound by cell<br />

surface receptors in vivo and disposed <str<strong>on</strong>g>of</str<strong>on</strong>g> by receptor-mediated endocytosis and lysosomal degradati<strong>on</strong>. Thus,<br />

a model is emerging in which ECs patrol extracellular spaces, bind to damaged proteins when present and<br />

mediate their rapid clearance and disposal. Disease pathologies associated with protein aggregati<strong>on</strong> may arise<br />

when this system is overloaded.<br />

50


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

1D_05_S<br />

(poster secti<strong>on</strong> A1, poster board #36, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EXTRACELLULAR HSP72: A DOUBLE-EDGED SWORD FOR HEALTH<br />

M<strong>on</strong>ika Fleshner<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Colorado, Dept <str<strong>on</strong>g>of</str<strong>on</strong>g> Integrative Physiology, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Integrative Physiology, Campus Box 354,<br />

80309 Boulder, United States<br />

Envir<strong>on</strong>mental or emoti<strong>on</strong>al challenge triggers a cascading series <str<strong>on</strong>g>of</str<strong>on</strong>g> physiological resp<strong>on</strong>ses which are<br />

collectively termed the “stress resp<strong>on</strong>se”. The stress resp<strong>on</strong>se can be assessed at the behavioral, neural,<br />

horm<strong>on</strong>al, immunological and single cell, levels and evolved to benefit an organism’s chance <str<strong>on</strong>g>of</str<strong>on</strong>g> survival<br />

during times <str<strong>on</strong>g>of</str<strong>on</strong>g> acute challenge. The stress resp<strong>on</strong>se has been studied <str<strong>on</strong>g>for</str<strong>on</strong>g> many years, however, its impact <strong>on</strong><br />

specifically immune functi<strong>on</strong> has <strong>on</strong>ly recently been appreciated. Acute activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress resp<strong>on</strong>se has<br />

both inhibitory and stimulatory effects <strong>on</strong> immunity. The focus <str<strong>on</strong>g>of</str<strong>on</strong>g> this presentati<strong>on</strong> is <strong>on</strong> a novel mechanism<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> the immunostimulatory effects <str<strong>on</strong>g>of</str<strong>on</strong>g> stress. Specifically, we propose that an endogenous, ubiquitous cellular<br />

stress protein, heat shock protein 72, when found in the extracellular envir<strong>on</strong>ment may c<strong>on</strong>tribute to stressinduced<br />

potentiati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> innate immunity. We develop the hypothesis that the release <str<strong>on</strong>g>of</str<strong>on</strong>g> extracellular heat<br />

shock protein 72 (eHsp72) is a normal feature <str<strong>on</strong>g>of</str<strong>on</strong>g> the acute stress resp<strong>on</strong>se that can have either positive or<br />

negative c<strong>on</strong>sequences <str<strong>on</strong>g>for</str<strong>on</strong>g> host defense depending <strong>on</strong> several factors, including the nature <str<strong>on</strong>g>of</str<strong>on</strong>g> the eHsp72<br />

(naked versus antigen-associated), and host health status (absence or presence <str<strong>on</strong>g>of</str<strong>on</strong>g> pre-existing inflammatory<br />

disease). Thus, stress-induced eHsp72 release may be a double-edged sword <str<strong>on</strong>g>for</str<strong>on</strong>g> host defense.<br />

1D_06_S<br />

(poster secti<strong>on</strong> A1, poster board #37, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HSP70 IS SECRETED FROM CELLS IN VESICLES THAT RESEMBLE LIPID RAFTS<br />

A. De Maio 1, 2 , M. Rodríguez 1 , T. Frey 2 , M. Gehrman 3 , D. Nino 1 , J. C. Diaz 4 , C. Steinem 5 ,<br />

N. Arispe 4 , G. Mult<str<strong>on</strong>g>of</str<strong>on</strong>g>f 3 , V. Vega 1<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Surgery, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Cali<str<strong>on</strong>g>for</str<strong>on</strong>g>nia San Diego, La Jolla, Cali<str<strong>on</strong>g>for</str<strong>on</strong>g>nia, USA;<br />

2Johns Hopkins University, Baltimore, Maryland, USA;<br />

3Klinikum Rechts der Isar, Technical University, Munich, Germany;<br />

4Uni<str<strong>on</strong>g>for</str<strong>on</strong>g>med Services University <str<strong>on</strong>g>of</str<strong>on</strong>g> the Health Sciences, Bethesda, Maryland, USA;<br />

5Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Organic and Biomolecular Chemistry, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Goettingen, Göttingen, Germany<br />

Heat shock proteins (hsp) play a major role in a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular processes during normal c<strong>on</strong>diti<strong>on</strong>s as well<br />

as during the restorati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> homeostasis after stress. Recently, Hsp70, the major stress-induced hsp, has been<br />

found in the extracellular medium. Moreover, Hsp70 has been observed to activate cells <str<strong>on</strong>g>of</str<strong>on</strong>g> the immune<br />

system. Thus, circulating Hsp70 is thought to act as a danger signal that triggers the resp<strong>on</strong>se to injury. We<br />

have previously reported that Hsp70 is capable <str<strong>on</strong>g>of</str<strong>on</strong>g> interacting with lipid membranes and has a high selectivity<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> phosphatidylserine (PS). The incorporati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 within PS-c<strong>on</strong>taining membranes was enhanced by<br />

the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> spingolipids (GM1) and cholesterol, which are major comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g> lipid rafts. In fact,<br />

Hsp70 was detected in the lipid raft fracti<strong>on</strong> isolated from Trit<strong>on</strong> X-100 solubilized HepG2 cells after heat<br />

shock (HS). The presence <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 in this fracti<strong>on</strong> was reduced by depleti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular cholesterol by<br />

treatment with β-cyclodextrin. Hsp70 was visualized within the plasma membrane <str<strong>on</strong>g>of</str<strong>on</strong>g> HepG2 cells after HS,<br />

co-localizing with lipid raft markers. Analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> the extracellular medium <str<strong>on</strong>g>of</str<strong>on</strong>g> HepG2 cells after heat shock<br />

revealed the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 within membranes that can be isolated by high speed centrifugati<strong>on</strong>. These<br />

51


23-26 August 2007,<br />

Budapest, Hungary<br />

membranes c<strong>on</strong>taining Hsp70 were rich in GM1 and cholesterol, but depleted <str<strong>on</strong>g>of</str<strong>on</strong>g> other cellular comp<strong>on</strong>ents,<br />

such as actin. Moreover, the Hsp70 within these extracellular membranes was resistant to Trit<strong>on</strong> X-100<br />

solubilizati<strong>on</strong>. Thus, extracellular membranes c<strong>on</strong>taining Hsp70 resemble lipid rafts. We propose that part <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the extracellular Hsp70, which may be involved in the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> immune cells, is present in the membrane<br />

derived from lipid rafts.<br />

52


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

1D_01_P<br />

(poster secti<strong>on</strong> A1, poster board #38, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PROTECTION OF CELLS: A POSSIBLE ROLE FOR EXTRA-CELLULAR HSP70<br />

Helen Williams, Claire Hunter-Lavin, John H. H. Williams<br />

Chester <str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g> Stress <str<strong>on</strong>g>Research</str<strong>on</strong>g>, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biological Sciences, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Chester, Parkgate Road, Chester,<br />

United Kingdom, CH1 4BJ, e-mail: helen.williams@chester.ac.uk<br />

Hsps have been typically regarded as intra-cellular molecules that mediate a range <str<strong>on</strong>g>of</str<strong>on</strong>g> essential housekeeping<br />

and cytoprotective functi<strong>on</strong>s but increasingly there has been accumulating evidence which suggests that these<br />

molecules have importance as extra-cellular proteins. Hsps have been detected <strong>on</strong> both the cell surface and<br />

outside <str<strong>on</strong>g>of</str<strong>on</strong>g> cells under physiological c<strong>on</strong>diti<strong>on</strong>s. We added soluble Hsp70 (bovine and recombinant) to U937<br />

cells and erythrocytes to determine whether this Hsp70 provided added protecti<strong>on</strong> to these cells. A time and<br />

c<strong>on</strong>centrati<strong>on</strong> dependent effect was observed in both cell types. A <strong>on</strong>e hour treatment with 0.1 µg/ml Hsp70<br />

protected U937 cells from apoptosis induced by heat stress at 42.6°C, determined by annexin-V, caspase-3<br />

activity and visually using fluorescence microscopy. One hour incubati<strong>on</strong> with 0.5 µg/ml Hsp70 protected<br />

U937 cells from necrosis induced by 46.2°C heat shock, measured by propidium iodide, and 1 hr <str<strong>on</strong>g>of</str<strong>on</strong>g> 2 µg/ml<br />

Hsp70 maintained cellular viability measured by MTS assay compared to c<strong>on</strong>trol populati<strong>on</strong>s. Erythrocytes<br />

required 3 hr <str<strong>on</strong>g>of</str<strong>on</strong>g> 10µg/ml Hsp70 to protect against cell lysis induced by heat shock at 42°C, treatment with<br />

BSA showed no effect. These data support a possible role <str<strong>on</strong>g>for</str<strong>on</strong>g> extra-cellular Hsp70 in the protecti<strong>on</strong> against<br />

heat-induced cell death. Further characterisati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> this resp<strong>on</strong>se will be determined by flow cytometry and<br />

fluorescence microscopy.<br />

1D_02_P<br />

(poster secti<strong>on</strong> A1, poster board #39, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE ROLE OF CLUSTERIN IN EXTRACELLULAR PROTEIN QUALITY CONTROL<br />

A. R. Wyatt 1 , M. R. Wils<strong>on</strong> 1 , A. Katsifis 2 , I. Greguric 2 , F. Mattner 2 , P. Bergh<str<strong>on</strong>g>of</str<strong>on</strong>g>er 2<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biological Sciences, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Woll<strong>on</strong>g<strong>on</strong>g, Northfields Avenue, Woll<strong>on</strong>g<strong>on</strong>g, NSW, Australia<br />

2Radiopharmaceuticals <str<strong>on</strong>g>Research</str<strong>on</strong>g>, The <str<strong>on</strong>g>Australian</str<strong>on</strong>g> Nuclear Science and Technology Organisati<strong>on</strong> (ANSTO),<br />

Lucas Heights, NSW, Australia<br />

Processes to attain and maintaining the correct three-dimensi<strong>on</strong>al shape, or ‘native c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>’ <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins<br />

are vital. However, certain c<strong>on</strong>diti<strong>on</strong>s including thermal and oxidative stress cause proteins to unfold and<br />

aggregate. Intracellular and/or extracellular protein aggregates have been identified in a large number <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

diseases, including Alzheimer’s disease, arthritis and type II diabetes. While several intracellular quality c<strong>on</strong>trol<br />

mechanisms <str<strong>on</strong>g>for</str<strong>on</strong>g> the folding state <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins have been characterized, corresp<strong>on</strong>ding mechanisms <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

extracellular protein quality c<strong>on</strong>trol have yet to be identified. Clusterin is an extracellular chaper<strong>on</strong>e that can<br />

stabilise proteins and prevents their precipitati<strong>on</strong> during exposure to high temperatures or oxidative stress.<br />

We have dem<strong>on</strong>strated that clusterin stabilizes proteins by <str<strong>on</strong>g>for</str<strong>on</strong>g>ming high molecular weight (HMW) complexes<br />

(> 4 x 10 7 Da) with them. Using an animal model, the fate <str<strong>on</strong>g>of</str<strong>on</strong>g> blood-borne 123 I -HMW complexes was<br />

investigated. 123 I -HMW complexes were rapidly cleared from circulati<strong>on</strong> and targeted primarily to the liver<br />

and spleen. In the absence <str<strong>on</strong>g>of</str<strong>on</strong>g> specific extracellular proteolytic mechanisms it appears likely that extracellular<br />

proteins are targeted and transported intracellularly <str<strong>on</strong>g>for</str<strong>on</strong>g> degradati<strong>on</strong>. Receptor-mediated endocytosis is <strong>on</strong>e<br />

possible route <str<strong>on</strong>g>for</str<strong>on</strong>g> the clearance <str<strong>on</strong>g>of</str<strong>on</strong>g> HMW complexes. Clusterin affinity chromatography <str<strong>on</strong>g>of</str<strong>on</strong>g> cell membrane<br />

53


23-26 August 2007,<br />

Budapest, Hungary<br />

proteins from liver and spleen may identify potential receptors <str<strong>on</strong>g>for</str<strong>on</strong>g> HMW complexes. These findings suggest<br />

an important role <str<strong>on</strong>g>for</str<strong>on</strong>g> clusterin in extracellular protein quality c<strong>on</strong>trol.<br />

54<br />

1D_03_P<br />

(poster secti<strong>on</strong> A1, poster board #40, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE EXTRACELLULAR PROTEASE INHIBITOR ALPHA 2 –MACROGLOBULIN<br />

HAS CHAPERONE-LIKE PROPERTIES<br />

Justin J. Yerbury, Katie French, Mark R. Wils<strong>on</strong><br />

School <str<strong>on</strong>g>of</str<strong>on</strong>g> Biological Sciences, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Woll<strong>on</strong>g<strong>on</strong>g, Northfields Avenue, Woll<strong>on</strong>g<strong>on</strong>g, NSW 2522, Australia. e-<br />

mail: jyerbury@uow.edu.au<br />

α 2 -Macroglobulin (α 2 M) is a secreted glycoprotein, found at high levels in human blood, best known <str<strong>on</strong>g>for</str<strong>on</strong>g> its<br />

ability to inhibit a large array <str<strong>on</strong>g>of</str<strong>on</strong>g> proteases by a unique trapping method. Protease binding promotes a change<br />

in structure <str<strong>on</strong>g>of</str<strong>on</strong>g> α 2 M yielding an “activated” c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> which exposes a binding site <str<strong>on</strong>g>for</str<strong>on</strong>g> the low density<br />

lipoprotein receptor. This process facilitates the clearance <str<strong>on</strong>g>of</str<strong>on</strong>g> α 2 M-protease complexes from the body. We<br />

report here that α 2 M also has potent chaper<strong>on</strong>e properties, making it the first known protein to combine<br />

both these activities. We show that α 2 M inhibits the heat induced aggregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> citrate synthase and creatine<br />

phosphokinase, and the oxidative stress induced aggregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> lysozyme and α-synuclein. Protease-mediated<br />

activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> α 2 M abolishes its chaper<strong>on</strong>e-like activity. However, we show that native α 2 M is able to <str<strong>on</strong>g>for</str<strong>on</strong>g>m<br />

soluble complexes with stressed proteins and then subsequently become activated by interacting with a<br />

protease. This activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> α 2 M in complex with misfolded proteins provides a potential mechanism <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

in vivo clearance <str<strong>on</strong>g>of</str<strong>on</strong>g> α 2 M/stressed protein/ protease complexes. We propose that α 2 M is a member <str<strong>on</strong>g>of</str<strong>on</strong>g> a small<br />

group <str<strong>on</strong>g>of</str<strong>on</strong>g> abundant extracellular proteins with chaper<strong>on</strong>e properties that patrol extracellular spaces <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

unfolded/misfolded proteins and facilitate their disposal.<br />

1D_04_P<br />

(poster secti<strong>on</strong> A1, poster board #41, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

BACTERIAL BINDING AND OPSONIZING EFFECT OF EXTRACELLULAR HSP70<br />

Minh Tu Nguyen, Tibor Károly Fábián, Mahavir Singh, Péter Csermely, Csaba Sőti<br />

Semmelweis University Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Chemistry, Budapest, P.O.Box 260; H-1444 Hungary<br />

e-mail: minhtu@freemail.hu<br />

Stress (or heat shock) proteins <str<strong>on</strong>g>for</str<strong>on</strong>g>m an ancient defense system <str<strong>on</strong>g>of</str<strong>on</strong>g> our cells. They were c<strong>on</strong>sidered to be<br />

intracellular, but cells under stressful c<strong>on</strong>diti<strong>on</strong>s can express stress proteins <strong>on</strong> their surface and release them<br />

to the extracellular space, where these proteins funti<strong>on</strong> as a „danger signal” and induce apoptosis <str<strong>on</strong>g>of</str<strong>on</strong>g> target<br />

cells as well as activate both humoral and cellular immune resp<strong>on</strong>ses. One <str<strong>on</strong>g>of</str<strong>on</strong>g> the most important stress<br />

protein implicated in extracellular signalling is Hsp70, the inducible cytosolic is<str<strong>on</strong>g>of</str<strong>on</strong>g>orm <str<strong>on</strong>g>of</str<strong>on</strong>g> the 70 kDa heat<br />

shock protein family. In the present experiments we investigated the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> extracellular Hsp70 <strong>on</strong> the<br />

interacti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> bacteria with the immun system.Our results dem<strong>on</strong>strated that extracellular Hsp70 was able to<br />

bind both to the surface <str<strong>on</strong>g>of</str<strong>on</strong>g> Gram-positive Streptococcus mutans and mitis and Gram-negative Escherichia<br />

coli bacteria. Acidic milieu (pH 5.5) and high temperature (42°C) facilitated while ATP partially inhibited the<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the Hsp70-bacterium complex. We found that extracellular Hsp70 exerted an ops<strong>on</strong>izing effect,


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

it was able to activate the killing activity <str<strong>on</strong>g>of</str<strong>on</strong>g> polymorph<strong>on</strong>uclear neutrophil leukocytes. Our results provide a<br />

novel mechanism by which extracellular Hsp70 may induce the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the innate immune resp<strong>on</strong>se.<br />

These findings may lead to the deeper understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> the relati<strong>on</strong>ship between stress and immunity, as well<br />

as may promote the development <str<strong>on</strong>g>of</str<strong>on</strong>g> immune stimulatory drugs.<br />

1D_05_P<br />

(poster secti<strong>on</strong> A1, poster board #42, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EXTRACELLULAR HSP70-INDUCED SIGNALING EVENTS IN A431 CARCINOMA<br />

CELLS<br />

Ant<strong>on</strong> L. Evd<strong>on</strong>in, Natalia D. Medvedeva<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Cytology, RAS, S-Petersburg, Russia, e-mail: evd<strong>on</strong>in@mail.ru<br />

Recent studies dem<strong>on</strong>strated that Hsp70 is being released into blood or the c<strong>on</strong>diti<strong>on</strong>ed medium <str<strong>on</strong>g>of</str<strong>on</strong>g> cultured<br />

cells under stressful c<strong>on</strong>diti<strong>on</strong>s. Exogenously added purified Hsp70 was shown to induce signal transducti<strong>on</strong><br />

in m<strong>on</strong>ocytes and macrophages. However, whether Hsp70 can induce signal transducti<strong>on</strong> events in n<strong>on</strong>immune<br />

cells remains unclear. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to investigate the signaling events triggered by<br />

extracellular Hsp70 in carcinoma cells. At initial steps <str<strong>on</strong>g>of</str<strong>on</strong>g> heat stress Hsp70 was shown to appear in the<br />

c<strong>on</strong>diti<strong>on</strong>ed medium <str<strong>on</strong>g>of</str<strong>on</strong>g> heated A431 carcinoma cells due to its secreti<strong>on</strong>. Both heat shock and extracellular<br />

Hsp70 induce associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> TLR2/4 with downstream adaptor protein MyD88 which c<strong>on</strong>firms the<br />

activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> TLR2/4. Simultaneously, heat shock, extracellular Hsp70 and c<strong>on</strong>diti<strong>on</strong>ed medium from heated<br />

A431 cells stimulate ligand-independent EGFR activati<strong>on</strong> and activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> EGFR-dependent signaling<br />

pathways including Erk1/2, PLCγ1 and STAT3. The depleti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 from such medium abolished<br />

EGFR transactivati<strong>on</strong>. The neutralizing antibody to TLR2 and 4 attenuated both EGFR and ERK1/2<br />

phosphorylati<strong>on</strong> which c<strong>on</strong>firms the cross talk between TLRs and EGFR signaling systems in A431 cells.<br />

Moreover, extracellular Hsp70 and heat shock induce associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> TLR2 and 4 with EGFR. While the<br />

protective functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> intracellular Hsp70 is well documented, the present study provide evidence that Hsp70<br />

secreted from cells at initial steps <str<strong>on</strong>g>of</str<strong>on</strong>g> heat stress might play the same functi<strong>on</strong> by mediating EGFR<br />

transactivati<strong>on</strong>, which is known to prevent stress-induced apoptosis.<br />

1D_06_P<br />

(poster secti<strong>on</strong> A1, poster board #43, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HDJ2 IN HUMAN SYNOVIAL FLUIDS<br />

Claudia Lehr, Rita Rzepka, Christoph Schick, Andrea Zgaga-Griesz, Inga Melchers<br />

Clinical <str<strong>on</strong>g>Research</str<strong>on</strong>g> Unit <str<strong>on</strong>g>for</str<strong>on</strong>g> Rheumatology, University Medical Center, 79106 Freiburg, Germany,<br />

e-mail: inga.melchers@uniklinik-freiburg.de<br />

In healthy human joints a thin layer <str<strong>on</strong>g>of</str<strong>on</strong>g> synovial fluid (SF) fills the joint space and covers the surface <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

synovial membrane and cartilage. During various joint diseases, mainly arthritis, the fluid volume increases.<br />

The resulting joint effusi<strong>on</strong> is clinically apparent and can be aspirated <str<strong>on</strong>g>for</str<strong>on</strong>g> therapy and analyses. The J-protein<br />

Hdj2 (DNAJA1, Hsj2, Hsdj, dj-2) is <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> ~40 known human Hsp40 chaper<strong>on</strong>s. Its highly c<strong>on</strong>served J-<br />

domain shows great homology to E. coli DnaJ and cooperates with HSP70 members. Earlier we described an<br />

enhanced expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsc70 (but not <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70) and several J-proteins in synovial tissue (ST) from patients<br />

with RA, but not in ST from patients with osteoarthritis (OA) (Kurzik-Dumke et al. Arthr Rheum. 1999,<br />

42:210-220; Schick et al. Arthr Rheum 2004, 50:88-93). We now show that also Hdj2 is overexpressed in ST<br />

55


23-26 August 2007,<br />

Budapest, Hungary<br />

from patients with RA, but not from patients with OA. In additi<strong>on</strong>, we collected SF from 136 patients with<br />

joint effusi<strong>on</strong>s: 51 patients with rheumatoid arthritis (RA) and 85 patients with reactive arthritis, psoriasis<br />

arthritis or another disease. These SFs were tested <str<strong>on</strong>g>for</str<strong>on</strong>g> the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> Hdj2 using a specific m<strong>on</strong>ocl<strong>on</strong>al<br />

antibody (KA2A5.6) in immunoblots. We detected Hdj2 in 62.5 % <str<strong>on</strong>g>of</str<strong>on</strong>g> all SFs, in different amounts. SFs <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

RA patients c<strong>on</strong>tained Hdj2 more frequently than SFs from patients with other diseases (72.6 % vs. 56.6 %),<br />

but the amounts <str<strong>on</strong>g>of</str<strong>on</strong>g> Hdj2 within positive SFs <str<strong>on</strong>g>of</str<strong>on</strong>g> both groups did not differ significantly from each other.<br />

Clinical and laboratory parameters (e. g. sex, age, durati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> disease, markers <str<strong>on</strong>g>of</str<strong>on</strong>g> inflammati<strong>on</strong>,<br />

autoantibodies, etc.) are collected and compared with the presence and quantity <str<strong>on</strong>g>of</str<strong>on</strong>g> extracellular Hdj2.<br />

56


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

1E. OTHER CHAPERONE RELATES TOPICS (HSP60, HSP90, COCHAPERONES,<br />

HSP CRYSTALLOGRAPHY AND STRUCTURES, STRESS OF SINGLE MOLECULES)<br />

1E_01_P<br />

(poster secti<strong>on</strong> A1, poster board #44, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE INTERACTION OF HELICAL PROTRUSIONS IS IMPORTANT IN THE<br />

PROTEIN FOLDING CYCLE OF GROUP II CHAPERONINS<br />

Taro Kanzaki 1 , Ryo Iizuka 2 , Kazunobu Takahashi 3 , Kosuke Maki 3 , Noriyuki Ishii 4 ,<br />

Muhamad Sahlan 1 , Masahiro Furutani 5 , Kunihiro Kuwajima 3 , Masafumi Yohda 1<br />

1Tokyo Univ. <str<strong>on</strong>g>of</str<strong>on</strong>g> Agri. & Technol., Tokyo;<br />

2Univ. <str<strong>on</strong>g>of</str<strong>on</strong>g> Tokyo, Tokyo;<br />

3Inst. <str<strong>on</strong>g>for</str<strong>on</strong>g> Mol. Sci., Aichi;<br />

4Adv. Ind. Sci. & Technol., Ibaraki;<br />

5Minase Res. Lab., Sekisui Chem. Co. Ltd, Osaka, Japan<br />

e-mail: kanzaki@bel.bio.tuat.ac.jp<br />

Group II chaper<strong>on</strong>ins, found in the archaeal and eukaryotic cytosol, functi<strong>on</strong> independently <str<strong>on</strong>g>of</str<strong>on</strong>g> the c<str<strong>on</strong>g>of</str<strong>on</strong>g>actor<br />

corresp<strong>on</strong>ding to GroES <str<strong>on</strong>g>of</str<strong>on</strong>g> group I chaper<strong>on</strong>ins. Rather, the helical protrusi<strong>on</strong> at the tip <str<strong>on</strong>g>of</str<strong>on</strong>g> the apical<br />

domain, which <str<strong>on</strong>g>for</str<strong>on</strong>g>ms a built-in lid <str<strong>on</strong>g>of</str<strong>on</strong>g> the central cavity, substitutes <str<strong>on</strong>g>for</str<strong>on</strong>g> the c<str<strong>on</strong>g>of</str<strong>on</strong>g>actor. We have been studying<br />

the protein folding mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> group II chaper<strong>on</strong>ins using the chaper<strong>on</strong>in from a hyperthermophilic<br />

archaeum, Themococcus sp. strain KS-1. Previous studies have shown that ATP drives the c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>al<br />

change <str<strong>on</strong>g>of</str<strong>on</strong>g> group II chaper<strong>on</strong>ins, from the open-lid, substrate binding c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>, to the close-lid<br />

c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> to encapsulate unfolded protein in the central cavity. However, the details <str<strong>on</strong>g>of</str<strong>on</strong>g> the ATP-driven<br />

reacti<strong>on</strong> remain obscure. It has been c<strong>on</strong>sidered that the c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>al change is driven in the cooperative<br />

manner. However, the mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> the cooperati<strong>on</strong> between subunits has not been studied. To elucidate<br />

this issue, we c<strong>on</strong>structed and characterized the chaper<strong>on</strong>in complexes which are composed <str<strong>on</strong>g>of</str<strong>on</strong>g> wild and<br />

mutated subunits in the ordered fashi<strong>on</strong> by c<strong>on</strong>necting them. Although the complexes c<strong>on</strong>taining mutati<strong>on</strong>s<br />

around ATP binding sites retained protein folding activity, the complex with the helical protrusi<strong>on</strong> deleti<strong>on</strong><br />

had lost ATP dependent c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>al change and also protein folding activity. Thus, we c<strong>on</strong>cluded that<br />

not cooperative acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ATP hydrolysis but the interacti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> helical protrusi<strong>on</strong>s in the ring is important in<br />

the protein folding cycle <str<strong>on</strong>g>of</str<strong>on</strong>g> group II chaper<strong>on</strong>ins.<br />

1E_02_P<br />

(poster secti<strong>on</strong> A1, poster board #45, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

GENE STRUCTURE AND SPATIAL DISTRIBUTION OF HSP60 CHAPERONIN OF<br />

LUCILIA CUPRINA: IMPLICATIONS IN GERM CELL DIFFERENTIATION AND<br />

NUCLEAR FUNCTIONS<br />

P. V. J. Reddy, Sunita Sharma, P. K. Tiwari<br />

School <str<strong>on</strong>g>of</str<strong>on</strong>g> Studies in Zoology, Jiwaji University, Gwalior – 474 011. India<br />

e-mail: pk_tiwari@hotmail.com<br />

The HSP60 chaper<strong>on</strong>in is widely appreciated <str<strong>on</strong>g>for</str<strong>on</strong>g> its role in protein folding. However, the detailed<br />

in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <strong>on</strong> its cellular functi<strong>on</strong>s still remained elusive. The present study is a follow-up <str<strong>on</strong>g>of</str<strong>on</strong>g> our studies <strong>on</strong><br />

57


23-26 August 2007,<br />

Budapest, Hungary<br />

the functi<strong>on</strong>al characterizati<strong>on</strong> and significance <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP60 protein in sheep blowfly, Lucilia cuprina, which<br />

revealed interesting results. The immunocytochemical and immun<str<strong>on</strong>g>of</str<strong>on</strong>g>lourescence studies showed the<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Lucilia HSP60 protein to be generally c<strong>on</strong>stitutive, whose level rises up<strong>on</strong> heat stress. In a few<br />

cell types, it appears tissue- or developmental stage-specific and even heat inducible. More striking is its<br />

presence in several nuclei <str<strong>on</strong>g>of</str<strong>on</strong>g> larval and adult polytene cell types while gene or protein analysis revealed an<br />

apparent absence <str<strong>on</strong>g>of</str<strong>on</strong>g> NLS, str<strong>on</strong>gly suggesting its involvement in both cytoplasmic and nuclear functi<strong>on</strong>s. The<br />

expressi<strong>on</strong> pattern in germline tissues is more interesting, particularly in ovarian tissues. The oocytes showed<br />

a stage-specific expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the protein, varying from 1st – 14th stages. The spatial pattern <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP60<br />

expressi<strong>on</strong> in mature eggs indicates a distinct role <str<strong>on</strong>g>for</str<strong>on</strong>g> it. The complete L. cuprina hsp60 gene was cl<strong>on</strong>ed<br />

using genomic PCR strategy and RACE. The major porti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the coding sequence <str<strong>on</strong>g>of</str<strong>on</strong>g> the hsp60 gene was<br />

isolated using the primers designed from Drosophila melanogaster cDNA sequence. The sequence analysis<br />

revealed a 2190 bps l<strong>on</strong>g sequence with a 1730 bps coding regi<strong>on</strong> and 5’ and 3’ UTRs <str<strong>on</strong>g>of</str<strong>on</strong>g> 189 and 271 bases<br />

respectively. Sequence alignment <str<strong>on</strong>g>of</str<strong>on</strong>g> nucleotide and amino acid sequences c<strong>on</strong>firmed a significant homology<br />

am<strong>on</strong>g divergent species. The in-silico analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> the protein structure revealed significant structural<br />

homology with GroEL. The real time PCR analysis showed that the copy number and expressi<strong>on</strong> levels <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

hsp60 gene are almost ten fold lesser than its compatriot Drosophila. RNA-RNA in-situ hybridizati<strong>on</strong>s<br />

showed transcript expressi<strong>on</strong> to be more in differentiating cell types than the differentiated cell types, further<br />

pointing its functi<strong>on</strong>al significance to be differential during various life cycle stages. Details <str<strong>on</strong>g>of</str<strong>on</strong>g> the possible<br />

mechanism/s <str<strong>on</strong>g>of</str<strong>on</strong>g> the nuclear localisati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP60 and its functi<strong>on</strong>al significance in cell nuclei and in<br />

differentiating (germ) cells will be discussed.<br />

1E_03_P<br />

(poster secti<strong>on</strong> A1, poster board #46, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HSP90 IN C. ELEGANS AND BRUGIA: HIGHLY CONSERVED YET DIFFERENT<br />

Eileen Devaney, Vicki Gillan, Kirsty Maitland, Nik A. I. I. Nik Him<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Glasgow, Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Veterinary Infecti<strong>on</strong> and Immunity, G61 1QH, Bearsden Road, Glasgow, UK<br />

e-mail: e.devaney@vet.gla.ac.uk<br />

The molecular architecture <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90 is highly c<strong>on</strong>served throughout evoluti<strong>on</strong>. However, C. elegans Hsp90<br />

is unique am<strong>on</strong>gst eukaryotes because <str<strong>on</strong>g>of</str<strong>on</strong>g> its resistance to Geldanamcyin. This finding was originally<br />

interpreted as an example <str<strong>on</strong>g>of</str<strong>on</strong>g> adaptive evoluti<strong>on</strong> as both C. elegans and the micro-organism that synthesizes<br />

GA live in the same ecological niche <str<strong>on</strong>g>of</str<strong>on</strong>g> the soil. In c<strong>on</strong>trast Hsp90 from the parasitic nematode Brugia<br />

pahangi specifically binds to GA. Using a comparative genomics approach we have investigated whether the<br />

GA-resistant phenotype <str<strong>on</strong>g>of</str<strong>on</strong>g> C. elegans is unique or is shared with other nematodes. From our analysis to date<br />

additi<strong>on</strong>al nematodes that express either a GA-resistant or a GA-sensitive Hsp90 have been identified and we<br />

are using this in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> in an attempt to dissect the molecular basis <str<strong>on</strong>g>of</str<strong>on</strong>g> GA resistance in C. elegans. In<br />

additi<strong>on</strong> we are expressing the GA sensitive Bp-Hsp90 in C. elegans to determine whether this c<strong>on</strong>fers GA<br />

sensitivity <strong>on</strong> the free-living species. Our results suggest that additi<strong>on</strong>al factors bey<strong>on</strong>d the primary amino<br />

acid sequence and predicted structure determine the GA sensitivity <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90.<br />

58


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

1E_04_P<br />

(poster secti<strong>on</strong> A1, poster board #47, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

AMINO ACID SUBSTITUTIONS OBSERVED IN HUMAN HSP90 ISOFORMS THAT<br />

IMPEDE THE DIMERIC INTERACTION<br />

Nemoto Takayuki<br />

Nagasaki University, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Oral Molecular Biology, 852-8588, 1-7-1 Sakamoto, Nagasaki, Japan<br />

e-mail: tnemoto@nagasaki-u.ac.jp<br />

It has been proposed that Hsp90 captures a client protein like a molecular clamp. The dimeric structure <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Hsp90 mediated by the C-terminal regi<strong>on</strong> appears to be essential as a joint <str<strong>on</strong>g>of</str<strong>on</strong>g> the clamp. We here examined<br />

the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> the amino acid substituti<strong>on</strong>s present between human Hsp90 is<str<strong>on</strong>g>of</str<strong>on</strong>g>orms, i.e., Hsp90α and<br />

Hsp90β, and a single nucleotide polymorphism (SNP) found in Caucasians <strong>on</strong> the dimeric interacti<strong>on</strong>. The<br />

dimer-<str<strong>on</strong>g>for</str<strong>on</strong>g>ming potential <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90β is less than that <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90α due to the 16 amino acid substituti<strong>on</strong>s at the<br />

561-685 amino-acid regi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the C-terminus. Bacterial two-hybrid system dem<strong>on</strong>strated, am<strong>on</strong>g the 16<br />

amino acids, the c<strong>on</strong>versi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> Thr566 and Ala629 <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90α to Ala558 and Met621 were primarily<br />

resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> impeded dimerizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90β Yeast expressi<strong>on</strong> system revealed that the SNP<br />

(Gln488>His) <str<strong>on</strong>g>of</str<strong>on</strong>g> the Hsp90α observed in Caucasians disrupts the functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90MacLean, M.J., et al.,<br />

2006). To investigate its molecular mechanism, we evaluated the domain-domain interacti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90α with<br />

the missense mutati<strong>on</strong>. Human Hsp90α 401-732 interacts with both Hsp90α 1-400 and 401-732 through the<br />

intra-molecular and dimeric interacti<strong>on</strong>s, respectively. The two-hybrid system revealed that the mutati<strong>on</strong><br />

selectively decreased the latter interacti<strong>on</strong>. The present study defined the amino acid substituti<strong>on</strong>s affecting<br />

the dimerizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90 and dem<strong>on</strong>strated that the dimeric interacti<strong>on</strong> is prerequisite <str<strong>on</strong>g>for</str<strong>on</strong>g> the in vivo<br />

functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90.<br />

1E_06_P<br />

(poster secti<strong>on</strong> A1, poster board #48, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

KNOCKDOWN OF THE HSP90 CO-CHAPERONE CDC37 CHEMOSENSITIZES AND<br />

ARRESTS THE GROWTH OF PROSTATE CANCER CELLS BY INHIBITING<br />

MULTIPLE SIGNALING CASCADES<br />

Phillip J. Gray, Jr., Mary Ann Stevens<strong>on</strong>, Stuart K. Calderwood<br />

Beth Israel Deac<strong>on</strong>ess Medical Center, Harvard Medical School, Bost<strong>on</strong>, MA 02215<br />

e-mail: pjgray@bidmc.harvard.edu<br />

In the fight against prostate cancer, the sec<strong>on</strong>d leading cause <str<strong>on</strong>g>of</str<strong>on</strong>g> cancer death in men, delineati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

complex molecular interacti<strong>on</strong>s within the prostate cancer cell will be critical to improving outcomes <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

patients. The HSP90 chaper<strong>on</strong>e system is already known to interact with and maintain a wide variety <str<strong>on</strong>g>of</str<strong>on</strong>g> cell<br />

signaling pathways comm<strong>on</strong>ly dysregulated in cancer. An important part <str<strong>on</strong>g>of</str<strong>on</strong>g> this system is the kinase and<br />

steroid receptor targeting subunit Cdc37. Using RNA interference by lentivirus-delivered shRNA c<strong>on</strong>structs<br />

we were able to determine the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> Cdc37 knockdown <strong>on</strong> prostate cancer cells. Our data indicate that<br />

loss <str<strong>on</strong>g>of</str<strong>on</strong>g> Cdc37 functi<strong>on</strong> induces growth arrest and sensitizes prostate cancer cells to chemotherapeutic drugs<br />

such as Paclitaxel. Although loss <str<strong>on</strong>g>of</str<strong>on</strong>g> Cdc37 functi<strong>on</strong> does not significantly alter the intracellular levels <str<strong>on</strong>g>of</str<strong>on</strong>g> most<br />

known HSP90/Cdc37 clients, it does markedly inhibit their activity and ability to functi<strong>on</strong> in signal<br />

59


23-26 August 2007,<br />

Budapest, Hungary<br />

transducti<strong>on</strong>. We show that loss <str<strong>on</strong>g>of</str<strong>on</strong>g> Cdc37 leads to reduced flux through multiple signaling pathways<br />

including the Erk pathway, the Akt pathway, the mTOR pathway and the androgen receptor pathway. We<br />

have also discovered synergistic interacti<strong>on</strong> between the knockdown <str<strong>on</strong>g>of</str<strong>on</strong>g> Cdc37 and the inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP90<br />

by the anticancer drug 17AAG. Thus Cdc37 is important <str<strong>on</strong>g>for</str<strong>on</strong>g> maintaining the tumor phenotype in cancer cells<br />

and represents a novel target in the search <str<strong>on</strong>g>for</str<strong>on</strong>g> multi-targeted therapies based <strong>on</strong> the HSP90 chaper<strong>on</strong>e<br />

system.<br />

1E_07_P<br />

(poster secti<strong>on</strong> A1, poster board #49, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

INTERACTION OF PREFOLDIN WITH GROUP II CHAPERONIN IN THE<br />

PRESENCE OF VARIOUS NUCLEOTIDES<br />

Tamotsu Zako 1,* , Yosuke Murase 1, 2 , Andrei Zhukov 3 , Robert Karlss<strong>on</strong> 3 , Masafumi Shimizu 4 ,<br />

Ryo Iizuka 2 , Mizuo Maeda 1 , Masafumi Yohda 2<br />

1RIKEN<br />

2Tokyo Univ. <str<strong>on</strong>g>of</str<strong>on</strong>g> Agri. and Tech,<br />

3BIAcore AB,<br />

4Tokyo Univ. <str<strong>on</strong>g>of</str<strong>on</strong>g> Tech., Bioengineering Lab. RIKEN, 2-1 Hirosawa, Wako, Saitama 351-0198 Japan,<br />

e-mail: zako@riken.jp<br />

Prefoldin is a molecular chaper<strong>on</strong>e that captures a protein-folding intermediate and transfers it to a group II<br />

chaper<strong>on</strong>in <str<strong>on</strong>g>for</str<strong>on</strong>g> correct folding. Previously we have shown that the binding rate <str<strong>on</strong>g>of</str<strong>on</strong>g> prefoldin to chaper<strong>on</strong>in<br />

correlates with the transfer rate <str<strong>on</strong>g>of</str<strong>on</strong>g> a substrate between them and electrostatic interacti<strong>on</strong> is important <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

their interacti<strong>on</strong>. However, the manner in which the prefoldin interacts with its substrates and cooperates<br />

with the chaper<strong>on</strong>in is poorly understood. In this study, we examined affinities <str<strong>on</strong>g>of</str<strong>on</strong>g> Pyrococcus prefoldin<br />

(PhPFD) and a chaper<strong>on</strong>in mutant from Thermococcus sp. strain KS-1 (CPNαK250E/K256E) in the<br />

presence <str<strong>on</strong>g>of</str<strong>on</strong>g> various nucleotides using fluorescent anisotropy and SPR sensor. CPNαK250E/K256E takes an<br />

open c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> in the nucleotide-free or ADP bound states and changes to a closed c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> up<strong>on</strong><br />

binding <str<strong>on</strong>g>of</str<strong>on</strong>g> ATP. Our results indicate that PhPFD binds more tightly to CPNαK250E/K256E in the<br />

nucleotide-free state or in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> ADP than in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> AMP-PNP, unhydrolyzable ATP<br />

analogue. These results agree well with the substrate protein transfer model suggesting the prefoldin binds to<br />

the open-state chaper<strong>on</strong>in. The kinetic experiments using BIAcore T100 have shown that PhPFD-<br />

CPNαK250E/K256E interacti<strong>on</strong> involves two steps. The fast encounter step is followed by a slower docking<br />

step whereby the complex undergoes a slow c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>al trans<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>.<br />

60


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

1E_08_P<br />

(poster secti<strong>on</strong> A1, poster board #50, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CHARACTERIZATION OF THE ACTIVATION MECHANISM OF HTRA PROTEASE<br />

FROM ESCHERICHIA COLI<br />

Anna Sobiecka-Szkatula 1 *, Joanna Skorko-Gl<strong>on</strong>ek 1 , Andrea Scire 2 , Fabio Tanfani 2 ,<br />

Barbara Lipinska 1<br />

1 University <str<strong>on</strong>g>of</str<strong>on</strong>g> Gdansk, Gdansk, Poland<br />

2 Universita Politecnica delle Marche, Anc<strong>on</strong>a, Italy<br />

e-mail: sobiecka@biotech.ug.gda.pl<br />

HtrA from E. coli is a periplasmic serine protease resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> degradati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> misfolded proteins. The<br />

proteolytic activity <str<strong>on</strong>g>of</str<strong>on</strong>g> HtrA is regulated in a temperature-dependent manner. According to the HtrA’s crystal<br />

structure below 30°C the access to the active center is restricted by three interacting loops: LA, L1 and L2. At<br />

higher temperatures the protease is activated and this process most probably requires the disrupti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

loop trio. The way this process occurs is still not clear. To examine the importance <str<strong>on</strong>g>of</str<strong>on</strong>g> the LA-L1-L2<br />

interacti<strong>on</strong> <strong>on</strong> HtrA stability we introduced a tryptophan residue into each <str<strong>on</strong>g>of</str<strong>on</strong>g> the loops. As the wt HtrA does<br />

not c<strong>on</strong>tain tryptophan, the introduced Trp served as internal probe <str<strong>on</strong>g>for</str<strong>on</strong>g> studying c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>al changes<br />

within the loops. Using Fourier Trans<str<strong>on</strong>g>for</str<strong>on</strong>g>m-Infrared Spectroscopy we c<strong>on</strong>firmed that the mutati<strong>on</strong>s did not<br />

affect the sec<strong>on</strong>dary structure or the thermal stability <str<strong>on</strong>g>of</str<strong>on</strong>g> the mutants; thus the proteins were suitable <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

further analysis. The c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>al changes induced by temperature were studied by circular dichroism<br />

(CD) technique at wavelengths 260-320 nm. We m<strong>on</strong>itored the CD spectra at temperature range 25- 50oC<br />

and focused particularly <strong>on</strong> the spectrum <str<strong>on</strong>g>of</str<strong>on</strong>g> tryptophan (wavelength range 280 –300 nm). We observed that<br />

significant changes in the CD signal intensity occurred at temperature ranges 30-32,5°C and 37–45°C. The<br />

observed changes correlate well with the changes <str<strong>on</strong>g>of</str<strong>on</strong>g> the HtrA proteolytic activity: at 30°C HtrA becomes<br />

proteolytically active and at temperature above 40°C the activity significantly rises to reach maximum at 50-<br />

55°C. The obtained results were verified further by spectr<str<strong>on</strong>g>of</str<strong>on</strong>g>luorimetric methods.<br />

1E_09_P<br />

(poster secti<strong>on</strong> A1, poster board #51, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ANTI-KU-POSITIVE AUTOIMMUNE DISEASES: POTENTIAL FUNGAL<br />

TRIGGERING<br />

F. Guarneri, M. Vaccaro, C. Guarneri<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Dermatology - University <str<strong>on</strong>g>of</str<strong>on</strong>g> Messina (Italy)<br />

AOU Policlinico “G. Martino” - Via C<strong>on</strong>solare Valeria - Gazzi – Messina (Italy)<br />

e-mail: f.guarneri@tiscali.it<br />

Molecular mimicry (MM) has been postulated as a trigger <str<strong>on</strong>g>of</str<strong>on</strong>g> autoimmune diseases (AID). Ku is a heterodimer<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> 70 kDa (p70) and 80 kDa (p80) protein subunits and has a key role in multiple processes, particularly in<br />

stress c<strong>on</strong>diti<strong>on</strong>s: DNA repair, chromosome maintenance, transcripti<strong>on</strong> regulati<strong>on</strong>, V(D)J recombinati<strong>on</strong>. Ku<br />

is also part <str<strong>on</strong>g>of</str<strong>on</strong>g> a group <str<strong>on</strong>g>of</str<strong>on</strong>g> DNA-associated antigens targeted by autoantibodies in systemic lupus<br />

erythematosus and related disorders. It is found in 5% <str<strong>on</strong>g>of</str<strong>on</strong>g> sclerodermic patients, and is suggestive <str<strong>on</strong>g>of</str<strong>on</strong>g> systemic<br />

sclerosis/polymyositis overlap syndrome. Since a possible role <str<strong>on</strong>g>of</str<strong>on</strong>g> MM in the generati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> autoimmunity to<br />

this antigen has been suggested, we used in silico techniques to identify potentially resp<strong>on</strong>sible microbial<br />

61


23-26 August 2007,<br />

Budapest, Hungary<br />

proteins. We used BLAST to compare amino acid sequences <str<strong>on</strong>g>of</str<strong>on</strong>g> p70 and p80, with all known sequences (as <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

April 18, 2007) <str<strong>on</strong>g>of</str<strong>on</strong>g> bacteria (n=5,229,868), viruses (n=629,582) and fungi (n=511,126). The cut-<str<strong>on</strong>g>of</str<strong>on</strong>g>f <str<strong>on</strong>g>for</str<strong>on</strong>g> the E<br />

parameter was 0.001. We found no homologies with bacterial or viral proteins. Homologous fungal proteins<br />

were 51 <str<strong>on</strong>g>for</str<strong>on</strong>g> p70 and 24 <str<strong>on</strong>g>for</str<strong>on</strong>g> p80. In both cases, <strong>on</strong>ly 12 bel<strong>on</strong>ged to human pathogens (A. clavatus, A.<br />

fumigatus, A. nidulans, A. niger, A. terreus, C. glabrata, C. globosum, C. immitis, C. ne<str<strong>on</strong>g>of</str<strong>on</strong>g>ormans, N. fischeri,<br />

Y. lipolytica, and D. hansenii <str<strong>on</strong>g>for</str<strong>on</strong>g> p70 and C. albicans <str<strong>on</strong>g>for</str<strong>on</strong>g> p80). Amino acid identity range was 20-30% and 20-<br />

27%, similarity range was 39-50% and 40-48%, E values were between 2x10-66 and 3x10-4 and between<br />

2x10-35 and 3x10-4, respectively. p70-homologous segments spanned at least <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the 3 T-cell epitopes <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

this molecule, while p80-homologous segments spanned at least <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the 3 T-cell epitopes <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

autoantigen in 8 cases (in the others, overlap was partial). Our data suggest that some fungi could trigger anti-<br />

Ku-positive AID via MM. Further research is needed to verify this hypothesis.<br />

1E_10_P<br />

(poster secti<strong>on</strong> A1, poster board #52, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SHEDDING OF MEMBRANE VESICLES CONTAINING HSP70 AND FGF-2 FROM A6<br />

STEM CELLS<br />

F. Geraci, G. Turturici, M. E. Candela, M. L.Vittorelli, S. Taverna, M. Salam<strong>on</strong>e, G. Giudice,<br />

G. Sc<strong>on</strong>zo<br />

Dipartimento di Biologia Cellulare e dello Sviluppo, Università di Palermo, Italy<br />

Our previous studies showed that A6 stem cells synthesize the inducible <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 without any stress,<br />

with an HSF1 independent pathway. In order to find a possible role <str<strong>on</strong>g>for</str<strong>on</strong>g> this basal expressi<strong>on</strong>, we observed its<br />

localizati<strong>on</strong> in the cell. Our results indicated that some <str<strong>on</strong>g>of</str<strong>on</strong>g> the HSP70 present inside the cell co-localized with<br />

actin micr<str<strong>on</strong>g>of</str<strong>on</strong>g>ilament network. The most interesting result was that HSP70 exists in two different <str<strong>on</strong>g>for</str<strong>on</strong>g>ms: a<br />

major Trit<strong>on</strong> X-100 soluble <str<strong>on</strong>g>for</str<strong>on</strong>g>m and a minor detergent-insoluble <str<strong>on</strong>g>for</str<strong>on</strong>g>m associated to detergent resistant<br />

microdomains. On the basis <str<strong>on</strong>g>of</str<strong>on</strong>g> both our findings and previous studies we might hypothesize that the basally<br />

expressed HSP70 might be released in the extracellular space through these microdomains. For this reas<strong>on</strong><br />

we tried to dem<strong>on</strong>strate whether or not HSP70 was released in the extracellular space. We found that A6<br />

stem cells were able to produce vesicles which were observed by c<strong>on</strong>focal microscope using fluorescinc<strong>on</strong>jugated<br />

phalloidine <strong>on</strong> cells grown in a three-dimensi<strong>on</strong>al matrix gel. Vesicle shedding was increased by<br />

serum replacement and it reached a peak three hours later. A6 cells are also able to produce vesicles in a<br />

serum free medium. Vesicle size, (∼1 micr<strong>on</strong>), may exclude the exosome origin. Immunoblot assays<br />

dem<strong>on</strong>strated the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> a large amount <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 in the vesicles, and its presence was c<strong>on</strong>firmed by<br />

immun<str<strong>on</strong>g>of</str<strong>on</strong>g>luorescence assays. An increase in HSP70 level was evident after heat shock. Immunoblot assays<br />

also dem<strong>on</strong>strated that these vesicles c<strong>on</strong>tained FGF-2. Its presence was c<strong>on</strong>firmed by functi<strong>on</strong>al analysis. In<br />

fact, vesicles obtained from A6 cells were able to induce uPA up-regulati<strong>on</strong>, a typical resp<strong>on</strong>se elicited by<br />

FGF-2, in bovine fetal aortic endothelial cells.<br />

62


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

1E_11_P<br />

(poster secti<strong>on</strong> A1, poster board #53, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

JJJ1, A NOVEL CYTOPLASMIC PRE-60S FACTOR INVOLVED IN THE RECYCLING<br />

OF PRERIBOSOMAL SHUTTLING PROTEINS IN SACCHAROMYCES CEREVISIAE<br />

Emilie Demoinet 2 , Alain Jacquier 1 , Georges Lutfalla 2 , Micheline From<strong>on</strong>t-Racine 1<br />

Departement <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, UMR5235, DAA, Place E. Bataill<strong>on</strong>, 34095 M<strong>on</strong>tpellier cedex 5, France<br />

The ribosome is <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the most important cellular macromolecular structures both by its size and by the<br />

energy that a cell c<strong>on</strong>sumes <str<strong>on</strong>g>for</str<strong>on</strong>g> its biogenesis. The biogenesis occurs from the nucleus to the cytoplasm. In<br />

additi<strong>on</strong> to the ribosomal proteins, about 80 out <str<strong>on</strong>g>of</str<strong>on</strong>g> the 200 preribosomal factors are involved in the 60S large<br />

subunit maturati<strong>on</strong>. In c<strong>on</strong>trast to the ribosomal proteins, which bind early and remain tightly associated to<br />

the mature particle, the preribosomal factors are bound <strong>on</strong>ly transiently. Some <str<strong>on</strong>g>of</str<strong>on</strong>g> them recycle into the<br />

nucleus, while others, like Arx1, Alb1, Tif6, shuttle between the nucleus and the cytoplasm. A few other<br />

factors like Rei1 are strictly cytoplasmic and participate to the dissociati<strong>on</strong>/recycling <str<strong>on</strong>g>of</str<strong>on</strong>g> the nuclear pre60S<br />

shuttling factors.<br />

Here, we investigate the role <str<strong>on</strong>g>of</str<strong>on</strong>g> Jjj1, an Hsp40 chaper<strong>on</strong>e. We show that Jjj1 is a pre-60S factor involved in<br />

the last cytoplasmic steps <str<strong>on</strong>g>of</str<strong>on</strong>g> the 60S subunit biogenesis.<br />

We found that Jjj1 and Rei1 are biochemically associated to the same complex and are two-hybrid partners.<br />

The absence <str<strong>on</strong>g>of</str<strong>on</strong>g> Jjj1 like the absence <str<strong>on</strong>g>of</str<strong>on</strong>g> Rei1, results in a defect in the amount <str<strong>on</strong>g>of</str<strong>on</strong>g> the large subunit, the<br />

appariti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> half-mers and leads to a cold sensitive phenotype. We could show that Rei1 and Jjj1 are<br />

genetically linked and that the deleti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ARX1 or ALB1 genes restored the wild-type phenotype <str<strong>on</strong>g>of</str<strong>on</strong>g> jjj1∆ as<br />

previously dem<strong>on</strong>strated <str<strong>on</strong>g>for</str<strong>on</strong>g> rei1∆. These results indicate that the two proteins Rei1 and Jjj1 act in the same<br />

pathway. Furthermore, even if the overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> REI1 is able to partially restore the growth defect <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

jjj1∆ strain, the reverse is not true, suggesting that their functi<strong>on</strong>s are not fully equivalent.<br />

We will present results showing that even if Jjj1 and Rei1 share several comm<strong>on</strong> characteristics, each protein<br />

is involved in a specific functi<strong>on</strong>. Altogether, these results suggest that <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the essential roles <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

cytoplasmic preribosomal factors would be to recycle the shuttling preribosomal factors.<br />

1E_12_P<br />

(poster secti<strong>on</strong> A1, poster board #54, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

IN VITRO ASSEMBLY OF HSP90 COMPLEXES IN YEAST CELL EXTRACTS<br />

Gary Flom, Jill L. Johns<strong>on</strong><br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Microbiology, Molecular Biology and Biochemistry, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Idaho, Moscow Id. 83844-3052<br />

e-mail: jilljohn@uidaho.edu<br />

The ATP-dependent molecular chaper<strong>on</strong>e Hsp90 and partner co-chaper<strong>on</strong>e proteins are required <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

folding and activity <str<strong>on</strong>g>of</str<strong>on</strong>g> diverse cellular client proteins, including steroid horm<strong>on</strong>e receptors and multiple<br />

<strong>on</strong>cogenic kinases. Hsp90 undergoes nucleotide-dependent c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>al changes but little is known about<br />

how these changes are coupled to client protein activati<strong>on</strong>. We established an assay to m<strong>on</strong>itor the interacti<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> wild-type and mutant Hsp90 with Sti1, Sba1 and Cpr6 in yeast cell extracts. Wild-type Hsp90 bound Sti1 in<br />

a nucleotide-independent manner, while Sba1 and Cpr6 specifically and independently interacted with Hsp90<br />

in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> the n<strong>on</strong>-hydrolyzable analog <str<strong>on</strong>g>of</str<strong>on</strong>g> ATP, AMP-PNP. Mutati<strong>on</strong>s throughout Hsp90 had<br />

dramatic effects <strong>on</strong> the interacti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> each <str<strong>on</strong>g>of</str<strong>on</strong>g> these three co-chaper<strong>on</strong>e proteins. These studies indicate that<br />

cycling <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90 between the nucleotide-free, open c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> and the ATP-bound, closed c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong><br />

63


23-26 August 2007,<br />

Budapest, Hungary<br />

is influenced by residues both within and outside the N-terminal ATPase domain and that these<br />

c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>al changes have dramatic effects <strong>on</strong> interacti<strong>on</strong> with co-chaper<strong>on</strong>e proteins. Our recent ef<str<strong>on</strong>g>for</str<strong>on</strong>g>ts<br />

focus <strong>on</strong> extending these studies to include analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> how nucleotide and Hsp90 mutati<strong>on</strong>s affects<br />

additi<strong>on</strong>al interacting proteins, including the Aha1 and Ppt1 co-chaper<strong>on</strong>es and candidate client proteins that<br />

interact with Hsp90.<br />

1E_13_P<br />

(poster secti<strong>on</strong> A1, poster board #55, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

IDENTIFICATION AND CHARACTERIZATION OF A NOVEL HSP40 FAMILY<br />

MEMBER PROTEIN, DNAJB8<br />

Kenjiro Kamiguchi, Alisa Sokolovskaya, Toshihiko Torigoe, Yoshihiko Hirohashi,<br />

Katsuya Nakanishi, Kanako Hirose, Noriyuki Sato<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Pathology, Sapporo Medical University, Minami-1, Nishi-17, Sapporo 060-8556, Japan,<br />

e-mail: kkamigu@sapmed.ac.jp<br />

Molecular chaper<strong>on</strong>es have evolved diverse tertiary and quaternary structures to stabilize n<strong>on</strong>-native<br />

polypeptides and facilitate their transiti<strong>on</strong> to the native state. Hsp40/DNAJ co-chaper<strong>on</strong>es are characterized<br />

by the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> a highly c<strong>on</strong>served J-domain that is required <str<strong>on</strong>g>for</str<strong>on</strong>g> interacti<strong>on</strong> with the ATPase domain <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Hsp70. More than <str<strong>on</strong>g>for</str<strong>on</strong>g>ty Hsp40 homologues have been identified in human and they are classified into three<br />

groups according to their domain structure. We report here that the isolati<strong>on</strong> and characterizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a novel<br />

human Hsp40 family member protein, DNAJB8 (JB8). JB8 was identified by searching the EST database <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

proteins that c<strong>on</strong>tained a J-domain. We first examined the distributi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> human JB8 mRNA in tissues. It<br />

was str<strong>on</strong>gly expressed in testis and several tumors, but not in heart, lung, liver, muscles, intestines, and<br />

leukocytes. Next, the subcellular localizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> JB8 was studied by scanning laser c<strong>on</strong>focal microscopy <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

chimera with the fluorescent protein (EGFP) transiently expressed in HeLa cells. JB8-EGFP was localized<br />

mainly to the cytosol, and probably to the centrosome. The centrosome is the major microtubule organizing<br />

center <str<strong>on</strong>g>of</str<strong>on</strong>g> animal cells. The centrosome duplicates <strong>on</strong>ce every cell cycle. Abrogati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> regulatory<br />

mechanisms governing centrosome duplicati<strong>on</strong> leads to generati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> amplified centrosomes (more than two<br />

centrosomes), which in turn leads to mitotic aberrati<strong>on</strong> and unequal segregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> chromosomes. It is<br />

reported that Hsp70 could suppress the abnormal centrosome amplificati<strong>on</strong> in cancer cells. Because JB8 is<br />

str<strong>on</strong>gly associated with Hsp/Hsc70 examined by the immunoprecipitati<strong>on</strong> assay, we examined whether JB8<br />

can suppress the abnormal centrosome amplificati<strong>on</strong>. Overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> JB8, but not DNAJC8 or HEDJ, in<br />

HeLa cells could str<strong>on</strong>gly suppress the abnormal centrosome amplificati<strong>on</strong>. Thus, these data suggest that JB8<br />

can suppress the malignant potential in cancer cells.<br />

1E_14_P<br />

(poster secti<strong>on</strong> A1, poster board #52, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE ROLE OF HSP70 GENE NUMBER IN FERTILITY<br />

Levente Kovács<br />

Babes-Bolyai University, Cluj-Napoca, str. Clinicilor 5, Romania<br />

The Hsp70 (heat-shock protein with the weight 70kDa) has various functi<strong>on</strong> within the cell: helps to the<br />

newly emerged proteins to find their native c<strong>on</strong>figurati<strong>on</strong>, repairs the impaired structure <str<strong>on</strong>g>of</str<strong>on</strong>g> the other proteins<br />

64


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

during the stress, and takes part in several metabolical pathways. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> our project was to find out the<br />

relati<strong>on</strong>ship between the hsp70 genes and the fertility using fruit fly Drosophila melanogaster as a model<br />

organism. Using genetical tools, we made mutants with 2, 3, 5, 7 and 8 hsp70 genes from those flies which<br />

have basicly 10 hsp70 genes (10 is the normal number <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp70 in the Drosophila’s genome).Testing the<br />

fertility <str<strong>on</strong>g>of</str<strong>on</strong>g> these mutants we find out that the fertility decreases with the number <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp70 genes. Moreover,<br />

we examined a change in the reproductive system: accumulated mature eggs in the ovarium. Our experiments<br />

dem<strong>on</strong>strated that the fertility str<strong>on</strong>gly depends <strong>on</strong> the number <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp70 genes in the organism.<br />

65


23-26 August 2007,<br />

Budapest, Hungary<br />

66<br />

1F. PHARMACOLOGICAL MODULATORS OF STRESS PROTEIN EXPRESSION<br />

1F_02_P<br />

(poster secti<strong>on</strong> A1, poster board #53, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STRESS PROTEIN ACTIVATION BY THE OMEGA-3 FATTY<br />

ACID DOCOSAHEXAENOIC ACID IN HUMAN COLON CANCER CELLS<br />

Hilde Bremseth 1* , Anne Gøril Lundemo 2 , Caroline Hild Jakobsen 2 , Gro Leite Størvold 2,3 ,<br />

Hans Einar Krokan 1 , Svanhild Arentz Schønberg 2<br />

1 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Cancer <str<strong>on</strong>g>Research</str<strong>on</strong>g> and Molecular Medicine, NTNU<br />

Erling Skjalgss<strong>on</strong>s gate 1, N-7006 Tr<strong>on</strong>dheim, Norway<br />

2 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Laboratory Medicine, Children’s and Women’s Health, NTNU<br />

Erling Skjalgss<strong>on</strong>s gate 1, N-7006 Tr<strong>on</strong>dheim, Norway<br />

3Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Genetics, Ullevål University Hospital, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Oslo, 0318 Oslo,<br />

Norway<br />

e-mail: hilde.bremseth@ntnu.no<br />

Docosahexaenoic acid (comm<strong>on</strong>ly known as DHA; 22:6(ω-3)) is an omega-3 essential fatty acid. We have<br />

earlier shown that DHA affects the proliferati<strong>on</strong> and survival <str<strong>on</strong>g>of</str<strong>on</strong>g> col<strong>on</strong> cancer cells. In order to evaluate the<br />

molecular mechanisms involved in the anti-cancer acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> this fatty acid, gene expressi<strong>on</strong> analysis were<br />

per<str<strong>on</strong>g>for</str<strong>on</strong>g>med at several time points. The results showed activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a stress resp<strong>on</strong>se after treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

cells with DHA. Heat shock protein 70 (HSP70) and the Nrf2-dependent stress protein heme oxygenase-1<br />

(HO-1) were both found to be highly up-regulated at the protein level. These results suggest that stress<br />

proteins are key mediators or modulators <str<strong>on</strong>g>of</str<strong>on</strong>g> the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> DHA. Genes encoding redox enzymes were also<br />

found to be up-regulated after DHA treatment. The work presented here will try to answer if the observed<br />

stress protein activati<strong>on</strong> in col<strong>on</strong> cancer cells treated with DHA is related to changes in the cellular redox<br />

state and involves modificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> redox-sensitive transcripti<strong>on</strong> factors.<br />

1F_03_P<br />

(poster secti<strong>on</strong> A1, poster board #54, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HANS SELYE’S “DISEASE OF ADAPTATION” - TYPE 2 DIABETES MELLITUS - A<br />

BREAKDOWN IN THE STRESS RESPONSE<br />

Philip L. Hooper, Paul L. Hooper<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Colorado Health Sciences Center, Glen Haven, CO, USA<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> New Mexico, Albuquerque, NM, USA<br />

e-mail: phoopermd@gmail.com<br />

Hans Selye identified type 2 diabetes as a “disease <str<strong>on</strong>g>of</str<strong>on</strong>g> adaptati<strong>on</strong>”, a pathology that occurs as the stress<br />

resp<strong>on</strong>se itself breaks down in the face <str<strong>on</strong>g>of</str<strong>on</strong>g> prol<strong>on</strong>ged, chr<strong>on</strong>ic stress. His writings p<strong>on</strong>dered the etiology <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

such diseases, suggesting that there was a finite energy <str<strong>on</strong>g>of</str<strong>on</strong>g> adaptati<strong>on</strong> that, when depleted, resulted in disease,<br />

early aging and ultimately death. Our thesis is that type 2 diabetes is indeed the product <str<strong>on</strong>g>of</str<strong>on</strong>g> a breakdown in the<br />

stress resp<strong>on</strong>se that results, not from general energetic c<strong>on</strong>straints, but from a specific self-perpetuating cycle<br />

that cripples the body’s ability to resp<strong>on</strong>d to stress via loss <str<strong>on</strong>g>of</str<strong>on</strong>g> insulin signaling. This breakdown occurs in the<br />

following way: chr<strong>on</strong>ic metabolic stress (from high fat diet, inactivity, aging, genetic predispositi<strong>on</strong>) leads to


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

the producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cytokines and counter-regulatory horm<strong>on</strong>es (cortisol, epinephrine, growth horm<strong>on</strong>e),<br />

which impair insulin signaling. Reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> insulin signaling activates glycogen synthase kinase-3 (GSK-3)<br />

which deactivates Heat Shock Factor-1, resulting in lower heat shock proteins (Hsps)-- a state which further<br />

promotes cytokine activity and augments defects in insulin signaling. Pro<str<strong>on</strong>g>of</str<strong>on</strong>g> <str<strong>on</strong>g>of</str<strong>on</strong>g> c<strong>on</strong>cept <str<strong>on</strong>g>for</str<strong>on</strong>g> this thesis is<br />

supported by these observati<strong>on</strong>s: 1) Reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> pain and stress in surgery improves outcomes. 2) Insulin<br />

infusi<strong>on</strong> at the time <str<strong>on</strong>g>of</str<strong>on</strong>g> surgery or in critically ill patients (with no diabetes) raises Hsp levels and improves<br />

morbidity and mortality. 3) Agents that improve insulin signaling (like GSK-3 inhibitors) raise Hsp levels,<br />

thus reducing inflammati<strong>on</strong> which further improves insulin signaling and survival. 4) When insulin signaling is<br />

damaged (pre-diabetes and diabetes), tissue Hsp levels are low and tissues are vulnerable to stress. Thus,<br />

Hans’ c<strong>on</strong>cern that the stress resp<strong>on</strong>se itself can lead to disease and death is valid, not due to energy<br />

exhausti<strong>on</strong>, but due to loss <str<strong>on</strong>g>of</str<strong>on</strong>g> cytoprotective Hsps and unabated inflammati<strong>on</strong>.<br />

1F_04_P<br />

(poster secti<strong>on</strong> A1, poster board #55, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECT OF GELDANAMYCIN ON TRANSCRIPTION OF HEAT-SHOCK PROTEIN<br />

90 ISOFORMS, HSP90ALPHA AND HSP90BETA, IN BRAIN OF GOLDFISH<br />

Nao Kagawa, Kohta Morishige<br />

Kinki University, Osaka, Japan. e-mail: kagawa@life.kindai.ac.jp<br />

Geldanamycin (GA) is a specific inhibitor <str<strong>on</strong>g>of</str<strong>on</strong>g> chaper<strong>on</strong>ing functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90 protein. We recently reported<br />

that intracerebral GA treatment drastically reduced basal level <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90 protein in the brain <str<strong>on</strong>g>of</str<strong>on</strong>g> goldfish<br />

(Carassius auratus). To understand mechanisms involved in this finding, we examined mRNA levels <str<strong>on</strong>g>of</str<strong>on</strong>g> two<br />

is<str<strong>on</strong>g>of</str<strong>on</strong>g>orms <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90, α and β, after GA administrati<strong>on</strong> in the brain <str<strong>on</strong>g>of</str<strong>on</strong>g> goldfish at a dose <str<strong>on</strong>g>of</str<strong>on</strong>g> 0.05 or 1.0 µg/gbody<br />

weight. Hsp90α and Hsp90β are stress-inducible and c<strong>on</strong>stitutive protein, respectively. Levels <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Hsp90α mRNA, Hsp90β mRNA and glyceraldehyde-3-phosphate-dehydrogenase (GAPDH) mRNA in the<br />

brain were determined by RT-PCR method at allotted time from 4 to 72 h after GA treatment. The OD ratio<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90α mRNA or Hsp90β mRNA to GAPDH mRNA was calculated as an index <str<strong>on</strong>g>of</str<strong>on</strong>g> transcripti<strong>on</strong> level<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> Hsp90α or Hsp90β gene. The OD ratio <str<strong>on</strong>g>for</str<strong>on</strong>g> Hsp90α after GA treatment at 1.0 and that at 0.05 µg/g both<br />

showed a 1.5 fold significant increase in comparis<strong>on</strong> with GA-untreated c<strong>on</strong>trol as determined at 12 and 20 h<br />

post-treatment; then, they peaked 20 and 28 h after treatment, respectively. Thereafter, the ratio values both<br />

decreased to the c<strong>on</strong>trol level at 36 h post-treatment. In c<strong>on</strong>trast, the OD ratios <str<strong>on</strong>g>for</str<strong>on</strong>g> Hsp90β were almost<br />

c<strong>on</strong>stant throughout the time course used after treatment without significant deviati<strong>on</strong> from the c<strong>on</strong>trol<br />

value. These results suggest that GA caused reducti<strong>on</strong> in the abundance <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90 protein in the brain <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

goldfish is a post-translati<strong>on</strong> event, most probably, by proteosome-mediated degradati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> GA-bound<br />

Hsp90α protein, which in turn upregulate transcripti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90α gene.<br />

67


23-26 August 2007,<br />

Budapest, Hungary<br />

68<br />

1F_05_P<br />

(poster secti<strong>on</strong> A1, poster board #56, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DESIGN OF NOVEL STRESS PROTECTORS BASED ON SHORT<br />

SYNTHETIC PEPTIDES<br />

E. V. Navolotskaya 1 , Y. A. Kovalitskaya 1 , A. A. Kolobov 2 , V. B. Sadovnikov 1<br />

1Branch <str<strong>on</strong>g>of</str<strong>on</strong>g> Shemyakin and Ovchinnikov Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Bioorganic Chemistry, Pushchino, Moscow Regi<strong>on</strong>, Russia;<br />

2State <str<strong>on</strong>g>Research</str<strong>on</strong>g> Center <str<strong>on</strong>g>for</str<strong>on</strong>g> Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Highly Pure Biopreparati<strong>on</strong>s, St. Petersburg, Russia<br />

The work is directed towards the soluti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the most important problems <str<strong>on</strong>g>of</str<strong>on</strong>g> modern biology and<br />

medicine – the creati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> novel effective and safe agents capable <str<strong>on</strong>g>of</str<strong>on</strong>g> elevating adaptati<strong>on</strong> potential <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

human body under the influence <str<strong>on</strong>g>of</str<strong>on</strong>g> various extreme factors. The objective <str<strong>on</strong>g>of</str<strong>on</strong>g> the work is synthesis <str<strong>on</strong>g>of</str<strong>on</strong>g> a set <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

short corticotropin-like peptides and study <str<strong>on</strong>g>of</str<strong>on</strong>g> their effect <strong>on</strong> n<strong>on</strong>-specific resistance <str<strong>on</strong>g>of</str<strong>on</strong>g> laboratory animals<br />

subjected to various stress factors. It was synthesized 75 <str<strong>on</strong>g>of</str<strong>on</strong>g> corticotropin-like peptides, which c<strong>on</strong>tained from<br />

13 to 2 amino acid residues. The ability <str<strong>on</strong>g>of</str<strong>on</strong>g> the synthesized peptides to inhibit the specific binding <str<strong>on</strong>g>of</str<strong>on</strong>g> tritium<br />

labeled corticotropin fragment 11-24 to rat adrenal cortex in vitro was investigated. On the base <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

obtained results there were selected 6 peptides (KKRR, KKRRP, VKKPGSSVKV, KKRRLLLL,<br />

LLKKRRLLLL, ac-KKRR-NH 2 ) with the highest inhibitory activity <str<strong>on</strong>g>for</str<strong>on</strong>g> in vivo tests. The influence <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

selected peptides <strong>on</strong> the level <str<strong>on</strong>g>of</str<strong>on</strong>g> glucocorticoids and catecholamines in the adrenals and blood <str<strong>on</strong>g>of</str<strong>on</strong>g> rats in the<br />

experiments <strong>on</strong> acute hemorrhage and hypobaric hypoxia were studied. It was established that intravenous<br />

injecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> peptides KKRR, KKRRP or ac-KKRR-NH 2 at the dose <str<strong>on</strong>g>of</str<strong>on</strong>g> 1 µg/kg could correct the changes in<br />

the c<strong>on</strong>tent <str<strong>on</strong>g>of</str<strong>on</strong>g> corticoster<strong>on</strong>e, adrenaline and noradrenaline in the adrenals and plasma <str<strong>on</strong>g>of</str<strong>on</strong>g> rats that were<br />

subjected to hemorrhagic shock or hypobaric hypoxia. It has been shown that the stress-protective activity <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

these peptides is mediated by the corticotropin receptor in cortex <str<strong>on</strong>g>of</str<strong>on</strong>g> the adrenals.<br />

1F_06_P<br />

(poster secti<strong>on</strong> A1, poster board #57, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PROTECTIVE EFFECT OF A NOVEL MOLECULAR CHAPERONE INDUCER,<br />

PAEONIFLORIN, ON THE HCL-INDUCED GASTRIC MUCOSAL INJURY<br />

Kenzo Ohtsuka, Daisuke Kawashima, Midori Asai<br />

Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Cell & Stress Biology, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Envir<strong>on</strong>mental Biology,<br />

Chubu University, 1200 Matsumoto-cho, Kasugai, Aichi 487-8501, Japan<br />

e-mail: kohtsuka@isc.chubu.ac.jp<br />

Moderate overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock proteins (HSPs) or molecular chaper<strong>on</strong>es could c<strong>on</strong>fer cells and<br />

tissues stress tolerance and provide beneficial effects <strong>on</strong> various pathological states associated with protein<br />

misfolding and protein aggregati<strong>on</strong>. Recently, we found a novel chaper<strong>on</strong>e inducing compound, pae<strong>on</strong>iflorin,<br />

which is <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the major c<strong>on</strong>stituents <str<strong>on</strong>g>of</str<strong>on</strong>g> a herbal medicine derived from Pae<strong>on</strong>ia lactiflora Pall. Pae<strong>on</strong>iflorin<br />

could induce Hsp70, Hsp40, and Hsp27 in cultured HeLa cells. Treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> cells with pae<strong>on</strong>iflorin resulted<br />

in enhanced phosphorylati<strong>on</strong> and acquisiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> DNA binding ability <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock factor 1 (HSF1), as well<br />

as the <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> characteristic HSF1 granules in the nucleus, suggesting that the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these HSPs<br />

by pae<strong>on</strong>iflorin is mediated by the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF1. Also, thermotolerance was induced by the treatment<br />

with pae<strong>on</strong>iflorin. Pae<strong>on</strong>iflorin had no apparent toxic effect at c<strong>on</strong>centrati<strong>on</strong>s as high as 500 M (Cell Stress<br />

& Chaper<strong>on</strong>es 9: 387-389, 2004).


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Also, we investigated the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> pae<strong>on</strong>iflorin <strong>on</strong> the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 in mouse stomach and the HClinduced<br />

gastric mucosal injury. Administrati<strong>on</strong> (i.p.) <str<strong>on</strong>g>of</str<strong>on</strong>g> pae<strong>on</strong>iflorin resulted in the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 in<br />

mouse stomach detected by western blot and immunohistochemical staining. Severe gastric mucosal injury<br />

was caused 3 hr after oral administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HCl. Prior administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> pae<strong>on</strong>iflorin could protect the HClinduced<br />

gastric mucosal injury as dem<strong>on</strong>strated by an overview and histological HE staining. The extent <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

protecti<strong>on</strong> was well correlated with the expressi<strong>on</strong> level <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70. No apparent systemic side effect <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

pae<strong>on</strong>iflorin was observed so far. Hsp70 was also induced in liver, heart and brain by pae<strong>on</strong>iflorin<br />

(manuscript in preparari<strong>on</strong>). From these results, it is suggested that pae<strong>on</strong>iflorin and pae<strong>on</strong>iflorin-c<strong>on</strong>taining<br />

herbal medicines might be used clinically as chaper<strong>on</strong>e inducers <str<strong>on</strong>g>for</str<strong>on</strong>g> the preventi<strong>on</strong> and treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> various<br />

pathological states, such as stress ulcers and ischemia-induced injuries, and <str<strong>on</strong>g>of</str<strong>on</strong>g> diseases associated with protein<br />

misfolding and protein aggregati<strong>on</strong>. (This work was supported in part by a Grant-in-Aid <str<strong>on</strong>g>for</str<strong>on</strong>g> the High-Tech<br />

<str<strong>on</strong>g>Research</str<strong>on</strong>g> Center Establishment Project (2002-2006, Chubu University) from the Japanese Ministry <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Educati<strong>on</strong>, Culture, Sports, Science and Technology.)<br />

1F_07_P<br />

(poster secti<strong>on</strong> A1, poster board #58, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ADAPTOGENS – MODIFIERS OF STRESS RESPONSE<br />

1 A. Panossian*, G. Wikman, 2 M. Hambartsumyan, A. Hovhanissyan<br />

1Swedish Herbal Institute <str<strong>on</strong>g>Research</str<strong>on</strong>g> and Development, Prinsgatan 12, SE-413 05 Göteborg, Sweden<br />

2“ExLab” Expert Analytical Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Armenia Drug Agency, Komitas Ave. 49/4,<br />

375051 Yerevan, Armenia,<br />

e-mail <str<strong>on</strong>g>of</str<strong>on</strong>g> corresp<strong>on</strong>ding author: ap@shi.se<br />

Adaptogens possess anti-fatigue and anti-stress activities that can increase mental and physical working<br />

per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance against a background <str<strong>on</strong>g>of</str<strong>on</strong>g> fatigue or stress. A characteristic feature <str<strong>on</strong>g>of</str<strong>on</strong>g> adaptogens is that they act<br />

as eustressors or challengers and give rise to stimulating or stress-ag<strong>on</strong>ising effects following single<br />

administrati<strong>on</strong>. The stress-protective acti<strong>on</strong> achieved by multiple administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> adaptogens is not,<br />

there<str<strong>on</strong>g>for</str<strong>on</strong>g>e, the result <str<strong>on</strong>g>of</str<strong>on</strong>g> inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> an organism, but <str<strong>on</strong>g>of</str<strong>on</strong>g> adaptive changes in the<br />

organism in resp<strong>on</strong>se to the repeated stress-ag<strong>on</strong>istic properties <str<strong>on</strong>g>of</str<strong>on</strong>g> the drug. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> the present study was<br />

to ascertain which mediators <str<strong>on</strong>g>of</str<strong>on</strong>g> stress resp<strong>on</strong>se are significantly involved in the mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

adaptogens, and to determine their relevance as biochemical markers <str<strong>on</strong>g>for</str<strong>on</strong>g> evaluating anti-stress effects in<br />

rabbits subjected to immobilisati<strong>on</strong> stress. Blood levels <str<strong>on</strong>g>of</str<strong>on</strong>g> stress-activated protein kinase (SAPK/JNK), the<br />

phosphorylated kinase p-SAPK/p-JNK, nitric oxide (NO), cortisol, testoster<strong>on</strong>e, prostaglandin E2,<br />

leukotriene B4 and thromboxane B2 were determined in groups <str<strong>on</strong>g>of</str<strong>on</strong>g> animals prior to daily oral administrati<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> placebo or rhodioloside or extracts <str<strong>on</strong>g>of</str<strong>on</strong>g> Eleutherococcus senticosus, Schisandra chinensis, Rhodiola rosea,<br />

Bry<strong>on</strong>ia alba or Panax ginseng over a 7 day period. Ten minutes after the final treatment, animals were<br />

immobilised <str<strong>on</strong>g>for</str<strong>on</strong>g> 2 hours and blood levels <str<strong>on</strong>g>of</str<strong>on</strong>g> the markers re-determined. In the placebo group, <strong>on</strong>ly p-<br />

SAPK/p-JNK, NO and cortisol were increased significantly (by 200-300% cf basal levels) following<br />

immobilisati<strong>on</strong> stress, whilst in animals that had received multiple doses <str<strong>on</strong>g>of</str<strong>on</strong>g> adaptogens/stress-protectors, the<br />

levels <str<strong>on</strong>g>of</str<strong>on</strong>g> NO and cortisol remained practically unchanged after acute stress. It is known that NO can str<strong>on</strong>gly<br />

inhibit the producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular energy through two mechanisms: (i) inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>drial respirati<strong>on</strong><br />

by reversible and irreversible inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cytochrome P450, and (ii) the inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> glycolysis through<br />

modificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the SH-groups <str<strong>on</strong>g>of</str<strong>on</strong>g> glyceraldehyde-3-phosphate dehydrogenase. The inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> stressinduced<br />

NO producti<strong>on</strong> dem<strong>on</strong>strated in the present study may provide an explanati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the energy<br />

bursting effects <str<strong>on</strong>g>of</str<strong>on</strong>g> adaptogens that result in a prol<strong>on</strong>ged phase <str<strong>on</strong>g>of</str<strong>on</strong>g> endurance and postp<strong>on</strong>ed fatigue.<br />

69


23-26 August 2007,<br />

Budapest, Hungary<br />

Rhodioloside and extracts <str<strong>on</strong>g>of</str<strong>on</strong>g> S. chinensis and R. rosea (adaptogens c<strong>on</strong>taining <strong>on</strong>ly phenolic compounds)<br />

were the most active inhibitors <str<strong>on</strong>g>of</str<strong>on</strong>g> stress-induced p-SAPK/p-JNK. B. alba and P. ginseng (stress-protectors<br />

c<strong>on</strong>taining <strong>on</strong>ly cortisol-like substances) exerted little effect <strong>on</strong> p-SAPK/p-JNK levels. It is suggested that the<br />

inhibitory effects <str<strong>on</strong>g>of</str<strong>on</strong>g> R. rosea and S. chinensis <strong>on</strong> SAPK/JNK activati<strong>on</strong> may be associated with their antidepressant<br />

activity as well as their positive effects <strong>on</strong> mental per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance under stress.<br />

1F_08_P<br />

(poster secti<strong>on</strong> A1, poster board #59, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

INHIBITION OF ER STRESS-INDUCED XBP1 ACTIVATION BY TRIENE-<br />

ANSAMYCINS<br />

Etsu Tashiro, Yushi Futamura, Naoka Hir<strong>on</strong>iwa, Masaya Imoto<br />

Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Science and Technology, Keio University, Yokohama 223-8522, Japan<br />

e-mail: tashiro@bio.keio.ac.jp<br />

Accumulati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> unfolded proteins in ER cause ER stress, leading to activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> several transcripti<strong>on</strong><br />

factors. Am<strong>on</strong>g them, XBP1 is <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the major transcripti<strong>on</strong> factors <str<strong>on</strong>g>of</str<strong>on</strong>g> ER stress. Under ER stress<br />

c<strong>on</strong>diti<strong>on</strong>, a 26-nt intr<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> XBP1 mRNA is spliced out by activated IRE1, leading to a frame shift. This<br />

splicing event creates translati<strong>on</strong>al frameshift in XBP1 mRNA to produce an active transcripti<strong>on</strong> factor.<br />

Recently, XBP1 is c<strong>on</strong>sidered to be much correlated with tumor progressi<strong>on</strong>, because it has been reported<br />

that Xbp1-knockdown cells did not <str<strong>on</strong>g>for</str<strong>on</strong>g>m tumors in mice and the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> XBP1 was increased in a<br />

several types <str<strong>on</strong>g>of</str<strong>on</strong>g> tumor cells. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, XBP1 inhibitor would be a new class <str<strong>on</strong>g>of</str<strong>on</strong>g> anti-cancer drug. To screen<br />

inhibitors <str<strong>on</strong>g>of</str<strong>on</strong>g> ER stress-induced XBP1 activati<strong>on</strong>, we first c<strong>on</strong>structed pcDNA3/XBP1-luc plasmid that is<br />

fused the luciferase cDNA downstream <str<strong>on</strong>g>of</str<strong>on</strong>g> XBP1 cDNA. Then, we transfected pcDNA3/XBP1-luc plasmid<br />

into HeLa cells to generate HeLa/XBP1-luc cells that are stably transcribed XBP1-luciferase mRNA. Under<br />

normal c<strong>on</strong>diti<strong>on</strong>, the XBP1-luciferase mRNA is not spliced and its translati<strong>on</strong> is terminated at the stop<br />

cod<strong>on</strong> near the splice site <str<strong>on</strong>g>of</str<strong>on</strong>g> XBP1 mRNA, resulting in that the luciferase protein is not produced. In<br />

c<strong>on</strong>trast, under ER stress c<strong>on</strong>diti<strong>on</strong>, the 26-nt intr<strong>on</strong> is excised, leading to frame shift <str<strong>on</strong>g>of</str<strong>on</strong>g> XBP1-luciferase<br />

mRNA and its translati<strong>on</strong> is then terminated at the stop cod<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the luciferase mRNA. Thus, we can easily<br />

detect the XBP1 activati<strong>on</strong> as the luciferase reporter signals in HeLa/XBP1-luc cells. In the course <str<strong>on</strong>g>of</str<strong>on</strong>g> our<br />

screening <str<strong>on</strong>g>for</str<strong>on</strong>g> the inhibitor <str<strong>on</strong>g>of</str<strong>on</strong>g> thapsigargin-induced XBP1 activati<strong>on</strong> from microbial origin by using<br />

HeLa/XBP1-luc cells, we found and isolated natural small molecules, triene-ansamycins such as trienomycin<br />

A. Triene-ansamycins showed a marked inhibitory activity toward thapsigargin-induced XBP1 activati<strong>on</strong>, as<br />

evaluated by both luciferase assay and RT-PCR. Here, we will present the isolati<strong>on</strong>, structure determinati<strong>on</strong>,<br />

and biological activities <str<strong>on</strong>g>of</str<strong>on</strong>g> Triene-ansamycins.<br />

70


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

1F_09_P<br />

(poster secti<strong>on</strong> A1, poster board #60, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECT OF DOBUTAMINE AND NOR-EPINEPHRINE ON TNF-Α AND IL-6<br />

RELEASE OF WHOLE BLOOD AFTER LPS STIMULATION<br />

V. Papandreou, P. Myrianthefs, G. Fildissis, J. Portolos, G. Baltopoulos<br />

Athens University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Nursing ICU at « KAT » Hospital, Nikis 2, 14561, Kifissia, Greece<br />

e-mail: vpapandr@gmail.com<br />

The purpose <str<strong>on</strong>g>of</str<strong>on</strong>g> the study was to examine the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> dobutamine (D) and nor-epinephrine (NE) <strong>on</strong><br />

cytokine release <str<strong>on</strong>g>of</str<strong>on</strong>g> whole blood after ex vivo LPS stimulati<strong>on</strong>. Heparinized blood samples were collected<br />

from 10 healthy volunteers, (mean age 33.6±1.5, all men) at 08.00 AM and transferred to the lab <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

processing. One hundred µL <str<strong>on</strong>g>of</str<strong>on</strong>g> blood was added in 900 µL <str<strong>on</strong>g>of</str<strong>on</strong>g> RPMI 1640 culture medium <str<strong>on</strong>g>for</str<strong>on</strong>g> final volume<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> 1 ml and placed in plastic culture dishes. Samples were then added to wells and maintained at 37 o C in a<br />

5% CO 2 atmosphere and LPS was added (500 pg/ml). After incubati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 4 hours the culture plates were<br />

centrifuged (1800 rpm, 5 min) and supernatants were collected <str<strong>on</strong>g>for</str<strong>on</strong>g> cytokine measurements using the ELISA<br />

method. We studied the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> D and NE at different doses (10 -6 , 10 -5 , 10 -4 ) <strong>on</strong> cytokine producti<strong>on</strong> after<br />

whole blood LPS stimulati<strong>on</strong>. We repeated the experiment by adding metoprolol (b-blocker) at a dose <str<strong>on</strong>g>of</str<strong>on</strong>g> 10 -5 .<br />

Total WBCs were 6730 ± 242.2; lymphocytes were 2461 ± 395.7 and m<strong>on</strong>ocytes 359.2 ± 38.5 per µl.<br />

Dobutamine and NE significantly (p


72<br />

23-26 August 2007,<br />

Budapest, Hungary


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

CORE MODULE 2<br />

STRESS AND CELLULAR<br />

FUNCTIONS<br />

73


23-26 August 2007,<br />

Budapest, Hungary<br />

SYMPOSIA<br />

• 2A. Cell stress, Hsps and apoptosis (August 25 th Saturday morning)<br />

(Organizer and chair: Seamus Martin)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

• 2B. Membrane-regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress resp<strong>on</strong>se (August 25 th Saturday afterno<strong>on</strong>)<br />

(Organizer and chair: László Vígh and John L. Harwood)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

• 2C. N<strong>on</strong>-coding RNAs and cellular stress (August 26 th Sunday morning)<br />

(Organizer and chair: Subhash C. Lakhotia)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

• 2D. ER stress (August 25 th Saturday afterno<strong>on</strong>)<br />

(Organizers and chairs: Gábor Bánhegyi and Amy S. Lee)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

• 2E. Hypoxia (August 24 th Friday afterno<strong>on</strong>)<br />

(Organizer and chair: Jacques Pouyssegur and Cormac T. Taylor)<br />

Poster: August 24 th Friday, Poster secti<strong>on</strong> B1, Building B<br />

• 2F. Quality c<strong>on</strong>trol and stress-related protein networks (August 25 th Saturday<br />

morning)<br />

(Organizer and chair: Wolfgang Voos)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

• 2G. Stress <str<strong>on</strong>g>of</str<strong>on</strong>g> bacteria (August 24 th Friday morning)<br />

(Organizer and chair: María M. Tavío)<br />

Poster: August 24 th Saturday, Poster secti<strong>on</strong> B1, Building B<br />

POSTER SYMPOSIA<br />

• 2H. Oxidative stress<br />

August 25 th Saturday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

• 2I.Stress <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>dria<br />

August 25 th Saturday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

• 2J. Stress signaling<br />

August 25 th Saturday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

• 2K. Gene expressi<strong>on</strong> regulati<strong>on</strong> under stress<br />

August 25 th Saturday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

• 2L. Yeast stress<br />

August 25 th Saturday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

• 2M. Other cellular stress related topics (Stress-related protein degradati<strong>on</strong>, Stress and cell<br />

movement, Stress proteins and the cell cycle, Extremophiles)<br />

August 25 th Saturday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

74


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2A. CELL STRESS, HSPS AND APOPTOSIS<br />

(SEAMUS MARTIN)<br />

2A_01_S<br />

P53 FAMILY IN APOPTOSIS<br />

Ele<strong>on</strong>ora Lapi 1 , Silvia Di Agostino 1 , Hilah Gal 3 , Sara D<strong>on</strong>zelli 1 , Eytan Domany 4 , Gide<strong>on</strong> Rechavi 5 ,<br />

Sabrina Strano 1 , David Givol 3 , Xin Lu 5 , Giovanni Blandino 1,2<br />

1Regina Elena Cancer Institute, Rome , Italy, e-mail: blandino@ifo.it<br />

2Rome Oncogenomic Center, Rome, Italy.<br />

3 Weizmann Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Science, Rehovot , Israel.<br />

4 Weizmann Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Science, Rehovot , Israel.<br />

5 Sheba Medical Center, Tel-Hashomer, Israel.<br />

6Ludwig Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Cancer <str<strong>on</strong>g>Research</str<strong>on</strong>g>, L<strong>on</strong>d<strong>on</strong>, UK<br />

The p53 family is known to be involved in the transcripti<strong>on</strong>al c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> growth arrest and apoptosis. Despite<br />

the recent identificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> specific p73-target genes by genome-wide expressi<strong>on</strong> pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ile techniques, p73-<br />

mediated apoptosis occurs mostly through the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a set <str<strong>on</strong>g>of</str<strong>on</strong>g> genes that were originally found to be<br />

activated by p53. This suggests that promoter selectivity by both p53 and p73 might be the result <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

biochemical events such as post-translati<strong>on</strong>al modificati<strong>on</strong>s and specific protein-protein interacti<strong>on</strong>s.<br />

We have already shown that the transcripti<strong>on</strong>al co-activator Yes-associated protein (YAP) interacts with p73<br />

and determines p73 gene targeting in resp<strong>on</strong>se to DNA damage. We have also found that YAP localizes into<br />

the PML nuclear bodies and requires PML to exert its functi<strong>on</strong> as a specific co-activator <str<strong>on</strong>g>of</str<strong>on</strong>g> p73. Here we<br />

show the existence <str<strong>on</strong>g>of</str<strong>on</strong>g> a pro-apoptotic auto-regulatory feedback loop, during the apoptotic resp<strong>on</strong>se, between<br />

p73, YAP and PML. We dem<strong>on</strong>strate that the p73/YAP complex is required <str<strong>on</strong>g>for</str<strong>on</strong>g> PML inducti<strong>on</strong> after<br />

cisplatin treatment and that PML exerts a vital role in the executi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the apoptotic process regulating YAP<br />

stability. YAP is becoming a very intriguing protein due to its critical role in regulating p73 accumulati<strong>on</strong> and<br />

functi<strong>on</strong> following DNA damage, but very little is known about its regulati<strong>on</strong>. Here we show that YAP is<br />

polyubiquitinylated and degraded through the ubiquitin-proteasome pathway. We also show that YAP and<br />

PML physically interact and that PML regulates YAP half-life, preventing its ubiquitinylati<strong>on</strong> and subsequent<br />

degradati<strong>on</strong>.<br />

2A_02_S<br />

HEAT SHOCK PROTEINS ORCHESTRATE DE DECISION DIFFERENTIATION<br />

VERSUS APOPTOSIS<br />

Celine Didelot, Jean-Antoine Ribeil, Yael Zermati, D. Lanneau, Olivier Hermine, Carmen Garrido<br />

INSERM UMR 866, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Dij<strong>on</strong>, 21000, France and Necker Hospital, PARIS 75743<br />

Cedex 15, France<br />

Heat shock proteins, particularly HSP90, HSP70 and HSP27, are well known regulators <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis by<br />

interfering with key apoptotic proteins Apoptosis and cell differentiati<strong>on</strong> are two physiological processes that<br />

share comm<strong>on</strong> features like chromatin c<strong>on</strong>densati<strong>on</strong> and the need <str<strong>on</strong>g>of</str<strong>on</strong>g> the proteases called caspases. Little is<br />

known about the role <str<strong>on</strong>g>of</str<strong>on</strong>g> HSPs in the differentiati<strong>on</strong> process. Here we show that HSP70, during terminal<br />

blood red cells differentiati<strong>on</strong> and at the <strong>on</strong>set <str<strong>on</strong>g>of</str<strong>on</strong>g> caspase activati<strong>on</strong>, translocates into the nucleus where<br />

colocalizes and interacts with GATA-1, a ttranscripti<strong>on</strong> factor essential <str<strong>on</strong>g>for</str<strong>on</strong>g> red cells progenitors<br />

75


23-26 August 2007,<br />

Budapest, Hungary<br />

(erythroblasts) differentiati<strong>on</strong>. In vitro and in vivo assays dem<strong>on</strong>strate that HSP70 inhibits caspase-3-mediated<br />

proteolysis <str<strong>on</strong>g>of</str<strong>on</strong>g> GATA-1, allowing the differentiati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the erythroblasts. If the amount <str<strong>on</strong>g>of</str<strong>on</strong>g> nuclear HSP70 is<br />

reduced, GATA-1 is cleaved and the cells die by apoptosis (Ribeil et al, Nature 2007).<br />

Another HSP needed in the differentiati<strong>on</strong> process is HSP90, and more specifically the is<str<strong>on</strong>g>of</str<strong>on</strong>g>orm beta. During<br />

m<strong>on</strong>ocytic and epithelial cells’ differentiati<strong>on</strong>, HSP90β accompanies the protein c-IAP1 (inhibitor <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

apoptosis protein-1) that is translocated from the nucleus to the cytosol. This translocati<strong>on</strong> is needed <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

cells to differentiate (Plenchette et al, Blood 2004). Depleti<strong>on</strong> or neutralizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP90β blocks c-IAP1<br />

cytosolic translocati<strong>on</strong> and the differentiati<strong>on</strong> process. We c<strong>on</strong>clude that HSPs, like HSP70 or HSP90β, by<br />

their presence in a given cellular compartment and their cytoprotective properties, direct the cells to<br />

differentiate.<br />

2A_03_S<br />

NOVEL MECHANISMS OF HEAT SHOCK-INDUCED APOPTOSIS<br />

Rania S. Miller<strong>on</strong>, Indra M. Mahajan, Shawn B. Bratt<strong>on</strong><br />

Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmacology and Toxicology, College <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmacy, The University <str<strong>on</strong>g>of</str<strong>on</strong>g> Texas at Austin<br />

Adaptive resp<strong>on</strong>ses to mild heat shock are am<strong>on</strong>g the most widely c<strong>on</strong>served and studied in nature. More<br />

intense heat shock, however, induces apoptosis through mechanisms that remain largely unknown. Recently,<br />

we have observed that heat shock activates an apical protease, which stimulates mitoch<strong>on</strong>drial outer<br />

membrane permeabilizati<strong>on</strong> (MOMP) and processing <str<strong>on</strong>g>of</str<strong>on</strong>g> the effector caspase-3 in a zVAD-FMK (polycaspase<br />

inhibitor) and Bcl-2-inhibitable manner. Surprisingly, however, heat shock did not require any <str<strong>on</strong>g>of</str<strong>on</strong>g> the known<br />

initiator caspases (-2, -8, -9, or -12) or their activating complexes to promote apoptotic cell death, but instead<br />

relied up<strong>on</strong> the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> an apparently novel apical protease with caspase-like activity. Current ef<str<strong>on</strong>g>for</str<strong>on</strong>g>ts in<br />

the laboratory are now focused <strong>on</strong> unraveling the mechanisms through which heat shock induces MOMP,<br />

and our most recent findings will be discussed.<br />

2A_04_S<br />

LYSOSOMAL CONTROL OF TUMOR CELL DEATH BY HSP70<br />

Thomas Kirkegaard-Sørensen, Nicole Fehrenbacher, Jesper Nylandsted, Marja Jäättelä<br />

Apoptosis Department and <str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g> Genotoxic Stress, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Cancer Biology, Danish Cancer Society,<br />

Strandboulevarden 49, DK-2100 Copenhagen, Denmark<br />

e-mail: mj@cancer.dk<br />

Tumor invasi<strong>on</strong> and metastasis are associated with altered lysosomal trafficking and increased expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the lysosomal proteases termed cathepsins. Emerging experimental evidence suggest that such alterati<strong>on</strong>s in<br />

lysosomes may <str<strong>on</strong>g>for</str<strong>on</strong>g>m an “Achilles heel” <str<strong>on</strong>g>for</str<strong>on</strong>g> cancer cells by sensitizing them to death pathways involving<br />

lysosomal membrane permeabilizati<strong>on</strong> and the release <str<strong>on</strong>g>of</str<strong>on</strong>g> cathepsins into the cytosol. Here, I will highlight our<br />

recent unpublished results <strong>on</strong> cancer-related changes in the compositi<strong>on</strong> and functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> lysosomes, focusing<br />

<strong>on</strong> the mechanisms by which lysosomal Hsp70 inhibits cancer cell death and emerges as a putative target <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

future cancer therapy.<br />

76


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2A_05_S<br />

UNDERSTANDING THE DEMOLITION PHASE OF APOPTOSIS<br />

Seamus J. Martin<br />

Molecular Cell Biology Laboratory, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Genetics, The Smurfit Institute, Trinity College, Dublin 2, Ireland<br />

e-mail: martinsj@tcd.ie<br />

Apoptosis (programmed cell death) is coordinated by a family <str<strong>on</strong>g>of</str<strong>on</strong>g> cysteine proteases—the caspases—that<br />

dismantle cells by targeting numerous proteins <str<strong>on</strong>g>for</str<strong>on</strong>g> limited proteolysis. The mammalian caspase family<br />

c<strong>on</strong>tains 13 members, some <str<strong>on</strong>g>of</str<strong>on</strong>g> which participate in apoptosis. Caspases normally exist as dormant precursor<br />

enzymes in healthy cells but can be activated at the <strong>on</strong>set <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis via a number <str<strong>on</strong>g>of</str<strong>on</strong>g> distinct activati<strong>on</strong><br />

pathways. Here we discuss the caspase activati<strong>on</strong> pathways that are initiated by the cytotoxic T cell/Natural<br />

Killer cell protease granzyme B, as well as by cytotoxic drugs and diverse stress stimuli. How caspase<br />

activati<strong>on</strong> results in the c<strong>on</strong>trolled demoliti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the cell will also be explored.<br />

77


23-26 August 2007,<br />

Budapest, Hungary<br />

78<br />

2A_01_P<br />

(poster secti<strong>on</strong> A2, poster board #75, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

OSTEOPROTEGERIN AS AN ANTI-APOPTOTIC PROTEIN<br />

Nina C. Dempsey, Claire Hunter-Lavin, Michael J. Marshall, John H. H. Williams<br />

Chester <str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g> Stress <str<strong>on</strong>g>Research</str<strong>on</strong>g>, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biological Science, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Chester,<br />

Parkgate Road, Chester, United Kingdom, CH1 4BJ<br />

e-mail: n.dempsey@chester.ac.uk<br />

The role <str<strong>on</strong>g>of</str<strong>on</strong>g> OPG in b<strong>on</strong>e turnover through its interacti<strong>on</strong> with RANKL is well established, however this<br />

protein has now been shown to have anti-apoptotic properties which may c<strong>on</strong>tribute to the survival <str<strong>on</strong>g>of</str<strong>on</strong>g> cancer<br />

cells. As c<strong>on</strong>firmati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> this potential role we show data that OPG can interact with both TRAIL and TNFα<br />

using ELISA and BIAcore. We have then dem<strong>on</strong>strated that OPG will protect cells against TRAIL or TNFα<br />

induced apoptosis.<br />

We have then focused <strong>on</strong> the release <str<strong>on</strong>g>of</str<strong>on</strong>g> OPG under stress c<strong>on</strong>diti<strong>on</strong>s from MG63 cells and looked at the<br />

existence <str<strong>on</strong>g>of</str<strong>on</strong>g> OPG feedback mechanisms. We show that a number <str<strong>on</strong>g>of</str<strong>on</strong>g> stressors that induce apoptosis also<br />

result in an increase in the release <str<strong>on</strong>g>of</str<strong>on</strong>g> OPG from cells. These stressors include – UV exposure and exposure to<br />

TNFα or TRAIL. The increase in release <str<strong>on</strong>g>of</str<strong>on</strong>g> OPG occurs prior to indicators <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis – caspase 3<br />

activati<strong>on</strong> or Annexin V binding. Interestingly, inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis by elevated temperature coincides with<br />

a total cessati<strong>on</strong> in OPG release.<br />

Producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> OPG can be regulated by a feedback mechanism as release can be increased by lowering the<br />

ambient c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> OPG, and c<strong>on</strong>versely that by increasing this ambient c<strong>on</strong>centrati<strong>on</strong>, the release <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

OPG is inhibited. The changes are mirrored by alterati<strong>on</strong>s in gene expressi<strong>on</strong>.<br />

We discuss the potential role <str<strong>on</strong>g>of</str<strong>on</strong>g> OPG in the treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> tumours and strategies that can be used to bypass<br />

this protecti<strong>on</strong>.<br />

2A_02_P<br />

(poster secti<strong>on</strong> A2, poster board #76, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HYPEROSMOTIC STRESS RESPONSE<br />

Roberta R. Alfieri, Mara B<strong>on</strong>elli, Andrea Cavazz<strong>on</strong>i, Angelo F. Borghetti, Pier Giorgio Petr<strong>on</strong>ini<br />

Dipartimento di Medicina Sperimentale Università degli Studi di Parma Via Volturno 39 Parma<br />

e-mail: roberta.alfieri@unipr.it<br />

Many types <str<strong>on</strong>g>of</str<strong>on</strong>g> mammalian cells can survive a moderately hypert<strong>on</strong>ic envir<strong>on</strong>ment due to a specific adaptati<strong>on</strong><br />

process that results in the cellular accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> compatible osmolytes. This adaptati<strong>on</strong> process, involves<br />

early resp<strong>on</strong>ses, occurring over millisec<strong>on</strong>ds to minutes, and later resp<strong>on</strong>ses, requiring hours to days. The<br />

virtually instantaneous reducti<strong>on</strong> in cell volume due to the osmotic efflux <str<strong>on</strong>g>of</str<strong>on</strong>g> water induced by acute<br />

hypert<strong>on</strong>ic stress is rapidly corrected by what is referred to as regulatory volume increase (RVI). This early<br />

process is mediated by pre-existing i<strong>on</strong> transport systems that produce increases in the intracellular<br />

c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> potassium, sodium and chloride i<strong>on</strong>s, and the accompanying influx <str<strong>on</strong>g>of</str<strong>on</strong>g> water causes RVI.<br />

The later phase is characterised by increased producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock proteins (HSPs) and either the<br />

synthesis or the uptake and cellular accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> compatible osmolytes. In mammalian cells the latter<br />

include neutral amino acids or their derivatives, polyols such as sorbitol and myo-inositol, and methylamines<br />

such as betaine. The usual explanati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> this phenomen<strong>on</strong> is the need to replace the early cellular<br />

accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> inorganic i<strong>on</strong>s with small organic molecules that do not affect cell functi<strong>on</strong> even at relatively


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

high intracellular c<strong>on</strong>centrati<strong>on</strong>s. Accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> compatible osmolytes within the cell thus maintains<br />

intracellular water homeostasis without impairing normal biochemical functi<strong>on</strong>s such as protein synthesis.<br />

Cells do not adapt and die by apoptosis when hypert<strong>on</strong>ic medium has been depleted <str<strong>on</strong>g>of</str<strong>on</strong>g> these molecules.<br />

Compatible osmolytes thus enable cells to survive under hypert<strong>on</strong>ic c<strong>on</strong>diti<strong>on</strong>s, protecting them from<br />

apoptosis and modulating the adaptive resp<strong>on</strong>se.<br />

2A_04_P<br />

(poster secti<strong>on</strong> A2, poster board #77, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

GLUTATHIONE S-TRANSFERASE PI REGULATES UV-INDUCED JNK SIGNALING<br />

IN SH-SY5Y CELLS<br />

M. Castro-Caldas 1,2 , E. Rodrigues 1 , M. C. Lechner 1 , M. J. Gama 1<br />

1UBMBE. Faculdade de Farmácia, Universidade de Lisboa<br />

2SABT. Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa<br />

Activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> c-Jun N-terminal kinase (JNK) signaling pathway is a key event in neur<strong>on</strong>al apoptosis. Previous<br />

studies dem<strong>on</strong>strated that in SH-SY5Y cells UV-induced apoptosis is associated with activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> JNK. The<br />

cellular mechanisms underlying the c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> JNK activity be<str<strong>on</strong>g>for</str<strong>on</strong>g>e and immediately after stress are not<br />

completely understood. Under resting c<strong>on</strong>diti<strong>on</strong>s the basal activity <str<strong>on</strong>g>of</str<strong>on</strong>g> JNK is low, since JNK is kept inactive<br />

by the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> <strong>on</strong>e or more repressors. Inactivati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> JNK may be mediated by binding through proteinprotein<br />

interacti<strong>on</strong>s to n<strong>on</strong>-substrate proteins, including Glutathi<strong>on</strong>e S-Transferase pi (GSTpi). GSTpi<br />

bel<strong>on</strong>gs to a multigene family <str<strong>on</strong>g>of</str<strong>on</strong>g> isozymes catalyzing detoxificati<strong>on</strong> reacti<strong>on</strong>s. Although it has previously been<br />

shown that over expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> GSTpi protects cells from JNK-mediated apoptosis, the mechanisms<br />

underlying regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> JNK signaling by GSTpi in neur<strong>on</strong>al cells have never been described. In this work,<br />

SH-SY5Y cells were treated with UV to evaluate the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> JNK signaling by GSTpi. The relative<br />

c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> the GSTpi, p-JNK/JNK and apoptotic proteins were estimated by Western blot. Direct<br />

interacti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> GSTpi and JNK was determined by co-immunoprecipitati<strong>on</strong> assays. Evaluati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> GSTpi<br />

dimers and multimeric complexes <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> was per<str<strong>on</strong>g>for</str<strong>on</strong>g>med by SDS-PAGE under n<strong>on</strong>-reducing c<strong>on</strong>diti<strong>on</strong>s.<br />

Our results show that UV treatment induces apoptosis in SH-SY5Y cells and the transient activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> JNK.<br />

Furthermore, the increase <str<strong>on</strong>g>of</str<strong>on</strong>g> JNK enzymatic activity correlates with changes <str<strong>on</strong>g>of</str<strong>on</strong>g> GSTpi-JNK complexes and<br />

the c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> GSTpi multimer <str<strong>on</strong>g>for</str<strong>on</strong>g>ms. Taken together our results suggest that GSTpi may act as a<br />

regulator <str<strong>on</strong>g>of</str<strong>on</strong>g> the UV-induced cellular stress resp<strong>on</strong>se, c<strong>on</strong>trolling JNK activity by protein-protein interacti<strong>on</strong>s.<br />

2A_05_P<br />

(poster secti<strong>on</strong> A2, poster board #78, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE ROLE OF HSP72 IN NEURONAL CELL SURVIVAL AND THE RESISTANCE<br />

OF NEURONE-LIKE SH-SY5Y CELLS TO APOPTOTIC INSULTS<br />

L. Cheng 1* , D. J. Smith 1 , P. Nagley 1 , R. L. Anders<strong>on</strong> 2<br />

1 Dept <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry and Molecular Biology, M<strong>on</strong>ash University, Victoria, Australia<br />

2 Peter MacCallum Cancer <str<strong>on</strong>g>Centre</str<strong>on</strong>g>, Victoria, Australia<br />

*e-mail: Lesley.Cheng@med.m<strong>on</strong>ash.edu.au<br />

Heat shock proteins have been implicated in neur<strong>on</strong>al cell survival. We studied here the anti-apoptotic effects<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> heat-inducible hsp72 in the human SH-SY5Y neuroblastoma cell line, propagated in an undifferentiated<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g>m and able to be differentiated into neur<strong>on</strong>e-like cells using retinoic acid with brain-derived neurotrophic<br />

factor. Mild heat stress (43C <str<strong>on</strong>g>for</str<strong>on</strong>g> 30 min) was applied to induce hsp72, subsequently referred to as thermal<br />

79


23-26 August 2007,<br />

Budapest, Hungary<br />

pre-c<strong>on</strong>diti<strong>on</strong>ing treatment. It was observed that thermal pre-c<strong>on</strong>diti<strong>on</strong>ing protects cells against apoptosis<br />

induced by a subsequent treatment with staurosporine (50 nM). Neur<strong>on</strong>e-like SH-SY5Y cells displayed<br />

reduced Bax activati<strong>on</strong>, cytochrome c release and nuclear fragmentati<strong>on</strong> when thermally pre-c<strong>on</strong>diti<strong>on</strong>ed<br />

compared to n<strong>on</strong>-pre-c<strong>on</strong>diti<strong>on</strong>ed c<strong>on</strong>trol cells (all m<strong>on</strong>itored by immunocytochemistry and c<strong>on</strong>focal<br />

microscopy). The suggesti<strong>on</strong> that hsp72 may be involved in blocking apoptosis and that the block by hsp72<br />

may be upstream <str<strong>on</strong>g>of</str<strong>on</strong>g> Bax was tested by c<strong>on</strong>structing stable transfectants over-expressing hsp72 (5YH72.1).<br />

Such cells maintained levels <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp72 comparable to those seen in untransfected undifferentiated SH-SY5Y<br />

cells exposed to thermal pre-c<strong>on</strong>diti<strong>on</strong>ing; similar levels <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp72 were also found in neur<strong>on</strong>e-like<br />

untransfected cells without heat shock, c<strong>on</strong>diti<strong>on</strong>s that also induce hsp72. 5YH72.1 cells showed enhanced<br />

thermotolerance, at significantly higher temperatures than neur<strong>on</strong>e-like untransfected cells (themselves more<br />

thermotolerant than their undifferentiated counterparts). Moreover, neur<strong>on</strong>e-like 5YH72.1 cells treated with<br />

50 nM <str<strong>on</strong>g>of</str<strong>on</strong>g> staurosporine failed almost completely to display Bax activati<strong>on</strong> and nuclear fragmentati<strong>on</strong>.<br />

Undifferentiated 5YH72.1 cells were also protected but to a lesser extent against the molecular effects <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

staurosporine treatment. The data support the propositi<strong>on</strong> that hsp72 is resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> the thermoprotecti<strong>on</strong><br />

observed in SH-SY5Y cells. Further, hsp72 acts upstream <str<strong>on</strong>g>of</str<strong>on</strong>g> the mitoch<strong>on</strong>dria to prevent apoptosis in these<br />

cells when expressed in moderately high quantities.<br />

2A_06_P<br />

(poster secti<strong>on</strong> A2, poster board #79, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

UPREGULATION OF ANTI-APOPTOTIC BCL-2 BY A NOVEL ENDOPLASMIC<br />

RETICULUM PROTEIN SIGMA-1 RECEPTOR<br />

Teruo Hayashi, Tsung-Ping Su<br />

Intramural <str<strong>on</strong>g>Research</str<strong>on</strong>g> Program, NIDA, NIH, DHHS, 333 Cassell Drive, Baltimore, MD 21224<br />

e-mail: thayashi@intra.nida.nih.gov<br />

Extreme cellular stress induces apoptosis, a deliberate life-relinquishment <str<strong>on</strong>g>of</str<strong>on</strong>g> a cell. Bcl-2 serves as a powerful<br />

antidote against apoptotic cell death by preventing the mitoch<strong>on</strong>drial permeability transiti<strong>on</strong> that causes the<br />

release <str<strong>on</strong>g>of</str<strong>on</strong>g> caspase activators. Although the level <str<strong>on</strong>g>of</str<strong>on</strong>g> Bcl-2 has been shown to be altered under a variety <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

cellular stresses, signaling pathways that regulate the Bcl-2 expressi<strong>on</strong> are not fully understood. Sigma-1<br />

receptors (Sig-1Rs) are endoplasmic reticulum (ER) proteins that are activated by steroids and psychotropic<br />

drugs. Sig-1Rs express highly in the nervous system as well as in carcinomas. Activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Sig-1Rs is known<br />

to promote a robust neuroprotective acti<strong>on</strong>, whereas inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Sig-1Rs promotes cell death such as seen<br />

in cancer cells. Recent studies have proposed, there<str<strong>on</strong>g>for</str<strong>on</strong>g>e, the Sig-1R as a potential therapeutic target in<br />

treatments <str<strong>on</strong>g>of</str<strong>on</strong>g> ischemia and cancer. Here, we reported that the Sig-1R is a novel ER protein regulating Bcl-2<br />

expressi<strong>on</strong>. Sig-1Rs co-localized with Bcl-2 at the ER, but not at mitoch<strong>on</strong>dria in CHO cells. Overexpressi<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> Sig-1Rs significantly increased Bcl-2 proteins in mitoch<strong>on</strong>dria, whereas knockdown <str<strong>on</strong>g>of</str<strong>on</strong>g> Sig-1Rs by siRNAs<br />

caused a prominent downregulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Bcl-2. RT-PCR and Northern blotting revealed that knockdown <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Sig-1Rs downregulated the bcl-2 mRNA, indicating a transcripti<strong>on</strong>al activati<strong>on</strong> and/or mRNA degradati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

bcl-2 by Sig-1Rs. In keeping with these findings, knockdown <str<strong>on</strong>g>of</str<strong>on</strong>g> Sig-1Rs potentiated cell death when CHO<br />

cells were under apoptotic stimuli. Our findings suggest that the novel ER protein Sig-1R is intrinsically<br />

regulating the level <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>drial Bcl-2, and, thus, Sig-1R ag<strong>on</strong>ists such as pregnenol<strong>on</strong>e sulfate and<br />

DHEAS may exhibit cell protective acti<strong>on</strong>, a least in part, by increasing the expressi<strong>on</strong> and thus augmenting<br />

the anti-apoptotic activity <str<strong>on</strong>g>of</str<strong>on</strong>g> Bcl-2. (Sp<strong>on</strong>sored by IRP/NIDA/NIH/DHHS)<br />

80


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2A_07_P<br />

(poster secti<strong>on</strong> A2, poster board #80, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HTRA2 IS UP-REGULATED IN THE HEAT-STRESSED RAT TESTIS<br />

Tetsuo Hayashi, Nobuyuki Ishii, Toshiya Terao, Makoto Morozumi, Takumi Yamada<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Urology, Saitama Medical Center, Saitama Medical University, Kawagoe, Saitama, Japan<br />

Aim: The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> the present study was to elucidate the role <str<strong>on</strong>g>of</str<strong>on</strong>g> the high temperature requirement A 2 (HtrA2)<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> the germ cell loss in the heat-stressed testis.<br />

Methods: We examined the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HtrA2, caspase-9 activity and proteolytic activity <str<strong>on</strong>g>of</str<strong>on</strong>g> HtrA2 in the rat<br />

testis and their in vivo resp<strong>on</strong>ses to experimental cryptorchid treatment.<br />

Results: Northern analysis revealed the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HtrA2 mRNA peaked at days 1 and 7 after cryptorchid<br />

treatment. While the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HtrA2 mRNA was recognized in the spermatog<strong>on</strong>ium, spermatocytes and<br />

some spermatids in normal adult rat testis, the experimental cryptorchidism treatment resulted in a marked<br />

increase in its signal intensity in spermatocytes and some spermatids and the layers <str<strong>on</strong>g>of</str<strong>on</strong>g> spermatog<strong>on</strong>ium and<br />

early primary spermatocytes became negative at days 1 and 7 after the treatment. However, the<br />

spermatog<strong>on</strong>ium, Sertoli cells and interstitial cells appeared to have str<strong>on</strong>g intensities at days 14, 28 and 56<br />

after the treatment. Western analysis revealed the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HtrA2 protein peaked at days 2 and 28,<br />

Caspase-9 activity peaked at day 2 and HtrA2 proteolytic activity peaked at day 28. C<strong>on</strong>sequently, the first<br />

peak <str<strong>on</strong>g>of</str<strong>on</strong>g> HtrA2 mRNA expressi<strong>on</strong> was followed by the peak <str<strong>on</strong>g>of</str<strong>on</strong>g> caspase-9 activity and the sec<strong>on</strong>d peak was<br />

followed by the peak <str<strong>on</strong>g>of</str<strong>on</strong>g> proteolytic activity.<br />

C<strong>on</strong>clusi<strong>on</strong>: These findings suggest the probabilities that the heat stress results in germ cell death by caspaseindependent<br />

manner with the elevati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proteolytic activity <str<strong>on</strong>g>of</str<strong>on</strong>g> HtrA2 as well as caspase-dependent manner<br />

with the elevati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> caspase-9 activity.<br />

2A_08_P<br />

(poster secti<strong>on</strong> A2, poster board #81, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

FUNCTIONAL STUDIES ON UBIQUITIN RECEPTOR PROTEIN, HFAF1, IN<br />

STRESS RESPONSES<br />

Jejean Lee, Young-Mee Kim, K<strong>on</strong>g-Joo Lee<br />

Center <str<strong>on</strong>g>for</str<strong>on</strong>g> Cell Signaling and Drug Discovery <str<strong>on</strong>g>Research</str<strong>on</strong>g>, College <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmacy and Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Life & Pharmaceutical<br />

Sciences, Ewha Womans University, Seoul 120-750, South Korea<br />

e-mail: kjl@ewha.ac.kr<br />

Human Fas-associated protein, hFAF1, was identified as Fas-associating molecule and a member <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

apoptosis signaling complex. We identified that hFAF1 acts as a scaffolding protein by unveiling the<br />

interacting proteins and newly found N-terminal ubiquitin-associated (UBA) domain. N-terminal UBA<br />

domain was identified to recruit K48- and K63-linked polyubiquitinated proteins and plays a role to<br />

accumulate the ubiquitinating proteins as a ubiquitin receptor. One ubiquitin like (UBL) domain interacts<br />

with Hsp70 and negatively regulates its chaper<strong>on</strong>e activity. C-terminal ubiquitin regulatory X (UBX) domain<br />

was identified to interact with AAA ATPase p97/VCP which is involved in the ubiquitin-proteasome<br />

pathway. These interacti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> hFAF1 to two chaper<strong>on</strong>e proteins (Hsp70 and VCP) suggest that FAF1 as a<br />

ubiquitin receptor plays important roles in stress resp<strong>on</strong>se and apoptotic cell death. In this study, we will<br />

dem<strong>on</strong>strate the regulati<strong>on</strong> mechanism by examining the post-translati<strong>on</strong>al modificati<strong>on</strong>s using proteomic<br />

tools, and the biological functi<strong>on</strong>s by examining the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> VCP binding defect mutants and RNAi <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

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23-26 August 2007,<br />

Budapest, Hungary<br />

hFAF1 in resp<strong>on</strong>se to stress. This can suggest the biological role and regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ubiquitin receptor hFAF1<br />

in stress-induced ubiquitin-proteasome system.<br />

2A_09_P<br />

(poster secti<strong>on</strong> A2, poster board #82, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PPARγ AGONISTS AND HSP70 RENDERS THE RESISTANCE TO APOPTOSIS IN γ-<br />

IRRADIATED CANCER CELLS<br />

Piotr Pierzchalski 1 , Agata Krawiec 1 , Wiesław Pawlik 1 , Maciej G<strong>on</strong>ciarz 2<br />

1Jagiell<strong>on</strong>ian University Medical College, Grzegórzecka 16, 31-531 Kraków, POLAND<br />

2 Endoscopy Unit, St. Barbara District Hospital, Plac Medyków 1, 41-200 Sosnowiec, POLAND<br />

The involvement <str<strong>on</strong>g>of</str<strong>on</strong>g> peroxisome proliferator-activated receptors (PPARs) in the cancer cell apoptosis is a<br />

generally accepted fact. However, some reports indicate that the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> PPARγ is directly resp<strong>on</strong>sible<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> carcinogenesis.<br />

It is well known that the high level <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock proteins (HSPs) in cancer cells is associated with metastasis,<br />

the poor prognosis and the resistance to radio as well as chemotherapy. HSP70 as a part <str<strong>on</strong>g>of</str<strong>on</strong>g> the most<br />

important systems <str<strong>on</strong>g>for</str<strong>on</strong>g> maintaining the viability <str<strong>on</strong>g>of</str<strong>on</strong>g> the cell, is known to counteract against the apoptosis. We<br />

report here the involvement <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 in anti-apoptotic acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> activated PPARγ in γ-irradiated human<br />

col<strong>on</strong> cancer cells.<br />

We have used Caco-2 cells (human col<strong>on</strong> adenocarcinoma) as an experimental model. In this system PPAR-γ<br />

ag<strong>on</strong>ists induced nuclear translocati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> PPAR-γ as well as HSF-1. This translocati<strong>on</strong> was followed by the<br />

increase <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 mRNA and protein expressi<strong>on</strong>.<br />

Cells subjected to γ-radiati<strong>on</strong> (phot<strong>on</strong>s) with therapeutic dose <str<strong>on</strong>g>of</str<strong>on</strong>g> 2,5 Gy, manifested pattern <str<strong>on</strong>g>of</str<strong>on</strong>g> PARP<br />

degradati<strong>on</strong> typical <str<strong>on</strong>g>for</str<strong>on</strong>g> apoptosis, showing both the native 112 KD and digested 85 KD <str<strong>on</strong>g>for</str<strong>on</strong>g>ms. It suggests<br />

activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> caspases 3 or 6. Stimulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the cultures with PPARγ ag<strong>on</strong>ists prior to the irradiati<strong>on</strong><br />

eliminated altogether the process <str<strong>on</strong>g>of</str<strong>on</strong>g> PPAR-γ nuclear translocati<strong>on</strong> and PARP degradati<strong>on</strong>. PPARγ remained<br />

in the complexes with AKT-1 in cytoplasmic as well as in nuclear pool. However this treatment did not<br />

affect HSF-1 translocati<strong>on</strong> and HSP70 expressi<strong>on</strong>.<br />

According to our elucidati<strong>on</strong>, in γ-irradiated cells nuclear translocati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> PPARγ is abolished and PPARγ-<br />

AKT-1 complexes are c<strong>on</strong>served in which PPARγ remains insensitive <str<strong>on</strong>g>for</str<strong>on</strong>g> its ag<strong>on</strong>ists treatment. Most likely,<br />

at the same time PPARγ ag<strong>on</strong>ists directly activate HSP 70. The process is undisturbed by the γ-irradiati<strong>on</strong><br />

what renders the col<strong>on</strong> cancer cells resistance to apoptosis.<br />

82


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2A_10_P<br />

(poster secti<strong>on</strong> A2, poster board #82, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

RELATIONSHIP BETWEEN APOPTOSIS AND HSP70 EXPRESSION IN<br />

LYMPHOCYTES EXPOSED TO STRESS MEDIATORS<br />

A. M. Sapozhnikov, E. I. Kovalenko, D. A. Murashko, E. A. Alekperov, O. A. Shustova<br />

Shemyakin & Ovchinnikov Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Bioorganic Chemistry, Moscow, Russia<br />

e-mail: amsap@mail.ru<br />

It has been shown that adrenaline in c<strong>on</strong>trast to glucocorticoids can activate immune system. However<br />

existing data indicate that catecholamines like glucocorticoids induce apoptosis in lymphocytes. Earlier we<br />

dem<strong>on</strong>strated that lymphoid cell apoptosis is accompanied by increase <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP expressi<strong>on</strong>. In this work we<br />

carried out a study <str<strong>on</strong>g>of</str<strong>on</strong>g> effect <str<strong>on</strong>g>of</str<strong>on</strong>g> adrenaline and dexamethas<strong>on</strong>e <strong>on</strong> apoptosis and HSP70 expressi<strong>on</strong> in cultures<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> mouse lymphocytes. Obtained results c<strong>on</strong>firmed apoptogenic effect <str<strong>on</strong>g>of</str<strong>on</strong>g> catecholamine and dexamethas<strong>on</strong>e.<br />

However, the stress mediators had opposite effects <strong>on</strong> level <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 expressi<strong>on</strong>: adrenaline enhanced<br />

intracellular c<strong>on</strong>tent <str<strong>on</strong>g>of</str<strong>on</strong>g> the protein whereas dexamethas<strong>on</strong>e had small but significant inhibitory acti<strong>on</strong> <strong>on</strong><br />

HSP70 expressi<strong>on</strong>. The catecholamine-induced increase <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 expressi<strong>on</strong> was mediated by α-<br />

adrenoreceptor because the effect was suppressed by phospholipase C inhibitor but was not decreased by<br />

PKA inhibitor. In additi<strong>on</strong>, we have revealed that α-adrenergic antag<strong>on</strong>ist prazosin reduce adrenaline-induced<br />

HSP70 expressi<strong>on</strong> while β-adrenergic antag<strong>on</strong>ist propranolol decrease the apoptogenic effect <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

catecholamine. Phenylephrine – specific α-adrenergic ag<strong>on</strong>ist had no apoptogenic effect in c<strong>on</strong>trast to<br />

adrenaline interacting with both α- and β-adrenoreceptors. We suppose that the difference between<br />

immunomodulating activity <str<strong>on</strong>g>of</str<strong>on</strong>g> catecholamines and glucocorticoids may be c<strong>on</strong>nected with their opposite<br />

influence <strong>on</strong> lymphocyte HSP system. The results suggest that effect <str<strong>on</strong>g>of</str<strong>on</strong>g> catecholamines <strong>on</strong> apoptosis and<br />

HSP70 expressi<strong>on</strong> are initiated by the same horm<strong>on</strong>al signal perceived by means <str<strong>on</strong>g>of</str<strong>on</strong>g> different adrenoreceptors.<br />

2A_11_P<br />

(poster secti<strong>on</strong> A2, poster board #83, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EXPRESSION OF HEAT SHOCK PROTEINS IN PRIMARY PORCINE MYOTUBES<br />

EXPOSED TO STRESSORS<br />

I. K. Straadt 1* , J. F. Young 1 , H. C. Bertram 1 , N. Gregersen 2 , P. Bross 2 , N. Oksbjerg 1<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Aarhus, 1 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Food Science, 2 <str<strong>on</strong>g>Research</str<strong>on</strong>g> Unit <str<strong>on</strong>g>for</str<strong>on</strong>g> Molecular Medicine, DK<br />

*e-mail: ida.straadt@agrsci.dk<br />

Stress exposure to animals prior to slaughter is causing big variati<strong>on</strong>s in the meat quality. Using primary<br />

porcine myotubes as a model, exposure to different stressors was investigated. When myotubes were exposed<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> 1 h to 50-250µM H 2 O 2 the peak in expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> mRNA <str<strong>on</strong>g>for</str<strong>on</strong>g> HSP70 and HO1, measured 18 h after stress<br />

exposure, was seen at a c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 200 µM H 2 O 2 . At 250 µM the expressi<strong>on</strong> decreased substantially,<br />

indicating that the cells were no l<strong>on</strong>ger capable <str<strong>on</strong>g>of</str<strong>on</strong>g> expressing heat shock proteins at a high level. Also the<br />

appearance <str<strong>on</strong>g>of</str<strong>on</strong>g> the cells evidenced the toxic effect <str<strong>on</strong>g>of</str<strong>on</strong>g> H 2 O 2 levels above 200 µM. When testing the viability<br />

this was c<strong>on</strong>firmed as the survival <str<strong>on</strong>g>of</str<strong>on</strong>g> myotubes exposed to more than 100 µM H 2 O 2 <str<strong>on</strong>g>for</str<strong>on</strong>g> 1 h showed a<br />

decreased cell viability determined by WST-1. Based <strong>on</strong> these observati<strong>on</strong>s a level <str<strong>on</strong>g>of</str<strong>on</strong>g> 100 µM H 2 O 2 was<br />

chosen <str<strong>on</strong>g>for</str<strong>on</strong>g> investigating the development in expressi<strong>on</strong> levels <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock proteins over a time period in the<br />

myotubes. A significant increase in expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> both HSP70 and HO1 mRNA was observed after exposure<br />

83


23-26 August 2007,<br />

Budapest, Hungary<br />

to H 2 O 2 <str<strong>on</strong>g>for</str<strong>on</strong>g> 1 h and measuring expressi<strong>on</strong> up to 18 h after exposure. No change in the cell viability<br />

determined by WST-1 was observed when the myotubes were exposed to heat (42°C or 45°C) <str<strong>on</strong>g>for</str<strong>on</strong>g> 1 h.<br />

However, comparing cells exposed to 42°C and 45°C the expressi<strong>on</strong> was approximately 10 times higher when<br />

exposed to 45°C. No change in the expressi<strong>on</strong> level <str<strong>on</strong>g>of</str<strong>on</strong>g> HO1 mRNA was observed when exposed to heat.<br />

Hence, primary porcine cells are more sensitive to H 2 O 2 compared to myotubes and muscle cell lines from<br />

other species. In c<strong>on</strong>trast to this, primary porcine muscle cells are not very sensitive to heat stress when<br />

determining the cell viability by WST-1.<br />

2A_12_P<br />

(poster secti<strong>on</strong> A2, poster board #84, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

APOPTOSIS AND HEAT SHOCK PROTEIN HSP70 IN AGED MAMMALIAN<br />

OOCYTES<br />

P. Esp<strong>on</strong>da 1 , H. Díaz 2<br />

1Centro de Investigaci<strong>on</strong>es Biológicas, CSIC, Madrid, Spain<br />

2Facultad de Ciencias, Universidad de Valparaíso, Valparaíso, Chile<br />

The fertile life <str<strong>on</strong>g>of</str<strong>on</strong>g> the mammalian is short and a few hours after ovulati<strong>on</strong> it loses its capacity to be fertilized.<br />

Ageing affect oocyte fertilizing ability and aged females produce numerous infertile and/or abnormal oocytes.<br />

We used normal and aged mouse oocytes to analyze initial apoptosis (using Anexin V), late apoptosis (using<br />

the TUNEL method) and the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress protein HSP70 (using antibodies and RT-PCR). We<br />

induced the ovulati<strong>on</strong> by horm<strong>on</strong>es (PMSG-HCG) to collect oocytes from young females (3 m<strong>on</strong>ths old).<br />

Gametes were also aged in the oviduct after ovulati<strong>on</strong> during 20 hours (postovulatory ageing) and other<br />

oocytes were collected from 12-15 m<strong>on</strong>ths old females (preovulatory ageing).<br />

Anexin V showed that a high percentage (39.2%) <str<strong>on</strong>g>of</str<strong>on</strong>g> initial apoptosis appeared in oocytes ovulated by old<br />

females, and in oocytes aged in the oviduct during 20 hours (31.6%). TUNEL results showed that the high<br />

percentages (56.5%) <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis occurred when oocytes were aged in the oviduct during 20 hours. Oocytes<br />

from young or old females showed a similar degree <str<strong>on</strong>g>of</str<strong>on</strong>g> damage (12.9-16.1%). Nevertheless, young oocytes<br />

recovered from females that were not treated with horm<strong>on</strong>es, show a lower percentage <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis (3.7%).<br />

TUNEL procedure was also applied to histological secti<strong>on</strong>s from ovaries <str<strong>on</strong>g>of</str<strong>on</strong>g> different ages (3; 11; 14 and 24<br />

m<strong>on</strong>ths old). Results showed that apoptotic cells were progressively increased in the insterticial tissue <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

organ.<br />

Normal oocytes do not showed the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress protein HSP70, but this protein appeared in the<br />

cytoplasm after an exposure to 40ºC during 4 hours, and in oocytes after postovulatory ageing. RT-PCR<br />

showed HSP70 gene expressi<strong>on</strong> in aged oocytes.<br />

These results indicated that ageing increase apoptosis <str<strong>on</strong>g>of</str<strong>on</strong>g> the mouse oocyte and <str<strong>on</strong>g>of</str<strong>on</strong>g> the ovarian tissue.<br />

Apoptosis and HSP70 are principally produced by envir<strong>on</strong>mental c<strong>on</strong>diti<strong>on</strong>s, as the l<strong>on</strong>g storage <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

oocyte in the oviduct or the employ <str<strong>on</strong>g>of</str<strong>on</strong>g> horm<strong>on</strong>es used to induce ovulati<strong>on</strong>.<br />

84


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2A_13_P<br />

(poster secti<strong>on</strong> A2, poster board #85, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STRESS HORMONES INDUCE IN CANCER CELLS ATP DEPLETION VIA<br />

PERTURBATION OF OXIDATIVE PHOSPHORYLATION<br />

Eliezer Flescher<br />

Tel Aviv University, Tel Aviv 69978 Israel, e-mail: flascher@post.tau.ac.il<br />

The jasm<strong>on</strong>ate family <str<strong>on</strong>g>of</str<strong>on</strong>g> plant stress horm<strong>on</strong>es exhibits selective cytotoxic activities towards cancer cell lines<br />

as well as leukemic cells from chr<strong>on</strong>ic lymphocytic leukemia patients. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this project was to elucidate<br />

the jasm<strong>on</strong>ate mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> acti<strong>on</strong>. We found that jasm<strong>on</strong>ates induce a rapid depleti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ATP in cancer<br />

cells, preceding any signs <str<strong>on</strong>g>of</str<strong>on</strong>g> cell death. Furthermore, we found a positive correlati<strong>on</strong> between the<br />

susceptibility <str<strong>on</strong>g>of</str<strong>on</strong>g> a given cell type to the cytotoxic effect <str<strong>on</strong>g>of</str<strong>on</strong>g> jasm<strong>on</strong>ates and the degree <str<strong>on</strong>g>of</str<strong>on</strong>g> ATP depleti<strong>on</strong><br />

induced in that cell. The two major sources <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular ATP, oxidative phosphorylati<strong>on</strong> and glycolysis,<br />

determine the steady state levels <str<strong>on</strong>g>of</str<strong>on</strong>g> ATP. Experiments using modulators <str<strong>on</strong>g>of</str<strong>on</strong>g> ATP synthesis via glycolysis or<br />

oxidative phosphorylati<strong>on</strong> suggest that the latter is the pathway suppressed by jasm<strong>on</strong>ates. C<strong>on</strong>sequently, the<br />

direct effects <str<strong>on</strong>g>of</str<strong>on</strong>g> jasm<strong>on</strong>ates <strong>on</strong> mitoch<strong>on</strong>dria were evaluated. Jasm<strong>on</strong>ates induced cytochrome c release and<br />

swelling in mitoch<strong>on</strong>dria isolated from cancer cells but not from normal <strong>on</strong>es. Thus, the selectivity <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

jasm<strong>on</strong>ates against cancer cells is rooted at the mitoch<strong>on</strong>drial level, and probably exploits differences between<br />

mitoch<strong>on</strong>dria from normal versus cancer cells. The permeability transiti<strong>on</strong> pore complex (PTPC) regulates<br />

movement <str<strong>on</strong>g>of</str<strong>on</strong>g> compounds across the mitoch<strong>on</strong>drial membrane. Abnormally l<strong>on</strong>g opening <str<strong>on</strong>g>of</str<strong>on</strong>g> this pore can be<br />

associated with cytochrome c escape into the cytosol, resulting eventually in cell death. Jasm<strong>on</strong>ate-induced<br />

release <str<strong>on</strong>g>of</str<strong>on</strong>g> cytochrome c from mitoch<strong>on</strong>dria isolated from cancer cells was inhibited by inhibitors <str<strong>on</strong>g>of</str<strong>on</strong>g> PTPC<br />

opening, suggesting that the mitoch<strong>on</strong>drial permeability transiti<strong>on</strong> induced by jasm<strong>on</strong>ates is PTPC-mediated.<br />

These findings positi<strong>on</strong> jasm<strong>on</strong>ates as promising anti-cancer drugs acting via energetic depleti<strong>on</strong> in neoplastic<br />

cells.<br />

2A_14_P<br />

(poster secti<strong>on</strong> A2, poster board #86, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

OXIDATIVE STRESS AND APOPTOSIS IN CARCINOGENESIS: MOLECULAR<br />

PLAYERS AND INTERACTIONS<br />

Claudia Campanella 1 , Nella M. Ardizz<strong>on</strong>e 1 , Ant<strong>on</strong>ella M<strong>on</strong>talbano 1 , Ant<strong>on</strong>ella Marino Gammazza 1 ,<br />

Anna Ribbene 1 , Anna M. Czarnecka 2 , Valentina Di Felice 1 , Marianna Bellafiore 1 , Giovanni Zummo 1 ,<br />

Francesco Cappello 1 , Everly C<strong>on</strong>way de Macario 3 , Alberto J. L. Macario 3<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Experimental Medicine, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Palermo, Italy<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Genetics, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Warsaw, Poland<br />

3Center <str<strong>on</strong>g>of</str<strong>on</strong>g> Marine Biotechnology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Maryland, Baltimore, MD, USA<br />

Oxidative stress (OS) and apoptosis are major determinants <str<strong>on</strong>g>of</str<strong>on</strong>g> cell fate in normal and tumor cells. The<br />

c<strong>on</strong>necti<strong>on</strong> between OS and apoptosis is not fully understood; the molecular networks involved have not<br />

been elucidated yet. Our work’s objectives are to identify network comp<strong>on</strong>ents and determine their<br />

interacti<strong>on</strong>s and the c<strong>on</strong>sequences <str<strong>on</strong>g>of</str<strong>on</strong>g> the interacti<strong>on</strong>s in tumor cells. The l<strong>on</strong>g-term goal is to understand at<br />

the molecular level the c<strong>on</strong>tributi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> OS and apoptosis to carcinogenesis and thus identify precise targets<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> anti-tumor therapy. We used a tumour cell line (NCI-H292 human airway mucoepidermoid carcinoma)<br />

85


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Budapest, Hungary<br />

and measured, be<str<strong>on</strong>g>for</str<strong>on</strong>g>e and after OS inducti<strong>on</strong> with various doses <str<strong>on</strong>g>of</str<strong>on</strong>g> H 2 O 2 , molecules known or suspected to<br />

participate in the cell’s resp<strong>on</strong>se to OS, including apoptosis: Hsp60, Hsp10, Hsp70, p53, procaspase C3 (p-<br />

C3), caspase 3 (C3), and p21. Various complementary methods were applied to assess cell viability and<br />

apoptosis, and to determine the cellular localizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the molecules investigated and their quantities at the<br />

levels <str<strong>on</strong>g>of</str<strong>on</strong>g> mRNA and protein. Occurrence <str<strong>on</strong>g>of</str<strong>on</strong>g> the bimolecular complexes Hsp60/p-C3 and Hsp60/p53 was<br />

also investigated be<str<strong>on</strong>g>for</str<strong>on</strong>g>e and after OS inducti<strong>on</strong>. Several findings were made; most c<strong>on</strong>spicuous <str<strong>on</strong>g>of</str<strong>on</strong>g> all were the<br />

presence <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp60/p-C3 complexes and the lack <str<strong>on</strong>g>of</str<strong>on</strong>g> C3 after OS inducti<strong>on</strong> despite cell progressi<strong>on</strong> through<br />

apoptosis. The link between Hsp60/p-C3 complex <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> and C3 absence is currently under study.<br />

2A_15_P<br />

(poster secti<strong>on</strong> A2, poster board #87, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

INTERACTIONS BETWEEN IMPORTANT REGULATORY PROTEINS AND THE<br />

STRESS RESPONSE PROTEIN, ΑB CRYSTALLIN<br />

Joy G. Ghosh † , Ananth K. Shenoy Jr. † , John I. Clark †‡<br />

Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Biological Structure † & Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Ophthalmology ‡ , Univ. <str<strong>on</strong>g>of</str<strong>on</strong>g> Washingt<strong>on</strong>, Seattle, WA 98195, USA<br />

αB crystallin, the archetype <str<strong>on</strong>g>of</str<strong>on</strong>g> small heat shock proteins (sHSPs), is an ‘unfolding resp<strong>on</strong>se protein’ that<br />

recognizes, binds, and stabilizes structurally compromised proteins during or after stress. Protein pin arrays<br />

identified interactive sequences in αB crystallin <str<strong>on</strong>g>for</str<strong>on</strong>g> twelve regulatory proteins including EGF, FGF-2, IGF-1,<br />

NGF-β, TGF-β, VEGF, insulin, β-catenin, caspase-3, caspase-8, Bcl-2, and Bcl-x L , which are important in<br />

cellular differentiati<strong>on</strong>, proliferati<strong>on</strong>, signalling, and apoptosis. Seven αB crystallin sequences had str<strong>on</strong>g<br />

interacti<strong>on</strong>s with FGF-2, NGF-β, VEGF, insulin, and β-catenin and mapped to the β3-β8-β9 interface and<br />

surface domains in the N- and C-termini. This is the first report in which interactive sequences <str<strong>on</strong>g>for</str<strong>on</strong>g> regulatory<br />

proteins were identified in a sHSP. The remaining seven proteins did not interact with αB crystallin. The αB<br />

crystallin interactive sequences <str<strong>on</strong>g>for</str<strong>on</strong>g> regulatory proteins overlap with sequences <str<strong>on</strong>g>for</str<strong>on</strong>g> complex assembly,<br />

chaper<strong>on</strong>e activity, and filament stabilizati<strong>on</strong>. The positi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the interactive sequences in the structure <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

αB crystallin complex is c<strong>on</strong>sistent with a dynamic functi<strong>on</strong>al mechanism to coordinate exposure <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

interactive surfaces <strong>on</strong> αB crystallin <str<strong>on</strong>g>for</str<strong>on</strong>g> the recogniti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> target proteins. Taken together, the results suggest<br />

the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> sHSPs includes the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> key comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular differentiati<strong>on</strong>, proliferati<strong>on</strong>,<br />

signalling, and apoptotic pathways in stress and n<strong>on</strong>-stress c<strong>on</strong>diti<strong>on</strong>s.<br />

86


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2A_16_P<br />

(poster secti<strong>on</strong> A2, poster board #88, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HEAT SHOCK PROTEIN 70 STABILIZES LYSOSOMAL MEMBRANES THROUGH<br />

BINDING TO THE LYSOSOMAL LIPID LBPA/BMP<br />

Thomas Kirkegaard-Sørensen 1 , Jesper Nylandsted 1 , Irina Moilanen 2 , Paavo Kinnunen 2 , Anke Roth 5 ,<br />

Carmen Garrido 3 , Jean Gruenberg 4 , K<strong>on</strong>rad Sandh<str<strong>on</strong>g>of</str<strong>on</strong>g>f 6 , Marja Jäättelä 1<br />

1Danish Cancer Society, Denmark,<br />

2University <str<strong>on</strong>g>of</str<strong>on</strong>g> Helsinki, Finland,<br />

3INSERM U517, Faculté de Pharmacie, France,<br />

4 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Geneva, Switzerland,<br />

5Institut für Chemie, Humboldt Universität zu Berlin, Germany,<br />

6Kekule-Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Organic Chemistry and Biochemistry, University <str<strong>on</strong>g>of</str<strong>on</strong>g> B<strong>on</strong>n, Germany<br />

The major stress-inducible Heat shock protein 70 (Hsp70) is a potent survival protein that c<strong>on</strong>fers<br />

cytoprotecti<strong>on</strong> against numerous death-inducing stimuli. In cancer cells a fracti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 localizes to the<br />

lysosomal membranes and this localizati<strong>on</strong> associates with a cytoprotective effect. Here we provide direct<br />

evidence <str<strong>on</strong>g>for</str<strong>on</strong>g> an interacti<strong>on</strong> between Hsp70 and the lysosomal membrane lipid LBPA/BMP. By targeting<br />

rHsp70 to the lysosomes via endocytosis and challenging the integrity <str<strong>on</strong>g>of</str<strong>on</strong>g> lysosomal membranes with<br />

photooxidati<strong>on</strong> and anti-cancer drugs, we show that lysosomes loaded with rHsp70 showed a marked<br />

resistance towards all these stimuli. The interacti<strong>on</strong> between rHsp70 and lysosomal membranes is mediated<br />

through a pH-dependent, high-affinity binding to the ani<strong>on</strong>ic lysosomal lipid lysobisphosphatidic<br />

acid/ Bis(m<strong>on</strong>o-acylglycero)phosphate (LBPA/BMP). LBPA/BMP is a lipid co-factor <str<strong>on</strong>g>for</str<strong>on</strong>g> the enzyme acid<br />

sphingomyelinase (aSMase) and the binding <str<strong>on</strong>g>of</str<strong>on</strong>g> rHsp70 to LBPA/BMP directly antag<strong>on</strong>izes its activity,<br />

providing an explanati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> the cytoprotective effect <str<strong>on</strong>g>of</str<strong>on</strong>g> lysosome-associated Hsp70. Remarkably, an<br />

antibody towards LBPA/BMP reverses the protective effect <str<strong>on</strong>g>of</str<strong>on</strong>g> rHsp70, hereby revealing a possible target <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

future cancer therapy.<br />

2A_17_P<br />

(poster secti<strong>on</strong> A2, poster board #89, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HEAT SHOCK PROTEIN 70 PEPTIDE APTAMERS: A NOVEL APPROACH FOR<br />

ANTI-CANCER CHEMOTHERAPY<br />

Anne-Laure Rérole, Elise Schmitt, Marc Bickle, Guido Kroemer, Pierre Colas, Carmen Garrido<br />

INSERM U-866, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine and Pharmacy, 21033 Dij<strong>on</strong>, France<br />

e-mail: cgarrido@u-bourgogne.fr, alaure.rerole@gmail.com<br />

C<strong>on</strong>stitutively high HSP expressi<strong>on</strong> is a property <str<strong>on</strong>g>of</str<strong>on</strong>g>, and essential <str<strong>on</strong>g>for</str<strong>on</strong>g> the survival <str<strong>on</strong>g>of</str<strong>on</strong>g> at least some cancers.<br />

Neutralizing HSPs is there<str<strong>on</strong>g>for</str<strong>on</strong>g>e an attractive strategy <str<strong>on</strong>g>for</str<strong>on</strong>g> anti-cancer therapy. Clinical trials using specific<br />

HSP90 inhibitor such as 17AAG are currently being per<str<strong>on</strong>g>for</str<strong>on</strong>g>med with encouraging results. Similarly, we and<br />

other groups have reported that HSP70 anti-sense c<strong>on</strong>structs or a c<strong>on</strong>struct bearing the HSP70 binding<br />

domain <str<strong>on</strong>g>of</str<strong>on</strong>g> AIF, have chemosensitizing properties, and may even kill cancer cell lines (in the c<strong>on</strong>text <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

adenoviral infecti<strong>on</strong>). With the objective <str<strong>on</strong>g>of</str<strong>on</strong>g> obtaining small molecules that inhibit HSP70, we have selected a<br />

collecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> eighteen peptide aptamers (with variable regi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> 8 or 13 aminoacids) <str<strong>on</strong>g>for</str<strong>on</strong>g> their ability to bind<br />

HSP70 in a yeast two-hybrid screening. Two <str<strong>on</strong>g>of</str<strong>on</strong>g> these peptide aptamers (A8 and A18) str<strong>on</strong>gly increased the<br />

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Budapest, Hungary<br />

sensitivity <str<strong>on</strong>g>of</str<strong>on</strong>g> the cells to apoptosis induced by cisplatin in vitro and in vivo (mouse melanoma model). These<br />

two aptamers associated with the ATP-binding domain <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 and show a str<strong>on</strong>g specificity <str<strong>on</strong>g>for</str<strong>on</strong>g> their<br />

target since their chemosensitizing effect was lost in cells in which inducible HSP70 genes had been deleted.<br />

In vivo, A8 and A18 displayed anti-tumorigenic properties in the absence <str<strong>on</strong>g>of</str<strong>on</strong>g> additi<strong>on</strong>al treatment. This antitumor<br />

effect is most probably related to the inflitrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> T lymphocytes and macrophages observed in the<br />

tumors expressing these aptamers. Altogether, these data indicate the potential interest <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 peptide<br />

aptamers as lead compounds <str<strong>on</strong>g>for</str<strong>on</strong>g> the development <str<strong>on</strong>g>of</str<strong>on</strong>g> a novel type <str<strong>on</strong>g>of</str<strong>on</strong>g> anti-cancer agent.<br />

88


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2B. MEMBRANE-REGULATION OF THE STRESS RESPONSE<br />

(LÁSZLÓ VÍGH, JOHN L. HARWOOD)<br />

2B_01_S<br />

CELL SIGNALLING OF STRESS VIA CERAMIDE AND ITS METABOLITES<br />

Ant<strong>on</strong>io Gómez-Muñoz, Patricia Gangoiti, María H. Granado, M<strong>on</strong>ika G<strong>on</strong>zález<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry and Molecular Biology, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Science and Technology,<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> the Basque Country, 48080-Bilbao, Spain<br />

e-mail: ant<strong>on</strong>io.gomez@ehu.es<br />

Cellular stress has been defined as the threat <str<strong>on</strong>g>of</str<strong>on</strong>g> damage to macromolecules. Since many lipids, enzymes and<br />

signalling pathways c<strong>on</strong>tribute to the cellular stress resp<strong>on</strong>se it is necessary to identify the key players that are<br />

located at major nodes within the stress resp<strong>on</strong>se network. Many types <str<strong>on</strong>g>of</str<strong>on</strong>g> stresses including UV or i<strong>on</strong>izing<br />

radiati<strong>on</strong>, oxidative stress, chemotherapeutic drugs, or starvati<strong>on</strong> cause DNA or protein damage. This can<br />

result in growth arrest, apoptosis, or inflammatory resp<strong>on</strong>ses. One <str<strong>on</strong>g>of</str<strong>on</strong>g> the mechanisms involved in these<br />

acti<strong>on</strong>s is the sphingomyelin pathway. Ceramide, the central molecule in this pathway, is an important sec<strong>on</strong>d<br />

messenger that engages different downstream effectors depending <strong>on</strong> the c<strong>on</strong>comitant activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> other<br />

sec<strong>on</strong>d messengers and the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> enzymes that c<strong>on</strong>vert ceramide to other related metabolites such as<br />

sphingosine, sphingosine 1-phosphate (S1P) or ceramide 1-phosphate (C1P). Whilst ceramide is proapoptotic<br />

and can induce cell cycle arrest, S1P or C1P are anti-apoptotic and have mitogenic properties.<br />

Ceramide and C1P can be interc<strong>on</strong>verted in cells by kinase and phosphatase activities. An appropriate balance<br />

between the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> these metabolites is crucial <str<strong>on</strong>g>for</str<strong>on</strong>g> cell and tissue homeostasis. Switching this balance<br />

towards accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> <strong>on</strong>e or the other can result in metabolic dysfuncti<strong>on</strong> or disease. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, the<br />

activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the enzymes that are involved in C1P and ceramide metabolism must be efficiently coordinated to<br />

ensure normal cell functi<strong>on</strong>ing.<br />

2B_02_S<br />

TEMPERATURE STRESS: REACTING AND ADAPTING – LESSONS FROM<br />

POIKILOTHERMS<br />

John L. Harwood<br />

School <str<strong>on</strong>g>of</str<strong>on</strong>g> Biosciences, Cardiff University, Cardiff CF10 3US, UK<br />

Abstract: Acanthamoeba castellanii is a comm<strong>on</strong> soil- or water-born protozo<strong>on</strong> that feeds <strong>on</strong> bacteria by<br />

phagocytosis. A. castellanii can grow between 4 and 32°C and has to adapt quickly to chilling in order to<br />

survive. We have identified a ∆ 12-fatty acid desaturase as key to low temperature adaptati<strong>on</strong>. The activity <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

this enzyme is mainly increased through gene expressi<strong>on</strong> and new protein synthesis. Interestingly, the activity<br />

can also be altered independently by dissolved oxygen levels. In additi<strong>on</strong>, we have identified a gene <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

∆ 12-desaturase which, when expressed in yeast, catalyses ∆ 15-desaturati<strong>on</strong> also. Moreover, it is also capable<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> producing very unusual n-1 polyunsaturated products.<br />

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23-26 August 2007,<br />

Budapest, Hungary<br />

2B_03_S<br />

MULTIPLE SIGNALLING PATHWAYS ACTIVATED BY HEAT SHOCK<br />

Jacques Landry<br />

<str<strong>on</strong>g>Centre</str<strong>on</strong>g> de recherche en cancérologie de l’Université Laval, L’Hôtel-Dieu de Québec, Québec, Canada, G1R 2J6,<br />

e-mail: jacques.landry@med.ulaval.ca<br />

Mild heat shock (HS) activates numerous signalling pathways the functi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> which either in survival or<br />

death processes are poorly understood. The origin(s) <str<strong>on</strong>g>of</str<strong>on</strong>g> the signal is(are) also undefined. Changes in protein<br />

c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> is a likely target <str<strong>on</strong>g>of</str<strong>on</strong>g> the acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock, but physical/mechanical perturbati<strong>on</strong>s <strong>on</strong> some<br />

higher order structure are probably also c<strong>on</strong>tributing signals to at least some pathways. In this short<br />

presentati<strong>on</strong> we will describe the numerous protein kinase pathways that are activated in a seemingly specific<br />

manner by heat shock focussing <strong>on</strong> the difference between heat shock and other stress and <strong>on</strong> the possible<br />

homeostatic functi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> these pathways in the global cell resp<strong>on</strong>se.<br />

2B_04_S<br />

ROLES OF MOLECULAR CHAPERONES IN QUALITY CONTROL OF<br />

MEMBRANES AND MEMBRANE ASSOCIATING PROTEINS IN PROKARYOTES<br />

Hitoshi Nakamoto<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry and Molecular Biology, Saitama University, Saitama 338-8570, Japan.<br />

e-mail: nakamoto@post.saitama-u.ac.jp<br />

Molecular chaper<strong>on</strong>es play important roles in protein quality c<strong>on</strong>trol. Ample evidence has accumulated to<br />

show that they associate with membranes although they do not c<strong>on</strong>tain transmembrane domains or signal<br />

sequences. Here, I present an overview <str<strong>on</strong>g>of</str<strong>on</strong>g> functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> molecular chaper<strong>on</strong>es, especially those from<br />

cyanobacteria, in quality c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> membrane and membrane-associating proteins.<br />

In c<strong>on</strong>trast to heterotrophic organisms such E. coli, cyanobacteria have layers <str<strong>on</strong>g>of</str<strong>on</strong>g> green membranes called<br />

thylakoid membranes where photosynthesis takes place. Thylakoid membranes possess membrane-embedded<br />

protein complexes such as photosystem II as well as peripheral soluble protein complexes such as<br />

phycobilisomes. Both photosystem II and phycobilisomes are thermolabile elements <str<strong>on</strong>g>of</str<strong>on</strong>g> the thylakoid<br />

membrane.<br />

Small Hsp, GroEL (Hsp60), DnaK (Hsp70), and HtpG (Hsp90) have been shown to associate with<br />

(thyakoid) membranes. Am<strong>on</strong>g them, small Hsp has been studied most extensively in terms <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular<br />

localizati<strong>on</strong> and physiological relevance <str<strong>on</strong>g>of</str<strong>on</strong>g> small Hsp thylakoid associati<strong>on</strong>.<br />

Genetic studies indicated that small Hsp c<strong>on</strong>fers thermostability to photosystem II and light-harvesting<br />

phycocyanins, the major comp<strong>on</strong>ent <str<strong>on</strong>g>of</str<strong>on</strong>g> phycobilisomes. C<strong>on</strong>stitutive expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a small Hsp in<br />

cyanobacterial cells stabilized subcellular structures such as thylakoid membranes under elevated temperature<br />

or intensive light stress. These results are c<strong>on</strong>sistent with in vitro studies by Vígh’s group that showed that<br />

small Hsp possesses an ability to stabilize the lipid phase <str<strong>on</strong>g>of</str<strong>on</strong>g> membranes. Thus, small Hsp possesses not <strong>on</strong>ly<br />

an activity to protect proteins located either in cytosol or in membranes, but also an ability to stabilize<br />

membranes in vivo.<br />

90


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2B_05_S<br />

CHARACTERIZATION OF SPECIFIC PLASMA MEMBRANE PERTURBATIONS<br />

WHICH ARE SUFFICIENT TO INITIATE OR REFINE THE HEAT SHOCK<br />

PROTEIN RESPONSE<br />

László Vigh<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, Biol. Res. Centr., Hung. Acad. Sci. Szeged, Hungary<br />

e-mail: vigh@brc.hu<br />

Cancer, diabetes, neurodegenerative and other diseases are known to be associated with abnormal HSP levels<br />

and characteristic membrane defects. The present study aims to establish a mechanism <str<strong>on</strong>g>for</str<strong>on</strong>g> the c<strong>on</strong>necti<strong>on</strong><br />

between changes <str<strong>on</strong>g>of</str<strong>on</strong>g> lipid compositi<strong>on</strong>, fluidity- and microdomain organizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> plasma membrane and<br />

altered expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSPs in mouse B16(F10) melanoma cells. Exposure <str<strong>on</strong>g>of</str<strong>on</strong>g> cells to various membrane<br />

fluidizers or drug candidates which interact specifically with certain membrane lipids, up-regulate the heat<br />

shock protein expressi<strong>on</strong> without inducing protein-unfolding. The isothermal membrane perturbati<strong>on</strong><br />

activated HSP inducti<strong>on</strong> is shown to be mediated via HSF1. M<strong>on</strong>itoring the surface membrane microdomains<br />

by c<strong>on</strong>focal- and ultrasensitive single molecular microscopy, we established a relati<strong>on</strong>ship between specific<br />

distributi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> lipid rafts and the c<strong>on</strong>comitant changes in the level, pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ile and cellular distributi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSPs.<br />

Compounds, that target specific membrane microdomains and capable to modulate stress protein signals with<br />

c<strong>on</strong>siderable therapeutic benefit, will be discussed.<br />

91


23-26 August 2007,<br />

Budapest, Hungary<br />

92<br />

2B_01_P<br />

(poster secti<strong>on</strong> A2, poster board #90, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

COMMON PHYSICOCHEMICAL PROPERTIES OF POLAR LIPIDS UNDERLYING<br />

THERMAL ADAPTATION OF MARINE HYDROBIONTS AND THEIR<br />

VULNERABILITY TO HIGH TEMPERATURE<br />

Nina Sanina<br />

Far Eastern Nati<strong>on</strong>al University, Vladivostok, Russia<br />

Changes <str<strong>on</strong>g>of</str<strong>on</strong>g> thermotropic behavior and fatty acid compositi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the major polar lipids from different species<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> marine invertebrates and macrophytes underlying their warm-acclimati<strong>on</strong> and seas<strong>on</strong>al acclimatizati<strong>on</strong><br />

were studied by GLC, DSC and polarizing microscopy to estimate the capacity <str<strong>on</strong>g>of</str<strong>on</strong>g> these phylogenetically<br />

different ectothermic organisms to adapt in c<strong>on</strong>diti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> global temperature changes. Comm<strong>on</strong> strategy <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

thermal adaptati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> studied animals and plants was revealed in spite <str<strong>on</strong>g>of</str<strong>on</strong>g> substantially different compositi<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> polar lipids and their fatty acid (FA) pools. The seas<strong>on</strong>al changes <str<strong>on</strong>g>of</str<strong>on</strong>g> thermotropic behavior and FA<br />

compositi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> phosphatidylcholine (PC) (or DGTS) and phosphatidylethanolamine (PE) were c<strong>on</strong>sisted<br />

with homeoviscouse adaptati<strong>on</strong> c<strong>on</strong>cepti<strong>on</strong>. However Т max <str<strong>on</strong>g>of</str<strong>on</strong>g> photosynthetic lipids (glycolipids and<br />

phosphatidylglycerol) did not change or even increased from summer to winter that may induce the lowtemperature<br />

inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> photosynthesis. Irrespective to different thermotropic behavior, the unsaturati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

both lipid groups rose due with increasing n-3/n-6 PUFAs ratio. The reciprocal c<strong>on</strong>versi<strong>on</strong> n-6↔n-3 was<br />

shown to regulate not <strong>on</strong>ly the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> their derivatives but the viscosity <str<strong>on</strong>g>of</str<strong>on</strong>g> membrane lipids. The most<br />

share <str<strong>on</strong>g>of</str<strong>on</strong>g> n-6 in PC and PE seems to c<strong>on</strong>nect with enhanced role <str<strong>on</strong>g>of</str<strong>on</strong>g> plasma membrane lipids in<br />

communicati<strong>on</strong> with envir<strong>on</strong>ment in summer. Whereas increased level <str<strong>on</strong>g>of</str<strong>on</strong>g> n-3 in photosynthetic lipids<br />

appears to maintain photosynthesis and respirati<strong>on</strong> at low temperatures. High vulnerability <str<strong>on</strong>g>of</str<strong>on</strong>g> hydrobi<strong>on</strong>ts to<br />

the sharp temperature elevati<strong>on</strong> in winter or superoptimal temperatures in summer were shown to be caused<br />

by the absence <str<strong>on</strong>g>of</str<strong>on</strong>g> effective compensati<strong>on</strong> in physical state <str<strong>on</strong>g>of</str<strong>on</strong>g> membrane lipids and proximity <str<strong>on</strong>g>of</str<strong>on</strong>g> their<br />

isotropic transiti<strong>on</strong> to these temperatures.<br />

2B_02_P<br />

(poster secti<strong>on</strong> A2, poster board #91, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ABNORMAL INTERACTION OF MUTANT HSP22 (HSPB8) WITH THE RNA<br />

HELICASE DDX20 (GEMIN3, DP103)<br />

Xiankui Sun 1 , Jean-M. F<strong>on</strong>taine 1 , Adam D. Hoppe 2 , Serena Carra 3 , Cheryl DeGuzman 1 ,<br />

Jody L. Martin 4 , Stephanie Sim<strong>on</strong> 5 , Patrick Vicart 5 , Michael J. Welsh 1 , Jacques Landry 3 ,<br />

Rainer Benndorf 1<br />

Departments <str<strong>on</strong>g>of</str<strong>on</strong>g> 1 Cell and Developmental Biology;<br />

2Microbiology and Immunology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Michigan Medical School, Ann Arbor, MI, USA;<br />

3 <str<strong>on</strong>g>Centre</str<strong>on</strong>g> de recherche en cancérologie de l'Université Laval, L'Hôtel-Dieu de Québec, Québec, Canada;<br />

4Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Cardiovascular Institute, Loyola University Chicago, Maywood, IL, USA;<br />

5EA 300 Stress et pathologies du cytosquelette, Université Paris 7, France<br />

e-mail: rbenndo@umich.edu<br />

Eight mutati<strong>on</strong>s in the small heat shock proteins (sHSP) Hsp22 and Hsp27 have been associated with the<br />

motor neur<strong>on</strong> diseases (MND) distal hereditary motor neuropathy and Charcot-Marie-Tooth disease. Hsp22<br />

and Hsp27 interact with each other, suggesting that these two etiologic factors may act in the same pathway.<br />

In an ef<str<strong>on</strong>g>for</str<strong>on</strong>g>t to learn about the role <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp22 in MND, we screened a human cDNA library by the yeast two-


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

hybrid method <str<strong>on</strong>g>for</str<strong>on</strong>g> potential binding proteins. One identified protein was the RNA helicase Ddx20, a core<br />

comp<strong>on</strong>ent <str<strong>on</strong>g>of</str<strong>on</strong>g> the survival-<str<strong>on</strong>g>of</str<strong>on</strong>g>-motor neur<strong>on</strong> (SMN) complexes. This interacti<strong>on</strong> was verified by independent<br />

methods including FRET. Both mutant Hsp22 <str<strong>on</strong>g>for</str<strong>on</strong>g>ms showed abnormally increased binding to Ddx20.<br />

Interestingly, Ddx20 itself binds to the SMN protein, and mutati<strong>on</strong>s in the SMN1 gene cause spinal muscular<br />

atrophy, another MND. Thus, these protein interacti<strong>on</strong> data have linked the etiologic factors Hsp22, Hsp27,<br />

and SMN, and mutati<strong>on</strong>s in any <str<strong>on</strong>g>of</str<strong>on</strong>g> these genes cause the various <str<strong>on</strong>g>for</str<strong>on</strong>g>ms <str<strong>on</strong>g>of</str<strong>on</strong>g> MND. SMN complexes are<br />

involved in RNP processing. The mutant Hsp22/Ddx20 interacti<strong>on</strong> was sensitive to treatment with RNase<br />

suggesting involvement <str<strong>on</strong>g>of</str<strong>on</strong>g> RNA in this interacti<strong>on</strong> and a potential role <str<strong>on</strong>g>of</str<strong>on</strong>g> sHSPs in RNP processing.<br />

93


23-26 August 2007,<br />

Budapest, Hungary<br />

94<br />

2C. NON-CODING RNAS AND CELLULAR STRESS<br />

(SUBHASH C. LAKHOTIA)<br />

2C_01_S<br />

REGULATION OF THE Σ E ENVELOPE STRESS RESPONSE BY SMALL RNAS<br />

Poul Valentin-Hansen, Jesper Johansen, Anders Aamann Rasmussen<br />

Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry and Molecular Biology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Southern Denmark, Denmark<br />

e-mail: valentine@bmb.sdu.dk<br />

It has become clear that all types <str<strong>on</strong>g>of</str<strong>on</strong>g> cells c<strong>on</strong>tain a wealth <str<strong>on</strong>g>of</str<strong>on</strong>g> small, n<strong>on</strong>-coding RNAs (sRNAs) that have<br />

important roles in regulating gene expressi<strong>on</strong> at the post-transcripti<strong>on</strong>al level. In the bacterium Escherichia coli<br />

genome-wide searches have led to the identificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ∼100 sRNAs and many <str<strong>on</strong>g>of</str<strong>on</strong>g> these are known or<br />

believed to act by base-pairing to modify the translati<strong>on</strong> and/or the stability <str<strong>on</strong>g>of</str<strong>on</strong>g> target mRNAs. Generally, the<br />

regulati<strong>on</strong> relies <strong>on</strong> short stretches <str<strong>on</strong>g>of</str<strong>on</strong>g> base pairing, which allows recogniti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> multiple target mRNAs by a<br />

single sRNA as well as the recogniti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a single target by multiple sRNAs. Regulatory RNA molecules<br />

acting in this manner require the RNA chaper<strong>on</strong>e Hfq, which promotes the pairing <str<strong>on</strong>g>of</str<strong>on</strong>g> complementary RNA<br />

molecules. Another feature <str<strong>on</strong>g>of</str<strong>on</strong>g> the chromosomally encoded sRNAs is that they are tightly regulated at the<br />

transcripti<strong>on</strong>al level, and frequently expressed as part <str<strong>on</strong>g>of</str<strong>on</strong>g> global regulatory networks which functi<strong>on</strong> in<br />

resp<strong>on</strong>se to envir<strong>on</strong>mental stress signals. - Here, I will focus <strong>on</strong> the identificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> two highly c<strong>on</strong>served<br />

σ E -dependent sRNAs, RybB and MicA, and discuss recent progress in our understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> the σ E -signalling<br />

system, which m<strong>on</strong>itors the bacterial cell envelope. Studies in E. coli and Salm<strong>on</strong>ella dem<strong>on</strong>strate that MicA<br />

and RybB act within the envelope stress resp<strong>on</strong>se and serve dual roles. First, when inducing stress occurs,<br />

they facilitate rapid shut-<str<strong>on</strong>g>of</str<strong>on</strong>g>f <str<strong>on</strong>g>of</str<strong>on</strong>g> the synthesis <str<strong>on</strong>g>of</str<strong>on</strong>g> multiple outer membrane proteins. Moreover, in unstressed<br />

cells, the two sRNAs functi<strong>on</strong> within a surveillance loop to maintain envelope homeostasis and to achieve<br />

autoregulati<strong>on</strong>.<br />

2C_02_S<br />

STRESS-INDUCED NUCLEAR BODIES AND TRANSCRIPTION OF REPEATED<br />

SEQUENCES<br />

Caroline Jolly, Angéline Eymery, Sabrina Fritah, Claire Vourc’h<br />

Institut Albert B<strong>on</strong>niot, CRI INSERM-UJF U823, Team “Stress and Dynamics <str<strong>on</strong>g>of</str<strong>on</strong>g> Genome Organizati<strong>on</strong>”,<br />

Site Santé La Tr<strong>on</strong>che, BP170, 38042 Grenoble cedex 9, France<br />

e-mail: caroline.jolly@ujf-grenoble.fr<br />

Exposure <str<strong>on</strong>g>of</str<strong>on</strong>g> cells to stress induces dramatic changes in gene expressi<strong>on</strong>, activating the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> certain<br />

genes such as those encoding the heat shock proteins or HSPs, and inactivating others. In paralell to the<br />

activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp gene expressi<strong>on</strong>, we have shown that heat shock also induces the <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> particular<br />

nuclear structures termed nuclear stress bodies or nSBs. These structures <str<strong>on</strong>g>for</str<strong>on</strong>g>m principally <strong>on</strong> the<br />

pericentromeric regi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> human chromosome 9 (9q12) through a direct binding <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF1 with satellite III<br />

repeated sequences. We have shown that heat shock induces the transcripti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these repeated sequences<br />

into n<strong>on</strong>-coding RNAs termed satellite III transcripts. This transcripti<strong>on</strong> is RNA-polymerase II- and HSF1-<br />

dependent. The functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the satellite III transcripts is still unknown, but several hypotheses can be<br />

c<strong>on</strong>sidered. Since they remain associated with chromosome 9 <str<strong>on</strong>g>for</str<strong>on</strong>g> several hours after synthesis, they may play<br />

a role in chromatin structure. Alternatively, since several splicing factors remain associated to the sat III


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

transcripts, they could play a role in the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> alternative splicing, a functi<strong>on</strong> which is indeed altered<br />

during heat exposure. I will present our latest findings c<strong>on</strong>cerning satellite III transcripts and discuss their<br />

possible functi<strong>on</strong> during stress exposure.<br />

2C_03_S<br />

EUKARYOTIC RNA THERMOSENSOR<br />

Ilya Shamovsky, June Hyung Lee, Maria Vera, K<strong>on</strong>stantin Shatalin, Evgeny Nudler<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, New York University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, New York, NY, 10016, USA<br />

The heat shock transcripti<strong>on</strong> factor (HSF1) plays a central role in the heat shock (HS) resp<strong>on</strong>se in eukaryotes<br />

by inducing the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock proteins (HSPs) and other cytoprotective proteins. HSF1 is present<br />

in unstressed mammalian cells in an inactive m<strong>on</strong>omeric <str<strong>on</strong>g>for</str<strong>on</strong>g>m and becomes activated by heat and other stress<br />

stimuli. HSF1 activati<strong>on</strong> involves trimerizati<strong>on</strong> and acquisiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a site-specific DNA-binding activity, which<br />

is negatively regulated by interacti<strong>on</strong> with certain HSPs. We have shown that HSF1 activati<strong>on</strong> by HS is an<br />

active process that is mediated by a rib<strong>on</strong>ucleoprotein complex c<strong>on</strong>taining translati<strong>on</strong> el<strong>on</strong>gati<strong>on</strong> factor<br />

eEF1A and a novel n<strong>on</strong>-coding RNA, which we termed HSR1 (Heat Shock RNA-1). Both HSR1 and eEF1A<br />

are required <str<strong>on</strong>g>for</str<strong>on</strong>g> HSF1 activati<strong>on</strong> in vitro. Antisense olig<strong>on</strong>ucleotides or siRNA against HSR1 impair the HS<br />

resp<strong>on</strong>se in vivo, rendering mammalian cells thermosensitive. We also show that n<strong>on</strong>-coding RNAs<br />

homologous to mammalian HSR1 are present in other eukaryotic species including Xenopus, Drosophila and<br />

C.elegans. HSR1 is c<strong>on</strong>stitutively expressed in all these organisms and its homologues are functi<strong>on</strong>ally<br />

interchangeable. Our results suggest a general model <str<strong>on</strong>g>for</str<strong>on</strong>g> eukaryotic HS genes activati<strong>on</strong> whereby HSR1<br />

serves as a cellular thermosensor that determines the temperature threshold <str<strong>on</strong>g>for</str<strong>on</strong>g> the HS resp<strong>on</strong>se; while<br />

eEF1A links HSP expressi<strong>on</strong> to major cellular perturbati<strong>on</strong>s during HS, such as translati<strong>on</strong>al shutdown and<br />

cytoskelet<strong>on</strong> collapse. The central role <str<strong>on</strong>g>of</str<strong>on</strong>g> HSR1 during HS implies that targeting this RNA could serve as a<br />

new therapeutic mode <str<strong>on</strong>g>for</str<strong>on</strong>g> cancer, inflammati<strong>on</strong> and other c<strong>on</strong>diti<strong>on</strong>s associated with HSF1 deregulati<strong>on</strong>.<br />

2C_04_S<br />

NON-CODING HSRω RNA AND POST-TRANSCRIPTIONAL PROCESSING IN<br />

STRESSED CELLS<br />

Moushami Mallik, Subhash C. Lakhotia<br />

Cytogenetics Laboratory, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Zoology, Banaras Hindu University, Varanasi 221 005, India<br />

e-mail: lakhotia@bhu.ac.in<br />

Every cell needs a large variety and number <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins <str<strong>on</strong>g>for</str<strong>on</strong>g> transcripti<strong>on</strong> and processing <str<strong>on</strong>g>of</str<strong>on</strong>g> nascent transcripts<br />

(splicing, other RNA processing, transport etc). Stress or n<strong>on</strong>-permissive c<strong>on</strong>diti<strong>on</strong>s, which largely inhibit<br />

transcripti<strong>on</strong>al and RNA processing machineries, result in high surplus <str<strong>on</strong>g>of</str<strong>on</strong>g> unengaged RNA processing<br />

proteins. Since these proteins are not degraded but must be available up<strong>on</strong> recovery, they need to be<br />

reversibly sequestered. Heat stress induced <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> nuclear stress bodies/granules in human cells and<br />

clustering <str<strong>on</strong>g>of</str<strong>on</strong>g> the varieties <str<strong>on</strong>g>of</str<strong>on</strong>g> nuclear speckles (IGCs, paraspeckles, omega speckles etc) in different cell types<br />

appear to reflect such sequestrati<strong>on</strong>. A major focus <str<strong>on</strong>g>of</str<strong>on</strong>g> studies in our laboratory is <strong>on</strong> the developmentally<br />

expressed and stress-inducible n<strong>on</strong>-coding hsrω gene in Drosophila. The large nuclear transcript <str<strong>on</strong>g>of</str<strong>on</strong>g> this gene,<br />

hsrω-n, is known to be required <str<strong>on</strong>g>for</str<strong>on</strong>g> organizing the unengaged nuclear hnRNPs and related RNA-binding<br />

proteins in nucleoplasmic omega speckles in nearly every cell type <str<strong>on</strong>g>of</str<strong>on</strong>g> Drosophila. Using transgenic lines<br />

95


23-26 August 2007,<br />

Budapest, Hungary<br />

designed to either over-express or ablate the hsrω-n transcripts, we rec<strong>on</strong>firm that the large nuclear hsrω-n<br />

transcript is required <str<strong>on</strong>g>for</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> omega speckles and show that this n<strong>on</strong>-coding transcript plays crucial<br />

roles in normal development and is also essential <str<strong>on</strong>g>for</str<strong>on</strong>g> survival <str<strong>on</strong>g>of</str<strong>on</strong>g> the organism following stress. We believe<br />

that such n<strong>on</strong>-coding RNA species per<str<strong>on</strong>g>for</str<strong>on</strong>g>m a wide range <str<strong>on</strong>g>of</str<strong>on</strong>g> functi<strong>on</strong>s in cells through their ability to interact<br />

with a great variety <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins and thus act as “hubs” to integrate complex networks <str<strong>on</strong>g>of</str<strong>on</strong>g> gene activity during<br />

development and under c<strong>on</strong>diti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> stress.<br />

96


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2C_01_P<br />

(poster secti<strong>on</strong> A2, poster board #92, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HEAT-INDUCED GLOBAL DEACETYLATION OF CORE HISTONES INVOLVES<br />

HDAC1 AND HDAC2<br />

Sabrina Fritah, Cyril Boyault, Susanna Chiocca, Saadi Khochbin, Caroline Jolly, Claire Vourc’h<br />

CRI Inserm U823, Institut Albert B<strong>on</strong>niot, Grenoble, France<br />

e-mail: Sabrina.fritah@e.ujf-grenoble.fr<br />

Heat shock induces major changes in gene expressi<strong>on</strong>. Besides the up-regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> specific genes, am<strong>on</strong>g<br />

which hsp genes, heat shock also induces a general shut-down <str<strong>on</strong>g>of</str<strong>on</strong>g> gene expressi<strong>on</strong> underlined by a global<br />

chromatin remodelling. While the mechanisms involved in the heat-induced activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp genes have<br />

been investigated in details, the molecular events associated with the global down-regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the genome<br />

are poorly understood. By using a combinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> in situ and molecular approaches, we have shown that heat<br />

shock induces a massive and reversible deacetylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> core hist<strong>on</strong>es affecting specific epigenetic marks <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

hist<strong>on</strong>es H3 and H4. This deacetylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> core hist<strong>on</strong>es is a c<strong>on</strong>served phenomen<strong>on</strong>. Moreover it is<br />

correlated with the intensity <str<strong>on</strong>g>of</str<strong>on</strong>g> stress. We have characterized the molecular mechanisms underlying the heatinduced<br />

deacetylati<strong>on</strong>, and have identified HDAC1 (Hist<strong>on</strong>e Deacetylase 1) and HDAC2 as the key actors <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

this event. Interestingly, we found that HDAC1 and HDAC2 are complexed with HSF1 (Heat shock factor<br />

1), in a stress dependent manner. We thus identify HDAC1 and HDAC2 as novel important actors <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

heat shock resp<strong>on</strong>se. Our last results c<strong>on</strong>cerning the mechanisms involved in the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HDACs<br />

activity during stress will be also presented and discussed.<br />

97


23-26 August 2007,<br />

Budapest, Hungary<br />

98<br />

2D. ER STRESS<br />

(GÁBOR BÁNHEGYI, AMY S. LEE)<br />

2D_01_S<br />

STRESS IN-AND-OUT<br />

Gábor Bánhegyi<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Chemistry, Semmelweis University, 1444 Budapest, POB 260, Hungary<br />

e-mail: banhegyi@puskin.sote.hu<br />

Glucocorticoids are main actors in the pathomechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> stress. In the original c<strong>on</strong>text <str<strong>on</strong>g>for</str<strong>on</strong>g>mulated by Hans<br />

Selye in his stress theory, glucocorticoids are produced in the adrenal cortex up<strong>on</strong> the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

hypothalamic − pituitary − adrenal axis. Their increased producti<strong>on</strong> mediates alarm reacti<strong>on</strong>s in acute stress,<br />

facilitating metabolic alterati<strong>on</strong>s in the general adaptati<strong>on</strong> syndrome allowing the individual to attempt<br />

countermeasures such as the „fight or flight” resp<strong>on</strong>se. Recent observati<strong>on</strong>s show that active glucocorticoids can<br />

be also <str<strong>on</strong>g>for</str<strong>on</strong>g>med from their inactive counterparts in various tissues, including liver and adipose tissue. This<br />

prereceptorial activati<strong>on</strong> takes place in the lumen <str<strong>on</strong>g>of</str<strong>on</strong>g> the endoplasmic reticulum (ER) and depends <strong>on</strong> the<br />

activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the glucose-6-phosphate transporter − hexose-6-phosphate dehydrogenase − 11β-hydroxysteroid<br />

dehydrogenase type 1 triad. Increased prereceptorial glucocorticoid activati<strong>on</strong> is accompanied by the signs <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

ER stress in certain human diseases (obesity, metabolic syndrome, type 2 diabetes). These diseases are more<br />

comm<strong>on</strong> am<strong>on</strong>g socio-ec<strong>on</strong>omically disadvantaged individuals and are associated with lifestyle factors and<br />

chr<strong>on</strong>ic stress. It has been recently suggested that the ER can functi<strong>on</strong> as a sensor <str<strong>on</strong>g>for</str<strong>on</strong>g> electr<strong>on</strong> d<strong>on</strong>ors and<br />

acceptors, i.e. nutrients and oxygen. In the current social envir<strong>on</strong>ment <str<strong>on</strong>g>of</str<strong>on</strong>g> high energy input and minimal<br />

physical activity, the ER encounters a nutrient (electr<strong>on</strong>) overload, leading to a redox imbalance in the lumen,<br />

which is the most frequent cause <str<strong>on</strong>g>of</str<strong>on</strong>g> ER stress and c<strong>on</strong>sequent apoptosis. Furthermore, reductive effects<br />

favor the increased prereceptorial glucocorticoid activati<strong>on</strong>. In c<strong>on</strong>clusi<strong>on</strong>, glucocorticoid resp<strong>on</strong>se can be<br />

initiated by an aut<strong>on</strong>omous sensing <str<strong>on</strong>g>of</str<strong>on</strong>g> (nutrient) stress in cellular level beside the central neuroendocrine<br />

mechanism.<br />

2D_02_S<br />

ER STRESS INDUCTION OF UPR REGULATOR GRP78: ROLE IN DEVELOPMENT<br />

AND DISEASE<br />

Amy S. Lee, Peter Baumeister, Changhui Mao, Miao Wang, Risheng Ye, Min Ni, Y<strong>on</strong>g Fu, Jianze Li,<br />

Dezheng D<strong>on</strong>g<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry and Molecular Biology and the USC/Norris Comprehensive Cancer Center, Keck School<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine <str<strong>on</strong>g>of</str<strong>on</strong>g> the University <str<strong>on</strong>g>of</str<strong>on</strong>g> Southern Cali<str<strong>on</strong>g>for</str<strong>on</strong>g>nia, 1441 Eastlake Ave., Los Angeles, CA 90089-9176.<br />

e-mail: amylee@usc.edu<br />

Mammalian cells have evolved multiple adaptive pathways, referred to as the unfolded protein resp<strong>on</strong>se<br />

(UPR), that allow them to resp<strong>on</strong>d to perturbati<strong>on</strong>s in endoplasmic reticulum (ER) homeostasis. One major<br />

pro-survival mechanism is mediated by the ER chaper<strong>on</strong>e GRP78/BiP, an anti-apoptotic protein which also<br />

regulates ER stress signaling. To probe the physiologic functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> GRP78/BiP, mouse models were<br />

recreated targeting the Grp78 allele. This lead to the discovery that complete depleti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> GRP78 results in<br />

early embry<strong>on</strong>ic lethality due to proliferati<strong>on</strong> defects and apoptosis <str<strong>on</strong>g>of</str<strong>on</strong>g> the inner cell mass which is the<br />

precursor <str<strong>on</strong>g>of</str<strong>on</strong>g> embry<strong>on</strong>ic stem cells. Our results show that reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> GRP78 level by half is sufficient to


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

maintain cellular homeostasis during development with no major c<strong>on</strong>sequence in ER stress signaling. This<br />

implies that an elevated GRP78 level is more critically needed in cells undergoing physiological or<br />

pathological stress, as exemplified by protecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> vulnerable neur<strong>on</strong>al cells and allowing cancer cells to<br />

evade the host defense system and cancer therapies. In this lecture, we will discuss the c<strong>on</strong>sequence <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

c<strong>on</strong>diti<strong>on</strong>al knockout <str<strong>on</strong>g>of</str<strong>on</strong>g> GRP78 in specific neur<strong>on</strong>al cells. Due to hypoxic c<strong>on</strong>diti<strong>on</strong>s and glucose deprivati<strong>on</strong><br />

caused by poor vascularizati<strong>on</strong>, the microenvir<strong>on</strong>ment <str<strong>on</strong>g>of</str<strong>on</strong>g> tumors represents physiological ER stress and the<br />

UPR is activated <str<strong>on</strong>g>for</str<strong>on</strong>g> tumor cell survival. In this lecture, we will discuss how GRP78 deficiency will affect<br />

cancer progressi<strong>on</strong> and the underlying mechanisms resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> the phenomen<strong>on</strong>.<br />

2D_04_S<br />

MANAGING AND EXPLOITING STRESS IN THE ANTIBODY FACTORY<br />

Roberto Sitia<br />

Università Vita-Salute San Raffaele Scientific Institute, 20132 Milano, Italy, Fax: +39 02 2643 4723<br />

e-mail: r.sitia@hsr.it<br />

Up<strong>on</strong> encounter with antigen, l<strong>on</strong>g-lived B lymphocytes differentiate into short-lived plasma cells, the<br />

terminal effectors <str<strong>on</strong>g>of</str<strong>on</strong>g> the humoral immune resp<strong>on</strong>se. Plasma cells are specialized in immunoglobulin (Ig)<br />

secreti<strong>on</strong>, each <str<strong>on</strong>g>of</str<strong>on</strong>g> them being capable <str<strong>on</strong>g>of</str<strong>on</strong>g> releasing thousands molecules per sec<strong>on</strong>d. How do plasma cells<br />

achieve such an efficiency How do they cope with metabolic and redox imbalances that exuberant protein<br />

secreti<strong>on</strong> can cause Is plasma cell death linked to Ig producti<strong>on</strong>, such as to limit antibody resp<strong>on</strong>ses We<br />

have dissected terminal plasma cell differentiati<strong>on</strong> through dynamic imaging, proteomics and genomic<br />

analyses. Our results show that waves <str<strong>on</strong>g>of</str<strong>on</strong>g> functi<strong>on</strong>ally related proteins are produced to increase the capacity <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the antibody factory, and shed some light in the signalling pathways utilised to orchestrate massive de novo ER<br />

biogenesis. As to the mechanisms that lead to plasma cell death, we showed that in the late phases <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

plasmacytic differentiati<strong>on</strong>, when antibody producti<strong>on</strong> becomes maximal, proteasomal activity unexpectedly<br />

decreases. The excessive load <str<strong>on</strong>g>for</str<strong>on</strong>g> the reduced proteolytic capacity correlates with accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

polyubiquitinated proteins, stabilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> endogenous proteasomal substrates (including Xbp1s, Ik-Bα and<br />

Bax), <strong>on</strong>set <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis, and sensitizati<strong>on</strong> to proteasome inhibitors. A developmental program seems<br />

there<str<strong>on</strong>g>for</str<strong>on</strong>g>e to link plasma cell death to protein producti<strong>on</strong>, explaining the peculiar sensitivity <str<strong>on</strong>g>of</str<strong>on</strong>g> normal and<br />

malignant plasma cells to proteasome inhibitors.<br />

2D_05_S<br />

(poster secti<strong>on</strong> A2, poster board #93, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ENDOPLASMATIC RETICULUM AND ENERGY STRESS RESPONSE<br />

MECHANISMS IN INTESTINAL EPITHELIAL CELLS UNDER CHRONIC<br />

INFLAMMATION: INHIBITORY EFFECTS OF INTERLEUKIN 10<br />

A. Messlik, P. A. Ruiz, S. C. Kim, R. B. Sartor, D. Haller*<br />

Technical University <str<strong>on</strong>g>of</str<strong>on</strong>g> Munich, Experimental Nutriti<strong>on</strong> Medicine, Am Forum 5, 85350 Freising, Germany<br />

*e-mail: haller@wzw.tum.de; Ph<strong>on</strong>e: ++49-(0)8161-712026; fax: ++49-(0)8161-712097<br />

The initiati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> endoplasmic reticulum (ER) stress resp<strong>on</strong>ses and energy deficiency in intestinal epithelial<br />

cells (IEC) may c<strong>on</strong>tribute to the pathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic intestinal inflammati<strong>on</strong>. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was<br />

99


23-26 August 2007,<br />

Budapest, Hungary<br />

to characterize anti-inflammatory mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> interleukin 10 (IL10) using functi<strong>on</strong>al epithelial cell<br />

proteomics.<br />

Proteome analysis from primary IEC <str<strong>on</strong>g>of</str<strong>on</strong>g> Enterococcus faecalis- and Escherichia coli m<strong>on</strong>oassociated IL10 deficient<br />

(IL10-/-) mice revealed increased expressi<strong>on</strong> levels <str<strong>on</strong>g>of</str<strong>on</strong>g> the glucose-regulated ER stress proteins (grp)78 under<br />

c<strong>on</strong>diti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> experimental colitis. Interestingly, the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ER stress resp<strong>on</strong>se mechanisms under<br />

c<strong>on</strong>diti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic inflammati<strong>on</strong> was associated with decreased expressi<strong>on</strong> levels <str<strong>on</strong>g>of</str<strong>on</strong>g> the mitoch<strong>on</strong>drial<br />

creatine kinase and increased activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the AMP kinase system in primary IEC, suggesting dysregulati<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the cellular energy homeostasis. Most importantly, IL10 inhibited grp78 expressi<strong>on</strong> in IL10 receptor<br />

rec<strong>on</strong>stituted epithelial cells. Chromatin immunoprecipitati<strong>on</strong> analysis revealed that IL10-mediated p38<br />

signaling inhibited TNF-induced recruitment <str<strong>on</strong>g>of</str<strong>on</strong>g> the ER-derived activating transcripti<strong>on</strong> factor (ATF)-6 to the<br />

grp78 promotor likely through the blockade <str<strong>on</strong>g>of</str<strong>on</strong>g> ATF-6 nuclear translocati<strong>on</strong>.<br />

The failure <str<strong>on</strong>g>of</str<strong>on</strong>g> energy homeostasis in primay IEC from inflamed IL10-/- mice was associated ER stress<br />

resp<strong>on</strong>ses in the intestinal epithelium. In additi<strong>on</strong>, IL10 inhibits inflammati<strong>on</strong>-induced ER stress resp<strong>on</strong>se<br />

mechanisms by modulating ATF-6 nuclear recruitment to the grp78 gene promotor.<br />

2D_06_S<br />

(poster secti<strong>on</strong> A2, poster board #94, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PROTEASOMAL DEGRADATION IS TRANSIENTLY ARRESTED DURING<br />

INHIBITION OF TRANSLATION IN ER STRESS<br />

Marina Shenkman, Gerardo Z. Lederkremer<br />

Tel Aviv University, Cell and Developmental Biology<br />

The unfolded protein resp<strong>on</strong>se (UPR) activates transcripti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> genes involved in proteasomal degradati<strong>on</strong>.<br />

However, we found that in its early stages the UPR leads to a transient inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proteasomal disposal <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

cytosolic substrates (p53 and p27Kip1) and <str<strong>on</strong>g>of</str<strong>on</strong>g> those targeted to ER-associated degradati<strong>on</strong> (uncleaved<br />

precursor <str<strong>on</strong>g>of</str<strong>on</strong>g> asialoglycoprotein receptor H2a). Degradati<strong>on</strong> resumed so<strong>on</strong> after the protein synthesis arrest<br />

that occurs in early UPR subsided. C<strong>on</strong>sistently, also protein synthesis inhibitors blocked<br />

ubiquitin/proteasomal degradati<strong>on</strong>. Ubiquitinati<strong>on</strong> was inhibited during the translati<strong>on</strong> block, suggesting<br />

short-lived E3 ubiquitin ligases as candidate depleted proteins. This was indeed the case <str<strong>on</strong>g>for</str<strong>on</strong>g> p53 whose E3<br />

ligase, MDM2, when overexpressed, restored the degradati<strong>on</strong>, whereas a mutant MDM2 in its acidic domain<br />

restored the ubiquitinati<strong>on</strong> but not completely the degradati<strong>on</strong>. Inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proteasomal degradati<strong>on</strong> early in<br />

UPR may prevent depleti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> essential short-lived factors during the translati<strong>on</strong> arrest. Stabilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> p27<br />

through this mechanism may explain the cell cycle arrest in G1 when translati<strong>on</strong> is blocked by inhibitors or by<br />

the UPR.<br />

100


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2D_01_P<br />

(poster secti<strong>on</strong> A2, poster board #95, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

APOLIPOPROTEIN B MISFOLDING AND ER STRESS: EVIDENCE THAT<br />

GLUCOSAMINE-INDUCED MISFOLDING OF APOB LEADS TO ER STRESS AND<br />

ER-ASSOCIATED PROTEASOMAL DEGRADATION<br />

Wei Qiu, Rita Kohen, Angela Rutlege, Khosrow Adeli<br />

Molecular Structure and Functi<strong>on</strong>, The Hospital <str<strong>on</strong>g>for</str<strong>on</strong>g> Sick Children, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Tor<strong>on</strong>to,<br />

Tor<strong>on</strong>to, Ontario, M5G 1X8, Canada<br />

Glucosamine treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> HepG2 cells increased levels <str<strong>on</strong>g>of</str<strong>on</strong>g> the ER chaper<strong>on</strong>es, 78-kDa glucose-regulated<br />

protein (Grp78) and Grp94, in a dose-dependent manner and led to significant decreases in both cellular and<br />

secreted apolipoprotein B100 (apoB100) by up to 97% (p


23-26 August 2007,<br />

Budapest, Hungary<br />

interacti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Ero1b with PDI. However, the Gly to Ser mutati<strong>on</strong> abolishes disulfide-dependent PDI-Ero1β<br />

heterodimers. Both the Gly to Ser and His to Tyr mutati<strong>on</strong>s make Ero1b susceptible to misoxidati<strong>on</strong> and<br />

aggregati<strong>on</strong>, particularly during a temperature or redox stress. We c<strong>on</strong>clude that the Ero FAD binding<br />

domain is critical <str<strong>on</strong>g>for</str<strong>on</strong>g> c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>al stability, allowing Ero proteins to withstand stress c<strong>on</strong>diti<strong>on</strong>s that cause<br />

client proteins to misfold.<br />

102<br />

2D_04_P<br />

(poster secti<strong>on</strong> A2, poster board #97, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE APOPTOTIC EFFECT OF N-3 FATTY ACIDS ON THE LEUKEMIA CELL LINE<br />

HL60 IS LIKELY TO BE MEDIATED THROUGH ER STRESS<br />

J. E. Slagsvold, S. A. Schønberg<br />

Dept <str<strong>on</strong>g>of</str<strong>on</strong>g> Laboratory Medicine, Women’s and Children’s Health, the Norwegian University <str<strong>on</strong>g>of</str<strong>on</strong>g> Science and Technology,<br />

N-7489 Tr<strong>on</strong>dheim, Norway, e-mail: slagsvol@stud.ntnu.no<br />

Background - Numerous studies indicate an inverse relati<strong>on</strong>ship between dietary intake <str<strong>on</strong>g>of</str<strong>on</strong>g> n-3<br />

polyunsaturated fatty acids (PUFAs) and cancer. Cell culture studies show that n-3 PUFAs inhibit cell<br />

proliferati<strong>on</strong> and induce programmed cell death, apoptosis. However, the molecular mechanisms behind<br />

these effects are complex and unclear. Aim – To investigate signaling pathways involved in EPA-induced<br />

growth arrest/apoptosis in HL-60 cells. Materials and methods – Gene expressi<strong>on</strong> pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iling <str<strong>on</strong>g>of</str<strong>on</strong>g> HL60 cells<br />

treated with EPA was per<str<strong>on</strong>g>for</str<strong>on</strong>g>med by Affymetrix GeneChip System and key markers <str<strong>on</strong>g>of</str<strong>on</strong>g> relevant pathways<br />

verified at protein level by Western blotting. Results - Several transcripts involved in ER homeostasis, cell<br />

cycle and apoptosis were affected. Transcripts involved in the unfolded protein resp<strong>on</strong>se (UPR) pathway,<br />

chaper<strong>on</strong>es, folding proteins and transcripts involved in apoptosis, were upregulated, whereas cyclin D1 and<br />

cellcycle/progressi<strong>on</strong> transcripts were downregulated. Proteins in the UPR like eIF2α-P, ATF4, SQSTM-1<br />

and cyclin D1 were verified by western blotting. C<strong>on</strong>clusi<strong>on</strong> - The data clearly show that the n-3 PUFA EPA<br />

induces apoptosis in HL 60 cells and that the mechanism may be mediated through ER stress.<br />

2D_05_P<br />

(poster secti<strong>on</strong> A2, poster board #98, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ERAD AND ER QUALITY CONTROL COMPARTMENTALIZATION DURING ER<br />

STRESS<br />

Bella Groisman, Edward Avezov, Julia Leitman, Efrat R<strong>on</strong>, Gerardo Z. Lederkremer<br />

Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Cell <str<strong>on</strong>g>Research</str<strong>on</strong>g> and Immunology, Tel Aviv University, e-mail: bellagr@post.tau.ac.il<br />

Protein homeostasis in mammalian cells is tightly c<strong>on</strong>trolled by protein synthesis and degradati<strong>on</strong>. An<br />

efficient quality c<strong>on</strong>trol machinery is required <str<strong>on</strong>g>for</str<strong>on</strong>g> proper cell functi<strong>on</strong>, which is particularly important in<br />

c<strong>on</strong>diti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> stress. Disrupti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> homeostasis, e.g by restricti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> oxygen or nutrient supply or interfering<br />

with calcium balance in the ER results in overloading <str<strong>on</strong>g>of</str<strong>on</strong>g> the ER, causing ER stress and activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a<br />

protective cellular mechanism termed the Unfolded Protein Resp<strong>on</strong>se (UPR). During the UPR there is an<br />

inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> genes encoding <str<strong>on</strong>g>for</str<strong>on</strong>g> chaper<strong>on</strong>es and E3 ubiquitin ligases targeted to reduce ER protein overload<br />

through ER-Associated Degradati<strong>on</strong> (ERAD).<br />

Using a model ERAD glycoprotein substrate and its n<strong>on</strong>-glycosylated versi<strong>on</strong> we found that several <str<strong>on</strong>g>of</str<strong>on</strong>g> these<br />

UPR induced factors, such as the putative mannose lectin EDEM and the E3 RING ubiquitin ligase HRD1


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

are required <str<strong>on</strong>g>for</str<strong>on</strong>g> degradati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> both substrates, whereas the cytosolic E3 member <str<strong>on</strong>g>of</str<strong>on</strong>g> the F-box family Fbs2<br />

participates in degradati<strong>on</strong> <strong>on</strong>ly <str<strong>on</strong>g>of</str<strong>on</strong>g> the glycoprotein. We found that up<strong>on</strong> ER stress HRD1 and Fbs2<br />

accumulate al<strong>on</strong>g with the ERAD substrate in the pericentriolar ER-derived quality c<strong>on</strong>trol compartment<br />

(ERQC), previously discovered in our lab. Other additi<strong>on</strong>al key factors implicated in ER quality c<strong>on</strong>trol and<br />

the ER stress resp<strong>on</strong>se, are also found in the ERQC and their localizati<strong>on</strong> depends <strong>on</strong> the functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the branches <str<strong>on</strong>g>of</str<strong>on</strong>g> the UPR. These and other findings suggest that protein recruitment to in the ERQC is<br />

involved in coping with ER stress.<br />

2D_06_P<br />

(poster secti<strong>on</strong> A2, poster board #99, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

LOW-IRON DIET ABROGATES CHRONIC INTESTINAL INFLAMMATION IN<br />

TNFDARE/WT MICE<br />

T. Werner, N. Schulz, G. Kollias, K. Schuemann, D. Haller *<br />

Technical University <str<strong>on</strong>g>of</str<strong>on</strong>g> Munich, Experimental Nutriti<strong>on</strong>al Medicine, Am Forum 5, 85350 Freising, Germany<br />

*e-mail: haller@wzw.tum.de; Ph<strong>on</strong>e: +49-(0)8161-712026; Fax: +49-(0)8161-712102<br />

Patients suffering from intestinal bowel disease (IBD) have decreased anti-oxidant and glutathi<strong>on</strong>e levels<br />

which are indicative <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidative stress. High c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> ir<strong>on</strong> have been shown to increase the<br />

oxidative status <strong>on</strong> the cell leading to the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> endoplasmic reticulum (ER) stress and protein<br />

oxidative damage. We studied the role <str<strong>on</strong>g>of</str<strong>on</strong>g> low-dietary ir<strong>on</strong> <strong>on</strong> molecular mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> intestinal epithelial<br />

cell (IEC) ER stress resp<strong>on</strong>ses under c<strong>on</strong>diti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic experimental ileitis in TNFDARE/WT mice.<br />

Histological analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> normal-ir<strong>on</strong> fed mice showed severe intestinal inflammati<strong>on</strong> with a score <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

7.67 ± 1.53 (pathological range from 0-10), whereas low-ir<strong>on</strong> fed mice presented a significant reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

inflammati<strong>on</strong> in proximal and distal ileum segments with a score <str<strong>on</strong>g>of</str<strong>on</strong>g> 2.30 +/- 0.68. Western blot analysis in<br />

primary IEC from TNFDARE/WT and wild type mice showed reduced NF-κB activati<strong>on</strong> and MAPK<br />

signalling including phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> RelA, cJun and Erk. The activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ER stress resp<strong>on</strong>ses including<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> glucose regulated protein 78, grp58, phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> eukaryotic initiati<strong>on</strong> factor 2 alpha as<br />

well as cleavage <str<strong>on</strong>g>of</str<strong>on</strong>g> caspase 12 and 3 were decreased under low-ir<strong>on</strong> diet. Furthermore, proteome analysis<br />

identified 40 differentially regulated proteins including the ir<strong>on</strong>-regulated pro-inflammatory mediator<br />

intelectin.<br />

This study presents evidence <str<strong>on</strong>g>for</str<strong>on</strong>g> the beneficial effects <str<strong>on</strong>g>of</str<strong>on</strong>g> low-ir<strong>on</strong> intake in a mouse model <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic ileitis<br />

including regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidative and ER stress resp<strong>on</strong>se mechanisms at the epithelial cell level and there<str<strong>on</strong>g>for</str<strong>on</strong>g>e<br />

pointing to possible therapeutic use <str<strong>on</strong>g>of</str<strong>on</strong>g> a low-ir<strong>on</strong> diet in patients <str<strong>on</strong>g>of</str<strong>on</strong>g> IBD.<br />

103


23-26 August 2007,<br />

Budapest, Hungary<br />

104<br />

2D_07_P<br />

(poster secti<strong>on</strong> A2, poster board #100, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

TRANSCRIPTIONAL PROFILING REVEALS INTEGRATION OF ER- AND<br />

OSMOTIC-STRESS PATHWAYS<br />

André S. T. Irsigler 1, , Maximiller D. L. Costa 1 , Ping Zhang 2 , Pedro A. Braga 1 , Ralph E. Dewey 2 ,<br />

Rebecca S. Bost<strong>on</strong> 3 , Elizabeth P. B. F<strong>on</strong>tes 1<br />

1Departamento de Bioquímica e Biologia Molecular, BIOAGRO, Universidade Federal de Viçosa,<br />

36571.000 Viçosa, Minas Gerais, Brazil<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Crop Science<br />

3Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Biology, North Carolina State University, Raleigh, North Carolina 27695<br />

Despite the potential <str<strong>on</strong>g>of</str<strong>on</strong>g> the endoplasmic reticulum (ER) stress resp<strong>on</strong>se to accommodate adaptive pathways,<br />

its integrati<strong>on</strong> with other envir<strong>on</strong>mental-induced resp<strong>on</strong>ses is poorly understood in plants. Here, we<br />

per<str<strong>on</strong>g>for</str<strong>on</strong>g>med global expressi<strong>on</strong> pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iling <strong>on</strong> soybean leaves exposed to polyethylene glycol treatment or to<br />

unfolded protein resp<strong>on</strong>se (UPR) inducers to identify integrated networks between osmotic and ER stressinduced<br />

adaptive resp<strong>on</strong>ses. The results unmasked the major branches <str<strong>on</strong>g>of</str<strong>on</strong>g> the ER-stress resp<strong>on</strong>se, which<br />

includes enhancing protein folding and degradati<strong>on</strong> in the ER, as well as specific osmotically regulated<br />

changes linked to cellular resp<strong>on</strong>ses induced by dehydrati<strong>on</strong>. However, a small proporti<strong>on</strong> (5.5%) <str<strong>on</strong>g>of</str<strong>on</strong>g> total upregulated<br />

genes represented a shared resp<strong>on</strong>se that seemed to integrate the two signaling pathways. These coregulated<br />

genes were c<strong>on</strong>sidered downstream targets based <strong>on</strong> similar inducti<strong>on</strong> kinetics and a synergistic<br />

resp<strong>on</strong>se to the combinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> osmotic- and ER-stress-inducing treatments. Genes in this integrated<br />

pathway with the str<strong>on</strong>gest synergistic inducti<strong>on</strong> encoded proteins with diverse roles. Two <str<strong>on</strong>g>of</str<strong>on</strong>g> them c<strong>on</strong>tained<br />

a plant-specific development and cell death (DCD) domain while another had homology to proteins with an<br />

ubiquitin-associated (UBA) domain. A NAC domain-c<strong>on</strong>taining protein exhibited robust early kinetics <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

inducti<strong>on</strong> c<strong>on</strong>sistent with a role as a transfactor. This integrated pathway diverged further from characterized<br />

ER-specific branches <str<strong>on</strong>g>of</str<strong>on</strong>g> UPR as downstream targets were inversely regulated by osmotic stress. Collectively,<br />

our results describe a novel branch <str<strong>on</strong>g>of</str<strong>on</strong>g> the ER stress resp<strong>on</strong>se that integrates the osmotic signal to potentiate<br />

transcripti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> shared target genes.<br />

2D_08_P<br />

(poster secti<strong>on</strong> A2, poster board #101, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

REGULATION OF ERGIC-53 GENE EXPRESSION IN RESPONSE TO<br />

ENDOPLASMIC RETICULUM STRESS<br />

Maurizio Renna, Stefano B<strong>on</strong>atti, § Giuseppina Amodio, § Silvia Franceschelli, Annalisa Fasano,<br />

Paolo Rem<strong>on</strong>delli<br />

Dipartimento di Scienze Farmaceutiche, Università degli Studi di Salerno, Fisciano-Salerno, Italy I-84034;<br />

§ Dipartimento di Biochimica e Biotecnologie Mediche, Università degli Studi di Napoli „Federico I”I, Naples,<br />

Italy I-80131<br />

Accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> unfolded proteins within the endoplasmic reticulum (ER) activates the unfolded protein<br />

resp<strong>on</strong>se (UPR) also known as the ER stress resp<strong>on</strong>se. We dem<strong>on</strong>strated that, in resp<strong>on</strong>se to ER stress<br />

induced by either nitric oxide (NO) or by thapsigargin (TG), the ER Golgi intermediate compartment protein<br />

ERGIC-53 and the adaptor protein MCFD2 accumulate by a transcripti<strong>on</strong>al mechanism. Interestingly,


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

transcripti<strong>on</strong>al activati<strong>on</strong> by NO involved <strong>on</strong>ly the ATF6α-dependent UPR pathway, while TG induced both<br />

the ATF6α− and the XBP1-mediated pathways. In additi<strong>on</strong>, the accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> both transporters occurred<br />

simultaneously to the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ER-stress dependent apoptosis, suggesting that these proteins may<br />

participate to the events that will eventually decide the fate <str<strong>on</strong>g>of</str<strong>on</strong>g> the cell. To investigate the molecular events<br />

that regulate UPR-mediated inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the gene we have analysed the transcripti<strong>on</strong>al regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

ERGIC-53. We found that the ERGIC-53 promoter c<strong>on</strong>tains a single cis-acting element that mediates<br />

inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the gene by thapsigargin and other ER stress-causing agents. This ER-stress-resp<strong>on</strong>se element<br />

proved to retain a novel structure and to be highly c<strong>on</strong>served in mammalian ERGIC-53 genes. The ERSE<br />

identified c<strong>on</strong>tains a 5’-end CCAAT sequence that c<strong>on</strong>stitutively binds NFY/CBF and, 9 nucleotides away, a<br />

3’-end regi<strong>on</strong> (5’-CCCTGTTGG-CCATC-3’) that is equally important <str<strong>on</strong>g>for</str<strong>on</strong>g> ER stress-mediated inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the gene. This sequence is the binding site <str<strong>on</strong>g>for</str<strong>on</strong>g> endogenous YY1 at the 5’-CCCTGTTGG-3’ part and <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

undefined factors at the CCATC 3-end. ATF6/YY1 but not XBP1, interacted with the ERGIC-53<br />

regulatory regi<strong>on</strong> and activated ERGIC-53 ERSE dependent transcripti<strong>on</strong>. Finally, we report that inducti<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> ER stress impaired the anterograde transport <str<strong>on</strong>g>of</str<strong>on</strong>g> reporter glycoproteins and alters the dinamics and the<br />

morphology <str<strong>on</strong>g>of</str<strong>on</strong>g> post-ER compartments. Our results suggest that increased levels <str<strong>on</strong>g>of</str<strong>on</strong>g> cargo receptor proteins<br />

might have a functi<strong>on</strong> either in the quality c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> protein folding in the endoplasmic reticulum or in the<br />

homeostasis <str<strong>on</strong>g>of</str<strong>on</strong>g> the intermediate compartment and Golgi complex during cell stress.<br />

2D_09_P<br />

(poster secti<strong>on</strong> A2, poster board #102, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CHRONIC AGGREGATION OF AN ER PROTEIN: EFFECTS ON PROTEIN<br />

HOMEOSTASIS AND THE CYTOSOLIC STRESS RESPONSE<br />

Diána Papp, Péter Csermely, Ákos Putics, Csaba Sőti<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Chemistry, Semmelweis University, Budapest, POBox 260, Hungary<br />

e-mail: pappdia@yahoo.com<br />

Protein c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>al diseases are important models to understand the role <str<strong>on</strong>g>of</str<strong>on</strong>g> protein homeostasis in the<br />

ageing <str<strong>on</strong>g>of</str<strong>on</strong>g> postmitotic tissues. One such model is alpha1-antitrypsin (AAT) deficiency, characterized by the<br />

accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a misfolding mutant (PiZ) inside the lumen <str<strong>on</strong>g>of</str<strong>on</strong>g> the hepatic endoplasmic reticulum (ER). The<br />

mutant protein harbors a Glu-Lys change, which induces a disrupti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a salt bridge, accompanied by an<br />

inability to pass the ER quality c<strong>on</strong>trol. Both human patients and PiZ transgenic mice have similar symptoms<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> hepatic failure, culminating in cirrhosis and hepatocellular carcinoma.<br />

The accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> misfolded proteins al<strong>on</strong>g the secretory pathway induces the unfolded protein resp<strong>on</strong>se<br />

(UPR). However, work from our and from other laboratories using either PiZ transgenic mice or<br />

overexpressing cell lines could neither detect inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ER chaper<strong>on</strong>es, nor the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> UPR<br />

mediators. Transgenic mice showed an interacti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> PiZ with protein disulfide isomerase (PDI), and a<br />

decreased protein disulfide reductase activity. Moreover, showing a crosstalk between the ER and the<br />

cytosolic compartment, both antioxidant and protein repair enzymes (thioredoxin and methi<strong>on</strong>ine sulfoxide<br />

reductase A) and the heat shock proteins (Hsp70 and Hsp90) were upregulated.<br />

To further elucidate the role <str<strong>on</strong>g>of</str<strong>on</strong>g> an ER protein in the general protein homeostasis at the cellular level, we<br />

engineered a transgenic liver cell line with inducible expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the wild type alpha-1-antitripsyn and the<br />

PiZ mutant. Results <str<strong>on</strong>g>of</str<strong>on</strong>g> our <strong>on</strong>going studies will be presented.<br />

105


23-26 August 2007,<br />

Budapest, Hungary<br />

106<br />

2E. HYPOXIA<br />

(JACQUES POUYSSEGUR, CORMAC T. TAYLOR)<br />

2E_01_S<br />

GAS SENSING IN THE NERVOUS SYSTEM: HYPOXIA AND POTASSIUM<br />

CHANNELS<br />

P. J. Kemp, S. P. Brazier, N. Baban, D. Riccardi, C. T. Müller, 1 S. J. Williams<br />

School <str<strong>on</strong>g>of</str<strong>on</strong>g> Biosciences, Cardiff University, UK<br />

e-mail: Kemp@cf.ac.uk.<br />

Whether part <str<strong>on</strong>g>of</str<strong>on</strong>g> the normal intra-uterine development, at high altitude or in pathological c<strong>on</strong>diti<strong>on</strong>s, hypoxia<br />

is <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the most comm<strong>on</strong> stresses to which an organism may be exposed and the ability to adapt to such<br />

changes in blood gases is crucial <str<strong>on</strong>g>for</str<strong>on</strong>g> optimal delivery <str<strong>on</strong>g>of</str<strong>on</strong>g> molecular oxygen to respiring tissues. The principal<br />

sensory comp<strong>on</strong>ent <str<strong>on</strong>g>of</str<strong>on</strong>g> this homeostatic mechanism is the carotid body. Ideally situated in the bifurcati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the comm<strong>on</strong> carotid artery, they resp<strong>on</strong>d muliplicatively to hypoxia, hypercapnia, pH and hypoglycaemia. At<br />

the cellular level, hypoxia promotes inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> plasma membrane the Ca 2+ -activated, K + channel (BK Ca ) <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

carotid body glomus cells which leads to Ca 2+ influx and transmitter release. Functi<strong>on</strong>al proteomics has<br />

recently dem<strong>on</strong>strated that hemeoxygenase-2 (HO-2) is an O 2 sensor linking hypoxia to BK Ca inhibiti<strong>on</strong><br />

(Williams et al., 2004 Science 306, 2093-2097), 2004; a process which depends up<strong>on</strong> carb<strong>on</strong> m<strong>on</strong>oxide (CO) as<br />

the sec<strong>on</strong>d messenger. The mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> such gas/channel interacti<strong>on</strong>s is complex, and may involve<br />

interacti<strong>on</strong>s with heme (Jaggar et al. Circ Res 97, 805-812, 2005. Using chemical and molecular modificati<strong>on</strong>s<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the BK Ca α-subunit in combinati<strong>on</strong> with channel chimera studies, we are now beginning to appreciate the<br />

kinetic and structural basis <str<strong>on</strong>g>of</str<strong>on</strong>g> the dynamic regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> BK Ca by endogenous producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> CO. Taken<br />

together, we have proposed a model <str<strong>on</strong>g>of</str<strong>on</strong>g> how HO-2 functi<strong>on</strong>s as a sensor <str<strong>on</strong>g>of</str<strong>on</strong>g> acute reducti<strong>on</strong>s in envir<strong>on</strong>mental<br />

O 2 . Thus, in normoxia, the protein partnership <str<strong>on</strong>g>of</str<strong>on</strong>g> HO-2 and BK Ca optimizes the permissive effect <str<strong>on</strong>g>of</str<strong>on</strong>g> CO.<br />

However, during the stress <str<strong>on</strong>g>of</str<strong>on</strong>g> hypoxia, the balance between intracellular heme c<strong>on</strong>centrati<strong>on</strong> and the<br />

evoluti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular CO is altered, thereby promoting channel inhibiti<strong>on</strong> and activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the carotid body.<br />

2E_02_S<br />

OXYGEN, A SOURCE OF LIFE AND STRESS – WHEN HYPOXIA MEETS CANCER<br />

E. Berra, M. C. Brahimi-Horn, J. Chiche, F. Dayan, R. Garcia-Medina, A. Ginouvès, N. Mazure,<br />

D. Roux, M-P. Sim<strong>on</strong>, J. Pouysségur<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Signaling, Developmental Biology and Cancer <str<strong>on</strong>g>Research</str<strong>on</strong>g>, CNRS-UMR 6543, <str<strong>on</strong>g>Centre</str<strong>on</strong>g> Antoine<br />

Lacassagne, 33 Avenue de Valombrose, 06189 Nice, France<br />

e-mail: pouysseg@unice.fr<br />

The development <str<strong>on</strong>g>of</str<strong>on</strong>g> an oxygen-rich atmosphere has been <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the most important events in the history <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

life <strong>on</strong> Earth. Oxygen with its excepti<strong>on</strong>al reactivity, represents the far most potent natural polluti<strong>on</strong> <strong>on</strong> our<br />

planet, promoting Life or Death. About 2.4 billi<strong>on</strong> years ago the high amount <str<strong>on</strong>g>of</str<strong>on</strong>g> dissolved and free oxygen,<br />

produced by photosynthesis, in the oceans and atmosphere has driven to extincti<strong>on</strong> most anaerobic<br />

organisms. Over the past 500 milli<strong>on</strong> years, oxygen levels fluctuated between 15 and 35% imposing c<strong>on</strong>stant<br />

stress <strong>on</strong> and subsequent stringent evoluti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> living organisms.<br />

During mammalian embry<strong>on</strong>ic development or in the c<strong>on</strong>text <str<strong>on</strong>g>of</str<strong>on</strong>g> tumor expansi<strong>on</strong>, proliferating cells rapidly<br />

outstrip the supply <str<strong>on</strong>g>of</str<strong>on</strong>g> nutrients. Although cells sense and resp<strong>on</strong>d to variati<strong>on</strong>s in c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> all


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

nutrients, oxygen sensing has emerged as a central c<strong>on</strong>trol mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> vasculogenesis. Whereas a decrease<br />

in the pO 2 (hypoxic stress) induces angiogenesis, an increase in pO 2 (hyperoxic stress) induces vascular<br />

pruning. Oxygen c<strong>on</strong>centrati<strong>on</strong>s ‘sculpt’ the blood vascular network <str<strong>on</strong>g>of</str<strong>on</strong>g> vertebrates. At the heart <str<strong>on</strong>g>of</str<strong>on</strong>g> this<br />

regulatory system is the Hypoxia-Inducible Factor, HIF, which interestingly c<strong>on</strong>trols, am<strong>on</strong>g other gene<br />

products, the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> VEGF-A and Angiopoïetin-2 (Ang-2), two key angiogenic factors. This finding<br />

has there<str<strong>on</strong>g>for</str<strong>on</strong>g>e placed the hypoxia-signaling pathway at the <str<strong>on</strong>g>for</str<strong>on</strong>g>efr<strong>on</strong>t <str<strong>on</strong>g>of</str<strong>on</strong>g> nutriti<strong>on</strong>al c<strong>on</strong>trol. Rapidly activated<br />

up<strong>on</strong> a hypoxic stress, HIF induces a vast array <str<strong>on</strong>g>of</str<strong>on</strong>g> gene products inducing cell-, tissue-, and organismalsurvival.<br />

Am<strong>on</strong>g the HIF-c<strong>on</strong>trolled functi<strong>on</strong>s are inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ATP-c<strong>on</strong>suming processes (protein and lipid<br />

synthesis), inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>drial respirati<strong>on</strong> (to save O 2 ), increase in anaerobic glucose metabolism,<br />

regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> intracellular pH, increased angiogenesis and cell migrati<strong>on</strong>, and so HIF has become recognized<br />

as a str<strong>on</strong>g promoter <str<strong>on</strong>g>of</str<strong>on</strong>g> tumor growth. This pro-<strong>on</strong>cogenic feature is <strong>on</strong>ly <strong>on</strong>e facet <str<strong>on</strong>g>of</str<strong>on</strong>g> the dual acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

HIF. Besides being a ‘guardian’ <str<strong>on</strong>g>of</str<strong>on</strong>g> oxygen homeostasis, HIF is capable <str<strong>on</strong>g>of</str<strong>on</strong>g> inducing pro-apoptotic gene<br />

products (BNIP3, BNIP3L) that are in fact pro-survival by inducing autophagy. The molecular mechanism<br />

leading to this survival process that is strictly c<strong>on</strong>trolled by a drop in pO 2 will be presented.<br />

2E_03_S<br />

HYPOXIA AND INFLAMMATION<br />

Cormac T. Taylor<br />

UCD C<strong>on</strong>way Institute, University College Dublin, Belfield, Dublin 4, Ireland. e-mail: cormac.taylor@ucd.ie<br />

The chemical reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> molecular oxygen which occurs during mitoch<strong>on</strong>drial oxidative phosphorylati<strong>on</strong> is<br />

the main source <str<strong>on</strong>g>of</str<strong>on</strong>g> metabolic energy <str<strong>on</strong>g>for</str<strong>on</strong>g> virtually all eukaryotic cells. Decreased tissue oxygen supply (leading<br />

to hypoxic stress) is a comm<strong>on</strong> feature in a range <str<strong>on</strong>g>of</str<strong>on</strong>g> disease states where inflammati<strong>on</strong> occurs including<br />

inflammatory bowel disease (IBD), arthritis, cancer, atherosclerosis and stroke. Recent studies indicate that<br />

hypoxia promotes inflammatory signaling pathways through specific mechanisms involving altered<br />

hydroxylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> specific residues <strong>on</strong> key transcripti<strong>on</strong> factors including (but likely not limited to) the hypoxia<br />

inducible factor (HIF-1) and nuclear factor kappa B (NF-κB). Thus it appears that hypoxia plays an<br />

important modulatory role in inflammatory disease development. It appears that a family <str<strong>on</strong>g>of</str<strong>on</strong>g> prolyl- and<br />

asparaginyl-hydroxylases are key comm<strong>on</strong> oxygen sensors in c<strong>on</strong>ferring hypoxic sensitivity to these pathways.<br />

These hydroxylases are absolutely dependent up<strong>on</strong> the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> molecular oxygen <str<strong>on</strong>g>for</str<strong>on</strong>g> activity and are thus<br />

inhibited in hypoxia leading to derepressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> transcripti<strong>on</strong>al effectors. A greater understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

oxygen sensing and signaling mechanisms leading to the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these transcripti<strong>on</strong>al resp<strong>on</strong>ses to<br />

hypoxia will allow the development <str<strong>on</strong>g>of</str<strong>on</strong>g> novel therapeutics in a range <str<strong>on</strong>g>of</str<strong>on</strong>g> disease states where hypoxia and<br />

inflammati<strong>on</strong> are co-incidental events.<br />

107


23-26 August 2007,<br />

Budapest, Hungary<br />

108<br />

2E_04_S<br />

STRESS REGULATED BHLH/PAS TRANSCRIPTION FACTORS: THE DIOXIN<br />

RECEPTOR AND HYPOXIA INDUCIBLE FACTORS<br />

M. L. Whitelaw, S. Linke, A. Chapman-Smith, T. Wallis, K. Dave, J. Gorman, D. J. Peet<br />

School <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular and Biomedical Sciences and ARC <str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g> the Molecular Genetics <str<strong>on</strong>g>of</str<strong>on</strong>g> Development,<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Adelaide, South Australia<br />

e-mail: murray.whitelaw@adelaide.edu.au<br />

The basic Helix-Loop-Helix / Per-Arnt-Sim (bHLH/PAS) family <str<strong>on</strong>g>of</str<strong>on</strong>g> transcripti<strong>on</strong> factors per<str<strong>on</strong>g>for</str<strong>on</strong>g>m essential<br />

functi<strong>on</strong>s during early development and help maintain homeostasis in the adult. For example, the Hypoxia<br />

Inducible Factors (HIF-1a and HIF-2a proteins) play a major role in angiogenesis and cellular adapti<strong>on</strong> to low<br />

oxygen stress. At normoxia, two oxygen dependent hydroxylases posttranslati<strong>on</strong>ally modify specific proline<br />

and asparagine residues <str<strong>on</strong>g>of</str<strong>on</strong>g> the HIFs, severely dampening their activity. During hypoxia, the HIFs exhibit<br />

dramatic increases in both protein stability and intrinsic transactivati<strong>on</strong> capacity, due to attenuati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these<br />

two hydroxylases. The Dioxin Receptor (or Aryl hydrocarb<strong>on</strong> receptor) resp<strong>on</strong>ds to the stress <str<strong>on</strong>g>of</str<strong>on</strong>g> xenobiotic<br />

infiltrati<strong>on</strong> by inducing a battery <str<strong>on</strong>g>of</str<strong>on</strong>g> genes <str<strong>on</strong>g>for</str<strong>on</strong>g> xenobiotic metabolism. The Dioxin Receptor (DR) is also the<br />

mediator <str<strong>on</strong>g>of</str<strong>on</strong>g> toxic resp<strong>on</strong>ses to dioxins and PCBs. Both HIFs and the DR need to heterodimerise with a<br />

central bHLH/PAS partner protein, termed Arnt, to <str<strong>on</strong>g>for</str<strong>on</strong>g>m active transcripti<strong>on</strong> factor complexes. The PAS<br />

domain provides a critical protein interacti<strong>on</strong> surface during dimerisati<strong>on</strong> and in the case <str<strong>on</strong>g>of</str<strong>on</strong>g> the DR, functi<strong>on</strong>s<br />

as a signal regulated domain. During our studies <str<strong>on</strong>g>of</str<strong>on</strong>g> stress induced activati<strong>on</strong> mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> the HIFs and<br />

DR, we have found recurring themes <str<strong>on</strong>g>of</str<strong>on</strong>g> posttranslati<strong>on</strong>al modificati<strong>on</strong> and an important role <str<strong>on</strong>g>of</str<strong>on</strong>g> the PAS<br />

domain in allowing the bHLH to bind to n<strong>on</strong>-can<strong>on</strong>ical E-box sequences <str<strong>on</strong>g>of</str<strong>on</strong>g> DNA. The presentati<strong>on</strong> will<br />

present data to illustrate and expand up<strong>on</strong> these themes.<br />

2E_05_S<br />

(poster secti<strong>on</strong> B1, poster board #228, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SPECIFIC FKBP38 INHIBITOR REDUCES HYPOXIA-INDUCED APOPTOSIS IN<br />

VENTRICULAR MYOCYTES FROM ADULT AND NEONATAL RAT HEARTS<br />

S. Rau 2 , B. Husse 1 , D. Wildemann 2 , M. Gekle 1 , G. Fischer 2<br />

1Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Physiology, University Halle and 2 Max-Planck <str<strong>on</strong>g>Research</str<strong>on</strong>g> Unit <str<strong>on</strong>g>for</str<strong>on</strong>g> Enzymology <str<strong>on</strong>g>of</str<strong>on</strong>g> Protein Folding, Halle,<br />

Germany<br />

e-mail: britta.husse@medizin.uni-halle.de<br />

Hypoxia/reoxygenati<strong>on</strong> causes cell death <str<strong>on</strong>g>of</str<strong>on</strong>g> cardiomyocytes by a mitoch<strong>on</strong>dri<strong>on</strong>-dependent pathway. The<br />

Ca 2+ /CaM activated FK506-binding protein 38 (FKBP38) can interact with Bcl-2 through its PPIase active<br />

site and participates in the promoti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis. This study investigated the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> specific FKBP38<br />

inhibiti<strong>on</strong> with DM-CHX (N-(N',N'-dimethylcarboxamidomethyl)-cycloheximide) <strong>on</strong> the<br />

hypoxia/reoxygenati<strong>on</strong>-induced apoptosis. Ventricular myocytes from adult or ne<strong>on</strong>atal rates were cultured<br />

and subjected to hypoxic c<strong>on</strong>diti<strong>on</strong>s (0.2% O 2 ) <str<strong>on</strong>g>for</str<strong>on</strong>g> 18 or 24 hrs (adult/ne<strong>on</strong>atal) followed by a reoxygenati<strong>on</strong><br />

period (21% O 2 ) <str<strong>on</strong>g>of</str<strong>on</strong>g> 24 hrs. Hypoxic c<strong>on</strong>diti<strong>on</strong> was proved by HIF1-alpha expressi<strong>on</strong> using western blots.<br />

Apoptotic cell analysis was determined by using the ViaCount assay (Guava Technologies). In adult myocytes,<br />

hypoxia caused 34% and hypoxia/reoxygenati<strong>on</strong> 31% apoptotic cells. Applicati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> DM-CHX (5µM)<br />

resulted in 22% (hypoxia) and 11% (hypoxia/reoxygenati<strong>on</strong>) apoptotic cells. In ne<strong>on</strong>atal myocytes, under<br />

both c<strong>on</strong>diti<strong>on</strong>s 64% apoptotic cells were analysed, reduced to 34% after DM-CHX (5 µM) treatment. As


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

positive c<strong>on</strong>trol, caspase-inhibiti<strong>on</strong> and cyclosporin A showed apoptosis inhibiti<strong>on</strong> in both types <str<strong>on</strong>g>of</str<strong>on</strong>g> myocytes.<br />

Hypoxia (2.5-fold) and hypoxia/reoxygenati<strong>on</strong> (1.4-fold) caused injury <str<strong>on</strong>g>of</str<strong>on</strong>g> adult myocytes measured by relative<br />

lactate dehydrogenase activity, which was reduced by DM-CHX (5 µM) treatment. Our results suggest that<br />

DM-CHX, a specific inhibitor <str<strong>on</strong>g>of</str<strong>on</strong>g> FKBP38, reduces apoptosis in cardiomyocytes in a dose-dependent manner.<br />

Such a specific drug could be used to decrease the loss <str<strong>on</strong>g>of</str<strong>on</strong>g> myocytes after damaged injury resulting in an<br />

improved cardiac functi<strong>on</strong>.<br />

2E_06_S<br />

(poster secti<strong>on</strong> B1, poster board #229, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

NF-KB FUNCTIONS THE DIVERSITY OF CELLULAR IGF-I/IGFBP-1 EXPRESSION<br />

BY HYPOXIA IN TIBETAN PLATEAU MAMMALS<br />

X. Q. Chen, S. J. Wang, Y. Liu, J. Z. Du<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Physiology, Divi. Neurobiology and Physiology, School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Zhejiang University, Hangzhou<br />

310058, China<br />

e-mail: dujz@cls.zju.edu.cn<br />

Ochot<strong>on</strong>a curz<strong>on</strong>iae, Microtus oec<strong>on</strong>omus and Myospalax baileyi are all native mammals that reside at<br />

Qinghai-Tibetan plateau in China and well acclimatized to envir<strong>on</strong>mental hypoxia. The present paper<br />

addresses the NF-kB’s, a nuclear transcripti<strong>on</strong>al factor, involvement in hypoxia stress-induced diversity <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

IGF-I/IGFBP-1 expressi<strong>on</strong> in hepatic and brain cells <str<strong>on</strong>g>of</str<strong>on</strong>g> Tibetan Plateau mammals. The IGF-I/IGFBP-1<br />

from the prefr<strong>on</strong>tal cortex and the liver cells was tested 6 h after hypoxia exposure (by CoCl 2 injecti<strong>on</strong> i.p. 20,<br />

40 mg/kg or by normobaric hypoxia, 16.0%, 10.8%, 8.0 %O 2 ) <str<strong>on</strong>g>of</str<strong>on</strong>g> the Plateau native mammals and mice.<br />

PDTC, an inhibitor <str<strong>on</strong>g>of</str<strong>on</strong>g> NF-κB, was used and preinjected be<str<strong>on</strong>g>for</str<strong>on</strong>g>e the hypoxia to evaluated NF-kB acti<strong>on</strong>. The<br />

results showed that 1) the IGF-I expressi<strong>on</strong> in mice hepatic cells <str<strong>on</strong>g>of</str<strong>on</strong>g> M. oec<strong>on</strong>omus and M. baileyi markedly<br />

increased after the hypoxia exposure, but there was no resp<strong>on</strong>se in the liver <str<strong>on</strong>g>of</str<strong>on</strong>g> O.curz<strong>on</strong>iae; 2) the IGFBP-1<br />

expressi<strong>on</strong> in mice hepatic cells <str<strong>on</strong>g>of</str<strong>on</strong>g> O. curz<strong>on</strong>iae and M. baileyi markedly enhanced, but no resp<strong>on</strong>se occurred in<br />

M. oec<strong>on</strong>omus after the hypoxia; 3) PDTC pretreated be<str<strong>on</strong>g>for</str<strong>on</strong>g>e hypoxia reversed the hypoxia-enhanced IGF-I in<br />

M. oec<strong>on</strong>omus and M. baileyi; 4) PDTC treatment also reversed the hypoxia-enhanced IGFBP-1 in O. curz<strong>on</strong>iae<br />

and M. baileyi; 5) hypoxia increased the IGF-I mRNA in brain <str<strong>on</strong>g>of</str<strong>on</strong>g> M. oec<strong>on</strong>omus and O. curz<strong>on</strong>iae but not <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

mice; 6) hypoxia did not induce changes <str<strong>on</strong>g>of</str<strong>on</strong>g> IGF-I levels in the brain cells <str<strong>on</strong>g>of</str<strong>on</strong>g> both plateau mammals and<br />

laboratory mice. The data suggest that 1) different pattern in IGF-I/IGFBP-1 expressi<strong>on</strong> induced by hypoxia<br />

represents a diversities in horm<strong>on</strong>e regulati<strong>on</strong> and cell protecti<strong>on</strong> from damage in Tibetan native mammals; 2)<br />

NF-κB mediates the transcripti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> IGF-I/IGFBP-1 in liver cells subjecting to hypoxia; Together, the<br />

diversity <str<strong>on</strong>g>of</str<strong>on</strong>g> target-gene phenotype expressi<strong>on</strong> may c<strong>on</strong>tribute to the multi-model in cell protecti<strong>on</strong> from<br />

hypoxia damage.Acknowledgement: This work was supported by the NSFC: Projects (No.<br />

30393130;30470648; 30570227), and by The Nati<strong>on</strong>al Basic <str<strong>on</strong>g>Research</str<strong>on</strong>g> Program “973” No. 2006CB504100).<br />

109


23-26 August 2007,<br />

Budapest, Hungary<br />

2E_01_P<br />

(poster secti<strong>on</strong> B1, poster board #230, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PRENATAL HYPOXIA STRESS INDUCED THE HIGH SENSITIVITY OF HPA IN<br />

ADULT OFFSPRING RATS<br />

X. Q Chen,* J. Z. Du*, J. M. Fan<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Physiology, Divi. Neurobiology and Physiology, School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Zhejiang University,<br />

New Campus, Hangzhou, 310058, China<br />

*e-mail: dujz@cls.zju.edu.cn; chewyg@zju.edu.cn<br />

The hypothalamo-pituitary-adrenal (HPA) axis is affected intensively by envir<strong>on</strong>ment stress in early life. We<br />

invistigated whether CRFR1 or CRFR2 are involved or not in modificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the <str<strong>on</strong>g>of</str<strong>on</strong>g>fspring’s HPA activity<br />

and behavior resp<strong>on</strong>se after maternal hypoxia stress (MHS). The pregnant mother rats were exposed to<br />

different stressors: simulated hypobaric hypoxia (5km altitude, 10.8% O 2 ), restraint, cold (4ºC), and in<br />

combinati<strong>on</strong> stress (Hypoxia+Restraint, Hypoxia+Cold, Hypoxia+Restraint+Cold) throughout the pregnant<br />

period 4h per day <str<strong>on</strong>g>for</str<strong>on</strong>g> 22 days, and the growth parameters <str<strong>on</strong>g>of</str<strong>on</strong>g> postnatal rats, HPA axis activity and<br />

neuroendocrine functi<strong>on</strong> during growing and late age <str<strong>on</strong>g>of</str<strong>on</strong>g> puberty rats were determinated, anxiety-like behavior<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> adult male rats was evaluated by elevated-plus maze (EPM), and the changes <str<strong>on</strong>g>of</str<strong>on</strong>g> CRFR1 mRNA and<br />

CRFR2 mRNA expressi<strong>on</strong> in pituitary <str<strong>on</strong>g>of</str<strong>on</strong>g> both late age <str<strong>on</strong>g>of</str<strong>on</strong>g> puberty and adult male rats were measured by in<br />

situ hybridizaiti<strong>on</strong>. The plasma corticoster<strong>on</strong>, ACTH, were tested using RIA. the NE, and DA levels in Locus<br />

Coeruleus (LC) determinated by HPLC. The research found that MHS significantly decreased the birth<br />

weight and delayed the growth rate in postnatal rats, enhanced the basal activity <str<strong>on</strong>g>of</str<strong>on</strong>g> HPA axis, the expressi<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> CRFR1 mRNA /CRFR2 mRNA in pituitary <str<strong>on</strong>g>of</str<strong>on</strong>g> late age <str<strong>on</strong>g>of</str<strong>on</strong>g> puberty rats, induced anxiety-like behavior,<br />

elevated the level <str<strong>on</strong>g>of</str<strong>on</strong>g> NE and DA in the LC. These data suggest that perinatal stress significantly infuluences<br />

HPA axis activity and behavior <str<strong>on</strong>g>of</str<strong>on</strong>g> adult <str<strong>on</strong>g>of</str<strong>on</strong>g>fspring rat, and the CRFR1 and CRFR2 may play a role in<br />

modificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these activities by pregnent maternal hypoxia and combnated stress. Acknowledgement:This<br />

work was supported by the Nati<strong>on</strong>al Science Foundati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> China: Major Project (No. 30393130) and<br />

Projects (Nos. 30270232; 30470648; 30570227), and by the China Ministry <str<strong>on</strong>g>of</str<strong>on</strong>g> Science and Technology (The<br />

Nati<strong>on</strong>al Basic <str<strong>on</strong>g>Research</str<strong>on</strong>g> Program “973” No. 2006CB504100).<br />

2E_02_P<br />

(poster secti<strong>on</strong> B1, poster board #231, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DEFENSE OF THERMAL AND OXYGEN STRESS IN DAPHNIA MAGNA<br />

Bettina Zeis, Sabine Zumbrägel, Rüdiger J. Paul<br />

Institut für Zoophysiologie, Hindenburgplatz 55, 48143 Münster, Germany<br />

e-mail: zeis@uni-muenster.de<br />

Small plankt<strong>on</strong>ic animals like Daphnia experience high fluctuati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> envir<strong>on</strong>mental oxygen c<strong>on</strong>tent and<br />

temperature, resulting in changes <str<strong>on</strong>g>of</str<strong>on</strong>g> oxygen supply or demand, respectively. Hypoxia or exposure to high<br />

temperatures may lead to the <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> oxygen radicals (reactive oxygen species, ROS) and changes in the<br />

cellular redox state. This may activate a biphasic stress resp<strong>on</strong>se system. A fast comp<strong>on</strong>ent <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress<br />

resp<strong>on</strong>se aims to reduce radical damage until cellular homeostasis is restored by the slower comp<strong>on</strong>ent<br />

(Kültz, 2005, Annu Rev Physiol 67:225).<br />

110


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

In the microcrustacean Daphnia magna, hemoglobin (Hb) is the central effector protein <str<strong>on</strong>g>of</str<strong>on</strong>g> the homeostasis<br />

resp<strong>on</strong>se to restore aerobic metabolism after oxygen- or temperature-induced impairment <str<strong>on</strong>g>of</str<strong>on</strong>g> tissue<br />

oxygenati<strong>on</strong>. Differential expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a set <str<strong>on</strong>g>of</str<strong>on</strong>g> Hb orthologs enhances both, c<strong>on</strong>centrati<strong>on</strong> and oxygen<br />

affinity <str<strong>on</strong>g>of</str<strong>on</strong>g> the respiratory protein. We studied the inducti<strong>on</strong> process <str<strong>on</strong>g>of</str<strong>on</strong>g> Hb following oxygen or temperature<br />

stress <strong>on</strong> the mRNA and protein level. We also detected changes in the oxygen-sensitive transcripti<strong>on</strong> factor<br />

HIF (hypoxia-inducible factor), which is under c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> ROS.<br />

Analysing the radical defense system under defined oxygen and temperature c<strong>on</strong>diti<strong>on</strong>s (Po 2 : 2, 21, and<br />

60 kPa at 20 °C and 10, 20, and 30 °C at normoxia: 21 kPa), we found that the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> catalase was slightly<br />

increased under hypoxia and at elevated temperature. The c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the redox buffer glutathi<strong>on</strong>e<br />

(GSH, GSSG) was increased at hyperoxic c<strong>on</strong>diti<strong>on</strong>s and high temperatures; the same was true <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

activity <str<strong>on</strong>g>of</str<strong>on</strong>g> glutathi<strong>on</strong>e-S-transferases. Comparing thresholds <str<strong>on</strong>g>for</str<strong>on</strong>g> the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress resp<strong>on</strong>se system<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> different cl<strong>on</strong>al lines allows us to discriminate genetically determined comp<strong>on</strong>ents from acclimati<strong>on</strong><br />

effects.<br />

2E_03_P<br />

(poster secti<strong>on</strong> B1, poster board #232, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HYPOXIC STRESS DISRUPTS CELL PROLIFERATION AND APOPTOSIS IN NON-<br />

CANCER TISSUES VIA TELOMERASE: A WHOLE FISH ANIMAL MODEL<br />

D. W. T. Au*, E. X. H. Chen, R. M. K. Yu, M. W. L. Chiang, H. O. L. Mok, B. W. P. Yip,<br />

R. Y. C. K<strong>on</strong>g<br />

Biology & Chemistry Department, City University <str<strong>on</strong>g>of</str<strong>on</strong>g> H<strong>on</strong>g K<strong>on</strong>g, 83 Tat Chee Avenue, Kowlo<strong>on</strong>, H<strong>on</strong>g K<strong>on</strong>g<br />

e-mail: bhdwtau@cityu.edu.hk<br />

We have employed the small fish species marine medaka (Oryzia melastigma) as a vertebrate model <str<strong>on</strong>g>for</str<strong>on</strong>g> studying<br />

whole animal resp<strong>on</strong>se under chr<strong>on</strong>ic hypoxia, with special interest to investigate regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> telomerase<br />

(TERT) expressi<strong>on</strong> via hypoxia-inducible factor and to link telomerase expressi<strong>on</strong> to cell proliferati<strong>on</strong> and<br />

apotptosis in n<strong>on</strong>-tumor tissues in vivo under hypoxia. The O. melastigma exhibits telomere length resembling<br />

those <str<strong>on</strong>g>of</str<strong>on</strong>g> humans. The full-length TERT cDNA, omTERT, encodes a protein <str<strong>on</strong>g>of</str<strong>on</strong>g> 1086 amino acids and<br />

c<strong>on</strong>tains all <str<strong>on</strong>g>of</str<strong>on</strong>g> the functi<strong>on</strong>al motifs that are c<strong>on</strong>served in other vertebrate TERTs. The omTERT gene is<br />

ubiquitously expressed in all fish tissues under normoxia, where highest expressi<strong>on</strong> was observed in g<strong>on</strong>ads<br />

and the lowest in liver. In vivo expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> omTERT was significantly upregulated in testis and liver in<br />

resp<strong>on</strong>se to hypoxia (at 96 h and 48 h, respectively), where c<strong>on</strong>comitant inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the hypoxia inducible<br />

factor-1α (omHIF-1α) and erythropoietin (omEpo) genes was also observed. Analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> the 5′-flanking<br />

sequence <str<strong>on</strong>g>of</str<strong>on</strong>g> the omTERT gene identified two HRE (hypoxia-resp<strong>on</strong>sive element; nt. −283 and −892) cores.<br />

Overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the HIF-1α induced omTERT promoter activity as dem<strong>on</strong>strated using transient<br />

transfecti<strong>on</strong> assays. The in vivo effects <str<strong>on</strong>g>of</str<strong>on</strong>g> TERT expressi<strong>on</strong> <strong>on</strong> tissue homeostasis (involving a balance <str<strong>on</strong>g>of</str<strong>on</strong>g> cell<br />

proliferati<strong>on</strong> and apoptosis) in whole vertebrate are poorly known. We have optimized a fixati<strong>on</strong> and tissue<br />

processing protocol <str<strong>on</strong>g>for</str<strong>on</strong>g> the adult medaka fish which has proven successful <str<strong>on</strong>g>for</str<strong>on</strong>g> quantitative in situ<br />

hybridizati<strong>on</strong> (Q-ISH) and immuno-histochemisty (Q-IHC). This whole fish model allows us to study the coexpressi<strong>on</strong><br />

pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iles <str<strong>on</strong>g>of</str<strong>on</strong>g> omTERT gene and TERT protein in multiple organs <str<strong>on</strong>g>of</str<strong>on</strong>g> marine medaka simultaneously,<br />

and to relate TERT expressi<strong>on</strong> to cell proliferati<strong>on</strong> (by PCNA) and apoptosis (TUNEL) in vivo. We found<br />

that hypoxia induces cell/tissue specific changes in expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> omTERT mRNA and protein, telomerase<br />

activity, cell proliferati<strong>on</strong>, apoptosis in adult medaka. The findings <str<strong>on</strong>g>of</str<strong>on</strong>g> our whole fish model will shed light <strong>on</strong><br />

hypoxic stress and telomerase biology in vertebrates in vivo.<br />

111


23-26 August 2007,<br />

Budapest, Hungary<br />

112<br />

2E_04_P<br />

(poster secti<strong>on</strong> B1, poster board #233, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CLONING AND EVOLUTION OF HIF-1Α CDNA FROM TIBETAN VERTEBRATES:<br />

PANTHOLOPS HODGSONI, MYOSPALAX BAILEYI, MYOSPALAX CANSUS,<br />

MICROTUS OECONOMUS, GYMNOCYPRIS PRZEWALSKII, AND EPO<br />

RESPONSIVNESS UNDER HYPOXIA<br />

X. Q. Chen a *, J. Z. Du a *, Y. B. Cao a , Y. Liu a , X-C. Chen b , G. Cai c , P. H. Xiao c<br />

aDept.Physiology, School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Zhejiang University, Hangzhou 310058, China<br />

bNorthwest Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Plateau Biology, The Chinese Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences<br />

cQinghai Kekexili Nati<strong>on</strong>al Nature Reserve,<br />

*e-mail: dujz@cls.zju.edu.cn<br />

Hypoxia-inducible factor 1α(HIF-1α) is an essential mediator <str<strong>on</strong>g>of</str<strong>on</strong>g> oxygen homeostasis. Tibetan<br />

antelope(P.hodgs<strong>on</strong>i), Plateau pika(O.curz<strong>on</strong>iae), Plateau Zokor(M.baileyi,M.cansus), Root vole(M. oec<strong>on</strong>omus), and<br />

naked carp(G.przewalskii) are native vertebrates at Qinghai-Tibetan plateau and well acclimatized to hypoxia.<br />

To better understand the adaptive mechanisms to hypoxia, cDNA encoding HIF-1α was isolated and<br />

characterized from those vertebrates. The deduced HIF-1α sequences <str<strong>on</strong>g>of</str<strong>on</strong>g> P. hodgs<strong>on</strong>i, M. oec<strong>on</strong>omus, M. baileyi<br />

and M.cansus showed 90-99% identities with those <str<strong>on</strong>g>of</str<strong>on</strong>g> the human, rat, yak, respectively; G.przewalskii 89-57%<br />

with that <str<strong>on</strong>g>of</str<strong>on</strong>g> rainbow trout and comm<strong>on</strong> carp. The c<strong>on</strong>servati<strong>on</strong>al and phylogenic clustering <str<strong>on</strong>g>of</str<strong>on</strong>g> vertebrates<br />

HIF-1α sequences was c<strong>on</strong>sistent with the vertebrate classificati<strong>on</strong>. We estimated cod<strong>on</strong>s under positive<br />

selecti<strong>on</strong>. All positive selecti<strong>on</strong> site were not in key domain, but between the key domain <str<strong>on</strong>g>of</str<strong>on</strong>g> TAD-N and<br />

TAD-C(594L, 600L,604T,605E,606E,607L,610V,615T,628T,646Q), and another was 8E in M. baileyi. The<br />

evidence show that plateau animals could have peculiarities <str<strong>on</strong>g>for</str<strong>on</strong>g> hypoxia, which linked to changes <str<strong>on</strong>g>of</str<strong>on</strong>g> Tibetan<br />

Plateau growing. HIF-1 increased in cortex and liver <str<strong>on</strong>g>of</str<strong>on</strong>g> mice, M. Baileyi, and M. oec<strong>on</strong>omus under hypoxia not in<br />

O. curz<strong>on</strong>iae; EPO increased in cortex and kidney <str<strong>on</strong>g>of</str<strong>on</strong>g> mice under hypoxia, but <strong>on</strong>ly in kidney <str<strong>on</strong>g>of</str<strong>on</strong>g> M. oec<strong>on</strong>omus<br />

under hypoxia. Additi<strong>on</strong>ally, under CoCl2, EPO increased in cortex and kidney <str<strong>on</strong>g>of</str<strong>on</strong>g> three Tibetan mammals not<br />

in mice. The differences <str<strong>on</strong>g>of</str<strong>on</strong>g> HIF and EPO in Tibetan animals and lowland mice suggest diversive strategies<br />

are involved in hypoxia. HIF-1 may play an role in mice and M. Baileyi not in Tibetan's O.curz<strong>on</strong>iae <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

hypoxia. The work was supported by the NSFC: (No. 30393130;30470648; 30570227 and by The Program<br />

“973” No. 2006CB504100).<br />

2E_05_P<br />

(poster secti<strong>on</strong> B1, poster board #234, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

COMBINED CASPASE AND CALPAIN INHIBITION OF PC12 AND NGF TREATED<br />

CELLS AFTER GLUCOSE AND OXYGEN DEPRIVATION INDUCED STRESS<br />

C. Pourzitaki 1 , I. Klagas 1 , A. Sioga 2 , A. Kaidoglou 2 , A. Kritis 3<br />

1Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmacology,<br />

2Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Histology and Embryology and<br />

3Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Physiology, Medical School, Aristotle University, Greece<br />

e-mail: Kritis@med.auth.gr<br />

Hypoxia-induced dysfuncti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the central nervous system may be caused by neur<strong>on</strong>al cell death or by<br />

changes in its neurochemical implicated in the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cell death. PC12 cells may serve as a useful model<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> studying the interacti<strong>on</strong> between hypoxia, and cell death. Programmed cell death (apoptosis) plays an<br />

important role in a wide variety <str<strong>on</strong>g>of</str<strong>on</strong>g> physiological processes, as well as in pathological cellular insults.


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Abnormal elevati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> intracellular Ca 2 is thought to be a critical trigger <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>al damage associated with<br />

hypoxia or ischemia. Several lines <str<strong>on</strong>g>of</str<strong>on</strong>g> evidence suggest that calcium-activated proteolysis plays a pivotal role in<br />

hypoxic-ischemic neur<strong>on</strong>al damage. Indeed, calpain-mediated proteolysis appears to be <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the earliest<br />

biochemical changes occurring after a dense ischemic challenge. In the present study, we tested the effects <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

z-VAD-fmk as a specific caspase inhibitor combined with MDL28170 a new and more permeable calpain<br />

inhibitor during oxygen and glucose deprivati<strong>on</strong> in naïve and NGF treated PC12 cells. Our results show that<br />

both protease inhibitors c<strong>on</strong>fer comparable cell death resistance to both naive and NGF treated PC12 cells at<br />

a c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 10µΜ. It seems that <strong>on</strong>ce the death signal is released the cell has to employ either<br />

apoptosis or necrosis in order to effect death. Since both classes <str<strong>on</strong>g>of</str<strong>on</strong>g> proteases are implicated in cell death in<br />

our system under the c<strong>on</strong>diti<strong>on</strong>s studied, we hypothesize that the two processes, necrosis and apoptosis, are<br />

not exclusive to themselves and that at some stages they can share functi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> their machinery to <strong>on</strong>e<br />

another.<br />

2E_06_P<br />

(poster secti<strong>on</strong> B1, poster board #235, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

BIOLGICAL MARKERS OF STRESS IN COLONIC HEALING AFTER PORTAL<br />

ISCHEMIA-REPERFUSION. EXPERIMENTAL STUDY<br />

A. Schanaider, M. Rolim, G. Cotta-Pereira, E. C. A. Eleutherio, P. C. Silva, C. B. Aratanha<br />

Article and <str<strong>on</strong>g>Research</str<strong>on</strong>g> from the Center <str<strong>on</strong>g>of</str<strong>on</strong>g> Experimental Surgery <str<strong>on</strong>g>of</str<strong>on</strong>g> the Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Surgery –<br />

Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine <str<strong>on</strong>g>of</str<strong>on</strong>g> Federal University <str<strong>on</strong>g>of</str<strong>on</strong>g> Rio de Janeiro<br />

PURPOSE: To develop the knowledge about the pathophysiology <str<strong>on</strong>g>of</str<strong>on</strong>g> col<strong>on</strong>ic healing in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

portal c<strong>on</strong>gesti<strong>on</strong> and to identify biological markers <str<strong>on</strong>g>of</str<strong>on</strong>g> this oxidative stress.<br />

METHODS: Seventy male Wistar albino adult young rats (Rattus novergicus albinus), weighting 250-350g were<br />

enrolled. Anesthetic procedure c<strong>on</strong>sists <str<strong>on</strong>g>of</str<strong>on</strong>g> Sev<str<strong>on</strong>g>of</str<strong>on</strong>g>lurane inhalati<strong>on</strong>. Four groups <str<strong>on</strong>g>of</str<strong>on</strong>g> experiment were<br />

established: G1 - C<strong>on</strong>trol (n = 10), G2 - Col<strong>on</strong>ic anastomosis (n = 20); G3- Total portal vein occlusi<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> 30<br />

minutes (n =20) G4- Associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> both procedures as described <str<strong>on</strong>g>for</str<strong>on</strong>g> groups 2 and 3. Blood samples from all<br />

groups were obtained 24 hours and 5 days following surgery in order to assess levels <str<strong>on</strong>g>of</str<strong>on</strong>g> mal<strong>on</strong>dialdehyde<br />

(MDA) and protein carb<strong>on</strong>yl groups by TBA and Slot-blotting reacti<strong>on</strong>, respectively Intestinal fragments<br />

stained with Red picrosirius red were also evaluated with polarizati<strong>on</strong>. Data were analyzed statistically by<br />

Anova and Tukey tests.<br />

RESULTS: Levels <str<strong>on</strong>g>of</str<strong>on</strong>g> MDA (Figure 1A,B) and carb<strong>on</strong>yl protein (Figure 2A,B) increased significantly either<br />

after 24 hours (p < 0.002) or 5 days (p < 0,0001), <strong>on</strong>ly in those rats bel<strong>on</strong>ging to group. Five animals <str<strong>on</strong>g>of</str<strong>on</strong>g> G4<br />

presented suture dehiscence. In G2 at day 5 it was observed collagens types III and I (Figure 3A,B). There<br />

were slight inflammatory infiltrate and no extra cellular matrix producti<strong>on</strong> at G3. G4 showed no collagen<br />

producti<strong>on</strong> (Figure 4A,B).<br />

CONCLUSION: Portal reperfusi<strong>on</strong> interferes with col<strong>on</strong>ic anastomosis healing and biological marker<br />

produced may be useful to characterize the oxidative stress.<br />

113


23-26 August 2007,<br />

Budapest, Hungary<br />

114<br />

2F. QUALITY CONTROL AND STRESS-RELATED PROTEIN NETWORKS<br />

(WOLFGANG VOOS)<br />

2F_01_S<br />

THYLAKOID PROTEASES IN HIGHER PLANTS - ROLES IN PROTEIN QUALITY<br />

CONTROL<br />

Zach Adam<br />

The Hebrew University, The Robert H. Smith Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Sciences and Genetics in Agriculture, Rehovot<br />

76100, Israel. e-mail: zach@agri.huji.ac.il<br />

The dependence <str<strong>on</strong>g>of</str<strong>on</strong>g> photosynthesis <strong>on</strong> light is obvious. The higher the light intensity, the higher is the rate <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

photosynthesis, up to a level where light energy is no l<strong>on</strong>ger limiting and photosynthesis remains c<strong>on</strong>stant.<br />

However, further increase in the intensity <str<strong>on</strong>g>of</str<strong>on</strong>g> light may lead to a decrease in photosynthesis rate, a<br />

phenomen<strong>on</strong> known as ‘photoinhibiti<strong>on</strong>’. Photoinhibiti<strong>on</strong> is attributed to oxidative damage, primarily to<br />

photosystem II (PSII) and its reacti<strong>on</strong> center protein D1. A number <str<strong>on</strong>g>of</str<strong>on</strong>g> mechanisms have been evolved<br />

during evoluti<strong>on</strong> to minimize oxidative damage, but if PSII is damaged after all, a PSII repair cycle operates<br />

to allow photosynthesis to proceed. A key comp<strong>on</strong>ent <str<strong>on</strong>g>of</str<strong>on</strong>g> this cycle is the proteolytic removal <str<strong>on</strong>g>of</str<strong>on</strong>g> damaged D1<br />

protein, prior to its replacement by a newly synthesized <strong>on</strong>e. Degradati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the D1 protein has been a<br />

central questi<strong>on</strong> in the field <str<strong>on</strong>g>of</str<strong>on</strong>g> photosynthesis <str<strong>on</strong>g>for</str<strong>on</strong>g> the past 20 years or so, but <strong>on</strong>ly in recent years the identity<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the proteases involved has started to unravel. Recombinant FtsH, a thylakoid ATP-dependent<br />

metalloprotease, was first shown to participate in D1 degradati<strong>on</strong> in an in vitro study. Later <strong>on</strong>, in vivo analysis<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> Arabidopsis FtsH mutants revealed that they were more sensitive to photoinhibiti<strong>on</strong> than wild type, and<br />

that damaged D1 protein was stabilized in them. Further analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> different mutants suggested that the<br />

chloroplast FtsH complex is composed <str<strong>on</strong>g>of</str<strong>on</strong>g> two essential types <str<strong>on</strong>g>of</str<strong>on</strong>g> subunits, each <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> them is encoded by<br />

two redundant genes. More recently, analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> knock-down mutants <str<strong>on</strong>g>of</str<strong>on</strong>g> the lumenal serine protease Deg1<br />

suggested that this protease is also involved in the process <str<strong>on</strong>g>of</str<strong>on</strong>g> D1 degradati<strong>on</strong>. Wider implicati<strong>on</strong>s to<br />

questi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> chloroplast biogenesis and maintenance will be discussed.<br />

2F_02_S<br />

PROTEIN FOLDING, QUALITY CONTROL AND DEGRADATION IN THE ER: THE<br />

ROLE OF N-GLYCANS<br />

Tatiana Soldà, Tito Cali, Carmela Galli, Maurizio Molinari<br />

Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> <str<strong>on</strong>g>Research</str<strong>on</strong>g> in Biomedicine, Bellinz<strong>on</strong>a, Switzerland,<br />

e-mail: maurizio.molinari@irb.unisi.ch<br />

The ER is the site <str<strong>on</strong>g>of</str<strong>on</strong>g> folding and assembly <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins destined <str<strong>on</strong>g>for</str<strong>on</strong>g> the plasma membrane, the secretory and<br />

endocytic organelles and the extracellular space. Most <str<strong>on</strong>g>of</str<strong>on</strong>g> the proteins synthesized in the ER are covalently<br />

modified by co-translati<strong>on</strong>al additi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> pre-assembled glucose 3 -mannose 9 -N-acetylglucosamine 2 - (Glc 3 -Man 9 -<br />

NAcGlc 2 ) core oligosaccharides. Protein-bound oligosaccharides are exposed to several ER-glycanases that<br />

sequentially remove terminal glucose or mannose residues. Rapid generati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a m<strong>on</strong>o-glucosylated (Glc 1 -<br />

Man 9 -GlcNAc 2 ) trimming intermediate is required to enter the calnexin chaper<strong>on</strong>e system in which protein<br />

folding progresses with highest efficiency. Removal and re-additi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the innermost glucose residue activate<br />

cycles <str<strong>on</strong>g>of</str<strong>on</strong>g> dissociati<strong>on</strong>/re-associati<strong>on</strong> with calnexin that may facilitate, and in some cases is required <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

acquisiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the polypeptide’s native structure. Slower removal <str<strong>on</strong>g>of</str<strong>on</strong>g> terminal α1,2-b<strong>on</strong>ded mannose residues


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

from N-linked glycans occurs up<strong>on</strong> persistent polypeptide retenti<strong>on</strong> in the ER, which is symptom <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

defective folding. Substrate de-mannosylati<strong>on</strong> eventually interrupts futile folding attempts, results in substrate<br />

exclusi<strong>on</strong> from the calnexin chaper<strong>on</strong>e system and promotes retro-translocati<strong>on</strong> into the cytosol <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

degradati<strong>on</strong> operated by the 26S proteasome. De-glucosylati<strong>on</strong> and de-mannosylati<strong>on</strong> activities must be<br />

tightly regulated because the N-glycan compositi<strong>on</strong> will determine if the associated protein will be subjected<br />

to folding-attempts in the ER lumen or if it will be retro-translocated into the cytosol and degraded.<br />

2F_03_S<br />

PROTEIN QUALITY CONTROL AND DEGRADATION AT THE ENDOPLASMIC<br />

RETICULUM<br />

Robert Gauss, Ernst Jarosch, Christian Hirsch, Thomas Sommer<br />

Max-Delbrück-Center <str<strong>on</strong>g>for</str<strong>on</strong>g> Molecular Medicine, 13092 Berlin, Germany<br />

A quality c<strong>on</strong>trol system surveys the lumen <str<strong>on</strong>g>of</str<strong>on</strong>g> the endoplasmic reticulum (ER) <str<strong>on</strong>g>for</str<strong>on</strong>g> terminally misfolded<br />

proteins. Polypeptides singled-out by this system are ultimately degraded by the cytosolic ubiquitin<br />

proteasome pathway. This process is termed ER-associated protein degradati<strong>on</strong> (ERAD). A central ERAD<br />

comp<strong>on</strong>ent is the ubiquitin ligase Hrd1/Der3. This ligase <str<strong>on</strong>g>for</str<strong>on</strong>g>ms a complex with its partner protein Hrd3 and<br />

with the ER-membrane protein Der1. Our data imply that Hrd3 is the major substrate receptor <str<strong>on</strong>g>of</str<strong>on</strong>g> this<br />

heterogenic ligase complex in the ER-lumen. Although Hrd3 and Der1 bind to soluble substrate proteins<br />

independently, both proteins are essential to trigger substrate dislocati<strong>on</strong>. At the cytosolic face <str<strong>on</strong>g>of</str<strong>on</strong>g> the ER the<br />

Hrd1-complex associates with the AAA-ATPase Cdc48/p97. Cdc48p binding depends <strong>on</strong> it’s membrane<br />

receptor Ubx2, but most importantly also <strong>on</strong> substrate processing by the Hrd1-complex, suggesting that<br />

ubiquitinati<strong>on</strong> precedes substrate mobilizati<strong>on</strong> by the Cdc48/p97-complex.In additi<strong>on</strong>, we were able to detect<br />

an interacti<strong>on</strong> between the ER quality c<strong>on</strong>trol lectin Yos9p and Hrd3p. We have identified designated regi<strong>on</strong>s<br />

in the luminal domain <str<strong>on</strong>g>of</str<strong>on</strong>g> Hrd3p that interact with Yos9p and Hrd1p. Binding <str<strong>on</strong>g>of</str<strong>on</strong>g> misfolded proteins occurs<br />

via Hrd3p, suggesting that Hrd3p recognises proteins which deviate from their native c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> while<br />

Yos9p ensures that <strong>on</strong>ly terminally misfolded polypeptides are degraded.<br />

2F_04_S<br />

PROTEASE AND CHAPERONE FUNCTIONS IN THE MAINTENANCE OF<br />

MITOCHONDRIAL PROTEIN HOMEOSTASIS<br />

Claudia Leidhold, Tom Bender, Wolfgang Voos<br />

Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Biochemistry and Molecular Biology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Freiburg, Herrmann-Herder-Str. 7, D-79104<br />

Freiburg, Germany<br />

Apart from supplying ATP, mitoch<strong>on</strong>dria are involved in crucial biosynthetic and signaling processes in a<br />

eukaryotic cell. Mitoch<strong>on</strong>drial protein homeostasis is determined by the import <str<strong>on</strong>g>of</str<strong>on</strong>g> newly synthesized proteins<br />

and proteolytic removal <str<strong>on</strong>g>of</str<strong>on</strong>g> excess or damaged polypeptides. Damaged polypeptides, generated under<br />

envir<strong>on</strong>mental stress c<strong>on</strong>diti<strong>on</strong>s, are first recognized by chaper<strong>on</strong>es, stabilized and refolded to the functi<strong>on</strong>al<br />

state. If this fails, the proteins are transferred to the proteolytic system <str<strong>on</strong>g>for</str<strong>on</strong>g> their removal. The coordinated<br />

activities <str<strong>on</strong>g>of</str<strong>on</strong>g> chaper<strong>on</strong>es and proteases <str<strong>on</strong>g>for</str<strong>on</strong>g>m a protein quality c<strong>on</strong>trol system that is required <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

maintenance <str<strong>on</strong>g>of</str<strong>on</strong>g> organellar functi<strong>on</strong>. Mitoch<strong>on</strong>drial proteases bel<strong>on</strong>g to the AAA+ protein family and can be<br />

separated into soluble and membrane-integrated types. We used a proteome analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> isolated mitoch<strong>on</strong>dria<br />

to determine the native substrate selectivity <str<strong>on</strong>g>of</str<strong>on</strong>g> these proteases. We were able to identify a group <str<strong>on</strong>g>of</str<strong>on</strong>g> specific<br />

115


23-26 August 2007,<br />

Budapest, Hungary<br />

substrates <str<strong>on</strong>g>for</str<strong>on</strong>g> the matrix protease Pim1 that were distinguished by an intrinsic low structural stability and the<br />

presence <str<strong>on</strong>g>of</str<strong>on</strong>g> small molecule c<str<strong>on</strong>g>of</str<strong>on</strong>g>actors. Cells lacking mitoch<strong>on</strong>drial proteases showed a higher sensitivity to<br />

high levels <str<strong>on</strong>g>of</str<strong>on</strong>g> reactive oxygen species (ROS). A specific subgroup <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>drial proteins showed<br />

enhanced degradati<strong>on</strong> rates in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> ROS. Enzymes c<strong>on</strong>taining Fe/S cluster exhibited a high<br />

sensitivity to increased ROS levels. Interestingly, proteins that bel<strong>on</strong>ged to the ROS detoxificati<strong>on</strong> system<br />

showed the highest relative degradati<strong>on</strong> rates. We c<strong>on</strong>clude that the protein quality c<strong>on</strong>trol system c<strong>on</strong>tributes<br />

prominently to the maintenance <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>drial protein functi<strong>on</strong>s under stress c<strong>on</strong>diti<strong>on</strong>s.<br />

2F_05_S<br />

(poster secti<strong>on</strong> A2, poster board #103, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

GENETIC DISSECTION OF THE PROTEIN QUALITY CONTROL IN<br />

ESCHERICHIA COLI<br />

Elena García-Fruitós 1 , M<strong>on</strong>ica Martínez-Al<strong>on</strong>so 1 , Nuria G<strong>on</strong>zàlez-M<strong>on</strong>talbán 1 , Minoska Valli 2,3 ,<br />

Diethard Mattanovich 2,3 , Ant<strong>on</strong>io Villaverde 1*<br />

1 Institut de Biotecnologia i de Biomedicina and Departament de Genètica i de Microbiologia, Universitat Autònoma<br />

de Barcel<strong>on</strong>a, Bellaterra, Barcel<strong>on</strong>a, Spain<br />

2 University <str<strong>on</strong>g>of</str<strong>on</strong>g> Natural Resources and Applied Life Sciences Vienna, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biotechnology, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Applied Microbiology, Vienna, Austria<br />

3 School <str<strong>on</strong>g>of</str<strong>on</strong>g> Bioengineering, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Applied Sciences FH-Campus Vienna, Austria<br />

The protein quality c<strong>on</strong>trol system is an evoluti<strong>on</strong>ary c<strong>on</strong>served complex mechanism based <strong>on</strong> a network <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

cellular proteins with overlapping foldase, disaggregase and protease activities. In bacterial cells, the<br />

coordinated activity <str<strong>on</strong>g>of</str<strong>on</strong>g> all these elements promote proper protein folding or digesti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> folding-reluctant,<br />

potentially toxic species, what is expected to keep misfolding-pr<strong>on</strong>e protein species in a soluble state and cells<br />

free from aggregates. However, recent insights <strong>on</strong> the biology <str<strong>on</strong>g>of</str<strong>on</strong>g> protein misfolding and aggregati<strong>on</strong> str<strong>on</strong>gly<br />

suggest that solubility and c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>al quality are not matching events, since at least in some examples,<br />

protein aggregates might c<strong>on</strong>tain properly folded species. By using Escherichia coli cell models, we have finely<br />

explored both protein solubility and quality, through the fluorescence emissi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> green fluorescent protein<br />

(GFP) engineered variants, in mutant cell strains lacking defined comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g> the quality c<strong>on</strong>trol system,<br />

including main chaper<strong>on</strong>es, small heat-shock proteins and proteases. In absence <str<strong>on</strong>g>of</str<strong>on</strong>g> either chaper<strong>on</strong>es DnaK,<br />

ClpB or ClpA and proteases ClpP or L<strong>on</strong>, GFP-producing cells are significantly more fluorescent than wild<br />

type cells (up to more than two fold), while the solubility <str<strong>on</strong>g>of</str<strong>on</strong>g> GFP is clearly higher in wild type cells exhibiting<br />

a fully functi<strong>on</strong>al quality c<strong>on</strong>trol network (up to around two fold). In all the identified mutants, the enhanced<br />

emissi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> recombinant cells is clearly linked to an elevated intracellular c<strong>on</strong>tent <str<strong>on</strong>g>of</str<strong>on</strong>g> highly fluorescent GFP<br />

molecules, resulting from inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> its proteolytic degradati<strong>on</strong> and the significant expansi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> its half life<br />

(from 2 to up to 7 h, at 37ºC). Interestingly, the excess <str<strong>on</strong>g>of</str<strong>on</strong>g> functi<strong>on</strong>al protein is overstocked as highly<br />

fluorescent inclusi<strong>on</strong> bodies c<strong>on</strong>taining properly folded protein species. Overall, these results indicate that the<br />

E. coli quality c<strong>on</strong>trol system is governed by an over-committed, chaper<strong>on</strong>e-mediated proteolytic machinery<br />

that acts <strong>on</strong> protein species that are either functi<strong>on</strong>al or can reach functi<strong>on</strong>al <str<strong>on</strong>g>for</str<strong>on</strong>g>ms when proteolytically<br />

stabilized. Intriguingly, the occurrence <str<strong>on</strong>g>of</str<strong>on</strong>g> molecular determinants <str<strong>on</strong>g>of</str<strong>on</strong>g> aggregati<strong>on</strong> does not require complete<br />

protein unfolding, and in fact, solubility and fluorescence emissi<strong>on</strong> are inversely correlated. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e,<br />

selected genetic deficiencies in the quality c<strong>on</strong>trol system dramatically enhance the intracellular pool <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

biologically active although insoluble, misfolding-pr<strong>on</strong>e proteins, a fact that might be specially relevant in the<br />

c<strong>on</strong>text <str<strong>on</strong>g>of</str<strong>on</strong>g> high quality recombinant protein producti<strong>on</strong>.<br />

116


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2F_06_S<br />

(poster secti<strong>on</strong> A2, poster board #104, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

LECTIN-DEFICIENT CALRETICULIN RETAINS FULL FUNCTIONALITY AS A<br />

CHAPERONE AND QUALITY CONTROL COMPONENT DURING THE<br />

BIOGENESIS OF CLASS I HISTOCOMPATIBILITY MOLECULES<br />

Breanna S. Ireland, Ulf Brockmeier, David B. Williams<br />

Depts. <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry and Immunology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Tor<strong>on</strong>to, Tor<strong>on</strong>to, Canada, M5S 1A8,<br />

e-mail: david.williams@utor<strong>on</strong>to.ca<br />

Calreticulin (Crt) is a soluble chaper<strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the endoplasmic reticulum (ER) that interacts with newly<br />

synthesized glycoproteins through a lectin site with specificity <str<strong>on</strong>g>for</str<strong>on</strong>g> Glc 1 Man 9 GlcNAc 2 oligosaccharides as well<br />

as through a polypeptide binding site that recognizes n<strong>on</strong>-native protein c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mers. The relative<br />

c<strong>on</strong>tributi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> each site to the overall functi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> Crt remains unknown. To address this issue, we created<br />

two point mutants, D317A and Y128A, that ablate the lectin functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Crt but do not alter its tertiary<br />

structure. We then examined their abilities to support the biogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> mouse class I histocompatibility<br />

molecules. Class I molecules functi<strong>on</strong> to present peptide antigens to cytotoxic T cells. They c<strong>on</strong>sist <str<strong>on</strong>g>of</str<strong>on</strong>g> a<br />

glycosylated transmembrane heavy chain, a soluble subunit termed β 2 -microglobulin and an 8-9 residue<br />

peptide ligand. In Crt-deficient cells, the surface expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> class I molecules is reduced 2- to 3-fold,<br />

loading <str<strong>on</strong>g>of</str<strong>on</strong>g> peptide ligands is inefficient, and peptide-deficient class I molecules are prematurely exported from<br />

the ER (defective quality c<strong>on</strong>trol). We expressed wild type Crt as well as both lectin-deficient Crt mutants in<br />

Crt-deficient cells. Remarkably, the lectin-deficient mutants were just as effective as wild type Crt in<br />

upregulating class I surface expressi<strong>on</strong>, enhancing peptide loading and preventing premature export from the<br />

ER. The mutants were also capable <str<strong>on</strong>g>of</str<strong>on</strong>g> binding to many newly synthesized glycoproteins in additi<strong>on</strong> to class I.<br />

We c<strong>on</strong>clude that in the absence <str<strong>on</strong>g>of</str<strong>on</strong>g> lectin-based interacti<strong>on</strong>s, Crt can utilize polypeptide-based interacti<strong>on</strong>s to<br />

effect its chaper<strong>on</strong>e and quality c<strong>on</strong>trol functi<strong>on</strong>s.<br />

117


23-26 August 2007,<br />

Budapest, Hungary<br />

118<br />

2F_01_P<br />

(poster secti<strong>on</strong> A2, poster board #105, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE STRESS PROTEOME: 2D-PAGE OF YEAST MITOCHONDRIA UNDER<br />

DIFFERENT OXIDATIVE STRESS CONDITIONS<br />

Claudia Leidhold, Tom Bender, Wolfgang Voos<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry and Molecular Biology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Freiburg, Germany<br />

e-mail: Wolfgang.Voos@biochemie.uni-freiburg.de<br />

Besides fulfilling further important metabolic functi<strong>on</strong>s, mitoch<strong>on</strong>dria are also the place <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular<br />

respirati<strong>on</strong>. During this process, as a side reacti<strong>on</strong>, a c<strong>on</strong>tinuous producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> reactive oxygen species (ROS)<br />

occurs. Because <str<strong>on</strong>g>of</str<strong>on</strong>g> their proximity to the place <str<strong>on</strong>g>of</str<strong>on</strong>g> ROS <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>, mitoch<strong>on</strong>drial proteins are at high risk <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

being irreversibly damaged and misfolded. This implies an important functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proteolysis in mitoch<strong>on</strong>drial<br />

stress defense. Indeed, yeast cells lacking Pim1, the major protease <str<strong>on</strong>g>of</str<strong>on</strong>g> the mitoch<strong>on</strong>drial matrix, show a<br />

significantly higher sensitivity to ROS than wild-type cells. In order to get a general overview about<br />

mitoch<strong>on</strong>drial proteins affected by oxidative stress we per<str<strong>on</strong>g>for</str<strong>on</strong>g>med an extensive proteome analysis. We<br />

compared the yeast mitoch<strong>on</strong>drial 2D-pattern after treatment with stress-inducing chemicals, such as H 2 O 2 ,<br />

antimycin A or menadi<strong>on</strong>e, to that <str<strong>on</strong>g>of</str<strong>on</strong>g> untreated mitoch<strong>on</strong>dria. Under each stress c<strong>on</strong>diti<strong>on</strong> we identified a<br />

number <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins as candidate substrates <str<strong>on</strong>g>for</str<strong>on</strong>g> proteolysis, with some <str<strong>on</strong>g>of</str<strong>on</strong>g> them even being sensitive to all<br />

c<strong>on</strong>diti<strong>on</strong>s tested. Proteins affected the most by ROS are either themselves involved in oxidative stress<br />

defense or c<strong>on</strong>tain oxidant-sensitive structures such as Fe-S-clusters. In vitro degradati<strong>on</strong> assays with selected<br />

candidate proteins could c<strong>on</strong>firm the results obtained from 2D studies. For most <str<strong>on</strong>g>of</str<strong>on</strong>g> the proteins degraded in<br />

a stress-specific manner we could identify Pim1 as the protease resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> their turnover.<br />

2F_02_P<br />

(poster secti<strong>on</strong> A2, poster board #106, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CHAPERONE-DEPENDENT AND INDEPENDENT ROLE OF N-GLYCAN IN CFTR<br />

FOLDING, STABILITY AND MEMBRANE TRAFFICKING<br />

R. Glozman, T. Okiy<strong>on</strong>eda, H. Barriere, G. L. Lukacs<br />

Hospital <str<strong>on</strong>g>for</str<strong>on</strong>g> Sick Children, Cell Biology, Tor<strong>on</strong>to, Ontario, Canada<br />

e-mail: glukacs@sickkids.ca<br />

Although N-glycan-mediated recruitment <str<strong>on</strong>g>of</str<strong>on</strong>g> lectin chaper<strong>on</strong>es (e.g. calnexin/calreticulin) can assist the<br />

folding and suppress the aggregati<strong>on</strong> tendency <str<strong>on</strong>g>of</str<strong>on</strong>g> newly synthesized glycoproteins at the ER, the glycandependent<br />

stabilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> plasma membrane protein remains elusive. Here we used the cystic fibrosis<br />

transmembrane c<strong>on</strong>ductance regulator (CFTR), a PKA-activated chloride channel, to investigate the role <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

N-glycan in its biogenesis and trafficking. CFTR glycosylati<strong>on</strong> was prevented by genetic and pharmacological<br />

means, while chaper<strong>on</strong>e-CFTR interacti<strong>on</strong> was disrupted in calnexin-depleted cells. Calnexin downregulati<strong>on</strong><br />

by siRNA or glucosidase I inhibiti<strong>on</strong> decreased CFTR folding efficiency by ~30% in the ER. In c<strong>on</strong>trast,<br />

n<strong>on</strong>-glycosylated CFTR folding was attenuated by >80%, suggesting a chaper<strong>on</strong>e-independent role <str<strong>on</strong>g>of</str<strong>on</strong>g> N-<br />

glycans in the channel c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>al maturati<strong>on</strong>. The structural destabilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cell-surface resident, n<strong>on</strong>glycosylated<br />

CFTR was documented by limited proteolysis and c<strong>on</strong>ceivable c<strong>on</strong>stitutes the trigger <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

ubiquitinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the destabilized channel. Ubiquitin c<strong>on</strong>jugati<strong>on</strong> signals to reroute the channel from the<br />

c<strong>on</strong>stitutive recycling pathway towards lysosomal degradati<strong>on</strong> by the ubiquitin-dependent endosomal sorting<br />

machinery. This mechanism appears to account <str<strong>on</strong>g>for</str<strong>on</strong>g> the >4-fold accelerated turnover <str<strong>on</strong>g>of</str<strong>on</strong>g> the mutant CFTR in


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

post-Golgi compartments. Our results, jointly, indicate that N-glycans are critical determinant <str<strong>on</strong>g>of</str<strong>on</strong>g> CFTR<br />

c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>al maturati<strong>on</strong>/stability in a chaper<strong>on</strong>e-independent manner and outline a possible paradigm <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

the rapid degradati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> glycosylati<strong>on</strong>-deficient membrane proteins from the cell surface.<br />

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23-26 August 2007,<br />

Budapest, Hungary<br />

120<br />

2G. STRESS OF BACTERIA<br />

(MARÍA M. TAVÍO)<br />

2G_01_S<br />

VARIATION IN STRESS RESPONSES WITHIN A BACTERIAL SPECIES AND THE<br />

INDIRECT COSTS OF STRESS RESISTANCE<br />

Thomas Ferenci a , Beny Spira<br />

a School <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular and Microbial Biosciences, The University <str<strong>on</strong>g>of</str<strong>on</strong>g> Sydney, NSW 2006 Australia<br />

ABSTRACT: Bacteria can exhibit high levels <str<strong>on</strong>g>of</str<strong>on</strong>g> resistance to <strong>on</strong>e or more envir<strong>on</strong>mental stresses such as<br />

temperature, osmolarity, radiati<strong>on</strong>, pH, starvati<strong>on</strong> as well as resistance to noxious chemicals and antibiotics.<br />

Yet evoluti<strong>on</strong> has not optimized stress resistance in all bacteria to all stresses. Even within a species like<br />

Escherichia coli, stress resistance is not c<strong>on</strong>stant between strains, suggesting that selecti<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> stress resistance is<br />

under counter-selecti<strong>on</strong> in some envir<strong>on</strong>ments. The trade-<str<strong>on</strong>g>of</str<strong>on</strong>g>fs associated with stress resistance in E. coli are<br />

due to more than the direct cost <str<strong>on</strong>g>of</str<strong>on</strong>g> resistance mechanisms. A significant indirect cost is that high stress<br />

resistance is associated with a reduced ability to compete <str<strong>on</strong>g>for</str<strong>on</strong>g> poor growth substrates like acetate or even good<br />

substrates like glucose at sub-optimal c<strong>on</strong>centrati<strong>on</strong>s. High stress resistance also decreases the ability to use<br />

inorganic nutrients like phosphate. This trade-<str<strong>on</strong>g>of</str<strong>on</strong>g>f between self preservati<strong>on</strong> and nutriti<strong>on</strong>al competence, called<br />

the SPANC balance, is likely to be major selective influence in natural populati<strong>on</strong>s. Another cost <str<strong>on</strong>g>of</str<strong>on</strong>g> high<br />

stress resistance in E. coli is an elevated mutati<strong>on</strong> rate and the increased generati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> deleterious mutati<strong>on</strong>s.<br />

Directi<strong>on</strong>al adaptati<strong>on</strong>s in SPANC balance and mutati<strong>on</strong> rate are envir<strong>on</strong>ment-dependent. The most<br />

comm<strong>on</strong> variati<strong>on</strong>s in SPANC are due to polymorphisms in the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> global regulators RpoS and ppGpp<br />

between different strains. High levels favour stress resistance, lower levels allow better nutriti<strong>on</strong>. The intimate<br />

associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> RpoS/ppGpp with stress resistance and SPANC balancing influences numerous cellular<br />

processes and bacterial properties, including virulence.<br />

2G_02_S<br />

MECHANISMS OF TRANSLATION REGULATION DURING COLD-SHOCK IN<br />

ESCHERICHIA COLI<br />

Claudio O. Gualerzi, Anna Maria Giuliodori<br />

Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Genetics, Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology MCA, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Camerino, 62032 Camerino (MC), Italy<br />

e-mail: claudio.gualerzi@unicam.it<br />

Cold-shock (cs) translati<strong>on</strong>al bias, namely the c<strong>on</strong>diti<strong>on</strong> which favors translati<strong>on</strong> at low temperature <str<strong>on</strong>g>of</str<strong>on</strong>g> cs<br />

mRNAs, is <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the main mechanisms by which Escherichia coli cells ensure the selective expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> its cs<br />

genes after cold stress. The bias is partly due to intrinsic features <str<strong>on</strong>g>of</str<strong>on</strong>g> cs mRNAs, which make them pr<strong>on</strong>e to<br />

translati<strong>on</strong> at low temperature, and to a cold stress-induced transient increase <str<strong>on</strong>g>of</str<strong>on</strong>g> the Initiati<strong>on</strong> Factors<br />

(IFs)/ribosome ratio. In this study we have undertaken the task <str<strong>on</strong>g>of</str<strong>on</strong>g>: i) identifying the mechanism generating<br />

the stoichiometric imbalance <str<strong>on</strong>g>of</str<strong>on</strong>g> the IFs/ ribosome ratio; ii) unraveling the role <str<strong>on</strong>g>of</str<strong>on</strong>g> the IFs in the translati<strong>on</strong><br />

bias; iii) elucidating the sec<strong>on</strong>dary structure <str<strong>on</strong>g>of</str<strong>on</strong>g> the paradigm cs mRNA, namely the E. coli cspA mRNA and<br />

iv) detecting possible temperature-dependent variati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> its structure.<br />

The results obtained indicate that: i) transcripti<strong>on</strong> and translati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> infA and infC which encode IF1 and IF3,<br />

respectively, are activated de novo by cs while ribosomal subunits assembly is slowed down; ii) at low<br />

temperature IF3 stimulates the rate <str<strong>on</strong>g>of</str<strong>on</strong>g> “30S initiati<strong>on</strong> complex” <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> with cs mRNAs while inducing


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

the <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> n<strong>on</strong>-productive 70S initiati<strong>on</strong> complexes with n<strong>on</strong>-cs mRNAs; iii) the increased level <str<strong>on</strong>g>of</str<strong>on</strong>g> IF1<br />

and IF3 during cs is essential to provide a sufficient pool <str<strong>on</strong>g>of</str<strong>on</strong>g> dissociated 30S ribosomal subunits capable <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

“70S initiati<strong>on</strong> complex” <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> and iv) the structure <str<strong>on</strong>g>of</str<strong>on</strong>g> cspA mRNA, as determined by chemical and<br />

enzymatic probing, changes up<strong>on</strong> temperature down-shift exposing the translati<strong>on</strong> initiati<strong>on</strong> regi<strong>on</strong>.<br />

2G_03_S<br />

IMPLICATION OF STRESS IN THE LOSS OF VIRULENCE FACTORS IN<br />

UROPATHOGENIC ESCHERICHIA COLI<br />

S. M. Soto, J. Vila<br />

Microbiology Department, Hospital Clinic <str<strong>on</strong>g>of</str<strong>on</strong>g> Barcel<strong>on</strong>a. Villarroel 170. 08036. Barcel<strong>on</strong>a, Spain<br />

e-mail: sarasotog@yahoo.es<br />

Escherichia coli is by far the most comm<strong>on</strong> cause <str<strong>on</strong>g>of</str<strong>on</strong>g> urinary tract infecti<strong>on</strong>s (UTI). Uropathogenic E. coli<br />

(UPEC) strains possess several virulence determinants that allow them to col<strong>on</strong>ize the urinary tract, avoid<br />

host defenses, and cause damage to the uroepithelium, which may, in some cases, lead to passage <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

bacterium into the blood-stream. Several genes encoding urovirulence factors, such as hemolysin, cytotoxic<br />

necrotizing factor type 1 (cnf1), P-pili F13 (pap), S-family adhesins, ir<strong>on</strong> systems, some capsule factors and the<br />

autotransporter toxin sat1 are located in the chromosome and/or plasmids <str<strong>on</strong>g>for</str<strong>on</strong>g>ming clusters named<br />

pathogenicity islands (PAIs). Several studies have dem<strong>on</strong>strated that quinol<strong>on</strong>e resistant E. coli strains have<br />

fewer virulent factors than quinol<strong>on</strong>e susceptible strains. Thus, the aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this work was to study the possible<br />

relati<strong>on</strong>ship between bacterial stress produced by quinol<strong>on</strong>es and the loss <str<strong>on</strong>g>of</str<strong>on</strong>g> virulence factors located in PAIs<br />

in UPEC. Three UPEC quinol<strong>on</strong>e-susceptible and hemolytic clinical strains were submitted to subinhibitory<br />

c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> cipr<str<strong>on</strong>g>of</str<strong>on</strong>g>loxacin. A sample <str<strong>on</strong>g>of</str<strong>on</strong>g> the well showing growth at the highest quinol<strong>on</strong>e c<strong>on</strong>centrati<strong>on</strong><br />

was spread <strong>on</strong>to large blood Columbia agar plates. The n<strong>on</strong>hemolytic col<strong>on</strong>ies were analyzed to determine<br />

the loss <str<strong>on</strong>g>of</str<strong>on</strong>g> the hemolysin gene (hly) and other factors related to PAIs. The three strains lost hemolytic<br />

capacity between passages 1-4 in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> cipr<str<strong>on</strong>g>of</str<strong>on</strong>g>loxacin. The loss rate was between 1x10 -4 and 5x10 -3 .<br />

No col<strong>on</strong>ies without hemolytic capacity were found after 15 passages <str<strong>on</strong>g>of</str<strong>on</strong>g> wild-type strains using antimicrobialfree<br />

culture medium. In c<strong>on</strong>clusi<strong>on</strong>, these findings suggest that quinol<strong>on</strong>es produce bacterial stress the<br />

resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> which is a loss <str<strong>on</strong>g>of</str<strong>on</strong>g> virulence factors.<br />

2G_04_S<br />

OSMOTIC STRESS AND OTHER STRESSORS AS INDUCERS OF MULTIDRUG<br />

RESISTANCE<br />

María M. Tavío<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Las Palmas de Gran Canaria. Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Health Sciences. Las Palmas G.C.- Spain,<br />

e-mail: mtavio@dcc.ulpgc.es<br />

In order to survive under and adapt to different c<strong>on</strong>diti<strong>on</strong>s bacteria have systems that are able to sense and<br />

resp<strong>on</strong>d to envir<strong>on</strong>mental stimuli. A complex network <str<strong>on</strong>g>of</str<strong>on</strong>g> regulatory systems ensures a coordinated and<br />

effective answer to different stresses that can act <strong>on</strong> a bacterium simultaneously. Hyperosmolarity and some<br />

chemicals as fluoroquinol<strong>on</strong>es, salicylate, n<strong>on</strong>-antimicrobial medicaments as diazepam, anti-inflammatory<br />

drugs, am<strong>on</strong>g others, can induce an increased active efflux and organic solvent tolerance, loss <str<strong>on</strong>g>of</str<strong>on</strong>g> porins and<br />

multidrug resistance, both in wild type strains and clinical isolates <str<strong>on</strong>g>of</str<strong>on</strong>g> enterobacteria. Besides the role <str<strong>on</strong>g>of</str<strong>on</strong>g> efflux<br />

systems in multidrug resistance phenotypes, they seem to have a natural functi<strong>on</strong> exporting signals <str<strong>on</strong>g>for</str<strong>on</strong>g> cell–<br />

121


23-26 August 2007,<br />

Budapest, Hungary<br />

cell communicati<strong>on</strong>. AcrAB-TolC is an efflux system that exudes fluoroquinol<strong>on</strong>es and is up-regulated by<br />

SdiA, a quorum-sensing transcripti<strong>on</strong>al regulator. Another transcripti<strong>on</strong>al regulators that are also involved in<br />

bacterial stress resp<strong>on</strong>se such as marA or soxS, also activate AcrAB-TolC. Sigma factors, and the twocomp<strong>on</strong>ent<br />

systems CpxAR and BaeSR are also key pieces in the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> gene expressi<strong>on</strong> in resp<strong>on</strong>se to<br />

stress c<strong>on</strong>diti<strong>on</strong>s. Thus, the resp<strong>on</strong>se regulator CpxR can lead to an increase <str<strong>on</strong>g>of</str<strong>on</strong>g> the mRNA level <str<strong>on</strong>g>of</str<strong>on</strong>g> several<br />

drug exporter genes and the mutati<strong>on</strong> level as well as to a diminished OmpF assembly. From this point <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

view the development <str<strong>on</strong>g>of</str<strong>on</strong>g> intrinsic multidrug resistance might be understood as part <str<strong>on</strong>g>of</str<strong>on</strong>g> the bacterial resp<strong>on</strong>se<br />

to stress. The in vitro inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> multidrug resistance has been associated with high levels <str<strong>on</strong>g>of</str<strong>on</strong>g> inducers, those<br />

that are close to their minimal inhibitory c<strong>on</strong>centrati<strong>on</strong>s. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, bacterial resp<strong>on</strong>se to osmotic stress<br />

might be linked to multidrug resistance phenotypes.<br />

2G_05_S<br />

(poster secti<strong>on</strong> B1, poster board #236, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

FUNCTION OF DNAK IN STREPTOCOCCUS INTERMEDIUS<br />

Toshifumi Tomoyasu, Atsushi Tabata, Hideaki Nagamune<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Technology and Science, The University <str<strong>on</strong>g>of</str<strong>on</strong>g> Tokushima Graduate School. Japan<br />

e-mail: tomoyasu@bio.tokushima-u.ac.jp<br />

Streptococcus intermedius is an anaerobe and bel<strong>on</strong>gs to the anginosus group <str<strong>on</strong>g>of</str<strong>on</strong>g> streptococci (AGS), which<br />

c<strong>on</strong>stitute a part <str<strong>on</strong>g>of</str<strong>on</strong>g> the normal flora <str<strong>on</strong>g>of</str<strong>on</strong>g> the human oral cavity as well as the upper respiratory,<br />

gastrointesti<strong>on</strong>al, and female urogenital tracts. AGS are recognized as opportunistic pathogens that cause<br />

purulent infecti<strong>on</strong>s and abscess <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>. We have previously reported that DnaK chaper<strong>on</strong>e c<strong>on</strong>trols the<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> flagella and several pathogenic factors in Salm<strong>on</strong>ella Typhimurium. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, we c<strong>on</strong>structed a<br />

dnaK null mutant from S. intermedius in order to investigate the possible role <str<strong>on</strong>g>of</str<strong>on</strong>g> DnaK in pathogenicity. The<br />

generati<strong>on</strong> time <str<strong>on</strong>g>of</str<strong>on</strong>g> a dnaK null mutant from S. intermedius was approximately twice that <str<strong>on</strong>g>of</str<strong>on</strong>g> the parent strain.<br />

Similar to other gram-negative bacteria, the dnaK null mutant exhibited a thermosensitive phenotype and<br />

could not grow above 40°C. However, the dnaK null mutant did not show acid and H 2 O 2 sensitivity, which is<br />

characteristic <str<strong>on</strong>g>of</str<strong>on</strong>g> gram-negative bacteria. Interestingly, GroEL accumulati<strong>on</strong> was observed in the dnaK null<br />

mutant. The genome sequences from AGS revealed that the heat shock resp<strong>on</strong>se, including expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

groESL oper<strong>on</strong>, appears to be c<strong>on</strong>trolled by the HrcA heat shock gene transcripti<strong>on</strong>al repressor. Our result<br />

suggests that DnaK might regulate the activity or cellular amount <str<strong>on</strong>g>of</str<strong>on</strong>g> HrcA. Neither the dnaK mutant nor the<br />

parent strain showed a significant difference with regard to the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> cytolysin (Intermedilysin) and<br />

hyalur<strong>on</strong>idase, and in the ability to <str<strong>on</strong>g>for</str<strong>on</strong>g>m bi<str<strong>on</strong>g>of</str<strong>on</strong>g>ilms. These data indicate that DnaK in S. intermedius plays a role<br />

in the fundamental functi<strong>on</strong>s <str<strong>on</strong>g>for</str<strong>on</strong>g> living (e.g., growth, thermoresistance, and heat shock regulati<strong>on</strong>) but has less<br />

functi<strong>on</strong>ality in the modulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> pathogenic factors.<br />

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3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2G_06_S<br />

(poster secti<strong>on</strong> B1, poster board #237, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE ROLE OF HEAT SHOCK PROTEINS IBPA/B IN PROTECTION OF E.COLI<br />

PROTEINS AGAINST THERMAL DENATURATION UNDER ANAEROBIC<br />

GROWTH CONDITIONS<br />

Ewelina Matuszewska, Dorota Kuczynska-Wisnik, Joanna Kwiatkowska, Elzbieta Ratajczak,<br />

Ewa Laskowska<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Gdansk, Kladki 24, Gdansk, Poland<br />

e-mail: ewelinam@biotech.ug.gda.pl<br />

The E. coli IbpA/B proteins bel<strong>on</strong>g to a group <str<strong>on</strong>g>of</str<strong>on</strong>g> molecular chaper<strong>on</strong>es which under heat shock protect<br />

polypeptides from irreversible denaturati<strong>on</strong> and facilitate their refolding to the native structure in cooperati<strong>on</strong><br />

with the ATP-dependent chaper<strong>on</strong>es: DnaK/DnaJ/GrpE and ClpB. The role <str<strong>on</strong>g>of</str<strong>on</strong>g> this chaper<strong>on</strong>e machinery<br />

has been thoroughly investigated, however it is very little known about their functi<strong>on</strong> under anaerobic<br />

c<strong>on</strong>diti<strong>on</strong>s, despite the fact that the gastrointestinal tracts are a natural anoxic envir<strong>on</strong>ment <str<strong>on</strong>g>for</str<strong>on</strong>g> E. coli. We<br />

have focused our interest <strong>on</strong> the role <str<strong>on</strong>g>of</str<strong>on</strong>g> IbpA/B in protecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> alcohol dehydrogenase (AdhE). The AdhE<br />

is a Fe 2+ dependent protein with three enzymatic activities: alcohol dehydrogenase, acetaaldehyde –CoA<br />

dehydrogenase and pyruvate <str<strong>on</strong>g>for</str<strong>on</strong>g>mate lyase deactivase. AdhE c<strong>on</strong>verts acetyl-coenzym A to acetaldehyde and<br />

then to ethanol in reacti<strong>on</strong>s coupled to oxidati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> two NADH molecules. The adhE gene is very highly<br />

expressed under anaerobic c<strong>on</strong>diti<strong>on</strong>s. We dem<strong>on</strong>strated that IbpA/B in cooperati<strong>on</strong> with DnaK system<br />

prevented heat - inactivati<strong>on</strong> and aggregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> AdhE under anaerobic c<strong>on</strong>diti<strong>on</strong>s, both in vitro and in vivo.<br />

ClpB inhibited aggregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular proteins in anaerobically growing E. coli submitted to heat stress, but<br />

was not required <str<strong>on</strong>g>for</str<strong>on</strong>g> the AdhE protecti<strong>on</strong>.<br />

123


23-26 August 2007,<br />

Budapest, Hungary<br />

2G_01_P<br />

(poster secti<strong>on</strong> B1, poster board #238, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE THIOREDOXIN SYSTEM IN THE PSYCHROPHILIC EUBACTERIUM<br />

PSEUDOALTEROMONAS HALOPLANKTIS<br />

R. Cotugno 1 , G. Salom<strong>on</strong>e 1 , M. R. Ruocco 1 , P. Grimaldi 1 , F. Cecere 1 , P. Falasca 2 , G. Raimo 2 ,<br />

M. Masullo 3 , E. De Vendittis 1<br />

1Dept. Biochemistry & Medical Biotechnologies, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Naples Federico II,<br />

2Dept. Science & Technology <str<strong>on</strong>g>of</str<strong>on</strong>g> the Envir<strong>on</strong>ment & Territory, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Molise,<br />

3Dept. Pharmacobiological Sciences, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Catanzaro Magna Graecia, Italy<br />

e-mail: cotugnoroberta@libero.it<br />

The thioredoxin system, involved in the preservati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the reduced state <str<strong>on</strong>g>of</str<strong>on</strong>g> cytoplasmic proteins, includes<br />

two ubiquitous key comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g> the intracellular redox balance: thioredoxin (Trx) and thioredoxin<br />

reductase (TrxR). Either Trx or TrxR c<strong>on</strong>tain a c<strong>on</strong>served CXXC sequence, switching between disulfide and<br />

free dithiol. Trx is a small m<strong>on</strong>omeric protein; TrxR is a NADPH-dependent homodimeric flavoenzyme. The<br />

redox cycle in the Trx/TrxR system also involves oscillati<strong>on</strong> between oxidized and reduced <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> FAD,<br />

using NADPH as electr<strong>on</strong> d<strong>on</strong>or.<br />

This report describes the biochemical characterizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the Trx/TrxR system in Pseudoalterom<strong>on</strong>as haloplanktis<br />

(Ph), a psychrophilic eubacterium isolated from marine Antarctic sediments. PhTrxR and PhTrx were obtained<br />

as recombinant His-tagged proteins, or isolated from P. haloplanktis cells. The activity <str<strong>on</strong>g>of</str<strong>on</strong>g> recombinant PhTrxR<br />

was evaluated by both DTNB- and thioredoxin-reducti<strong>on</strong> methods and compared with that <str<strong>on</strong>g>of</str<strong>on</strong>g> endogenous<br />

PhTrxR. After exogenous FAD capti<strong>on</strong>, recombinant PhTrxR shows the same activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the endogenous<br />

enzyme. The thermal denaturati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the recombinant enzyme was followed by fluorescence melting curves<br />

in the temperature range 5-75°C. Heat inactivati<strong>on</strong> experiments gave a half-life <str<strong>on</strong>g>of</str<strong>on</strong>g> 10 min at 60°C. However,<br />

when studying the thermophilicity with the DTNB-reducti<strong>on</strong> method, maximum activity was reached at<br />

30°C. Like the endogenous counterpart, recombinant PhTrx reduces the insulin disulfides in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

either DTT or PhTrxR/NADPH.<br />

2G_03_P<br />

(poster secti<strong>on</strong> B1, poster board #239, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ESCHERICHIA COLI HEAT SHOCK PROTEINS IBPA/B ARE INVOLVED IN<br />

RESISTANCE TO OXIDATIVE STRESS INDUCED BY COPPER IONS<br />

Ewelina Matuszewska, Joanna Kwiatkowska, Dorota Kuczyńska-Wiśnik, Ewa Laskowska<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Gdańsk, Kładki 24, Gdańsk, Poland<br />

e-mail: lasko@biotech.ug.gda.pl<br />

E. coli IbpA/B proteins in cooperati<strong>on</strong> with other molecular chaper<strong>on</strong>es systems prevent the aggregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

thermally denatured proteins and support their refolding to the native state. Several data suggest that IbpA/B<br />

participate in the protecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> E. coli cells against oxidative stress: IbpA/B inhibits inactivati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> some<br />

E. coli enzymes by superoxide radicals in vitro. Furthermore, overproduced IbpA/B increase E. coli resistance<br />

to superoxide stress. We dem<strong>on</strong>strate that the IbpA/B participate in the protecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> E. coli against oxidative<br />

stress induced by copper i<strong>on</strong>s. The transiti<strong>on</strong> metal copper is essential to a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular functi<strong>on</strong>s,<br />

however even moderately increased level <str<strong>on</strong>g>of</str<strong>on</strong>g> copper may be toxic <str<strong>on</strong>g>for</str<strong>on</strong>g> the cell. The toxicity results from the<br />

124


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Fent<strong>on</strong> or Haber-Weiss reacti<strong>on</strong> in which copper i<strong>on</strong>s catalyze the producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> OH radicals from<br />

hydroperoxide. We show that the lack <str<strong>on</strong>g>of</str<strong>on</strong>g> IbpA/B causes increased E. coli sensitivity to copper i<strong>on</strong>s. IbpA/B<br />

proteins inhibited copper-catalyzed oxidati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a model enzyme – alcohol dehydrogenase (AdhE) both in<br />

vivo and in vitro. We suggest that the overall ability <str<strong>on</strong>g>of</str<strong>on</strong>g> IbpA/B to protect cells from copper -induced damage<br />

may result from the metal chelati<strong>on</strong> and direct binding to the protected proteins. Similar activity has been<br />

proposed <str<strong>on</strong>g>for</str<strong>on</strong>g> alfa-crystallin, a mammalian molecular chaper<strong>on</strong>e homologous to the E. coli IbpA/B.<br />

2G_04_P<br />

(poster secti<strong>on</strong> B1, poster board #240, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STARVATION INDUCED PROTEIN FROM MYCOBACTERIUM SMEGMATIS<br />

Saraswathi Ramachandran, Dipankar Chatterji<br />

MBU, IISc, Bangalore 560012, India, e-mail: rs@mbu.iisc.ernet.in<br />

Stati<strong>on</strong>ary phase is characterized by scarcity <str<strong>on</strong>g>of</str<strong>on</strong>g> nutrients, which is essentially the c<strong>on</strong>diti<strong>on</strong> in which wild-type<br />

bacteria are c<strong>on</strong>sidered to be found in nature.<br />

A remarkable feature <str<strong>on</strong>g>of</str<strong>on</strong>g> bacteria is their ability to remain viable during prol<strong>on</strong>ged periods <str<strong>on</strong>g>of</str<strong>on</strong>g> starvati<strong>on</strong> and<br />

their capacity <str<strong>on</strong>g>for</str<strong>on</strong>g> rapid resumpti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> growth when nutrients become available again.<br />

Bacteria have evolved sophisticated mechanisms towards this end, some <str<strong>on</strong>g>of</str<strong>on</strong>g> the dramatic examples being<br />

spore-<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> and multicellular organizati<strong>on</strong> in certain bacteria. In c<strong>on</strong>trast, many other bacteria, am<strong>on</strong>g<br />

them E.coli and Mycobacterium sp., resp<strong>on</strong>d by entering a stress-induced program <str<strong>on</strong>g>of</str<strong>on</strong>g> lowered metabolic activity<br />

and increased resistance to various kinds <str<strong>on</strong>g>of</str<strong>on</strong>g> envir<strong>on</strong>mental assaults.<br />

The transiti<strong>on</strong> from exp<strong>on</strong>ential growth to stati<strong>on</strong>ary phase c<strong>on</strong>diti<strong>on</strong>s is accompanied by physiological and<br />

morphological changes al<strong>on</strong>g with changes in gene-expressi<strong>on</strong> patterns, with a generalized shutdown <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

protein synthesis and up-regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> genes with important functi<strong>on</strong>s during the stati<strong>on</strong>ary phase.<br />

The Dps (DNA binding Protein from Starved cells) protein binds to DNA without an apparent sequencespecificity<br />

and is c<strong>on</strong>sidered to protect DNA by direct binding, as well as by its ferroxidati<strong>on</strong> activity that<br />

protects the cell from Fent<strong>on</strong>-reacti<strong>on</strong> mediated damage.<br />

Up<strong>on</strong> entry into carb<strong>on</strong>-limited stati<strong>on</strong>ary phase, M.smegmatis also undergoes physiological changes. In order<br />

to study the stati<strong>on</strong>ary phase resp<strong>on</strong>se in more detail, we decided to focus <strong>on</strong> the M.smegmatis Dps (DNA-<br />

Binding Protein from Starved Cells) that was identified by previous work in the lab as a protein whose<br />

expressi<strong>on</strong> was up regulated under c<strong>on</strong>diti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> carb<strong>on</strong> deprivati<strong>on</strong>.<br />

The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> the present study is to investigate the in vivo functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Dps in Mycobacterium smegmatis as well as<br />

its structural features.<br />

125


23-26 August 2007,<br />

Budapest, Hungary<br />

2G_05_P<br />

(poster secti<strong>on</strong> B1, poster board #241, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ANTIBACTERIAL EFFECT OF SCUTELLARIAE RADIX ON WILD TYPE AND<br />

MUTANTS OF E.COLI<br />

Ju-Young Jung 1 , Chi-Y<strong>on</strong>g Eom 2 , Se<strong>on</strong>g-Kyu Park 3 , Jae-Ri Jung 4 , Je<strong>on</strong>g-Han Suh 1 ,<br />

H<strong>on</strong>g-Yeoul Kim 1,4*<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, College <str<strong>on</strong>g>of</str<strong>on</strong>g> Oriental Medicine, Kyung Hee University, Seoul, Korea (Rep.),<br />

2Metabolome Analysis Team, KBSI Seoul Center, 126-16 Anam-D<strong>on</strong>g, Korea University, Seoul, Korea (Rep.),<br />

3Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Prescripti<strong>on</strong>ology, College <str<strong>on</strong>g>of</str<strong>on</strong>g> Oriental Medicine, Kyung Hee University, Seoul, Korea (Rep.),<br />

4HelixPharms Co., Ltd., Seoul, Korea (Rep.) *e-mail: hyk@khu.ac.kr<br />

Inorganic polyphosphate(poly P), a linear polymer <str<strong>on</strong>g>of</str<strong>on</strong>g> hundreds <str<strong>on</strong>g>of</str<strong>on</strong>g> phosphate residues linked by high-energy<br />

phosphoanhydride b<strong>on</strong>ds, is having been found in all microbes, fungi, plants and animals examined. In E. coli,<br />

and enzyme resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> polyP synthesis is the polyphosphate kinase (ppK), exopolyphospatase (ppX),<br />

which hydrolysed the terminal residues <str<strong>on</strong>g>of</str<strong>on</strong>g> poly P, was discovered at same oper<strong>on</strong>. Natural herbal extract <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Scutellariae Radix (Scutellaria baicalensis George) have been used as antibacterial substance against infected<br />

bacteria. Scutellariae Radix extract showed remarkable antimicrobial activity against S. typhimurium and S.<br />

typhimurium ∆ppK, but little attenti<strong>on</strong> has been paid to the selective antibacterial activity <str<strong>on</strong>g>of</str<strong>on</strong>g> pathogenic and<br />

n<strong>on</strong>pathogenic bacteria. This study suggests the possible mechanism(s) <str<strong>on</strong>g>of</str<strong>on</strong>g> antibacterial activity in terms <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

relati<strong>on</strong>ship between ppK and other relating genes, sspA(stringent starvati<strong>on</strong> protein A) and sspB, sspA,<br />

B oper<strong>on</strong> and other ppK-related genes.<br />

126


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2H. OXIDATIVE STRESS<br />

2H_01_P<br />

(poster secti<strong>on</strong> A2, poster board #107, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STRESS RESPONSES IN PROCHLOROCOCCUS: OXIDATIVE PROTEOLYSIS OF<br />

GLUTAMINE SYNTHETASE AND DIFFERENTIAL EXPRESSION OF CLPP1, FTSH1<br />

AND LON GENES<br />

G. Gómez-Baena 1 , A. López-Lozano 1 , O. A. Rangel 1 , R.P. D<strong>on</strong>alds<strong>on</strong> 2 , J. M. García-Fernández 1 ,<br />

J. Díez 1<br />

1Departamento de Bioquímica y Biología Molecular. Universidad de Córdoba. Campus Rabanales. Edificio Severo<br />

Ochoa. 14071-Córdoba, Spain, e-mail: v52gobag@uco.es<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biological Sciences. George Washingt<strong>on</strong> University. 2023 G Street, NW, 20052, Washingt<strong>on</strong>, D.C.<br />

USA, e-mail: robd<strong>on</strong>@gwu.edu<br />

Prochlorococcus is a marine cyanobacterium resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> a significant part <str<strong>on</strong>g>of</str<strong>on</strong>g> the global primary producti<strong>on</strong> as<br />

well as <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the most abundant organisms <strong>on</strong> Earth. Hence, the knowledge <str<strong>on</strong>g>of</str<strong>on</strong>g> nutrient assimilati<strong>on</strong> in<br />

Prochlorococcus could provide important in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> about the physiology <str<strong>on</strong>g>of</str<strong>on</strong>g> this amazing organism and its<br />

ability to cope with limitati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> nutrients and light in the oceanic envir<strong>on</strong>ment. The main limiting nutrient in<br />

the ocean is nitrogen, and glutamine synthetase (GS) catalyzes its incorporati<strong>on</strong> into carb<strong>on</strong> backb<strong>on</strong>es,<br />

c<strong>on</strong>necting the nitrogen and carb<strong>on</strong> metabolism. Our work is focused <strong>on</strong> the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> this important<br />

enzyme under stress c<strong>on</strong>diti<strong>on</strong>s. Our results show that the degradati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> GS is regulated by metal-catalyzed<br />

oxidati<strong>on</strong> in Prochlorococcus. The proteolysis OF GS IS mediated by proteases that recognize the oxidized <str<strong>on</strong>g>for</str<strong>on</strong>g>m<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the enzyme. Starvati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> essential nutrients (such a nitrogen and ir<strong>on</strong>) and changes in the redox and<br />

energetic state <str<strong>on</strong>g>of</str<strong>on</strong>g> the cell seem to promote the oxidative modificati<strong>on</strong> and degradati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> GS. On the other<br />

hand, we have measured the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the clpP1, ftsH1 and l<strong>on</strong> genes, encoding <str<strong>on</strong>g>for</str<strong>on</strong>g> three proteases, under<br />

different stress c<strong>on</strong>diti<strong>on</strong>s, finding that ClpP and La proteases seem to play an important role in the resp<strong>on</strong>se<br />

to stress c<strong>on</strong>diti<strong>on</strong>s in Prochlorococcus.<br />

2H_02_P<br />

(poster secti<strong>on</strong> A2, poster board #108, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HYDROGEN PEROXIDE ENZYMATIC DETOXIFYCATION IN SWINE<br />

FOLLICULAR FLUID: FOLLICULAR SIZE-DEPENDENT CHANGES<br />

Giuseppina Basini, Sim<strong>on</strong>a Bussolanti, Sujen Ele<strong>on</strong>ora Santini, Francesca Grasselli<br />

Dipartimento di Produzi<strong>on</strong>i Animali, Biotecnologie Veterinarie, Qualità e Sicurezza degli Alimenti, Sezi<strong>on</strong>e di<br />

Fisiologia Veterinaria, Università di Parma, 43100 Parma, Italy<br />

e-mail: basini@unipr.it<br />

Follicular fluid (FF) is produced during folliculogenesis, which begins in the ovarian cortex with the<br />

recruitment <str<strong>on</strong>g>of</str<strong>on</strong>g> primordial follicles and ends in ovulati<strong>on</strong> or atresia. FF may be regarded as a biological<br />

“window” reflecting metabolic and horm<strong>on</strong>al processes occurring in the microenvir<strong>on</strong>ment <str<strong>on</strong>g>of</str<strong>on</strong>g> the maturing<br />

oocytes be<str<strong>on</strong>g>for</str<strong>on</strong>g>e ovulati<strong>on</strong> and also a predictor <str<strong>on</strong>g>of</str<strong>on</strong>g> the successful fertilizati<strong>on</strong>. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, the knowledge <str<strong>on</strong>g>of</str<strong>on</strong>g> its<br />

compositi<strong>on</strong> deserves special attenti<strong>on</strong>. Oxidative metabolism is essential <str<strong>on</strong>g>for</str<strong>on</strong>g> energy producti<strong>on</strong> and is<br />

unavoidably associated with the generati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> reactive oxygen species (ROS). The objective <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was<br />

there<str<strong>on</strong>g>for</str<strong>on</strong>g>e to evaluate, in FF collected from small (5 mm) swine<br />

127


23-26 August 2007,<br />

Budapest, Hungary<br />

follicles, hydrogen peroxide (H 2 O 2 ) c<strong>on</strong>centrati<strong>on</strong> and the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the enzymes involved in H 2 O 2 removal,<br />

glutathi<strong>on</strong>e peroxidase (GSH-Px) and catalase (CAT). Our data evidenced that H 2 O 2 levels significantly<br />

(p


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2H_04_P<br />

(poster secti<strong>on</strong> A2, poster board #110, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ISOTHIOCYANATES ENHANCE CYTOTOXIC EFFECT OF PEROXYNITRITE ON<br />

PMNS: A CORRELATION WITH NITROSATIVE STRESS BIOMARKER<br />

N. Trakranrungsie 1 , P. Yatmark 1 , Y. Maneerat 2 , K. Kirtikara 3<br />

1Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Veterinary Science, Mahidol University, Thailand<br />

2Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Tropical Medicine, Mahidol University, Thailand<br />

3Nati<strong>on</strong>al Center <str<strong>on</strong>g>for</str<strong>on</strong>g> Genetic Engineering and Biotechnology, Thailand<br />

e-mail: vsntk@mahidol.ac.th<br />

Isothiocyanates (ITCs), a class <str<strong>on</strong>g>of</str<strong>on</strong>g> phytochemicals available from many cruciferous vegetables, have been<br />

reported to have a dichotomous modulating effect <strong>on</strong> oxidative stress. In additi<strong>on</strong>, it has been suggested that<br />

ITCs may display a more cytotoxicity to tumor cells than normal cells. Although the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> ITCs are<br />

intensively studied in cancer cell lines, the experiments <strong>on</strong> normal cells appear limited. The present study<br />

there<str<strong>on</strong>g>for</str<strong>on</strong>g>e investigated the modulatory role <str<strong>on</strong>g>of</str<strong>on</strong>g> allyl and benzyl isothiocyanates (AITC and BITC, respectively)<br />

<strong>on</strong> peroxynitrite-induced cytotoxic effect in polymorph<strong>on</strong>uclear neutrophils (PMNs) and simultaneously<br />

evaluated the <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 8-isoprostane and nitrotyrosine as biomarkers <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidative and nitrosative stress.<br />

3-Morpholinsydn<strong>on</strong>imine (SIN-1), a peroxynitrite d<strong>on</strong>or, reduced PMNs viability but increased 8-isoprostane<br />

and nitrotyrosine levels in a c<strong>on</strong>centrati<strong>on</strong>-dependent manner. AITC or BITC al<strong>on</strong>e at c<strong>on</strong>centrati<strong>on</strong> ≤ 3<br />

µM did not cause the cytotoxic effect as compared to untreated c<strong>on</strong>trol cells. On the c<strong>on</strong>trary, pretreatment<br />

with AITC or BITC (.003, .03, .3, 3 µM) further enhanced cytotoxicity in SIN-1-treated cells. It was also<br />

observed that the increased <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> nitrotyrosine, but not 8-isoprostane, was depended <strong>on</strong> AITC or<br />

BITC c<strong>on</strong>centrati<strong>on</strong> and correlated with the loss <str<strong>on</strong>g>of</str<strong>on</strong>g> PMNs viability. The results suggest that these ITCs<br />

could exacerbate the cytotoxic effect <str<strong>on</strong>g>of</str<strong>on</strong>g> peroxynitrite in isolated milk PMNs, in part, by enhancing nitrosative<br />

stress.<br />

2H_05_P<br />

(poster secti<strong>on</strong> A2, poster board #111, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HSP70 AND HSP32 (HO-1) ARE ESSENTIAL FOR THE FOLATE INDUCED<br />

PROTECTION AGAINST HOMOCYSTEINE INDUCED DAMAGE IN<br />

ENDOTHELIAL CELLS<br />

Catherine S. Birch, John H. H. Williams<br />

Chester <str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g> Stress <str<strong>on</strong>g>Research</str<strong>on</strong>g>, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Chester, Chester, CH1 4BJ UK<br />

e-mail: john.williams@chester.ac.uk<br />

Homocysteine (Hcy) is an independent risk factor <str<strong>on</strong>g>for</str<strong>on</strong>g> vascular disease and Alzheimer’s disease. Hcy promotes<br />

its harmful effect by promoting the generati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> reactive oxygen species (ROS) within the vasculature<br />

leading to oxidative damage and initiating inflammatory resp<strong>on</strong>ses which ultimately lead to endothelial<br />

dysfuncti<strong>on</strong>. Folate treatment is an established therapeutic strategy to reduce Hcy in patients suffering with<br />

vascular disorders. Endothelial cells counteract the harmful effect <str<strong>on</strong>g>of</str<strong>on</strong>g> inflammati<strong>on</strong> by upregulating a set <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

protective genes including the heat shock proteins (Hsps). Hsps are a highly c<strong>on</strong>served group <str<strong>on</strong>g>of</str<strong>on</strong>g> inducible<br />

stress proteins involved in the protecti<strong>on</strong> and c<strong>on</strong>servati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> protein synthesis. Hsp70 is upregulated when<br />

cells are subjected to stressful stimuli, Hsp32 (HO-1) is highly inducible when faced with oxidative stress.<br />

129


23-26 August 2007,<br />

Budapest, Hungary<br />

Data from qPCR show that Hsp70 and Hsp32 (HO-1) are upregulated in a dose-dependent manner when<br />

cells are exposed to Hcy. MTS viability assays indicate that folate (5-MTHF) protects cells from Hcy-induced<br />

damage. Inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 (quercetin) and Hsp32 (Sn(IX) protoporphyrin) reduce the protective effect <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

5-MTHF. RNAi targeted to Hsp70 and Hsp32 (HO-1) support this data. Combined treatment (quercetin plus<br />

Sn(IX) protoporphyrin) totally inhibit the protective effect <str<strong>on</strong>g>of</str<strong>on</strong>g> 5-MTHF. Combined treatment with siRNA <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

Hsp70 and Hsp32 also totally prevent 5-MTHF protecti<strong>on</strong>.<br />

130<br />

2H_06_P<br />

(poster secti<strong>on</strong> A2, poster board #112, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE BENEFICIAL EFFECTS OF LIPOIC ACID ON PROTEIN OXIDATION AND<br />

TOTAL ANTIOXIDANT CAPACITY IN EXPERIMENTAL ADRIAMYCIN<br />

(DOXORUBICIN) INDUCED OXIDATIVE CARDIOTOXICITY AND<br />

NEPHROTOXICITY<br />

Pinar Atukeren 1 , Zeynep Ozturk 1 , Ezel Uslu 1 , Ibrahim Bayrak 2 , M. Koray Gumustas 1 ,<br />

Merve Aldikactioglu 3 , Selin Sert 3 , Murat Yassa 3 , Tuncay Altug 2<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry,<br />

2Animal Producti<strong>on</strong> and <str<strong>on</strong>g>Research</str<strong>on</strong>g> Laboratory,<br />

3Cerrahpasa Medical School, Cerrahpasa Medical Faculty, Istanbul University, Istanbul, Turkey<br />

e-mail: p_atukeren@yahoo.com<br />

Doxorubicin (DOX) is a widely used anticancer agent whose clinical use is limited <strong>on</strong> account <str<strong>on</strong>g>of</str<strong>on</strong>g> its toxicity<br />

which have been associated with reactive oxygen species (ROS).Lipoic acid (LA) has cytoprotective potential<br />

which has previously been explained by its antioxidant properties distinguishing from other antioxidants:LA<br />

neutralizes free radicals in both the fatty and watery regi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> cells,and functi<strong>on</strong>s in both its reduced and<br />

oxidized <str<strong>on</strong>g>for</str<strong>on</strong>g>ms.The present study aimed to investigate the protective efficacy <str<strong>on</strong>g>of</str<strong>on</strong>g> LA <strong>on</strong> DOX induced<br />

oxidative damage in heart and kidney tissues.40 rats were divided into five groups:1st;c<strong>on</strong>trol, 2nd;<strong>on</strong>ly DOX<br />

injected, 3rd;<strong>on</strong>ly LA administered (50mg/kg/day,20 days), 4th;LA administered <str<strong>on</strong>g>for</str<strong>on</strong>g> 20 days+DOX injected,<br />

5th; single dose LA (50mg/kg) injected same time with DOX injecti<strong>on</strong>.DOX was injected to the rats<br />

(4mg/kg) 48 hours be<str<strong>on</strong>g>for</str<strong>on</strong>g>e sacrifizati<strong>on</strong>.In heart and kidney tissues; protein oxidati<strong>on</strong> was assesed by the levels<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> protein carb<strong>on</strong>yls (PC),also thiobarbituric acid reacting substances (TBARS) were determined and<br />

antioxidant status <str<strong>on</strong>g>of</str<strong>on</strong>g> tissues were determined by evaluating total antioxidant capacity (TAC) and superoxide<br />

dismutase (SOD) activity.PC and TBARS levels were elevated in both tissues, in DOX injected groups when<br />

compared with the c<strong>on</strong>trol and LA pretreatment restored TAC levels and SOD activity.The study has<br />

highlighted the beneficial effects <str<strong>on</strong>g>of</str<strong>on</strong>g> lipoic acid pretreatment in reversing the damages caused by doxorubicin.<br />

2H_07_P<br />

(poster secti<strong>on</strong> A2, poster board #113, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

OXIDATIVE STRESS DURING PERIPARTUM PERIOD IN MARES: FIELD STUDY<br />

F. Righi, S. E. Santini, E. Gervasini, S. Bussolati, G. Basini, A. Quarantelli<br />

Dipartimento di Produzi<strong>on</strong>i Animali, Biotecnologie Veterinarie, Qualità e Sicurezza degli Alimenti. Università di<br />

Parma. 43100 Parma, Italy; e-mail: federico.righi@unipr.it<br />

Oxidative stress has been shown to occur in dairy cattle around calving, but there is no current data <strong>on</strong><br />

similar phenomen<strong>on</strong> in periparturient mares. In a previous study we found a decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> Ferric Reducing<br />

Ability (FRAP) in mares’ sera around parturiti<strong>on</strong>, suggesting n<strong>on</strong>-enzymatic antioxidant molecule


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

c<strong>on</strong>sumpti<strong>on</strong> during this period. Since this mechanism could lead to redox imbalance, the aim <str<strong>on</strong>g>of</str<strong>on</strong>g> the present<br />

study was to investigate the possible insurgence <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidative stress in mares around foaling by analyzing the<br />

same samples. Levels <str<strong>on</strong>g>of</str<strong>on</strong>g> Reactive Oxygen Molecules (ROMs) were measured by colorimetric method <strong>on</strong><br />

blood samples collected from 17 Thoroughbred mares at days -24 (T1), +1 (T2) and +9 (T3) relative to<br />

parturiti<strong>on</strong>. No significant variati<strong>on</strong> was detected in ROMs c<strong>on</strong>centrati<strong>on</strong> at the intervals c<strong>on</strong>sidered. These<br />

findings seem to exclude the occurrence <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidative stress during the peripartum period in mares. Further<br />

research is needed to better define the mechanisms involved in oxidative status regulati<strong>on</strong> around foaling.<br />

2H_08_P<br />

(poster secti<strong>on</strong> A2, poster board #114, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

BRAIN OXIDATIVE STRESS IN HYPERTENSIVE RATS<br />

M. R. Garrido, A. Csibi, M. Pastorello, J. A. Silva, C. Ciangherotti, A. Israel<br />

School <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmacy, Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Neuropeptides, Universidad Central de Venezuela, Caracas, Venezuela;<br />

e-mail: mrgarrido11@hotmail.com<br />

Chr<strong>on</strong>ic exposure to stress alters prooxidant-antioxidant balance. Genetic hypertensi<strong>on</strong> (SHR) is a model <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

chr<strong>on</strong>ic stress which is known to be accompanied with increased oxidative stress and decreased antioxidant<br />

enzyme activity. Angiotensin II (ANG) influences blood pressure via its ability to stimulate the NAD(P)Hoxidase<br />

and produce reactive oxygen species (ROS) such as superoxide (0 2 .- ), which is sequentially<br />

metabolized by the enzymes superoxide dismutase and catalase. In turn, ROS can activate the mitogenactivated<br />

protein kinase (MAPK) pathway that is associated with growth and cellular differentiati<strong>on</strong>. Several<br />

indirect marker <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidative stress are increased in the brains <str<strong>on</strong>g>of</str<strong>on</strong>g> SHR compared with those <str<strong>on</strong>g>of</str<strong>on</strong>g> Wistar-Kyoto<br />

(WKY) rats. There is no direct evidence, however, <str<strong>on</strong>g>of</str<strong>on</strong>g> changes in oxidative stress and MAPK in the brain in<br />

hypertensi<strong>on</strong>. Thus, we investigates the status <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidative stress in hypothalamus (HYP) and sub<str<strong>on</strong>g>for</str<strong>on</strong>g>nical<br />

organ (SFO) <str<strong>on</strong>g>of</str<strong>on</strong>g> WKY and SHR rats, and the involvement <str<strong>on</strong>g>of</str<strong>on</strong>g> ANG II, by measuring the enzymatic activity<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> catalase (CAT) and the expressi<strong>on</strong> and activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> MAPKs. Male rats, 200-220g, normotensive WKY or<br />

SHR were sacrificed and tissues microdissected under stereomicroscopic c<strong>on</strong>trol. CAT activity was<br />

determined spectrophotometrically and ERKs activati<strong>on</strong> by Western blot analysis. In SHR brain structures<br />

such as HYP and SFO, CAT activity was decreased when compared with WKY. Likewise, the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

MAPKs and its activati<strong>on</strong> was diminished in the SHR rats. In both structures ANG II (10 -7 M) decreased<br />

CAT activity. Our results indicate an alterati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidative stress and MAPK activati<strong>on</strong> in the brain <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

hypertensive rats (Grant CDCH PG 06-30-5222-2003).<br />

131


23-26 August 2007,<br />

Budapest, Hungary<br />

132<br />

2H_09_P<br />

(poster secti<strong>on</strong> A2, poster board #115, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ADRENOMEDULLIN, NITRIC OXIDE AND SUPEROXIDE DISMUTASE IN<br />

EXPERIMENTAL STRESS ULCER FORMATION<br />

Yesim Temiz 1 , Hakan Ekmekci 2 , Ezel Uslu 1 , Tuncay Altug 3 , M. Koray Gumustas 1<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, Cerrahpasa Medical Faculty, Istanbul University, Istanbul, Turkey<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Pediatric Heamatology/Oncology, B<strong>on</strong>e Marrow Transplantati<strong>on</strong> Unit, Istanbul Medical Faculty,<br />

Istanbul University, Istanbul, Turkey<br />

3Animal Producti<strong>on</strong> and <str<strong>on</strong>g>Research</str<strong>on</strong>g> Laboratory, Cerrahpasa Medical Faculty, Istanbul University, Istanbul, Turkey<br />

e-mail: pr<str<strong>on</strong>g>of</str<strong>on</strong>g>gumustas@yahoo.com<br />

Stress ulcer is a comm<strong>on</strong> cause <str<strong>on</strong>g>of</str<strong>on</strong>g> gastrointestinal bleeding and oxygen derived free radicals play important<br />

role in the pathogenesis.Adrenomedullin (ADM) is a vasodilator peptide; together with nitric oxide (NO) and<br />

endothelin, it is <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the secretuar product <str<strong>on</strong>g>of</str<strong>on</strong>g> endothel and its vasodilator effect is supposed to depend <strong>on</strong><br />

NO and effective <str<strong>on</strong>g>of</str<strong>on</strong>g> protecting the gastric mucosa.Calcium dobesilate (Doxium © ) is an angioprotective agent.<br />

In our study, we investigated the ADM and NO levels in experimentally induced stress model, in rats. We<br />

sought TBARS levels and SOD activity together with the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> calcium dobesilate as well. In ulcerative<br />

group; plasma ADM and NO,and in gastric mucosa NO levels were significantly increased. Elevated TBARS<br />

levels and decreased SOD activity were found in gastric mucosa.On the other hand, pretreated with calcium<br />

dobesilate, showed lower plasma ADM and elevated NO levels compared to ulcerative animals. No<br />

difference were found in comparis<strong>on</strong> between two groups <str<strong>on</strong>g>for</str<strong>on</strong>g> NO levels. But TBARS levels were lower in<br />

calcium dobesilate pretreated group’s gastric mucosa. Our results suggests that, in stress ulcer, elevated levels<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> ADM and NO, may generated to protect organism to a resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> increased oxidative stress and<br />

decreased gastric mucosal blood flow. Although these results supports that calcium dobesilate has an<br />

antioxidant and angioprotective properties, it is needed to work <str<strong>on</strong>g>for</str<strong>on</strong>g> further <strong>on</strong> this molecule.<br />

2H_10_P<br />

(poster secti<strong>on</strong> A2, poster board #116, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ROLE OF OXIDATIVE STRESS IN THE NATRIURESIS INDUCED BY CENTRAL<br />

ADMINISTRATION OF ANGIOTENSIN II<br />

A. Israel, J. Arzola, M. Varela, S. De Jesús, Y. Toro<br />

Unidad de Neuropéptidos, School <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmacy, Universidad Central de Venezuela<br />

e-mail: astern88@hotmail.com<br />

The brain renin angiotensin system has an important role in the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> arterial blood pressure and fluid<br />

and electrolyte metabolism. Abundant evidence now suggest that a key mechanism by which central<br />

angiotensin II (ANG) influences blood pressure it’s via its ability to produce reactive oxygen species (ROS)<br />

such as superoxide (0 2 .- ). Cerebroventricular (IVT) ANG administrati<strong>on</strong> to c<strong>on</strong>scious rats produces<br />

antidiuresis and natriuresis, through stimulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> AT 1 receptor. The role <str<strong>on</strong>g>of</str<strong>on</strong>g> ROS in these acti<strong>on</strong>s is<br />

unknown. Thus, we assessed the involvement <str<strong>on</strong>g>of</str<strong>on</strong>g> brain NAD(P)H-oxidase and ROS by the use <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

selective NAD(P)H-oxidase inhibitor, Apocynin (APO) and the superoxide dismutase mimetic, Tempol<br />

(TEM) administered IVT. Male Sprague-Dawley rats (220 g) were IVT-cannulated and subjected to the<br />

following IVT-treatments: saline (5 µl), APO (33 µg/kg/5µl); TEM (50 µmol/10µl). After 30 minutes <str<strong>on</strong>g>of</str<strong>on</strong>g>


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

each treatment, animals received an IVT-injecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> either ANG (50 pmol/5µl) or saline (5 or 10 µl).<br />

Animals were water loaded (20 ml/kg, p.o.) and placed into metabolic cages. Urine was collected at 1, 3 and<br />

6 hr. Urinary Na + and K + was determined by flame photometry. ANG-IVT reduced urine volume at 1 hr and<br />

increased sodium and potassium excreti<strong>on</strong> at 1, 3 y 6 hrs. Central administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> APO or TEMP<br />

significantly reduced ANG-induced natriuretic effect (P


23-26 August 2007,<br />

Budapest, Hungary<br />

134<br />

2H_12_P<br />

(poster secti<strong>on</strong> A2, poster board #118, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SOCIAL STRESS INCREASES URINARY EXCRETION OF BIOPYRRINS,<br />

OXIDATIVE METABOLITES OF BILIRUBIN, IN MICE<br />

Keiko Motoyama 1 , Tomoya Miyashita 1 , Tokio Yamaguchi 2 , Satoru Hirabayashi 1 ,<br />

Hirotaka Naito 1 , Norio Ohashi 1 , Keiko Unno 3 , Hiroyuki Sakakibara 1 , Kayoko Shimoi 1,*<br />

1Graduate School <str<strong>on</strong>g>of</str<strong>on</strong>g> Nutriti<strong>on</strong>al and Envir<strong>on</strong>mental Sciences, and 3 Graduate School <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmaceutical Sciences,<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Shizuoka, 52-1 Yada, Suruga-ku, Shizuoka 422-8526, Japan, 2 Medical <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute, Tokyo<br />

Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8510, Japan<br />

*e-mail: shimoi@smail.u-shizuoka-ken.ac.jp (K. Shimoi)<br />

In this study, we investigated whether or not urinary excreti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> biopyrrins, oxidative metabolites <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

bilirubin, in mice could be used as a biomarker <str<strong>on</strong>g>for</str<strong>on</strong>g> psychosocial stress. Male BALB/c mice (4 weeks old)<br />

were housed 5 per cage <str<strong>on</strong>g>for</str<strong>on</strong>g> 10 days. After acclimatizati<strong>on</strong>, mice were exposed to two kinds <str<strong>on</strong>g>of</str<strong>on</strong>g> psychosocial<br />

stress; isolati<strong>on</strong> (1 mouse per cage) and c<strong>on</strong>fr<strong>on</strong>tati<strong>on</strong> (2 mice per partiti<strong>on</strong>ed cage <str<strong>on</strong>g>for</str<strong>on</strong>g> 10 days and then<br />

partiti<strong>on</strong> was removed). Mouse blood, urine, and liver were collected after 7 and 30 days <str<strong>on</strong>g>of</str<strong>on</strong>g> stress. Serum<br />

levels <str<strong>on</strong>g>of</str<strong>on</strong>g> corticoster<strong>on</strong>e and urinary levels <str<strong>on</strong>g>of</str<strong>on</strong>g> biopyrrins were determined by EIA and ELISA, respectively. An<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> heme oxygenase-1 (HO-1), which is known to be induced by oxidative stress, was analyzed<br />

using the methods <str<strong>on</strong>g>of</str<strong>on</strong>g> RT-PCR and western blot in the mouse liver. Adrenal hypertrophy and significant<br />

increases in serum c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> corticoster<strong>on</strong>e were observed in mice exposed to these types <str<strong>on</strong>g>of</str<strong>on</strong>g> social<br />

stress <str<strong>on</strong>g>for</str<strong>on</strong>g> 7 and 30 days. Both <str<strong>on</strong>g>of</str<strong>on</strong>g> the social stress <str<strong>on</strong>g>for</str<strong>on</strong>g> 7 days markedly induced the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HO-1 in the<br />

liver, and significantly increased the urinary excreti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> biopyrrins. These phenomen<strong>on</strong>s decreased after<br />

30 days, although they were still rather high compared to the c<strong>on</strong>trol group. These results suggested that<br />

social stress causes oxidative stress and that biopyrrins could be useful biomarkers <str<strong>on</strong>g>of</str<strong>on</strong>g> psychosocial stress.<br />

2H_13_P<br />

(poster secti<strong>on</strong> A2, poster board #119, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

BIOCHEMICAL EVALUATION OF THE ROS AND SIALIC ACID LEVELS IN<br />

CARDIAC AND RENAL TISSUES OF ADRIAMYCIN- ADMINISTRATED RATS AND<br />

THE POSSIBLE ROLE OF LIPOIC ACID TREATMENT<br />

Zeynep Ozturk, Pınar Atukeren, Ezel Uslu, İbrahim Bayrak, Hüseyin Sönmez, Özlem Armay,<br />

Merve Aldıkaçtıoğlu, Murat Yassa, Tuncay Altuğ<br />

e-mail: zozturk72@hotmail.com<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, Cerrahpasa Medical Faculty, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Istanbul<br />

Experimental Animals Breeding and <str<strong>on</strong>g>Research</str<strong>on</strong>g> Laboratory, Cerrahpasa Medical Faculty, University <str<strong>on</strong>g>of</str<strong>on</strong>g> İstanbul<br />

Adriamycin (doxorubicin) is an anthracycline antibiotic that has been used <str<strong>on</strong>g>for</str<strong>on</strong>g> more than 30 years <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> a wide variety <str<strong>on</strong>g>of</str<strong>on</strong>g> cancers. It is widely accepted that oxidative stress and the producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> free<br />

radicals are involved in doxorubicin acti<strong>on</strong>, both in terms <str<strong>on</strong>g>of</str<strong>on</strong>g> antitumor effects and cardiotoxicity. Sialic acid;<br />

plays an important role in cellular functi<strong>on</strong>s and also influences c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> glycoproteins which is found<br />

<strong>on</strong> the cell membrane.Free radicals are potentially dangerous products <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular metabolism. Excessive<br />

amounts <str<strong>on</strong>g>of</str<strong>on</strong>g> reactive oxygen species (ROS) can start some lethal chain reacti<strong>on</strong>s such as ROS that induces<br />

lipid peroxidati<strong>on</strong>. Alpha-lipoic acid is a powerful antioxidant. In this study we investigated the ROS and


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

sialic acid levels in cardiac and renal tissues <str<strong>on</strong>g>of</str<strong>on</strong>g> doxorubucin- administrated rats and the possible role <str<strong>on</strong>g>of</str<strong>on</strong>g> lipoic<br />

acid.40 Wistar albino rats were divided into five groups: c<strong>on</strong>trol, ADR group (4mg/kg intraperit<strong>on</strong>eal), Acute<br />

-lipoic acid and ADR (single dose 50mg/kg lipoic acid,4mg/kg ADR intraperit<strong>on</strong>eal), <strong>on</strong>ly lipoic acid<br />

(50mg/kg,intraperit<strong>on</strong>eal), the last group lipoic acid (50mg/kg, intraperit<strong>on</strong>eal) and ADR (4mg/kg daily<br />

intraperit<strong>on</strong>eal). Sialic acid and TBARS levels were elevated in ADR, LA, LA+DOX groups as compared<br />

with the c<strong>on</strong>trol group in the cardiac tissue. In renal tissue, sialic acid levels declined, in LA and acute<br />

LA+ADR, whereas TBARS levels were elevated in ADR and LA+ADR group.<br />

2H_14_P<br />

(poster secti<strong>on</strong> A2, poster board #120, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ACTIVATION OF CELLULAR ANTIOXIDANT DEFENCE SYSTEMS AFTER<br />

EXOGENOUS HEAT STRESS IN MEN<br />

Il<strong>on</strong>a Pokora 1 , Katarzyna Kempa 2 , Elżbieta Kimsa 2 , Zbigniew Pokora 3<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Physiology, Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Physical Educati<strong>on</strong>, Katowice, Poland, e-mail: i.pokora@awf.katowice.pl<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Physical Educati<strong>on</strong>, Katowice, Poland<br />

3Department <str<strong>on</strong>g>of</str<strong>on</strong>g> General and Molecular Biology, and Genetics, Silesian Medical University, Katowice, Poland<br />

The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> our study was to examine changes in the blood and cellular antioxidant defense system in<br />

resp<strong>on</strong>se to exogenous heat stress (induced by sauna bath) in young men. At rest prior and after sauna bath<br />

tympanic (T ty ) and skin temperatures (T sk ) were measured and body heat storage (S) was calculated. Be<str<strong>on</strong>g>for</str<strong>on</strong>g>e<br />

sauna bath, then 60 min, 120 min and 24h post sauna, capillary blood samples were drawn <str<strong>on</strong>g>for</str<strong>on</strong>g> determinati<strong>on</strong>s<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g>: hemoglobin c<strong>on</strong>tent (HB), hematocrit (HCT), glucose (GLU), blood carb<strong>on</strong> dioxide (pCO 2 ), pH and<br />

blood oxygen (pO 2 ), respectively. The changes in blood (%BV), plasma (%PV) and cell (%CV) volumes were<br />

calculated. Additi<strong>on</strong>ally, venous blood samples were withdrawn to determine plasma: cortisol level, activity <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

creatine kinase (CK-NAC). Blood antioxidant potential was assessed by determinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> antioxidant enzyme<br />

activities: glutathi<strong>on</strong>e peroxidase (GSH-Px), superoxide dismutase (SOD), catalase (CAT) and glutathi<strong>on</strong>e<br />

reductase (GR). Exogenous heat stress induced an increase in Tty (∆+2.1 o C) and plasma cortisol level,<br />

decreased cell (-.05%), blood (-2.97%) and plasma (-4.36%) volumes. Cell volume started to increase after 2h<br />

recovery. CK-NAC activity was increased provided evidence slight increase in muscle cell membranes<br />

permeability. There was a decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> activity <str<strong>on</strong>g>of</str<strong>on</strong>g> GSH-Px and CAT as well as an increase in activities <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

SOD and GR immediately post sauna. The observati<strong>on</strong>s in this study imply that exogenous heat stress was<br />

associated with activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular and blood antioxidant defense system.<br />

135


23-26 August 2007,<br />

Budapest, Hungary<br />

2H_15_P<br />

(poster secti<strong>on</strong> A2, poster board #121, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PROTECTIVE EFFECTS OF LIPID OXIDATION PRODUCTS AGAINST NEURONAL<br />

CELL DEATH INDUCED BY GLUTAMATE AND 6-HYDROXYDOPAMINE:<br />

INDUCTION OF ADAPTIVE RESPONSE AND ENHANCEMENT OF CELL<br />

TOLERANCE<br />

Yoshiro Saito, Zhi-Hua Chen, Yasukazu Yoshida, Etsuo Niki<br />

Human Stress Signal Res Ctr, Nat Inst Adv Ind Sci Tech (AIST), Ikeda Osaka, JAPAN<br />

e-mail: yoshiro-saito@aist.go.jp<br />

There is increasing evidence to suggest that reactive oxygen species, including a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> lipid peroxidati<strong>on</strong><br />

products, can induce an adaptive resp<strong>on</strong>se and enhance cell tolerance; however, the details <str<strong>on</strong>g>of</str<strong>on</strong>g> the underlying<br />

molecular mechanisms have not been clarified. In the present study using both PC12 cells and cultured<br />

cortical neur<strong>on</strong>s, we investigated the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> lipid oxidati<strong>on</strong> products such as 15-deoxy-∆ 12,14 -prostaglandin J 2<br />

(15d-PGJ 2 ), 4-hydroxy-2-n<strong>on</strong>enal (4-HNE) and 7-hydroxycholesterol (7-OHCh) against the cell death<br />

induced by 6-hydroxydopamine (6-OHDA) and glutamate. Pretreatment with these lipid oxidati<strong>on</strong> products<br />

at sublethal c<strong>on</strong>centrati<strong>on</strong>s resulted in a significant protective effect against subsequent oxidative stress, and<br />

15d-PGJ 2 , in particular, exhibited a complete protective effect against glutamate-induced neur<strong>on</strong>al cell death.<br />

Pretreatment with 15d-PGJ 2 increased the intracellular glutathi<strong>on</strong>e (GSH) as well as the gene expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

glutamate-cysteine ligase (GCL), the rate-limiting enzyme <str<strong>on</strong>g>of</str<strong>on</strong>g> GSH synthesis. 15d-PGJ 2 protected cells from<br />

glutamate-induced GSH depleti<strong>on</strong>, while the inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular GSH synthesis by buthi<strong>on</strong>ine sulfoximine<br />

abolished the adaptive resp<strong>on</strong>se induced by 15d-PGJ 2 . These findings indicate that at low levels, 15d-PGJ 2<br />

acts as a potent survival mediator against glutamate-induced insults via the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> an adaptive resp<strong>on</strong>se<br />

primarily through the up-regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the intracellular GSH synthesis. Furthermore, we observed that the<br />

adaptive resp<strong>on</strong>se induced by 4-HNE was mediated through the elevated thioredoxin reductase activity in an<br />

NF-E2-related factor 2 (Nrf2)-dependent manner, while 7-OHCh exhibited the protective effect via the<br />

increase <str<strong>on</strong>g>of</str<strong>on</strong>g> GSH c<strong>on</strong>tents in the Nrf2-independent manner. The possible physiological role <str<strong>on</strong>g>of</str<strong>on</strong>g> adaptive<br />

resp<strong>on</strong>se induced by lipid oxidati<strong>on</strong> products via different mechanism, will be discussed.<br />

2H_16_P<br />

(poster secti<strong>on</strong> A2, poster board #122, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SIMULTANEOUS EVALUATION OF OXIDATIVE AND NITROSATIVE STRESS<br />

BIOMARKERS IN PEROXYNITRITE-INDUCED CYTOTOXIC EFFECT ON<br />

POLYMORPHONUCLEAR NEUTROPHILS<br />

N. Trakranrungsie 1 , P. Yatmark 1 , Y. Maneerat 2 , K. Kirtikara 3<br />

1Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Veterinary Science, Mahidol University, Thailand<br />

2Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Tropical Medicine, Mahidol University, Thailand<br />

3Nati<strong>on</strong>al Center <str<strong>on</strong>g>for</str<strong>on</strong>g> Genetic Engineering and Biotechnology, Thailand<br />

e-mail: vspym@mahidol.ac.th<br />

Peroxynitrite is a str<strong>on</strong>g oxidant and capable <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidizing and nitrating several macromolecules. Peroxynitrite<br />

reacti<strong>on</strong> can be reflected via the <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidized and nitrated species such as isoprostanes and<br />

nitrotyrosine as observed in many animal and human c<strong>on</strong>diti<strong>on</strong>s associated with oxidative/ nitrosative<br />

136


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

damage. Since the functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> neutrophils as the first line <str<strong>on</strong>g>of</str<strong>on</strong>g> defence can be affected by oxidative/<br />

nitrosative stress, the objective <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to evaluate the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> peroxynitrite in milk-derived<br />

polymorph<strong>on</strong>uclear neutrophils (PMNs) and c<strong>on</strong>comitantly m<strong>on</strong>itor the <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 8-isoprostane and<br />

nitrotyrosine as biomarkers <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidative and nitrosative stress, respectively. 3-Morpholinsydn<strong>on</strong>imine (SIN-<br />

1), a peroxynitrite d<strong>on</strong>or, reduced PMNs viability in a c<strong>on</strong>centrati<strong>on</strong>-dependent manner with estimated EC 50<br />

value <str<strong>on</strong>g>of</str<strong>on</strong>g> 375 µM. Meanwhile, the increased levels <str<strong>on</strong>g>of</str<strong>on</strong>g> 8-isoprostane and nitrotyrosine were observed as<br />

c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> SIN-1 increased. The peroxynitrite inhibitors, glutathi<strong>on</strong>e, melat<strong>on</strong>in and uric acid,<br />

markedly and c<strong>on</strong>centrati<strong>on</strong> dependently attenuated the cytotoxicity <str<strong>on</strong>g>of</str<strong>on</strong>g> SIN-1, in which a str<strong>on</strong>g inverse<br />

relati<strong>on</strong>ship between 8-isoprostane and nitrotyrosine levels and cell survival was evident. The data imply that<br />

the cytotoxic effect <str<strong>on</strong>g>of</str<strong>on</strong>g> peroxynitrite <strong>on</strong> PMNs and the inhibitory acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> known peroxynitrite inhibitors<br />

were mediated via both oxidative and nitrosative mechanisms.<br />

2H_17_P<br />

(poster secti<strong>on</strong> A2, poster board #123, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STABILITY OF ESCHERICHIA COLI CYTOCHROME BD COMPLEXED WITH<br />

PHYSIOLOGICALLY RELEVANT LIGANDS<br />

V.B. Borisov 1 , E. Forte 2 , P. Sarti 2 , M. Brunori 2 , A.A. K<strong>on</strong>stantinov 1 , A. Giuffre 2<br />

1Belozersky Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Physico-Chemical Biology, Lom<strong>on</strong>osov Moscow State University, Moscow 119992, Russia.<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemical Sciences and CNR Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular Biology and Pathology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Rome<br />

“La Sapienza”, Rome I-00185, Italy<br />

e-mail: bor@genebee.msu.su<br />

Escherichia coli cytochrome bd c<strong>on</strong>tains three hemes, b 558 , b 595 and d, but no copper. Apart from its role in<br />

energy c<strong>on</strong>servati<strong>on</strong>, cytochrome bd plays other crucial physiological functi<strong>on</strong>s in bacteria, the enzyme being<br />

preferentially expressed under microaerobic and other stress c<strong>on</strong>diti<strong>on</strong>s. In pathogenic bacteria, mutati<strong>on</strong>s in<br />

cytochrome bd result in a reduced virulence, as if this enzyme were required <str<strong>on</strong>g>for</str<strong>on</strong>g> adaptati<strong>on</strong> to c<strong>on</strong>diti<strong>on</strong>s<br />

created by host immunity, and in particular <str<strong>on</strong>g>for</str<strong>on</strong>g> col<strong>on</strong>izati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> O 2 -poor envir<strong>on</strong>ments. As a unique feature,<br />

this oxidase is stable in a single-electr<strong>on</strong> reduced, “mixed valence” MV state, with heme d 2+ bound to O 2 , NO<br />

or CO. Stopped-flow ligand-exchange experiments at 20°C show that O 2 dissociates from the MV enzyme at<br />

k = 78 ± 0.5 s -1 . Possibly relevant to patho-physiology <str<strong>on</strong>g>of</str<strong>on</strong>g> E. coli infecti<strong>on</strong>, the NO dissociati<strong>on</strong> rate from<br />

heme d 2+ is unusually high, being 0.133 ± 0.005 s -1 <str<strong>on</strong>g>for</str<strong>on</strong>g> the fully reduced (R) enzyme and 0.036 ± 0.003 s -1 <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

the MV state. Likewise, the CO dissociati<strong>on</strong> rate is greater from R (6.0 ± 0.2 s -1 ) than from MV cytochrome<br />

bd (4.20 ± 0.34 s -1 ). These data show that the ligand dissociati<strong>on</strong> from heme d 2+ is unusually fast and<br />

c<strong>on</strong>trolled by the redox state <str<strong>on</strong>g>of</str<strong>on</strong>g> the hemes b (presumably b 595 ). Work partially supported by the Russian<br />

Foundati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> Basic <str<strong>on</strong>g>Research</str<strong>on</strong>g> (Grant 05-04-48096).<br />

137


23-26 August 2007,<br />

Budapest, Hungary<br />

138<br />

2H_18_P<br />

(poster secti<strong>on</strong> A2, poster board #124, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE EFFECTS OF DIFFERENT STRESS MODELS ON THE CUZN SUPEROXIDE<br />

DISMUTASE EXPRESSION IN RAT BRAIN<br />

Dragana Filipović, Miroslav Demajo, Marija B. Radojčić<br />

”VINČA” Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Nuclear Sciences, Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular Biology and Endocrinology,<br />

P.O.Box 522-090, 11000 Belgrade, Serbia<br />

Exposure to stress alters the normal body homeostasis and leads to the development <str<strong>on</strong>g>of</str<strong>on</strong>g> various pathological<br />

c<strong>on</strong>diti<strong>on</strong>s, which might involve alterati<strong>on</strong>s in the antioxidant defence system. In the present study we<br />

investigated the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> CuZn superoxide dismutase (CuZnSOD) in the cytosol <str<strong>on</strong>g>of</str<strong>on</strong>g> hippocampus and<br />

brain cortex <str<strong>on</strong>g>of</str<strong>on</strong>g> Wistar male rats exposed to 21 daily isolati<strong>on</strong> as chr<strong>on</strong>ic stressor, sole or in combinati<strong>on</strong> with<br />

2h acute stress <str<strong>on</strong>g>of</str<strong>on</strong>g> immobilizati<strong>on</strong> or cold (4 0 C). Expressi<strong>on</strong> pattern <str<strong>on</strong>g>of</str<strong>on</strong>g> the CuZnSOD in c<strong>on</strong>trol and stressed<br />

rats were followed at both, mRNA and protein levels, using RT-PCR and Western blot, respectively. Also,<br />

plasma corticoster<strong>on</strong>e was measured by ELISA kit.<br />

We found that CuZnSOD expressi<strong>on</strong> was increased in the hippocampus and brain cortex after both acute<br />

stressors. Opposite to that, chr<strong>on</strong>ic stress <str<strong>on</strong>g>of</str<strong>on</strong>g> isolati<strong>on</strong> led to negligible changes in hippocampal CuZnSOD<br />

level. The subsequent exposure <str<strong>on</strong>g>of</str<strong>on</strong>g> animals to acute stressors resulted in similar but a smaller increase in<br />

CuZnSOD, compared to the acute stressors, in both brain structures. These results suggest that, acute and<br />

chr<strong>on</strong>ic stressors most probably generate intracellular imbalance between producti<strong>on</strong> and eliminati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

reactive oxygen species (ROS). Relatively increased levels <str<strong>on</strong>g>of</str<strong>on</strong>g> CuZnSOD in these c<strong>on</strong>diti<strong>on</strong>s are required to<br />

remove the high level <str<strong>on</strong>g>of</str<strong>on</strong>g> ROS in order to protect against ROS damage in the hippocampus and brain cortex.<br />

The chr<strong>on</strong>ic neuroendocrine stress inactivatati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> CuZnSOD and potentially <str<strong>on</strong>g>of</str<strong>on</strong>g> other antioxidant defence<br />

enzymes may c<strong>on</strong>stitute a basis <str<strong>on</strong>g>for</str<strong>on</strong>g> propagati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> variety <str<strong>on</strong>g>of</str<strong>on</strong>g> pathological processes in the brain cells. In that<br />

view, special care should be payed to development <str<strong>on</strong>g>of</str<strong>on</strong>g> antioxidant therapeutics <str<strong>on</strong>g>for</str<strong>on</strong>g> antag<strong>on</strong>izing stress induced<br />

redox disbalances in neur<strong>on</strong>al cells.<br />

2H_19_P<br />

(poster secti<strong>on</strong> A2, poster board #125, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

OXIDATIVE STRESS MARKERS IN SURGICALLY TREATED PATIENTS WITH<br />

DRUG-RESISTANT EPILEPSY<br />

Jeffrey López*, María E. G<strong>on</strong>zález, Lourdes Lorigados, Lilia Morales, Gretel Riverón,<br />

Yosvany Bauza<br />

*Departamento de Neurología Experimental, Instituto de Neurología y Neurocirugía, Calle 29 # 139 esquina D,<br />

Vedado, Plaza, 11100 – La Habana. Cuba<br />

e-mail: erojas@infomed.sld.cu, jeffrey@neuro.ciren.cu<br />

Excitotoxic events related to epilepsy are closely c<strong>on</strong>nected to an excessive activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> glutamate receptors,<br />

which causes a pathological calcium entrance to the cell. The generati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> reactive oxygen species, i.e. an<br />

oxidative stress c<strong>on</strong>diti<strong>on</strong>, is <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the c<strong>on</strong>sequences <str<strong>on</strong>g>of</str<strong>on</strong>g> this extreme calcium influx. There are experimental<br />

evidences about the alterati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the redox status, in patients and in animal models <str<strong>on</strong>g>of</str<strong>on</strong>g> epilepsy. However,<br />

there are few reports <str<strong>on</strong>g>of</str<strong>on</strong>g> these variables in peripheral fluids, and n<strong>on</strong>e -that we could found- reporting the<br />

redox status evaluati<strong>on</strong> after surgical treatment. We have studied the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the antioxidant enzymes:<br />

superoxide dismutase, catalase and glutathi<strong>on</strong>e peroxidase, as well as the indicators <str<strong>on</strong>g>of</str<strong>on</strong>g> damage to lipids and


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

proteins: mal<strong>on</strong>dialdehyde and advanced oxidati<strong>on</strong> protein products, in the serum <str<strong>on</strong>g>of</str<strong>on</strong>g> patients suffering from<br />

drug-resistant temporal lobe epilepsy be<str<strong>on</strong>g>for</str<strong>on</strong>g>e and at different times after surgery. Pre-surgery patients<br />

presented increases in the indicators <str<strong>on</strong>g>of</str<strong>on</strong>g> damage to lipids and proteins, and alterati<strong>on</strong>s in the activities <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

antioxidant enzymes in respect to normal c<strong>on</strong>trol subjects. Post-surgery, patients showed a trend to<br />

normalizati<strong>on</strong> in all the studied variables except <str<strong>on</strong>g>for</str<strong>on</strong>g> superoxide dismutase. We c<strong>on</strong>clude that drug-resistant<br />

temporal lobe epilepsy is associated with an oxidative stress c<strong>on</strong>diti<strong>on</strong>, which is favorably modified by<br />

removing the epileptic foci.<br />

2H_20_P<br />

(poster secti<strong>on</strong> A2, poster board #126, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PROTECTION AGAINST RADIATION INDUCED OXIDATIVE STRESS IN SWISS<br />

ALBINO MICE BY MENTHA PIPERITA (LINN)<br />

Ravindra M. Samarth*, Meenakshi Panwar, Madhu Kumar, Ashok Kumar<br />

Radiati<strong>on</strong> & Cancer Biology Laboratory, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Zoology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Rajasthan, Jaipur-302 004,<br />

INDIA<br />

Purpose: To evaluate protecti<strong>on</strong> against radiati<strong>on</strong> induced oxidative stress in Swiss albino mice by Mentha<br />

piperita (Linn).<br />

Materials and Methods: Animals (Swiss albino mice) were given either double distilled water or leaf extract <str<strong>on</strong>g>of</str<strong>on</strong>g> M.<br />

piperita orally (1 g/kg b.wt./day) <strong>on</strong>ce a day <str<strong>on</strong>g>for</str<strong>on</strong>g> three c<strong>on</strong>secutive days, after 30 min <str<strong>on</strong>g>of</str<strong>on</strong>g> treatments were<br />

exposed to 8 Gy gamma radiati<strong>on</strong>. Mice were autopsied after 30 minutes <str<strong>on</strong>g>of</str<strong>on</strong>g> post-irradiati<strong>on</strong>. Biochemical<br />

parameters such as reduced glutathi<strong>on</strong>e c<strong>on</strong>tent, glutathi<strong>on</strong>e peroxidase, glutathi<strong>on</strong>e reductase, glutathi<strong>on</strong>e<br />

S-transferase, superoxide dismutase, catalase and lipid peroxidati<strong>on</strong> were studied in liver <str<strong>on</strong>g>of</str<strong>on</strong>g> Swiss albino mice.<br />

Results: Animals pretreated with leaf extract <str<strong>on</strong>g>of</str<strong>on</strong>g> M. piperita and exposed to 8.0 Gy gamma radiati<strong>on</strong> showed a<br />

significant increases in the activities <str<strong>on</strong>g>of</str<strong>on</strong>g> reduced glutathi<strong>on</strong>e c<strong>on</strong>tent (p


23-26 August 2007,<br />

Budapest, Hungary<br />

2H_21_P<br />

(poster secti<strong>on</strong> A2, poster board #127, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE ROLE OF CATALASES FROM THE SOIL COMAMONAS TERRIGENA IN THE<br />

RESPONSE TO OXIDATIVE STRESS<br />

Jana Godočíková, Mária Bučková, Marcel Zámocký, Bystrík Polek<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular Biology, Slovak Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences, Dúbravska cesta 21, SK-84551, Bratislava, Slovakia<br />

e-mail: bystrik.polek@savba.sk<br />

We have evaluated the role <str<strong>on</strong>g>of</str<strong>on</strong>g> m<strong>on</strong><str<strong>on</strong>g>of</str<strong>on</strong>g>uncti<strong>on</strong>al heme-c<strong>on</strong>taining catalase encoded by cat-1 gene from the soil<br />

bacterium Comam<strong>on</strong>as terrigena N3H in the resp<strong>on</strong>se to various <str<strong>on</strong>g>for</str<strong>on</strong>g>ms <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidative stress. Our results indicate<br />

that this c<strong>on</strong>stitutively expressed catalase represents the major source <str<strong>on</strong>g>for</str<strong>on</strong>g> the defence <str<strong>on</strong>g>of</str<strong>on</strong>g> Comam<strong>on</strong>as terrigena<br />

cells against toxic peroxides but the cells can express also a sec<strong>on</strong>d <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> catalase that is bigger and its<br />

regulati<strong>on</strong> is probably more complicated. The sequence analysis c<strong>on</strong>firmed the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> highly c<strong>on</strong>served<br />

catalase sequence motifs in two envir<strong>on</strong>mental strains <str<strong>on</strong>g>of</str<strong>on</strong>g> C.terrigena but in those strains that were not exposed<br />

to oxidative stress no such sequence motif could be detected. Obtained results underline the importance <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

catalase expressi<strong>on</strong> in the defence mechanism against oxidative stress in bacterial cells.<br />

2H_22_P<br />

(poster secti<strong>on</strong> A2, poster board #128, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CHARACTERISATION OF THE EFFECT OF OXIDATIVE STRESS ON APOPTOSIS,<br />

NECROSIS, AND CELLULAR LOCATION AND RELEASE OF HSP70<br />

Ola M. B. Altaie, Claire Hunter-Lavin, John H. H. Williams<br />

Chester <str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g> Stress <str<strong>on</strong>g>Research</str<strong>on</strong>g>, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biological Sciences, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Chester, Parkgate Road, Chester,<br />

CH1 4BJ. UK<br />

e-mail: john.williams@chester.ac.uk<br />

Previous studies show the release <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock protein 70 (Hsp70) from healthy and necrotic cells but not<br />

from apoptotic cells. Oxidative stress is known to induce cell death by apoptosis and necrosis. We have<br />

studied the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> a range <str<strong>on</strong>g>of</str<strong>on</strong>g> c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> hydrogen peroxide and homocysteine <strong>on</strong> apoptosis and<br />

necrosis in several cell lines, al<strong>on</strong>gside measurements <str<strong>on</strong>g>of</str<strong>on</strong>g> intracellular and cell surface Hsp70 and Hsp70<br />

release. Apoptosis in Jurkat cells peaked after a 6 hour incubati<strong>on</strong> with 0.01mM H 2 O 2 , as measured using<br />

Annexin-V and Caspase-3 activity. Necrosis was measured using propidium iodide and determined to be<br />

induced after a 6 hour incubati<strong>on</strong> with 1mM H 2 O 2 . Apoptosis and necrosis were also c<strong>on</strong>firmed visually using<br />

fluorescence microscopy. These optimal c<strong>on</strong>centrati<strong>on</strong>s were subsequently used <str<strong>on</strong>g>for</str<strong>on</strong>g> the investigati<strong>on</strong> into the<br />

effects <str<strong>on</strong>g>of</str<strong>on</strong>g> H 2 O 2 <strong>on</strong> surface and intracellular Hsp70, using flow cytometry, and Hsp70 release. The effects <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

anti-oxidants such as folic acid <strong>on</strong> the above measurements were also investigated.<br />

140


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2H_23_P<br />

(poster secti<strong>on</strong> A2, poster board #129, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

MENOPAUSE AND OXIDATIVE STRESS: EFFECT OF<br />

METHYLTETRAHYDROFOLATE SUPPLEMENTATION<br />

V. Sblendorio 1 , M. Cannoletta 2 , B. Palmieri 1 , A. Cagnacci 2<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> General Surgery, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Modena, Italy<br />

2Gynecology Unit, Policlinico <str<strong>on</strong>g>of</str<strong>on</strong>g> Modena, Italy<br />

Background: Oxidative stress is an imbalance between free radicals and antioxidant molecules.<br />

Menopause is <str<strong>on</strong>g>of</str<strong>on</strong>g>ten characterized by hot flashes, brain functi<strong>on</strong> reducti<strong>on</strong>, atherosclerosis and atrophic<br />

changes <str<strong>on</strong>g>of</str<strong>on</strong>g> the skin, comprehensive ageing phenomena triggered by oestrogens blood level drop. We support<br />

the c<strong>on</strong>cepts that oxygen stress c<strong>on</strong>tributes to menopause disease and that some <str<strong>on</strong>g>of</str<strong>on</strong>g> its physiopathological<br />

effects may be prevented and/or treated improving the antioxidant defence <str<strong>on</strong>g>of</str<strong>on</strong>g> postmenopausal women.<br />

Un<str<strong>on</strong>g>for</str<strong>on</strong>g>tunately, even if many compounds have been addressed to the market with the claim <str<strong>on</strong>g>of</str<strong>on</strong>g> an, in vitro,<br />

antioxidant activity, very few molecules have been evaluated in vivo, in terms <str<strong>on</strong>g>of</str<strong>on</strong>g> clinical effectiveness<br />

compared to oxidant status instrumental improvement. In this study we evaluated if administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

methyltetrahydr<str<strong>on</strong>g>of</str<strong>on</strong>g>olate can improve oxidative status.<br />

Methods: Global evaluati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the oxidative status by FORT (Free Oxygen Radicals Testing) and FORD (Free<br />

Oxygen Radicals Defence) tests in 21 healthy postmenopausal women be<str<strong>on</strong>g>for</str<strong>on</strong>g>e and after supplementati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

15 mg <str<strong>on</strong>g>of</str<strong>on</strong>g> methyltetrahydr<str<strong>on</strong>g>of</str<strong>on</strong>g>olate <str<strong>on</strong>g>for</str<strong>on</strong>g> 3 weeks.<br />

Results: We obtained a significant decrease in FORT (p= 0.0335), increase <str<strong>on</strong>g>of</str<strong>on</strong>g> FORD (p=0.0438) and<br />

antioxidant/free radicals ratio (p= 0.0389). There was no significant difference in glucose levels be<str<strong>on</strong>g>for</str<strong>on</strong>g>e and<br />

after methyltetrahydr<str<strong>on</strong>g>of</str<strong>on</strong>g>olate administrati<strong>on</strong>, but there was a significant decrease in insulin levels (p= 0.0074)<br />

and resistance as evaluated by HOMA (p= 0.0063).<br />

C<strong>on</strong>clusi<strong>on</strong>s: Although preliminary, these results indicate that methyltetrahydr<str<strong>on</strong>g>of</str<strong>on</strong>g>olate supplementati<strong>on</strong> may<br />

reduce oxidative stress and insulin resistance <str<strong>on</strong>g>of</str<strong>on</strong>g> postmenopausal women.<br />

141


23-26 August 2007,<br />

Budapest, Hungary<br />

2I. STRESS OF MITOCHONDRIA<br />

2I_01_P<br />

(poster secti<strong>on</strong> A2, poster board #130, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CARDIOPROTECTIVE EFFECTS OF POLYPHENOLS EXTRACTED FROM<br />

PROPOLIS AGAINST DOXORUBICIN TOXICITY<br />

M. Alyane, L. Benguedouar, M. Lahouel<br />

Laboratoire de Pharmacologie, Faculté des Sciences,<br />

Université de Jijel, 18000, Algérie<br />

e-mail: alyanen@yahoo.fr<br />

Propolis (bee glue) is <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the major hive products <str<strong>on</strong>g>of</str<strong>on</strong>g> bees and is rich in flav<strong>on</strong>oids, which are known <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

antioxidant activities. It is well known that the chemical properties <str<strong>on</strong>g>of</str<strong>on</strong>g> phenolic acids or flav<strong>on</strong>oids, in terms<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the availability <str<strong>on</strong>g>of</str<strong>on</strong>g> the phenolic hydrogens as hydrogen d<strong>on</strong>ating radical scavengers, predict their<br />

antioxidant properties.<br />

In this study, the flav<strong>on</strong>oids scavenging activity <str<strong>on</strong>g>of</str<strong>on</strong>g> propolis has been exploited to obtain protecti<strong>on</strong> against<br />

the peroxidative damage in rat heart mitoch<strong>on</strong>dria which was induced by the administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> an acute dose<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> doxorubicin (DXR 20mg kg −1 , i.p). The peroxidative lesi<strong>on</strong>s were evaluated biochemically and<br />

biophysically, 24 H after DXR administrati<strong>on</strong>.<br />

Abnormal biochemical changes in heart mitoch<strong>on</strong>dria from DXR treated rats including a marked increase in<br />

both mal<strong>on</strong>dialdehyde (MDA) and ani<strong>on</strong> superoxide producti<strong>on</strong>, decrease both <str<strong>on</strong>g>of</str<strong>on</strong>g> respiratory chain ratio<br />

(RCR= V3/V4) and P/O.<br />

Biophysically, collapse <str<strong>on</strong>g>of</str<strong>on</strong>g> membrane potential <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>dria, first <str<strong>on</strong>g>of</str<strong>on</strong>g> the two steps <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>drial death<br />

pathway, and swelling, resp<strong>on</strong>sible <str<strong>on</strong>g>of</str<strong>on</strong>g> fragmentati<strong>on</strong> and massive cytochrome c release, were observed.<br />

Pretreatment <str<strong>on</strong>g>of</str<strong>on</strong>g> rats with propolis extract, given per os (100mg/kg/day) during four days prior to DXR<br />

injecti<strong>on</strong>, substantially reduced the peroxidative damage in the heart mitoch<strong>on</strong>dria: we showed significant<br />

reducing both <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>drial MDA <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> and producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> superoxide ani<strong>on</strong>, restorati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> RCR<br />

and P/O, reducing <str<strong>on</strong>g>of</str<strong>on</strong>g> rate and the amplitude <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>drial swelling and finally restorati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the loss <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the mitoch<strong>on</strong>drial membrane potential. The data dem<strong>on</strong>strate that antioxidants from natural sources may be<br />

useful in the protecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cardiotoxicity in patients who receive doxorubicin.<br />

142


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2I_03_P<br />

(poster secti<strong>on</strong> A2, poster board #131, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE STUDY OF THE DAMAGE OF MITOCHONDRION INDUCED BY PRENATAL<br />

STRESS IN OFFSPRING HIPPOCAMPAL NEURONS<br />

Liang S<strong>on</strong>g 1 , Jian-bin Zheng 3 , Hui Li 2 , Zhi-r<strong>on</strong>g Suo 3 , Qing Cai 1 , Ning Jia 1 , Zhuan-li Bai 1 , Rui Chen 2 ,<br />

Qing-h<strong>on</strong>g Li 2 , Da-xin Cheng 2 , Zh<strong>on</strong>g-liang Zhu 1,4 *<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Physiology and Pathophysiology, Xi’an Jiaot<strong>on</strong>g University, School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Key laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Envir<strong>on</strong>ment and Gene Related to Diseases (Xi’an Jiaot<strong>on</strong>g University), Ministry <str<strong>on</strong>g>of</str<strong>on</strong>g> Educati<strong>on</strong>, Xi’an, Shaanxi<br />

710061, P. R. China;<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Pediatrics, Xi’an Jiaot<strong>on</strong>g University, First Hospital, Xi’an, Shaanxi 710061, P. R. China;<br />

3Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Analytical Science Northwest University, Xi’an, Shaanxi 710069, P. R. China;<br />

4Northwest University College <str<strong>on</strong>g>of</str<strong>on</strong>g> Life Science, Xi’an, Shaanxi 710069, P. R. China<br />

Objective: In recent years, more investigator were attracted by the research about the damage <str<strong>on</strong>g>of</str<strong>on</strong>g> hippocampus<br />

induced by prenatal stress. In present study, inner mitoch<strong>on</strong>drial membrane potential (IMMP) in<br />

hippocampal neur<strong>on</strong> and the c<strong>on</strong>tent <str<strong>on</strong>g>of</str<strong>on</strong>g> 8-hydroxy-2’-deoxyguanosine (8-OH-dG) in hippocampus <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g>fspring rats were measured to make clear the changes <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>drial functi<strong>on</strong> and explore the<br />

mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> the damage <str<strong>on</strong>g>of</str<strong>on</strong>g> hippocampus induced by prenatal stress.<br />

Methods: In this experiment the <str<strong>on</strong>g>of</str<strong>on</strong>g>fspring rats were divided into two groups, CON group and PNS<br />

group(those whose dams were exposed to restraint stress <strong>on</strong> days 14-20 <str<strong>on</strong>g>of</str<strong>on</strong>g> pregnancy). Offspring (1 m<strong>on</strong>th<br />

old) were anesthetized with ether and decapitated, and hippocampus were dissected and stored <str<strong>on</strong>g>for</str<strong>on</strong>g> later<br />

experiment: The mitoch<strong>on</strong>dri<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> hippocampal neur<strong>on</strong>s were labeled with specific fluorescent probe <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Rhodamine 123 to measure IMMP by flow cytometry; The extracted and purified mitoch<strong>on</strong>drial DNA <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

hippocampal neur<strong>on</strong>s were used to measure 8-OH-dG, a typical biomarker <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidative stress, by the method<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> high per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance liquid chromatography-electrochemical detecti<strong>on</strong> (HPLC-ECD).<br />

Results: Prenatal stress significantly lowered the IMMP compared with CON group (68.437±10.680 vs<br />

124.780±7.336P


23-26 August 2007,<br />

Budapest, Hungary<br />

2I_04_P<br />

(poster secti<strong>on</strong> A2, poster board #132, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EVIDENCE OF NMN-AT ACTIVITY, WHICH ALLOWS FOR NAD + SYNTHESIS<br />

FROM NMN AND ENDOGENOUS ATP IN MITOCHONDRIA ISOLATED FROM<br />

AGED-DEHYDRATED SLICES TUBERS OF HELIANTHUS TUBEROSUS<br />

Maria Luigia Pallotta Catello Di Martino*<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Health Sciences, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Animal, Plant and Enviromental Science*.<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Molise, 86100 Campobasso, Italy<br />

e-mail: pallotta@unimol.it<br />

Pyridine dinucleotides are key redox carriers in the soluble phase <str<strong>on</strong>g>of</str<strong>on</strong>g> all living cells, and both NAD + and<br />

NADP + play crucial roles in pro-oxidant and antioxidant metabolism. The capacity <str<strong>on</strong>g>of</str<strong>on</strong>g> cells to modulate the<br />

NAD(P)H/NAD(P) + ratio is thus critical not <strong>on</strong>ly <str<strong>on</strong>g>for</str<strong>on</strong>g> central redox c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> metabolism but also <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

preemptive management <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidative stress. Recent studies show that NAD + and NADP + can be utilized to<br />

produce a number <str<strong>on</strong>g>of</str<strong>on</strong>g> metabolites important in cell signalling. Other reacti<strong>on</strong>s that c<strong>on</strong>sume NAD + include<br />

protein de-acetylati<strong>on</strong> and poly ADP-ribosylati<strong>on</strong>. These developments bring a significant amount <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

additi<strong>on</strong>al interest to the investigati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular NAD + biosynthesis and regulati<strong>on</strong>. Relatively little is known<br />

about NAD + synthesis in plants. Comparative genomics suggests that plants <str<strong>on</strong>g>for</str<strong>on</strong>g>m NAD + through de novo<br />

synthesis from quinolinate, as well as by a salvage pathway that reuses nicotinamide released from NAD + .<br />

NADP + is produced by phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> NAD + The purpose <str<strong>on</strong>g>of</str<strong>on</strong>g> our work was to verify the presence <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

mitoch<strong>on</strong>drial Nicotinamide m<strong>on</strong><strong>on</strong>ucleotide adenylyl-transferase (NMN-AT), that catalyzes the reversible<br />

reacti<strong>on</strong> NMN+ ATP NAD + + PP i NMN was added to mitoch<strong>on</strong>dria isolated from aged-dehydrated slices<br />

tubers <str<strong>on</strong>g>of</str<strong>on</strong>g> Helianthus tuberosus and NAD(P) + synthesis was tested by means <str<strong>on</strong>g>of</str<strong>on</strong>g> HPLC experiments. The<br />

dependence <str<strong>on</strong>g>of</str<strong>on</strong>g> NAD(P) + synthesis rate <strong>on</strong> NMN c<strong>on</strong>centrati<strong>on</strong> was studied. Hyperbolic dependence <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

reacti<strong>on</strong> rate <strong>on</strong> NMN c<strong>on</strong>centrati<strong>on</strong> was found and was inhibited by PPi and AMP. The characterizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

NMN-AT provided compelling evidence that NAD + biosynthesis pathways may exist in plant mitoch<strong>on</strong>dria.<br />

144


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2J. STRESS SIGNALING<br />

2J_01_P<br />

(poster secti<strong>on</strong> A2, poster board #133, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE EFFECTS OF PRENATAL STRESS ON EXPRESSION OF NF-ΚB JNK AND<br />

P38MAPK IN THE HIPPOCAMPUS OF OFFSPRING RATS<br />

Hui Li 1 , Tian-bao S<strong>on</strong>g 2 , Zh<strong>on</strong>g-liang Zhu 3* , Rui Chen 1 , Da-xin Cheng 1 , Qing Cai 2 , Ning Jia 2 ,<br />

Liang S<strong>on</strong>g 2 , Qing-h<strong>on</strong>g Li 1<br />

1 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Paediatrics, First Affiliated Hospital <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical College <str<strong>on</strong>g>of</str<strong>on</strong>g> Xi’an Jiaot<strong>on</strong>g University, Xi’an, Shaan<br />

xi 710061, China<br />

2 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Physiology and Pathophysiology, Medical College <str<strong>on</strong>g>of</str<strong>on</strong>g> Xi’an Jiaot<strong>on</strong>g University, Xi’an, Shaan xi<br />

710061, China<br />

3 College Life <str<strong>on</strong>g>of</str<strong>on</strong>g> Science Northwest University, Xi’an, Shaan xi 710069, China<br />

*Pr<str<strong>on</strong>g>of</str<strong>on</strong>g>essor Zhu Zh<strong>on</strong>g-liang, College Life <str<strong>on</strong>g>of</str<strong>on</strong>g> Science, Northwest University, Xi’an, Shaanxi 710069, China<br />

e-mail: zlzhu@mail.xjtu.edu.cn<br />

Substantive data has shown that maternal factors, like anxiety, depressi<strong>on</strong> and some negative stresses, can<br />

affect <str<strong>on</strong>g>of</str<strong>on</strong>g>fspring development. Previous studies revealed that PNS could cause impairment <str<strong>on</strong>g>of</str<strong>on</strong>g> spatial learning,<br />

inhibit cell proliferati<strong>on</strong> in the dentate gyrus and hippocampus and reduce the number <str<strong>on</strong>g>of</str<strong>on</strong>g> hippocampal<br />

neur<strong>on</strong>s in <str<strong>on</strong>g>of</str<strong>on</strong>g>fspring rats. JNK and p38MAPK are members <str<strong>on</strong>g>of</str<strong>on</strong>g> serine/thre<strong>on</strong>ine protein kinases(MAPK).<br />

MAPK not <strong>on</strong>ly regulates cell proliferati<strong>on</strong>, differentiati<strong>on</strong> and survival, but also plays important roles in<br />

synaptic plasticity and memory <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>. Nuclear factor (NF-κB) is involved in regulating cell proliferati<strong>on</strong>,<br />

differentiati<strong>on</strong> and survival and plays important roles in LTP <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> as well.<br />

In this essay, the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> NF-κB, p38MAPK and JNK in hippocampus <str<strong>on</strong>g>of</str<strong>on</strong>g> was determined by<br />

establishing the model <str<strong>on</strong>g>of</str<strong>on</strong>g> PNS in order to explore the cell signal transducti<strong>on</strong> pathways c<strong>on</strong>cerned. Animals<br />

were divided three groups: c<strong>on</strong>trol (CON), mid-term stress (MS), late term stress (LS). The pregnant rats<br />

were exposed to the restraint stress (3 times in a day, 45 minutes each time) during the mid-term or<br />

late-term gestati<strong>on</strong> stage. Western-blotting and immunohistochemistry techniques were used to examine the<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> NF-κB and p38MAPK/JNK in hippocamous.The results as follows. Both MS and LS<br />

decreased the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> p65 in female <str<strong>on</strong>g>of</str<strong>on</strong>g>fspring, and there was a significant difference between MS and<br />

LS groups (P


23-26 August 2007,<br />

Budapest, Hungary<br />

2J_02_P<br />

(poster secti<strong>on</strong> A2, poster board #134, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ONCOGENIC RAS, VIA PI3K AND MAPK PATHWAYS, TRIGGERS ENDOPLASMIC<br />

RETICULUM STRESS-INDUCED APOPTOSIS<br />

Jinjin Guo, LeaAnn Collins, Zhi-Xi<strong>on</strong>g, J. Xiao<br />

Beth Israel Deac<strong>on</strong>ess Medical Center, Radiati<strong>on</strong> Oncology<br />

Oncogenic Ras induces apoptosis up<strong>on</strong> protein kinase C (PKC) suppressi<strong>on</strong>. The mechanisms by which Ras<br />

regulates apoptosis remain unclear. In this study, we used three V12-Ha-ras effector loop mutants to dissect<br />

Ras downstream signaling in murine NIH3T3 cells and tested the roles <str<strong>on</strong>g>of</str<strong>on</strong>g> each mutant and the three possible<br />

pairwise combinati<strong>on</strong>s in the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis. The expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a single ras mutant can not activate<br />

programmed cell death after treatment with GO6976 (a PKC inhibitor). The c<strong>on</strong>current activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> PI3K<br />

and MAPK pathways increases the susceptibility <str<strong>on</strong>g>of</str<strong>on</strong>g> the cells to apoptosis in resp<strong>on</strong>se to PKC suppressi<strong>on</strong>.<br />

We also showed that although such co-activati<strong>on</strong> al<strong>on</strong>e moderately upregulates ROS producti<strong>on</strong>, c<strong>on</strong>current<br />

suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> PKC causes a dramatic elevati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ROS which leads to the accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> unfolded<br />

proteins in the endoplasmic reticulum (ER) and subsequent inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis. Thus, our study suggests<br />

that under the c<strong>on</strong>diti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> PKC abrogati<strong>on</strong>, <strong>on</strong>cogenic Ras, via the PI3K and MAPK pathways, perturbs the<br />

state <str<strong>on</strong>g>of</str<strong>on</strong>g> the cellular redox, which signals to the ER stress-regulated apoptotic machinery <str<strong>on</strong>g>for</str<strong>on</strong>g> the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

apoptosis.<br />

2J_03_P<br />

(poster secti<strong>on</strong> A2, poster board #135, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

AMYLOID Β-PROTEIN POTENTIATES TUNICAMYCIN-INDUCED NEURONAL<br />

DEATH IN ORGANOTYPIC HIPPOCAMPAL SLICE CULTURES<br />

Yoshihisa Ito, Toru Imai, Yasuhiro Kosuge, Kumiko Ishige<br />

<str<strong>on</strong>g>Research</str<strong>on</strong>g> Unit <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmacology, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Clinical Pharmacy, College <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmacy, Nih<strong>on</strong> University, 7-7-1<br />

Narashinodai, Funabashi-shi, Chiba 274-8555, Japan<br />

e-mail: yoshiito@pha.nih<strong>on</strong>-u.ac.jp<br />

We have assessed amyloid β-protein (Aβ)-induced neurotoxicity, with and without added tunicamycin (TM),<br />

an inhibitor <str<strong>on</strong>g>of</str<strong>on</strong>g> N-glycosylati<strong>on</strong> in the endoplasmic reticulum (ER), in organotypic hippocampal slice cultures<br />

(OHCs). In the OHCs cultured <str<strong>on</strong>g>for</str<strong>on</strong>g> 3 weeks, there was little neurotoxicity after treatment with Aβ 25-35 (25 µM)<br />

al<strong>on</strong>e <str<strong>on</strong>g>for</str<strong>on</strong>g> 48 h. However, with TM al<strong>on</strong>e, c<strong>on</strong>centrati<strong>on</strong>-dependent neur<strong>on</strong>al death was observed at<br />

c<strong>on</strong>centrati<strong>on</strong>s between 20 and 80 µg/mL. When Aβ was combined with TM (Aβ+TM), cell death was more<br />

acute than with TM al<strong>on</strong>e. Western blot analysis revealed that calpain activity and the active <str<strong>on</strong>g>for</str<strong>on</strong>g>ms <str<strong>on</strong>g>of</str<strong>on</strong>g> caspase-<br />

12 and caspase-3 were increased after exposure to Aβ+TM as compared with exposure to TM al<strong>on</strong>e. In<br />

c<strong>on</strong>trast, the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> GRP94, GRP78 and CHOP were not changed in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> Aβ. Aβ potentiati<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> TM neurotoxicity was reversibly blocked by S-allyl-L-cysteine (SAC), an organosulfur compound purified<br />

from aged garlic extract, and the L-type calcium channel blocker, nifedipine, in a restricted neur<strong>on</strong>al area <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the OHCs. Simultaneously applied SAC also reversed the increases in calpain activity and the active <str<strong>on</strong>g>for</str<strong>on</strong>g>ms <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

caspase-12 and caspase-3 by Aβ +TM with no change in the increased levels <str<strong>on</strong>g>of</str<strong>on</strong>g> GRP 94 and 78 and CHOP.<br />

These data indicate that Aβ facilitates the calpain-caspase-12-caspase-3 pathway, thus potentiating TMinduced<br />

neur<strong>on</strong>al death in the hippocampus.<br />

146


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2J_04_P<br />

(poster secti<strong>on</strong> A2, poster board #136, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SIGNALING EVENTS INDUCED BY STRESS FACTORS IN CEREBRAL<br />

ENDOTHELIAL CELLS<br />

I. A. Krizbai 1 , A. E. Farkas 1 , I. Wilhelm 1 , P. Nagyőszi 1 , M. Széll 2 , P. O. Couraud 3 , B. Wechsler 3 ,<br />

I. A. Romero 4 , Z. Bálint 1 , G. Váró 1 ,<br />

1Biological <str<strong>on</strong>g>Research</str<strong>on</strong>g> <str<strong>on</strong>g>Centre</str<strong>on</strong>g>, Szeged, Hungary;<br />

2University <str<strong>on</strong>g>of</str<strong>on</strong>g> Szeged, Szeged, Hungary,<br />

3Institut Cochin, Paris, France;<br />

4The Open University, Milt<strong>on</strong> Keynes, UK<br />

Being located at the interface <str<strong>on</strong>g>of</str<strong>on</strong>g> blood and brain cerebral endothelial cells (CECs) are primary targets <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

different envir<strong>on</strong>mental stimuli. In our study we made an attempt to reveal novel elements <str<strong>on</strong>g>of</str<strong>on</strong>g> signaling<br />

activated by hyperosmotic, oxidative and calcium depleti<strong>on</strong> induced stress in CECs. Hyperosmotic stress<br />

elicited by mannitol has been successfully used to reversibily open the blood-brain barrier. We have shown<br />

that hyperosmotic c<strong>on</strong>diti<strong>on</strong>s induce protein phosphorylati<strong>on</strong> <strong>on</strong> both Ser/Thr and Tyr residues. Am<strong>on</strong>g the<br />

targets <str<strong>on</strong>g>of</str<strong>on</strong>g> protein tyrosine phosphorylati<strong>on</strong> are beta-catenin and possibly the ezrin/moesin complex.<br />

Furthermore, our results suggest that besides src the receptor tyrosine kinase Axl plays an important role in<br />

mediating the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> hyperosmosis. Activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Rho signaling my be involved in the stress resp<strong>on</strong>se<br />

induced by calcium depleti<strong>on</strong>. We have shown that besides changes in juncti<strong>on</strong>al protein expressi<strong>on</strong> and<br />

localizati<strong>on</strong> calcium removal induces significant changes in the morphological parameters <str<strong>on</strong>g>of</str<strong>on</strong>g> CECs as well as<br />

revealed by atomic <str<strong>on</strong>g>for</str<strong>on</strong>g>ce microscopy. These changes could be partially inhibited by the Rho-dependent kinase<br />

inhibitor Y27632 suggesting a role <str<strong>on</strong>g>for</str<strong>on</strong>g> ROCK in mediating the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> low calcium c<strong>on</strong>centrati<strong>on</strong> in CECs.<br />

Recent research has revealed that n<strong>on</strong>-coding RNAs are able to fulfill a wide range <str<strong>on</strong>g>of</str<strong>on</strong>g> regulatory functi<strong>on</strong>s in<br />

eukaryotic cells. In our studies we have observed, that the n<strong>on</strong>-coding RNA PRINS (Psoriasis Susceptibility-<br />

Related RNA Gene Induced by Stress) is rapidly upregulated severalfold by oxidative stress induced by<br />

DMNQ. Further studies are underway to elucidate the role <str<strong>on</strong>g>of</str<strong>on</strong>g> PRINS in the regulati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> endothelial<br />

processes. Our results suggest that multiple signaling pathways are activated in by different stress factors in<br />

CECs.<br />

2J_05_P<br />

(poster secti<strong>on</strong> A2, poster board #137, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE OXIDATION OF ASK1 AND ITS REDUCTION BY THIOREDOXIN-1<br />

REGULATE THE INDUCTION OF JNK AND APOPTOSIS BY H2O2<br />

Philippe. J. Nadeau, Steve. J. Charrette, Jacques Landry<br />

<str<strong>on</strong>g>Centre</str<strong>on</strong>g> de recherche en cancérologie de l’Université Laval, L’Hôtel-Dieu de Québec, Québec, Canada, G1R 2J6,<br />

e-mail: jacques.landry@med.ulaval.ca<br />

The oxidative stress resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> lower organisms is mediated by “redox sensors”. They c<strong>on</strong>sist in<br />

transcripti<strong>on</strong> factors or oxidases <str<strong>on</strong>g>of</str<strong>on</strong>g> transcripti<strong>on</strong> factors which, when activated by specific oxidati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

cysteine residus, induce an antioxidant resp<strong>on</strong>se. In mammals, the oxidative stress resp<strong>on</strong>se involves the<br />

activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> signaling pathways that induce many biological resp<strong>on</strong>ses including apoptosis. Apoptosis signal<br />

regulated kinase-1 (Ask1) lies upstream <str<strong>on</strong>g>of</str<strong>on</strong>g> a major redox-sensitive pathway involving Jun N-terminal kinase<br />

(JNK). We found that cell exposure to H 2 O 2 caused the rapid oxidati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Ask1 leading to its<br />

147


23-26 August 2007,<br />

Budapest, Hungary<br />

multimerizati<strong>on</strong> through the <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> interchain disulfide b<strong>on</strong>ds. Oxidized Ask1 was fully reduced within<br />

minutes after inducti<strong>on</strong> by H 2 O 2 . During this reducti<strong>on</strong>, the thiol-disulphide oxidoreductase thioredoxin-1<br />

(Trx1) became covalently associated with Ask1. Overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Trx1 accelerated the reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Ask1<br />

and a redox-inactive mutant <str<strong>on</strong>g>of</str<strong>on</strong>g> Trx1 (C35A) remained trapped with Ask1 blocking its reducti<strong>on</strong>. Preventing<br />

the oxidati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Ask1 either by overexpressing Trx1 or using an Ask1 mutant in which the sensitive cysteines<br />

were mutated (Ask1-∆Cys) impaired the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> JNK but not the phoshorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Ask1 by H 2 O 2 .<br />

Furthermore, oxidati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Ask1 was required <str<strong>on</strong>g>for</str<strong>on</strong>g> H 2 O 2 -induced Ask1-mediated apoptosis. Those results<br />

indicate that oxidati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Ask1 is not required <str<strong>on</strong>g>for</str<strong>on</strong>g> its own phosphorylati<strong>on</strong>/activati<strong>on</strong> but is essential <str<strong>on</strong>g>for</str<strong>on</strong>g> its<br />

functi<strong>on</strong> upstream <str<strong>on</strong>g>of</str<strong>on</strong>g> the H 2 O 2 -induced JNK signaling pathway. We suggest that Ask1 acts as a “redox<br />

sensor” in mammalian cells and we propose a new model <str<strong>on</strong>g>for</str<strong>on</strong>g> its activati<strong>on</strong> mechanism by H 2 O 2 .<br />

2J_06_P<br />

(poster secti<strong>on</strong> A2, poster board #138, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE TYROSINE KINASE C-ABL MEDIATES HEAT SHOCK ACTIVATION OF THE<br />

P38 AND JNK PATHWAYS<br />

Sébastien Ian Nadeau, Herman Lambert, Jacques Landry<br />

<str<strong>on</strong>g>Centre</str<strong>on</strong>g> de recherche du CHUQ-L'Hôtel-Dieu de Québec, Université Laval, Québec, Canada, GIR 2J6,<br />

e-mail: jacques.landry@med.ulaval.ca<br />

Heat shock (HS) rapidly induces the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> numerous signaling pathways. The triggering switches <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these pathways and their functi<strong>on</strong>s in the HS resp<strong>on</strong>se are still poorly understood. In this<br />

study, we show that their inducti<strong>on</strong> correlates with an increase in the total c<strong>on</strong>tent <str<strong>on</strong>g>of</str<strong>on</strong>g> phosphotyrosines.<br />

Accordingly, the c-Src and c-Abl tyrosine kinases (TK) are activated by HS. To evaluate the possible<br />

involvement <str<strong>on</strong>g>of</str<strong>on</strong>g> these TK, we analyzed the HS inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the p38 and JNK pathways after pretreatment<br />

with different TK inhibitors. Our results notably show that STI571, a TK drug which inhibits c-Abl and<br />

PDGFr, greatly reduces the HS activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the p38 and JNK pathways, but has no effect <strong>on</strong> H 2 O 2 -mediated<br />

p38 activati<strong>on</strong>. By using a specific inhibitor <str<strong>on</strong>g>of</str<strong>on</strong>g> PDGFr and siRNA <str<strong>on</strong>g>for</str<strong>on</strong>g> either PDGFr or c-Abl, we show that<br />

c-Abl and not PDGFr is involved. Moreover, the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a specific siRNA-resistant plasmid encoding<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> c-Abl rescues the HS activati<strong>on</strong>. Since an important fracti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> c-Abl is nuclear, we initially suggested that<br />

the HS signal might arise from the nucleus. This was further supported by the observati<strong>on</strong> that HS activates<br />

ATM, a known upstream activator <str<strong>on</strong>g>of</str<strong>on</strong>g> c-Abl, and induces H2AX phosphorylati<strong>on</strong>. However, cells deficient in<br />

ATM or in both DNA-PK and ATM were still pr<str<strong>on</strong>g>of</str<strong>on</strong>g>icient in HS-induced p38 activati<strong>on</strong>, whereas they could<br />

not support HS-induced H2AX phosphorylati<strong>on</strong>. From these last results, we infer that the HS signal instead<br />

comes from the cytoplasmic c-Abl. Indeed, the existence <str<strong>on</strong>g>of</str<strong>on</strong>g> an actin binding domain in c-Abl hints at a<br />

possible link between the early HS disrupti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> actin filaments and the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the p38 and JNK<br />

pathways.<br />

148


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2J_07_P<br />

(poster secti<strong>on</strong> A2, poster board #139, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE HSP40 CHAPERONE MDG1/ERDJ4 IS A NOVEL MODULATOR OF THE<br />

INTEGRATED STRESS RESPONSE<br />

M. El-Hadi, L. Merkel, F. Pröls<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Freiburg, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Anatomy and Cell Biology II, D-79104 Freiburg, Germany<br />

The Hsp40 chaper<strong>on</strong>e Mdg1/ERdj4 is localized in the ER compartment and is upregulated in cells subjected<br />

to various stresses like heat shock, hypoxia and ER stress. Recently it has been shown that expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

Mdg1 hamster homolog is significantly decreased in a high metastatic cell line in comparis<strong>on</strong> to a low<br />

metastatic cell line. Overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Mdg1 in these cell lines led to a cell cycle arrest. In this work we show<br />

that Mdg1 is largely upregulated in a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> tumors. As a target <str<strong>on</strong>g>of</str<strong>on</strong>g> the IRE1α-XBP1-pathway, Mdg1 is<br />

c<strong>on</strong>sidered to play a role in the sustained Unfolded Protein Resp<strong>on</strong>se (UPR), in which protein degradati<strong>on</strong>,<br />

cell cycle arrest or even apoptosis are induced. The investigati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the UPR signaling cascade revealed<br />

CHOP, which is part <str<strong>on</strong>g>of</str<strong>on</strong>g> the Integrated Stress Resp<strong>on</strong>se (ISR), as a downstream target <str<strong>on</strong>g>of</str<strong>on</strong>g> Mdg1. We further<br />

show that Mdg1 protein has the ability to interact with ATF4 protein indicating that CHOP transcripti<strong>on</strong><br />

might be specifically induced by Mdg1-ATF4 heterodimers. Mdg1 protein thus links the signaling pathways<br />

during sustained UPR with the ISR. Together with the findings that ATF4, CHOP and Mdg1 are highly<br />

expressed in tumor cells we c<strong>on</strong>clude that ATF4-Mdg1 heterodimers lead to cell cycle arrest via CHOP<br />

upregulati<strong>on</strong> thereby reducing the risk <str<strong>on</strong>g>of</str<strong>on</strong>g> metastasis <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>.<br />

2J_08_P<br />

(poster secti<strong>on</strong> A2, poster board #140, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE NEUTRAL SPHINGOMYELINASE IN THE HEAT-INDUCED APOPTOSIS<br />

Takeshi Yabu, Michiaki Yamashita<br />

Nati<strong>on</strong>al <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Fisheries Science<br />

A neutral sphingomyelinase (SMase) is c<strong>on</strong>sidered a major candidate <str<strong>on</strong>g>for</str<strong>on</strong>g> mediating stress inducible ceramide<br />

producti<strong>on</strong>. Here, we identified and characterized the neutral SMase that is activated and generated ceramide<br />

under stress c<strong>on</strong>diti<strong>on</strong>s. The enzyme was found to be a regulator <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis mediated by heat stress. A<br />

zebrafish cDNA cl<strong>on</strong>e resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> Mg 2+ -dependent neutral SMase activity by the expressi<strong>on</strong> cl<strong>on</strong>ing<br />

system was isolated. The cDNA cl<strong>on</strong>e encoded a polypeptide <str<strong>on</strong>g>of</str<strong>on</strong>g> 420 amino acid residues (putative molecular<br />

mass: 46.9 kDa) c<strong>on</strong>taining predicted two transmembrane domains at the C-terminal regi<strong>on</strong>. The bacterially<br />

expressed recombinant neutral SMase hydrolyzed a [choline-mythyl- 14 C]sphingomyelin optimally at pH 7.5 in<br />

dependent <strong>on</strong> Mg 2+ i<strong>on</strong>. The loss <str<strong>on</strong>g>of</str<strong>on</strong>g> neutral SMase functi<strong>on</strong> with anti-sense phosphothiate olig<strong>on</strong>ucleotides<br />

inhibited ceramide generati<strong>on</strong>, caspase-3 activati<strong>on</strong> and apoptotic cell death under the heat stress c<strong>on</strong>diti<strong>on</strong>s,<br />

indicating that the neutral SMase induces a ceramide mediated pro-apoptotic signaling pathway that executes<br />

the heat-induced apoptosis.<br />

149


23-26 August 2007,<br />

Budapest, Hungary<br />

2J_09_P<br />

(poster secti<strong>on</strong> A2, poster board #141, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE ACTIVATION AND REGULATION OF CA-CAM SIGNALLING SYSTEM ON<br />

THE ACTIVITY OF CA 2+ -ATPASE AND H + -ATPASE IN LEAVES OF LONGAN<br />

UNDER STIMULATED ACID RAIN STRESS<br />

Qiu D<strong>on</strong>gliang, Pan Tengfei, Li Y<strong>on</strong>gyu, Zheng Shan, Ma Cuilan<br />

College <str<strong>on</strong>g>of</str<strong>on</strong>g> Horticulture, Fujian Agriculture and Forestry University, Fuzhou 350002 P.R.China<br />

L<strong>on</strong>gan seedlings (Dimocarpus l<strong>on</strong>gana Lour. cv. Wul<strong>on</strong>gling) <str<strong>on</strong>g>of</str<strong>on</strong>g> <strong>on</strong>e-year age grown in pots were selected to<br />

study the activati<strong>on</strong> and regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Ca-CaM signalling system <strong>on</strong> the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> Ca 2+ -ATPase and H + -<br />

ATPase in leaves <str<strong>on</strong>g>of</str<strong>on</strong>g> l<strong>on</strong>gan under stimulated acid rain stress. Sulfuric acid and nitric acid were selected <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

preparati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> artificial acid rain, based <strong>on</strong> the mole rati<strong>on</strong> 1:5 <str<strong>on</strong>g>of</str<strong>on</strong>g> sulfuric acid to nitric acid in the<br />

precipitati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Southern Fujian, China. Diluti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> reagent grade acid was d<strong>on</strong>e with distilled water and<br />

determined by PHSJ-4 acidity analyzer. Three soluti<strong>on</strong> with pH5.6 (C<strong>on</strong>trol),pH3.5 and pH2.5 were prepared.<br />

A sprayer was used to apply the acid soluti<strong>on</strong>s and 10mmol Ca(NO 3 ) 2 , 2mmol/L CPZ and 5mmol/L EGTA<br />

to young trees. The study focused <strong>on</strong> the resp<strong>on</strong>ses <str<strong>on</strong>g>of</str<strong>on</strong>g> the c<strong>on</strong>tent <str<strong>on</strong>g>of</str<strong>on</strong>g> CaM, the activities <str<strong>on</strong>g>of</str<strong>on</strong>g> H + -ATPase and<br />

Ca 2+ -ATPase in leaves <str<strong>on</strong>g>of</str<strong>on</strong>g> l<strong>on</strong>gan to simulated acid rain stress, with an aim to study the exterior Ca 2+<br />

regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the impair by acid rain, and to probe into the mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> receiving and transferring the stress<br />

signals. The main results <str<strong>on</strong>g>of</str<strong>on</strong>g> the study were as following:<br />

The c<strong>on</strong>tent <str<strong>on</strong>g>of</str<strong>on</strong>g> calmodulin (CaM) was influenced by the intensity and durati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> simulated acid rain. The<br />

CaM c<strong>on</strong>tent fluctuated during stress, but the fluctuating amplitude decreased gradually while stress durati<strong>on</strong><br />

prol<strong>on</strong>ged. The activities <str<strong>on</strong>g>of</str<strong>on</strong>g> H + -ATPase and Ca 2+ -ATPase were also influenced by stress intensity and<br />

durati<strong>on</strong>. Both <str<strong>on</strong>g>of</str<strong>on</strong>g> Ca 2+ -ATPase and H + -ATPase in leaves were very sensitive to acid rain, and their activities<br />

were significantly inhibited under the acid rain stress <str<strong>on</strong>g>of</str<strong>on</strong>g> pH 3.5 and pH 2.5. The decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> H + -ATPase<br />

activities was caused by the breakage <str<strong>on</strong>g>of</str<strong>on</strong>g> Ca-CaM signal system under the acid rain stress.<br />

The CaM c<strong>on</strong>tent, activities <str<strong>on</strong>g>of</str<strong>on</strong>g> Ca 2+ -ATPase and H + -ATPase in leaves were activated by spraying 10 mmol/L<br />

Ca 2 , and reduced by spraying calmodulin inhibitor CPZ with c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 2 mmol/L, and the calcium<br />

inhibitor EGTA with 5 mmol/L. The results showed that the Ca-CaM signal system transferred the acid rain<br />

stress signals, the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> Ca 2+ -ATPase and H + -ATPase were activated and regulated by CaM, and the<br />

exterior Ca 2 could promote the signal transfer.<br />

150


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2K. GENE EXPRESSION REGULATION UNDER STRESS<br />

2K_01_P<br />

(poster secti<strong>on</strong> A2, poster board #142, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

REGULATION OF FOX GENES IN T LYMPHOCYTES BY NOREPINEPHRINE AND<br />

CAMP<br />

Daniel H. Davis, Lloyd Graf Jr., Rh<strong>on</strong>na L. Cohen, D<strong>on</strong>ald A. Chambers<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Illinois College <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, 1853 W. Polk St., Chicago IL, 60612, USA<br />

e-mail: d<strong>on</strong>c@uic.edu<br />

Catecholamines, e.g., norepinephrine (NE) have been identified as psychogenic stressors and associated with<br />

altered immune functi<strong>on</strong>. We have identified a norepinephrine/ß2-adrenergic receptor/cAMP/PKAdependent<br />

mRNA decay system in T lymphocytes. The Fox family <str<strong>on</strong>g>of</str<strong>on</strong>g> transcripti<strong>on</strong> factors has been<br />

implicated in regulating lymphocyte physiology. Foxm1b is reported to be a master gene related to cellular<br />

proliferati<strong>on</strong> and Foxo1 has been associated with the suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proliferati<strong>on</strong> and promoti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

apoptosis. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, we explored whether a PKA-selective cAMP analogue (8Br-cAMP) and NE could<br />

regulate BALB/c murine thymocyte cell lines, S49wt and S49kin- (PKA deficient), by affecting the genetic<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Foxm1b and Foxo1. Real-time PCR studies revealed a 2-3 fold upregulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Foxo1 mRNA<br />

in S49wt, but not kin- cells with 150uM NE or 600uM cAMP as early as 2 hrs after treatment. Foxm1b<br />

mRNA was downregulated in S49wt cells by greater than 2 fold by 8Br-cAMP after 6 hrs (at 24 hrs).<br />

Treatment with an Epac-selective cAMP (500uM) analogue resulted in no change in Foxo1 or Foxm1b<br />

expressi<strong>on</strong> in S49 wt or S49 kin- cells. These results associate PKA in modulating both increased Foxo1<br />

mRNA expressi<strong>on</strong> and decreased Foxm1b expressi<strong>on</strong> and c<strong>on</strong>sequent fox gene-mediated regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

lymphocyte physiology. Supported by NIH.<br />

2K_03_P<br />

(poster secti<strong>on</strong> A2, poster board #143, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STUDIES ON GLUTATHIONE S-TRANSFERASE ACTIVITIES AND GENE<br />

EXPRESSION LEVELS IN TRITICUM AESTIVUM CULTIVARS DURING<br />

POLYETHYLENE GLYCOL-INDUCED OSMOTIC STRESS<br />

Ágnes Gallé 1 , Jolán Csiszár 1 , Mária Secenji 2 , Irma Tari 1 , Adrienn Guóth 1 , Dénes Dudits 2 ,<br />

János Györgyey 2 , László Erdei 1<br />

1 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Physiology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Szeged, PO Box 654, H-6701 Szeged, Hungary<br />

2 Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Biology, Biological <str<strong>on</strong>g>Research</str<strong>on</strong>g> Center, Szeged, Hungary<br />

e-mail: gallea@bio.u-szeged.hu<br />

We investigated the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> osmotic stress <strong>on</strong> two drought tolerant wheat cultivars: Triticum aestivum cv.<br />

Kobomugi is a near isohydric Asian landrace, T.a. cv. Öthalom is an anisohydric, dehydati<strong>on</strong> tolerating<br />

Hungarian genotype. Osmotic stress treatment was applied gradually by polyethylene glycol (PEG 6000)<br />

treatment reaching 400 mOsm (-0.976 MPa) <strong>on</strong> <strong>on</strong>e-week-old plants under c<strong>on</strong>trolled c<strong>on</strong>diti<strong>on</strong>s. Changes in<br />

glutathi<strong>on</strong>e S-transferase (GST) and glutathi<strong>on</strong>e peroxidase (GSPX) activities and in their expressi<strong>on</strong> levels<br />

were determined during a 2-week-period as functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> time. GST isoenzymes represent a large and variable<br />

group <str<strong>on</strong>g>of</str<strong>on</strong>g> antioxidative enzymes, with several different activities and sequence patterns. Phylogenetic analysis<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> wheat GSTs was per<str<strong>on</strong>g>for</str<strong>on</strong>g>med in silico and using the tentative c<strong>on</strong>sensus sequences (TC) a dendogram was<br />

151


23-26 August 2007,<br />

Budapest, Hungary<br />

c<strong>on</strong>structed. According to the c<strong>on</strong>served sequences used <str<strong>on</strong>g>for</str<strong>on</strong>g> classificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> GST proteins, we could identify<br />

four groups <str<strong>on</strong>g>of</str<strong>on</strong>g> wheat GSTs (phi, zeta, theta and tau). Real Time PCR analysis with two group-specific primer<br />

showed a significant increase in the amount <str<strong>on</strong>g>of</str<strong>on</strong>g> GST transcripts two days after plants were exposed to<br />

400 mOsm PEG. The changes in the transcript levels were compared with the GST activities measured in the<br />

same times. According to our results the members <str<strong>on</strong>g>of</str<strong>on</strong>g> phi GSTs can be resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> the fast resp<strong>on</strong>se after<br />

the osmotic stress, while the tau GST group <str<strong>on</strong>g>for</str<strong>on</strong>g> the later enhancement <str<strong>on</strong>g>of</str<strong>on</strong>g> GST activity.<br />

2K_05_P<br />

(poster secti<strong>on</strong> A2, poster board #144, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

POST-TRANSCRIPTIONAL REGULATION OF ALTERED HEAT SHOCK RESPONSE<br />

IN LONG LIVED C. ELEGANS<br />

Gawain McColl, Aric Rogers, Alan E. Hubbard, Christopher D. Link, Silvestre Alavez,<br />

Pankaj Kapahi, Gord<strong>on</strong> J. Lithgow<br />

Buck Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Age <str<strong>on</strong>g>Research</str<strong>on</strong>g>, Novato USA, e-mail: gmccoll@buckinstitute.org<br />

The insulin-like signaling (ILS) pathway plays a major role in the modulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> lifespan in C. elegans. Altered<br />

stress resp<strong>on</strong>ses, HSP expressi<strong>on</strong> and resistance to a wide range <str<strong>on</strong>g>of</str<strong>on</strong>g> stressors are also associated with changes<br />

in ILS. The causal relati<strong>on</strong>ship <str<strong>on</strong>g>of</str<strong>on</strong>g> these phenotypes with altered aging is yet to be fully determined. The<br />

transcripti<strong>on</strong> factors DAF-16 and HSF-1 have both been purported as key regulators <str<strong>on</strong>g>for</str<strong>on</strong>g> these effects. Here<br />

we present new data that dem<strong>on</strong>strates that increased resistance to heat shock, due to lowered ILS, is not<br />

dependent <strong>on</strong> a transcripti<strong>on</strong>al resp<strong>on</strong>se, but does still require active translati<strong>on</strong>. Furthermore, we describe<br />

new genes under post-transcripti<strong>on</strong>al regulati<strong>on</strong> in l<strong>on</strong>g-lived ILS mutants following heat shock via a genome<br />

wide translati<strong>on</strong>al state array analysis (TSAA).<br />

2K_06_P<br />

(poster secti<strong>on</strong> A2, poster board #145, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

REGULATION OF THE NF-κB PATHWAY BY LOW LEVELS OF H2O2 AND IMPACT<br />

ON INFLAMMATION<br />

V. Oliveira-Marques 1* , L. Cyrne 1 , H. S. Marinho 1 , F. Antunes 1,2<br />

1Grupo de Bioquímica dos Oxidantes e Antioxidantes, Centro de Química e Bioquímica, Faculdade de Ciências da<br />

Universidade de Lisboa, Portugal<br />

2Instituto Bento Da Rocha Cabral, Lisboa, Portugal<br />

*e-mail: vmmarques@fc.ul.pt<br />

During the inflammatory resp<strong>on</strong>se, host cells are exposed to pro-inflammatory cytokines, such as TNF-α,<br />

and also hydrogen peroxide (H 2 O 2 ). It is known that the cytokines liberated by neutrophils and macrophages<br />

induce the NF-κB transcripti<strong>on</strong> factor in the surrounding cells thus promoting an inflammatory resp<strong>on</strong>se.<br />

MCF-7 and HeLa cells were exposed to moderate doses <str<strong>on</strong>g>of</str<strong>on</strong>g> H 2 O 2 using the steady-state approach: [H 2 O 2 ] ss . It<br />

is a c<strong>on</strong>trolled and calibrated method that allows applying lower c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> H 2 O 2 than those applied<br />

with the bolus approach, because H 2 O 2 c<strong>on</strong>sumpti<strong>on</strong> by cells is compensated by the c<strong>on</strong>tinuous producti<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> H 2 O 2 by glucose oxidase. TNF-α (0.37 ng/mL) and [H 2 O 2 ] ss (25 µM) acted synergistically to induce NFκB<br />

translocati<strong>on</strong> to the nucleus, while an antag<strong>on</strong>ism was observed when H 2 O 2 was delivered as a bolus<br />

152


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

additi<strong>on</strong> (1 mM). The differences in the results might be explained with the high oxidative level caused by the<br />

bolus additi<strong>on</strong>, which is far from the physiological c<strong>on</strong>diti<strong>on</strong>s and can disrupt cellular redox homeostasis. The<br />

synergistic translocati<strong>on</strong> to the nucleus, with [H 2 O 2 ] ss (12.5 µM) and TNF-α <str<strong>on</strong>g>for</str<strong>on</strong>g> 6 h, led to an up-regulati<strong>on</strong> in<br />

the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> some NF-κB-dependent genes with a pro-inflammatory role (e.g. IL-8, MCP-1, TNF-α),<br />

but also anti-inflammatory genes, such as Heme oxygenase-1 and IL-6. We propose a regulatory role <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

H 2 O 2 during inflammati<strong>on</strong>, through the up-regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> pro-inflammatory genes, that<br />

stimulate the inflammatory resp<strong>on</strong>se, and anti-inflammatory genes that prevent an excessive build up <str<strong>on</strong>g>of</str<strong>on</strong>g> a<br />

pro-inflammatory envir<strong>on</strong>ment.<br />

2K_07_P<br />

(poster secti<strong>on</strong> A2, poster board #146, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PKCα -DEPENDENT PHOSPHORYLATION OF THE MRNA STABILIZING<br />

FACTOR HUR: IMPLICATIONS FOR POSTTRANSCRIPTIONAL REGULATION OF<br />

CYCLOOXYGENASE-2<br />

Anke Doller * , Andrea Huwiler* † , Roswitha Müller * , Heinfried H. Radeke * , Wolfgang Eberhardt * ,<br />

Josef Pfeilschifter *<br />

* Pharmazentrum Frankfurt/ZAFES, Klinikum der Johann Wolfgang Goethe-Universität,<br />

Frankfurt am Main, Germany<br />

†Institut für Pharmakologie, Universität Bern, CH-3010 Bern, Switzerland<br />

In this study, we investigated the molecular mechanisms underlying the ATP analog adenosine 5´O-<br />

(thiotriphosphate) (ATPγS)-induced nucleo-cytoplasmic shuttling <str<strong>on</strong>g>of</str<strong>on</strong>g> the mRNA stabilizing factor HuR in<br />

human mesangial cells (hMC). Using synthetic protein kinase C (PKC) inhibitors and siRNA approaches we<br />

dem<strong>on</strong>strate that knock-down <str<strong>on</strong>g>of</str<strong>on</strong>g> PKCα efficiently blocked the ATP-dependent nuclear HuR export to the<br />

cytoplasm. The functi<strong>on</strong>al importance <str<strong>on</strong>g>of</str<strong>on</strong>g> PKCα in HuR shuttling is highlighted by the high cytosolic HuR<br />

c<strong>on</strong>tent detected in hMC stably overexpressing PKCα when compared to mock-transfected cells. The ATPinduced<br />

recruitment <str<strong>on</strong>g>of</str<strong>on</strong>g> HuR to the cytoplasm is preceded by a direct interacti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> PKCα with nuclear HuR<br />

and accompanied by increased Ser-phosphorylati<strong>on</strong> as dem<strong>on</strong>strated by co-immunoprecipitati<strong>on</strong><br />

experiments. Mapping <str<strong>on</strong>g>of</str<strong>on</strong>g> putative PKC target sites identified serines 158 and 221 being indispensable <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

HuR-phosphorylati<strong>on</strong> by PKCα . RNA pull-down assay and RNA EMSA dem<strong>on</strong>strated that the HuR<br />

shuttling by ATP is accompanied by an increased HuR-binding to COX-2 mRNA. Physiologically, the ATPdependent<br />

increase in RNA binding is linked with an augmentati<strong>on</strong> in COX-2 mRNA stability and<br />

subsequent increase in PGE 2 synthesis. Regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HuR via PKCα-dependent phosphorylati<strong>on</strong><br />

emphasizes the importance <str<strong>on</strong>g>of</str<strong>on</strong>g> posttranslati<strong>on</strong>al modificati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> stimulus-dependent HuR shuttling.<br />

153


23-26 August 2007,<br />

Budapest, Hungary<br />

154<br />

2K_08_P<br />

(poster secti<strong>on</strong> A2, poster board #147, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ZBP1 REGULATES MRNA STABILITY DURING CELLULAR STRESS<br />

Nadine Stöhr, Stefan Hüttelmaier<br />

NBL3 <str<strong>on</strong>g>Research</str<strong>on</strong>g> Group, Zentrum für Angewandte Medizinische und Humanbiologische Forschung,<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Martin-Luther-University, 06120 Halle, Germany<br />

An essential c<strong>on</strong>stituent <str<strong>on</strong>g>of</str<strong>on</strong>g> the integrated stress resp<strong>on</strong>se (ISR) is a reversible translati<strong>on</strong>al suppressi<strong>on</strong>. This<br />

mRNA silencing occurs in distinct cytoplasmic foci called stress granules (SGs) that transiently associate with<br />

processing bodies (PBs) typically serving as mRNA decay centers. How mRNAs are protected from<br />

degradati<strong>on</strong> in these structures and if SGs serve a physiological significant role remains largely elusive. In<br />

recent studies we identified that the Zipcode-binding protein 1 (ZBP1) regulates the cytoplasmic fate <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

specific mRNAs in n<strong>on</strong>-stressed cells, but also is a key regulator <str<strong>on</strong>g>of</str<strong>on</strong>g> mRNA turnover during the ISR. The<br />

associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ZBP1 with target mRNAs in SGs was not essential <str<strong>on</strong>g>for</str<strong>on</strong>g> mRNA-targeting to SGs. However,<br />

ZBP1 knock down induced a selective destabilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> target mRNAs during the ISR, while <str<strong>on</strong>g>for</str<strong>on</strong>g>ced<br />

expressi<strong>on</strong> increased mRNA stability. Our results indicate that although targeting <str<strong>on</strong>g>of</str<strong>on</strong>g> mRNAs to SGs is<br />

n<strong>on</strong>specific, the stabilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> mRNAs during cellular stress requires specific protein-mRNA interacti<strong>on</strong>s.<br />

These retain mRNAs in SGs and prevent premature decay in PBs or by the exosome. Hence, RNA-binding<br />

proteins (RBPs) are essential <str<strong>on</strong>g>for</str<strong>on</strong>g> translati<strong>on</strong>al adaptati<strong>on</strong> during cellular stress by modulating mRNA<br />

turnover.<br />

Based <strong>on</strong> our resent observati<strong>on</strong>s we establish a new stress-based screening procedure <str<strong>on</strong>g>for</str<strong>on</strong>g> RNA ligands and<br />

thereby identify molecular networks facilitating ZBP1 functi<strong>on</strong> in SGs. Moreover, we are interested in the<br />

physiological role <str<strong>on</strong>g>of</str<strong>on</strong>g> SGs intending to set the stage <str<strong>on</strong>g>for</str<strong>on</strong>g> the identificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> SGs in cancer cells and primary<br />

tumors.<br />

2K_09_P<br />

(poster secti<strong>on</strong> A2, poster board #148, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

GENE EXPRESSION AND REGULATION OF HEAT SHOCK PROTEINS IN<br />

TRYPANOSOMA CRUZI<br />

D. R. de Carvalho, R. A. Campos, M. Fernandes, R. Silva, E. R<strong>on</strong>dinelli, T. P. Ürményi<br />

Instituto de Bi<str<strong>on</strong>g>of</str<strong>on</strong>g>ísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, CCS, Bl. G, Cidade<br />

Universitária, Rio de Janeiro, RJ 21941-902, Brazil, e-mail: turmenyi@bi<str<strong>on</strong>g>of</str<strong>on</strong>g>.ufrj.br<br />

The understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> gene regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> molecular chaper<strong>on</strong>es will shed light <strong>on</strong> the mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> posttranscripti<strong>on</strong>al<br />

regulati<strong>on</strong> in trypanosomatids. The gene organizati<strong>on</strong> and gene expressi<strong>on</strong> pattern <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70,<br />

HSP60 and HSP10 genes <str<strong>on</strong>g>of</str<strong>on</strong>g> Trypanosoma cruzi have been previously characterized by our group and others,<br />

and characterizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> gene structure and expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ClpA/HSP100 are currently under way. Our aim is<br />

to study the mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> gene regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these proteins, and to that end the presence and functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

heat shock-resp<strong>on</strong>sive elements in the corresp<strong>on</strong>ding mRNAs are being investigated. Plasmids c<strong>on</strong>taining<br />

the chloranfenicol acetyltransferase (CAT) reporter gene under the c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> the 18S rRNA promoter were<br />

c<strong>on</strong>structed in which the CAT gene is flanked by segments <str<strong>on</strong>g>of</str<strong>on</strong>g> intergenic regi<strong>on</strong>s c<strong>on</strong>taining either the 5' or 3'<br />

UTR <str<strong>on</strong>g>of</str<strong>on</strong>g> the HSP70 and Rab7 mRNAs. CAT assays <str<strong>on</strong>g>of</str<strong>on</strong>g> transiently transfected T. cruzi cells show that heat<br />

shock-resp<strong>on</strong>sive elements are present in both the 5’ and 3’ UTRs <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 mRNA. A similar series <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

reporter plasmids are being c<strong>on</strong>structed with HSP10 and HSP60 sequences. CAT mRNA levels in transfected


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

cells are being determined to assess the c<strong>on</strong>tributi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> mRNA stability and its translati<strong>on</strong> to the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the CAT enzyme. We are also determining the half-life <str<strong>on</strong>g>of</str<strong>on</strong>g> the endogenous HSP10, HSP60, HSP70 and<br />

ClpA/HSP100 mRNAs under stressing and n<strong>on</strong>-stressing c<strong>on</strong>diti<strong>on</strong>s to further characterize gene regulati<strong>on</strong><br />

mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> these proteins. Supported by CNPq and FAPERJ.<br />

2K_10_P<br />

(poster secti<strong>on</strong> A2, poster board #149, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

GENOME-WIDE TRANSCRIPTIONAL CHANGES IN RESPONSE TO OXIDATIVE<br />

STRESS IN DROSOPHILA<br />

Chris G. Mah<strong>on</strong> 1 , T<strong>on</strong>y L. Parkes 2 , J. Tim Westwood 1<br />

1Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Cell and Systems Biology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Tor<strong>on</strong>to, Mississauga, ON and 2 Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, North Bay, ON<br />

Canada, e-mail: t.westwood@utor<strong>on</strong>to.ca<br />

Aerobic organisms have evolved a series <str<strong>on</strong>g>of</str<strong>on</strong>g> metabolic defenses whose role it is to c<strong>on</strong>tain and/or repair the<br />

deleterious effects <str<strong>on</strong>g>of</str<strong>on</strong>g> reactive oxygen species (ROS). One <str<strong>on</strong>g>of</str<strong>on</strong>g> the most prominent <str<strong>on</strong>g>of</str<strong>on</strong>g> these is Cu-Zn or<br />

cytosolic superoxide dismutase (cSOD1) which c<strong>on</strong>verts superoxide ani<strong>on</strong> radicals to hydrogen peroxide and<br />

oxygen. Previous genetic studies investigating the phenotypic c<strong>on</strong>sequences <str<strong>on</strong>g>of</str<strong>on</strong>g> a null mutati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> cSOD in<br />

Drosophila melanogaster have resulted in the characterizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a series <str<strong>on</strong>g>of</str<strong>on</strong>g> phenotypes that include: increased<br />

superoxide c<strong>on</strong>centrati<strong>on</strong> in cells; dramatic reducti<strong>on</strong> in l<strong>on</strong>gevity; male sterility and female semi-sterility; and<br />

locomotory and behavioural defects. It has been speculated that increases in superoxide ani<strong>on</strong>s in cells<br />

triggers a number <str<strong>on</strong>g>of</str<strong>on</strong>g> defense resp<strong>on</strong>ses. In this study, we used cSODn 108 null flies as well as flies in which<br />

cSOD activity was restored via transgenic rescue. Flies were collected at 0-24 and 48-72 hours post eclosi<strong>on</strong><br />

and total RNA extracted. Transcripti<strong>on</strong>al pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iles comparing cSOD null vs. transgenic rescue were obtained<br />

using l<strong>on</strong>g olig<strong>on</strong>ucleotide DNA microarrays from the Canadian Drosophila Microarray <str<strong>on</strong>g>Centre</str<strong>on</strong>g> (CDMC,<br />

www.flyarrays.com). Transcripts from more than 200 genes decreased in the cSOD null flies including genes<br />

involved in reproducti<strong>on</strong>, carbohydrate metabolism, lipid biosynthesis and proteases. More than 80 genes<br />

showed increased transcript levels including <strong>on</strong>es involved in lipid catabolism, DNA damage resp<strong>on</strong>se,<br />

apoptosis, and cell adhesi<strong>on</strong>. Unexpectedly, heat shock genes were not induced.<br />

2K_11_P<br />

(poster secti<strong>on</strong> A2, poster board #150, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

FUNCTIONAL STUDIES OF UBIQUITIN CARBOXY-TERMINAL HYDROLASE L1 NOVEL<br />

VARIANT USING PROTEOMICS AND FUNCTIONAL GENOMICS<br />

Sun Kim, Hyun Jung Kim, Hee-Jung Kim, K<strong>on</strong>g-Joo Lee<br />

The Center <str<strong>on</strong>g>for</str<strong>on</strong>g> Cell Signaling & Drug Discovery <str<strong>on</strong>g>Research</str<strong>on</strong>g>, College <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmacy and Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Life &<br />

Pharmaceutical Sciences, Ewha Womans University, Seoul 120-750, Korea, e-mail: kjl@ewha.ac.kr<br />

Ubiquitin Carboxy-terminal Hydrolase isozyme L1(UCH-L1) is a thiol protease that recognizes and<br />

hydrolyzes a peptide b<strong>on</strong>d at the carboxy-terminal <str<strong>on</strong>g>of</str<strong>on</strong>g> ubiquitin, and is involved in the processing <str<strong>on</strong>g>of</str<strong>on</strong>g> ubiquitin<br />

precursors and ubiquitinated proteins. It is normally expressed exclusively in neur<strong>on</strong>s and testis, and<br />

abnormal expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> UCH-L1 has been shown to correlate with several <str<strong>on</strong>g>for</str<strong>on</strong>g>ms <str<strong>on</strong>g>of</str<strong>on</strong>g> cancer, such as cancer <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the lung, col<strong>on</strong>, and pancreas. Mutati<strong>on</strong>s in the UCH-L1 gene have been reported to be linked to<br />

susceptibility to and protecti<strong>on</strong> from Parkins<strong>on</strong>’s disease. Although the correlati<strong>on</strong> between UCH-L1 and<br />

155


23-26 August 2007,<br />

Budapest, Hungary<br />

disease is quite reported, the molecular mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> biological functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> UCH-L1 is poorly understood.<br />

To find out the biological functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> UCH-L1, Proteomics combining 2D-gel electrophoresis and ESI-q-<br />

TOF tandem MS were employed and four variants <str<strong>on</strong>g>of</str<strong>on</strong>g> UCH-L1 were identified. Subsequently, differentially<br />

regulated genes or proteins in cells overexpressing <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the four variants were identified by proteomics and<br />

cDNA microarray, and c<strong>on</strong>firmed by western blot analysis and real-time quantitative RT-PCR. The results<br />

dem<strong>on</strong>strate that UCH-L1 has heterogeneous populati<strong>on</strong>s and each variant plays significantly different<br />

biological roles.<br />

2K_12_P<br />

(poster secti<strong>on</strong> A2, poster board #151, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

COMPARATIVE GENOMICS RESPONSE TO HEAVY METAL STRESS IN THE<br />

CILIATE TETRAHYMENA THERMOPHILA<br />

Virginia Campos, Ana Martín-G<strong>on</strong>zález, Juan C. Gutiérrez<br />

Dpto. Microbiología-III. Facultad de Biología. Universidad Complutense (UCM). Madrid 28040. Spain.<br />

e-mail: juancar@bio.ucm.es<br />

Gene expressi<strong>on</strong> libraries from the ciliate protozoa Tetrahymena thermophila, were obtained after expositi<strong>on</strong> to<br />

Cd (5 mg/L, 3h or 24h) or arsenate (As +5 ) (7.5 mg/L, 1h). All isolated cDNA molecules presented a polyA<br />

tail, c<strong>on</strong>firming the mRNA nature <str<strong>on</strong>g>of</str<strong>on</strong>g> all inserts. From the Cd-gene expressi<strong>on</strong> library 109 cl<strong>on</strong>es were<br />

analysed, and 112 cl<strong>on</strong>es from the As-gene library. Gene expressi<strong>on</strong> results show important differences<br />

between both heavy metal (Cd / As) cell resistance mechanisms, and also between both Cd gene libraries<br />

after different treatment times (3h or 24h). From the different gene libraries we can point out the following<br />

identified protein groups; transporters proteins (ABC, ATPases, etc.) 4.6% in Cd3h, 2.4% in Cd24h and 2.6%<br />

in As1h library, protein-kinases (5.5% Cd3h, 4.8% Cd24h, 3.5% As1h), enzymes involved in protein<br />

degradati<strong>on</strong> ( 6.4% Cd3h, 4.8% Cd24h, 6.3% As1h), oxidative stress protein ( 2.7% Cd3h, 7.3% Cd24h,<br />

18.7% As1h), DNA binding proteins (H4 hist<strong>on</strong>e, NgoA) 6.4% Cd3h, 26.8% Cd24h, 7.1 As1h). About this<br />

last group, the NgoA protein (exclusive <str<strong>on</strong>g>of</str<strong>on</strong>g> this ciliate) is the most abundant (2 cl<strong>on</strong>es in Cd3h, 11 cl<strong>on</strong>es in<br />

Cd24h and 7 cl<strong>on</strong>es in As1h), especially after Cd l<strong>on</strong>g exposure. Metallothi<strong>on</strong>eins are also induced ( 1.8%<br />

Cd3h, 14.6% Cd24h, 0.9% As1h), and am<strong>on</strong>g the 3 CdMTs reported in this ciliate (Díaz et al., 2007) the<br />

MTT5 is the most induced, which agrees with the quantitative RT-PCR results (MTT5 >> MTT1 > MTT3).<br />

Several <str<strong>on</strong>g>of</str<strong>on</strong>g> these genes might be selected to elaborate, in the next future, DNA macro- or micro-arrays to<br />

detect the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> these envir<strong>on</strong>mental pollutants, or a general cellular stress, from soil or aquatic<br />

samples.<br />

156


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2K_13_P<br />

(poster secti<strong>on</strong> A2, poster board #152, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ANALYSIS OF GENE EXPRESSION AND REGULATION OF A NOVEL, STRESS-<br />

INDUCIBLE TRANSCRIPT IN THE MOUSE<br />

C. Liebl, M. V. Schmidt, J. Schülke, D. Trümbach, K. Ganea, V. Sterlemann, T. Rein, M. B. Müller<br />

Max Planck Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychiatry, Munich<br />

e-mail: liebl@mpipsykl.mpg.de<br />

We studied the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> 24 hours maternal deprivati<strong>on</strong> <strong>on</strong> gene expressi<strong>on</strong> in the paraventricular nucleus<br />

(PVN) <str<strong>on</strong>g>of</str<strong>on</strong>g> 9 day-old mice using microarray technology. Am<strong>on</strong>g the str<strong>on</strong>gest regulated genes we discovered a<br />

novel transcript (interim name MPIP-101), which showed a pr<strong>on</strong>ounced upregulati<strong>on</strong> in the maternal<br />

deprived group. This gene codes <str<strong>on</strong>g>for</str<strong>on</strong>g> a short hypothetical protein <str<strong>on</strong>g>of</str<strong>on</strong>g> unknown functi<strong>on</strong> that has also a human<br />

ortholog. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> the current study is the functi<strong>on</strong>al analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> the expressi<strong>on</strong> and regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> this gene<br />

in ne<strong>on</strong>atal and adult mouse brain. In the ne<strong>on</strong>ate , GR-antag<strong>on</strong>ist treatment prevented an maternal<br />

deprivati<strong>on</strong> induced upregulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> MPIP101 mRNA in the PVN, indicating a GR dependant gene<br />

regulati<strong>on</strong>. This finding is in line with a promoter analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> this gene, which revealed several functi<strong>on</strong>al<br />

GRE sites. In c<strong>on</strong>diti<strong>on</strong>al CRHR1 knockout mice the maternal deprivati<strong>on</strong> induced upregulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> MPIP-<br />

101 was significantly reduced, suggesting an additi<strong>on</strong>al regulatory mechanism via CRH mediated pathways. In<br />

adult mice MPIP-101 is mostly expressed in the CA3 regi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the hippocampus, in the cortex and the<br />

cerebellum, with very little expressi<strong>on</strong> in the PVN under basal c<strong>on</strong>diti<strong>on</strong>s. Dexamethas<strong>on</strong>e treatment,<br />

restraint stress and food deprivati<strong>on</strong> result in a pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ound inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> gene expressi<strong>on</strong> in the PVN,<br />

c<strong>on</strong>firming the data <str<strong>on</strong>g>of</str<strong>on</strong>g> the ne<strong>on</strong>ate. In <strong>on</strong>going studies we analyze the cellular localizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> MPIP-101, its<br />

interacti<strong>on</strong> with other proteins and co-localizati<strong>on</strong> with central stress markers.<br />

2K_14_P<br />

(poster secti<strong>on</strong> A2, poster board #153, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE ROLE OF SUPRAMAMMILLARY CELLS PROJECTING THE HIPPOCAMPUS<br />

IN STRESS RESPONSE<br />

Wo<strong>on</strong>g-Ki Choi, Jin-Kyung Oh, Ji-Hyun Kim, Kwang-Ho Pyun, Insop Shim<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Integrative Medicine, College <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, The Catholic University <str<strong>on</strong>g>of</str<strong>on</strong>g> Korea, Seoul 137-701,<br />

South Korea<br />

The hypothalamus-pituitary-adrenocortex (HPA) axis is the central mediator <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress resp<strong>on</strong>se system.<br />

Supramammillary (SuM) regi<strong>on</strong> is relatively unique am<strong>on</strong>g hypothalamic structures in that it sends a large,<br />

direct, projecti<strong>on</strong> to the hippocampal <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>. In a previous study, we have shown that mild stress could<br />

activate supramammillary cells which project to the hippocampus. However, role <str<strong>on</strong>g>of</str<strong>on</strong>g> these cell populati<strong>on</strong>s in<br />

modulating stress resp<strong>on</strong>se is not known. The present study examined the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> stress <strong>on</strong> different<br />

populati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> supramammillary cells which is projecting to the hippocampus by injecting the fluorescent<br />

retrograde tracer fluorogold (FG) into the hippocampus and utilizing immunohistochemistry <str<strong>on</strong>g>of</str<strong>on</strong>g> dopamine,<br />

GABA, nitiric oxide (NO), acetylcholine, CRF and serot<strong>on</strong>in. Immobilizati<strong>on</strong> stress (2hr) produced a marked<br />

increase in these supramammillary cells. Supramammillary areas c<strong>on</strong>tain serot<strong>on</strong>ergic, cholinergic and<br />

corticotropin releasing cells projecting to the hippocampus. Dopaminergic, GABAnergic and nitric oxide<br />

positive cells <str<strong>on</strong>g>of</str<strong>on</strong>g> supramammillary areas hardly projecting to the hippocampus. 53% <str<strong>on</strong>g>of</str<strong>on</strong>g> 5-HT cells, 31% <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

ChAT cells, 56% <str<strong>on</strong>g>of</str<strong>on</strong>g> CRF cells were double stained with retrograde cells from the hippocampus. These results<br />

157


23-26 August 2007,<br />

Budapest, Hungary<br />

suggest that the supramammillary areas c<strong>on</strong>tain distinct cell populati<strong>on</strong>s which are very differentially<br />

resp<strong>on</strong>sive to stress and that serot<strong>on</strong>ergic cells projecting to the hippocampus may play an important role in<br />

modulating stress-related behaviors.<br />

2K_15_P<br />

(poster secti<strong>on</strong> A2, poster board #154, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECT OF A PATHOGENIC STRESSOR ON HUMAN HSP10<br />

Giampiero La Rocca 1 , Everly C<strong>on</strong>way de Macario 2 , Rita Anzal<strong>on</strong>e 1 , Sim<strong>on</strong>a Corrao 1 ,<br />

Francesca Magno 1 , Giovanni Zummo 1 , Francesco Cappello 1 , Alberto J. L. Macario 2 , Felicia Farina 1<br />

1Dipartimento di Medicina Sperimentale, Università di Palermo, Italy; 2 Center <str<strong>on</strong>g>of</str<strong>on</strong>g> Marine Biotechnology,<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Maryland Biotechnology Institute, Baltimore, MD, USA<br />

The impact <str<strong>on</strong>g>of</str<strong>on</strong>g> cigarette smoke extract (CSE) <strong>on</strong> the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the chaper<strong>on</strong>es Hsp10, Hsp60, Hsp70, and<br />

Hsp90 was studied using a human lung fibroblast (HFL-1) cell line, a model system to study lung pathology<br />

associated with cigarette smoking, by proteomics using 2D-IPG, silver stain and Western blotting, and at the<br />

level <str<strong>on</strong>g>of</str<strong>on</strong>g> mRNA by RT-PCR. Chaper<strong>on</strong>omics was used <str<strong>on</strong>g>for</str<strong>on</strong>g> gene and protein identificati<strong>on</strong>. The four<br />

chaper<strong>on</strong>es were expressed in unstressed HFL-1 cells at levels that changed in cells exposed to CSE <str<strong>on</strong>g>for</str<strong>on</strong>g> 24 h.<br />

Quali- and quantitative changes in the four chaper<strong>on</strong>es occurred in the stressed cells with the changes in<br />

Hsp10 being the most striking. While three Hsp10 isoelectric variants were detected in unstressed cells, <strong>on</strong>ly<br />

<strong>on</strong>e, the most basic, was still present in cells exposed to CSE and its mass was changed as if post-translati<strong>on</strong>al<br />

modificati<strong>on</strong>s had been caused by the stressor. The three Hsp10 protein variants, which were detected <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

first time in this work, present in unstressed cells could be due, <str<strong>on</strong>g>for</str<strong>on</strong>g> example, to posttranslati<strong>on</strong>al<br />

modificati<strong>on</strong>s <strong>on</strong> a single product <str<strong>on</strong>g>of</str<strong>on</strong>g> the known human hsp10 gene, or to differences in primary structure <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the variants. The latter is the most likely scenario as indicated by our data; the variants could be the product<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the can<strong>on</strong>ical hsp10 gene and the two hsp10 paralogs present in the human genome. Current research aims<br />

at elucidating the nature <str<strong>on</strong>g>of</str<strong>on</strong>g> the variants and the mechanism by which two <str<strong>on</strong>g>of</str<strong>on</strong>g> them become undetectable in<br />

stressed cells.<br />

158


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2L. YEAST STRESS<br />

2L_01_P<br />

(poster secti<strong>on</strong> A2, poster board #155, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

OPTICAL TWEEZERS COMBINED WITH A MICROFLUIDIC DEVICE FOR<br />

STUDIES OF STRESS RESPONSE IN SINGLE CELLS<br />

Emma Erikss<strong>on</strong> a , Elzbieta Petelenz b , Stefan Hohmann b , Mattias Goksör a<br />

a Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Physics, Göteborg University, SE-41296 Göteborg, Sweden, e-mail: emma.erikss<strong>on</strong>@physics.gu.se<br />

b Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Cell and Molecular Biology, Göteborg University, Sweden<br />

Traditi<strong>on</strong>ally biological experiments are per<str<strong>on</strong>g>for</str<strong>on</strong>g>med <strong>on</strong> large populati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> cells. However, such experiments<br />

cannot reveal behaviours <strong>on</strong> a subcellular level. Optical tweezers have proven to be a useful tool <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

c<strong>on</strong>ducting single cell studies. By combining optical tweezers and a micr<str<strong>on</strong>g>of</str<strong>on</strong>g>luidic device, we have developed a<br />

plat<str<strong>on</strong>g>for</str<strong>on</strong>g>m where we rapidly can change the envir<strong>on</strong>ment <str<strong>on</strong>g>of</str<strong>on</strong>g> a single cell. In our micr<str<strong>on</strong>g>of</str<strong>on</strong>g>luidic device different<br />

media, flowing in separate channels, are combined into a single channel. In such small geometries the flow is<br />

laminar, which means that different media <strong>on</strong>ly mix due to diffusi<strong>on</strong>. C<strong>on</strong>sequently a sharp c<strong>on</strong>centrati<strong>on</strong><br />

gradient <str<strong>on</strong>g>of</str<strong>on</strong>g> a substance can be established. By moving a cell across the gradient, using the optical tweezers, we<br />

are able to change the envir<strong>on</strong>ment <str<strong>on</strong>g>of</str<strong>on</strong>g> a single cell in a fracti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a sec<strong>on</strong>d. During the experiment the cell is<br />

m<strong>on</strong>itored using fluorescence microscopy. Our method is universal, enabling experiments <strong>on</strong> many types <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

cells with different stress media. The system can be expanded to include holographic optical tweezers, thus<br />

enabling several cells to be studied and analyzed simultaneously. The experimental plat<str<strong>on</strong>g>for</str<strong>on</strong>g>m is dem<strong>on</strong>strated<br />

<strong>on</strong> yeast cells (Saccharomyces cerevisiae). We have m<strong>on</strong>itored the diameter <str<strong>on</strong>g>of</str<strong>on</strong>g> yeast cells that are moved between a<br />

flow <str<strong>on</strong>g>of</str<strong>on</strong>g> pure growth medium and a sodium chloride c<strong>on</strong>taining flow. In additi<strong>on</strong> we have also studied yeast<br />

cells where GFP is fused to a protein involved in the induced stress resp<strong>on</strong>se. In this way the spatial<br />

localizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> both the Hog1 protein and the Yap1p protein were followed during osmotic and oxidative<br />

stress, respectively.<br />

2L_02_P<br />

(poster secti<strong>on</strong> A2, poster board #156, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PHYSIOLOGICAL CHARACTERIZATION OF CPR1 ENCODING PEPTIDYL-PROYL<br />

CIS-TRANS ISOMERASE IN THE YEAST SACCHAROMYCES CEREVISIAE KNU5377<br />

UNDER THE MENADIONE-INDUCED OXIDATIVE STRESS<br />

Il-Sup Kim a , Sunhye Kwak a , Sang-O Kw<strong>on</strong> a , Hyunsook Lee a,b , Ilsoo Mo<strong>on</strong> b , Ingnyol Jin a*<br />

aDepartment <str<strong>on</strong>g>of</str<strong>on</strong>g> Microbiology, Kyungpook Nati<strong>on</strong>al University, Daegu 702-701, Korea<br />

bDepartment <str<strong>on</strong>g>of</str<strong>on</strong>g> Anatomy, D<strong>on</strong>gguk University Medical School, Ky<strong>on</strong>gju 780-714, Korea<br />

Cpr1p coding cytosolic cyclophilin (Cyp)-related peptidyl-proyl cis-trans isomerase was str<strong>on</strong>gly induced<br />

during ethanol fermentati<strong>on</strong> at 40ºC in Saccharomyces cerevisiae KNU5377. The cpr1∆ mutant in S. cerevisiae<br />

KNU5377Y, not a laboratory strain such as S. cerevisiae BY4741, showed the stress sensitivity to manadi<strong>on</strong>e<br />

(MD). Cell viability in the cpr1∆ mutant decreased up to 20% under 0.4 mM c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> MD. Under the<br />

stress c<strong>on</strong>diti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 0.4 mM MD, markers <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidative damage such as the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> hydroperoxide, carb<strong>on</strong>yl<br />

c<strong>on</strong>tent and mal<strong>on</strong>dialdehyde was higher in the cpr1∆ mutant than in the wild type <str<strong>on</strong>g>of</str<strong>on</strong>g> KNU5377. But there<br />

was no difference between the wild type and the cpr1∆ mutant in a laboratory strain <str<strong>on</strong>g>of</str<strong>on</strong>g> S. cerevisiae BY4741<br />

even in the 0.2 mM MD stress. This cpr1∆ mutant KNU5377 in these stress c<strong>on</strong>diti<strong>on</strong>s repressed the<br />

159


23-26 August 2007,<br />

Budapest, Hungary<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> various antioxidant enzymes including Cu/Zn superoxide dismutase (Sod1p), thioredoxin 2<br />

(Trx2p), thioredoxin reductase (Tlr1p), thioredoxin peroxidase (Tsa1p), glucose-6-phosphate dehydrogenase<br />

(G6PDH), cytosolic isocitrate dehydrogenase (cIDH), alcohol dehydrogenase (Adhp), and Hsp104.<br />

Interestingly, the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Sod1 was reduced in two cpr1∆ mutants <str<strong>on</strong>g>of</str<strong>on</strong>g> KNU5377 and BY4741. The<br />

down-regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> G6PDH and cIDH in the cpr1∆ mutant <str<strong>on</strong>g>of</str<strong>on</strong>g> KNU5377 caused a reducti<strong>on</strong> in the cellular<br />

NADPH level. These repressed proteins in the cpr1∆ mutant <str<strong>on</strong>g>of</str<strong>on</strong>g> KNU5377 were recovered after treatment <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

such inhibitor as PMSF under the MD stress. In additi<strong>on</strong>, the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Cpr1p in KNU5377 was<br />

regulated by HSF1 under the MD stress. These data indicates that the CPR1 in KNU5377 has an effect <strong>on</strong><br />

the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> antioxidant enzymes and heat shock proteins during the MD stress and is regulated by<br />

HSF1.<br />

2L_03_P<br />

(poster secti<strong>on</strong> A2, poster board #157, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CHANGES OF PROTEOME AND CAROTENOID PRODUCTION<br />

IN STRESSED RED YEASTS<br />

I. Marova 1 , A. Halienova 1 , M. Carnecka 1 , V. Hanusova 2 , V. Hezinova 1<br />

1Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Chemistry, BUT, Brno, Czech Republic, e-mail: marova@fch.vutbr.cz<br />

2Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Chemical and Food Technology, STU, Bratislava, Slovak Republic<br />

Envir<strong>on</strong>mental stress surrounding yeast cells evokes various changes in their behaviour in order to survive. In<br />

this work, protein and metabolic pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iles <str<strong>on</strong>g>of</str<strong>on</strong>g> some carotenogenic yeasts grown in optimal and stress<br />

c<strong>on</strong>diti<strong>on</strong>s were compared. Different 11 carotenogenic yeast strains were cultivated under osmotic, oxidative<br />

and metal stress. Produced carotenoids and ergosterol were measured using HPLC//MS. Glycerol was<br />

analysed using Boehringer kit. Proteins were analyzed by PAGE-SDS, 1D micr<str<strong>on</strong>g>of</str<strong>on</strong>g>luidic system and by 2D<br />

electrophoresis. Under all stress c<strong>on</strong>diti<strong>on</strong>s expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> some protein fracti<strong>on</strong>s was changed. Osmotic stress<br />

led to overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> some specific protein fracti<strong>on</strong>s in most <str<strong>on</strong>g>of</str<strong>on</strong>g> studied yeast strains. These proteomic<br />

pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iles were different from protein pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iles obtained at oxidative as well as metal stress. Presence <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

exogenous stress led to important overproducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> pigments as well as <str<strong>on</strong>g>of</str<strong>on</strong>g> lipidic substances (ergosterol,<br />

glycerol). Producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> carotenoids by R. glutinis cells was about 5-6x higher under oxidative stress, while<br />

producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ergosterol increased more than 10-x. Salt and metal stress led also to slight increase <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

carotenoid producti<strong>on</strong>. Combinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> stress factors in cultivati<strong>on</strong> media induced significant increase <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

beta-carotene <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> mainly in S. roseus (230mg/g in medium with 2% NaCl and 5mM H 2 O 2 ) and in salt<br />

stressed R. glutinis ( 200mg/g). Producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> carotenoids under exogenous stress changed simultaneously<br />

with ergosterol producti<strong>on</strong>, but inversely to glycerol producti<strong>on</strong>.<br />

160


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2L_04_P<br />

(poster secti<strong>on</strong> A2, poster board #158, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

BIPHASIC MODULATION OF THE CELLULAR STRESS RESPONSE BY<br />

HISTAMINE H3R AND H4R RECEPTOR ANTAGONISTS IN EUKARYOTIC CELLS<br />

K. Papamichael 1 , B. Delitheos 1 , R. L. Thurm<strong>on</strong>d 2 , E. Tiligada 1<br />

1Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmacology, Medical School University <str<strong>on</strong>g>of</str<strong>on</strong>g> Athens, Greece; 2 Johns<strong>on</strong> & Johns<strong>on</strong> Pharmaceutical <str<strong>on</strong>g>Research</str<strong>on</strong>g><br />

& Development, L.L.C., San Diego, CA, USA<br />

The highly c<strong>on</strong>served inducible cellular stress resp<strong>on</strong>se (CSR) to adverse envir<strong>on</strong>mental/microenvir<strong>on</strong>mental<br />

c<strong>on</strong>diti<strong>on</strong>s, like heat and drugs, is partly coupled to heat shock protein (hsps) expressi<strong>on</strong>. Am<strong>on</strong>gst others,<br />

histamine (HI), a key inflammatory mediator, interacts with diverse comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g> the CSR. This study<br />

aimed at exploring the role <str<strong>on</strong>g>of</str<strong>on</strong>g> H 3 R and H 4 R antag<strong>on</strong>ists, thioperamide (THIO) and JNJ7777120, respectively,<br />

<strong>on</strong> the CSR in yeast, an established experimental model, which lacks known HR but possesses HI<br />

metabolizing systems. The resp<strong>on</strong>se was evaluated by determining microscopically the viability <str<strong>on</strong>g>of</str<strong>on</strong>g> postlogarithmic<br />

phase grown cell cultures after heat shock (HS) at 53 o C <str<strong>on</strong>g>for</str<strong>on</strong>g> 30min. The effects <str<strong>on</strong>g>of</str<strong>on</strong>g> THIO and<br />

JNJ7777120 were investigated following administrati<strong>on</strong> through to the post-logarithmic phase or <str<strong>on</strong>g>for</str<strong>on</strong>g> 2h<br />

(prec<strong>on</strong>diti<strong>on</strong>ing) prior to HS. Inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> thermotolerance after thermal prec<strong>on</strong>diti<strong>on</strong>ing at 37°C <str<strong>on</strong>g>for</str<strong>on</strong>g> 2h<br />

served as positive c<strong>on</strong>trol. The expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> hsps was determined by western blotting. Similarly to HI,<br />

administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> either THIO or JNJ7777120 induced biphasic effects <strong>on</strong> cell viability, with significant<br />

increases at 0.03-0.3mM and decreases at 0.3-3mM. The presence <str<strong>on</strong>g>of</str<strong>on</strong>g> cycloheximide, a de novo protein synthesis<br />

inhibitor, reversed the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> THIO or JNJ7777120 prec<strong>on</strong>diti<strong>on</strong>ing. Preliminary investigati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp104,<br />

hsp70 and hsp60 expressi<strong>on</strong> showed a comparable pattern. The ability <str<strong>on</strong>g>of</str<strong>on</strong>g> the HR antag<strong>on</strong>ists to induce a<br />

biphasic CSR in yeast, al<strong>on</strong>g with the de novo protein synthesis c<strong>on</strong>tributi<strong>on</strong> in prec<strong>on</strong>diti<strong>on</strong>ing with the agents<br />

and the alterati<strong>on</strong>s in hsp expressi<strong>on</strong> point to the need <str<strong>on</strong>g>for</str<strong>on</strong>g> elucidating the potential clinical implicati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

this phenotype.<br />

2L_05_P<br />

(poster secti<strong>on</strong> A2, poster board #159, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

TRANSCRIPTION FACTORS SPT3P AND MED3P IN RECOVERY OF YEAST CELLS<br />

AFTER THERMAL INSULT<br />

M. Simola, A.-L. Hänninen, L. Seppä, P. Auvinen, M. Makarow<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Biotechnology, P.O.Box 56, FIN-00014 University <str<strong>on</strong>g>of</str<strong>on</strong>g> Helsinki, Finland<br />

e-mail: mari.j.simola@helsinki.fi<br />

Up<strong>on</strong> subjecti<strong>on</strong> to heat shock temperature 37 o C, S. cerevisiae cells acquire thermotolerance, which enables<br />

survival after exposure to 50 o C. We found earlier that when cells grown at 24 o C, adapted at 37 o C and treated<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> 20 min at 50 o C are shifted back to 24 o C, expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> chaper<strong>on</strong>es Hsp104 (cytosol), Kar2p/BiP and<br />

Lhs1p (ER) and Hsp78 (mitoch<strong>on</strong>dria) is up-regulated after 2-4 h <str<strong>on</strong>g>of</str<strong>on</strong>g> recovery at 24 o C. We designated this<br />

novel mechanism delayed up-regulati<strong>on</strong> (DUR), and found it to be mediated by Hsf1p via heat shock<br />

elements. Analysing global gene expressi<strong>on</strong> we found here that producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> transcripti<strong>on</strong> factors Spt3p and<br />

Med3p (Pgd1p/Hrs1p), essential <str<strong>on</strong>g>for</str<strong>on</strong>g> thermotolerance, also was subject to DUR. Spt3p is a member <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

SAGA complex (Spt-Ada-Gcn5-acetyltransferase complex) and Med3p is a subunit <str<strong>on</strong>g>of</str<strong>on</strong>g> the Mediator complex.<br />

HAC1 u (uninduced) mRNA is transcribed under normal c<strong>on</strong>diti<strong>on</strong>s. When unfolded proteins accumulate in<br />

the ER, HAC1 u is spliced by the ER-membrane kinase Ire1p to HAC1 i (induced) mRNA, which is translated<br />

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23-26 August 2007,<br />

Budapest, Hungary<br />

into the transcripti<strong>on</strong> factor Hac1p, causing increased expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ER chaper<strong>on</strong>es. We found that<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HAC1 mRNA is subject to DUR. Producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HAC1 i was str<strong>on</strong>gly up-regulated during<br />

recovery at 24 o C, due to increased transcripti<strong>on</strong> and splicing <str<strong>on</strong>g>of</str<strong>on</strong>g> HAC1 u mRNA. In the absence <str<strong>on</strong>g>of</str<strong>on</strong>g> Spt3p or<br />

Med3p, DUR <str<strong>on</strong>g>of</str<strong>on</strong>g> Hac1p producti<strong>on</strong> was diminished and delayed. C<strong>on</strong>sequently, a heat-denatured cytosolic<br />

reporter enzyme failed to be effectively reactivated, and refolding <str<strong>on</strong>g>of</str<strong>on</strong>g> heat-denatured secretory proteins in the<br />

ER to secreti<strong>on</strong>-competent c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>s was delayed. Moreover, in the spt3∆ med3∆ mutant the cell shape<br />

and bud <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> were str<strong>on</strong>gly affected.<br />

2L_06_P<br />

(poster secti<strong>on</strong> A2, poster board #160, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

MRNA MISTRANSLATION IN YEAST: THE ROLE OF A CONSTITUTIVE STRESS<br />

RESPONSE IN CELL ADAPTATION AND EVOLUTION<br />

Raquel M. Silva 1 , João A. Paredes 1 , Tatiana Lima-Costa 1 , Marian G. Koerkamp 2 , Michel Perrot 3 ,<br />

Hélian Boucherie 3 , Frank Holstege 2 , Manuel A. S. Santos 1<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology and CESAM, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Aveiro, 3810-193 Aveiro, Portugal.<br />

e-mail: msantos@bio.ua.pt<br />

2UMC Utrecht, Department <str<strong>on</strong>g>for</str<strong>on</strong>g> Physiological Chemistry, Postbus 80042 3508 TA, Utrecht, The Netherlands<br />

3IBGC, UPR CNRS9026, 1, Rue Camille Saint-Saëns, 33700, Bordeaux, France<br />

Translati<strong>on</strong> fidelity assures the producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> stable and functi<strong>on</strong>al proteomes and, there<str<strong>on</strong>g>for</str<strong>on</strong>g>e, mistranslati<strong>on</strong><br />

events that result in the synthesis <str<strong>on</strong>g>of</str<strong>on</strong>g> aberrant proteins are <str<strong>on</strong>g>of</str<strong>on</strong>g>ten linked to disease. In normal physiological<br />

c<strong>on</strong>diti<strong>on</strong>s mistranslati<strong>on</strong> errors are low, however, the error frequency increases under stress. In order to<br />

understand this phenotype at the molecular level, we have induced c<strong>on</strong>stitutive mistranslati<strong>on</strong> in the<br />

eukaryotic model Saccharomyces cerevisiae, using tRNA engineering methodologies.<br />

Transcriptome and proteome characterizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the engineered strains shows that mistranslati<strong>on</strong> extensively<br />

and permanently remodels gene expressi<strong>on</strong>. Additi<strong>on</strong>ally, mistranslating cells accumulate trehalose and<br />

glycogen and increase proteasome activity, suggesting that these cells resp<strong>on</strong>d to the producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> aberrant<br />

proteins by inducing mechanisms that promote protein folding, stabilizati<strong>on</strong> and degradati<strong>on</strong>.<br />

Remarkably, mRNA mistranslati<strong>on</strong> in yeast impaired sexual reproducti<strong>on</strong>, induced ploidy alterati<strong>on</strong>s, and<br />

triggered stress cross-protecti<strong>on</strong> that generated adaptive advantages under envir<strong>on</strong>mental challenging<br />

c<strong>on</strong>diti<strong>on</strong>s. This study highlights unanticipated roles <str<strong>on</strong>g>for</str<strong>on</strong>g> mRNA mistranslati<strong>on</strong> in the evoluti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> novel<br />

phenotypes and speciati<strong>on</strong>.<br />

162


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2M. OTHER CELLULAR STRESS RELATED TOPICS (STRESS-RELATED PROTEIN<br />

DEGRADATION, STRESS AND CELL MOVEMENT, STRESS PROTEINS AND THE<br />

CELL CYCLE, EXTREMOPHILES)<br />

2M_01_P<br />

(poster secti<strong>on</strong> A2, poster board #161, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ARSENITE-ELICITED SKP2 PROTEOLYSIS REQUIRES LEUPEPTIN-SENSITIVE<br />

PROTEASES AND APC/C CDH1 BUT NOT 26S PROTEASOME<br />

Yen-An Tang, Ju-Pi Li, Jia-Ling Yang 1<br />

Molecular Carcinogenesis Laboratory, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Biotechnology & Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Life Sciences,<br />

Nati<strong>on</strong>al Tsing Hua University, Hsinchu 30013, Taiwan<br />

1e-mail: jlyang@life.nthu.edu.tw<br />

Skp2, an F-box protein <str<strong>on</strong>g>of</str<strong>on</strong>g> SCF E3 ubiquitin ligase, targets key cell cycle regulators <str<strong>on</strong>g>for</str<strong>on</strong>g> degradati<strong>on</strong> including<br />

p27 and p21 to timely c<strong>on</strong>trol the progressi<strong>on</strong> from late G 1 until mitotic entry, while its protein level is keep<br />

low via the APC/C Cdh1 and 26S proteasome mediated ubiquitinati<strong>on</strong>-degradati<strong>on</strong> pathway at G 0 -G 1 . Here we<br />

show a novel mechanism <str<strong>on</strong>g>for</str<strong>on</strong>g> the Skp2 destructi<strong>on</strong> under arsenite stress. Arsenite decreased cellular Skp2<br />

protein and mRNA levels in a dose- and time-dependent manner. Under protein synthesis inhibiti<strong>on</strong>, arsenite<br />

markedly induced Skp2 ubiquitinati<strong>on</strong> and degradati<strong>on</strong>. Depleti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Cdh1 using small interfering RNA<br />

lowered the arsenite-induced Skp2 ubiquitinati<strong>on</strong> and degradati<strong>on</strong>; however, co-treatment with 26S<br />

proteasome inhibitors MG132, ALLN, or ALLM did not avoid the Skp2 degradati<strong>on</strong> under arsenite.<br />

Intriguingly, co-treatment with leupeptin, an inhibitor <str<strong>on</strong>g>of</str<strong>on</strong>g> lysosomal proteases, rescued the arsenite-induced<br />

Skp2 destructi<strong>on</strong>. The Skp2 down-regulati<strong>on</strong> was parallel to increases in p21 and p27 levels, causing a marked<br />

delay <str<strong>on</strong>g>of</str<strong>on</strong>g> S entry in arsenite-treated G 1 cells. These results suggest that the arsenite-elicited ubiquitinated-Skp2<br />

generated by APC Cdh1 is not a target <str<strong>on</strong>g>of</str<strong>on</strong>g> 26S proteasome, while Skp2 proteolysis under this stress requires<br />

lysosomal proteases. This study also suggests that Skp2 down-regulati<strong>on</strong> may be a key c<strong>on</strong>tributor to the G 1<br />

arrests caused by arsenite.<br />

2M_02_P<br />

(poster secti<strong>on</strong> A2, poster board #162, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

MICROELASTIC GRADIENT GELS TO INDUCE CELLULAR MECHANOTAXIS<br />

Satoru Kidoaki 1* , Takehisa Matsuda 2<br />

1 Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Biomolecular Chemistry, Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Materials Science and Engineering, Kyushu University, Fukuoka<br />

812-8581, Japan, e-mail: kidoaki@ms.ifoc.kyushu-u.ac.jp<br />

2 Genome Biotechnology Laboratory, Kanazawa Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Technology, Ishikawa 924-0838, Japan<br />

The understanding and realizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> directi<strong>on</strong>al cell movement towards a harder regi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a cell culture<br />

substrate surface, so-called mechanotaxis, might provide a solid basis <str<strong>on</strong>g>for</str<strong>on</strong>g> a functi<strong>on</strong>al artificial extracellular<br />

matrix, enabling manipulati<strong>on</strong> and elucidati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cell motility. The photolithographic surface microelasticity<br />

patterning method was developed <str<strong>on</strong>g>for</str<strong>on</strong>g> fabricating a cell-adhesive hydrogel with a microelasticity-gradient<br />

(MEG) surface using photocurable styrenated gelatin to investigate the c<strong>on</strong>diti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> surface elasticity to<br />

induce mechanotaxis as a basis <str<strong>on</strong>g>for</str<strong>on</strong>g> such substrate-elasticity-dependent c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> cell motility. Patterned<br />

MEG gels c<strong>on</strong>sisting <str<strong>on</strong>g>of</str<strong>on</strong>g> different absolute surface elasticities and elasticity jumps were prepared. Surface<br />

elasticity and its two-dimensi<strong>on</strong>al distributi<strong>on</strong> were characterized by microindentati<strong>on</strong> tests using atomic <str<strong>on</strong>g>for</str<strong>on</strong>g>ce<br />

163


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Budapest, Hungary<br />

microscopy (AFM). From analyses <str<strong>on</strong>g>of</str<strong>on</strong>g> trajectories <str<strong>on</strong>g>of</str<strong>on</strong>g> 3T3 cell movement <strong>on</strong> each prepared MEG gel, two<br />

critical criteria <str<strong>on</strong>g>of</str<strong>on</strong>g> the elasticity jump and the absolute elasticity to induce mechanotaxis were identified: 1) a<br />

high elasticity ratio between the hard regi<strong>on</strong> and the s<str<strong>on</strong>g>of</str<strong>on</strong>g>t <strong>on</strong>e, and 2) elasticity <str<strong>on</strong>g>of</str<strong>on</strong>g> the s<str<strong>on</strong>g>of</str<strong>on</strong>g>ter regi<strong>on</strong> to provide<br />

medium motility. Design <str<strong>on</strong>g>of</str<strong>on</strong>g> these c<strong>on</strong>diti<strong>on</strong>s was found to be necessary <str<strong>on</strong>g>for</str<strong>on</strong>g> fabricating an artificial<br />

extracellular matrix to c<strong>on</strong>trol or manipulate cell motility.<br />

2M_03_P<br />

(poster secti<strong>on</strong> A2, poster board #163, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

APOPTOSIS OF BLOOD'S CELLS IN FAMILIES OF PATIENT WITH NEUROTIC,<br />

STRESS-RELATED AND SOMATOFORM DISORDERS<br />

Lyudmila A. Ryadovaya, E. V. Gutkevich, S. A. Ivanova, E. M. Epanchinceva, O. V. Aksy<strong>on</strong>ova<br />

Mental Health <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute, Russian Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Sciences<br />

Objective: To establish processes <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis in families <str<strong>on</strong>g>of</str<strong>on</strong>g> patients with mental disorders. Methods: 55<br />

families <str<strong>on</strong>g>of</str<strong>on</strong>g> patients with dissociative (c<strong>on</strong>versi<strong>on</strong>) disorders and 36 families <str<strong>on</strong>g>of</str<strong>on</strong>g> patients with adjustment<br />

disorders. We estimate <str<strong>on</strong>g>of</str<strong>on</strong>g> processes <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis at receptor and cell-like levels <str<strong>on</strong>g>for</str<strong>on</strong>g> the patients and relatives<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the first degree <str<strong>on</strong>g>of</str<strong>on</strong>g> relati<strong>on</strong>ship. Results: We have observed statistically significant increase <str<strong>on</strong>g>of</str<strong>on</strong>g> expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

a receptor CD95 in patients with dissociative (c<strong>on</strong>versi<strong>on</strong>) disorders in comparis<strong>on</strong> with c<strong>on</strong>trol<br />

(14,73±0,85% and 11,79±0,44%, accordingly, p


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2M_04_P<br />

(poster secti<strong>on</strong> A2, poster board #164, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CDC20 DEGRADATION REQUIRES P38 MAPK AND CDH1-INDEPENDENT APC/C<br />

ACTIVITIES DURING CADMIUM-INDUCED MITOTIC ARREST<br />

Ai-Hsin Yen, Jia-Ling Yang 1<br />

Molecular Carcinogenesis Laboratory, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Biotechnology & Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Life Sciences, Nati<strong>on</strong>al Tsing Hua<br />

University, Hsinchu 30013, Taiwan<br />

1e-mail: jlyang@life.nthu.edu.tw<br />

Cdc20, an activator <str<strong>on</strong>g>of</str<strong>on</strong>g> the anaphase promoting complex/Cyclosome (APC/C) E3 ligase (APC/C Cdc20 ), timely<br />

targets key mitotic regulators <str<strong>on</strong>g>for</str<strong>on</strong>g> destructi<strong>on</strong> via ubiquitinati<strong>on</strong>, while its destructi<strong>on</strong> triggered by Cdh1-<br />

dependent APC/C (APC/C Cdh1 ) is essential <str<strong>on</strong>g>for</str<strong>on</strong>g> mitotic exit. We report here the mechanism involved in<br />

unscheduled Cdc20 destructi<strong>on</strong> during cadmium stress. Cadmium dose- and time-dependently decreased the<br />

Cdc20 levels in CL3 human n<strong>on</strong>-small-cell lung carcinoma cells at exp<strong>on</strong>ential growth and early G2 phase.<br />

This Cdc20 down-regulati<strong>on</strong> was due mainly to a decrease in protein half-life mediated via the ubiquitinproteasome<br />

pathway. Cdc20 destructi<strong>on</strong> in cadmium-treated early G2 cells was correlated to mitotic arrest,<br />

which was rescued by blockage <str<strong>on</strong>g>of</str<strong>on</strong>g> p38 MAPK activati<strong>on</strong> using SB202190; whereas, JNK or ERK activati<strong>on</strong><br />

were dispensable. The cadmium-elicited Cdc20 degradati<strong>on</strong> was also suppressed in cells stably expressed a<br />

dominant negative <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> p38 MAPK or knockdown <str<strong>on</strong>g>of</str<strong>on</strong>g> p38α via siRNA. Forced expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a<br />

c<strong>on</strong>stitutive active MKK6 and p38 markedly induced Cdc20 destructi<strong>on</strong>. Intriguingly, Cdc27 (an APC/C<br />

subunit) but not Cdh1 (an APC/C activator) depleti<strong>on</strong> markedly suppressed the Cdc20 destructi<strong>on</strong> under<br />

cadmium or overexpressing p38 signaling. Together, these results suggest that stress-induced Cdc20<br />

ubiquitinati<strong>on</strong> and proteolysis requires the p38 MAPK signaling and a Cdh1-independent APC/C activity.<br />

Such a regulati<strong>on</strong> is necessary <str<strong>on</strong>g>for</str<strong>on</strong>g> preventing the progressi<strong>on</strong> from metaphase to anaphase after cadmium<br />

stress.<br />

2M_05_P<br />

(poster secti<strong>on</strong> A2, poster board #165, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

MECHANISMS OF TRANSLATION REGULATION DURING COLD-SHOCK IN<br />

ESCHERICHIA COLI<br />

Anna Maria Giuliodori, Paolo Marchi, Mara Giangrossi, Anna Brandi, Claudio O. Gualerzi,<br />

Cynthia L. P<strong>on</strong><br />

Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Genetics, Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology MCA, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Camerino, 62032 Camerino (MC), Italy<br />

e-mail: annamaria.giuliodori@unicam.it<br />

Cold-shock (cs) translati<strong>on</strong>al bias, namely the c<strong>on</strong>diti<strong>on</strong> which favors translati<strong>on</strong> at low temperature <str<strong>on</strong>g>of</str<strong>on</strong>g> cs<br />

mRNAs, is <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the main mechanisms by which Escherichia coli cells ensure the selective expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> its cs<br />

genes after cold stress. The bias is partly due to intrinsic features <str<strong>on</strong>g>of</str<strong>on</strong>g> cs mRNAs, which make them pr<strong>on</strong>e to<br />

translati<strong>on</strong> at low temperature, and to a cold stress-induced transient increase <str<strong>on</strong>g>of</str<strong>on</strong>g> the Initiati<strong>on</strong> Factors<br />

(IFs)/ribosome ratio. In this study we have undertaken the task <str<strong>on</strong>g>of</str<strong>on</strong>g>: i) identifying the mechanism generating<br />

the stoichiometric imbalance <str<strong>on</strong>g>of</str<strong>on</strong>g> the IFs/ ribosome ratio; ii) unraveling the role <str<strong>on</strong>g>of</str<strong>on</strong>g> the IFs in the translati<strong>on</strong><br />

bias; iii) elucidating the sec<strong>on</strong>dary structure <str<strong>on</strong>g>of</str<strong>on</strong>g> the paradigm cs mRNA, namely the E. coli cspA mRNA and<br />

iv) detecting possible temperature-dependent variati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> its structure.<br />

165


23-26 August 2007,<br />

Budapest, Hungary<br />

The results obtained indicate that: i) transcripti<strong>on</strong> and translati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> infA and infC which encode IF1 and IF3,<br />

respectively, are activated de novo by cs while ribosomal subunits assembly is slowed down; ii) at low<br />

temperature IF3 stimulates the rate <str<strong>on</strong>g>of</str<strong>on</strong>g> “30S initiati<strong>on</strong> complex” <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> with cs mRNAs while inducing<br />

the <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> n<strong>on</strong>-productive 70S initiati<strong>on</strong> complexes with n<strong>on</strong>-cs mRNAs; iii) the increased level <str<strong>on</strong>g>of</str<strong>on</strong>g> IF1<br />

and IF3 during cs is essential to provide a sufficient pool <str<strong>on</strong>g>of</str<strong>on</strong>g> dissociated 30S ribosomal subunits capable <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

“70S initiati<strong>on</strong> complex” <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> and iv) the structure <str<strong>on</strong>g>of</str<strong>on</strong>g> cspA mRNA, as determined by chemical and<br />

enzymatic probing, changes up<strong>on</strong> temperature down-shift exposing the translati<strong>on</strong> initiati<strong>on</strong> regi<strong>on</strong>.<br />

2M_06_P<br />

(poster secti<strong>on</strong> A2, poster board #166, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECTS OF SOME STRESS FACTORS ON THE BLOOD ANTIOXIDANT DEFENSE<br />

SYSTEM IN C57BL/6 MICE<br />

I. V. Mikhaylov, A. A. Baizhumanov, M. Ya. Akhalaya<br />

Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Radiati<strong>on</strong> Biophysics, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biophysics, Biology Faculty, M.V. Lom<strong>on</strong>osov Moscow State<br />

University, Moscow, Russia<br />

e-mail: evanmih@gmail.com<br />

Intracellular factors determining the resistance to adverse envir<strong>on</strong>mental events probably play a role in the<br />

adaptive resp<strong>on</strong>se. Any moderate adverse event activates defense reserves <str<strong>on</strong>g>of</str<strong>on</strong>g> the organism via stimulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

oxidati<strong>on</strong> processes, which in turn activate antioxidant system and improve general organism’s resistance to<br />

stress factors. We studied the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> short-term swimming in cold water (13°C) <strong>on</strong> parameters <str<strong>on</strong>g>of</str<strong>on</strong>g> the blood<br />

antioxidant system (SOD, catalase, ceruloplasmin (CP), and n<strong>on</strong>protein thiols). The test parameters <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

antioxidant protecti<strong>on</strong> increased 1 h after cold exposure and remained high 1 day after treatment. These<br />

changes were accompanied by an increase in the adaptive capacity. After swimming in cold water the<br />

resistance <str<strong>on</strong>g>of</str<strong>on</strong>g> animals to another stress factor (administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> epinephrine) was higher compared to<br />

c<strong>on</strong>trols. The similar changes <str<strong>on</strong>g>of</str<strong>on</strong>g> blood antioxidant systems were observed after investigati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> another stress<br />

exposure (immobilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> animals during 30 min.). But the n<strong>on</strong>specific reacti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the organism to<br />

envir<strong>on</strong>mental factors may be accompanied by the appearance <str<strong>on</strong>g>of</str<strong>on</strong>g> stress markers which can evaluate role <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

different stress factors <strong>on</strong> adaptive resp<strong>on</strong>se. It is well known the immunosuppressive acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ultraviolet B<br />

irradiati<strong>on</strong> (UVB) <strong>on</strong> living systems. In our experiments in vivo we showed the different modulati<strong>on</strong> acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

UVB-irradiati<strong>on</strong> <strong>on</strong> the immobilizati<strong>on</strong> stress-induced changes <str<strong>on</strong>g>of</str<strong>on</strong>g> SOD and CP in blood plasma <str<strong>on</strong>g>of</str<strong>on</strong>g> animals.<br />

The combined exposure <str<strong>on</strong>g>of</str<strong>on</strong>g> immobilizati<strong>on</strong> and UVB provided to stimulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> SOD activity as compared to<br />

<strong>on</strong>ly acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> immobilizati<strong>on</strong>, but abolished the stress-induced increase in plasma CP level. The different<br />

sensitivity <str<strong>on</strong>g>of</str<strong>on</strong>g> SOD and CP to UVB-irradiati<strong>on</strong> was observed in in vitro experiments: the dose-dependent<br />

decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> CP was shown whereas SOD activity did not changed. We c<strong>on</strong>clude that plasma CP level may be<br />

used as marker <str<strong>on</strong>g>of</str<strong>on</strong>g> UVB-induced immunosuppressive acti<strong>on</strong>.<br />

166


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2M_07_P<br />

(poster secti<strong>on</strong> A2, poster board #167, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

MOLECULAR CHAPERONES IN DIAPAUSE EMBRYOS OF THE EXTREMOPHILE<br />

ARTEMIA FRANCISCANA<br />

Svetla Bojikova-Fournier, Yan Hu, Zhijun Qiu, Tania S. Villeneuve, Yu Sun, Tao Chen,<br />

Reinout Am<strong>on</strong>s, Thomas H. MacRae<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, Dalhousie University, Halifax, N.S. B3H 4J1, Canada<br />

Diapause destined embryos <str<strong>on</strong>g>of</str<strong>on</strong>g> the extremophile crustacean, Artemia franciscana, encyst as gastrulae (cysts),<br />

exhibiting pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ound reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> metabolic activity and extreme stress tolerance. Resistance to stress is<br />

thought to depend partly <strong>on</strong> the developmentally regulated synthesis <str<strong>on</strong>g>of</str<strong>on</strong>g> three small heat shock proteins<br />

(sHSPs), <strong>on</strong>ly <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> which is stress inducible. The sHSPs <str<strong>on</strong>g>for</str<strong>on</strong>g>m poly-disperse oligomers and functi<strong>on</strong> as<br />

molecular chaper<strong>on</strong>es. The best characterized Artemia sHSP, termed p26, c<strong>on</strong>fers thermal tolerance <strong>on</strong><br />

transfected mammalian cells and inhibits apoptosis. Molecular analyses by site-directed mutagenesis revealed<br />

the importance <str<strong>on</strong>g>of</str<strong>on</strong>g> individual p26 amino acid residues, including a highly c<strong>on</strong>served arginine, in chaper<strong>on</strong>e<br />

activity, and suggested that β-strand 7 has an important role in oligomerizati<strong>on</strong>. Artemin, a ferritin<br />

homologue, is also abundantly up-regulated in diapause destined Artemia embryos. Artemin lacks the di-ir<strong>on</strong><br />

ferroxidase center characteristic <str<strong>on</strong>g>of</str<strong>on</strong>g> ferritin and is enriched in cysteines that may promote protein stability by<br />

disulphide b<strong>on</strong>d <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>. Interestingly, artemin and ferritin both functi<strong>on</strong> as molecular chaper<strong>on</strong>es in vitro<br />

and artemin c<strong>on</strong>fers thermal and oxidative tolerance <strong>on</strong> transfected mammalian cells. Artemin appears to<br />

have evolved from ferritin, losing the ability to bind ir<strong>on</strong>, but retaining chaper<strong>on</strong>e activity. As a c<strong>on</strong>sequence<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> substantial accumulati<strong>on</strong> in diapause embryos, artemin has the potential to c<strong>on</strong>tribute significantly to stress<br />

resistance. The available evidence collectively favors the hypothesis that molecular chaper<strong>on</strong>es play critical<br />

roles during diapause in A. franciscana, and by extrapolati<strong>on</strong>, in other organisms.<br />

167


168<br />

23-26 August 2007,<br />

Budapest, Hungary


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

MODULE 3<br />

SYSTEM LEVEL<br />

INTERDISCIPLINARY<br />

APPROACHES<br />

169


23-26 August 2007,<br />

Budapest, Hungary<br />

SYMPOSIA<br />

• 3A. Hans Selye Centennial Symposium (August 24th Friday morning)<br />

(Organizer and chair: George P. Chrousos)<br />

Poster: August 24 th Saturday, Poster secti<strong>on</strong> A1, Building A groundfloor<br />

POSTER SYMPOSIA<br />

• 3B. Fr<strong>on</strong>tiers in stress research<br />

August 25 th Saturday, Poster secti<strong>on</strong> A1, Building A groundfloor<br />

• 3C. Chaper<strong>on</strong>e networks and stress proteomics<br />

August 25 th Saturday, Poster secti<strong>on</strong> A1, Building A groundfloor<br />

170


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

3A. HANS SELYE CENTENNIAL SYMPOSIUM<br />

(GEORGE P. CHROUSOS)<br />

3A_01_S<br />

AN INTEGRATED VIEW OF THE STRESS RESPONSE AND STRESS-RELATED<br />

BEHAVIORAL AND / OR SOMATIC DISORDERS<br />

George P. Chrousos<br />

Athens University Medical School, Athens, Greece<br />

Stress activates the central and peripheral comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress system, i.e., the hypothalamic-pituitaryadrenal<br />

(HPA) axis and the arousal/sympathetic system. The principal effectors <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress system are<br />

corticotropin-releasing horm<strong>on</strong>e (CRH), arginine vasopressin, the proopiomelanocortin-derived peptides<br />

alpha -melanocyte- stimulating horm<strong>on</strong>e and beta-endorphin, the glucocorticoids, and the catecholamines<br />

norepinephrine and epinephrine. The developing brain undergoes rapid growth and is characterized by high<br />

turnover <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>al c<strong>on</strong>necti<strong>on</strong>s during the prenatal and early extrauterine life. These processes and, hence,<br />

brain plasticity, slow down during childhood and puberty and plateau in young adulthood. Horm<strong>on</strong>al acti<strong>on</strong>s<br />

in early life, and to a much lesser extent later, can be organizati<strong>on</strong>al, i.e., can have effects that last <str<strong>on</strong>g>for</str<strong>on</strong>g> l<strong>on</strong>g<br />

periods <str<strong>on</strong>g>of</str<strong>on</strong>g> time, frequently <str<strong>on</strong>g>for</str<strong>on</strong>g> the entire life <str<strong>on</strong>g>of</str<strong>on</strong>g> the individual. Horm<strong>on</strong>es <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress system and sex<br />

steroids have such effects, which influence the behavior and certain physiologic functi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> individuals <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

life.<br />

Exposure <str<strong>on</strong>g>of</str<strong>on</strong>g> the developing brain to severe and/or prol<strong>on</strong>ged stress may result in hyperactivity/<br />

hyperreactivity <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress system, with resultant amygdala hyperfuncti<strong>on</strong> (fear reacti<strong>on</strong>), decreased activity<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the hippocampus (defective glucocorticoid negative feedback, cogniti<strong>on</strong>) and the mesocorticolimbic<br />

dopaminergic system (dysthymia, novelty seeking, addictive behaviors), hyperactivati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the HPA axis<br />

(hypercortisolism), suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> reproductive, growth, thyroid and immune functi<strong>on</strong>s, and changes in pain<br />

and fatigue percepti<strong>on</strong>. These changes may be accompanied by disturbed childhood, adolescent and adult<br />

behaviors, including excessive fear (“inhibited child syndrome”) and addictive behaviors, dysthymia and/or<br />

depressi<strong>on</strong> and gradual development <str<strong>on</strong>g>of</str<strong>on</strong>g> comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g> the metabolic syndrome X, including visceral obesity,<br />

diabetes mellitus type 2 and essential hypertensi<strong>on</strong>. Prenatal stress exerted during the period <str<strong>on</strong>g>of</str<strong>on</strong>g> sexual<br />

differentiati<strong>on</strong> may be accompanied by impairment <str<strong>on</strong>g>of</str<strong>on</strong>g> this process, with behavioral and/or somatic sequelae.<br />

The vulnerability <str<strong>on</strong>g>of</str<strong>on</strong>g> individuals to develop varying degrees and/or comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g> the above life-l<strong>on</strong>g<br />

syndrome is defined by as yet unidentified genetic factors, which account <str<strong>on</strong>g>for</str<strong>on</strong>g> up to 60% <str<strong>on</strong>g>of</str<strong>on</strong>g> the variance.<br />

CRH has marked kindling and glucocorticoids have str<strong>on</strong>g c<strong>on</strong>solidating properties, hence both <str<strong>on</strong>g>of</str<strong>on</strong>g> these<br />

horm<strong>on</strong>es are crucial in the development <str<strong>on</strong>g>of</str<strong>on</strong>g>, and can each al<strong>on</strong>e produce, the above syndrome. CRH and<br />

glucocorticoids may act in synergy, as in acoustic startle, while glucocorticoids may suppress or stimulate<br />

CRH, as in the hypothalamus and amygdala respectively. A CRH receptor type 1 antag<strong>on</strong>ist antalarmin<br />

inhibits both the development and expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> c<strong>on</strong>diti<strong>on</strong>ed fear in rats and has anxiolytic properties in<br />

m<strong>on</strong>keys. Pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ound stressors, such as those from sexual abuse, rape, etc., may elicit comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g> poststress<br />

the syndrome in older children, adolescents and adults as well. Most frequently, chr<strong>on</strong>ic dysthymia<br />

and/or depressi<strong>on</strong> may ensue, associated with gastrointestinal complaints and/or the premenstrual tensi<strong>on</strong><br />

syndrome. A lesser proporti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> individuals may develop the classic posttraumatic stress disorder<br />

characterized by hypocortisolism and intrusive and avoidance symptoms; in younger individuals it may<br />

present as dissociative pers<strong>on</strong>ality disorder.<br />

171


23-26 August 2007,<br />

Budapest, Hungary<br />

172<br />

3A_02_S<br />

STRESS AND ADAPTATION: CHANGES<br />

IN THE SYMPATHOADRENAL SYSTEM ACTIVITY<br />

R. Kvetnansky<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Experimental Endocrinology, Slovak Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences, Bratislava, Slovakia<br />

According to Hans Selye stress is an unspecific resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> the organism to any demand made up<strong>on</strong> it.<br />

However, the specific activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> two comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g> the sympathoadrenal system (adrenomedullary and<br />

sympath<strong>on</strong>eural) by various stressors has been shown.<br />

The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> the present work was to investigate changes in enzymes involved in norepinephrine and<br />

epinephrine (EPI) synthesis - tyrosine hydroxylase (TH), dopamine-β-hydroxylase (DBH) and<br />

phenylethanolamine N-methyltransferase (PNMT) – their gene expressi<strong>on</strong>, immunoprotein levels and<br />

activities in the adrenal medulla (AM), sympathetic ganglia, hearts and brains <str<strong>on</strong>g>of</str<strong>on</strong>g> rats and mice after a single or<br />

repeated exposure to various stressors. Immobilizati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> 2h (IMO), cold 4ºC (COLD), administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

insulin 5IU (INS) or 2-deoxyglucose 500 mg/kg (2DG) were used. A single exposure to IMO, COLD, INS<br />

or 2DG was found to induce increases in TH, DBH and PNMT mRNA levels in the studied organs.<br />

Increased transcripti<strong>on</strong> rate is resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> stress-induced TH and PNMT gene expressi<strong>on</strong>. Repeated<br />

exposure to these stressors elevated besides mRNA also activity and immunoprotein levels <str<strong>on</strong>g>of</str<strong>on</strong>g> the enzymes.<br />

Various stressors regulate TH, DBH and PNMT gene expressi<strong>on</strong> by different transcripti<strong>on</strong>al mechanisms.<br />

PNMT gene expressi<strong>on</strong> is mainly regulated by HPA axis and in corticoliberine gene knock-out mice is<br />

basically reduced especially after stress exposure. One day cold exposure elevated TH mRNA levels in AM,<br />

however, 28 day cold exposure did not show any changes. Cold-adapted rats, however, resp<strong>on</strong>ded to<br />

heterotypic novel stressors (IMO, INS, or 2DG) by exaggerated resp<strong>on</strong>ses.<br />

Thus, our data suggest an adaptati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> TH and PNMT gene expressi<strong>on</strong> during l<strong>on</strong>g-term exposure to<br />

stressors. An exposure <str<strong>on</strong>g>of</str<strong>on</strong>g> adapted rats to novel stressors induces exaggerated resp<strong>on</strong>ses. It is the readiness <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the l<strong>on</strong>g-term stressed organism to overresp<strong>on</strong>d, already at the level <str<strong>on</strong>g>of</str<strong>on</strong>g> expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> genes, to the changed<br />

quality <str<strong>on</strong>g>of</str<strong>on</strong>g> stressor, that we c<strong>on</strong>sider as an important adaptive phenomen<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the sympathoadrenal system.<br />

3A_03_S<br />

CHOLINESTERASE MODULATIONS OF MAMMALIAN STRESS AND ANXIETY<br />

REACTIONS<br />

Herm<strong>on</strong>a Soreq<br />

The Hebrew University <str<strong>on</strong>g>of</str<strong>on</strong>g> Jerusalem; Safra Campus-Givat Ram; Jerusalem 91904; Israel<br />

Stress and anxiety disorders present a major mental health problem, but their putative involvement in the<br />

initiati<strong>on</strong> and/or progressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> neurodegenerative diseases is being debated. Recent research in the lab<br />

focuses <strong>on</strong> the molecular mechanism(s) underlying anxiety-induced changes in cholinergic neurotransmissi<strong>on</strong>.<br />

These mechanisms modulate the motor c<strong>on</strong>trol over movement, regulate working memory, and activate<br />

brain-to-body communicati<strong>on</strong> through the neur<strong>on</strong>-immune interface modifying blood cells compositi<strong>on</strong> and<br />

platelet producti<strong>on</strong>. Importantly, stress-associated changes were found in the expressi<strong>on</strong> pattern <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

acetylcholinesterase ACHE gene, which encodes the acetylcholine hydrolyzing enzyme AChE. AChE is not<br />

<strong>on</strong>e, but a combinatorial series <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins having indistinguishable enzymatic activity yet with variant N- and<br />

C-termini due to alternate promoter usage and 3'-alternative splicing. Differentially induced under stress, they<br />

show distinct n<strong>on</strong>-hydrolytic properties, interact with variant-specific protein partners and induce inverse


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

signaling cascades. Surprisingly, transcripti<strong>on</strong>al and post-transcripti<strong>on</strong>al regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> AChE pre-mRNA not<br />

<strong>on</strong>ly protects blood and nerve cells from acute dangers, but may also entail l<strong>on</strong>g-term advantages. Specifically,<br />

causal involvement <str<strong>on</strong>g>of</str<strong>on</strong>g> both AChE and its closely related enzyme butyrylcholinesterase (BChE) in the<br />

progressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Alzheimer's and Parkins<strong>on</strong>'s diseases, anticipates future therapeutic needs <str<strong>on</strong>g>for</str<strong>on</strong>g> drugs targeting<br />

specific cholinesterases or the corresp<strong>on</strong>ding RNA transcripts.<br />

3A_04_S<br />

(poster secti<strong>on</strong> A1, poster board #1, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

KEEPING ALL TOGETHER IN A FRAGMENTED WORLD: THE EVIDENCE BASE<br />

OF THE SALUTOGENIC RESEARCH<br />

M<strong>on</strong>ica Erikss<strong>on</strong>, Bengt Lindström<br />

Folkhälsan <str<strong>on</strong>g>Research</str<strong>on</strong>g> Center, Health Promoti<strong>on</strong> <str<strong>on</strong>g>Research</str<strong>on</strong>g> Program, Paasikivigatan 4,<br />

FIN-00250 Helsinki, Finland<br />

This c<strong>on</strong>ference focuses <strong>on</strong> linking knowledge from different areas <str<strong>on</strong>g>of</str<strong>on</strong>g> stress research particularly exploring<br />

the system level. Thus trying to make sense <str<strong>on</strong>g>of</str<strong>on</strong>g> our fragmented world. Social trends point to a major upset <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the traditi<strong>on</strong>al social structures such as the rupture <str<strong>on</strong>g>of</str<strong>on</strong>g> local and intimate networks because <str<strong>on</strong>g>of</str<strong>on</strong>g> migrati<strong>on</strong> into<br />

urban areas, changed functi<strong>on</strong> and structure <str<strong>on</strong>g>of</str<strong>on</strong>g> family networks, changes in the patterns <str<strong>on</strong>g>of</str<strong>on</strong>g> working life and<br />

communicati<strong>on</strong> and in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> technology. At the same time there are c<strong>on</strong>cerns regarding increasing<br />

violence, alienati<strong>on</strong>, a decreasing physical and mental health. All this makes it difficult to find and run a<br />

coherent life. Besides the research <str<strong>on</strong>g>of</str<strong>on</strong>g> Selye there are other theories and frameworks c<strong>on</strong>tributing to stress<br />

research. One <str<strong>on</strong>g>of</str<strong>on</strong>g> them is the salutogenic theory developed by Aar<strong>on</strong> Ant<strong>on</strong>ovsky in the late 1970s. This<br />

presentati<strong>on</strong> focuses <strong>on</strong> the salutogenic approach such as Aar<strong>on</strong> Ant<strong>on</strong>ovsky <str<strong>on</strong>g>for</str<strong>on</strong>g>mulated his salutogenic<br />

theory “Sense <str<strong>on</strong>g>of</str<strong>on</strong>g> Coherence” (SOC) as a global life orientati<strong>on</strong> to view the world as comprehensible,<br />

manageable and meaningful. He claimed that the way people view their life has a positive influence <strong>on</strong> their<br />

health.[1, 2] The salutogenesis has become an established c<strong>on</strong>cept and c<strong>on</strong>sidered as a paradigm shift from<br />

the pathogenic focus <strong>on</strong> risk factors <str<strong>on</strong>g>for</str<strong>on</strong>g> disease to the salutogenic focus <strong>on</strong> the strengths and determinants<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> health. How do people manage the lack <str<strong>on</strong>g>of</str<strong>on</strong>g> c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> the life The answer was <str<strong>on</strong>g>for</str<strong>on</strong>g>mulated in terms <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

SOC and General Resistance Resources. The SOC is a resource that enables people to manage tensi<strong>on</strong>, to<br />

reflect about their external and internal resources (GRRs), to identify and mobilize them, to promote effective<br />

coping by finding soluti<strong>on</strong>s and resolve tensi<strong>on</strong> in a health promoting manner.<br />

The findings from an <strong>on</strong>going review <str<strong>on</strong>g>of</str<strong>on</strong>g> the salutogenic research 1992-2003 by the authors is presented.[3-7]<br />

The review c<strong>on</strong>sists <str<strong>on</strong>g>of</str<strong>on</strong>g> 458 scientific articles and 13 doctoral theses. To date the SOC questi<strong>on</strong>naire has been<br />

used in at least 33 languages in 32 countries all over the World. The SOC scale seems to be a reliable, valid,<br />

and cross culturally applicable instrument measuring health. SOC tends to increase with age. SOC is str<strong>on</strong>gly<br />

related to perceived good health, especially mental health, and QoL. The str<strong>on</strong>ger the SOC the better the<br />

perceived health in general. SOC seems to have a main, moderating or mediating role in the explanati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

health. It seems to be able to predict health and quality <str<strong>on</strong>g>of</str<strong>on</strong>g> life. The applicability <str<strong>on</strong>g>of</str<strong>on</strong>g> the SOC c<strong>on</strong>cept in<br />

practice <strong>on</strong> both an individual, a group and <strong>on</strong> a societal level is discussed. With the evidence base <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

salutogenic research <strong>on</strong> hands the c<strong>on</strong>clusi<strong>on</strong> is that the Salutogenesis is a valuable approach <str<strong>on</strong>g>for</str<strong>on</strong>g> individuals<br />

and groups in order to build capacity and coherence and healthy societies.<br />

173


23-26 August 2007,<br />

Budapest, Hungary<br />

3A_05_S<br />

(poster secti<strong>on</strong> A1, poster board #2, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

GLYCOBIOLOGY OF STRESS<br />

Olga Gornik, Gordan Lauc<br />

1University <str<strong>on</strong>g>of</str<strong>on</strong>g> Zagreb, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmacy and Biochemistry, Zagreb, Croatia<br />

e-mail: glauc@pharma.hr<br />

Psychological stress is associated with numerous diseases, but molecular mechanisms linking stress to the<br />

development <str<strong>on</strong>g>of</str<strong>on</strong>g> disease are <strong>on</strong>ly starting to be understood. Stress alert is c<strong>on</strong>veyed by horm<strong>on</strong>al signals<br />

throughout the body, yet a particular cell resp<strong>on</strong>se to a horm<strong>on</strong>al signal is not determined by the signal itself,<br />

but by the molecular compositi<strong>on</strong>, energy c<strong>on</strong>tent, and by the physiological role and current status <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

target cell. Stress induced changes in glycoc<strong>on</strong>jugate structures and expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> their receptors lectins<br />

appear to be an important molecular c<strong>on</strong>sequences <str<strong>on</strong>g>of</str<strong>on</strong>g> stress experience. At the moment <strong>on</strong>ly several<br />

fragments <str<strong>on</strong>g>of</str<strong>on</strong>g> the glycobiological mechanisms involved in the physiological resp<strong>on</strong>se to psychological stress<br />

are known, but the complete picture is slowly emerging. Corticosteroids affect activity <str<strong>on</strong>g>of</str<strong>on</strong>g> at least <strong>on</strong>e<br />

glycosyltransferase both in vitro and in vivo. Altered activity <str<strong>on</strong>g>of</str<strong>on</strong>g> glycosyltransferases results in different<br />

carbohydrate structures attached to glycoproteins, and these changes have been dem<strong>on</strong>strated both in<br />

humans and in experimental animals. A change in the carbohydrate structures attached to a glycoprotein is a<br />

well-established way to change its structural and functi<strong>on</strong>al properties, and recently this was shown to be <strong>on</strong>e<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the mechanisms that c<strong>on</strong>trol activity <str<strong>on</strong>g>of</str<strong>on</strong>g> membrane receptors. Although this type <str<strong>on</strong>g>of</str<strong>on</strong>g> glycosylati<strong>on</strong>-mediated<br />

receptor modulati<strong>on</strong> in stress still has to be proven, it is a very interesting hypothesis. On the other hand, new<br />

glycoc<strong>on</strong>jugate structures could also represent novel signals <strong>on</strong> the cell surface that could alter interacti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the cell with neighboring cells. Stress is also known to be associated with the appearance <str<strong>on</strong>g>of</str<strong>on</strong>g> novel lectins that<br />

could be receptors <str<strong>on</strong>g>for</str<strong>on</strong>g> either novel, or also “normal” glycoc<strong>on</strong>jugate structures, translating their structures<br />

into molecular functi<strong>on</strong>s. Although most <str<strong>on</strong>g>of</str<strong>on</strong>g> this is still largely speculative, hopefully more will be known so<strong>on</strong><br />

about the molecular role <str<strong>on</strong>g>of</str<strong>on</strong>g> glycoc<strong>on</strong>jugates, their lectin receptors, and glycosyltransferases in the<br />

physiological resp<strong>on</strong>se to psychological stress.<br />

174


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

3A_01_P<br />

(poster secti<strong>on</strong> A1, poster board #3, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SEVERE STRESS AND ITS IMPLICATIONS: THE NEED TO RECOGNISE CONTRA-<br />

VITAL PATHOPHYSIOLOGICAL RESPONSES<br />

C. P. Arun<br />

Kellogg College, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Ox<str<strong>on</strong>g>for</str<strong>on</strong>g>d, e-mail: arunpeter@yahoo.com<br />

INTRODUCTION For animals in the wild, predati<strong>on</strong>, not old age is the most comm<strong>on</strong> mode <str<strong>on</strong>g>of</str<strong>on</strong>g> death.<br />

Nature appears to provide <str<strong>on</strong>g>for</str<strong>on</strong>g> the almost inevitable event <str<strong>on</strong>g>of</str<strong>on</strong>g> death from some <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> trauma by allowing a<br />

quick final exit. REVIEW OF LITERATURE Our previous work (Arun, 2004) presented a c<strong>on</strong>trarian view<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> pathophysiology, asserting that the catecholamine resp<strong>on</strong>se (that lasts <str<strong>on</strong>g>for</str<strong>on</strong>g> <strong>on</strong>ly 24 hours) in acute stress<br />

plays a deciding role in the survival <str<strong>on</strong>g>of</str<strong>on</strong>g> the animal exposed to severe stress and could actually hasten death if<br />

the situati<strong>on</strong> is hopeless. HYPOTHESIS We propose the use <str<strong>on</strong>g>of</str<strong>on</strong>g> the term ‘c<strong>on</strong>tra-vital’(= deleterious to life)<br />

to qualify acti<strong>on</strong>s taken by an organism’s organ systems to accelerate its own death. Our view is at variance<br />

with c<strong>on</strong>temporary scientific thinking that reflexes exist <strong>on</strong>ly to promote the organism’s survival. From the<br />

point <str<strong>on</strong>g>of</str<strong>on</strong>g> view <str<strong>on</strong>g>of</str<strong>on</strong>g> the individual animal, initiati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the c<strong>on</strong>tra-vital resp<strong>on</strong>se is beneficial when death is<br />

inevitable since it would diminish suffering. From a sociobiology standpoint, a quick death <str<strong>on</strong>g>of</str<strong>on</strong>g> a severely<br />

injured member is advantageous to the herd since rather than a futile wait <str<strong>on</strong>g>for</str<strong>on</strong>g> an irrecoverably injured animal<br />

to recover, the herd can move <strong>on</strong>, accepting the animal’s death as a fact. DISCUSSION The systemic<br />

resp<strong>on</strong>se to sepsis appears to incorporate a c<strong>on</strong>tra-vital reflex and this will be elaborated in this paper. The<br />

recent worldwide initiative ‘Surviving Sepsis’ campaign is a tacit recogniti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the existence <str<strong>on</strong>g>of</str<strong>on</strong>g> c<strong>on</strong>tra-vital<br />

resp<strong>on</strong>ses. CONCLUSIONS Physiological systems appear to have a self-destruct mechanism available to<br />

accept defeat in the face <str<strong>on</strong>g>of</str<strong>on</strong>g> insuperable stress. There is a need <str<strong>on</strong>g>for</str<strong>on</strong>g> the scientific community to mature in its<br />

thinking, fight the taboo <str<strong>on</strong>g>of</str<strong>on</strong>g> studying death and recognize that c<strong>on</strong>tra-vital reflexes are a reality in severe stress.<br />

175


23-26 August 2007,<br />

Budapest, Hungary<br />

176<br />

3B. FRONTIERS IN STRESS RESEARCH<br />

3B_01_P<br />

(poster secti<strong>on</strong> A1, poster board #4, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PSYCHOPHYSIOLOGICAL EFFECTS OF NEUREXAN ® ON STRESS-INDUCED<br />

ELECTROPSYCHOGRAMS. A DOUBLE BLIND, RANDOMIZED, PLACEBO-<br />

CONTROLLED STUDY IN HUMAN VOLUNTEERS<br />

Wilfried Dimpfel<br />

Justus Liebig University Giessen c/o NeuroCode AG, D 35578 Wetzlar, Sportparkstr. 9, Germany<br />

e-mail: w.dimpfel@neurocode-ag.com<br />

Objective. There is evidence <str<strong>on</strong>g>for</str<strong>on</strong>g> the effectiveness <str<strong>on</strong>g>of</str<strong>on</strong>g> natural drugs in treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> anxiety. However, there is a<br />

need <str<strong>on</strong>g>for</str<strong>on</strong>g> further prospective, double blind, randomized, placebo c<strong>on</strong>trolled clinical studies which dem<strong>on</strong>strate<br />

efficacy.<br />

Methods: A preparati<strong>on</strong> c<strong>on</strong>taining (am<strong>on</strong>gst others) low dose extracts <str<strong>on</strong>g>of</str<strong>on</strong>g> passiflora incarnata and avena sativa<br />

(Neurexan ® ) was tested in 30 healthy human volunteers in a randomised, placebo c<strong>on</strong>trolled cross-over trial.<br />

Eligible subjects aged 30 – 60 years were assessed under c<strong>on</strong>diti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> relaxati<strong>on</strong> and under experimental<br />

stress-inducti<strong>on</strong> (calculati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> mathematical tasks with financial reward or punishment according to<br />

per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance). After recording <str<strong>on</strong>g>of</str<strong>on</strong>g> the electropsychogram (electrical activity in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> emoti<strong>on</strong>al<br />

provocati<strong>on</strong>s) frequency analysis using Fast Fourier Trans<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> and analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> source density was<br />

per<str<strong>on</strong>g>for</str<strong>on</strong>g>med 1 to 4 hours after administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 4 tablets verum or placebo. Task dependent increase <str<strong>on</strong>g>of</str<strong>on</strong>g> beta 2<br />

power was used as a surrogate parameter <str<strong>on</strong>g>of</str<strong>on</strong>g> a stress induced anxiety. The same experimental procedure was<br />

repeated <str<strong>on</strong>g>for</str<strong>on</strong>g> each subject <strong>on</strong> the ‘crossed-over’ medicati<strong>on</strong> after an interval <str<strong>on</strong>g>of</str<strong>on</strong>g> at least <strong>on</strong>e week.<br />

Results: In the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> verum, circadian enhancement <str<strong>on</strong>g>of</str<strong>on</strong>g> alpha 2 waves was blunted. Beta 2 power was<br />

statistically significantly lower during the sec<strong>on</strong>d and third hour after administrati<strong>on</strong> in comparis<strong>on</strong> to<br />

placebo. Values returned to baseline values 5 hours after administrati<strong>on</strong>. The preparati<strong>on</strong> was very well<br />

tolerated.<br />

C<strong>on</strong>clusi<strong>on</strong>: Recording <str<strong>on</strong>g>of</str<strong>on</strong>g> electrical activity during a stress-related emoti<strong>on</strong>al situati<strong>on</strong> revealed a smaller<br />

increase <str<strong>on</strong>g>of</str<strong>on</strong>g> beta 2 power under verum compared to placebo. This is indicative <str<strong>on</strong>g>of</str<strong>on</strong>g> better coping with task<br />

related stress whilst <strong>on</strong> Neurexan ® .<br />

3B_03_P<br />

(poster secti<strong>on</strong> A1, poster board #5, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SECOND-HAND STRESS: NEUROBIOLOGICAL EVIDENCE FOR A HUMAN<br />

ALARM PHEROMONE<br />

L.R. Mujica-Parodi, Helmut H. Strey, Blaise Frederick, Robert Savoy, David Cox,<br />

Bosky Ravindranath, Yevgeny Botanov<br />

Laboratory <str<strong>on</strong>g>for</str<strong>on</strong>g> the Study <str<strong>on</strong>g>of</str<strong>on</strong>g> Emoti<strong>on</strong> and Cogniti<strong>on</strong>, Departments <str<strong>on</strong>g>of</str<strong>on</strong>g> Biomedical Engineering and Psychiatry,<br />

St<strong>on</strong>y Brook University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Health Sciences Center T18, St<strong>on</strong>y Brook, NY 11794-8181 USA<br />

Pherom<strong>on</strong>es functi<strong>on</strong> as airborne chemical signals released by an individual into the envir<strong>on</strong>ment that, even<br />

in the absence <str<strong>on</strong>g>of</str<strong>on</strong>g> c<strong>on</strong>scious percepti<strong>on</strong>, affect the physiology or behavior <str<strong>on</strong>g>of</str<strong>on</strong>g> other members <str<strong>on</strong>g>of</str<strong>on</strong>g> the same<br />

species. While almost all clinical research in the area has c<strong>on</strong>centrated <strong>on</strong> human reproductive pherom<strong>on</strong>es,<br />

n<strong>on</strong>-human mammals are known to also possess alarm pherom<strong>on</strong>es, which rapidly transmit warning <str<strong>on</strong>g>of</str<strong>on</strong>g> danger


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

to others <str<strong>on</strong>g>of</str<strong>on</strong>g> the same species via olfacti<strong>on</strong>. The existence <str<strong>on</strong>g>of</str<strong>on</strong>g> alarm pherom<strong>on</strong>es is well-established in<br />

mammals, with animals exposed to odors secreted by acutely stressed c<strong>on</strong>specifics expressing physiological<br />

and behavioral changes that are indistinguishable from their reacti<strong>on</strong>s to predators, including increased neural<br />

activity in the amygdala, analgesia, increased hiding and avoidance, freezing and rearing, hyperthermia, and<br />

air-sampling. Using functi<strong>on</strong>al MRI, we dem<strong>on</strong>strate here the first direct evidence <str<strong>on</strong>g>for</str<strong>on</strong>g> a human alarm<br />

pherom<strong>on</strong>e, with humans showing brain activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the amygdala and hypothalamus—the primary brain<br />

regi<strong>on</strong>s resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> the fear resp<strong>on</strong>se—during inhalati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> sweat taken from a sample <str<strong>on</strong>g>of</str<strong>on</strong>g> first-time<br />

skydivers, with exercise sweat as a c<strong>on</strong>trol. The fMRI subjects were unable to identify the samples, and rated<br />

the odors <str<strong>on</strong>g>for</str<strong>on</strong>g> both c<strong>on</strong>diti<strong>on</strong>s as mild and n<strong>on</strong>-aversive. The results were found <str<strong>on</strong>g>for</str<strong>on</strong>g> breathing the fear sweat,<br />

but not sniffing it, most likely reflecting the fact that the neural circuitry associated with sniffing is explicitly<br />

tied to c<strong>on</strong>scious odor detecti<strong>on</strong>, while pherom<strong>on</strong>es are hypothesized to operate unc<strong>on</strong>sciously. Both males<br />

and females showed equivalent neural activati<strong>on</strong>, thereby ruling out the influence <str<strong>on</strong>g>of</str<strong>on</strong>g> sexual pherom<strong>on</strong>es in<br />

accounting <str<strong>on</strong>g>for</str<strong>on</strong>g> our results. Our findings indicate that there may be a hidden biological comp<strong>on</strong>ent to human<br />

social dynamics, in which emoti<strong>on</strong>al stress is, quite literally, “c<strong>on</strong>tagious.”<br />

3B_04_P<br />

(poster secti<strong>on</strong> A1, poster board #6, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

OVER-EXPRESSION OF THE ESTROGEN RECEPTOR BETA ASSOCIATED<br />

PROTEIN HSP27 IS ATHEROPROTECTIVE<br />

Edward R. O’Brien, Xiaoling Zhoa, Katey Rayner, Y<strong>on</strong>g-Xiang Chen<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Ottawa Heart Institute, CANADA<br />

e-mail: eobrien@ottawaheart.ca<br />

Recently, we discovered that HSP27 is an estrogen receptor beta associated protein that represses estrogen<br />

mediated transcripti<strong>on</strong> (Miller et al, ATVB 2005). Moreover, in human cor<strong>on</strong>ary arteries the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

HSP27 decreases with the progressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> atherosclerotic disease stage and serum levels are >3-fold higher in<br />

normal c<strong>on</strong>trols compared to patients with angiographic evidence <str<strong>on</strong>g>of</str<strong>on</strong>g> cor<strong>on</strong>ary artery disease. Purpose: To<br />

determine if HSP27 is protective against the development <str<strong>on</strong>g>of</str<strong>on</strong>g> atherosclerosis. Methods: Mice over-expressing<br />

human HSP27 (hHSP27) were crossbred with atherosclerosis-pr<strong>on</strong>e apoE null mice (apoE) to yield<br />

hHSP27apoE mice (n=9 female, 5 male) and compared to apoE mice (n=7 female, 6 male) <str<strong>on</strong>g>for</str<strong>on</strong>g> aortic lesi<strong>on</strong><br />

development. All mice were fed a cholesterol supplemented diet <str<strong>on</strong>g>for</str<strong>on</strong>g> 4 weeks and euthanized at age 10 weeks.<br />

Results: All mice were viable, <str<strong>on</strong>g>of</str<strong>on</strong>g> similar body weight and had equally elevated cholesterol levels<br />

(approximately 1100 mg/dL). Quantitative histomorphological studies <str<strong>on</strong>g>of</str<strong>on</strong>g> en face aortic specimens revealed<br />

that the percentage aortic area with overt atherosclerosis was similar in both groups <str<strong>on</strong>g>of</str<strong>on</strong>g> male mice. However,<br />

am<strong>on</strong>gst female mice there was a 41% reducti<strong>on</strong> in atherosclerotic lesi<strong>on</strong> area in the hHSP27apoE vs. apoE<br />

mice (p


23-26 August 2007,<br />

Budapest, Hungary<br />

178<br />

3B_05_P<br />

(poster secti<strong>on</strong> A1, poster board #7, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CHARACTERIZATION OF AUTONOMIC NERVOUS SYSTEM (ANS) STABILITY<br />

AND VARIATIONS BY THE ANS SPOT<br />

Markku Paloheimo<br />

Eye Hospital, Helsinki University Hospital, Box 220, 00029 HUS, Finland, and<br />

GE Medical, <str<strong>on</strong>g>Research</str<strong>on</strong>g>, Helsinki, Finland<br />

e-mail: markku.paloheimo@hus.fi<br />

The aut<strong>on</strong>omic nervous system enables all <str<strong>on</strong>g>of</str<strong>on</strong>g> our body systems to operate in an external envir<strong>on</strong>ment that is<br />

both physically and emoti<strong>on</strong>ally challenging. A Google search using words “stress aut<strong>on</strong>omic nervous<br />

system” yielded 43,500 hits. Am<strong>on</strong>g the tests <str<strong>on</strong>g>of</str<strong>on</strong>g> parasympathetic cardiovagal regulati<strong>on</strong> include heart rate<br />

analysis, heart rate variati<strong>on</strong> with deep breathing, variati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> beat-to-beat blood pressure and heart rate (HR),<br />

and the Valsalva ratio. Tests <str<strong>on</strong>g>of</str<strong>on</strong>g> sympathetic adrenergic vascular regulati<strong>on</strong> include blood pressure analysis in<br />

various situati<strong>on</strong>s, the Valsalva manoeuvre, sustained handgrip, mental stress, and cold water immersi<strong>on</strong>.<br />

Tests <str<strong>on</strong>g>of</str<strong>on</strong>g> sympathetic cholinergic sudomotor functi<strong>on</strong> include the sympathetic skin resp<strong>on</strong>se, quantitative<br />

sudomotor ax<strong>on</strong> reflex test, sweat box testing, and quantificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> sweat imprints.<br />

We have developed a method to display neural or horm<strong>on</strong>al β-adrenergic resp<strong>on</strong>ses <str<strong>on</strong>g>of</str<strong>on</strong>g> heart rate and<br />

peripheral cholinergic neural vascular resp<strong>on</strong>ses <str<strong>on</strong>g>of</str<strong>on</strong>g> finger pulse amplitude to characterize the aut<strong>on</strong>omic state<br />

and variati<strong>on</strong>s during general anaesthesia and sedati<strong>on</strong>. This was d<strong>on</strong>e by displaying pulse-to-pulse intervals<br />

(in ms) <strong>on</strong> the X-axis and subsequent plethysmographic pulse wave amplitudes <strong>on</strong> the Y-axis (see Figures).<br />

The data is obtained from Datex-Ohmeda m<strong>on</strong>itors via serial interface and the source <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

plethysmographic signal is a comm<strong>on</strong> pulse oximetry sensor <strong>on</strong> the subject’s finger. The number <str<strong>on</strong>g>of</str<strong>on</strong>g> traced<br />

observati<strong>on</strong>s can be set (Number <str<strong>on</strong>g>of</str<strong>on</strong>g> points). Six most recent data is shown as red dots and the older <strong>on</strong>es in<br />

white, which <str<strong>on</strong>g>for</str<strong>on</strong>g>ms a trend. Low sympathetic activity is characterized by high pulse amplitudes and low heart<br />

rates (l<strong>on</strong>g intervals). Changes in aut<strong>on</strong>omic activity is shown as a movement <str<strong>on</strong>g>of</str<strong>on</strong>g> the spot; vagal activati<strong>on</strong> is<br />

seen as pulse rate slowing and increase in sympathetic activity is heralded by a decrease in pulse<br />

plethysmographic amplitude and increase in heart rate. During anticholinergic and β-blocking medicati<strong>on</strong>,<br />

heart rate variability is decreased, but these drugs do not affect pulse amplitude variability.<br />

This n<strong>on</strong>invasive method has been used to follow aut<strong>on</strong>omic state and resp<strong>on</strong>ses during general anaesthesia,<br />

c<strong>on</strong>scious sedati<strong>on</strong>, intensive care and psychological testing. A Surgical Stress Index (see Google) displayed<br />

<strong>on</strong> a scale <str<strong>on</strong>g>of</str<strong>on</strong>g> 0 -100 is under development.<br />

3B_08_P<br />

(poster secti<strong>on</strong> A1, poster board #8, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

O-GLCNAC, A NEW PARADIGM FOR REGULATING STRESS-SIGNALING<br />

NETWORKS<br />

Natasha E. Zachara 1 , Henrik Molina 1,2 , Akilesh Pandey 1,2 , Gerald W. Hart 1<br />

1. Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biological Chemistry, Johns Hopkins University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Baltimore, MD 21218.<br />

2. McKusick-Nathans Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Genetic Medicine, Johns Hopkins University, Baltimore, MD 21287.<br />

e-mail: nzachara@jhmi.edu<br />

In resp<strong>on</strong>se to numerous <str<strong>on</strong>g>for</str<strong>on</strong>g>ms <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular stress, levels <str<strong>on</strong>g>of</str<strong>on</strong>g> the O-GlcNAc protein modificati<strong>on</strong> are elevated<br />

rapidly and dynamically <strong>on</strong> myriad nucleocytoplasmic proteins. Increased O-GlcNAc levels are linked to<br />

stress tolerance in both cell culture and animal models, suggesting that this phenomen<strong>on</strong> is a survival


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> cells. Elevated levels <str<strong>on</strong>g>of</str<strong>on</strong>g> O-GlcNAc have been linked to changes in Hsp70 and Hsp40 protein<br />

levels, as well as altered capacitive calcium entry, suggesting at least two mechanisms by which O-GlcNAc<br />

may protect cells. These, and other data, suggest a new paradigm in the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> stress mediated signal<br />

transducti<strong>on</strong> pathways, further investigati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> which will elucidate new roles <str<strong>on</strong>g>for</str<strong>on</strong>g> O-GlcNAc in diverse<br />

clinical and cellular settings such as Ischemia-reperfusi<strong>on</strong> injury, neurodegenerative diseases and aging. In<br />

order to better understand the mechanism(s) by which O-GlcNAc modulates the cellular resp<strong>on</strong>se to stress,<br />

we have determined which proteins are dynamically O-GlcNAc modified in resp<strong>on</strong>se to cellular stress using<br />

Stable Isotope Labeling with Amino Acids in Cell Culture. We have identified 27 O-GlcNAc modified<br />

proteins, <str<strong>on</strong>g>of</str<strong>on</strong>g> which 19 exhibited an increase in O-GlcNAc protein modificati<strong>on</strong> in resp<strong>on</strong>se to heat shock.<br />

These proteins are predominantly involved in signal transducti<strong>on</strong>, transcripti<strong>on</strong> and vesicle transport.<br />

Currently, we are examining the role <str<strong>on</strong>g>of</str<strong>on</strong>g> glycosylati<strong>on</strong> <strong>on</strong> these proteins, and determining how this c<strong>on</strong>tributes<br />

to O-GlcNAc mediated stress tolerance. Under a licensing agreement between Covance and Johns Hopkins<br />

University, GWH receives a share <str<strong>on</strong>g>of</str<strong>on</strong>g> royalties <strong>on</strong> sales <str<strong>on</strong>g>of</str<strong>on</strong>g> CTD110.6.<br />

3B_09_P<br />

(poster secti<strong>on</strong> A1, poster board #9, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CHROMOSOME ABERRATIONS, DNA REPARATION, MICRONUCLEI,<br />

PROLIFERATION AND APOPTOSIS IN LYMPHOCYTES’ CULTURES OF HUMAN<br />

UNDER EMOTIONAL STRESS<br />

F. I. Ingel<br />

A.N.Sysin State <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Human Ecology and Envir<strong>on</strong>mental Hygiene <str<strong>on</strong>g>of</str<strong>on</strong>g> Russian Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical<br />

Sciences, Moscow, Russia<br />

e-mail: faina_ingel@uk2.net<br />

Our experiments <strong>on</strong> mice (Ingel et al, 1993 - 1999) dem<strong>on</strong>strated that emoti<strong>on</strong>al stress (immobilizati<strong>on</strong>)<br />

induced chromosome aberrati<strong>on</strong>s in b<strong>on</strong>e marrow and thymus in comm<strong>on</strong> dynamics during 25 days, and,<br />

additi<strong>on</strong>ally, increased susceptibility <str<strong>on</strong>g>of</str<strong>on</strong>g> genome <str<strong>on</strong>g>of</str<strong>on</strong>g> these cells to standard mutagen cyclophosphamide. So we<br />

supposed that the same effects may be induced with human emoti<strong>on</strong>al stress in stage <str<strong>on</strong>g>of</str<strong>on</strong>g> disadaptati<strong>on</strong>. Then<br />

303 adults and 248 children from different industrial towns <str<strong>on</strong>g>of</str<strong>on</strong>g> Russia and settlements in Kazakhstan were<br />

tested <str<strong>on</strong>g>for</str<strong>on</strong>g> emoti<strong>on</strong>al stress expressi<strong>on</strong> (Cholms-Ray’, Taylor’, Akkles’ scales – <str<strong>on</strong>g>for</str<strong>on</strong>g> adults and Luscher’ test –<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> children) in parallel with medical and biochemical (blood, urine) studies, PCR analysis (GSTM1, GSTT1,<br />

CYP1A1, CYP1A2, PON 54) and cytogenetical investigati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> peripheral blood cell in cultures (without<br />

and with additi<strong>on</strong>al load <str<strong>on</strong>g>of</str<strong>on</strong>g> mutagen in vitro). Results dem<strong>on</strong>strated that level <str<strong>on</strong>g>of</str<strong>on</strong>g> stress expressi<strong>on</strong> positively<br />

correlated with: a) industrial c<strong>on</strong>tacts with toxic and mutagenic compounds or residing in z<strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> ecological<br />

disaster (Aral Sea Basin) as well as blood c<strong>on</strong>taminati<strong>on</strong> (dioxin, r = 0,75-0,87; p≤0,001); b) frequencies <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

chromosome aberrati<strong>on</strong>s and micr<strong>on</strong>uclei in cultivated blood lymphocytes (r = 0,74-0,78; p≤0,001); c)<br />

coefficient <str<strong>on</strong>g>of</str<strong>on</strong>g> UV-induced DNA reparati<strong>on</strong> (r = 0,86; p≤0,00001); d) radiosensitivity <str<strong>on</strong>g>of</str<strong>on</strong>g> cells in culture<br />

(r=0,081; p=0,001); e) proliferati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cells in cultures <str<strong>on</strong>g>of</str<strong>on</strong>g> peripheral blood (r = 0,75; p=0,000001); f) speed<br />

and symmetry <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular divisi<strong>on</strong> (r=0,56 – 0,62; p= 0,0001); g) apoptosis <str<strong>on</strong>g>of</str<strong>on</strong>g> cells in culture <str<strong>on</strong>g>of</str<strong>on</strong>g> adults (r =<br />

0,45; p=0,002). Negative correlati<strong>on</strong> between stress expressi<strong>on</strong> and apoptosis <str<strong>on</strong>g>of</str<strong>on</strong>g> lymphocytes in culture was<br />

detected <str<strong>on</strong>g>for</str<strong>on</strong>g> children (r=-0,42; p=0,003). Additi<strong>on</strong>ally, disadaptative stress expressi<strong>on</strong> was significantly<br />

associated with mutati<strong>on</strong> in tested genes <str<strong>on</strong>g>of</str<strong>on</strong>g> detoxificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> xenobiotics. C<strong>on</strong>clusi<strong>on</strong>s: the most <str<strong>on</strong>g>of</str<strong>on</strong>g> results <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

human and animals were identical - emoti<strong>on</strong>al stress induced mutati<strong>on</strong>s and increased susceptibility <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

genome to envir<strong>on</strong>mental mutagens. So, chr<strong>on</strong>ic emoti<strong>on</strong>al disadaptati<strong>on</strong> shows the same cytogenetic<br />

properties as full carcinogen.<br />

179


23-26 August 2007,<br />

Budapest, Hungary<br />

3C. CHAPERONE NETWORKS AND STRESS PROTEOMICS<br />

3C_01_P<br />

(poster secti<strong>on</strong> A1, poster board #10, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE IMPACT OF THE YEAST RAC-SSB CO-TRANSLATIONAL SYSTEM OF<br />

CHAPERONES ON THE CELLULAR ACTIVITY OF MUTATED PROTEINS<br />

Katarzyna Tomala, Ryszard Kor<strong>on</strong>a<br />

Jagiell<strong>on</strong>ian University, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Envir<strong>on</strong>mental Sciences, Address: Gr<strong>on</strong>ostajowa 7, 30-387 Kraków, Poland;<br />

e-mail: tomala@eko.uj.edu.pl<br />

A large fracti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> sp<strong>on</strong>taneous mutati<strong>on</strong>s lead to destabilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> protein structure. This results in exposure<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> hydrophobic patches <strong>on</strong> the protein surface. It is thus very likely that mutant proteins are more frequently<br />

bound by the molecular chaper<strong>on</strong>es. The subsequent processing <str<strong>on</strong>g>of</str<strong>on</strong>g> excessively unstable proteins is poorly<br />

known. The more frequent binding by the chaper<strong>on</strong>es may help to overcome the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> mutati<strong>on</strong> and<br />

secure functi<strong>on</strong>ing <str<strong>on</strong>g>of</str<strong>on</strong>g> the protein in the cell. Alternatively, the excessive activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the chaper<strong>on</strong>es<br />

c<strong>on</strong>centrated <strong>on</strong> a defective polypeptide may c<strong>on</strong>stitute a signal that it has to be degraded. Our research was<br />

focused <strong>on</strong> the RAC-Ssb system <str<strong>on</strong>g>of</str<strong>on</strong>g> co-translati<strong>on</strong>al chaper<strong>on</strong>es in yeast. We asked whether its role is to<br />

uphold folding efficiency <str<strong>on</strong>g>of</str<strong>on</strong>g> mutated proteins or rather sort them out and exclude from the cellular<br />

metabolism. We prepared a library <str<strong>on</strong>g>of</str<strong>on</strong>g> mutagenized ADE2 gene whose inactivity or malfuncti<strong>on</strong>ing results in<br />

red pigmentati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the yeast col<strong>on</strong>ies. We then screened <str<strong>on</strong>g>for</str<strong>on</strong>g> mutati<strong>on</strong>s producing red color in strains in<br />

which RAC-Ssb was inactivated by gene deleti<strong>on</strong>. No such synthetic phenotypes were found indicating that<br />

mutati<strong>on</strong>al destabilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Ade2p is not aggravated in the absence <str<strong>on</strong>g>of</str<strong>on</strong>g> the chaper<strong>on</strong>e activity. In a sec<strong>on</strong>d<br />

screen, we first identified an array <str<strong>on</strong>g>of</str<strong>on</strong>g> mutati<strong>on</strong>ally destabilized variants <str<strong>on</strong>g>of</str<strong>on</strong>g> Ade2p proteins. This was marked<br />

by their thermosensitivity. In case <str<strong>on</strong>g>of</str<strong>on</strong>g> all mutants, their growth <strong>on</strong> media lacking adenine was better when the<br />

RAC-Ssb was inactivated. This indicates that the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> chaper<strong>on</strong>es leads to a decline <str<strong>on</strong>g>of</str<strong>on</strong>g> a cellular pool <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

active Ade2(ts) molecules. Current experiments show that the defective protein is present in the cell in <str<strong>on</strong>g>for</str<strong>on</strong>g>m<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> aggregates. We suggest that the RAC-Ssb can have a role in sequestering defective proteins from the<br />

cytosol and restricting their unc<strong>on</strong>trolled interacti<strong>on</strong>s with other elements <str<strong>on</strong>g>of</str<strong>on</strong>g> the cellular metabolism.<br />

3C_02_P<br />

(poster secti<strong>on</strong> A1, poster board #11, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

MOLECULAR STUDY OF ACID SHOCK PROTEINS IN A DIARRHEAGENIC<br />

ENTEROAGGREGATIVE ESCHERICHIA COLI (EAEC)<br />

Punit Kaur, Puneet Badesha<br />

Postgraduate Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical educati<strong>on</strong> & <str<strong>on</strong>g>Research</str<strong>on</strong>g>, India<br />

e-mail: puneetbadesha@yahoo.com<br />

EAEC is an emerging pathotype <str<strong>on</strong>g>for</str<strong>on</strong>g> persistent pediatric diarrhea characterized by an aggregative adherence<br />

(AA) pattern to HEp-2 cells. Its pathogenesis is poorly understood. Like other enterics, it faces acidic<br />

c<strong>on</strong>diti<strong>on</strong>s in stomach and small intestine. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, the adaptive strategies like acid tolerance resp<strong>on</strong>ses<br />

(ATRs) are induced to ensure its survival during the disease progressi<strong>on</strong>. Outer membrane proteins (OMPs)<br />

are the key molecules that interface cell with the envir<strong>on</strong>ment and are important in adhesi<strong>on</strong>, invasi<strong>on</strong> and<br />

intracellular survival <str<strong>on</strong>g>of</str<strong>on</strong>g> pathogens in the host. Most notable pH-resp<strong>on</strong>se regulators like Fur, PhoP, OmpR<br />

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2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

and RpoS are reported and the rpoS, an alternative sigma factor is well known <str<strong>on</strong>g>for</str<strong>on</strong>g> the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ATRs in<br />

S. typhimurium and EHEC.<br />

ATR in EAEC was studied by growing cells at variable pH range (2.0-7.4). Cell survival was observed till pH<br />

4.0 but the preadapted cells (adapted at mild acidic pH 5.0 <str<strong>on</strong>g>for</str<strong>on</strong>g> 2 hours), survived two fold better than that <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the n<strong>on</strong>-adapted cells (pH 4.0). Phenotypic characterizati<strong>on</strong> under acid stress c<strong>on</strong>diti<strong>on</strong>s was checked in vitro.<br />

At pH 4.0, clump <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> was totally abolished. The bi<str<strong>on</strong>g>of</str<strong>on</strong>g>ilm score was


23-26 August 2007,<br />

Budapest, Hungary<br />

182<br />

3C_04_P<br />

(poster secti<strong>on</strong> A1, poster board #13, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

LOCAL AND SYSTEMIC CHANGES IN PROTEIN EXPRESSION FOLLOWING<br />

BIOTIC STRESS IN EUROPEAN BEACH (FAGUS SYLVATICA)<br />

Cristina-Maria Valcu, Katja Schlink<br />

Technical University <str<strong>on</strong>g>of</str<strong>on</strong>g> Munich, Secti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Forest Genetics, Am Hochanger 13, D-85354 Freising,<br />

e-mail: valcu@wzw.tum.de<br />

Pathogen and herbivore attack induced changes in protein expressi<strong>on</strong> patterns were investigated at proteome<br />

level in European beach (Fagus sylvatica) saplings. Two model systems were employed: (1) infecti<strong>on</strong> with the<br />

root pathogen Phytophthora citricola which induces root necrosis and plant wilting and (2) wounding as elicitor<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the resp<strong>on</strong>se to herbivore attack. The plant defence resp<strong>on</strong>se was investigated at both local and systemic<br />

level. Protein expressi<strong>on</strong> patterns were characterised <str<strong>on</strong>g>for</str<strong>on</strong>g> root and leaf samples by means <str<strong>on</strong>g>of</str<strong>on</strong>g> two-dimensi<strong>on</strong>al<br />

electrophoresis at three hours after wounding and at different stages <str<strong>on</strong>g>of</str<strong>on</strong>g> root infecti<strong>on</strong>. Infecti<strong>on</strong> experiments<br />

were per<str<strong>on</strong>g>for</str<strong>on</strong>g>med in two experimental setups involving in vitro infecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> saplings as well as infecti<strong>on</strong>s in soil.<br />

Within the first system we investigated both local and systemic resp<strong>on</strong>se to pathogen infecti<strong>on</strong> while the latter<br />

experiments were used <str<strong>on</strong>g>for</str<strong>on</strong>g> the study <str<strong>on</strong>g>of</str<strong>on</strong>g> systemic resp<strong>on</strong>se in c<strong>on</strong>diti<strong>on</strong>s close to the natural <strong>on</strong>es. Wounding<br />

experiments were carried out in similar c<strong>on</strong>diti<strong>on</strong>s in order to enable the direct comparis<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> plant reacti<strong>on</strong><br />

to the two types <str<strong>on</strong>g>of</str<strong>on</strong>g> stress. The local resp<strong>on</strong>se to wounding was characterised both <str<strong>on</strong>g>for</str<strong>on</strong>g> roots and leaves <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

plants grown in liquid system, while the systemic resp<strong>on</strong>se was studied in leaf samples from plants grown in<br />

liquid culture or in soil. Protein spots up-regulated or down-regulated in resp<strong>on</strong>se to infecti<strong>on</strong> or wounding<br />

were further identified by means <str<strong>on</strong>g>of</str<strong>on</strong>g> mass spectrometry. We were able to identify several proteins specifically<br />

regulated in resp<strong>on</strong>se to pathogen or herbivore attack. Additi<strong>on</strong>ally, a significant overlap <str<strong>on</strong>g>of</str<strong>on</strong>g> the two defence<br />

pathways known to exist in plants was c<strong>on</strong>firmed.<br />

3C_05_P<br />

(poster secti<strong>on</strong> A1, poster board #14, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

A PHYSIOLOGICAL AND HOMOLOGY BASED PROTEOMICS<br />

CHARACTERIZATION OF SALINITY-INDUCED GLYCOLATE METABOLISM IN<br />

NITROGEN-FIXING ANABAENA DOLIOLUM<br />

Ashish Kumar Srivastava 1,2 , Lal Chand Rai 2<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Botany, Mizoram University, Aizawl-796001, India<br />

2Molecular Biology Secti<strong>on</strong>, Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Algal Biology, Center <str<strong>on</strong>g>of</str<strong>on</strong>g> Advanced Study in Botany, Banaras Hindu<br />

University, Varanasi-221005, India<br />

This study provides first hand physiological and proteomic in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <strong>on</strong> the occurrence <str<strong>on</strong>g>of</str<strong>on</strong>g> glycolate<br />

metabolism in fresh water nitrogen fixing cyanobacterium Anabaena doliolum LCR1 under salinity. A reducti<strong>on</strong><br />

in growth, O 2 -evoluti<strong>on</strong>, photosystem II, carb<strong>on</strong> fixati<strong>on</strong>, chlorophyll and NADPH c<strong>on</strong>tent and an increase<br />

in intracellular Na + , photosystem I, respirati<strong>on</strong> and ATP c<strong>on</strong>tent was observed after 150 mM NaCl treatment<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> 1 and 24h. In view <str<strong>on</strong>g>of</str<strong>on</strong>g> the alterati<strong>on</strong> in the above menti<strong>on</strong>ed physiological parameters, the proteomic study<br />

(using two dimensi<strong>on</strong>al gel electrophoresis and MALDI-TOF MS/MS) was d<strong>on</strong>e. Of the 201 protein spots in<br />

c<strong>on</strong>trol <strong>on</strong>ly 90 sustained the high dose <str<strong>on</strong>g>of</str<strong>on</strong>g> salinity. Of these protein spots, six showed significant and<br />

reproducible alterati<strong>on</strong>s, which showed homology with Fe-SOD, SOD, phycocyanin alpha chain, EF-Tu,<br />

RuBisCO and phosphoribulokinase <str<strong>on</strong>g>of</str<strong>on</strong>g> Nostoc PCC7120. Thus the physiological and proteomics data jointly


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

suggests that the organism is deriving carb<strong>on</strong> from a different source; <strong>on</strong>e such source could be the glycolate<br />

metabolism. To examine the alternate route <str<strong>on</strong>g>of</str<strong>on</strong>g> carb<strong>on</strong>, phosphoglyceric acid and the intermediates <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

glycolate pathway were measured. Evidence <strong>on</strong> the operati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> this pathway came <str<strong>on</strong>g>for</str<strong>on</strong>g>m the: (i) enhanced<br />

accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> phosphoglyceric acid and inhibited phosphoenolpyruvate carboxykinase activity, (ii) the<br />

increased activity <str<strong>on</strong>g>of</str<strong>on</strong>g> enzyme glycolate oxidase, and (iii) increase in the intermediates <str<strong>on</strong>g>of</str<strong>on</strong>g> the C2 glycolate cycle<br />

(photorespirati<strong>on</strong>) i.e. glycine, serine and amm<strong>on</strong>ia. In view <str<strong>on</strong>g>of</str<strong>on</strong>g> the producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> amm<strong>on</strong>ia, the possible<br />

generati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> serine from PGA through n<strong>on</strong>-photorespiratory pathway is ruled out. However, this pathway<br />

generates peroxide whose scavenging enzymes are sensitive to salt stress; there<str<strong>on</strong>g>for</str<strong>on</strong>g>e the organism fails to<br />

acclimatize to salt. Taking recourse to the above, occurrence <str<strong>on</strong>g>of</str<strong>on</strong>g> salt-induced glycolate metabolism<br />

(photorespirati<strong>on</strong>) in A. doliolum is proposed.<br />

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2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

MODULE 4<br />

PLANT STRESS<br />

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SYMPOSIA<br />

• 4A. Plant oxidative stress (August 25th Saturday morning)<br />

(Organizer and chair: Imre Vass)<br />

Poster: August 24 th Friday, Poster secti<strong>on</strong> B2, Building B<br />

• 4B. Biotic stress and disease resistance in plants (August 25th Saturday afterno<strong>on</strong>)<br />

(Organizer and chair: Terry Graham)<br />

Poster: August 24 th Friday, Poster secti<strong>on</strong> B2, Building B<br />

• 4C. Stress signaling in plants (August 24th Friday afterno<strong>on</strong>)<br />

(Organizer and chair: Csaba K<strong>on</strong>cz)<br />

Poster: August 24 th Friday, Poster secti<strong>on</strong> B2, Building B<br />

• 4D. L<strong>on</strong>g-term acclimati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> plants and ecosystems to elevated CO2 (August 26th<br />

Sunday morning)<br />

(Organizers and chairs: Zoltán Tuba, and Michal V. Marek)<br />

Poster: August 24 th Friday, Poster secti<strong>on</strong> B2, Building B<br />

POSTER SYMPOSIA<br />

• 4E. Abiotic stress<br />

August 24 th Friday, Poster secti<strong>on</strong> B2, Building B<br />

• 4F. Drought and desiccati<strong>on</strong> stresses in ecosystems<br />

August 24 th Friday, Poster secti<strong>on</strong> B2, Building B<br />

• 4G. Heavy metal stress<br />

August 24 th Friday, Poster secti<strong>on</strong> B2, Building B<br />

186


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

4A. PLANT OXIDATIVE STRESS<br />

(IMRE VASS)<br />

4A_01_S<br />

EVOLUTION OF DESICCATION TOLERANCE: GENOMIC ASPECTS<br />

Dorothea Bartels<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> B<strong>on</strong>n<br />

The Scrophulariaceae are <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the most diverse plant families and have been shown to be polyphyletic.<br />

Several Scrophulariaceae which bel<strong>on</strong>g to the genera Craterostigma and Lindernia desiccati<strong>on</strong> tolerant and are<br />

there<str<strong>on</strong>g>for</str<strong>on</strong>g>e <str<strong>on</strong>g>of</str<strong>on</strong>g> particular interest. Craterostigma plantagineum has been used extensively to identify molecular<br />

mechanisms, which are involved in the acquisiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> desiccati<strong>on</strong> tolerance. These studies revealed a large<br />

number <str<strong>on</strong>g>of</str<strong>on</strong>g> genes which are up-regulated in resp<strong>on</strong>se to dehydrati<strong>on</strong>. The genes can approximately be divided<br />

into two groups. One group encodes proteins which are directly involved in protecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular structures.<br />

The other group <str<strong>on</strong>g>of</str<strong>on</strong>g> genes encodes genes which are involved in regulating the resp<strong>on</strong>se to dehydrati<strong>on</strong>. The<br />

dehydrati<strong>on</strong>/rehydrati<strong>on</strong> cycle is also characterized by a massive sugar c<strong>on</strong>versi<strong>on</strong> from octulose to sucrose.<br />

C. plantagineum bel<strong>on</strong>gs to the Linderniae which comprises desiccati<strong>on</strong> tolerant and n<strong>on</strong>-desiccati<strong>on</strong> tolerant<br />

plants. Using molecular markers a phylogenetic tree was c<strong>on</strong>structed to determine the molecular relati<strong>on</strong>ships<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the different plant species. It appears that desiccati<strong>on</strong> tolerant plants cluster together. Comparative studies<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> these plant species identified c<strong>on</strong>served genomic structures and very recently evolved mobile signals in<br />

stress-relevant genes.<br />

4A_02_S<br />

FUNCTIONS OF TOCOPHEROLS AND OTHER LIPID-SOLUBLE ANTIOXIDANTS<br />

IN PLANTS UNDER LIGHT- OR METAL-INDUCED OXIDATIVE STRESS<br />

Michel Havaux 1 , Valérie Collin 1 , Pascal Rey 1 , Christian Triantaphylidès 1 , Michel Matringe 2<br />

1CEA/Cadarache, IBEB/SBVME, Laboratoire d’Ecophysiologie Moléculaire des Plantes,<br />

F-13108 Saint-Paul-lez-Durance, France; e-mail: michel.havaux@cea.fr<br />

2CEA/Grenoble, iRTSV, Laboratoire de Physiologie Cellulaire Végétale, F-38054 Grenoble, France<br />

The term ‘vitamin E’ describes the beneficial biological activity <str<strong>on</strong>g>of</str<strong>on</strong>g> a group <str<strong>on</strong>g>of</str<strong>on</strong>g> structurally related compounds,<br />

the tocochromanols, in animals and humans. Those compounds are composed <str<strong>on</strong>g>of</str<strong>on</strong>g> a chromanol head group<br />

and a prenyl side chain. Natural vitamin E includes four tocopherols and four tocotrienols, which are<br />

synthesized exclusively by oxygenic photosynthetic organisms. In leaves <str<strong>on</strong>g>of</str<strong>on</strong>g> vascular plants, α-tocopherol is<br />

the predominant <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> vitamin E. A detailed analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> tocochromanol distributi<strong>on</strong> in chloroplasts isolated<br />

from young tobacco leaves showed that α-tocopherol is predominantly located in the thylakoid membranes.<br />

The protective role <str<strong>on</strong>g>of</str<strong>on</strong>g> vitamin E and the functi<strong>on</strong>al interacti<strong>on</strong>s between vitamin E and other plastid<br />

antioxidants (e.g. xanthophyll carotenoids) were studied using Arabidopsis and tobacco mutant/transgenic<br />

plants that lack or over-accumulate vitamin E c<strong>on</strong>stituents and/or carotenoids. This genetic approach was<br />

combined with the use <str<strong>on</strong>g>of</str<strong>on</strong>g> new biochemical and biophysical methods that allow characterizati<strong>on</strong>,<br />

quantificati<strong>on</strong> and imaging <str<strong>on</strong>g>of</str<strong>on</strong>g> lipid peroxidati<strong>on</strong> in vivo. Both tocopherols and tocotrienols were found to<br />

protect thylakoid membranes against photooxidative stress. We also found that vitamin E and the<br />

xanthophyll zeaxanthin have overlapping functi<strong>on</strong>s, with lack <str<strong>on</strong>g>of</str<strong>on</strong>g> vitamin E being compensated by an<br />

increased level <str<strong>on</strong>g>of</str<strong>on</strong>g> zeaxanthin and vice versa. Lack <str<strong>on</strong>g>of</str<strong>on</strong>g> both compounds resulted in a very photosensitive<br />

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phenotype. Vitamin E was also found to be essential <str<strong>on</strong>g>for</str<strong>on</strong>g> tolerance <str<strong>on</strong>g>of</str<strong>on</strong>g> Arabidopsis towards oxidative stress<br />

induced by stress c<strong>on</strong>diti<strong>on</strong>s different from high light, such as heavy metals.<br />

4A_03_S<br />

THE GENETIC BASIS OF SINGLET OXYGEN-MEDIATED SIGNALING OF STRESS<br />

RESPONSES IN PLANTS<br />

Rasa Meskauskiene, Klaus Apel<br />

Swiss Federal Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Technology (ETH Zurich), Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Sciences, Universitaetstr.2, CH - 8092<br />

Zurich, Switzerland<br />

e-mail: kapel@ethz.ch<br />

The evoluti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> aerobic metabolic processes such as respirati<strong>on</strong> and photosynthesis unavoidably leads to the<br />

producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> reactive oxygen species (ROS) in mitoch<strong>on</strong>dria, chloroplasts and peroxisomes. A comm<strong>on</strong><br />

feature am<strong>on</strong>g the different ROS types is their capacity to cause oxidative damage by inactivating e.g.<br />

proteins, nucleic acids and lipids. These cytotoxic properties explain the evoluti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> complex arrays <str<strong>on</strong>g>of</str<strong>on</strong>g> ROS<br />

scavengers. In plants, chloroplasts and peroxisomes are the major sites <str<strong>on</strong>g>of</str<strong>on</strong>g> ROS producti<strong>on</strong>. Various abiotic<br />

stress c<strong>on</strong>diti<strong>on</strong>s may limit the ability <str<strong>on</strong>g>of</str<strong>on</strong>g> a plant to use light energy <str<strong>on</strong>g>for</str<strong>on</strong>g> photosynthesis. Under such stress<br />

c<strong>on</strong>diti<strong>on</strong>s hyper-reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the photosynthetic electr<strong>on</strong> transport chain and photo-inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

photosynthesis may occur even at moderate light intensities, <str<strong>on</strong>g>of</str<strong>on</strong>g>ten causing damages that have been<br />

interpreted as unavoidable c<strong>on</strong>sequences <str<strong>on</strong>g>of</str<strong>on</strong>g> injuries inflicted up<strong>on</strong> plants by toxic levels <str<strong>on</strong>g>of</str<strong>on</strong>g> ROS. However,<br />

this paradigm needs to be modified. Stress resp<strong>on</strong>ses triggered by ROS are not <strong>on</strong>ly due to physicochemical<br />

damages but may also be caused by the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> genetically determined stress resp<strong>on</strong>se programs. We’ll<br />

present results <str<strong>on</strong>g>of</str<strong>on</strong>g> our work showing that ROS may act as signals during stress whose specificities seem to<br />

depend <strong>on</strong> the chemical identity <str<strong>on</strong>g>of</str<strong>on</strong>g> a given ROS and its intracellular site <str<strong>on</strong>g>of</str<strong>on</strong>g> generati<strong>on</strong>.<br />

4A_04_S<br />

MOLECULAR MECHANISMS OF PHOTOINHIBITION OF PHOTOSYSTEM II<br />

Imre Vass, Krisztián Cser, Otilia Cheregi<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Biology, Biological <str<strong>on</strong>g>Research</str<strong>on</strong>g> Center, Hungarian Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences,<br />

6726 Szeged, Temesvári krt. 62, Hungary<br />

Corresp<strong>on</strong>ding author: Imre Vass. Ph<strong>on</strong>e: +36-62-599-700, Fax: +36-62-433-434, e-mail: imre@brc.hu<br />

Light-induced decline <str<strong>on</strong>g>of</str<strong>on</strong>g> photosynthetic activity, generally called as photoinhibiti<strong>on</strong>, is a general phenomen<strong>on</strong><br />

in all oxygenic photosynthetic organisms under c<strong>on</strong>diti<strong>on</strong>s when the metabolic processes can not keep up<br />

with the electr<strong>on</strong> flow produced by the primary photoreacti<strong>on</strong>s. Although light-induced damage occurs in all<br />

pigmented photosynthetic complexes the main site <str<strong>on</strong>g>of</str<strong>on</strong>g> photodamage is Photosystem II. The main factors,<br />

which are resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> the light sensitivity <str<strong>on</strong>g>of</str<strong>on</strong>g> Photosystem II are excited pigment molecules, oxygen,<br />

manganese, as well as electr<strong>on</strong> d<strong>on</strong>ors with high oxidizing potential. Photosystem II can be efficiently<br />

protected from photodamage by the combinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> harmless dissipati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> absorbed light energy, n<strong>on</strong>radiative<br />

charge recombinati<strong>on</strong> and repair <str<strong>on</strong>g>of</str<strong>on</strong>g> damaged reacti<strong>on</strong> center complexes making possible the safe<br />

utilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> light, the highly energetic substrate <str<strong>on</strong>g>of</str<strong>on</strong>g> photosynthesis. The lecture will cover the principles and<br />

basic mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> photodamage <str<strong>on</strong>g>of</str<strong>on</strong>g> photosynthesis, and its repair.<br />

188


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

4A_05_S<br />

(poster secti<strong>on</strong> B2, poster board #259, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

MAMMALIAN BAX INITIATES PLANT CELL DEATH THROUGH ROS<br />

PRODUCTION AND ORGANELLE DESTRUCTION<br />

Maki Kawai-Yamada, Keiko Yoshinaga, Hir<str<strong>on</strong>g>of</str<strong>on</strong>g>umi Uchimiya<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular and Cellular Biosciences, The University <str<strong>on</strong>g>of</str<strong>on</strong>g> Tokyo, Japan<br />

e-mail: mkawai@iam.u-tokyo.ac.jp<br />

Relatively few endogenous plant genes that share sequence homology with the mammalian apoptotic genes<br />

have been identified to date. N<strong>on</strong>etheless, similarities in PCD exist in plants and animals. Mammalian<br />

proapoptotic gene Bax is known to cause cell death when expressed in yeast and plants. We examined<br />

transgenic plants expressing both Bax and organelle-targeted green fluorescent protein (GFP) to analyze the<br />

cellular event that occur during Bax-induced plant cell death. The results indicated that Bax induced temporal<br />

and special cell death events at the organelle level. Such events included i<strong>on</strong> leakage, DNA fragmentati<strong>on</strong> and<br />

cell shrinkage. The mitoch<strong>on</strong>dria changed morphologically from being bacilli-shaped to being round. The<br />

chloroplasts lost membrane functi<strong>on</strong> and their c<strong>on</strong>tents leaked out, followed by the disrupti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the vacuole.<br />

Light was not essential <str<strong>on</strong>g>for</str<strong>on</strong>g> Bax-induced i<strong>on</strong> leakage or organelle disrupti<strong>on</strong>, but <str<strong>on</strong>g>for</str<strong>on</strong>g> chlorosis. Furthermore,<br />

ROS producti<strong>on</strong> was involved in triggering cell death. To compare Bax-induced cell death and other ROSmediated<br />

plant cell death, Arabidopsis leaves expressing mitoch<strong>on</strong>drial-targeted GFP were treated with ROSinducing<br />

chemicals, such as hydrogen peroxide, paraquat and menadi<strong>on</strong>e. After 24h treatment, mitoch<strong>on</strong>dria<br />

showed morphological changes from a bacillus-like to a round shape. The size <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>dria decreased by<br />

half compared with c<strong>on</strong>trols. Such cellular events may cause energy depleti<strong>on</strong>, and resulted in plant cell death.<br />

4A_06_S<br />

(poster secti<strong>on</strong> B2, poster board #260, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

REACTIVE CARBONYL DETOXIFICATION AND STRESS RESISTANCE IN PLANTS<br />

Gábor V. Horváth, Zoltán Turóczy, Petra Kis, Katalin Török, Éva Hideg, László Sass, Dénes Dudits<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Biology, Biological <str<strong>on</strong>g>Research</str<strong>on</strong>g> Center HAS, Szeged, Hungary<br />

e-mail: hvg@brc.hu<br />

Productivity <str<strong>on</strong>g>of</str<strong>on</strong>g> plants is greatly affected by envir<strong>on</strong>mental stresses, there<str<strong>on</strong>g>for</str<strong>on</strong>g>e there is a c<strong>on</strong>tinuous need <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

the genetic improvement <str<strong>on</strong>g>of</str<strong>on</strong>g> stress tolerance in the agriculture. During oxidative stress rapid accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

reactive oxygen species (ROS) and reactive carb<strong>on</strong>yl species (RCS, carb<strong>on</strong>yl stress) significantly c<strong>on</strong>tributes to<br />

the damage <str<strong>on</strong>g>of</str<strong>on</strong>g> crop plants. Improvement <str<strong>on</strong>g>of</str<strong>on</strong>g> intracellular scavenging capacity <str<strong>on</strong>g>of</str<strong>on</strong>g> such compounds provenly<br />

lead to increased stress tolerance. Diverse enzymes can detoxify the reactive carb<strong>on</strong>yl species (eg. 4-hydoxyn<strong>on</strong>enal<br />

or methylglyoxal). Their importance is clearly dem<strong>on</strong>strated by the fact that overproducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

alkenal reductases, aldehyde dehydrogenases or the members <str<strong>on</strong>g>of</str<strong>on</strong>g> the glyoxalase enzyme family in transgenic<br />

plants led to improved tolarance to wide range <str<strong>on</strong>g>of</str<strong>on</strong>g> stress c<strong>on</strong>diti<strong>on</strong>s. We have isolated the MsALR aldose<br />

reductase homolog gene from Medicago sativa and showed the accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the transcript at higher levels in<br />

resp<strong>on</strong>se to different stress treatments. Transgenic tobacco plants ectopically expressing the MsALR cDNA<br />

were more tolerant to dehydrati<strong>on</strong> stress and recovered better from damages caused by water deficit than the<br />

untrans<str<strong>on</strong>g>for</str<strong>on</strong>g>med wild type plants and were more tolerant to heavy metal, salt, dehydrati<strong>on</strong> and UV-B stress.<br />

Results <str<strong>on</strong>g>of</str<strong>on</strong>g> enzyme activity measurements and in vivo assays <str<strong>on</strong>g>for</str<strong>on</strong>g> protecti<strong>on</strong> against methylglyoxal toxicity<br />

strengthened our hypothesis that <strong>on</strong>e important functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> aldo/keto reductase proteins is also the<br />

eliminati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> reactive aldehydes and the reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the c<strong>on</strong>sequences <str<strong>on</strong>g>of</str<strong>on</strong>g> carb<strong>on</strong>yl stress in plants.<br />

189


23-26 August 2007,<br />

Budapest, Hungary<br />

190<br />

4A_01_P<br />

(poster secti<strong>on</strong> B2, poster board #261, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CYTOCHROMES B561, NEW PLAYERS IN PLANT IRON METABOLISM AND<br />

OXIDATIVE STRESS METABOLISM<br />

Han Asard, Alajos Bérczi, Kim Van Beek, Daniel Griesen<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Antwerp, Antwerp, Belgium; Biological <str<strong>on</strong>g>Research</str<strong>on</strong>g> Center, Szeged, Hungary<br />

e-mail: han.asard@ua.ac.be<br />

Cytochromes b561 (Cyt b561) are a newly identified class <str<strong>on</strong>g>of</str<strong>on</strong>g> trans-membrane proteins, using ascorbate as an<br />

electr<strong>on</strong> d<strong>on</strong>or. These proteins have been dem<strong>on</strong>strated to transfer electr<strong>on</strong>s across the membrane in which<br />

they are embedded, but their physiological role remains unclear. We have identified four Cyt b561 is<str<strong>on</strong>g>of</str<strong>on</strong>g>orms<br />

(AtCytb1-4) in Arabidopsis and are characterizing their mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> acti<strong>on</strong> and using biochemical,<br />

molecular biological and physiological approaches. Several lines <str<strong>on</strong>g>of</str<strong>on</strong>g> evidence suggest that the plant Cyts b561<br />

are involved in ir<strong>on</strong> metabolism and in oxidative stress resp<strong>on</strong>ses: 1) A knock-out in <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the four Cyts<br />

b561 identified from Arabidopsis dem<strong>on</strong>strates a particular phenotype under ir<strong>on</strong> deficiency. 2) The<br />

recombinant AtCytb1 protein can be oxidized by ir<strong>on</strong> chelates. 3) The AtCytb1 gene appears upregulated<br />

under ir<strong>on</strong>-deficiency c<strong>on</strong>diti<strong>on</strong>s. And 4) In vivo experiments with AtCytb1 expressed in yeast dem<strong>on</strong>strate<br />

its ferric-reductase capability. Recent experiments however also dem<strong>on</strong>strate that the AtCytb1 knock-out<br />

plants show a particular phenotype under oxidative stress c<strong>on</strong>diti<strong>on</strong>s. Str<strong>on</strong>gly reduced root development is<br />

observed in the mutant plants when treated with paraquat. These results suggest that Cyts b561 may provide<br />

a link between plant ir<strong>on</strong> metabolism and oxidative stress phenomena, using ascorbate as the electr<strong>on</strong> d<strong>on</strong>or.<br />

4A_02_P<br />

(poster secti<strong>on</strong> B2, poster board #262, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SELENIUM-INDUCED OXIDATIVE STRESS IN COFFEE CELL SUSPENSION<br />

CULTURES<br />

Ricardo A. Azevedo 1 , Priscila L. Gratão 1 , Paulo Mazzafera 2 , Peter J. Lea 3 , Rui A. Gomes-Junior 4<br />

1ESALQ, Universidade de São Paulo, Brazil; 2 Universidade Estadual de Campinas, Brazil<br />

3University <str<strong>on</strong>g>of</str<strong>on</strong>g> Lancaster, UK, 4 Instituto Agr<strong>on</strong>ômico do Paraná, Brazil<br />

e-mail: raazeved@esalq.usp.br<br />

Selenium (Se) is an essential element <str<strong>on</strong>g>for</str<strong>on</strong>g> humans and animals that is required <str<strong>on</strong>g>for</str<strong>on</strong>g> key antioxidant reacti<strong>on</strong>s,<br />

but can be toxic at high c<strong>on</strong>centrati<strong>on</strong>s. We have investigated the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> Se in the <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> selenite <strong>on</strong><br />

c<str<strong>on</strong>g>of</str<strong>on</strong>g>fee cell suspensi<strong>on</strong> cultures over a 12 day period. The antioxidant defence systems were induced in c<str<strong>on</strong>g>of</str<strong>on</strong>g>fee<br />

cells grown in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> 0.05 and 0.5 mM sodium selenite (Na 2 SeO 3 ). Lipid peroxidati<strong>on</strong> and alterati<strong>on</strong>s<br />

in antioxidant enzymes were the main resp<strong>on</strong>ses observed, including a severe reducti<strong>on</strong> in ascorbate<br />

peroxidase activity, even at 0.05 mM sodium selenite. Ten superoxide dismutase (SOD) isoenzymes were<br />

detected and the two major Mn-SOD isoenzymes (bands V and VI) resp<strong>on</strong>ded more to 0.05 mM selenite.<br />

SOD band V exhibited a general decrease in activity after 12 h <str<strong>on</strong>g>of</str<strong>on</strong>g> treatment with 0.05 mM selenite, whereas<br />

band VI exhibited the opposite behavior and increased in activity. An extra isoenzyme <str<strong>on</strong>g>of</str<strong>on</strong>g> glutathi<strong>on</strong>e<br />

reductase (GR) was induced in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> selenite, which c<strong>on</strong>firmed our previous reports obtained with<br />

Cd (Gomes-Junior et al.: Chemosphere 65, 1330-1337, 2006) and Ni (Gomes-Junior et al.: Plant Physiology<br />

and Biochemistry 44, 420-429, 2006) indicating that this GR isoenzyme may have the potential to be a marker<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> oxidative stress in c<str<strong>on</strong>g>of</str<strong>on</strong>g>fee.


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

4A_03_P<br />

(poster secti<strong>on</strong> B2, poster board #263, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

RESPONSE OF BRAZILIAN CULTIVARS OF SOYBEAN (GLYCINE MAX) EXPOSE<br />

TO OZONE UNDER CONTROLLED CONDITIONS<br />

Patrícia Bulbovas a* , Silvia Ribeiro de Souza b , Marisa Domingos b , Ricardo Antunes Azevedo a<br />

a Departamento de Genética, Escola Superior de Agricultura Luiz de Queiroz, Universidade de São Paulo,<br />

Piracicaba CEP 13400-970, SP, Brasil<br />

b Instituto de Botânica, São Paulo, CEP 01061-970, SP, Brasil<br />

* e-mail: patibul@usp.br<br />

Oz<strong>on</strong>e (O 3 ) can enter the leaves and react with water <str<strong>on</strong>g>for</str<strong>on</strong>g>ming reactive oxygen species (ROS). ROS is able to<br />

cause physiological and biochemical damage to plants. This study was carried out to investigate the O 3 effects<br />

<strong>on</strong> two distinct Brazilian cultivars <str<strong>on</strong>g>of</str<strong>on</strong>g> soybean (Glycine max): ‘Tracajá’ and ‘Sambaíba’. Plants were grown in<br />

two chambers, <strong>on</strong>e with filtered air (FA) and the other with filtered air plus 40 ppb <str<strong>on</strong>g>of</str<strong>on</strong>g> oz<strong>on</strong>e (FA+O 3 ) <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

five days. Comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g> the antioxidative defense system such as ascorbic acid (AA), glutathi<strong>on</strong>e reductase<br />

(GR), ascorbate peroxidase (APX), guayacol peroxidase (GPX), catalase (CAT) and superoxide dismutase<br />

(SOD) were analyzed and the relative growth rate (RGR) and biomass producti<strong>on</strong> were also determined. APX<br />

and GPX in ‘Sambaíba’ and AA and GPX in ‘Tracajá’ decreased under FA+O 3 . AA and CAT in ‘Sambaíba’<br />

and CAT in ‘Tracajá’ exhibited similar levels under both treatments. GR in ‘Sambaíba’ and GR and APX<br />

increased in ‘Tracajá’ exposed to FA+O 3 . CAT and SOD activity staining by n<strong>on</strong>-denaturing PAGE revealed<br />

the same is<str<strong>on</strong>g>of</str<strong>on</strong>g>orms numbers <str<strong>on</strong>g>for</str<strong>on</strong>g> both cultivars, but different is<str<strong>on</strong>g>of</str<strong>on</strong>g>orms <str<strong>on</strong>g>of</str<strong>on</strong>g> GR. ‘Tracajá’exhibited variati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

some CAT and SOD is<str<strong>on</strong>g>of</str<strong>on</strong>g>orms in FA+O 3 . The root/shoot ratio in ‘Tracajá’ and root/shoot ratio, leaf RGR<br />

and root biomass producti<strong>on</strong> in ‘Sambaíba’ were lower under FA+O 3 . O 3 treatment induced distinct<br />

antioxidative resp<strong>on</strong>ses by the distinct antioxidant systems in resp<strong>on</strong>se to O 3 (AF+O 3 ). The antioxidant<br />

defense system variati<strong>on</strong>s compensated the lower O 3 interference <strong>on</strong> growth parameters and biomass<br />

producti<strong>on</strong>.<br />

4A_04_P<br />

(poster secti<strong>on</strong> B2, poster board #264, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

GLUTATHIONE HALF-CELL REDUCTION POTENTIAL: A UNIVERSAL STRESS<br />

MARKER FROM PLANTS TO HUMANS<br />

Ilse Kranner, Sim<strong>on</strong>a Birtić, Hugh W. Pritchard<br />

Seed C<strong>on</strong>servati<strong>on</strong> Department, Royal Botanic Gardens, Kew, Wakehurst Place, West Sussex RH 17 6TN, UK<br />

e-mail: i.kranner@kew.org<br />

Ageing phenomena and programmed cell death (PCD) are typically studied in human or animal cells rather<br />

than in plants. However, the seeds <str<strong>on</strong>g>of</str<strong>on</strong>g> higher plants represent excellent models <str<strong>on</strong>g>for</str<strong>on</strong>g> the study <str<strong>on</strong>g>of</str<strong>on</strong>g> ageing because<br />

viability loss can be easily induced experimentally. ‘Orthodox’ seeds are desiccati<strong>on</strong> tolerant and ‘recalcitrant’<br />

seeds are desiccati<strong>on</strong> sensitive. Lethal damage, as judged by germinati<strong>on</strong> tests, can be induced by artificial<br />

ageing in orthodox, and by drying in recalcitrant seeds. We have investigated mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> viability loss to<br />

find a reliable marker that quantifies ‘stress’. Oxidative damage has previously been correlated with<br />

degenerative processes and death, but how exactly this c<strong>on</strong>tributes to viability loss is unknown. We show in<br />

four species subjected to ageing or desiccati<strong>on</strong>, that seed viability decreased by 50% when the half-cell<br />

reducti<strong>on</strong> potential <str<strong>on</strong>g>of</str<strong>on</strong>g> glutathi<strong>on</strong>e (E GSSG/2GSH ), a major cellular antioxidant and redox buffer, increased to a<br />

191


23-26 August 2007,<br />

Budapest, Hungary<br />

z<strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> -180 to -160 mV, in agreement with a model that has been suggested <str<strong>on</strong>g>for</str<strong>on</strong>g> human cells (1). In a metaanalysis<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> data representative <str<strong>on</strong>g>of</str<strong>on</strong>g> 13 plant and fungal orders we show that stress generally becomes lethal<br />

when E GSSG/2GSH exceeds -160 mV. We put <str<strong>on</strong>g>for</str<strong>on</strong>g>ward that this change in E GSSG/2GSH is <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the ‘death triggers’<br />

that initiate PCD, finally causing inter-nucleosomal DNA fragmentati<strong>on</strong> in the final, or executi<strong>on</strong> phase, <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

PCD. E GSSG/2GSH is there<str<strong>on</strong>g>for</str<strong>on</strong>g>e a universal marker <str<strong>on</strong>g>of</str<strong>on</strong>g> plant cell viability and allows us to predict whether a seed<br />

will live, germinate and produce a new plant, or if it will die.<br />

192<br />

4A_05_P<br />

(poster secti<strong>on</strong> B2, poster board #265, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EXPRESSION ANALYSIS OF PHENYLALANINE AMMONIA LYASE IN RELATION<br />

TO OZONE STRESS IN YELLOW POPLAR PLANTS<br />

(LIRIODENDRON TULIPIFERA)<br />

A. Francini, E. Pellegrini, G. Lorenzini, C. Nali*<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Tree Science, Entomology and Plant Pathology “G. Scaramuzzi”, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Pisa, Via del<br />

Borghetto 80, 56124 Pisa, Italy.<br />

*e-mail: cristina.nali@agr.unipi.it<br />

Tropospheric oz<strong>on</strong>e is a widespread phytotoxic pollutant that causes significant damage to plants, but<br />

relatively little is known about oz<strong>on</strong>e stress in urban trees. The phenylpropanoid pathway is <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the most<br />

important pathways <str<strong>on</strong>g>for</str<strong>on</strong>g> the synthesis <str<strong>on</strong>g>of</str<strong>on</strong>g> natural products in plants such as phenols and lignin. In this study,<br />

the hypothesis that phenylalanine amm<strong>on</strong>ia lyase (PAL) may be affected by oz<strong>on</strong>e treatment (120 ppb, 5 h d -1<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> 45 c<strong>on</strong>secutive days) in Liriodendr<strong>on</strong> tulipifera (yellow poplar) plants was examined. At the end <str<strong>on</strong>g>of</str<strong>on</strong>g> exposure,<br />

the material showed characteristic symptoms <str<strong>on</strong>g>of</str<strong>on</strong>g> injury in the <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> severe minute roundish dark-blackish<br />

necrosis localised in the interveinal areas <str<strong>on</strong>g>of</str<strong>on</strong>g> the adaxial surface <str<strong>on</strong>g>of</str<strong>on</strong>g> leaves. Treated plants increase PAL activity<br />

after oz<strong>on</strong>e fumigati<strong>on</strong> (+43%) and total phenols (+41%) compared to c<strong>on</strong>trols. Total leaf RNA protocol<br />

was optimised <str<strong>on</strong>g>for</str<strong>on</strong>g> yellow poplar. Degenerate primers were designed <strong>on</strong> highly c<strong>on</strong>served sequences available<br />

<strong>on</strong> GenBank, that encode <str<strong>on</strong>g>for</str<strong>on</strong>g> this enzyme. The primers yielded 679 bp cDNA fragments. Sequences obtained<br />

were translated in aminoacids and compared in the GenBank. Blast results showed high homology at<br />

aminoacids level with PAL, c<strong>on</strong>firming that the cDNA fragments isolated effectively encode <str<strong>on</strong>g>for</str<strong>on</strong>g> this gene. In<br />

particular, the sequences showed high homologies with Ulmus pumila (93%) and with Pyrus comunis (96%). The<br />

northern analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> PAL gene expressi<strong>on</strong> lead to support the view <str<strong>on</strong>g>of</str<strong>on</strong>g> oz<strong>on</strong>e as an abiotic elicitor <str<strong>on</strong>g>of</str<strong>on</strong>g> defence<br />

resp<strong>on</strong>ses in yellow poplar.<br />

4A_06_P<br />

(poster secti<strong>on</strong> B2, poster board #266, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

INFLUENCE OF ANTIOXIDANTS ON SOME STAGES OF OXIDATIVE STRESS IN<br />

PLANT CELLS OF WHEAT AND BARLEY SEEDLINGS<br />

N. Shkute, M. Savicka, N. Zatinajchenko, N. Rjabovola<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> research and management <str<strong>on</strong>g>of</str<strong>on</strong>g> biological resources, Daugavpils University, Vienības iela 13, Daugavpils,<br />

LV-5401, Latvia<br />

The producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> superoxide is crucial <str<strong>on</strong>g>for</str<strong>on</strong>g> normal morphogenesis at early stages and is causal <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis<br />

and senescence at late stages <str<strong>on</strong>g>of</str<strong>on</strong>g> development <str<strong>on</strong>g>of</str<strong>on</strong>g> etiolated cereals seedlings (wheat, barley and others). It was


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

shown, that cyclic change <str<strong>on</strong>g>of</str<strong>on</strong>g> rate <str<strong>on</strong>g>of</str<strong>on</strong>g> superoxide producti<strong>on</strong> is correlate with changes <str<strong>on</strong>g>of</str<strong>on</strong>g> nuclear DNA<br />

synthesis and apoptotic fragmentati<strong>on</strong> in cells <str<strong>on</strong>g>of</str<strong>on</strong>g> some development and senescence separated organs (leaf,<br />

coleoptile) <str<strong>on</strong>g>of</str<strong>on</strong>g> this plants. It was found, that antioxidant defence systems: superoxide dismutase and<br />

peroxidase are activated in apoptotic cells <str<strong>on</strong>g>of</str<strong>on</strong>g> senescence organs. The influence <str<strong>on</strong>g>of</str<strong>on</strong>g> endogenic antioxidant<br />

(ascorbic acid) and synthetic antioxidant (α - I<strong>on</strong>ol) <strong>on</strong> superoxide producti<strong>on</strong>, <strong>on</strong> activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> antioxidant<br />

defence systems, <strong>on</strong> nuclear DNA synthesis and nuclear DNA apoptotic fragmentati<strong>on</strong> and <strong>on</strong> structure <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

cellular organelles was investigated. It was found, that cyclosporin A, inhibitor <str<strong>on</strong>g>of</str<strong>on</strong>g> the mammalian permeability<br />

transiti<strong>on</strong> pore, inhibits <str<strong>on</strong>g>of</str<strong>on</strong>g> peroxide <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> and nuclear DNA apoptotic fragmentati<strong>on</strong>. Taking this date<br />

mitoch<strong>on</strong>dria are likely to be involved in plant programmed cell death, induced by oxidative stress, but the<br />

molecular mechanisms may be different from those found in animals.<br />

4A_07_P<br />

(poster secti<strong>on</strong> B2, poster board #267, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SEASONAL VARIATION IN THE ACTIVITY OF ANTIOXIDANT ENZYMES<br />

PEROXIDASE, SUPEROXIDE DISMUTASE AND CATALASE IN AN OPEN AND A<br />

SHADED POPULATION OF IRIS PUMILAS<br />

Branka Tucić*, Ana Vuleta, Darka Šešlija<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Evoluri<strong>on</strong>ary Biology, Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Biological <str<strong>on</strong>g>Research</str<strong>on</strong>g> „Siniša Stanković“, Belgrade, Serbia,<br />

*e-mail: btucic@ibiss.bg.ac.yu<br />

Seas<strong>on</strong>al variati<strong>on</strong> in the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> peroxidase (POD, EC 1.11.1.7), superoxide dismutase (SOD, EC<br />

1.15.1.1), and catalase (CAT, EC 1.11.1.6) was determined in the leaves <str<strong>on</strong>g>of</str<strong>on</strong>g> two Iris pumila populati<strong>on</strong>s, <strong>on</strong>e<br />

naturally growing at an open dune site and the other in the understory <str<strong>on</strong>g>of</str<strong>on</strong>g> a Pinus stand. A repeated-measures<br />

pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ile analysis revealed that the average level, as well as the mean change in the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> these enzymes<br />

varied significantly between c<strong>on</strong>trasting light habitats. POD activity was significantly greater at exposed dune<br />

site than under vegetati<strong>on</strong> canopy, and reached its maximum in the summer but <strong>on</strong>ly in plants experiencing<br />

full sunlight. C<strong>on</strong>versely, in woodland understory, POD activity gradually increased from spring to autumn.<br />

The mean activity <str<strong>on</strong>g>of</str<strong>on</strong>g> SOD and CAT was c<strong>on</strong>sistently greater in plants inhabiting vegetati<strong>on</strong> shade compared<br />

to those exposed to full irradiance. Throughout the growing seas<strong>on</strong>, the variati<strong>on</strong> pattern <str<strong>on</strong>g>of</str<strong>on</strong>g> CAT activity was<br />

the same in both light habitats, while the resp<strong>on</strong>se curve <str<strong>on</strong>g>of</str<strong>on</strong>g> SOD activity changed the shape with<br />

envir<strong>on</strong>mental c<strong>on</strong>diti<strong>on</strong>s, particularly in the period from summer to autumn. At open dune site in the spring,<br />

the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> POD appeared to be inversely related to SOD, suggesting that plants with lower SOD<br />

producti<strong>on</strong> up-regulate their POD level to compensate <str<strong>on</strong>g>for</str<strong>on</strong>g> SOD reducti<strong>on</strong> under given envir<strong>on</strong>mental setup.<br />

The observed results imply that abiotic stress can disrupt the redox homeostasis in I. pumila plants, changing<br />

the balance between POD, SOD and CAT activities according to the extent <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidative stress in cells.<br />

193


23-26 August 2007,<br />

Budapest, Hungary<br />

194<br />

4A_08_P<br />

(poster secti<strong>on</strong> B2, poster board #268, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

BATTLE AGAINST REACTIVE COMPOUNDS-AKRS AND STRESS<br />

Zoltán Turóczy, Mátyás Cserháti, Petra Kis, Dénes Dudits, Gábor V. Horváth<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Biology, Biological <str<strong>on</strong>g>Research</str<strong>on</strong>g> Center HAS, Temesvári krt. 62, H-6726 Szeged, Hungary<br />

e-mail: turoczy@brc.hu<br />

Plants exposed to abiotic stress are subjected to oxidative damage. Reactive compounds produced under such<br />

c<strong>on</strong>diti<strong>on</strong>s significantly increase the cytotoxic effect <str<strong>on</strong>g>of</str<strong>on</strong>g> envir<strong>on</strong>mental stress factors. Aldo/keto reductases<br />

(AKRs) have been c<strong>on</strong>sidered as effective enzymes <str<strong>on</strong>g>for</str<strong>on</strong>g> the detoxificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> lipid peroxidati<strong>on</strong>- and<br />

glycolysis-derived reactive aldehydes. The promoters <str<strong>on</strong>g>of</str<strong>on</strong>g> targeted rice AKR genes were screened <str<strong>on</strong>g>for</str<strong>on</strong>g> stressrelated<br />

transcripti<strong>on</strong> factor binding sites (TFBS’s) by a new, enumerati<strong>on</strong> based method algorithm. QRT-PCR<br />

was used to determine the transcript pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ile <str<strong>on</strong>g>for</str<strong>on</strong>g> the selected genes in abscisic acid (ABA), H 2 O 2 , NaCl and<br />

mannitol treated rice cell suspensi<strong>on</strong>s. Am<strong>on</strong>g the genes, OsALR1 showed the highest inducibility and<br />

transcript level during the treatments, suggesting its important role in stress tolerance. Furthermore the Q-<br />

PCR experiments substantiated, that the number <str<strong>on</strong>g>of</str<strong>on</strong>g> motifs found in the promoters and the stress resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the genes were in close correlati<strong>on</strong>. In additi<strong>on</strong>, the photosynthetic parameters <str<strong>on</strong>g>of</str<strong>on</strong>g> the tobacco lines,<br />

overexpressing rice OsALR1 and OsALR4 were better then those <str<strong>on</strong>g>of</str<strong>on</strong>g> the wild type plants after paraquat and<br />

methylglyoxal (MG) treatments. The in vitro enzyme kinetic c<strong>on</strong>stants <str<strong>on</strong>g>of</str<strong>on</strong>g> the GST-OsALR1 fusi<strong>on</strong> protein<br />

revealed a high reducing activity <str<strong>on</strong>g>for</str<strong>on</strong>g> toxic aldehydes like MG in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> NADPH. Results <str<strong>on</strong>g>of</str<strong>on</strong>g> in vivo<br />

assays in E. coli <str<strong>on</strong>g>for</str<strong>on</strong>g> protecti<strong>on</strong> against MG toxicity have also made it clear, that <strong>on</strong>e important functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

these AKR proteins is the eliminati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> reactive aldehydes from plant cells expletively besides the glyoxalase<br />

system. This work was supported by NKFP Grant No. 4-064-2004, Gábor V. Horváth is grateful <str<strong>on</strong>g>for</str<strong>on</strong>g> the support <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

“János Bólyai” <str<strong>on</strong>g>Research</str<strong>on</strong>g> Fellowship.<br />

4A_09_P<br />

(poster secti<strong>on</strong> B2, poster board #269, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

INFLUENCE OF CADMIUM ON THE ANTIOXIDANT SYSTEM<br />

OF TOBACCO PLANTS<br />

A. I. Cardoso 1 , M. P. Mourato *1 , L. L. Martins 1 , A. P. Pinto 2 , A. M. Mota 3 ,<br />

M. L. G<strong>on</strong>çalves 3 , A. Varennes 1<br />

(1) Instituto Superior de Agr<strong>on</strong>omia, Technical University <str<strong>on</strong>g>of</str<strong>on</strong>g> Lisb<strong>on</strong>, Tapada da Ajuda, 1349-017 Lisboa, Portugal<br />

(2) Departamento de Química, Universidade de Évora. Colégio Luís António Verney 7000 Évora<br />

(3) Instituto Superior Técnico, Technical University <str<strong>on</strong>g>of</str<strong>on</strong>g> Lisb<strong>on</strong>, Av. Rovisco Pais, 1049-001 Lisboa, Portugal<br />

* e-mail: mmourato@isa.utl.pt<br />

In this work we studied the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> cadmium (0, 0.010, 0.025, 0.050, 0.100 mM) <strong>on</strong> enzymatic and n<strong>on</strong>enzymatic<br />

parameters <str<strong>on</strong>g>of</str<strong>on</strong>g> tobacco plants grown in hydrop<strong>on</strong>ic culture, in order to get a further insight in the<br />

detoxificati<strong>on</strong> processes and cadmium absorpti<strong>on</strong> mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> that plant. Tobacco has been chosen since<br />

it has been referred as a cadmium accumulator plant. Leaf dry weight percentage increased with cadmium<br />

c<strong>on</strong>centrati<strong>on</strong> indicating lower water absorpti<strong>on</strong> possibly due to affected root development. Chlorophyll a<br />

and b levels decreased markedly (down to 16 % <str<strong>on</strong>g>of</str<strong>on</strong>g> the c<strong>on</strong>trol value <str<strong>on</strong>g>for</str<strong>on</strong>g> the highest cadmium c<strong>on</strong>centrati<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> 0.100 mM) showing that the photosynthetic system was affected by cadmium. Hydrogen peroxide<br />

measurements showed no significant variati<strong>on</strong>s, which could indicate that the antioxidant system <str<strong>on</strong>g>of</str<strong>on</strong>g> this plant


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

is capable <str<strong>on</strong>g>of</str<strong>on</strong>g> quenching the possible excess <str<strong>on</strong>g>of</str<strong>on</strong>g> hydrogen peroxide induced by cadmium. In fact, both guaiacol<br />

peroxidase and ascorbate peroxidase activities were enhanced with increasing cadmium c<strong>on</strong>centrati<strong>on</strong> in<br />

nutrient soluti<strong>on</strong>. Another enzyme usually involved in the antioxidant system <str<strong>on</strong>g>of</str<strong>on</strong>g> plants, superoxide dismutase,<br />

showed no difference in its activity levels compared to the c<strong>on</strong>trol. A slight increase <strong>on</strong> mal<strong>on</strong>dialdehyde<br />

levels, both in leaves and roots, indicated that lipid peroxidati<strong>on</strong> should occur as a result <str<strong>on</strong>g>of</str<strong>on</strong>g> increased ROS<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> induced by cadmium toxicity.<br />

4A_10_P<br />

(poster secti<strong>on</strong> B2, poster board #270, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

INVESTIGATION OF ARSENATE PHYTOTOXICITY IN CUCUMBER PLANTS<br />

V. Czech 1 , E. Cseh 1 , F. Fodor 1 , P. Czövek 1 , J. Fodor 2<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Physiology and Molecular Plant Biology, L. Eötvös University, P.O. Box 120, H-1518<br />

Budapest, Hungary<br />

2 Plant Protecti<strong>on</strong> Institute, Hungarian Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences, H-1525 Budapest, P.O. Box 102<br />

The effect <str<strong>on</strong>g>of</str<strong>on</strong>g> arsenate <strong>on</strong> the stress-sensitive vegetable plant, cucumber (Cucumis sativus cv. Joker F1) grown<br />

in hydrop<strong>on</strong>ic culture (modified Hoagland soluti<strong>on</strong>, 10µM KH 2 PO 4 , 10µM FeCl 3 and 10µM KH 2 AsO 4 ) was<br />

investigated.<br />

The presence <str<strong>on</strong>g>of</str<strong>on</strong>g> arsenic in soils and drinking water and its c<strong>on</strong>sequent appearance in the food chain has a<br />

potential health damaging effect. The arsenic load <str<strong>on</strong>g>of</str<strong>on</strong>g> the Hungarian populati<strong>on</strong> in the various parts <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

country is substantially different, it exceeds 50 µg/l in the Southern part <str<strong>on</strong>g>of</str<strong>on</strong>g> the Great Plain. The arsenic<br />

allowance <str<strong>on</strong>g>of</str<strong>on</strong>g> drinking water in the European Uni<strong>on</strong> is 10 µg/l.<br />

As(V) treatment influences the water household <str<strong>on</strong>g>of</str<strong>on</strong>g> cucumber depending <strong>on</strong> its stage <str<strong>on</strong>g>of</str<strong>on</strong>g> development. When<br />

the first leaf is developing at the time <str<strong>on</strong>g>of</str<strong>on</strong>g> treatment turgor loss occurs first in the roots then in the shoot.<br />

As(V) inhibits transpirati<strong>on</strong>. This effect is significantly reduced by 100µM phosphate because As(V) is taken<br />

up through the phosphate transport system. Arsenate increases the water saturati<strong>on</strong> deficit <str<strong>on</strong>g>of</str<strong>on</strong>g> the leaf and<br />

reduces the exudati<strong>on</strong> rate <str<strong>on</strong>g>of</str<strong>on</strong>g> the root to 25% <str<strong>on</strong>g>of</str<strong>on</strong>g> the c<strong>on</strong>trol. As(V) also significantly reduces the dry matter<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> plant parts and causes an i<strong>on</strong> leakage in the roots leading up to 50% higher c<strong>on</strong>ductivity in the washing<br />

soluti<strong>on</strong>. These findings imply that the turgor loss is caused by the loss <str<strong>on</strong>g>of</str<strong>on</strong>g> membrane semipermeability.<br />

As(V) is reduced to As (III) in the roots that may lead to the <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> reactive oxygen species. Lipid<br />

peroxidati<strong>on</strong> occurs in the hypocotyls in which the mal<strong>on</strong>dialdehyde c<strong>on</strong>centrati<strong>on</strong> increases due to the<br />

oxidative stress. Cucumbers grown <strong>on</strong> Fe-ascorbate (10µM FeCl 3 + 30µM ascorbate) had healthy hypocotyls<br />

in spite <str<strong>on</strong>g>of</str<strong>on</strong>g> the As(V) treatment so that Fe-ascorbate prevented lipid peroxidati<strong>on</strong>. Further measurements <strong>on</strong><br />

the stress indicator H 2 O 2 level and the redox processes <str<strong>on</strong>g>of</str<strong>on</strong>g> ascorbate in the hypocotyls are discussed.<br />

195


23-26 August 2007,<br />

Budapest, Hungary<br />

196<br />

4B. BIOTIC STRESS AND DISEASE RESISTANCE IN PLANTS<br />

(TERRY GRAHAM)<br />

4B_01_S<br />

USE OF GENE SILENCING AND METABOLOMICS TO CHARACTERIZE<br />

INTERACTIVE STRESS AND DEFENSE PATHWAYS IN SOYBEAN<br />

T. L. Graham, M. Y. Graham, S. Subramanian, O. Yu<br />

Ohio State University, Columbus, OH, USA 43210<br />

e-mail: graham.1@osu.edu<br />

Induced resistance to pathogens can be specific to a given race <str<strong>on</strong>g>of</str<strong>on</strong>g> a pathogen or effective against a range <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

pathogens. The <str<strong>on</strong>g>for</str<strong>on</strong>g>mer type <str<strong>on</strong>g>of</str<strong>on</strong>g> resistance involves a <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> programmed cell death called the hypersensitive<br />

resp<strong>on</strong>se (HR). The latter type <str<strong>on</strong>g>of</str<strong>on</strong>g> resistance, called general or basal resistance, is <str<strong>on</strong>g>of</str<strong>on</strong>g>ten induced in plants by<br />

elicitors from the pathogen called pathogen associated molecular patterns (PAMPs). This <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> resistance<br />

has some mechanistic similarities to innate immunity in animals. In the work described here we have used<br />

RNAi gene silencing and metabolic pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iling to study the molecular bases <str<strong>on</strong>g>of</str<strong>on</strong>g> soybean resistance to the<br />

pathogen, Phytophthora sojae. Resistance resp<strong>on</strong>ses were examined in the c<strong>on</strong>text <str<strong>on</strong>g>of</str<strong>on</strong>g> <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the most highly<br />

characterized PAMPs, the cell wall glucan elicitor (WGE) from P. sojae. Silencing either <str<strong>on</strong>g>of</str<strong>on</strong>g> two biosynthetic<br />

enzymes <str<strong>on</strong>g>for</str<strong>on</strong>g> soybean is<str<strong>on</strong>g>of</str<strong>on</strong>g>lav<strong>on</strong>oids (is<str<strong>on</strong>g>of</str<strong>on</strong>g>lav<strong>on</strong>e synthase or chalc<strong>on</strong>e reductase) led to a complete loss <str<strong>on</strong>g>of</str<strong>on</strong>g> racespecific<br />

resistance and HR cell death in resp<strong>on</strong>se to P. sojae. C<strong>on</strong>sistent with this, WGE, the major pathogen<br />

elicitor <str<strong>on</strong>g>of</str<strong>on</strong>g> the is<str<strong>on</strong>g>of</str<strong>on</strong>g>lav<strong>on</strong>oids in soybean, induced an HR-like cell death in root tissues. Silencing <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

endoglucanase thought to release active elicitor fragments from WGE abolished both HR cell death and<br />

WGE-induced cell death. Silencing <str<strong>on</strong>g>of</str<strong>on</strong>g> a unique metallothi<strong>on</strong>ein gene (MMT) led to greatly enhanced WGEinduced<br />

cell death, suggesting a role <str<strong>on</strong>g>of</str<strong>on</strong>g> MMT in preventing runaway cell death. Finally, silencing <str<strong>on</strong>g>of</str<strong>on</strong>g> a<br />

pathogenesis-related protein, PR-1a, led to loss <str<strong>on</strong>g>of</str<strong>on</strong>g> expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> MMT and general resistance. Thus, our<br />

studies led to the revelati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> very interesting cross-talk between the cell death and general resistance<br />

pathways in soybean.<br />

4B_02_S<br />

ENGAGEMENT AND MODIFICATION OF THE PLANT HOST STRESS<br />

MACHINERY: A VIRULENCE STRATEGY OF THE PLANT PATHOGEN<br />

PSEUDOMONAS SYRINGAE<br />

Jean T. Greenberg<br />

The University <str<strong>on</strong>g>of</str<strong>on</strong>g> Chicago, Chicago, Il 60637, USA<br />

e-mail: jgreenbe@midway.uchicago.edu<br />

Pseudom<strong>on</strong>as syringae cause disease <strong>on</strong> a broad range <str<strong>on</strong>g>of</str<strong>on</strong>g> plant hosts. To have a successful infecti<strong>on</strong>, these<br />

bacteria utilize a specialized secreti<strong>on</strong> apparatus called the type III secreti<strong>on</strong> system (T3SS). P. syringae secrete<br />

a large group (20-40+) <str<strong>on</strong>g>of</str<strong>on</strong>g> effectors through the T3SS directly into plant cells. There is tremendous diversity in<br />

the repertoire <str<strong>on</strong>g>of</str<strong>on</strong>g> secreted effectors between different P. syringae strains. However, a few effectors are comm<strong>on</strong><br />

to all P. syringae strains. We have selected <strong>on</strong>e such comm<strong>on</strong> effector and <strong>on</strong>e rare effector to functi<strong>on</strong>ally<br />

analyze in detail, including their localizati<strong>on</strong> in plants and their interacti<strong>on</strong> with potential host targets. We<br />

have found that these effectors can modify host functi<strong>on</strong>s and alter their stress resp<strong>on</strong>ses. This points to an<br />

interesting intersecti<strong>on</strong> between bacterial virulence mechanisms and host stress-coping mechanisms.


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

4B_03_S<br />

THE NDR1-ACTIN CONNECTION: LINKING GENE-FOR-GENE RESISTANCE<br />

AND THE ACTIN CYTOSKELETON<br />

Brad Day, Miaoying Tian, Marina Varbanova, Caleb Knepper<br />

Michigan State University. Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Pathology. East Lansing, MI. USA, e-mail: bday@msu.edu<br />

Disease resistance in plants involves a molecular surveillance mechanism capable <str<strong>on</strong>g>of</str<strong>on</strong>g> resp<strong>on</strong>ding to a myriad<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> plant derived elicitors. Recent research in this area has revealed a complex genetic and biochemical<br />

network required <str<strong>on</strong>g>for</str<strong>on</strong>g> the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> innate immune resp<strong>on</strong>ses in plants. In total, the coordinated<br />

interacti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> bacterial effector proteins, plant chaper<strong>on</strong>es and resistance (R)-proteins c<strong>on</strong>tribute to the<br />

molecular and biochemical events which dictate host susceptibility and resistance. <str<strong>on</strong>g>Research</str<strong>on</strong>g> in our laboratory<br />

focuses <strong>on</strong> the identificati<strong>on</strong> and characterizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> protein-protein interacti<strong>on</strong>s which not <strong>on</strong>ly functi<strong>on</strong> in<br />

the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> resistance, but also in the negative regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> effector-triggered immunity in Arabidopsis<br />

thaliana. Using NDR1, a protein required <str<strong>on</strong>g>for</str<strong>on</strong>g> the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> R-protein mediated resistance in Arabidopsis, as<br />

a molecular and biochemical model <str<strong>on</strong>g>for</str<strong>on</strong>g> the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> resistance, we are working towards the elucidati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the dynamic linkages between effector-triggered immunity and the host actin cytoskelet<strong>on</strong>. To this end, we<br />

have begun characterizing several proteins required <str<strong>on</strong>g>for</str<strong>on</strong>g> reorganizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the actin cytoskelet<strong>on</strong> in plants, and<br />

have identified a genetic interacti<strong>on</strong> between bacterial effector acti<strong>on</strong> and the dynamics <str<strong>on</strong>g>of</str<strong>on</strong>g> the host<br />

cytoskelet<strong>on</strong>. Similarities between the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> Yersinia pestis effector proteins and P. syringae effectors will<br />

be discussed. Our working model suggests that the actin cytoskelet<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> plants may be a virulence target <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

pathogens.<br />

4B_04_S<br />

ELUCIDATION OF FUMONISIN B1-INDUCED CELL DEATH SIGNALLING IN<br />

ARABIDOPSIS THALIANA<br />

Daniel F. Tomé, John M. Hamilt<strong>on</strong>, Stephen Chivasa, Keith Lindsey, Ant<strong>on</strong>i R. Slabas<br />

Creative Gene Technology Ltd., Integrative Cell Biology Laboratory, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Durham, Durham DH1 3LE,<br />

United Kingdom. e-mail: j.m.hamilt<strong>on</strong>@durham.ac.uk<br />

Programmed Cell Death (pcd) is a ubiquitous process in plants that is very similar to apoptosis in animals. It<br />

occurs in various tissues during plant development, such as death <str<strong>on</strong>g>of</str<strong>on</strong>g> the tapetum cell layer in mature anthers,<br />

the sheading <str<strong>on</strong>g>of</str<strong>on</strong>g> leaves in autumn in perennial plants, and in the differentiati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> xylem. Pcd also occurs as a<br />

resp<strong>on</strong>se to stress such as pathogen attack. However, the mechanism by which plants initiate pcd and the key<br />

proteins involved are poorly understood. Recently, we reported that removal <str<strong>on</strong>g>of</str<strong>on</strong>g> extracellular ATP (eATP)<br />

from plant tissues initiates pcd. We also reported that fum<strong>on</strong>isin B1 (FB1)-induced pcd is mediated via eATP<br />

depleti<strong>on</strong> and can be blocked by additi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ATP. Rescue <str<strong>on</strong>g>of</str<strong>on</strong>g> cells from FB1-induced death by ATP is<br />

possible if ATP is supplied be<str<strong>on</strong>g>for</str<strong>on</strong>g>e the cells are irreversibly committed to death. Changes in gene expressi<strong>on</strong> at<br />

or around the time <str<strong>on</strong>g>of</str<strong>on</strong>g> commitment are <str<strong>on</strong>g>of</str<strong>on</strong>g> interest as these could be crucial players in cell death. We are now<br />

using 2-dimensi<strong>on</strong>al difference gel electrophoresis and DNA microarray technologies to identify the key<br />

comp<strong>on</strong>ents in this pathway, focusing <strong>on</strong> time points around commitment. We identified a number <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

differentially expressed genes and proteins associated with this death pathway. Reverse genetic screening<br />

using homozygous knockout mutants has allowed us to identify candidates that could prove critical to the<br />

executi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> pcd in plants. Moreover, the emerging data is beginning to give us insight into the mechanism<br />

by which eATP sustains cell viability in plants.<br />

197


23-26 August 2007,<br />

Budapest, Hungary<br />

4B_05_S<br />

(poster secti<strong>on</strong> B2, poster board #271, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DEHYDROASCORBATE UPTAKE IS IMPAIRED IN THE EARLY RESPONS OF<br />

ARABIDOPSIS PLANT CELL CULTURES TO CADMIUM<br />

Nele Horemans *a , Tine Raeymaekers * , Kim Van Beek * , David D’Haese * , Ann Cuypers ** ,<br />

Yves Guisez *<br />

* Plant Physiology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Antwerp, Groenenborgerlaan 171, 2020 Antwerp, Belgium,<br />

e-mail: nele.horemans@ua.ac.be<br />

** <str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g> Envir<strong>on</strong>mental Sciences, Hasselt University, Agoralaan Building D, 3590 Diepenbeek, Belgium<br />

The balance between antioxidants such as ascorbate (ASC) and glutathi<strong>on</strong>e and oxidative reactive oxygen<br />

species (ROS) is known to play a pivotal role in the resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> plant cells to abiotic stress. Here cell cultures<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> Arabidopsis thaliana were investigated <strong>on</strong> their resp<strong>on</strong>se to elevated levels <str<strong>on</strong>g>of</str<strong>on</strong>g> aluminum, zinc or cadmium.<br />

For Zn and Al no significant H 2 O 2 -accumulati<strong>on</strong> was detected. Cd, however, induced a rapid and<br />

c<strong>on</strong>centrati<strong>on</strong> dependent and initially extracellular H 2 O 2 -accumulati<strong>on</strong> that could be inhibited by DPI (20<br />

µM). RT-PCR analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> 3 Rboh genes showed an increased transcripti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Rboh F after 15 min. No change<br />

in ASC c<strong>on</strong>centrati<strong>on</strong> was observed during the first three hours after Cd-additi<strong>on</strong>. In c<strong>on</strong>trast glutathi<strong>on</strong>e<br />

levels completely diminished within <strong>on</strong>e hour. This drop could be attributed to an increase in phytochelatin 4.<br />

At the plasma membrane, Cd further induced a significant decrease in dehydroascorbate uptake activity (up to<br />

90% inhibiti<strong>on</strong> after four hours). This decrease is not present when cells are treated with LaCl 3, be<str<strong>on</strong>g>for</str<strong>on</strong>g>e<br />

exposure to CdCl 2 . LaCL 3 is a typical inhibitor <str<strong>on</strong>g>of</str<strong>on</strong>g> Ca channels and is known to prevent Cd uptake and Cd<br />

induced ROS producti<strong>on</strong> in plants. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e these results seem to indicate Cd uptake is a prerequisite <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

the change in DHA transport activity. However DPI did not prevent the drop in DHA uptake activity<br />

indicating that this resp<strong>on</strong>se seems to be independent <str<strong>on</strong>g>of</str<strong>on</strong>g> the H 2 O 2 -producti<strong>on</strong>. The possible functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

drop in DHA uptake in resp<strong>on</strong>se to Cd stress will be discussed.<br />

198


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

4B_01_P<br />

(poster secti<strong>on</strong> B2, poster board #272, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STRESS REACTIONS OCCURRING DURING BIRD CHERRY INFESTATION BY<br />

BIRD CHERRY-OAT APHID<br />

Hubert Sytykiewicz, Iw<strong>on</strong>a Łukasik, Bogumił Leszczyński<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Podlasie, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry and Molecular Biology,<br />

B. Prusa 12 St., 08-110 Siedlce, Poland<br />

e-mail: leszczb@ap.siedlce.pl<br />

Quite <str<strong>on</strong>g>of</str<strong>on</strong>g>ten biochemical interacti<strong>on</strong>s between host-plants and herbivorous insect pests are related to stress<br />

reacti<strong>on</strong>s within infested plant tissues and feeding herbivores. In the present paper we report <strong>on</strong> such dual<br />

stress reacti<strong>on</strong>s within tissues <str<strong>on</strong>g>of</str<strong>on</strong>g> the bird cherry-oat aphid (Rhopalosiphum padi L.) fed <strong>on</strong> the bird cherry<br />

(Prunus padus L.).The carried out experiments showed that the aphid infestati<strong>on</strong> had elucidated an increase in<br />

activity <str<strong>on</strong>g>of</str<strong>on</strong>g> leucylaminopeptidase and other proteinases isolated at pH 5 and pH 7 from the bird cherry leaves.<br />

It is important since these enzymes are involved in inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the plant defensive mechanisms towards<br />

various pathogens. It has been recognized that the <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the earliest plant resp<strong>on</strong>ses to herbivores involve<br />

generati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the reactive oxygen species and inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidative stress within insect tissues. It appears to<br />

be real <str<strong>on</strong>g>for</str<strong>on</strong>g> the studied aphid species since depleti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> total thiol groups and increase in lipid peroxidati<strong>on</strong><br />

was dem<strong>on</strong>strated within the bird cherry-oat aphid tissues exposured to plant phenolics. However, R. padi<br />

showed an ability to neutralize the free oxygen radicals with help <str<strong>on</strong>g>of</str<strong>on</strong>g> enzymatic and n<strong>on</strong>-enzymatic<br />

antioxidants. For example, an inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the aphid superoxide dismutase and catalase, neutralizing toxic<br />

oxygen <str<strong>on</strong>g>for</str<strong>on</strong>g>ms was found during the spring migrati<strong>on</strong> period, when the highest level <str<strong>on</strong>g>of</str<strong>on</strong>g> the proteinases activity<br />

within the bird cherry leaves was noted. The role <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress reacti<strong>on</strong>s in biochemical interacti<strong>on</strong>s between<br />

the bird cherry and the bird cherry-oat aphid is discussed.<br />

4B_02_P<br />

(poster secti<strong>on</strong> B2, poster board #273, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PROTEOMIC ANALYSIS OF LEAF RUST-INFECTED WHEAT<br />

V. Pós 1 , K. Manninger 2 , K. Halász 1 , É. Hunyadi-Gulyás 3 , E. Szájli 3 , M. Cserháti 4 , K. Medzihradszky 3 ,<br />

J. Györgyey 4 , N. Lukács 1<br />

1Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Physiology and Biochemistry, CUB, Budapest<br />

2Plant Protecti<strong>on</strong> Inst., Budapest<br />

3BRC Proteomics Res. Group, Szeged<br />

4BRC Inst. <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Biol., Szeged<br />

e-mail: noemi.lukacs@uni-corvinus.hu<br />

The extracellular matrix plays an important role in plant defense against biotic stress. Presuming that the<br />

genetic background <str<strong>on</strong>g>of</str<strong>on</strong>g> near-isogenic wheat lines differing in their resistance gene(s) has an influence <strong>on</strong> the<br />

protein compositi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> extracellular matrix, we analyzed their intercellular washing fluid (ICF) to identify<br />

fungal infecti<strong>on</strong>-associated changes in the protein pattern and to detect secreted proteins which may<br />

c<strong>on</strong>tribute to resistance reacti<strong>on</strong>. Seedlings <str<strong>on</strong>g>of</str<strong>on</strong>g> the susceptible ’Thatcher’ and the corresp<strong>on</strong>ding near-isogenic<br />

Lr1 and Lr9 wheat lines resistant to the pathotype 43722 <str<strong>on</strong>g>of</str<strong>on</strong>g> leaf rust (P. rec<strong>on</strong>dita fsp. tricii) were inoculated<br />

and apoplastic fluid was collected over a week p.i. Proteins were separated <strong>on</strong> 1- and 2D PAGE and<br />

identified by MALDI-TOF and LC-MS-MS. In the Lr1-resistant line three apoplastic proteins, a glucan-endo-<br />

199


23-26 August 2007,<br />

Budapest, Hungary<br />

1,3-beta-D-glycosidase (35413 Da, pI 8.8), a chitinase 1 enzyme (27077 Da, pI 8.7) and a pathogenesis-related<br />

protein, PR1.1 (17651 Da, pI 8.7) were induced earlier and reached higher c<strong>on</strong>centrati<strong>on</strong> than in the Thatcher<br />

line. In additi<strong>on</strong>, the ICF <str<strong>on</strong>g>of</str<strong>on</strong>g> both Lr1 and Lr9 showed much higher 1,3-endoglucanase activity and different<br />

inducti<strong>on</strong> kinetics as that <str<strong>on</strong>g>of</str<strong>on</strong>g> the ‘Thatcher’ line. Since it is well known that these proteins play an important<br />

role in the antifungal defence <str<strong>on</strong>g>of</str<strong>on</strong>g> resistant as well as <str<strong>on</strong>g>of</str<strong>on</strong>g> sensitive plants, their differential expressi<strong>on</strong> might be<br />

caused by differences in gene regulati<strong>on</strong>. Results <str<strong>on</strong>g>of</str<strong>on</strong>g> the promoter analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> the corresp<strong>on</strong>ding homologous<br />

genes in rice will be shown to point out potential comm<strong>on</strong> motifs in the promoter regi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the most<br />

homologous proteins.<br />

4B_03_P<br />

(poster secti<strong>on</strong> B2, poster board #274, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

TITLE: ROLE OF MAIZE PHENOLICS IN THE GENOTYPIC RESISTANCE TO<br />

GIBBERELLA STALK ROT (FUSARIUM GRAMINEARUM SCHWABE)<br />

Rogelio Santiago*, Lana M. Reid, John T. Arnas<strong>on</strong>, German Sandoya, Rosa A. Malvar<br />

*C<strong>on</strong>sejo Superior de Investigaci<strong>on</strong>es Científicas, Misión Biológica de Galicia.<br />

Carballeira nº 8- Salcedo, P<strong>on</strong>tevedra. 36143 SPAIN<br />

Teleph<strong>on</strong>e: +34 986 85 48 00, Fax: +34 986 84 13 72<br />

e-mail: rsantiago@mbg.cesga.es<br />

Six maize inbred lines known to represent a wide spectrum <str<strong>on</strong>g>of</str<strong>on</strong>g> susceptibility to Gibberella stalk rot were<br />

investigated <str<strong>on</strong>g>for</str<strong>on</strong>g> a relati<strong>on</strong>ship between their phenolic c<strong>on</strong>tents in the pith and their resistance to Gibberella<br />

stalk rot. The phenolic acid pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iles were evaluated from silking to grain maturity. Four different fracti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

phenolic compounds were extracted from inoculated and n<strong>on</strong>-inoculated (c<strong>on</strong>trol) pith tissues: insoluble cell<br />

wall-bound phenolics, free phenolics, soluble ester-bound phenolics, and soluble glycoside-bound phenolics.<br />

Analysis by HPLC revealed that p-coumaric and ferulic acid were the most abundant compounds in the<br />

soluble and cell wall bound fracti<strong>on</strong>s. The quantity <str<strong>on</strong>g>of</str<strong>on</strong>g> free, glycoside-bound and ester-bound phenolics in the<br />

pith was lower than the level required <str<strong>on</strong>g>for</str<strong>on</strong>g> inhibiti<strong>on</strong> to <str<strong>on</strong>g>of</str<strong>on</strong>g> Fusarium growth and/or mycotoxins producti<strong>on</strong>;<br />

however, significant negative correlati<strong>on</strong>s between diferulic acids c<strong>on</strong>tents in the cell walls and disease<br />

severity rating four days after inoculati<strong>on</strong> were found. According to these results previous studies showed<br />

significant negative correlati<strong>on</strong>s between disease severity and diferulic acid c<strong>on</strong>tents in the maize grain.<br />

Special attenti<strong>on</strong> should be there<str<strong>on</strong>g>for</str<strong>on</strong>g>e given to levels <str<strong>on</strong>g>of</str<strong>on</strong>g> diferulic acids during the early infecti<strong>on</strong> process.<br />

Diferulates may play a role in genotypic resistance <str<strong>on</strong>g>of</str<strong>on</strong>g> maize to Gibberella stalk rot as barriers pre<str<strong>on</strong>g>for</str<strong>on</strong>g>med prior<br />

to infecti<strong>on</strong>.<br />

200


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

4C. STRESS SIGNALING IN PLANTS<br />

(CSABA KONCZ)<br />

4C_01_S<br />

REGURATORY GENE NETWORK IN DROUGHT STRESS RESPONSE<br />

Kazuo Shinozaki 1 , Kazuko Yamaguchi-Shinozaki 2, 3<br />

1RIKEN Plant Science Center, Suehiro-cho, Tsurumi-ku, Yokohama, Japan, 2 Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular Physiology,<br />

Graduate School <str<strong>on</strong>g>of</str<strong>on</strong>g> Agricultural and Life Sciences, The University <str<strong>on</strong>g>of</str<strong>on</strong>g> Tokyo, Bunkyo-ku, Tokyo, Japan, 3 Biological<br />

Resources Divisi<strong>on</strong>, Japan Internati<strong>on</strong>al <str<strong>on</strong>g>Research</str<strong>on</strong>g> Center <str<strong>on</strong>g>for</str<strong>on</strong>g> Agricultural Sciences (JIRCAS), Tsukuba, Japan<br />

e-mail: sinozaki@rtc.riken.jp<br />

Drought stress induces a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> genes <str<strong>on</strong>g>of</str<strong>on</strong>g> which products functi<strong>on</strong> in drought stress tolerance and<br />

resp<strong>on</strong>se in plants. Many stress-inducible genes have been used to improve stress tolerance by gene transfer.<br />

In this c<strong>on</strong>gress, we present our recent studies <strong>on</strong> molecular mechanism in drought stress resp<strong>on</strong>se and<br />

tolerance. We have identified complex regulatory systems in stress-resp<strong>on</strong>sive gene expressi<strong>on</strong>: ABAdependent<br />

and ABA-independent systems. In <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the ABA-independent pathways, a cis-acting element<br />

(DRE/CRT) and its binding proteins, DREB2s, are important cis- and trans-acting elements in droughtresp<strong>on</strong>sive<br />

gene expressi<strong>on</strong>, respectively. DREB2 is also involved in heat stress resp<strong>on</strong>se. In the ABAdependent<br />

pathways, bZIP transcripti<strong>on</strong> factors (AREB/ABF) are involved in the major process. Protein<br />

phosphorylati<strong>on</strong> is important <str<strong>on</strong>g>for</str<strong>on</strong>g> the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> AREB proteins. The MYB/MYC and NAC transcripti<strong>on</strong><br />

factors are involved in ABA-resp<strong>on</strong>sive gene expressi<strong>on</strong> and jasm<strong>on</strong>ic acid resp<strong>on</strong>se In the ABA-dependent<br />

pathway, stress-inducible NCED3 is mainly involved in the ABA biosynthesis during drought stress. We<br />

analyzed metabolic pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iles regulated by ABA using T-DNA tagged mutant and with GC-MS and LC-MS.<br />

We analyzed the functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> CYP707A3 in the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ABA metabolism during stress resp<strong>on</strong>ses. We<br />

also report the functi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> SnRK2 protein kinases in drought and ABA resp<strong>on</strong>ses using mutants and<br />

transgenic overexpressors.<br />

4C_02_S<br />

STRESS-REGULATED SIRNAS AND MIRNAS<br />

Jian-Kang Zhu<br />

Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Integrative Genome Biology, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Botany and Plant Sciences, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Cali<str<strong>on</strong>g>for</str<strong>on</strong>g>nia, Riverside<br />

It is <str<strong>on</strong>g>of</str<strong>on</strong>g>ten presumed that RNA interference (RNAi) evolved as a cellular surveillance mechanism to silence<br />

“<str<strong>on</strong>g>for</str<strong>on</strong>g>eign” double stranded RNAs (dsRNAs) so that cells can defend against viral infecti<strong>on</strong> or transpos<strong>on</strong>s.<br />

However, recent work suggests a widespread occurrence <str<strong>on</strong>g>of</str<strong>on</strong>g> dsRNAs unrelated to viral replicati<strong>on</strong> or<br />

transpos<strong>on</strong>s. A significant porti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular dsRNAs can be <str<strong>on</strong>g>for</str<strong>on</strong>g>med by natural antisense transcripts (NATs).<br />

Approximately 10-20% <str<strong>on</strong>g>of</str<strong>on</strong>g> genes in plant and animal genomes encode cis-NATs, i.e. these genes overlap but<br />

are <strong>on</strong> the opposite strands <str<strong>on</strong>g>of</str<strong>on</strong>g> DNA. In Arabidopsis, dsRNAs generated from cis-NATs can be processed by<br />

Dicer-like enzymes into 21-24 nt nat-siRNAs which then direct the cleavage <str<strong>on</strong>g>of</str<strong>on</strong>g> complementary mRNAs.<br />

Evidence indicates that many cis-NAT genes are regulated by biotic and abiotic stresses, and they generate<br />

nat-siRNAs <strong>on</strong>ly under specific stress c<strong>on</strong>diti<strong>on</strong>s. The stress-induced nat-siRNAs are important comp<strong>on</strong>ents<br />

in the regulatory circuits leading to stress acclimati<strong>on</strong>. These results suggest that RNAi is not <strong>on</strong>ly critical <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

cellular surveillance but it is also an important cellular gene regulatory mechanism. In this presentati<strong>on</strong>, I will<br />

201


23-26 August 2007,<br />

Budapest, Hungary<br />

talk about abiotic stress-regulated nat-siRNAs and other endogenous siRNAs in Arabidopsis. In additi<strong>on</strong>, I<br />

will discuss the role <str<strong>on</strong>g>of</str<strong>on</strong>g> miRNAs in plant resp<strong>on</strong>ses to abiotic stresses.<br />

4C_03_S<br />

FROM MUTANTS TO GENES, PATHWAYS AND NETWORKS<br />

IN PLANT STRESS SIGNALING<br />

Hans J. Bohnert, Shis<strong>on</strong>g Ma, Qingqiu G<strong>on</strong>g, Pinghua Li<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Illinois at Urbana-Champaign, Plant Biology<br />

Arabidopsis thaliana transcript pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iles can report the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> abiotic and biotic stresses <strong>on</strong> tissue- and cellspecific<br />

gene expressi<strong>on</strong>. Organizing these datasets could reveal the structure and mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> resp<strong>on</strong>ses<br />

and cross-talk between pathways, and in which cells the plants perceive, signal, resp<strong>on</strong>d to and integrate<br />

envir<strong>on</strong>mental inputs.<br />

We have clustered Arabidopsis transcript pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iles <str<strong>on</strong>g>for</str<strong>on</strong>g> >30 treatments, comprising abiotic, biotic and chemical<br />

stresses. Ubiquitous stress resp<strong>on</strong>ses in Arabidopsis, similar to those <str<strong>on</strong>g>of</str<strong>on</strong>g> fungal and animal cells, employ genes<br />

in pathways related to MAP kinases, Snf1-related kinase, vesicle transport, mitoch<strong>on</strong>drial functi<strong>on</strong>s, and the<br />

fundamental transcripti<strong>on</strong> machinery. The induced resp<strong>on</strong>se to various stresses can be attributed to genes<br />

whose promoters are characterized by a small number <str<strong>on</strong>g>of</str<strong>on</strong>g> comm<strong>on</strong> regulatory motifs, while sec<strong>on</strong>dary motifs<br />

have also been identified. Most genes that are down-regulated by stress show distinct tissue-specific<br />

expressi<strong>on</strong> patterns and appear to be under strict developmental regulati<strong>on</strong>. The ABA-dependent<br />

transcriptome is delineated in the cluster structure, while functi<strong>on</strong>s dependent <strong>on</strong> reactive oxygen species are<br />

widely distributed, possibly indicating evoluti<strong>on</strong>ary pressures c<strong>on</strong>ferring distincti<strong>on</strong> to different stresses in<br />

time and space. Cell lineages in the root express stress-resp<strong>on</strong>sive genes at different levels. The intersecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

stress-resp<strong>on</strong>sive and cell-specific pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iles identified cell lineages affected by abiotic stress.<br />

In an extensi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these studies, the Graphical Gaussian Model (GGM) was used to assemble a gene network<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> the Arabidopsis transcriptome. Based <strong>on</strong> partial correlati<strong>on</strong> (pcor), GGM infers co-regulati<strong>on</strong> patterns<br />

between gene pairs c<strong>on</strong>diti<strong>on</strong>al <strong>on</strong> the behavior <str<strong>on</strong>g>of</str<strong>on</strong>g> other genes. We used ‘regularized’ GGM, coupled with<br />

iterative random samplings, to expand the network to cover the whole Arabidopsis genome (22,266 genes).<br />

This resulted in a network <str<strong>on</strong>g>of</str<strong>on</strong>g> ~18,000 interacti<strong>on</strong>s (edges) am<strong>on</strong>g ~7,000 genes (nodes) with high c<strong>on</strong>fidence<br />

(p < 2.2E-19), where the c<strong>on</strong>necti<strong>on</strong>s represented ~0.01% <str<strong>on</strong>g>of</str<strong>on</strong>g> all possible edges.<br />

When querying <str<strong>on</strong>g>for</str<strong>on</strong>g> selected genes, locally coherent sub-networks emerged that were predominantly related to<br />

metabolic functi<strong>on</strong>s and stress resp<strong>on</strong>ses. Sub-networks <str<strong>on</strong>g>for</str<strong>on</strong>g> sulfate, phosphate, nitrogen, carbohydrate,<br />

tryptophan, cell wall metabolism, and the cold stress resp<strong>on</strong>se were analyzed in detail. GGM recovered<br />

interacti<strong>on</strong>s with biological significance that escaped capture by Pears<strong>on</strong> correlati<strong>on</strong> networks, while<br />

eliminating ambiguous interacti<strong>on</strong>s inherent in the latter. GGM displayed many known co-regulati<strong>on</strong><br />

pathways as sub-networks and added novel comp<strong>on</strong>ents to known edges. Finally, the network rec<strong>on</strong>ciled<br />

individual sub-networks in a topology joined at the whole genome level, and provided a general framework<br />

that can instruct future studies <strong>on</strong> plant metabolism and stress resp<strong>on</strong>ses.<br />

202


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

4C_04_S<br />

REGULATORY ROLES OF AMP-ACTIVATED PROTEIN KINASES AND PRL1-CDC5<br />

SPLICEOSOME ASSEMBLY COMPLEX IN PLANT STRESS SIGNALLING<br />

Csaba K<strong>on</strong>cz, Dóra Szak<strong>on</strong>yi, Gergely Molnár, Mihály Horváth, Zsuzsa K<strong>on</strong>cz<br />

Max-Planck Institut für Züchtunsgs<str<strong>on</strong>g>for</str<strong>on</strong>g>shung, Carl-v<strong>on</strong>-Linne-Weg 10¸ 50829 Köln, Germany<br />

e-mail: k<strong>on</strong>cz@mpiz-koeln.mpg.de<br />

AMP-activated kinases (AMPKs) c<strong>on</strong>trol essential metabolic and signaling pathways in resp<strong>on</strong>se to stress<br />

stimuli. Trimeric AMPKs, called Snf1-related kinases (SnRK1s) in Arabidopsis, are <str<strong>on</strong>g>for</str<strong>on</strong>g>med by combinatorial<br />

assembly <str<strong>on</strong>g>of</str<strong>on</strong>g> 3 catalytic alpha (AKIN10, 11 and 12), 3 substrate targeting beta (AKINβ1, 2 and3) and <strong>on</strong>e<br />

AMP-binding gamma (AKINβγ/SNF4) subunits. SnRK1s undergo self-activati<strong>on</strong> by autophosphorylati<strong>on</strong>,<br />

their genes show different regulati<strong>on</strong>, and the stability <str<strong>on</strong>g>of</str<strong>on</strong>g> subunits is c<strong>on</strong>trolled by proteasomal degradati<strong>on</strong>.<br />

SnRK1α kinases are found in complex with the α7 subunit <str<strong>on</strong>g>of</str<strong>on</strong>g> proteasome and SKP1 subunit <str<strong>on</strong>g>of</str<strong>on</strong>g> SCF ubiquitin<br />

ligases suggesting a role in ubiquitinati<strong>on</strong>-dependent protein degradati<strong>on</strong>. Genetic dissecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> SnRK1<br />

signaling is hampered by the lack <str<strong>on</strong>g>of</str<strong>on</strong>g> akin10 and akin11 inserti<strong>on</strong> mutati<strong>on</strong>s and deficient male transmissi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

snrk1γ mutants. Inducible overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> SnRK1α AKIN10 results in hypersensitivity to the stress<br />

horm<strong>on</strong>e abscisic acid (ABA). AKIN10 phosphorylates and thereby stabilizes the bZIP transcripti<strong>on</strong> factor<br />

ABI5, a key regulator <str<strong>on</strong>g>of</str<strong>on</strong>g> germinati<strong>on</strong> resp<strong>on</strong>se to ABA. SnRK1α subunits interact with the nuclear WD-40<br />

repeat protein PRL1 (Pleiotropic Regulator Locus 1) that functi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> SnRK1-inhibitor in vitro. In additi<strong>on</strong> to<br />

other defects, prl1 mutants display ABA hypersensitivity, and hyperphosphorylati<strong>on</strong> and stabilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

ABI5. Stability <str<strong>on</strong>g>of</str<strong>on</strong>g> PRL1 is c<strong>on</strong>trolled by proteasomal degradati<strong>on</strong>. PRL1 is found in complex with the Myb3R<br />

factor CDC5 that immunoprecipitates the proteasome, CULLIN1, and several unknown ubiquitinated<br />

proteins. PRL1 and CDC5 are c<strong>on</strong>served subunits <str<strong>on</strong>g>of</str<strong>on</strong>g> Ntc (nineteen-complex) spliceosome-activating<br />

complex. Inactivati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> CDC5 results in prl1-like phenotype suggesting functi<strong>on</strong>al interdependence. Both<br />

prl1 and cdc5 mutati<strong>on</strong>s cause early flowering and changes in petal and stamen development. Steady-state<br />

mRNA level <str<strong>on</strong>g>of</str<strong>on</strong>g> FLC, a key repressor <str<strong>on</strong>g>of</str<strong>on</strong>g> flowering, is reduced and FLC pre-mRNA shows defective splicing<br />

in prl1. Floral defects <str<strong>on</strong>g>of</str<strong>on</strong>g> prl1 reflect aberrant splicing <str<strong>on</strong>g>of</str<strong>on</strong>g> pre-mRNAs <str<strong>on</strong>g>of</str<strong>on</strong>g> floral homeotic genes AP1, AP3, AG<br />

and PI. Alterati<strong>on</strong>s in the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> microRNAs c<strong>on</strong>trolling ABA resp<strong>on</strong>se, leaf development and flowering<br />

time in prl1 suggest implicati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> spliceosome activating complex and SnRK1s in the c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> pre-mRNA<br />

splicing and biogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> small inhibitory RNAs.<br />

4C_05_S<br />

(poster secti<strong>on</strong> B2, poster board #275, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CHARACTERIZATION OF BIOCHEMICAL PROPERTIES OF A RICE<br />

DEHYDRATION INDUCIBLE SUCROSE NONFERMENTING1 (SNF1)-RELATED<br />

PROTEIN KINASE 2 (SNRK2) FAMILY<br />

Sun Joo Kim 1 , Myung Hee Nam 2 , Dool Yi Kim 1 , Ye<strong>on</strong> Hee Lee 1 , Seok-Cheol Suh 1 , In-Sun Yo<strong>on</strong> 1 *<br />

1Cell and Genetics Divisi<strong>on</strong>, Nati<strong>on</strong>al Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Agricultural Biotechnology, Suw<strong>on</strong> 441-707, Republic <str<strong>on</strong>g>of</str<strong>on</strong>g> Korea.<br />

2Metabolome Analysis Team, Korea Basic Science Institute<br />

* e-mail: isyo<strong>on</strong>@rda.go.kr<br />

OSRK1 is a rice SnRK2 protein kinase activated by hyperosmotic stress and ABA. In the present study, we<br />

investigated regulatory mechanism and down stream targets <str<strong>on</strong>g>of</str<strong>on</strong>g> OSRK1. GST-fused recombinant OSRK1<br />

showed str<strong>on</strong>g substrate preference <str<strong>on</strong>g>for</str<strong>on</strong>g> rice bZIP transcripti<strong>on</strong> factors and uncomm<strong>on</strong> c<str<strong>on</strong>g>of</str<strong>on</strong>g>actor requirement<br />

203


23-26 August 2007,<br />

Budapest, Hungary<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> Mn 2+ over Mg 2+ . We observed positive relati<strong>on</strong>ship between autophosphorylati<strong>on</strong> level and kinase activity.<br />

Moreover, OSRK1 could transphosphorylate itself. By deleti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> C-terminus 73 amino acids or mutati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Ser-158 and Thr-159 to aspartic acids (Asp) in the activati<strong>on</strong> loop, the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> OSRK1 was dramatically<br />

decreased. Our data suggests that inter-molecular (auto)phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> catalytic domain <str<strong>on</strong>g>of</str<strong>on</strong>g> OSRK1 is<br />

important <str<strong>on</strong>g>for</str<strong>on</strong>g> enzyme activati<strong>on</strong>. In an attempt to investigate OSRK1 signaling comp<strong>on</strong>ents, we identified<br />

two putative calcium binding proteins by yeast two hybrid screening. In vitro OSRK1 activity was stimulated<br />

by those CBPs. By in gel kinase assays, we also showed that ca. 52 kDa and 61 kDa protein kinase activities<br />

were highly stimulated in resp<strong>on</strong>se to salt or ABA in roots <str<strong>on</strong>g>of</str<strong>on</strong>g> transgenic rice over-expressing OSRK1. They<br />

are likely to be downstream target kinases <str<strong>on</strong>g>for</str<strong>on</strong>g> OSRK1 signaling pathway. This work was supported by On-<br />

Site Cooperative Agriculture <str<strong>on</strong>g>Research</str<strong>on</strong>g> Project, RDA, Republic <str<strong>on</strong>g>of</str<strong>on</strong>g> Korea.<br />

4C_06_S<br />

(poster secti<strong>on</strong> B2, poster board #276, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DEAD-BOX HELICASES IN PLANT STRESS TOLERANCE<br />

Narendra Tuteja<br />

Internati<strong>on</strong>al <str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g> Genetic Engineering & Biotechnology, New Delhi<br />

e-mail: narendra@icgeb.res.in<br />

Abiotic stress is an increasing threat in reducing agricultural productivity worldwide. The high salinity stress<br />

impairs crop producti<strong>on</strong> <strong>on</strong> at least 20% <str<strong>on</strong>g>of</str<strong>on</strong>g> irrigated land. As salinity stress affects the cellular geneexpressi<strong>on</strong><br />

machinery, it is evident that molecules involved in nucleic acid processing including helicases, are<br />

likely to be affected as well. Helicases are <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the smallest molecular motors <str<strong>on</strong>g>of</str<strong>on</strong>g> biological system, which<br />

harness the chemical free energy <str<strong>on</strong>g>of</str<strong>on</strong>g> ATP hydrolysis to catalyze the opening <str<strong>on</strong>g>of</str<strong>on</strong>g> energetically stable duplex<br />

DNA (called DNA helicases) or unfold the sec<strong>on</strong>dary structures in RNA (called RNA helicases) and thereby<br />

are involved in almost all aspect <str<strong>on</strong>g>of</str<strong>on</strong>g> nucleic acid metabolism. Mostly all the helicases c<strong>on</strong>tain some c<strong>on</strong>served<br />

signature motifs including DEAD-box, which act as an engine to power DNA unwinding. In plants the role<br />

helicases in abiotic stress is just begening to emerge. Here we present the isolati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> two pea DNA helicases<br />

(PDH45 and PDH47) and their role in salinity stress tolerance. PDH45 and PDH47 both are homologous to<br />

eIF4A, involved in translati<strong>on</strong> initiati<strong>on</strong>, localized in nucleus and cytosol, and c<strong>on</strong>tain multiple activities<br />

including DNA and RNA unwinding, ATP-binding and ATPase. The transcript <str<strong>on</strong>g>of</str<strong>on</strong>g> both the genes were<br />

found to upregulated under salinity stress. However, both the proteins c<strong>on</strong>tain <strong>on</strong>ly 56% identity and differ in<br />

following properties: PDH45 is unipolar, active at <strong>on</strong>ly basic pH, expressed more in root than shoot and not<br />

regulated by ABA treatment; while PDH47 is a bipolar enzyme, active at both acidic and basic pH, expressed<br />

more in shoot than root and upregulated by the ABA treatment. The over-expressi<strong>on</strong> PDH45 in tobacco<br />

plants c<strong>on</strong>fers high salinity tolerance without yield loss, thus suggesting a new pathway <str<strong>on</strong>g>for</str<strong>on</strong>g> manipulating stress<br />

tolerance in crop plants.<br />

204


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

4C_01_P<br />

(poster secti<strong>on</strong> B2, poster board #277, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

INDUCTION OF LECTIN EXPRESSION IN RICE AS A REPONSE TO STRESS<br />

TREATMENTS<br />

Elke Fouquaert, Godelieve Gheysen, Els J. M. Van Damme<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular Biotechnology, Ghent University, Coupure Links 653, 9000 Gent, Belgium<br />

e-mail: Elke.Fouquaert@Ugent.be<br />

Numerous wild and cultivated plant species c<strong>on</strong>tain carbohydrate-binding proteins called lectins. Most plant<br />

lectins are involved in the recogniti<strong>on</strong> and binding <str<strong>on</strong>g>of</str<strong>on</strong>g> glycans from <str<strong>on</strong>g>for</str<strong>on</strong>g>eign organisms, and accordingly are<br />

believed to play a role in plant defense. However, in recent years evidence has accumulated that plants<br />

synthesize well-defined carbohydrate binding proteins up<strong>on</strong> exposure to stresses like drought, high salt,<br />

wounding, treatment with some plant horm<strong>on</strong>es or pathogen attack.<br />

As early as 1990 Claes et al. (1) dem<strong>on</strong>strated that a protein called SalT is specifically induced when rice plants<br />

are subjected to salt stress. Later SalT was identified as a dimeric mannose-binding jacalin-related lectin<br />

composed <str<strong>on</strong>g>of</str<strong>on</strong>g> 15 kDa subunits (2). More recently we have identified two more lectins in rice, bel<strong>on</strong>ging to the<br />

class <str<strong>on</strong>g>of</str<strong>on</strong>g> OSR40 proteins and a Galanthus nivalis-related lectin, respectively. Expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the OSR40 proteins<br />

is induced by salt stress or ABA treatment (3).<br />

Localizati<strong>on</strong> studies <str<strong>on</strong>g>of</str<strong>on</strong>g> the rice lectins dem<strong>on</strong>strated that they are exclusively located in the cytoplasm and the<br />

nucleus. The occurrence <str<strong>on</strong>g>of</str<strong>on</strong>g> the genes encoding the three different lectins in the rice genome was studied by<br />

screening different rice species and cultivars with a different genetic background. Lectin inducti<strong>on</strong> in<br />

resp<strong>on</strong>se to different stress c<strong>on</strong>diti<strong>on</strong>s was also analyzed.<br />

Since these lectins are synthesized <strong>on</strong>ly as a resp<strong>on</strong>se to specific physical, chemical and biotic stress factors, it<br />

can be assumed that they play a specific physiological role in the plant.<br />

4C_02_P<br />

(poster secti<strong>on</strong> B2, poster board #278, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECT OF LOW-CONCENTRATION STRESSORS IN BARLEY SEEDLINGS<br />

Erika Kovács 1,2 , Péter Nyitrai 1 , Pálma Czövek 1 , Mihály Óvári 3 , Ár<strong>on</strong> Keresztes 2<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Physiology and Molecular Plant Biology, Eötvös University, H-1518 Budapest<br />

P.O.B. 120, Hungary<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Anatomy, Eötvös University, H-1518 Budapest, P.O.B. 120, Hungary<br />

3Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Organic Chemistry, Eötvös University, H-1518 Budapest, P.O.B. 112, Hungary<br />

e-mail: k-erika@elte.hu<br />

The effects <str<strong>on</strong>g>of</str<strong>on</strong>g> some chemical stressors in high c<strong>on</strong>centrati<strong>on</strong> have been thoroughly investigated in plants, but<br />

in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> about the same stressors in low c<strong>on</strong>centrati<strong>on</strong> is very limited. In our experiments the effects <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

low-c<strong>on</strong>centrati<strong>on</strong> Cd 2+ (5x10 -8 M) and a herbicide, DCMU (10 -7 M) were investigated in barley seedlings.<br />

Chlorophyll accumulati<strong>on</strong> indicated a stimulative effect <str<strong>on</strong>g>of</str<strong>on</strong>g> both stressors in treated plants. Our aim was to<br />

reveal the mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> this stimulati<strong>on</strong>. The amount <str<strong>on</strong>g>of</str<strong>on</strong>g> active cytokinins increased in treated roots (prior to<br />

their transport to the leaves). This seems to be the key event in stimulati<strong>on</strong>. In order to identify the signaling<br />

pathway involved in the increased cytokinin synthesis <str<strong>on</strong>g>of</str<strong>on</strong>g> roots, the PIP 2 -IP 3 /DAG and MAPK pathways<br />

were tested with different inhibitors added together with the stressors to the nutrient soluti<strong>on</strong>. It seems that<br />

the signal is transmitted through DAG and protein kinase C to the MAPK pathway. It is known that higher<br />

205


23-26 August 2007,<br />

Budapest, Hungary<br />

amounts <str<strong>on</strong>g>of</str<strong>on</strong>g> Cd 2+ or DCMU cause oxidative stress in plants, up<strong>on</strong> which the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> antioxidant enzymes<br />

(e.g. SOD) increases, and mal<strong>on</strong>yl dialdehyde (MDA) might be finally produced by peroxidati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

membrane lipids. After applicati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Cd 2+ and DCMU, the amounts <str<strong>on</strong>g>of</str<strong>on</strong>g> SOD and MDA were measured to<br />

ascertain whether also low-c<strong>on</strong>centrati<strong>on</strong> stressors may cause oxidative stress. We found a slight transient<br />

increase in the amount <str<strong>on</strong>g>of</str<strong>on</strong>g> SOD in roots.<br />

4C_03_P<br />

(poster secti<strong>on</strong> B2, poster board #279, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CALCIUM SENSORS ARE REGULATED BY DIFFERENT STRESS SIGNALS<br />

Ana Lima, Edgar Cruz e Silva, Etelvina Figueira<br />

<str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g> Cell Biology, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Aveiro, Portugal<br />

e-mail: agusmaolima@gmail.com<br />

In order to adapt to envir<strong>on</strong>mental stresses, plants use diverse signalling strategies. Calcium functi<strong>on</strong>s as a<br />

versatile messenger in mediating resp<strong>on</strong>ses to biotic/abiotic stress signals and a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> developmental cues<br />

in plants. The signal-specific Ca signature is readily decoded by an array <str<strong>on</strong>g>of</str<strong>on</strong>g> Ca-binding proteins or Ca sensor.<br />

However, little is known about the specificity <str<strong>on</strong>g>of</str<strong>on</strong>g> calcium signatures and how the signal is decoded. Several<br />

families <str<strong>on</strong>g>of</str<strong>on</strong>g> Ca 2+ sensors have been identified in higher plants, such as calmodulin (CaM) and CBL (calcineurin<br />

B-like) proteins. Recent investigati<strong>on</strong>s have dem<strong>on</strong>strated that this protein variety may be are related to the<br />

specificity <str<strong>on</strong>g>of</str<strong>on</strong>g> stress signal percepti<strong>on</strong> and to the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the corresp<strong>on</strong>ding stress tolerance mechanisms.<br />

There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, the knowledge <str<strong>on</strong>g>of</str<strong>on</strong>g> which calcium sensors are triggered by specific stress signals is <str<strong>on</strong>g>of</str<strong>on</strong>g> major<br />

importance. Calcium-binding proteins were isolated from Vitis vinifera cells, after exposure to several stresses,<br />

with the aim to understand the specificity and regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> each individual stress. Vitis vinifera cell cultures<br />

were exposed to salt, heavy metal, heat and cold stresses. Calcium-binding proteins were purified by<br />

hydrophobic interacti<strong>on</strong> chromatography. Results dem<strong>on</strong>strated that exposure to different stresses induced<br />

alterati<strong>on</strong>s in the proteins eluted. Protein pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iles were separated by SDS-PAGE and Native PAGE in the<br />

presence and absence <str<strong>on</strong>g>of</str<strong>on</strong>g> calcium. The eluted proteins comprised not <strong>on</strong>ly calcium binding proteins, but also<br />

their targets, since <strong>on</strong>ly few proteins presented calcium-induced alterati<strong>on</strong>s in their c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>s.Results<br />

provide new insights in stress signal transducti<strong>on</strong> and corroborate the importance <str<strong>on</strong>g>of</str<strong>on</strong>g> calcium signaling in<br />

plant stress resp<strong>on</strong>ses.<br />

4C_04_P<br />

(poster secti<strong>on</strong> B2, poster board #280, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ADAPTATION MECHANISMS IN RICE (ORYZA SATIVA L.) UNDER SALT STRESS<br />

A. Kader, S. Lindberg<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Botany, Stockholm university, 106 91 Stockholm, Sweden<br />

e-mail: Sylvia.Lindberg@botan.su.se<br />

The project focuses <strong>on</strong> two important aspects <str<strong>on</strong>g>of</str<strong>on</strong>g> Na + toxicity in salt-tolerant rice cv. Pokkali and salt-sensitive<br />

cv. BRRI Dhan29, namely i) how Na + stress induces a change in cytosolic Ca 2+ , [Ca 2+ ] cyt , and pH, [pH] cyt , and<br />

ii) how cells could maintain a low cytosolic Na + and/or Na + /K + ratio. The salt-induced changes in [Ca 2+ ] cyt<br />

and [pH] cyt and their sources were m<strong>on</strong>itored in single rice protoplasts by fluorescence microscopy. The<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the transporter genes OsHKT1, OsHKT2 and OsVHA, which are thought to play a significant<br />

206


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

role in maintaining correct cytosolic Na + and or Na + /K + ratio, were examined in both rice cultivars under salt<br />

stress c<strong>on</strong>diti<strong>on</strong> by real time RT-PCR and in situ PCR. The results show that Na + must be sensed inside the<br />

cytosol, be<str<strong>on</strong>g>for</str<strong>on</strong>g>e any changes in [Ca 2+ ] cyt and [pH] cyt occur. Sensing <str<strong>on</strong>g>of</str<strong>on</strong>g> Na + induced different changes in [Ca 2+ ] cyt<br />

and [pH] cyt in the two rice cultivars with different sources <str<strong>on</strong>g>for</str<strong>on</strong>g> the changes. The [pH] cyt changes were coupled<br />

to different H + -ATPases in the two cultivars. The expressi<strong>on</strong> analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> OsHKT1, OsHKT2 and OsVHA<br />

showed variable and cell- specific inducti<strong>on</strong> in these cultivars under salt stress c<strong>on</strong>diti<strong>on</strong>. The important<br />

mechanism <str<strong>on</strong>g>for</str<strong>on</strong>g> salt tolerance in cv. Pokkali was to keep cytosolic Na + at a low level, by reducing Na + -influx<br />

(through down-regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> OsHKT1) and compartmentalizing cytosolic Na + into the vacuole (through the<br />

inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> vacuolar H + ATPase OsVHA, an energizer <str<strong>on</strong>g>for</str<strong>on</strong>g> the t<strong>on</strong>oplast Na + /H + antiporter). Pokkali might<br />

also induce increased uptake <str<strong>on</strong>g>of</str<strong>on</strong>g> K + through the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> OsHKT2, as evident in this study. Vacuolar<br />

compartmentalizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Na + is also present in salt-sensitive cv. BRRI Dhan29, but to a lesser extent and<br />

much later than in cv. Pokkali. The results suggest that the signaling and subsequent adaptive resp<strong>on</strong>ses in the<br />

salt-tolerant rice cv. Pokkali are different from that in the salt-sensitive cv. BRRI Dhan29.<br />

4C_06_P<br />

(poster secti<strong>on</strong> B2, poster board #281, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ALTERNATIVE ELECTRON DONORS SUBSTITUTE THE INACTIVATED<br />

OXYGEN-EVOLVING COMPLEX IN DIFFERENT PLANT SPECIES<br />

Szilvia Z. Tóth, Jos Thomas Puthur, Győző Garab<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Biology, Biological <str<strong>on</strong>g>Research</str<strong>on</strong>g> <str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>of</str<strong>on</strong>g> the Hungarian Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences, PO Box 521, H-6701<br />

Szeged, Hungary<br />

e-mail: Sztoth@brc.hu, jtputhur@rediffmail.com<br />

Electr<strong>on</strong> transport processes were investigated in different plant species in vivo under the c<strong>on</strong>diti<strong>on</strong>s in<br />

which the oxygen-evoluti<strong>on</strong> was fully inhibited by a heat pulse (48-51 °C, 40 s in water bath). This treatment<br />

induces no visible symptoms and the plants recover in 1-2 days. Under these c<strong>on</strong>diti<strong>on</strong>s, the K peak (~F 400 µs )<br />

appears in the fast chl a fluorescence (OJIP) transient reflecting partial Q A reducti<strong>on</strong>, which is due to a stable<br />

charge separati<strong>on</strong> resulting from the d<strong>on</strong>ati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> <strong>on</strong>e electr<strong>on</strong> by tyrozine Z. Additi<strong>on</strong>al fluorescence increase<br />

occurs in the 0.2-2 s time range <str<strong>on</strong>g>of</str<strong>on</strong>g> the fluorescence kinetics indicating additi<strong>on</strong>al Q A- accumulati<strong>on</strong>. Recently<br />

we have shown that this Q A- accumulati<strong>on</strong> was due to naturally occurring alternative electr<strong>on</strong> d<strong>on</strong>ors at the<br />

d<strong>on</strong>or side <str<strong>on</strong>g>of</str<strong>on</strong>g> photosystem II. The d<strong>on</strong>ati<strong>on</strong> probably originates from ascorbate, which provides electr<strong>on</strong>s<br />

with a halftime <str<strong>on</strong>g>of</str<strong>on</strong>g> ~30 ms in barley (Tóth et al. 2007, Biochim Biophys Acta 1767: 295-305). In this study, we<br />

investigated several other plant species, including Pisum sativum, Arabidopsis thaliana, Agropyr<strong>on</strong> el<strong>on</strong>gatum,<br />

Marchantia polymorpha, as well as the cyanobacterium Synechocystis PCC 6803 and the green alga Chlamydom<strong>on</strong>as<br />

reinhardtii. We established the functi<strong>on</strong>ing <str<strong>on</strong>g>of</str<strong>on</strong>g> the alternative electr<strong>on</strong> d<strong>on</strong>ors in all the investigated species.<br />

Preliminary data indicate that the inactivati<strong>on</strong> temperatures <str<strong>on</strong>g>of</str<strong>on</strong>g> the oxygen-evolving complex and the halftimes<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> electr<strong>on</strong> d<strong>on</strong>ati<strong>on</strong> may vary from species to species, and may also depend <strong>on</strong> the growth c<strong>on</strong>diti<strong>on</strong>s. We<br />

will discuss the physiological significance <str<strong>on</strong>g>of</str<strong>on</strong>g> this alternative electr<strong>on</strong> d<strong>on</strong>ati<strong>on</strong> to photosystem II in light and<br />

heat stress.<br />

207


23-26 August 2007,<br />

Budapest, Hungary<br />

4D. LONG-TERM ACCLIMATION OF PLANTS AND ECOSYSTEMS TO ELEVATED CO 2<br />

(ZOLTÁN TUBA, MICHAL V. MAREK)<br />

208<br />

4D_01_S<br />

WILL FOREST ECOSYSTEMS CONTINUE TO REMOVE CO2 FROM THE<br />

ATMOSPHERE AS CLIMATE CHANGES<br />

P.G. Jarvis*, S. Linder<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Edinburgh and Swedish University <str<strong>on</strong>g>of</str<strong>on</strong>g> Agricultural Sciences<br />

*e-mail: margaretsjarvis@aol.com<br />

In almost every <str<strong>on</strong>g>for</str<strong>on</strong>g>est we look today we find that there is a net removal <str<strong>on</strong>g>of</str<strong>on</strong>g> CO 2 from the atmosphere and<br />

carb<strong>on</strong> is accumulating in the soil and in the trees. A net gain <str<strong>on</strong>g>of</str<strong>on</strong>g> carb<strong>on</strong> by <str<strong>on</strong>g>for</str<strong>on</strong>g>est ecosystems implies a lack <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

balance between the processes removing CO 2 from the atmosphere and the processes returning CO 2 to the<br />

atmosphere. Ecosystem net primary producti<strong>on</strong> (NPP) is the difference between the gross photosynthetic producti<strong>on</strong><br />

(GPP) and the losses <str<strong>on</strong>g>of</str<strong>on</strong>g> carb<strong>on</strong> resulting from the autotrophic respirati<strong>on</strong> (R A ), i.e., NPP = GPP – R A . The net<br />

ecosystem producti<strong>on</strong> (NEP) is the difference between the net primary producti<strong>on</strong> (NPP) and the heterotrophic<br />

respirati<strong>on</strong> (R H ) associated with mineralisati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> organic matter in the soil, i.e., NEP = NPP - R H . For a <str<strong>on</strong>g>for</str<strong>on</strong>g>est<br />

at equilibrium, we would expect NEP to be zero (i.e., NPP = R H ). C<strong>on</strong>versely, when we measure NEP > 0,<br />

NPP must exceed R H . This world-wide disequilibrium in <str<strong>on</strong>g>for</str<strong>on</strong>g>ests today is the reas<strong>on</strong> why <str<strong>on</strong>g>for</str<strong>on</strong>g>ests are a large<br />

global carb<strong>on</strong> sink, removing from the atmosphere close to 40% <str<strong>on</strong>g>of</str<strong>on</strong>g> CO 2 emissi<strong>on</strong>s. It is comm<strong>on</strong>ly assumed,<br />

and has been shown by some models, that as atmospheric [C0 2 ] and surface temperature increase in the<br />

future, this current disequilibrium will reverse (i.e., RH > NPP). A key questi<strong>on</strong> is whether this is likely<br />

Whilst including a carb<strong>on</strong> cycle in GCMs and other models has been a major step <str<strong>on</strong>g>for</str<strong>on</strong>g>ward, a carb<strong>on</strong> cycle<br />

without an associated nitrogen cycle is unrealistic. Firstly, correlati<strong>on</strong> across sites shows that NPP is<br />

stimulated by the c<strong>on</strong>current depositi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> atmospheric N. Sec<strong>on</strong>dly soil warming experiments show that N<br />

released by decompositi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> litter and soil organic matter leads to increases in tree leaf area, uptake <str<strong>on</strong>g>of</str<strong>on</strong>g> CO 2<br />

and tree growth. Thirdly, after an initial enhancement <strong>on</strong> warming, R H settles down to the rate prior to the<br />

increase in temperature. For these reas<strong>on</strong>s, recent projecti<strong>on</strong>s by models linking the carb<strong>on</strong> and nitrogen<br />

cycles show NPP <str<strong>on</strong>g>of</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g>ests increasing over the next 100 years in parallel with R H , as atmospheric [CO 2 ] and<br />

surface temperature increase. Thus we may expect the <strong>on</strong>going removal <str<strong>on</strong>g>of</str<strong>on</strong>g> [CO 2 ] from the atmosphere by<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g>ests (NPP > RH) to be maintained.<br />

4D_02_S<br />

STRESS TO PLANTS AND THE ECOSYSTEM UNDER ELEVATED CO2<br />

Hartmut K. Lichtenthaler<br />

Botanical Institute, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Karlsruhe, Kaiserstr. 12, D-76133 Karlsruhe, Germany<br />

e-mail: hartmut.lichtenthaler@bio.uka.de<br />

The original stress c<strong>on</strong>cept <str<strong>on</strong>g>of</str<strong>on</strong>g> Hans Selye has also been extended to plants and is today well defined to<br />

describe the acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> unfavorable envir<strong>on</strong>mental c<strong>on</strong>straints and stressors <strong>on</strong> plants as well as the plants’<br />

resp<strong>on</strong>se via stress avoiding and stress coping mechanisms including special short-term acclimati<strong>on</strong>s and<br />

l<strong>on</strong>g-term adaptati<strong>on</strong>s. Several examples <str<strong>on</strong>g>of</str<strong>on</strong>g> stress coping mechanisms are presented. Elevated CO 2 is a new<br />

challenge <str<strong>on</strong>g>for</str<strong>on</strong>g> plants and the ecosystem. It may not be a real stress, but it is a c<strong>on</strong>tinuous strain. This requires<br />

particular acclimati<strong>on</strong> and adaptati<strong>on</strong> resp<strong>on</strong>ses some <str<strong>on</strong>g>of</str<strong>on</strong>g> which are known. In fact, the plants make use <str<strong>on</strong>g>of</str<strong>on</strong>g>


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

their general stress coping mechanisms either directly applied or in a modified <str<strong>on</strong>g>for</str<strong>on</strong>g>m to avoid or compensate<br />

possible negative effects <str<strong>on</strong>g>of</str<strong>on</strong>g> elevated CO 2 . One has to c<strong>on</strong>sider that <strong>on</strong> top <str<strong>on</strong>g>of</str<strong>on</strong>g> this all the classical abiotic,<br />

biotic and anthropogenic stressors are threatening plant growth and development also under elevated CO 2 .<br />

Thus, the knowledge <str<strong>on</strong>g>of</str<strong>on</strong>g> the general stress coping and stress avoiding resp<strong>on</strong>ses and tolerance mechanisms is<br />

needed to understand the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the plants’ metabolism. Emissi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> volatile isoprenoids, such as<br />

isoprene or methylbutenol or the accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> m<strong>on</strong>o- or diterpenes, are now understood as a possibility<br />

to regulate the internal cell metabolism and to protect the plants against photoinibiti<strong>on</strong> and photooxidati<strong>on</strong>.<br />

Measurements <str<strong>on</strong>g>of</str<strong>on</strong>g> chlorophyll fluorescence and particularly the imaging <str<strong>on</strong>g>of</str<strong>on</strong>g> chlorophyll fluorescence and the<br />

plants’ blue-green fluorescence provide excellent means to early detect and describe stress symptoms <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

plants and the ecosystem. Several examples are given.<br />

4D_03_S<br />

TWO-FACE CHARACTER OF THE ELEVATED CARBON DIOXIDE IMPACTS ON<br />

THE PLANT METABOLISM<br />

Michal V. Marek, Otmar Urban<br />

Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Plants ecological Physiology, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Systems Biology and Ecology, v.v.i. , Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

Czech Republic, Porici 3b, CZ-6030 00 Brno, Czech republic<br />

e-mail: emarek@usbe.cas.cz<br />

The rising atmospheric CO 2 c<strong>on</strong>centrati<strong>on</strong> (EC) is an important ecophysiological topic. Because <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

important CO 2 functi<strong>on</strong>s (effects <strong>on</strong> carboxylati<strong>on</strong>, RUBISCO activati<strong>on</strong>, stomatal resp<strong>on</strong>se) <strong>on</strong> assimilati<strong>on</strong>,<br />

it is possible to expect wide range <str<strong>on</strong>g>of</str<strong>on</strong>g> plants resp<strong>on</strong>ses to EC. Thus, positive stimulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> photosynthesis or<br />

the assimilati<strong>on</strong> depressi<strong>on</strong> - photosynthetic adjustment were observed and may be explained by (1) a<br />

decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> RuBPCO amount and/or activity, (2) diluti<strong>on</strong>/redistributi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> nitrogen and phosphorus mineral<br />

status <str<strong>on</strong>g>of</str<strong>on</strong>g> assimilatory apparatus, (3) starch, (4) decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> photosynthetically active pigments c<strong>on</strong>tent and<br />

diminuti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> light-harvesting complexes, and (5) differences in the new sinks-source status <str<strong>on</strong>g>of</str<strong>on</strong>g> the plant. All menti<strong>on</strong>ed<br />

resp<strong>on</strong>ses were observed <strong>on</strong> the example <str<strong>on</strong>g>of</str<strong>on</strong>g> a dense spruce stand exposed <str<strong>on</strong>g>for</str<strong>on</strong>g> the l<strong>on</strong>g-term affected EC. The<br />

pr<strong>on</strong>ounced vertical pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ile <str<strong>on</strong>g>of</str<strong>on</strong>g> the photosynthetically active radiati<strong>on</strong> led to the differentiati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

photosynthetic apparatus between the shaded (S) and sun needles (E). The prol<strong>on</strong>ged exposure to EC was<br />

resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> the apparent assimilatory activity stimulati<strong>on</strong> observed mainly in deeply S needles. In E<br />

needles some signals <strong>on</strong> a manifestati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the acclimati<strong>on</strong> depressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the assimilati<strong>on</strong> were found. The<br />

l<strong>on</strong>g-term effect <str<strong>on</strong>g>of</str<strong>on</strong>g> EC was resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> the decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> nitrogen c<strong>on</strong>tent. Moreover, the prol<strong>on</strong>ged<br />

exposure to EC did not cause any stimulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> electr<strong>on</strong> transport rate (ETR) <str<strong>on</strong>g>for</str<strong>on</strong>g> the E-needles but a str<strong>on</strong>gly<br />

positive effect <str<strong>on</strong>g>of</str<strong>on</strong>g> EC <strong>on</strong> ETR was observed <str<strong>on</strong>g>for</str<strong>on</strong>g> the S-needles. The analysis shows that the depressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

photosynthetic activity by l<strong>on</strong>g-term impact <str<strong>on</strong>g>of</str<strong>on</strong>g> EC is mainly caused by decreased RUBISCO carboxylati<strong>on</strong><br />

rate. All menti<strong>on</strong>ed modificati<strong>on</strong>s to photosynthetic assimilati<strong>on</strong> depend <strong>on</strong> time during the growing seas<strong>on</strong>.<br />

The str<strong>on</strong>g depressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> assimilati<strong>on</strong> observed in the autumn m<strong>on</strong>ths was the result <str<strong>on</strong>g>of</str<strong>on</strong>g> insufficient carb<strong>on</strong><br />

sink capacity.<br />

209


23-26 August 2007,<br />

Budapest, Hungary<br />

210<br />

4D_04_S<br />

GRASSLAND ECOLOGY UNDER ELEVATED AIR CO2 CONCENTRATIONS<br />

Zoltán Tuba 1,2 , Zoltán Nagy 2<br />

1Plant Ecological <str<strong>on</strong>g>Research</str<strong>on</strong>g> Group <str<strong>on</strong>g>of</str<strong>on</strong>g> Hungarian Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Botany and Plant Physiology, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Agricultural and Envir<strong>on</strong>mental Sciences, Szent István<br />

University, Gödöllő, H-2103 Gödöllő, Páter K. u. 1, Hungary,<br />

e-mail: tuba.zoltan@mkk.szie.hu<br />

Global climate change appears to be the greatest ecological problem <str<strong>on</strong>g>of</str<strong>on</strong>g> the future. For example the CO 2<br />

emitted into the atmosphere will have a l<strong>on</strong>g-term effect and this is <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the most influential ecological<br />

factor at global scale. Its increasing c<strong>on</strong>centrati<strong>on</strong> affects the plants directly, causing changes in their<br />

ecophysiological processes. C<strong>on</strong>sequently, the tolerance, reproducti<strong>on</strong>, distributi<strong>on</strong> and abundance <str<strong>on</strong>g>of</str<strong>on</strong>g> plant<br />

species will be altered. Species compositi<strong>on</strong> and diversity <str<strong>on</strong>g>of</str<strong>on</strong>g> plant communities, the vegetati<strong>on</strong> dynamics are<br />

all expected to be changed, too. C<strong>on</strong>sidering terrestrial ecosystems, grasslands are the sec<strong>on</strong>d largest<br />

vegetati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> after the <str<strong>on</strong>g>for</str<strong>on</strong>g>ests and their area is further increasing with the land use changes (clearance<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the <str<strong>on</strong>g>for</str<strong>on</strong>g>ests, urbanisati<strong>on</strong>, extensive agricultural practices and aband<strong>on</strong>ment <str<strong>on</strong>g>of</str<strong>on</strong>g> agricultural land). On a<br />

global scale grassllands cover 24% <str<strong>on</strong>g>of</str<strong>on</strong>g> the Earth's vegetated area. They occur over a broad range <str<strong>on</strong>g>of</str<strong>on</strong>g> climatic<br />

and soil c<strong>on</strong>diti<strong>on</strong>s and vary from intensively managed sown pastures to natural grassland communities.<br />

Despite their importance, the potential effects <str<strong>on</strong>g>of</str<strong>on</strong>g> climate change <strong>on</strong> grasslands have received much less<br />

attenti<strong>on</strong> than effects <strong>on</strong> other ecosystems such as <str<strong>on</strong>g>for</str<strong>on</strong>g>ests. The main objective <str<strong>on</strong>g>of</str<strong>on</strong>g> the proposed talk is to<br />

discuss the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> elevated air CO 2 c<strong>on</strong>cenrati<strong>on</strong>s <strong>on</strong> the structure (botanical, species compositi<strong>on</strong>, floral<br />

diversity, canopy and below ground/root architecture) and functi<strong>on</strong> (CO 2 ,and H 2 O exchange carb<strong>on</strong> cycling,<br />

dry matter producti<strong>on</strong> resp<strong>on</strong>ses, etc.) <str<strong>on</strong>g>of</str<strong>on</strong>g> grasslands and to quantify the carb<strong>on</strong> storage in grassland<br />

ecosystems.<br />

4D_05_S<br />

(poster secti<strong>on</strong> B2, poster board #282, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HOW DO PLANTS SURVIVE EXTREME CLIMATIC EVENTS<br />

Ivan Nijs<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Antwerp, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, Universiteitsplein 1, B-2610 Wilrijk, Belgium,<br />

e-mail: ivan.nijs@ua.ac.be<br />

Climatic extremes, such as heat waves and extreme droughts, are predicted to become more frequent and<br />

severe in a future atmosphere with more greenhouse gases. However, untill now nearly all experimental<br />

studies <strong>on</strong> the impact <str<strong>on</strong>g>of</str<strong>on</strong>g> climate warming <strong>on</strong> plants have c<strong>on</strong>centrated <strong>on</strong> changes in mean temperature, and<br />

studies <strong>on</strong> extremes are <strong>on</strong>ly beginning to emerge. Here I report <strong>on</strong> six recent field studies c<strong>on</strong>cerning the<br />

ecophysiological effects <str<strong>on</strong>g>of</str<strong>on</strong>g> heat waves, including three studies <strong>on</strong> temperate grassland and three <strong>on</strong> arctic<br />

tundra. In all experiments, the heat waves were generated with computer-c<strong>on</strong>trolled infrared irradiati<strong>on</strong> in the<br />

field. In some cases we allowed natural precipitati<strong>on</strong>, in others we excluded rainfall completely. We<br />

investigated the following questi<strong>on</strong>s: (1) is resistance to extremes in individuals plants coupled to specific<br />

plant traits (2) does resistance vary with the complexity <str<strong>on</strong>g>of</str<strong>on</strong>g> the community (3) do climatic extremes exert<br />

physiological after-effects l<strong>on</strong>g after the event has passed, e.g. <strong>on</strong> stomatal functi<strong>on</strong>ing, photosynthetic<br />

efficiency, chlorophyll synthesis/breakdown, etc. (4) does tundra react differently than temperate grasslands<br />

The main findings can be summarized as follows. In grassland species, morphological and ecophysiological


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

indicators that best explained plant resistance to extreme drought, were different from known indicators <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

resistance to moderate stress. Plant survival in complex grassland communities was different from survival in<br />

m<strong>on</strong>ocultures, which can be explained by differences in resource use (water). In arctic communities, heat<br />

waves alleviated stress during exposure, but stress symptoms aggravated after the heat waves had ended.<br />

Because plant resp<strong>on</strong>ses were species-specific in most communities, future shifts in species compositi<strong>on</strong><br />

when current regimes <str<strong>on</strong>g>of</str<strong>on</strong>g> extreme events change, can be anticipated.<br />

4D_06_S<br />

(poster secti<strong>on</strong> B2, poster board #283, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

AUTOMATIC PLANT LEAF TEMPERATURE MONITORING<br />

FOR STRESS DETECTION<br />

Laury Chaerle 1,* , Ilkka Lein<strong>on</strong>en 2,3 , Hamlyn G. J<strong>on</strong>es 2 , Dominique Van Der Straeten 1,* ,<br />

1Unit Plant Horm<strong>on</strong>e Signalling and Bio-imaging, Ghent University, K.L. Ledeganckstraat 35,<br />

B-9000 Gent, Belgium<br />

2Plant <str<strong>on</strong>g>Research</str<strong>on</strong>g> Unit, Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Envir<strong>on</strong>mental and Applied Biology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Dundee at SCRI, Invergowrie,<br />

Dundee DD2 5DA, Scotland, UK<br />

3Present address: School <str<strong>on</strong>g>of</str<strong>on</strong>g> Applied Sciences, Cranfield University, Bed<str<strong>on</strong>g>for</str<strong>on</strong>g>dshire MK43 0AL, UK<br />

e-mail: laury.chaerle@ugent.be, dominique.vanderstraeten@ugent.be<br />

Transpirati<strong>on</strong> and photosynthesis are the key parameters <str<strong>on</strong>g>of</str<strong>on</strong>g> plant development. Stress situati<strong>on</strong>s inevitably<br />

alter the kinetics <str<strong>on</strong>g>of</str<strong>on</strong>g> these key plant physiological processes. Measuring these parameters to compare and<br />

characterize agricultural per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance <str<strong>on</strong>g>of</str<strong>on</strong>g> plant cultivars is routinely d<strong>on</strong>e with portable gas analysis equipment,<br />

directly m<strong>on</strong>itoring water loss and CO 2 use. Since repeated measurements are needed, this procedure is<br />

tedious and c<strong>on</strong>sequently few leaves per plant can be measured. Thermal imaging reveals leaf temperature in<br />

a n<strong>on</strong>-c<strong>on</strong>tact way, and can be used effectively <str<strong>on</strong>g>for</str<strong>on</strong>g> m<strong>on</strong>itoring transpirati<strong>on</strong> as a functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> time. To achieve<br />

single-plant resoluti<strong>on</strong> in Arabidopsis, a plant growth-chamber based robotised imaging system was used. A<br />

number <str<strong>on</strong>g>of</str<strong>on</strong>g> Arabidopsis ecotypes selected <strong>on</strong> maximum genetic diversity was ranked <strong>on</strong> average rosette<br />

temperature. Due to both natural variati<strong>on</strong> in leaf temperature and the fact that plant leaf temperature<br />

changes during a day, rosettes cannot always be discerned from the background <str<strong>on</strong>g>of</str<strong>on</strong>g> well-watered substrate.<br />

There<str<strong>on</strong>g>for</str<strong>on</strong>g>e leaf area detecti<strong>on</strong> was based <strong>on</strong> co-located chlorophyll fluorescence images. In comparis<strong>on</strong> with<br />

direct transpirati<strong>on</strong> measurements, thermal imaging coupled to automated plant selecti<strong>on</strong> detected more<br />

significant temperature differences. This study proves the applicability <str<strong>on</strong>g>of</str<strong>on</strong>g> thermography under c<strong>on</strong>trolled<br />

c<strong>on</strong>diti<strong>on</strong>s to compare the transpirati<strong>on</strong>al resp<strong>on</strong>ses <str<strong>on</strong>g>of</str<strong>on</strong>g> different plant cultivars, with applicati<strong>on</strong>s in drought<br />

tolerance and disease resistance screening.<br />

211


23-26 August 2007,<br />

Budapest, Hungary<br />

4E. ABIOTIC STRESS<br />

4E_01_P<br />

(poster secti<strong>on</strong> B2, poster board #284, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STRUCTURE-FUNCTION STUDIES OF TOMATO ASR1, A WATER STRESS- AND<br />

SALT STRESS-REGULATED PLANT-SPECIFIC PROTEIN<br />

Dudy Bar-Zvi 1 , Zvia K<strong>on</strong>rad 1 , Slava Rom 1 , Dor<strong>on</strong> Shkolnik 1 , Natalia Shadrin 2 , Rodolfo Ghirlando 3 ,<br />

Yehuda Goldgur 2<br />

Departments <str<strong>on</strong>g>of</str<strong>on</strong>g> Life Sciences 1 , and Chemistry 2 , Ben-Guri<strong>on</strong> University, Beer-Sheva 84105, Israel<br />

3Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular Biology, Nati<strong>on</strong>al Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Diabetes and Digestive and Kidney Diseases,<br />

Nati<strong>on</strong>al Institutes <str<strong>on</strong>g>of</str<strong>on</strong>g> Health, USA<br />

e-mail: barzvi@bgu.ac.il<br />

ASR1 (abscisic acid stress ripening) is a low molecular weight plant specific protein encoded by an abioticstress<br />

regulated gene. The ASR1 protein possesses a zinc-dependent DNA-binding activity. The DNA<br />

binding site was suggested to reside in the central part <str<strong>on</strong>g>of</str<strong>on</strong>g> the polypeptide by using truncated <str<strong>on</strong>g>for</str<strong>on</strong>g>ms <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

protein. Two additi<strong>on</strong>al zinc-binding sites were shown to be localized at the amino terminus <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

polypeptide. ASR1 protein is presumed to be a natively unfolded protein using a number <str<strong>on</strong>g>of</str<strong>on</strong>g> predicti<strong>on</strong><br />

algorithms. The degree <str<strong>on</strong>g>of</str<strong>on</strong>g> order <str<strong>on</strong>g>of</str<strong>on</strong>g> ASR1 was determined experimentally using n<strong>on</strong>-tagged recombinant<br />

protein expressed in E. coli and purified to homogeneity. Purified ASR1 was shown to be unfolded using<br />

dynamic light scattering, gel filtrati<strong>on</strong>, microcalorimetry, circular dichroism and Fourier trans<str<strong>on</strong>g>for</str<strong>on</strong>g>m infrared<br />

(FTIR) spectrometry. The protein was shown to be m<strong>on</strong>omeric by analytical ultracentrifugati<strong>on</strong>. Additi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

zinc i<strong>on</strong>s resulted in a global change in the ASR1 structure, from m<strong>on</strong>omer to homodimer. Up<strong>on</strong> binding <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

zinc i<strong>on</strong>s, the protein becomes ordered as shown by FTIR and microcalorimetry, c<strong>on</strong>comitant with<br />

dimerizati<strong>on</strong>. Cytosolic ASR1 is unfolded whereas nuclear ASR1 is ordered. The effect <str<strong>on</strong>g>of</str<strong>on</strong>g> zinc binding <strong>on</strong><br />

ASR1 folding and dimerizati<strong>on</strong>, and activity <str<strong>on</strong>g>of</str<strong>on</strong>g> cytosolic and nuclear ASR1 protein in is discussed.<br />

4E_03_P<br />

(poster secti<strong>on</strong> B2, poster board #285, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

T22, A BIOCONTROL TRICHODERMA HARZIANUM STRAIN ENHANCED<br />

DROUGHT STRESS RESISTANCE OF PEACH ROOTSTOCK ‘PEMA’ (PRUNUS<br />

PERSICA X PRUNUS AMYGDALUS)<br />

P. Czövek 1 , I. Király 1 , I. Balla 2 , E. Szűcs 2<br />

1Eötvös University Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Physiology and Molecular Plant Biology<br />

H-1117, Budapest Pázmány sétány 1/C Hungary, e-mail: ikiraly@ludens.elte.hu<br />

2<str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Fruitgrowing and Ornamentals H-1223 Budapest, Park u. 2.<br />

Bioc<strong>on</strong>trol is an envir<strong>on</strong>mentally sound method used <str<strong>on</strong>g>for</str<strong>on</strong>g> the preventi<strong>on</strong> and c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> plant diseases.<br />

Advisedly selected fungal and bacterial strains are marketed worldwide promoting to spread <str<strong>on</strong>g>of</str<strong>on</strong>g> this<br />

inexpensive and effective technology. The effect <str<strong>on</strong>g>of</str<strong>on</strong>g> bioc<strong>on</strong>trol species is composed <str<strong>on</strong>g>of</str<strong>on</strong>g> (1) direct interacti<strong>on</strong><br />

with potential plant pathogens, (2) eliciting host-mediated defense reacti<strong>on</strong>s, (3) other beneficial effects as<br />

enhanced root growth. Trichoderma harzianum is a soil fungi living in the close vicinity <str<strong>on</strong>g>of</str<strong>on</strong>g> plant roots. As a<br />

hyper parasite it kills many pathogenic fungi generally by hydrolyzing mycelia walls. The T22 T. harzianum<br />

212


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

strain has also a str<strong>on</strong>g chitinase activity and many beneficial effects <strong>on</strong> the host plant, including enhanced<br />

root growth. We used the commercially available T22 strain to defend fruit trees against drought stress<br />

pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iting its effect <strong>on</strong> the plant defense reacti<strong>on</strong>s. The changes <str<strong>on</strong>g>of</str<strong>on</strong>g> the superoxide dismutase activity and the<br />

raise <str<strong>on</strong>g>of</str<strong>on</strong>g> mal<strong>on</strong>dialdehyde level as a c<strong>on</strong>sequence <str<strong>on</strong>g>of</str<strong>on</strong>g> far g<strong>on</strong>e lipid peroxidati<strong>on</strong> were m<strong>on</strong>itored during the<br />

stress treatment. The physiological state <str<strong>on</strong>g>of</str<strong>on</strong>g> the c<strong>on</strong>trol and stressed fruit trees was characterized also by the<br />

chlorophyll c<strong>on</strong>tent, the fresh weight-dry weight ratio and the stomatal c<strong>on</strong>ductancy. The results obtained<br />

clearly dem<strong>on</strong>strate that T22 treated fruit trees resisted well even the vigorous drought stress applied. The<br />

enhanced resistance correlated with the changing <str<strong>on</strong>g>of</str<strong>on</strong>g> the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the reactive oxygen scavenging system.<br />

This work was supported by GVOP-3.1.1-2004-05-0061/3.0.<br />

4E_04_P<br />

(poster secti<strong>on</strong> B2, poster board #286, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECTS OF POTASSIUM ON SALT TOLERANCE OF SEEDLING IRANIAN<br />

PISTACHIO<br />

Majid Fekri<br />

Vali-Asr University, Rafsanjan, Iran, e-mail: mjdfekri@yahoo.com<br />

The effects <str<strong>on</strong>g>of</str<strong>on</strong>g> two K levels (50 and 100 mg/kg K2SO4) and six treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> irrigati<strong>on</strong> with soluti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> NaCl<br />

(1, 2, 4, 8, 16 and 20 ds/m) <strong>on</strong> the growth, Chemical compositi<strong>on</strong> and plant water relati<strong>on</strong>ships seeding <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Iranian pistachio (Pistachio vera L .) were studied in an arid calcareous soil in a glasshouse experiment.<br />

Tolerant to salt was less in the low K level than high K level, probably because <str<strong>on</strong>g>of</str<strong>on</strong>g> higher uptake and /or<br />

transport <str<strong>on</strong>g>of</str<strong>on</strong>g> Cl and Na i<strong>on</strong>s, lesser osmoregulatory ability, and greater reducti<strong>on</strong>s in the top and root dry<br />

weights. The 50% reducti<strong>on</strong> the top and root dry weights were obtained, respectively, at an electrical<br />

c<strong>on</strong>ductivity <str<strong>on</strong>g>of</str<strong>on</strong>g> the saturati<strong>on</strong> extract (ECe) <str<strong>on</strong>g>of</str<strong>on</strong>g> 12 and 13 ds/m <str<strong>on</strong>g>for</str<strong>on</strong>g> low K level and 16 and 17.5 <str<strong>on</strong>g>for</str<strong>on</strong>g> high k<br />

level. Higher levels <str<strong>on</strong>g>of</str<strong>on</strong>g> K fertilizati<strong>on</strong> may be beneficial <str<strong>on</strong>g>for</str<strong>on</strong>g> Pistachio to tolerate to salt stress c<strong>on</strong>diti<strong>on</strong>s.<br />

4E_05_P<br />

(poster secti<strong>on</strong> B2, poster board #287, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ROLE OF PHOTOSYNTHETIC PERFORMANCE IN SALT STRESS ACCLIMATION<br />

OF TOMATO AFTER SALICYLIC ACID PRE-TREATMENT<br />

Katalin Gémes, Ágnes Szepesi, Adrienn Guóth, Irma Tari<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Physiology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Szeged, 6701 Szeged POB 654, Hungary<br />

e-mail: pigsy17@freemail.hu, tari@bio.u-szeged.hu<br />

Impositi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> salt stress reduced the net CO 2 assimilati<strong>on</strong> rate, chlorophyll (Chl) and carotenoid c<strong>on</strong>tents,<br />

stomatal c<strong>on</strong>ductance and biomass producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> tomato (Lycopersic<strong>on</strong> esculentum Mill. L. cv. Rio Fuego). Pretreatments<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> plants with 10 -4 M, but not with 10 -7 M salicylic acid (SA), could partially restore the CO 2<br />

fixati<strong>on</strong> rate, Chl a/b ratio, carotenoid levels under 100 mM NaCl exposure. Accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> soluble sugars,<br />

a biochemical marker <str<strong>on</strong>g>of</str<strong>on</strong>g> salinity tolerance <str<strong>on</strong>g>of</str<strong>on</strong>g> tomato, could be detected in pre-treated plants. After SA<br />

applicati<strong>on</strong>, c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> hexoses (glucose and fructose) remained high in the leaves and that <str<strong>on</strong>g>of</str<strong>on</strong>g> sucrose<br />

in the roots during salt stress. Both SA and salt stress increased the H 2 O 2 producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> tissues. Activities <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

superoxide dismutase and catalase (CAT), a H 2 O 2 generating and scavenging enzymes, respectively, decreased<br />

significantly under salt stress, but in 10 -4 M SA pre-treated plants, CAT activity was significantly induced both<br />

in the root and shoot tissues. The improved photosynthetic per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance, the accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> soluble sugars<br />

213


23-26 August 2007,<br />

Budapest, Hungary<br />

as compatible osmolytes and the effective eliminati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> H 2 O 2 by CAT c<strong>on</strong>tributed to the successful<br />

acclimati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 10 -4 M SA pre-treated tomato plants to high salinity.<br />

214<br />

4E_06_P<br />

(poster secti<strong>on</strong> B2, poster board #288, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

COMPARISON OF CHANGES IN PHOTOSYNTHESIS, CHLOROPHYLL<br />

FLUORESCENCE PARAMETERS AND ABSCISIC ACID LEVELS IN WHEAT<br />

CULTIVARS UNDER DROUGHT STRESS DURING GRAIN FILLING AND IN<br />

SEEDLINGS UNDER OSMOTIC STRESS<br />

A. Guóth 1 , I. Tari 1 , Á. Gallé 1 , J. Csiszár 1 , L. Cseuz 2 , L. Erdei 1<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Physiology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Szeged, 6701 Szeged, POB 654 Hungary<br />

2Cereal <str<strong>on</strong>g>Research</str<strong>on</strong>g> N<strong>on</strong>-Pr<str<strong>on</strong>g>of</str<strong>on</strong>g>it Company, 6701 Szeged, POB 391 Hungary<br />

e-mail: guotha@bio.u-szeged.hu<br />

We investigated the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> water deficit <strong>on</strong> the photosynthetic parameters and the abscisic acid (ABA)<br />

levels in wheat. A comparis<strong>on</strong> was made between the changes <str<strong>on</strong>g>of</str<strong>on</strong>g> the parameters menti<strong>on</strong>ed above, in seedling<br />

stage under osmotic stress and in reproductive growth phase under soil drought in two Hungarian wheat<br />

cultivars Triticum aestivum L. cv. MV Emese (resistant) and GK Élet (sensitive). The water status parameters,<br />

CO 2 assimilati<strong>on</strong>, chlorophyll a (chla) fluorescence, pigment c<strong>on</strong>tent and ABA levels were determined as a<br />

functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the development <str<strong>on</strong>g>of</str<strong>on</strong>g> water deficit. The tolerant genotype cv. Emese maintained higher water<br />

potential in upper leaves under osmotic or drought stress than the sensitive cv. Élet. In spite <str<strong>on</strong>g>of</str<strong>on</strong>g> this, the<br />

tolerant line exhibited an earlier ABA accumulati<strong>on</strong> in the grains (DPA 9) than the sensitive line (DPA 24).<br />

The characteristics <str<strong>on</strong>g>of</str<strong>on</strong>g> the CO 2 assimilati<strong>on</strong> and chla fluorescence parameters measured in flag leaf (Yield, qP,<br />

NPQ) did not differ between the two varieties and compared to the c<strong>on</strong>trol, the Yield changed similarly to<br />

the CO 2 fixati<strong>on</strong>. But in seedling stage under osmotic stress the CO 2 assimilati<strong>on</strong> declined significantly and in<br />

c<strong>on</strong>trast to the flag leaf the qP decreased and the NPQ increased significantly in both varieties, and the<br />

tendencies were the same in both genotypes. The chla fluorescence parameters and the efficiency <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

photosynthesis measured <strong>on</strong> the seedlings did not corresp<strong>on</strong>d to those measured <strong>on</strong> the flag leaf in the<br />

reproductive growth phase.-This work was supported by Grant No. NKFP 4/064/2004<br />

4E_07_P<br />

(poster secti<strong>on</strong> B2, poster board #289, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EVALUATION OF ABIOTIC STRESS TOLERANCE OF NICOTIANA TABACUM<br />

PLANTS BEARING AN ANTISENSE SUPPRESSOR OF THE PROLINE<br />

DEHYDROGENASE GENE<br />

A. V. Kochetov, Ya. S. Kolodyazhnaya, S. E. Titov, M. L. Komarova, A. V. Romanova, V. S. Koval,<br />

E. A. Trif<strong>on</strong>ova, V. K. Shumny<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Cytology and Genetics, Novosibirsk 630090, Russia; e-mail: ak@bi<strong>on</strong>et.nsc.ru<br />

Investigati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proline metabolism is important both to understand stress resp<strong>on</strong>se mechanisms and to<br />

improve crops. Earlier evaluati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> stress resistance <str<strong>on</strong>g>of</str<strong>on</strong>g> transgenic arabidopsis plants bearing antisense<br />

suppressor <str<strong>on</strong>g>of</str<strong>on</strong>g> proline dehydrogenase resulted in some c<strong>on</strong>troversial observati<strong>on</strong>s (Nanjo et al., 1999; Mani et<br />

al., 2002; Ribarits, 2007). We obtained tobacco plants bearing a fragment <str<strong>on</strong>g>of</str<strong>on</strong>g> arabidopsis proline


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

dehydrogenase gene in antisense orientati<strong>on</strong>. The usage <str<strong>on</strong>g>of</str<strong>on</strong>g> heterologous antisense suppressor resulted in a<br />

mild increase in proline c<strong>on</strong>tent in n<strong>on</strong>-stressed plants (by a factor <str<strong>on</strong>g>of</str<strong>on</strong>g> 2 to 3 in different lines). Under the<br />

stress c<strong>on</strong>diti<strong>on</strong>s (200 mM <str<strong>on</strong>g>of</str<strong>on</strong>g> NaCl) the c<strong>on</strong>tent <str<strong>on</strong>g>of</str<strong>on</strong>g> proline was increased to similar levels in both transgenic<br />

and c<strong>on</strong>trol plants. According to the results obtained transgenic plants were characterized by a mild increase<br />

in general (n<strong>on</strong>-specific) stress tolerance: (1) they survive better in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> NaCl or PEG6000 in<br />

growth media and c<strong>on</strong>tain a smaller quantity <str<strong>on</strong>g>of</str<strong>on</strong>g> MDA; (2) they tolerate higher c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> various<br />

heavy metals (Pb, Cd, Ni, Hg); (3) the detached leaves <str<strong>on</strong>g>of</str<strong>on</strong>g> transgenic plants lose water at a lower rate; (4) the<br />

electrolyte leakage experiments showed lower membrane permeability <str<strong>on</strong>g>of</str<strong>on</strong>g> cells <str<strong>on</strong>g>of</str<strong>on</strong>g> transgenic plants; (5) after<br />

the expositi<strong>on</strong> at high temperature, seeds <str<strong>on</strong>g>of</str<strong>on</strong>g> transgenic plants were characterized by a higher germinati<strong>on</strong> rate.<br />

It may be assumed that an increased level <str<strong>on</strong>g>of</str<strong>on</strong>g> proline under normal c<strong>on</strong>diti<strong>on</strong>s could partially alleviate the<br />

damage at stress <strong>on</strong>set and facilitate rapid inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> stress resp<strong>on</strong>se mechanisms. Transgenic plants were<br />

characterized by normal phenotype and development.<br />

4E_08_P<br />

(poster secti<strong>on</strong> B2, poster board #290, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

RESOURCE SHARING AMONG INTERCONNECTED RAMETS ENHANCES<br />

STRESS TOLERANCE IN IRIS PUMILA<br />

Sanja Manitašević 1 , Ana Vuleta*, Gordana Matić, Jadranka Dunđerski, Branka Tucić*<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry and *Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Evoluti<strong>on</strong>ary Biology, Institute <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

Biological <str<strong>on</strong>g>Research</str<strong>on</strong>g> „Siniša Stanković“, 11060 Belgrade, Serbia, 1 e-mail: manitas@ibiss.bg.ac.yu<br />

Physically c<strong>on</strong>nected ramets <str<strong>on</strong>g>of</str<strong>on</strong>g> cl<strong>on</strong>al plants are able to reciprocally exchange essential resources (i.e., water,<br />

assimilates and nutrients) if they are unevenly distributed over space. We tested the hypothesis that<br />

translocati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> resources between source and sink ramets increases stress tolerance <str<strong>on</strong>g>of</str<strong>on</strong>g> the whole cl<strong>on</strong>e<br />

growing in a patchy envir<strong>on</strong>ment. This study was c<strong>on</strong>ducted in an exposed populati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Iris pumila growing<br />

in the wild. In early spring, circle-shaped cl<strong>on</strong>al genotypes were cut into two equal parts with different<br />

integrati<strong>on</strong> status: C - with physically c<strong>on</strong>nected rhizome segments and NC - with ceased c<strong>on</strong>necti<strong>on</strong>s. One<br />

half <str<strong>on</strong>g>of</str<strong>on</strong>g> each cl<strong>on</strong>e was shaded with a neutral screen in such a way that both C and NC parts c<strong>on</strong>sisted <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

exposed (high-light stressed) and shaded (up to 50% <str<strong>on</strong>g>of</str<strong>on</strong>g> ambient irradiance) quarters. During the summer, the<br />

leaf tissues were sampled from each quarter within every cl<strong>on</strong>e. Variati<strong>on</strong> pattern <str<strong>on</strong>g>of</str<strong>on</strong>g> the following traits was<br />

examined: the level <str<strong>on</strong>g>of</str<strong>on</strong>g> stress proteins Hsp70 and Hsp90 (Hsp90a and Hsp90b is<str<strong>on</strong>g>of</str<strong>on</strong>g>orms), as well as relative<br />

water c<strong>on</strong>tent (RWC) and specific leaf area (SLA). While the value <str<strong>on</strong>g>of</str<strong>on</strong>g> SLA decreased with irradiance, Hsp90a<br />

expressi<strong>on</strong> and RWC increased, regardless <str<strong>on</strong>g>of</str<strong>on</strong>g> their integrati<strong>on</strong> status. C<strong>on</strong>versely, the amount <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 and<br />

Hsp90b dramatically elevated exclusively in the exposed quarter interc<strong>on</strong>nected with shaded <strong>on</strong>e. These<br />

results suggest that resource-sharing am<strong>on</strong>g interc<strong>on</strong>nected ramets could provide a powerful strategy in<br />

I. pumila to enhance the tolerance to stressful abiotic heterogeneity comm<strong>on</strong>ly occurring within its natural<br />

habitats.<br />

215


23-26 August 2007,<br />

Budapest, Hungary<br />

216<br />

4E_09_P<br />

(poster secti<strong>on</strong> B2, poster board #291, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ACCLIMATION AND TOLERANCE TO HEAT AND HIGH ILLUMINATION IN<br />

BRASSICA<br />

María José Quiles, Milagros Díaz, Virginia de Haro, Romualdo Muñoz<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Biology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Murcia, 30100 Espinardo, Murcia, Spain<br />

Two species <str<strong>on</strong>g>of</str<strong>on</strong>g> Brassica were used to study their acclimati<strong>on</strong> to heat and high illuminati<strong>on</strong> during the first<br />

stages <str<strong>on</strong>g>of</str<strong>on</strong>g> the development. One, B. fruticulosa, is a wild species from south-east Spain and is adapted to both<br />

heat and high light intensity in its natural habitat, while the other, B. oleracea, is an agricultural species that is<br />

widely cultivated throughout the wold. The B. fruticulosa plants grown under high illuminati<strong>on</strong> and heat<br />

showed a greater reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the foliar area and biomass than the B. oleracea plants in relati<strong>on</strong> with the<br />

respective c<strong>on</strong>trol plants,. The quantum yield <str<strong>on</strong>g>of</str<strong>on</strong>g> the PS II, the capacity <str<strong>on</strong>g>of</str<strong>on</strong>g> photosynthetic electr<strong>on</strong> transport<br />

and photochemical quenching, decreased in B. oleracea plants when grown under stress c<strong>on</strong>diti<strong>on</strong>s, indicating<br />

the inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> PS II. However, in B. fruticulosa the values <str<strong>on</strong>g>of</str<strong>on</strong>g> these parameters were similar to c<strong>on</strong>trol plants.<br />

PS I was more stable than PS II, probably because it plays a protective role though cyclic electr<strong>on</strong> flow. The<br />

PS I activity increased in B. oleracea and B. fruticulosa plants exposed to heat and high illuminati<strong>on</strong>. The<br />

activities <str<strong>on</strong>g>of</str<strong>on</strong>g> plastidial NADH deshydrogenase complex and terminal oxidase were much higher in B. fruticulosa<br />

than in B. oleracea, and both were stimulated in plants grown in stress c<strong>on</strong>diti<strong>on</strong>s. Acclimati<strong>on</strong> and tolerance to<br />

high illuminati<strong>on</strong> and heat <str<strong>on</strong>g>of</str<strong>on</strong>g> the photosystem activities was higher in the wild species than in the agricultural<br />

species, indicating that plant adaptati<strong>on</strong> to these stresses in natural c<strong>on</strong>diti<strong>on</strong>s favours subsequent<br />

acclimati<strong>on</strong>, and that, the chlororespirati<strong>on</strong> process is probably involved in both adaptati<strong>on</strong> and acclimati<strong>on</strong><br />

to heat and high illuminati<strong>on</strong>.<br />

4E_10_P<br />

(poster secti<strong>on</strong> B2, poster board #292, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECT OF SALINITY AND SOIL NITROGEN APPLICATION ON BIOCHEMICAL<br />

INDICES OF PISTACHIO<br />

A. Razavi Nasab, A. Tajabadi Pour, H. Shirani<br />

Soil Science Department, Agriculture College, Vali-Asr University, Rafsanjan, Iran,<br />

e-mail: azamrazavinasab@yahoo.com<br />

Soil salinity is an important growth limiting factor <str<strong>on</strong>g>for</str<strong>on</strong>g> most plants. Salinity inhibits plant growth by osmotic<br />

stress, nutriti<strong>on</strong>al imbalance and some specific toxicity and it is being progressively exacerbated by<br />

fertilizati<strong>on</strong>, especially in arid regi<strong>on</strong>s. In other hands, nitrogen (N) is an essential element <str<strong>on</strong>g>for</str<strong>on</strong>g> plants. It has a<br />

fundamental role in amino acids, proteins, nucleoic acids and lots <str<strong>on</strong>g>of</str<strong>on</strong>g> enzyme structure. The proper use <str<strong>on</strong>g>of</str<strong>on</strong>g> N<br />

fertilizers in all soils is important, but particularly so in saline soils, where N might reduce the adverse effects<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> salinity. In high salinity levels, the antag<strong>on</strong>istic relati<strong>on</strong>ships are attributed to the negative membrance<br />

potential <str<strong>on</strong>g>of</str<strong>on</strong>g> root cells and the negative charge <str<strong>on</strong>g>of</str<strong>on</strong>g> both nitrate and chloride. Most plants resp<strong>on</strong>d to salinity<br />

stress in their envir<strong>on</strong>ment by osmoregulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> their cellular c<strong>on</strong>tent. Synthesis and accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> organic<br />

solute <str<strong>on</strong>g>of</str<strong>on</strong>g> carbohydrate and N compounds (e.g. proline) are used <str<strong>on</strong>g>for</str<strong>on</strong>g> this purpose. The present study,<br />

there<str<strong>on</strong>g>for</str<strong>on</strong>g>e, was initiated to evaluate the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> N and salinity <strong>on</strong> proline and reducing sugars c<strong>on</strong>tent in<br />

pistachio. Pistachio seedlings were grown in potted soils exposed to four salinity levels (0, 800, 1600, and<br />

2400 mg NaCl kg -1 soil) and four N levels (0, 60, 120, and 180 mg kg -1 soil as urea). As the salinity levels<br />

increased, the leaf proline c<strong>on</strong>tent significantly increased while c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> reducing sugars decreased. A


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

significant increase in proline and reducing sugars was observed with N applicati<strong>on</strong>. The evidence presented<br />

in this study suggests that a functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proline under salt stress is that <str<strong>on</strong>g>of</str<strong>on</strong>g> osmoregulati<strong>on</strong>. Also, the results<br />

show that the provisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the optimal nutriti<strong>on</strong>al regimes, there<str<strong>on</strong>g>for</str<strong>on</strong>g>e, has an important c<strong>on</strong>tributi<strong>on</strong> to the<br />

determinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> pistachio reacti<strong>on</strong> to soil salinity.<br />

4E_11_P<br />

(poster secti<strong>on</strong> B2, poster board #293, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECT OF SALINITY AND SOIL ZINC AND PHOSPHOROUS APPLICATION ON<br />

BIOCHEMICAL INDICES OF PISTACHIO<br />

R. Shahriaripour, A. Tajabadipour, V. Mozaffari<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Soil Science, College <str<strong>on</strong>g>of</str<strong>on</strong>g> Agriculture, Rafsanjan University, Rafsanjan, Iran,<br />

e-mail: Roghayeh_shahriari2371@yahoo.com<br />

Salinity is a world-wide problem which adversely affects the growth <str<strong>on</strong>g>of</str<strong>on</strong>g> many plants through a series <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

interacting factors including some specific i<strong>on</strong> toxicities, i<strong>on</strong> imbalance and suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> water potential.<br />

Most plants resp<strong>on</strong>d to changing osmotic potentials in their external envir<strong>on</strong>ment by osmoregulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> their<br />

cellular c<strong>on</strong>tent. Synthesis and accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> organic solutes (e.g proline) are utilized <str<strong>on</strong>g>for</str<strong>on</strong>g> this purpose. The<br />

amount <str<strong>on</strong>g>of</str<strong>on</strong>g> proline accumulati<strong>on</strong> correlates well with the degree <str<strong>on</strong>g>of</str<strong>on</strong>g> stress in plants subjected to different salts.<br />

Soil fertilizati<strong>on</strong> appears to be beneficial in reducing the plant growth depressi<strong>on</strong> due to soluble salts. The<br />

present study, there<str<strong>on</strong>g>for</str<strong>on</strong>g>e, was initiated to evaluate the main and interactive effects <str<strong>on</strong>g>of</str<strong>on</strong>g> zinc, phosphorous and<br />

salinity levels <strong>on</strong> proline and reducing sugars c<strong>on</strong>tent in pistachio. Pistachio plants were grown in potted soils<br />

exposed to four salinity levels (0, 1000, 2000, 3000 mg NaCl Kg -1 soil), four phosphorous levels (0, 60, 120 and 180 mg P<br />

Kg -1 soil as Ca(H 2 PO 4 ) 2 ) and three zinc (Zn) levels (0, 5, 10 mg Zn Kg -1 soil as ZnSo 4 .7H 2 O). As the salinity levels increased,<br />

the leaf proline c<strong>on</strong>tent significantly increased. However, Zn applicati<strong>on</strong> declined the proline c<strong>on</strong>tent but the<br />

applicati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> phosphorous had not a significant effect <strong>on</strong> proline and reducing sugars c<strong>on</strong>tent. The reducing<br />

sugars c<strong>on</strong>centrati<strong>on</strong> was affected by salinity levels and Zn applicati<strong>on</strong>. A significant decrease in reducing<br />

sugars was observed with increasing salinity levels and decreasing Zn levels. The evidence presented in this<br />

study suggests that a functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proline in pistachio under salt stress is that <str<strong>on</strong>g>of</str<strong>on</strong>g> osmoregulati<strong>on</strong>. It seems that<br />

proline is not directly involved in osmoregulati<strong>on</strong>, but it plays an indirect role by increasing hydrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

protoplasm. The results <str<strong>on</strong>g>of</str<strong>on</strong>g> this study show that the provisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the optimal nutriti<strong>on</strong>al regimes, there<str<strong>on</strong>g>for</str<strong>on</strong>g>e,<br />

has an important c<strong>on</strong>tributi<strong>on</strong> to the determinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> pistachio reacti<strong>on</strong> to soil salinity.<br />

4E_12_P<br />

(poster secti<strong>on</strong> B2, poster board #294, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

INFLUENCE OF A NOVEL MITOCHONDRIAL PPR DOMAIN PROTEIN ON<br />

ABIOTIC STRESS RESPONSES OF ARABIDOPSIS THALIANA<br />

Laura Zsigm<strong>on</strong>d, Gyöngyi Székely, Csaba K<strong>on</strong>cz, László Szabados<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Biology, Biological <str<strong>on</strong>g>Research</str<strong>on</strong>g> Center, 6726-Szeged, Temesvári krt. 62, Hungary,<br />

e-mail: szabados@brc.hu<br />

Abiotic stress can activate the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> numerous genes through several abscisic acid (ABA) dependent<br />

and independent pathways. ABA plays important roles in dormancy, dessicati<strong>on</strong> c<strong>on</strong>trol, and osmotic stress<br />

resp<strong>on</strong>ses. We have identified a novel T-DNA tagged Arabidopsis mutant, which is characterized by<br />

hypersensitivity to high salinity, abscisic acid (ABA) and sugar treatments. The mutant was further<br />

217


23-26 August 2007,<br />

Budapest, Hungary<br />

characterized by enhanced proline accumulati<strong>on</strong>, increased tolerance to methylglyoxalate, and altered<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> numerous stress-resp<strong>on</strong>sive and other genes. The T-DNA inserti<strong>on</strong> disrupted the gene<br />

At3g16890, encoding an unknown mitoch<strong>on</strong>drial protein, composed <str<strong>on</strong>g>of</str<strong>on</strong>g> 14 pentatricopeptide (PPR) repeats.<br />

Overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the full length cDNA in the tagged mutant complemented the stress hypersensitivity and<br />

lead to increased salt tolerance in several transgenic lines. The encoded protein was localized in the<br />

mitoch<strong>on</strong>dria and was associated with complex III. in the mitoch<strong>on</strong>drial electr<strong>on</strong> transport system.<br />

Characterizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the mutant and the corresp<strong>on</strong>ding gene suggest a possible link between mitoch<strong>on</strong>drial<br />

electr<strong>on</strong> transport, stress resp<strong>on</strong>ses and ABA regulati<strong>on</strong> in higher plants.<br />

4E_13_P<br />

(poster secti<strong>on</strong> B2, poster board #295, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECT OF S-METHYLMETHIONINE ON MAIZE EXPOSED TO COLD STRESS<br />

CONDITIONS<br />

D. Szegő 1 , T. Visnovitz 1 , E. Páldi 2 , D. Lásztity 1 , Z. Szigeti 1 , I. Rácz 1<br />

1 Dept. Plant Physiol. Mol. Plant Biol., Eötvös L. Univ., Pázmány P. l. 1/C, H-1117 Budapest<br />

2 Agric. Res. Inst. Acad. Sci. Hung., Brunszvik s. 2., H-2462 Mart<strong>on</strong>vásár<br />

e-mail: raczil@ludens.elte.hu<br />

To improve stress- and disease-tolerance <str<strong>on</strong>g>of</str<strong>on</strong>g> cultivated plants and thus increase crop yield is a crucial problem<br />

in the agriculture. This problem is usually solved either by classical breeding or producing transgenic plants.<br />

Utilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> biologically active substances, possessing an effect <strong>on</strong> metabolism and thus ensuring the<br />

favourable property, provides alternative possibility. The natural intermediate compound S-methylmethi<strong>on</strong>ine<br />

(SMM) also bel<strong>on</strong>gs to biologically active substances. SMM has an important role in the sulphur metabolism,<br />

participating in methylati<strong>on</strong> reacti<strong>on</strong>s, in the biosynthesis <str<strong>on</strong>g>of</str<strong>on</strong>g> osmoprotectant sulphopropi<strong>on</strong>ates and through<br />

this, in stress- and disease-tolerance <str<strong>on</strong>g>of</str<strong>on</strong>g> plants. Physiological effect <str<strong>on</strong>g>of</str<strong>on</strong>g> SMM is similar to that <str<strong>on</strong>g>of</str<strong>on</strong>g> polyamines in<br />

some respects, which is probably due to the fact that SMM can take part in reacti<strong>on</strong> pathways leading to the<br />

biosynthesis <str<strong>on</strong>g>of</str<strong>on</strong>g> polyamines. In this work we studied the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> exogenously added SMM in maize (Zea mays<br />

L., cultivar Norma) under cold stress c<strong>on</strong>diti<strong>on</strong>s. Time course <str<strong>on</strong>g>of</str<strong>on</strong>g> changes in the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> polyamines<br />

(agmatine, putrescine, spermidine and spermine) were followed under cold, SMM and the combined<br />

treatments. To understand the molecular background <str<strong>on</strong>g>of</str<strong>on</strong>g> changes, expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> arginine decarboxylase,<br />

spermidine synthase and SAM-decarboxylase genes, which take part in the polyamine synthesis, were also<br />

investigated. Changes in expressi<strong>on</strong> levels <str<strong>on</strong>g>of</str<strong>on</strong>g> CBF1 transcripti<strong>on</strong> factor, which causes switching <strong>on</strong> several<br />

genes under cold stress c<strong>on</strong>diti<strong>on</strong>s, were also measured, so that we could get broader knowledge <str<strong>on</strong>g>of</str<strong>on</strong>g> stress<br />

avoiding role <str<strong>on</strong>g>of</str<strong>on</strong>g> the SMM.<br />

218


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

4E_14_P<br />

(poster secti<strong>on</strong> B2, poster board #296, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE EFFECT OF SOIL ZINC APPLICATION ON LIPID PEROXIDATION OF CELL<br />

MEMBRANE, PHENOLIC COMPOUNDS AND FLAVONOIDS IN PISTACHIO<br />

(PISTACIA VERA CV. GHAZVINI) UNDER SALINITY STRESS<br />

M. Talebi, V. Mozaffari, A. Tajabadipour<br />

Soil Sciences Department, Vali-Asr University, Rafsanjan, Iran, e-mail: msctalebi@yahoo.com<br />

Salt stress is c<strong>on</strong>sidered as <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the most important abiotic factors limiting plant growth and yield in many<br />

areas <str<strong>on</strong>g>of</str<strong>on</strong>g> the world. One <str<strong>on</strong>g>of</str<strong>on</strong>g> the main criteria <str<strong>on</strong>g>for</str<strong>on</strong>g> salt tolerance is cell membrane stability under stress. Reactive<br />

oxygen species (ROS) and lipid peroxidati<strong>on</strong> are c<strong>on</strong>sidered to be destructive to cell membrane under salt<br />

stress. Phenolic compounds and flav<strong>on</strong>oids in plants decrease negative effects <str<strong>on</strong>g>of</str<strong>on</strong>g> reactive oxygen species and<br />

increase stability <str<strong>on</strong>g>of</str<strong>on</strong>g> cell membrane. The objective <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to evaluate the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> zinc and salinity<br />

<strong>on</strong> some physiological and biochemical properties <str<strong>on</strong>g>of</str<strong>on</strong>g> pistachio (cv. Ghazvini). A factorial greenhouse<br />

experiment was carried out as completely randomized design with three replicati<strong>on</strong>s. Treatments were 4 levels<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> Zn (0, 5, 10 and 15 mg kg -1 soil) and 5 levels <str<strong>on</strong>g>of</str<strong>on</strong>g> salinity (0, 800, 1600, 2400 and 3200 mg NaCl kg -1 soil).<br />

The results indicated that by increasing salinity, lipid peroxidati<strong>on</strong> in leaves increased and levels <str<strong>on</strong>g>of</str<strong>on</strong>g> MDA (as<br />

an indicator <str<strong>on</strong>g>of</str<strong>on</strong>g> lipid peroxidati<strong>on</strong>) increased significantly in resp<strong>on</strong>se to NaCl. MDA c<strong>on</strong>centrati<strong>on</strong>s increased<br />

with soil salinity levels up to 1600 mg NaCl kg -1 soil, but decreased at higher salinity. Simultaneously,<br />

c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> other aldehyde increased with decreasing MDA. Applicati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 10 mg kg -1 zinc decreased<br />

over 33% MDA c<strong>on</strong>centrati<strong>on</strong> in leaves. Also, phenolic c<strong>on</strong>tent and flav<strong>on</strong>oids increased with increasing<br />

salinity. Applicati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 15 mg kg -1 <str<strong>on</strong>g>of</str<strong>on</strong>g> Zn significantly increased these compounds in comparis<strong>on</strong> to c<strong>on</strong>trol.<br />

In other words applicati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Zn increased phenolic compounds by 24% and flav<strong>on</strong>oids by 28%. The results<br />

presented in this paper suggest that zinc can increase the growth rate <str<strong>on</strong>g>of</str<strong>on</strong>g> salinity stress in pistachio in increase<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> phenolic compounds and flav<strong>on</strong>oids as an antistress compounds.<br />

4E_16_P<br />

(poster secti<strong>on</strong> B2, poster board #297, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PROTEIN MARKERS FOR REGENERATION CAPACITY FROM SALT STRESSED<br />

EMBRYOGENIC SUSPENSION CULTURES OF DACTYLIS GLOMERATA L<br />

Lyuben Zagorchev, Ivanka Tzacheva, Mariela Odjakova*<br />

Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, S<str<strong>on</strong>g>of</str<strong>on</strong>g>ia University<br />

8 Dragan Tzankov Blvd., S<str<strong>on</strong>g>of</str<strong>on</strong>g>ia, Bulgaria<br />

*e-mail: modjakova@bi<str<strong>on</strong>g>of</str<strong>on</strong>g>ac.uni-s<str<strong>on</strong>g>of</str<strong>on</strong>g>ia.bg<br />

Embryogenic suspensi<strong>on</strong> cultures <str<strong>on</strong>g>of</str<strong>on</strong>g> Dactylis glomerata were treated with different salt c<strong>on</strong>centrati<strong>on</strong>s - 85<br />

mM, 170 mM and 255 mM NaCl. While 85 mM NaCl enhances the level <str<strong>on</strong>g>of</str<strong>on</strong>g> somatic embryogenesis, higher<br />

salt c<strong>on</strong>centrati<strong>on</strong>s affect negatively the regenerati<strong>on</strong> potential. The protein patterns <str<strong>on</strong>g>of</str<strong>on</strong>g> cell wall-associated<br />

and culture media fracti<strong>on</strong>s showed significant alterati<strong>on</strong>s. Proteins, abundant in the cell wall at 85 mM NaCl<br />

and absent at higher salt c<strong>on</strong>centrati<strong>on</strong>s are potential markers <str<strong>on</strong>g>for</str<strong>on</strong>g> the regenerati<strong>on</strong> capacity. Many <str<strong>on</strong>g>of</str<strong>on</strong>g> these<br />

protein fracti<strong>on</strong>s appear in the culture media at 170 mM NaCl. A protein, with pI 9.2-9.4 and molecular<br />

weight 73 kD was chosen, as an antigen <str<strong>on</strong>g>for</str<strong>on</strong>g> screening the human semisynthetic single-chain Fv (scFv) phage<br />

library (Griffin.1) by 2D phage panning <str<strong>on</strong>g>for</str<strong>on</strong>g> selecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> specific m<strong>on</strong>ocl<strong>on</strong>al antibodies. Eight phage cl<strong>on</strong>es<br />

were selected in four rounds <str<strong>on</strong>g>of</str<strong>on</strong>g> screening. All scFvs were positive <strong>on</strong> Western blot analysis. Selected<br />

219


23-26 August 2007,<br />

Budapest, Hungary<br />

recombinant antibodies were obtained in soluble <str<strong>on</strong>g>for</str<strong>on</strong>g>m by expressi<strong>on</strong> in Escherichia coli HB2151. Western<br />

blot analysis against a 2D PAGE sample <str<strong>on</strong>g>of</str<strong>on</strong>g> extracellular proteins from cultures, treated with 170 mM NaCl,<br />

showed two fracti<strong>on</strong>s: the 73 kD protein, pI 9.2, used as antigen, plus a 33 kD, pI 8.3 protein. The 73 kD<br />

protein is found in the cell wall at 85 mM NaCl and absent at 170 mM. To our opini<strong>on</strong>, the selected<br />

antibodies can be used as potential markers <str<strong>on</strong>g>for</str<strong>on</strong>g> the regenerati<strong>on</strong> capacity <str<strong>on</strong>g>of</str<strong>on</strong>g> Dactylis glomerata cell cultures.<br />

4E_17_P<br />

(poster secti<strong>on</strong> B2, poster board #298, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

MEDICAGO TRUNCATULA PLANTS EXPRESSING THE ELIP-LIKE DSP22<br />

SHOWED IMPROVED CAPACITY TO RECOVER FROM A SEVERE WATER<br />

DEFICIT<br />

S. S. Araújo 1a) , A. S. Duque 1 , J. Marques da Silva 2,3 , D. Santos 1 , A. Bernardes da Silva 2,3 ,<br />

P. Fevereiro 1,2<br />

1 Plant Cell Biotechnology Laboratory, ITQB, Universidade Nova de Lisboa, Apt 127,<br />

2781-901 Oeiras, Portugal<br />

2 Departamento de Biologia Vegetal, FCUL, Campo Grande, 1749-016 Lisboa, Portugal<br />

3 Centro de Engenharia Biológica, FCUL, Campo Grande, 1749-016 Lisboa, Portugal<br />

a) e-mail: saraujo@itqb.unl.pt<br />

Medicago truncatula cv Jemal<strong>on</strong>g plants expressing the DSP22 showed improved capacity to recover from a<br />

severe water deficit episode after rewatering, when compared to the n<strong>on</strong> trans<str<strong>on</strong>g>for</str<strong>on</strong>g>med line. DSP22, an ELIPlike<br />

protein from Craterostigma plantagineum, is thought to protect the chloroplast against photo-oxidative<br />

damage during the dehydrati<strong>on</strong> and rehydrati<strong>on</strong> process so characteristic <str<strong>on</strong>g>of</str<strong>on</strong>g> this resurrecti<strong>on</strong> plant.<br />

Transgenic plants showed higher Φ PSII values under moderate water deprivati<strong>on</strong> and during rewatering when<br />

compared to the wild type. They also showed higher amounts <str<strong>on</strong>g>of</str<strong>on</strong>g> chlorophylls and lower Chl a.Chl b -1 ratio in<br />

both water regimes. Such results suggest that the protective roles <str<strong>on</strong>g>of</str<strong>on</strong>g> the ELIP-like DSP22 against water stress<br />

or during rewatering may be associated with the transient binding to photosynthetic pigments and/or with<br />

the preservati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> chloroplast protein complexes, such as the LHCII. Biomass accumulati<strong>on</strong> in the<br />

transgenic line was less affected by water deficit than in the n<strong>on</strong> transgenic lines. Transgenic M. truncatula<br />

plants showed evidences <str<strong>on</strong>g>of</str<strong>on</strong>g> a higher potential to withstand water stress than the n<strong>on</strong> transgenic, an aspect<br />

that could explain the better chances to recover from temporary water stress episodes, which is an important<br />

issue when addressing plant improvement toward water stress.<br />

220


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

4E_18_P<br />

(poster secti<strong>on</strong> B2, poster board #299, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ANALYSIS OF SALT AND DROUGHT TOLERANCE IN TRANSGENIC TOMATO<br />

OVEREXPRESSING THE ARABIDOPSIS TRANSCRIPTION FACTOR ATHB7<br />

R. Iannac<strong>on</strong>e, A. Petrozza, N. Armentano, G. Tadd<strong>on</strong>io, G. Sagace, H. Johanness<strong>on</strong>*,<br />

P. Engstrom*, F. Cellini<br />

Metap<strong>on</strong>tum Agrobios, Metap<strong>on</strong>to (MT), Italy; *Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Physiological Botany, Evoluti<strong>on</strong>ary Biology <str<strong>on</strong>g>Centre</str<strong>on</strong>g>,<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Uppsala, Villavagen 6, SE-752 36 Uppsala, Sweden<br />

e-mail: riannac<strong>on</strong>e@agrobios.it<br />

ATHB7 is a member <str<strong>on</strong>g>of</str<strong>on</strong>g> HD-zip protein and was dem<strong>on</strong>strated to be involved in drought resp<strong>on</strong>se in plants.<br />

ATHB7 is transcripti<strong>on</strong>ally induced by dehydrati<strong>on</strong> in post-germinative stages <str<strong>on</strong>g>of</str<strong>on</strong>g> the life cycle and is<br />

supposed to act as a negative regulator <str<strong>on</strong>g>of</str<strong>on</strong>g> growth (Ols<strong>on</strong>n et al. 2004). C<strong>on</strong>stitutive expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ATHB7 in<br />

Arabidopsis led to a suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> shoot el<strong>on</strong>gati<strong>on</strong> growth and retardati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> bolting.<br />

To assess the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> this gene in an heterologous system we obtained transgenic tomato plants<br />

overexpressing ATHB7 mRNA, by using a CaMV35S::ATHB7 c<strong>on</strong>struct. Transgenic T0 plants did not show<br />

any visible phenotypic alterati<strong>on</strong> and selected lines showed high level <str<strong>on</strong>g>of</str<strong>on</strong>g> expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ATHB7 RNA.<br />

Drought and salt tolerance was evaluated in T2 tomato progeny <str<strong>on</strong>g>of</str<strong>on</strong>g> four independent transgenic lines. Six<br />

replicates <str<strong>on</strong>g>for</str<strong>on</strong>g> each line were grown in greenhouse under normal agr<strong>on</strong>omic trial and under stress c<strong>on</strong>diti<strong>on</strong>.<br />

Drought tolerance was evaluated using <strong>on</strong>e third <str<strong>on</strong>g>of</str<strong>on</strong>g> irrigati<strong>on</strong> water respect to the not-stressed plants while<br />

salt tolerance was evaluated using irrigati<strong>on</strong> water c<strong>on</strong>taining 150 mM NaCl. Physiological and agr<strong>on</strong>omical<br />

data were collected during the trial and differences in stress resp<strong>on</strong>se were observed am<strong>on</strong>g transgenic lines<br />

and respect to the isogenic lines. No meaningful differences were observed between transgenic and n<strong>on</strong><br />

transgenic lines in the n<strong>on</strong>-stressed trials. The experiment was repeated using T3 progeny <strong>on</strong> the two best<br />

per<str<strong>on</strong>g>for</str<strong>on</strong>g>ming transgenic lines and the n<strong>on</strong> transgenic c<strong>on</strong>trol.<br />

Results <strong>on</strong> agr<strong>on</strong>omic per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance and physiological data <str<strong>on</strong>g>of</str<strong>on</strong>g> tested plants will be discussed.<br />

4E_19_P<br />

(poster secti<strong>on</strong> B2, poster board #300, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ROLE OF PHOTOSYNTHETIC PERFORMANCE IN SALT STRESS ACCLIMATION<br />

OF TOMATO AFTER SALICYLIC ACID PRE-TREATMENT<br />

Katalin Gémes, Ágnes Szepesi, Adrienn Guóth, Irma Tari<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Physiology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Szeged, 6701 Szeged POB 654, Hungary<br />

e-mail: pigsy17@freemail.hu, tari@bio.u-szeged.hu<br />

Impositi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> salt stress reduced the net CO 2 assimilati<strong>on</strong> rate, chlorophyll (Chl) and carotenoid c<strong>on</strong>tents,<br />

stomatal c<strong>on</strong>ductance and biomass producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> tomato (Lycopersic<strong>on</strong> esculentum Mill. L. cv. Rio Fuego). Pretreatments<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> plants with 10 -4 M, but not with 10 -7 M salicylic acid (SA), could partially restore the CO 2<br />

fixati<strong>on</strong> rate, Chl a/b ratio, carotenoid levels under 100 mM NaCl exposure. Accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> soluble sugars,<br />

a biochemical marker <str<strong>on</strong>g>of</str<strong>on</strong>g> salinity tolerance <str<strong>on</strong>g>of</str<strong>on</strong>g> tomato, could be detected in pre-treated plants. After SA<br />

applicati<strong>on</strong>, c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> hexoses (glucose and fructose) remained high in the leaves and that <str<strong>on</strong>g>of</str<strong>on</strong>g> sucrose<br />

in the roots during salt stress. Both SA and salt stress increased the H 2 O 2 producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> tissues. Activities <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

superoxide dismutase and catalase (CAT), a H 2 O 2 generating and scavenging enzymes, respectively, decreased<br />

significantly under salt stress, but in 10 -4 M SA pre-treated plants, CAT activity was significantly induced both<br />

221


23-26 August 2007,<br />

Budapest, Hungary<br />

in the root and shoot tissues. The improved photosynthetic per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance, the accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> soluble sugars<br />

as compatible osmolytes and the effective eliminati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> H 2 O 2 by CAT c<strong>on</strong>tributed to the successful<br />

acclimati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 10 -4 M SA pre-treated tomato plants to high salinity. – This work was supported by OTKA<br />

T038392.<br />

4E_20_P<br />

(poster secti<strong>on</strong> B2, poster board #301, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DEVELOPING SALT STRESS TOLERANCE IN THE LEGUME VIGNA MUNGO<br />

(BLACKGRAM) USING THE TRANSGENIC APPROACH<br />

Neera Bhalla Sarin 1* , C. P.Upadhyaya 1 , P. Bhomkar 1 , R. Rajwanshi 1 , Nishakant Pandey 1 , N. Shiva<br />

Prakash 1 , Mikhail Pooggin 2 , Thomas Hohn 2<br />

1 Jawaharlal Nehru University, New Delhi, India<br />

2 Botanical Institute, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Basel, Switzerland<br />

1* Corresp<strong>on</strong>ding author: Neera Bhalla Sarin, Pr<str<strong>on</strong>g>of</str<strong>on</strong>g>essor, SLS, JNU, New Delhi-110067<br />

e-mail: cerac<strong>on</strong>@vsnl.com<br />

The producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> pulses is facing a number <str<strong>on</strong>g>of</str<strong>on</strong>g> c<strong>on</strong>straints due to soil salinity, poor soil fertility and poor<br />

moisture retenti<strong>on</strong> by marginal and sub marginal soils. Blackgram [Vigna mungo (L) Hepper] is an important<br />

pulse <str<strong>on</strong>g>of</str<strong>on</strong>g> India providing a rich source <str<strong>on</strong>g>of</str<strong>on</strong>g> dietary protein to the largely vegetarian populati<strong>on</strong>. Adverse biotic<br />

and abiotic stresses limit the yield <str<strong>on</strong>g>of</str<strong>on</strong>g> Vigna mungo. Because <str<strong>on</strong>g>of</str<strong>on</strong>g> narrow genetic base in pure line selecti<strong>on</strong> and<br />

natural variati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> abiotic stress tolerance, a viable opti<strong>on</strong> is to genetically engineer blackgram <str<strong>on</strong>g>for</str<strong>on</strong>g> abiotic<br />

stress tolerance. Reproducible and efficient regenerati<strong>on</strong> systems <str<strong>on</strong>g>of</str<strong>on</strong>g> blackgram have been utilized <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

genetic trans<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Vigna mungo using a novel c<strong>on</strong>struct pnpt-lox-csm-GlyI in which the glyoxalaseI(gly1)<br />

gene is driven by the c<strong>on</strong>stitutive Cestrum yellow leaf curl virus (csm) promoter, and the selecti<strong>on</strong> marker<br />

(kanamycin) gene is flanked by lox sites, allowing its removal by the lox sequence specific Cre-recombinase.<br />

Southern blot analysis was d<strong>on</strong>e by including the left and the right border sequences <str<strong>on</strong>g>of</str<strong>on</strong>g> the gene c<strong>on</strong>struct<br />

which c<strong>on</strong>firmed the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> the desired gly1 gene in the transgenic plants. Single copy inserti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

glyI gene were found in the transgenic plants. Physiological tests <str<strong>on</strong>g>for</str<strong>on</strong>g> salt tolerance were per<str<strong>on</strong>g>for</str<strong>on</strong>g>med using leaf<br />

disc assays as well as whole plants grown under in vitro and in vivo c<strong>on</strong>diti<strong>on</strong>s. In leaf disc assays, the leaf discs<br />

from trans<str<strong>on</strong>g>for</str<strong>on</strong>g>med plants showed delayed senescence and retained higher amount <str<strong>on</strong>g>of</str<strong>on</strong>g> chlorophyll. Under in<br />

vitro as well as in vivo growth c<strong>on</strong>diti<strong>on</strong>s, the transgenic plants showed better growth as compared to the<br />

untrans<str<strong>on</strong>g>for</str<strong>on</strong>g>med c<strong>on</strong>trol plants when subjected to saline c<strong>on</strong>diti<strong>on</strong>s.<br />

The challenges and pitfalls <str<strong>on</strong>g>of</str<strong>on</strong>g> this investigati<strong>on</strong> will be discussed in order to find plausible soluti<strong>on</strong>s.<br />

222


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

4E_21_P<br />

(poster secti<strong>on</strong> B2, poster board #302, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CURRENT UNDERSTANDING ON THE MECHANISM OF RESISTANCE TO<br />

XENOBIOTIC STRESSOR BIPYRIDYL HERBICIDES IN HORSEWEED CONYZA<br />

CANADENSIS L. (CRONQ.)<br />

Zoltán Szigeti, Tamás Visnovitz, Vilmos Soós, Balázs Jóri, Dóra Szegő, Il<strong>on</strong>a Rácz, Demeter Lásztity<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Physiology and Molecular Plant Biology, Eötvös Loránd University H-1117 Budapest,<br />

Pázmány Péter Lane 1/c, Hungary; e-mail: bioszigeti@ludens.elte.hu<br />

Bipyridyl herbicides paraquat and diquat exert their phytotoxic effect by diverting electr<strong>on</strong>s from the<br />

photosynthetic electr<strong>on</strong> transport chain <strong>on</strong> the reducing side <str<strong>on</strong>g>of</str<strong>on</strong>g> PSI generating reactive oxygen radicals. To<br />

reveal mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> paraquat resistance, in our earlier experiments we studied uptake and intracellular<br />

localizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> paraquat, the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> selective transporter inhibitors <strong>on</strong> these processes, and paraquat<br />

induced changes <str<strong>on</strong>g>of</str<strong>on</strong>g> gene expressi<strong>on</strong> in susceptible and resistant biotypes. According to these results<br />

resistance seems to be a paraquat-induced mechanism, based <strong>on</strong> the sequestrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> paraquat into a<br />

metabolically inactive compartment, the vacuole. Participati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> not directly energized transporters are<br />

suggested in this process, howewer, the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> inhibitor <str<strong>on</strong>g>of</str<strong>on</strong>g> V type vacuolar ATPases refers to the energyrequirement<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the transport. Paraquat resistant biotype <str<strong>on</strong>g>of</str<strong>on</strong>g> horseweed exhibits a lower extent <str<strong>on</strong>g>of</str<strong>on</strong>g> resistance to<br />

the chemically related bipyridyl-compound diquat. To reveal similarities and differences in the mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

paraquat and diquat resistance, in the course <str<strong>on</strong>g>of</str<strong>on</strong>g> the present work we studied the uptake and physiological<br />

effect <str<strong>on</strong>g>of</str<strong>on</strong>g> diquat by fluorescence imaging and by determining changes in functi<strong>on</strong>al activity (based <strong>on</strong> changes<br />

in variable fluorescence Fv/Fm values), as well as the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> selective transporter inhibitors <strong>on</strong> intracellular<br />

sequestrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> diquat. Results <str<strong>on</strong>g>of</str<strong>on</strong>g> recent experiments indicate less effective vacuolar sequestrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> diquat<br />

as compared to paraquat in the resistant biotype.<br />

4E_22_P<br />

(poster secti<strong>on</strong> B2, poster board #303, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DEVELOPMENT OF EXPRESSION CASSETTES FOR STRESS-INDUCED AND<br />

ROOT-SPECIFIC EXPRESSION IN CEREALS<br />

Zoltán Zombori, Antal Mai, Katalin Török, Dénes Dudits, János Györgyey<br />

Biological <str<strong>on</strong>g>Research</str<strong>on</strong>g> Center <str<strong>on</strong>g>of</str<strong>on</strong>g> the H.A.S., Temesvári krt. 62. H-6726 Szeged, Hungary<br />

e-mail: arthur@brc.hu<br />

Abiotic stresses are very important factors that can reduce the crop producti<strong>on</strong>. Plant root is the primary<br />

organ <str<strong>on</strong>g>for</str<strong>on</strong>g> uptake <str<strong>on</strong>g>of</str<strong>on</strong>g> water and nutrients, there<str<strong>on</strong>g>for</str<strong>on</strong>g>e plays important role in tolerance to stresses like drought or<br />

salinity. Plants developing str<strong>on</strong>ger and deeper roots suffer less from water deficit. Genes facilitating<br />

development <str<strong>on</strong>g>of</str<strong>on</strong>g> efficient root system during drought stress can increase the survival <str<strong>on</strong>g>of</str<strong>on</strong>g> the plant. Fusi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

drought stress-related root-specific promoters to these genes may provide envir<strong>on</strong>ment friendly and efficient<br />

soluti<strong>on</strong> to improve roots <str<strong>on</strong>g>of</str<strong>on</strong>g> crop plants under stress c<strong>on</strong>diti<strong>on</strong>s. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> our work is to develop<br />

expressi<strong>on</strong> cassettes <str<strong>on</strong>g>for</str<strong>on</strong>g> cereals <str<strong>on</strong>g>for</str<strong>on</strong>g> the above purpose. Two promoters were selected, the rice Catalase B and<br />

the RSOsPR10 promoters. The CatB promoter is known to be root-specific, the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the RSOsPR10<br />

mRNA is high in salt and drought stress c<strong>on</strong>diti<strong>on</strong>s in root. Both promoters have stress-related transcripti<strong>on</strong><br />

factor binding sites and motifs (MYB, WRKY, DREB, LTRE, root motifs) in their sequence. The 1.6 kb<br />

CatB promoter and the 2 kb <str<strong>on</strong>g>of</str<strong>on</strong>g> the 5` flanking regi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the RSOsPR10 were cl<strong>on</strong>ed, and fusi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> these<br />

223


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Budapest, Hungary<br />

promoters to reporter genes (GUS, eGFP, YFP) were c<strong>on</strong>structed and trans<str<strong>on</strong>g>for</str<strong>on</strong>g>med into rice Nipp<strong>on</strong>bare cv.<br />

calli. On the regenerated T 0 rice plants, salt stress was per<str<strong>on</strong>g>for</str<strong>on</strong>g>med that revealed the RSOsPR10 promoter<br />

directing root-specific and stress-induced expressi<strong>on</strong> pattern <str<strong>on</strong>g>of</str<strong>on</strong>g> GFP in stress c<strong>on</strong>diti<strong>on</strong>s. Tests <strong>on</strong> the CatB<br />

trans<str<strong>on</strong>g>for</str<strong>on</strong>g>med calli, regenerati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> their T 0 plants are in progress. Independent T 1 lines <str<strong>on</strong>g>of</str<strong>on</strong>g> RSOsPR10<br />

transgenic rice plants will be generated <str<strong>on</strong>g>for</str<strong>on</strong>g> detailed characterizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the expressi<strong>on</strong> cassette.<br />

4E_23_P<br />

(poster secti<strong>on</strong> B2, poster board #304, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

TRANSCRIPT PROFILES MAY REFLECT DIFFERENT ADAPTATION STRATEGIES<br />

DURING DROUGHT STRESS IN WHEAT<br />

Maria Sečenji*, Ágnes Lendvai*, Pál Miskolczi*, Gábor Kocsy†, Nils Stein‡, Patrick Schweizer‡,<br />

Dénes Dudits*, János Györgyey*<br />

*Biological <str<strong>on</strong>g>Research</str<strong>on</strong>g> Center <str<strong>on</strong>g>of</str<strong>on</strong>g> the H.A.S., Temesvári krt.62., 6726 Szeged, Hungary<br />

†Agricultural <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> the H.A.S., Brunszvik u. 2., 2462 Mart<strong>on</strong>vásár, Hungary<br />

‡Institut für Pflanzengenetik und Kulturpflanzen<str<strong>on</strong>g>for</str<strong>on</strong>g>schung, Correnstraße 3., 06466 Gatersleben, Germany<br />

e-mail: arthur@brc.hu<br />

Drought is <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the major abiotic factors that limits agricultural crop producti<strong>on</strong>. Nevertheless, water<br />

deficit usually coincides with other abiotic stresses, adversely effecting plant growth. Plants can survive these<br />

adverse c<strong>on</strong>diti<strong>on</strong>s using different strategies. Plant resistance to water deficit may arise from escape,<br />

avoidance, or tolerance strategies. Plants usually apply a combinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these strategies under dry c<strong>on</strong>diti<strong>on</strong>s.<br />

These distinct strategies are caused by underlying transcripti<strong>on</strong>al alterati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> a high number <str<strong>on</strong>g>of</str<strong>on</strong>g> genes. To<br />

follow the transcripti<strong>on</strong>al changes under water deficit, cDNA macroarray was hybridized with root samples<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> two wheat genotypes, both displaying tolerance to water-deficit stress but following distinct strategies,<br />

derived from an experimental system based <strong>on</strong> reduced irrigati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> simulating stress c<strong>on</strong>diti<strong>on</strong>s. As a result,<br />

8.0 % <str<strong>on</strong>g>of</str<strong>on</strong>g> the genes were up-regulated and 8.5 % were down-regulated in the adaptive cultivar. In the<br />

“escaper” genotype, these ratios were 5.1 % and 4.8 %, respectively. After functi<strong>on</strong>al classificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> genes<br />

induced by water-deficit stress, three groups <str<strong>on</strong>g>of</str<strong>on</strong>g> genes displayed significant differences between the two<br />

genotypes: stress- and defense-related; signal transducti<strong>on</strong> comp<strong>on</strong>ents; cell organizati<strong>on</strong> and cell wall<br />

biogenesis-related genes. These results suggest a hypothetical elucidati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the molecular genetic background<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the different tolerance strategies <str<strong>on</strong>g>of</str<strong>on</strong>g> the two examined wheat genotypes.<br />

224


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

4F. DROUGHT AND DESICCATION STRESSES IN ECOSYSTEMS<br />

4F_01_P<br />

(poster secti<strong>on</strong> B2, poster board #305, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

RESPONSE OF ORNAMENTAL PERENNIALS TO DROUGHT STRESS IN URBAN<br />

CONDITIONS<br />

Wojciech K. Chylinski, Aleksandra J. Lukaszewska<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Ornamental Plants, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Horticulture and Landscape Architecture,<br />

Warsaw Agricultural University<br />

159 Nowoursynowska Str., 02-776 Warsaw, Poland<br />

e-mail: chylinski@mail.pt<br />

In comparis<strong>on</strong> to natural habitats (<str<strong>on</strong>g>for</str<strong>on</strong>g>ests, grasslands, wetlands and riparian z<strong>on</strong>es) cities and towns represent<br />

very young habitats (Wittig 2004). The presence <str<strong>on</strong>g>of</str<strong>on</strong>g> high anthropopressure and the extensive infrastructure <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

urban areas have a major impact <strong>on</strong> such envir<strong>on</strong>mental factors as climate, soil layer, hydrology, disturbance<br />

regime, and management practices (Sukopp and Heneke 1983; Sudnik-Wójcikowska and Galera 2005; Turner<br />

et al. 2005). The most exposed to drought, which is the most important abiotic stress in urban areas, are the<br />

herbaceous plants: perennials and annuals. In this trial a resp<strong>on</strong>se to water stress <str<strong>on</strong>g>of</str<strong>on</strong>g> three comm<strong>on</strong>ly used<br />

garden species, avens (Geum coccineum Sm.), heartleaf saxifrage (Bergenia cordifolia (Haw.) Sternb.) and st<strong>on</strong>ecrop<br />

(Sedum spectabile Bor.) was compared. The first avens is very herbaceous and pr<strong>on</strong>e to wilting, heartleaf<br />

saxifrage has thick rhizome and leathery, evergreen leaves and the last <strong>on</strong>e is a succulent species adapted to<br />

drought. During two vegetative seas<strong>on</strong>s (2005 and 2006) plants were subjected to water stress: drought was<br />

imposed by withhelding watering during 10-days cycles, separated by 10-days <str<strong>on</strong>g>of</str<strong>on</strong>g> normal watering.<br />

Amm<strong>on</strong>ium c<strong>on</strong>tent in leaves all the species increased significantly under stress but the range <str<strong>on</strong>g>of</str<strong>on</strong>g> increase was<br />

different. The reducti<strong>on</strong> in the a+b chlorophyll c<strong>on</strong>centrati<strong>on</strong> in leaves <str<strong>on</strong>g>of</str<strong>on</strong>g> avens was significantly time- and<br />

stress dependent while reacti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> st<strong>on</strong>ecrop and heartleaf saxifrage was less spectacular. The above results<br />

show that amm<strong>on</strong>ium and chlorophyll a+b c<strong>on</strong>tents merit further attenti<strong>on</strong> as possible indicators <str<strong>on</strong>g>of</str<strong>on</strong>g> plant<br />

resp<strong>on</strong>se to drought stress in ornamental plants but additi<strong>on</strong>al studies are needed be<str<strong>on</strong>g>for</str<strong>on</strong>g>e these parameters can<br />

be used to evaluate new plants <str<strong>on</strong>g>for</str<strong>on</strong>g> introducti<strong>on</strong> into urban growing c<strong>on</strong>diti<strong>on</strong>s, or as selecti<strong>on</strong> criteria in<br />

breeding <str<strong>on</strong>g>for</str<strong>on</strong>g> adaptati<strong>on</strong> to demanding growing c<strong>on</strong>diti<strong>on</strong>s.<br />

225


23-26 August 2007,<br />

Budapest, Hungary<br />

4G. HEAVY METAL STRESS<br />

4G_01_P<br />

(poster secti<strong>on</strong> B2, poster board #306, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

GENOTOXIC AND HISTOLOGIC EFFECT OF CHROMIUM EXPOSURE IN<br />

LACTUCA SATIVA<br />

Eleazar Rodriguez, André Ferreira, Mónica Almeida, Armando Costa, João Loureiro,<br />

C<strong>on</strong>ceição Santos<br />

CESAM & Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Aveiro, 3810-193 Aveiro, Portugal<br />

e-mail: eleazar@bio.ua.pt<br />

Chromium is a highly toxic n<strong>on</strong>-essential metal <str<strong>on</strong>g>for</str<strong>on</strong>g> microorganisms and plants. Due to its widespread<br />

industrial use, chromium (Cr) has become a serious pollutant in diverse envir<strong>on</strong>mental settings. Cr toxicity in<br />

plants is less studied than in animals, but it seems to depend <strong>on</strong> Cr valence state: Cr 6+ , more soluble and<br />

mobile, is more toxic than Cr 3+ . So, in this work we studied the genotoxic and histologic effects <str<strong>on</strong>g>of</str<strong>on</strong>g> this two<br />

Cr species in three Lactuca sativa L. cultivars (Póvoa, Teide and Twistil). For this, plantlets were grown <strong>on</strong><br />

solid matrix and watered with ½ Hoagland soluti<strong>on</strong> c<strong>on</strong>taining final c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> 0, 50, 150 and 300<br />

mg/L <str<strong>on</strong>g>of</str<strong>on</strong>g> Cr 3+ and Cr 6+ (supplied respectively as CrCl 3 .6H 2 O and as K 2 CrO 4 ). After 30 days, leaves and roots<br />

were sampled <str<strong>on</strong>g>for</str<strong>on</strong>g> histological and flow cytometric (FCM) analyses (0 and 300 mg/L). Nuclear DNA c<strong>on</strong>tent<br />

and the G 2 :G0/G 1 ratio were estimated, and changes in the full peak coefficient <str<strong>on</strong>g>of</str<strong>on</strong>g> variati<strong>on</strong> (mesure <str<strong>on</strong>g>of</str<strong>on</strong>g> DNA<br />

damage) <str<strong>on</strong>g>of</str<strong>on</strong>g> the G 0 /G 1 peak were recorded. While in roots, significant differences in nuclear DNA c<strong>on</strong>tent<br />

were detected between c<strong>on</strong>trol and exposed plants exposed to Cr 3+ (all cultivars), <str<strong>on</strong>g>for</str<strong>on</strong>g> leaves this was verified<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> plants exposed to Cr 6+ (cv. Póvoa). As <str<strong>on</strong>g>for</str<strong>on</strong>g> the G 2 :G0/G 1 ratio, a significant increase was observed in<br />

roots <str<strong>on</strong>g>of</str<strong>on</strong>g> plants exposed to Cr 6+ , whereas in leaves this ratio decreased (all cultivars).<br />

4G_02_P<br />

(poster secti<strong>on</strong> B2, poster board #307, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HEAVY METAL STRESS AND STRESS RESPONSES IN PISUM SATIVUM L<br />

B. Bartha, L. Erdei<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Physiology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Szeged, P.O.B. 654., 6701 Szeged, Hungary<br />

e-mail: barthab@bio.u-szeged.hu; erdei@bio.u-szeged.hu<br />

Several metals - such as copper – are essential microelements but in high c<strong>on</strong>centrati<strong>on</strong>s they damage plants<br />

by causing oxidative stress. The effect <str<strong>on</strong>g>of</str<strong>on</strong>g> copper was investigated in 2 weeks-old Pisum sativum, after <strong>on</strong>e-week<br />

treatment with different CuSO 4 c<strong>on</strong>centrati<strong>on</strong>s (5, 10, 25 and 50 µM). Growth parameters (root-shoot length<br />

and mass), the capacity <str<strong>on</strong>g>of</str<strong>on</strong>g> the heavy metal uptake, the root-to-shoot translocati<strong>on</strong> and the changes <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

macro element distributi<strong>on</strong> were measured. Catalase (CAT), guaiacol-peroxidase (GPX), glutathi<strong>on</strong>e reductase<br />

(GR) and glutathi<strong>on</strong>e-S-transferase (GST) activities were also determined and NO synthesis was detected in<br />

roots. Plants kept the metal i<strong>on</strong>s in their roots, while a minimal translocati<strong>on</strong> to the shoot was detected <strong>on</strong>ly<br />

in cases <str<strong>on</strong>g>of</str<strong>on</strong>g> 25 and 50 µM treatments. Cu 2+ caused growth inhibiti<strong>on</strong> in roots; it means 30-50% decrease in<br />

root length and mass compared to c<strong>on</strong>trol plants. GR activity decreased dramatically in root tissues while in<br />

shoot we observed increased enzyme activities. GST activities slightly decreased in both tissues. In plants<br />

treated with 25 µM Cu 2+ , time dependence <str<strong>on</strong>g>of</str<strong>on</strong>g> enzyme activities was measured. GPX activity increased in both<br />

226


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

roots and leaves; GR and CAT activities increased in shoots but decreased in roots. NO producti<strong>on</strong> was<br />

measured after 100 µM heavy metal treatment from 1 to 48 hours. On the basis <str<strong>on</strong>g>of</str<strong>on</strong>g> the time course, the Cu 2+ -<br />

induced NO producti<strong>on</strong> clearly showed a biphasic reacti<strong>on</strong>, namely, a fast burst <str<strong>on</strong>g>of</str<strong>on</strong>g> NO release followed by a<br />

slow increase. It is c<strong>on</strong>cluded that the immediate stress resp<strong>on</strong>se in the alarm phase is the decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> defence<br />

capacity accompanied by the appearance <str<strong>on</strong>g>of</str<strong>on</strong>g> NO in the root, while in the shoot acclimatizati<strong>on</strong> processes are<br />

initiated.<br />

4G_03_P<br />

(poster secti<strong>on</strong> B2, poster board #308, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CD-INDUCED CHANGES IN THE EXPRESSION OF CHLOROPHYLL-PROTEIN<br />

COMPLEXES IN POPULUS GLAUCA<br />

B. Basa, M. Gárd<strong>on</strong>yi, É. Sárvári*, L. Tamás<br />

Dept Plant Physiol & Mol Plant Biol, Eötvös Univ, Budapest, Hungary<br />

*e-mail: e_sarvari@ludens.elte.hu<br />

The molecular resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> chlorophyll(Chl)-protein pattern to Cd treatment was studied in poplar plants<br />

grown in hydrop<strong>on</strong>ics. The relati<strong>on</strong>ship between Chl-protein level and their mRNA level was aimed to<br />

examine. Plants were treated with 10 µM Cd(NO 3 ) 2 starting at their four-leaf stage. Structural and functi<strong>on</strong>al<br />

photosynthetic parameters were measured and RNA samples were extracted from developing and mature<br />

leaves during the two-week treatment. Chl-protein pattern analyzed by Deriphate-PAGE revealed that the<br />

most effected complex was the photosystem I including its light-harvesting antenna, LHCI. To investigate<br />

whether the reduced amount <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins was due to the change <str<strong>on</strong>g>of</str<strong>on</strong>g> the expressi<strong>on</strong> pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ile, the mRNA level <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the Lhca1-4 genes has been analyzed by quantitative real-time RT-PCR. In additi<strong>on</strong>, their stress-enhanced<br />

relatives Lhca5 and Lil1 gene (ELIPs early light induced proteins) were also studied. Transcripti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

rarely expressed Lhca5 slightly enhanced in mature leaves at the beginning <str<strong>on</strong>g>of</str<strong>on</strong>g> the Cd treatment. However, in<br />

young developing leaves high expressi<strong>on</strong> level was detected, which decreased remarkably during leaf<br />

development. In Cd-treated plants a str<strong>on</strong>g reducti<strong>on</strong> was dem<strong>on</strong>strated. Similar results were observed in the<br />

case <str<strong>on</strong>g>of</str<strong>on</strong>g> Lil1 gene, though its enhancement was str<strong>on</strong>ger in mature leaves in early phase <str<strong>on</strong>g>of</str<strong>on</strong>g> the treatment. The<br />

abundantly expressed Lhca genes (1-4) displayed identical transcripti<strong>on</strong>al behavior. In young expanding leaves<br />

Cd inhibited their transcripti<strong>on</strong> and mRNA levels decreased gradually, whereas in mature leaves Cd-induced<br />

inhibiti<strong>on</strong> was less pr<strong>on</strong>ounced. The results obtained at mRNA level were c<strong>on</strong>sistent with those at protein<br />

level. It is hypothesized that the reducti<strong>on</strong> in the amount <str<strong>on</strong>g>of</str<strong>on</strong>g> LHCI antenna under Cd stress was triggered by<br />

transcripti<strong>on</strong>al resp<strong>on</strong>se.<br />

227


23-26 August 2007,<br />

Budapest, Hungary<br />

4G_04_P<br />

(poster secti<strong>on</strong> B2, poster board #309, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ANTIOXIDANT AND MORPHOLOGIC ANALYSIS OF LYCOPERSICON<br />

ESCULENTUM CV MICRO-TOM PLANTS SUBJECTED TO CADMIUM STRESS<br />

1P. L. Gratão, 1 C. C. M<strong>on</strong>teiro, 2 M. L. Rossi, 2 A. P. M. Rodriguez, 1 L. E. P. Peres, 1* R. A. Azevedo<br />

1Lab. Genética Bioquímica de Plantas/Dep. Genética/ESALQ/USP/Piracicaba/SP/Brasil;<br />

2Lab. Biotecnologia Vegetal/CENA/USP/Piracicaba/SP/Brasil;<br />

1*Pesquisador Dep. Genética - ESALQ/USP/Piracicaba/SP/Brasil, e-mail: raazeved@esalq.usp.br<br />

The c<strong>on</strong>taminati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> envir<strong>on</strong>ments by heavy metal polluti<strong>on</strong> mainly by cadmium (Cd), which is c<strong>on</strong>sidered<br />

<strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the more toxic, is generated by mining, industrial activities, sewage sludge and phosphated fertilizers<br />

used in agriculture. Cd can induce excessive producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> reactive oxygen species (ROS) which can cause<br />

serious damage to cellular metabolism, leading to growth inhibiti<strong>on</strong>. A class <str<strong>on</strong>g>of</str<strong>on</strong>g> antioxidant enzymes acts in<br />

the scavenging <str<strong>on</strong>g>of</str<strong>on</strong>g> ROS and plants exposed to heavy metals may exhibit alterati<strong>on</strong> in enzyme activity. In this<br />

study we observed alterati<strong>on</strong>s in CAT and peroxidases activities in tomato cv Micro-Tom plants. SOD<br />

isoenzymes activities were also shown to be altered. Besides, morphologic analysis allowed the evaluati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Cd-induced damage. The results suggest that in tomato cv Micro-Tom the main defence system to Cd stress<br />

is variable during the heavy metal exposure period and its c<strong>on</strong>centrati<strong>on</strong>. The study <str<strong>on</strong>g>of</str<strong>on</strong>g> such resp<strong>on</strong>ses may<br />

allow the evaluati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> tolerance levels and specificity <str<strong>on</strong>g>of</str<strong>on</strong>g> the resp<strong>on</strong>se to levels <str<strong>on</strong>g>of</str<strong>on</strong>g> polluti<strong>on</strong> in the<br />

envir<strong>on</strong>ment. These data may be useful in breeding programs or biotechnological alternatives to produce<br />

and/or select tolerant plants that may be used in phytoremediati<strong>on</strong> in order to reduce the amount <str<strong>on</strong>g>of</str<strong>on</strong>g> heavy<br />

metals in c<strong>on</strong>taminated areas. Another important aspect is the identificati<strong>on</strong> or selecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> plants that<br />

accumulate very low amounts <str<strong>on</strong>g>of</str<strong>on</strong>g> Cd in the parts that are used <str<strong>on</strong>g>for</str<strong>on</strong>g> animal and human food.<br />

4G_05_P<br />

(poster secti<strong>on</strong> B2, poster board #310, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

KINETICS OF PHOTOSYNTHETIC RESPONSES AND DEVELOPMENT OF<br />

PROTECTIVE MECHANISMS DURING CD STRESS IN POPLAR<br />

P. Szegi* 1 , Á. Solti 1 , L. Gáspár 1 , I. Mészáros 2 , É. Sárvári 1<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Physiology & Molecular Plant Biology, Eötvös University, Budapest, Hungary<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Botany, Debrecen University, Debrecen, Hungary<br />

*e-mail: sheedy@mad.hu<br />

It is well known that Cd inhibits photosynthesis and induces oxidative stress. However, the time scale <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

appearance <str<strong>on</strong>g>of</str<strong>on</strong>g> symptoms and the <strong>on</strong>set <str<strong>on</strong>g>of</str<strong>on</strong>g> the protective mechanisms has been scarcely studied. To get<br />

in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <strong>on</strong> the kinetics <str<strong>on</strong>g>of</str<strong>on</strong>g> processes depending <strong>on</strong> leaf maturity, Cd effects <strong>on</strong> both photosynthetically<br />

competent and developing leaves <str<strong>on</strong>g>of</str<strong>on</strong>g> poplar plants (Populus alba v. Kopeczki, grown in hydrop<strong>on</strong>ics) were<br />

followed during the treatment (with 10 µM Cd(NO3)2) <str<strong>on</strong>g>of</str<strong>on</strong>g> plants being in four-leaf stage <str<strong>on</strong>g>for</str<strong>on</strong>g> a three-week<br />

period. In mature leaves, the actual quantum efficiency <str<strong>on</strong>g>of</str<strong>on</strong>g> photosystem (PS)II (∆F/Fm’) and n<strong>on</strong>photochemical<br />

quenching (NPQ) did not change. Reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> CO2 fixati<strong>on</strong> and stomatal c<strong>on</strong>ductance were<br />

the earliest resp<strong>on</strong>ding parameters starting to change from the day 2-3. Decrease in chlorophyll (Chl) c<strong>on</strong>tent,<br />

Chl a/b ratio, and in the amount <str<strong>on</strong>g>of</str<strong>on</strong>g> PSII core was slight (starting from day 3). The activity <str<strong>on</strong>g>of</str<strong>on</strong>g> ascorbate<br />

peroxidase (APX) decreased, and the amount <str<strong>on</strong>g>of</str<strong>on</strong>g> mal<strong>on</strong>dialdehyde (MDA) slightly increased from day 4<br />

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3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

compared to c<strong>on</strong>trols. In newly developed leaves most <str<strong>on</strong>g>of</str<strong>on</strong>g> the studied parameters were more str<strong>on</strong>gly affected.<br />

Lowered Chl c<strong>on</strong>tent, Chl a/b ratio (decrease in PSI>LHCII>PSII), and inhibited CO2 fixati<strong>on</strong> appeared<br />

early (from day 3), while reducti<strong>on</strong> in ∆F/Fm’, NPQ, APX activity and increase in MDA c<strong>on</strong>tent were<br />

observed from day 4. While CO2 fixati<strong>on</strong> was the most influenced parameter in mature leaves, the stage <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

photosynthetic structures was equally important in developing leaves. A recovery phase started <strong>on</strong> the sec<strong>on</strong>d<br />

week <str<strong>on</strong>g>of</str<strong>on</strong>g> treatment even in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> Cd.<br />

4G_06_P<br />

(poster secti<strong>on</strong> B2, poster board #311, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CD-INDUCED INHIBITION OF PHOTOSYNTHESIS CAN BE RECOVERED BY<br />

ELEVATED FE SUPPLY<br />

Á. Solti 1* , P. Szegi 1 , L. Gáspár 1 , L. Lévai 2 , Z. Szigeti 1 , É. Sárvári 1<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Physiology & Molecular Plant Biology, Eötvös University, Budapest, Hungary<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Botany & Crop Physiology, Debrecen University, Debrecen, Hungary<br />

*e-mail: sadambio@elte.hu<br />

Cd causes Fe deficiency-like symptoms, and thus inhibits photosynthesis. Surplus Fe supplied together with<br />

the Cd diminished or even abolished the effects. However, it was due to the decreased Cd translocati<strong>on</strong> into<br />

the shoots. To clarify how important is the Cd-induced Fe deficiency am<strong>on</strong>g the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> Cd <strong>on</strong><br />

photosynthesis we studied the recovery effect <str<strong>on</strong>g>of</str<strong>on</strong>g> surplus Fe supplied after the Cd symptoms had been<br />

developed. Hydrop<strong>on</strong>ically cultured poplar (Populus glauca v. Kopeczkii) plants were treated with 10 µM<br />

Cd(NO 3 ) 2 <str<strong>on</strong>g>for</str<strong>on</strong>g> a week, during which the Cd symptoms – including growth retardati<strong>on</strong>, decreased chlorophyll<br />

(Chl) synthesis (increased red/far-red fluorescence ratio), reduced Chl a/b ratio and photosynthetic activity<br />

(CO 2 fixati<strong>on</strong>, the effective photosystem II efficiency – ∆F/Fm’), increased blue-green fluorescence (BGF) –<br />

were developed. Next week normal (10 µM) or fivefold amount <str<strong>on</strong>g>of</str<strong>on</strong>g> Fe-citrate was given together with or<br />

without Cd into the nutrient soluti<strong>on</strong>. N<strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the recovery treatments restored the growth rate or BGF to<br />

the c<strong>on</strong>trol level. However, photosynthetic parameters were partially or totally recovered, though the Cd<br />

c<strong>on</strong>tent <str<strong>on</strong>g>of</str<strong>on</strong>g> leaves did not decrease. The recovery <str<strong>on</strong>g>of</str<strong>on</strong>g> red/far-red fluorescence started from the vein regi<strong>on</strong> and<br />

then developed into the farther parts. Kinetically, the higher the Fe c<strong>on</strong>tent, the faster the recovery was.<br />

C<strong>on</strong>cerning the dynamics <str<strong>on</strong>g>of</str<strong>on</strong>g> regenerati<strong>on</strong>, Chl a/b ratio and ∆F/Fm’ were the most sensitive parameters. In<br />

c<strong>on</strong>clusi<strong>on</strong>, the Cd induced Fe deficiency plays a key role in Cd effects <strong>on</strong> photosynthesis, and the defects can<br />

be repaired by extra Fe supply.<br />

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23-26 August 2007,<br />

Budapest, Hungary<br />

4G_07_P<br />

(poster secti<strong>on</strong> B2, poster board #312, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE INFLUENCE OF ELEUTHEROCOCCUS SENTICOSUS (RUPR. ET MAXIM. EX<br />

MAXIM) EXTRACT TO THE TOXICITY OF CADMIUM<br />

Virgilijus Zitkevičius 1 , Nijole Savickiene 1 , Alina Smalinskiene 2 , Vaiva Lesauskaite 2 ,<br />

Stanislovas Ryselis 3 , Oleg Abdrakhmanov 3 , Rima Kregzdyte 3 , Il<strong>on</strong>a Sadauskiene 3 , Le<strong>on</strong>id Ivanov 3 ,<br />

Arunas Savickas 4<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmaceutical Chemistry and Pharmacognosy, Kaunas University <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine<br />

2Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Cardiology, Kaunas University <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine<br />

3Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Biomedical <str<strong>on</strong>g>Research</str<strong>on</strong>g>, Kaunas University <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine<br />

4Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Drug Technology and Pharmaceutical Management, Kaunas University <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Kaunas,<br />

Lithuania<br />

e-mail: virgzit@takas.lt<br />

Cadmium is known to be human carcinogen based <strong>on</strong> sufficient evidence <str<strong>on</strong>g>of</str<strong>on</strong>g> carcinogenicity in humans. It<br />

disturbs the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> biochemical systems <str<strong>on</strong>g>of</str<strong>on</strong>g> cells. The sensitivity <str<strong>on</strong>g>of</str<strong>on</strong>g> cells or tissues to cadmium appears to<br />

be related, at least in part, to their ability to produce metallothi<strong>on</strong>ein (MT), a protective molecule that binds<br />

heavy metals, including cadmium. Activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the MT <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> in resp<strong>on</strong>se to cadmium exposure results<br />

in producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> metallothi<strong>on</strong>ein, which sequesters cadmium, thus limiting its genotoxic effects.<br />

Eleutherococcus senticosus (Rupr. et Maxim. ex Maxim) can modify cadmium influence and it`s toxicity. The aim<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the study was to evaluate the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> Eleutherococcus senticosus (Rupr. et Maxim. ex Maxim) extract <strong>on</strong> the<br />

accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cadmium in liver, kidney and blood and mitotic activity <str<strong>on</strong>g>of</str<strong>on</strong>g> liver cells after the chr<strong>on</strong>ic<br />

intoxicati<strong>on</strong> by cadmium. <str<strong>on</strong>g>Research</str<strong>on</strong>g> was supported by a Grant (No. G-71/06) <str<strong>on</strong>g>of</str<strong>on</strong>g> Lithuanian Foundati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

<str<strong>on</strong>g>Research</str<strong>on</strong>g> and Studies.<br />

Experiments were carried out <strong>on</strong> the white laboratory mice. Mice (n=57) were periodically i.p. injected <str<strong>on</strong>g>for</str<strong>on</strong>g> 6<br />

weeks with CdCl 2 (0.05 LD 50 Cd 2+ ) and Eleutherococcus senticosus (Rupr. et Maxim. ex Maxim) extract soluti<strong>on</strong>s<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> two different c<strong>on</strong>centrati<strong>on</strong>s (0.05 LD 50 and 0.1 LD 50 ) and their combinati<strong>on</strong>s. C<strong>on</strong>trol mice were injected<br />

with 0.9% saline. Cadmium c<strong>on</strong>centrati<strong>on</strong> in blood and tissue specimens was determined by atomic<br />

absorpti<strong>on</strong> spectrophotometer Perkin-Elmer/Zeeman 3030. The number <str<strong>on</strong>g>of</str<strong>on</strong>g> mitotic liver cells was counted in<br />

10 randomly selected reference areas (0.04 mm 2 ). Preparati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> extract from roots <str<strong>on</strong>g>of</str<strong>on</strong>g> Eleutherococcus senticosus<br />

(Rupr. et Maxim. ex Maxim) was made in the factory “Valentis” (Lithuania).<br />

Cd 2+ c<strong>on</strong>centrati<strong>on</strong> in blood <str<strong>on</strong>g>of</str<strong>on</strong>g> mice in group Eleutherococcus senticosus (Rupr. et Maxim. ex Maxim) (0.05)+Cd<br />

was 2,45-fold higher, there<str<strong>on</strong>g>for</str<strong>on</strong>g>e in group Eleutherococcus senticosus (Rupr. et Maxim. ex Maxim) (0.1)+Cd was<br />

1,72-fold higher comparing to cadmium group, in liver that was 1,61-fold and 1,43-fold, respectively, in<br />

kidney 1,72-fold and 1.90-fold, respectively. The mitotic activity <str<strong>on</strong>g>of</str<strong>on</strong>g> liver cells induced by Cd 2+ after injecti<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> Eleutherococcus senticosus (Rupr. et Maxim. ex Maxim) extract was the same as in c<strong>on</strong>trol group.<br />

L<strong>on</strong>g-term injecti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> extract <str<strong>on</strong>g>of</str<strong>on</strong>g> Eleutherococcus senticosus (Rupr. et Maxim. ex Maxim) (0.05 LD 50 and 0.1<br />

LD 50 ) combined with CdCl 2 (0.05 LD 50 ) leads to the significant increase <str<strong>on</strong>g>of</str<strong>on</strong>g> cadmium c<strong>on</strong>centrati<strong>on</strong> in blood<br />

and investigated organs <str<strong>on</strong>g>of</str<strong>on</strong>g> experimental mice. Eleutherococcus senticosus (Rupr. et Maxim. ex Maxim) decreases<br />

the mitotic activity <str<strong>on</strong>g>of</str<strong>on</strong>g> liver cells induced by cadmium.<br />

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3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

4G_08_P<br />

(poster secti<strong>on</strong> B2, poster board #313, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

GLUTATHIONE-S-TRANSFERASE GENES OF THE CILIATE TETRAHYMENA<br />

THERMOPHILA : A LARGE FAMILY OF KEY DEFENCE ENZYMES AGAINST<br />

DIFFERENT STRESSES<br />

Ruth Ortega, Ana Martín-G<strong>on</strong>zález, Juan C. Gutiérrez<br />

Dpto. Microbiología-III. Facultad de Biología. Universidad Complutense (UCM). Madrid 28040. Spain.<br />

e-mail: anamarti@bio.ucm.es<br />

Eukaryotic Glutathi<strong>on</strong>e-S-transferases (GSTs) comprise a large family <str<strong>on</strong>g>of</str<strong>on</strong>g> enzymes protecting cell from a wide<br />

range <str<strong>on</strong>g>of</str<strong>on</strong>g> biotic or abiotic stresses (xenobiotics, heavy metals, oxidative stress). After the complete<br />

macr<strong>on</strong>uclear genome sequence <str<strong>on</strong>g>of</str<strong>on</strong>g> the ciliate T. thermophila becomes published, 63 putative ORFs encoding<br />

GSTs have been detected. Based <strong>on</strong> the amino acid sequence similarity these GST genes have been assigned<br />

to the following phylogenetic classes; 7 to the class Omega (91% identity), 5 to the class Theta (86 %<br />

identity), 2 to the class Zeta 47 (65% identity), to the class Mu (90% identity) and 2 have not been assigned to<br />

any known class. Theta and Omega classes are present in all eukaryotic organisms, while the Mu class is<br />

mammalians specific. So, ciliates (T. thermophila and Paramecium tetraurelia) are the <strong>on</strong>ly eukaryotic<br />

microorganisms with Mu GSTs, which have 30-41% identity with human Mu GSTs. The structural and<br />

chromosomal locati<strong>on</strong> characteristics <str<strong>on</strong>g>of</str<strong>on</strong>g> these 63 ciliate GSTs are described and discussed. Likewise, a<br />

phylogenetic analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> all known ciliate GSTs is carried out. Several GST genes are clustered in the genome<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> T. thermophila, and this locati<strong>on</strong> is related with their expressi<strong>on</strong> capacity. The expressi<strong>on</strong> gene analysis, by<br />

RT-PCR, under different stress c<strong>on</strong>diti<strong>on</strong>s (heavy metals, oxidative stress) and the screening <str<strong>on</strong>g>of</str<strong>on</strong>g> other EST<br />

databases (starvati<strong>on</strong>, c<strong>on</strong>jugati<strong>on</strong>), have shown that; 1- about a 28% <str<strong>on</strong>g>of</str<strong>on</strong>g> Mu GSTs are expressed under the<br />

used stress c<strong>on</strong>diti<strong>on</strong>s. 2- about 50% expressed GST genes, from the same class, are not clustered. 3- several<br />

GST genes are differentially over-expressed under a specific stress with regard c<strong>on</strong>trol.<br />

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232<br />

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Budapest, Hungary


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

MODULE 5<br />

STRESS AT LEVEL OF THE<br />

ORGANISM<br />

233


23-26 August 2007,<br />

Budapest, Hungary<br />

SYMPOSIA<br />

• 5A. Stress, aging and cellular senescence (August 25th Saturday afterno<strong>on</strong>)<br />

(Organizers and chairs: Efstathios S. G<strong>on</strong>os, and Gord<strong>on</strong> J. Lithgow)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> B1, Building B<br />

• 5B. Hsp-s in cerebral protecti<strong>on</strong> (August 25th Saturday morning)<br />

(Organizer and chair: Ian R. Brown)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> B1, Building B<br />

• 5C. Heavy metal stress (August 25th Saturday morning)<br />

(Organizer and chair: Lawrence E. Hightower)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> B1, Building B<br />

• 5D. Stress and the immune resp<strong>on</strong>se (August 25th Saturday afterno<strong>on</strong>)<br />

(Organizer and chair: Tamas Bartfai)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> B1, Building B<br />

• 5E. ISN Symposium <strong>on</strong> stress endocrinology (August 24th Friday morning)<br />

(Organizer and chair: Francois Tr<strong>on</strong>che)<br />

Poster: August 24 th Friday, Poster secti<strong>on</strong> B1, Building B<br />

POSTER SYMPOSIA<br />

• 5F. Exercise stress<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> B1, Building B<br />

• 5G. Stress and nutriti<strong>on</strong><br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> B1, Building B<br />

• 5H. Stress <str<strong>on</strong>g>of</str<strong>on</strong>g> domestic animals and fish<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> B1, Building B<br />

• 5I. Stress and NO<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> B1, Building B<br />

• 5J. Other stress topics at level <str<strong>on</strong>g>of</str<strong>on</strong>g> the organism (Gravity stress, Stress <str<strong>on</strong>g>of</str<strong>on</strong>g> viral infecti<strong>on</strong>s,<br />

Acute phase resp<strong>on</strong>se, Hsp-s in development, Radiati<strong>on</strong> stress – electrosmog)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> B1, Building B<br />

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3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5A. STRESS, AGING AND CELLULAR SENESCENCE<br />

(EFSTATHIOS S. GONOS, GORDON J. LITHGOW)<br />

5A_01_S<br />

LONGEVITY ASSURANCE MOLECULAR PATHWAYS IN HUMAN CELLS<br />

Efstathios S. G<strong>on</strong>os<br />

Nati<strong>on</strong>al Hellenic <str<strong>on</strong>g>Research</str<strong>on</strong>g> Foundati<strong>on</strong>, Athens, Greece<br />

e-mail: sg<strong>on</strong>os@eie.gr<br />

Ageing and l<strong>on</strong>gevity are two multifactorial biological phenomena whose knowledge at molecular level is still<br />

limited. We have developed a cl<strong>on</strong>al senescence induced system and we have cl<strong>on</strong>ed several senescence<br />

associated genes. Analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> the functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the isolated genes, encoding <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

Clusterin/Apolipoprotein J (CLU), suggests that it is a novel survival factor. CLU is found over-expressed in<br />

vitro under a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> stress c<strong>on</strong>diti<strong>on</strong>s and in vivo in samples from patients suffering from various agerelated<br />

diseases as well as in primary tumours which have acquired chemotherapeutic drug resistance. In<br />

additi<strong>on</strong>, it has been dem<strong>on</strong>strated that inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> endogenous CLU expressi<strong>on</strong> by RNA interference<br />

induces growth retardati<strong>on</strong>, higher rates <str<strong>on</strong>g>of</str<strong>on</strong>g> endogenous cellular death and sensitizes human cells to stress<br />

(Cancer Res 64, 1834-1842, 2004). Recent findings indicate that effective and sustained CLU depleti<strong>on</strong> by<br />

siRNA induces late morphological alterati<strong>on</strong>s, growth arrest at the G1/S checkpoint and activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

mitoch<strong>on</strong>drial axis <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis that engages caspase-9. Moreover, CLU knock-down resulted in down<br />

regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the BH pro-survival (bcl-2 and bcl-XL) proteins and activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> p53 and its downstream<br />

targets, namely p21WAF1/CIP1 and bax.<br />

We have also attempted an overall molecular and biochemical approach regarding proteasome functi<strong>on</strong> in<br />

replicative senescence and cell survival. We have observed reduced levels <str<strong>on</strong>g>of</str<strong>on</strong>g> proteasomal peptidase activities<br />

coupled with increased levels <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidized and ubiquitinated proteins in senescent cells. We have found the<br />

catalytic subunits <str<strong>on</strong>g>of</str<strong>on</strong>g> the 20S complex and subunits <str<strong>on</strong>g>of</str<strong>on</strong>g> the 19S regulatory complex to be down-regulated in<br />

senescent cells. This is accompanied by a decrease in the level <str<strong>on</strong>g>of</str<strong>on</strong>g> both 20S and 26S complexes (J Biol Chem<br />

278, 28026-28037, 2003). In support, partial inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proteasomes in young cells by specific inhibitors<br />

induced a senescence-like phenotype. Stable over-expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> β subunits or POMP in human cell lines<br />

resulted in enhanced proteasome assembly and activities and increased cell survival following treatments with<br />

various oxidants. Moreover, stable over-expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> β5 subunit delayed senescence in human fibroblasts<br />

(J Biol Chem 280, 11840-11850, 2005). Finally in search <str<strong>on</strong>g>of</str<strong>on</strong>g> natural compounds that may activate proteasome,<br />

we have identified that the main c<strong>on</strong>stituent <str<strong>on</strong>g>of</str<strong>on</strong>g> olives, oleuropein, exerts stimulatory effects <strong>on</strong> proteasome.<br />

Importantly, c<strong>on</strong>tinuous treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> human fibroblasts cultures with oleuropein delays senescence by<br />

approximately 15%.<br />

5A_02_S<br />

CHEMICAL STRESS RESPONSE MIMETICS EXTEND LIFESPAN<br />

Gord<strong>on</strong> J. Lithgow<br />

The Buck Institute, 8001 Redwood Blvd., Novato, CA 94945, USA<br />

e-mail: glithgow@buckinstitute.org<br />

There are c<strong>on</strong>siderable mechanistic links between organismal stress resistance and aging 1 . We have shown<br />

previously that l<strong>on</strong>g-lived mutants <str<strong>on</strong>g>of</str<strong>on</strong>g> the nematode C. elegans are resistant to thermal stress and over-<br />

235


23-26 August 2007,<br />

Budapest, Hungary<br />

accumulate small heat shock proteins (shsps) which al<strong>on</strong>e can extend lifespan 2-4 . A large number <str<strong>on</strong>g>of</str<strong>on</strong>g> genes<br />

determine normal aging rate in the nematode and mutati<strong>on</strong>s in these genes tend to be highly pleiotropic<br />

including effects <strong>on</strong> organismal stress resistance and stress gene expressi<strong>on</strong>. We undertook genetic screen <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

mutati<strong>on</strong>s that c<strong>on</strong>ferred increased resistance to multiple stresses and were surprised when we tagged a gene<br />

shown in other organisms to affect checkpoint functi<strong>on</strong>s. Checkpoints are evoluti<strong>on</strong>arily c<strong>on</strong>served signal<br />

transducti<strong>on</strong> pathways that arrest cell divisi<strong>on</strong> in resp<strong>on</strong>se to DNA damage or stalled replicati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g>ks. We<br />

dem<strong>on</strong>strated that CID-1, CHK-1 or CDC-25 checkpoint proteins also determine organismal stress<br />

resistance and lifespan. This pathway and other signaling pathways have remarkable effects <strong>on</strong> lifespan; we<br />

have observed some mutati<strong>on</strong>s extend lifespan 500%.<br />

Each <str<strong>on</strong>g>of</str<strong>on</strong>g> these genetic modulators <str<strong>on</strong>g>of</str<strong>on</strong>g> aging are potential targets <str<strong>on</strong>g>for</str<strong>on</strong>g> chemical interventi<strong>on</strong>s. We will describe a<br />

series <str<strong>on</strong>g>of</str<strong>on</strong>g> screens <str<strong>on</strong>g>of</str<strong>on</strong>g> chemical compound libraries and show that chemical modulators <str<strong>on</strong>g>of</str<strong>on</strong>g> aging can be<br />

identified. These may open up new therapeutic avenues <str<strong>on</strong>g>for</str<strong>on</strong>g> age-related diseases.<br />

236<br />

5A_03_S<br />

AGE DEPENDENT COLLAPSE OF PROTEIN FOLDING HOMEOSTASIS<br />

Anat Ben-Zvi, Richard I. Morimoto<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, Molecular Biology, and Cell Biology, Rice Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Biomedical <str<strong>on</strong>g>Research</str<strong>on</strong>g>,<br />

Northwestern University, Evanst<strong>on</strong>, IL 60208, USA<br />

The balance between protein folding and degradati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g>ms the cellular protein-folding homeostasis, which<br />

can compensate <str<strong>on</strong>g>for</str<strong>on</strong>g> inherent, <str<strong>on</strong>g>of</str<strong>on</strong>g>f pathway misfolding. During the life span <str<strong>on</strong>g>of</str<strong>on</strong>g> an organism there are changes<br />

in expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular folding and degradati<strong>on</strong> comp<strong>on</strong>ents as well as an accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> damage proteins,<br />

however, the impact <str<strong>on</strong>g>of</str<strong>on</strong>g> these changes <strong>on</strong> the cellular folding homeostasis is not known. Using meta-stable<br />

proteins in C. elegans to probe <str<strong>on</strong>g>for</str<strong>on</strong>g> changes in the folding capacity, we find that there is an age dependent<br />

collapse <str<strong>on</strong>g>of</str<strong>on</strong>g> the cellular folding envir<strong>on</strong>ment. This drop in folding capacity is modified genetically, by the Hsf-<br />

1 and Daf-16 pathways promoting or postp<strong>on</strong>ing homeostasis decline. We find that both pathways c<strong>on</strong>tribute<br />

independently to protein folding homeostasis but down regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> either pathway is sufficient to disturb<br />

folding homeostasis. We suggest that the balance between misfolding loads and folding and turnover<br />

machinery determines this age dependent folding homeostasis collapse.<br />

5A_04_S<br />

MILD STRESS-INDUCED HORMESIS AND ANTI-AGING EFFECTS<br />

ON HUMAN CELLS<br />

Suresh Rattan<br />

Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Cellular Ageing, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular Biology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Aarhus, Denmark<br />

e-mail: rattan@mb.au.dk<br />

The phenomen<strong>on</strong> in which adaptive resp<strong>on</strong>ses to low doses <str<strong>on</strong>g>of</str<strong>on</strong>g> otherwise harmful c<strong>on</strong>diti<strong>on</strong>s improve the<br />

functi<strong>on</strong>al ability <str<strong>on</strong>g>of</str<strong>on</strong>g> cells and organisms is known as hormesis. Several physical, chemical and biological<br />

stressors exhibit hormetic effects, including heavy metals, pesticides, antibiotics, chemotherapeutic agents,<br />

ethanol, aldehydes, chlor<str<strong>on</strong>g>of</str<strong>on</strong>g>orm, pro-oxidants, hypergravity and i<strong>on</strong>izing radiati<strong>on</strong>. The key c<strong>on</strong>ceptual<br />

features <str<strong>on</strong>g>of</str<strong>on</strong>g> hormesis are the disrupti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> homeostasis, the modest overcompensati<strong>on</strong>, and the reestablishment<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> homeodynmaics. A critical comp<strong>on</strong>ent <str<strong>on</strong>g>of</str<strong>on</strong>g> the homeodynamic property <str<strong>on</strong>g>of</str<strong>on</strong>g> living systems is<br />

their capacity to resp<strong>on</strong>d to stress. Thermoregulati<strong>on</strong>, detoxificati<strong>on</strong>, cell proliferati<strong>on</strong>/apoptosis, DNA


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

repair, heat shock protein synthesis, protein turnover and antioxidative resp<strong>on</strong>ses are some <str<strong>on</strong>g>of</str<strong>on</strong>g> the main<br />

homeostatic resp<strong>on</strong>ses. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, it is observed that if cells and organisms are exposed to brief periods <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

mild stress so that their stress resp<strong>on</strong>se-induced gene expressi<strong>on</strong> is upregulated and the related pathways <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

maintenance and repair are stimulated, several anti-aging, health-improving and l<strong>on</strong>gevity-promoting<br />

hormetic effects occur. Extensive studies per<str<strong>on</strong>g>for</str<strong>on</strong>g>med in our labs have shown that repeated mild heat stress has<br />

anti-aging hormetic effects <strong>on</strong> various cellular and biochemical characteristics <str<strong>on</strong>g>of</str<strong>on</strong>g> human skin fibroblasts<br />

undergoing aging. These effects include the maintenance <str<strong>on</strong>g>of</str<strong>on</strong>g> chaper<strong>on</strong>e protein pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iles, reducti<strong>on</strong> in the<br />

accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidatively and glycoxidatively damaged proteins, stimulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the proteasomal activities<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> the degradati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> abnormal proteins, improved cellular resistance to ethanol, hydrogenperoxide and<br />

ultraviolet-B rays, and enhanced levels <str<strong>on</strong>g>of</str<strong>on</strong>g> various antioxidant enzymes. Further research will elucidate the<br />

mechanisms and possibilities <str<strong>on</strong>g>of</str<strong>on</strong>g> human applicati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> physical and mental stress as a beneficial challenge.<br />

5A_05_S<br />

(poster secti<strong>on</strong> B1, poster board #179, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

MITOCHONDRIAL J HAPLOTYPE AND ANTI-OXIDANT STATUS AND IN THE<br />

BELFAST ELDERLY LONGITUDINAL FREE-LIVING AGEING STUDY (BELFAST)<br />

Irene Maeve Rea 1 , Owen A. Ross 2, 5 , G Pooler Archbold 3 , Ian R. Young 4 , Martin Curran 2 ,<br />

Derek Middlet<strong>on</strong> 2<br />

1 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Geriatric Medicine, Queens University Belfast, 2 Northern Ireland Tissue Typing and Immunogenetics<br />

Laboratory, Belfast City Hospital, 3 Biochemistry Laboratory, Belfast City Hospital, 4 Nutriti<strong>on</strong> and Metabolism<br />

Group, Queens University Belfast, 5 Mayo Clinic, Jacks<strong>on</strong>ville, USA<br />

Mitoch<strong>on</strong>dria are the chief source <str<strong>on</strong>g>of</str<strong>on</strong>g> both energy and oxidants inside the cell. This dual role can damage cells<br />

despite an array <str<strong>on</strong>g>of</str<strong>on</strong>g> antioxidants available to mop up endogenous oxidants including hydrogen peroxide,<br />

superoxide and hydroxyl radicals. The mitoch<strong>on</strong>drial haplotype J seems related to ‘successful’ ageing in<br />

Italian, Finnish and Irish groups <str<strong>on</strong>g>of</str<strong>on</strong>g> n<strong>on</strong>agenarian/centenarians. In this study we questi<strong>on</strong>ed whether<br />

J haplotype octo/n<strong>on</strong>agenarians from the BELFAST study dem<strong>on</strong>strated enhanced anti-oxidant pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ile or<br />

other phenotype characteristics.<br />

Methods: Briefly, community-living, mentally alert (Folstein >27/30) octo/n<strong>on</strong>agenarian subjects were<br />

enlisted as part <str<strong>on</strong>g>of</str<strong>on</strong>g> BELFAST study. DNA typing <str<strong>on</strong>g>for</str<strong>on</strong>g> mitoch<strong>on</strong>drial haplotypes was collected and blood <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

serum antioxidants together with Blood pressure measurements. Serum uric acid, bilirubin, ceruloplasmin,<br />

glutathi<strong>on</strong>e, selenium and vitamins C, A, E and α-carotene were not significantly different between J and n<strong>on</strong><br />

J mitoch<strong>on</strong>drial haplotypes but whole blood and serum gluthati<strong>on</strong>e peroxidase (GSHpx) were lower in J<br />

haplotype octo/n<strong>on</strong>agenarians. Both systolic and diastolic blood pressure was significantly lower in J<br />

haplotype octo/n<strong>on</strong>agenarians.<br />

C<strong>on</strong>clusi<strong>on</strong>: BELFAST study octo/n<strong>on</strong>agenarian subjects, carrying the J haplotype, do not show enhanced<br />

antioxidant status <str<strong>on</strong>g>for</str<strong>on</strong>g> most <str<strong>on</strong>g>of</str<strong>on</strong>g> the major antioxidants, including uric acid and vitamin C, but show reduced<br />

whole blood and serum glutathi<strong>on</strong>e peroxidase status. Interestingly J mitoch<strong>on</strong>drial haplotype<br />

octo/n<strong>on</strong>agenarians did have lower systolic and diastolic blood pressure.<br />

237


23-26 August 2007,<br />

Budapest, Hungary<br />

5A_06_S<br />

(poster secti<strong>on</strong> B1, poster board #180, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

INFLUENCE OF AGE AND ETHANOL INTOXICATION ON HEME OXYGENASE 1<br />

EXPRESSION IN RAT LIVER<br />

Anna Lisa Furfaro, Stefania Patriarca, Emanuela Balbis, Cinzia Domenicotti, Damiano Cottalasso,<br />

Umberto Maria Marinari, Maria Adelaide Pr<strong>on</strong>zato, Nicola Traverso<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Experimental Medicine, Secti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> General Pathology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Genova, Via L. B. Alberti 2,<br />

16132, Genova, Italy<br />

e-mail:annalisa.furfaro@virgilio.it<br />

Many authors recognize that heme oxygenase 1 (HO-1) expressi<strong>on</strong> is a marker <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular resp<strong>on</strong>se to<br />

oxidative stress; since ageing is related to oxidative “wear and tear”, HO-1 may represent a candidate<br />

biomarker <str<strong>on</strong>g>of</str<strong>on</strong>g> ageing. In this study, we evaluated the hepatic expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HO-1 mRNA in 2.5-24 m<strong>on</strong>th-old<br />

rats; this expressi<strong>on</strong> was higher at 6 m<strong>on</strong>ths than at 2.5 m<strong>on</strong>ths <str<strong>on</strong>g>of</str<strong>on</strong>g> age, but thereafter increased no further.<br />

However, while 2.5 and 6-m<strong>on</strong>th-old rats resp<strong>on</strong>ded to acute ethanol intoxicati<strong>on</strong> by displaying increased<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> liver HO-1 mRNA, 18 m<strong>on</strong>th-old rats did not show any resp<strong>on</strong>se; this phenomen<strong>on</strong> suggests<br />

that during development and ageing the transcripti<strong>on</strong>al resp<strong>on</strong>se to oxidative stress is progressively impaired.<br />

This may be due to decreased transcripti<strong>on</strong>al ability to resp<strong>on</strong>d to stress in older animals, rather than by a<br />

reducti<strong>on</strong> in oxidative stress.<br />

238


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5A_01_P<br />

(poster secti<strong>on</strong> B1, poster board #181, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

AISA ("ANTI INFLAMMATORY SENESCENCE ACTIVES")<br />

A PLANT DERIVED APPROACH OF ANTI-STRESS<br />

Jean-François Biss<strong>on</strong>, Chantal Menut, Marc Moutet, Patrizia d’Alessio<br />

ETAP-54500 Vandoeuvre-les-Nancy; Laboratoire de Chimie Bio-Moléculaire-UMR 5032 ENSCM, 34296<br />

M<strong>on</strong>tpellier; EandC Biocitech France; AISA Therapeutics CHU Paul Brousse<br />

12, ave Paul Vaillant-Couturier 94807 Villejuif France<br />

e-mail: dalessio@vjf.inserm.fr<br />

Psycho-social stress induces arterial vasomotor alterati<strong>on</strong>s such as vasospams and transitory hypertensi<strong>on</strong>.<br />

Moreover, stress has a pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ound impact <strong>on</strong> vascular endothelium, inducing a systemic inflammati<strong>on</strong>. Starting<br />

from plant extracts, we have isolated through bio-guided selecti<strong>on</strong>, characterized and patented 4 m<strong>on</strong>oterpens,<br />

that we have named "AISA", Anti-Inflammatory Senescence Actives, targeting activated<br />

endothelium. In our in vitro plat<str<strong>on</strong>g>for</str<strong>on</strong>g>m, they inhibit the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> adhesi<strong>on</strong> molecules and the<br />

polymerizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> actin fibers, following the activati<strong>on</strong> by TNF-α. Inhibiti<strong>on</strong> is c<strong>on</strong>served at all replicative<br />

passages <str<strong>on</strong>g>of</str<strong>on</strong>g> vascular endothelial cells. In pre-clinical models, AISA 5203-L is well tolerated and has a good<br />

oral bio-availability. This molecule has shown c<strong>on</strong>vincing anti-inflammatory per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance in a rat TNBS<br />

colitis model, comparable to those obtained with ibupr<str<strong>on</strong>g>of</str<strong>on</strong>g>en. Using AISA-5203-L in a stress model, a<br />

substantial anti-stress activity could be documented by a FOB (Functi<strong>on</strong>al Observati<strong>on</strong> Battery) <str<strong>on</strong>g>of</str<strong>on</strong>g> 56 tests<br />

(patent pending).<br />

We would like to propose that a new anti-inflammatory molecule <str<strong>on</strong>g>of</str<strong>on</strong>g> plant origin interfering with cell<br />

replicative senescence would be resp<strong>on</strong>sible <str<strong>on</strong>g>of</str<strong>on</strong>g> relevant anti-stress effects. This hypothesis would permit the<br />

positi<strong>on</strong>ing <str<strong>on</strong>g>of</str<strong>on</strong>g> AISA molecules as candidates therapeutic applicati<strong>on</strong>s in multi-factorial diseases, such as<br />

hypertensi<strong>on</strong>, neuro-degenerative diseases, chr<strong>on</strong>ic pain syndrome, obesity and depressi<strong>on</strong>.<br />

5A_02_P<br />

(poster secti<strong>on</strong> B1, poster board #182, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

INTERACTION OF HSP-16.2 WITH HISTONES AFTER HEAT-SHOCK IN THE<br />

NEMATODE C. ELEGANS<br />

S. Alavez, G. J. Lithgow<br />

The Buck Institute, 8001 Redwood Blvd. Novato, CA 94945 USA. e-mail: salavez@buckinstitute.org<br />

Stress resp<strong>on</strong>se is a very significant determinate <str<strong>on</strong>g>of</str<strong>on</strong>g> the life history <str<strong>on</strong>g>of</str<strong>on</strong>g> C. elegans. As the first metazoan genome<br />

to be sequenced, C. elegans is a major model <str<strong>on</strong>g>for</str<strong>on</strong>g> molecular genetic studies <str<strong>on</strong>g>of</str<strong>on</strong>g> aging because <str<strong>on</strong>g>of</str<strong>on</strong>g> the availability<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> a range <str<strong>on</strong>g>of</str<strong>on</strong>g> single gene mutant lines with greatly extended lifespan. Interestingly, these l<strong>on</strong>g-lived variants<br />

are also stress tolerant. Am<strong>on</strong>g others, we dem<strong>on</strong>strated that at least <strong>on</strong>e chaper<strong>on</strong>e gene encoding heat<br />

shock protein -16 (hsp-16) works together with reduced insulin signaling to bring about lifespan extensi<strong>on</strong>.<br />

However, the mechanism by which hsp-16 increases lifespan or thermotolerance in this animal model<br />

remains unclear.<br />

We have now obtained an stable transgenic line that over-express the fusi<strong>on</strong> protein HSP-16.2::GFP under<br />

the c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp-16.2 promoter that presents a remarkable thermotolerance and a high expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

fusi<strong>on</strong> protein after heat-shock (HS) measured by westernblot. The lifespan <str<strong>on</strong>g>of</str<strong>on</strong>g> these over-expressing lines is<br />

extended (36%, p


23-26 August 2007,<br />

Budapest, Hungary<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> this transgene in multiple tissues. We have found a relocalizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP-16.2::GFP to the<br />

nuclei <str<strong>on</strong>g>of</str<strong>on</strong>g> several neur<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> the nerve ring and some intestinal cells after HS in this strain as revealed by<br />

DAPI and GFP colocalizati<strong>on</strong>. On line with this observati<strong>on</strong>, we have identified Hist<strong>on</strong>e-3 and Hist<strong>on</strong>e-4<br />

through MALDI-TOF and LC/MS mass spectrometry by coimmunoprecipitati<strong>on</strong> with an HSP-16.2 specific<br />

antibody and posterior separati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the proteins via 1-dimensi<strong>on</strong>al gel electrophoresis after HS.<br />

These results show that hist<strong>on</strong>es are targets <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP-16.2 during stress c<strong>on</strong>diti<strong>on</strong>s (HS) and suggest a nuclear<br />

role <str<strong>on</strong>g>of</str<strong>on</strong>g> chaper<strong>on</strong>es to regulate thermotolerance and maybe other stressing c<strong>on</strong>diti<strong>on</strong>s like aging.<br />

5A_03_P<br />

(poster secti<strong>on</strong> B1, poster board #183, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

LIPOIC ACID: HOW AN INCREASE IN RESPIRATION CAN DECREASE STRESS<br />

AND INCREASE LIFESPAN IN C. ELEGANS<br />

Olivier Descamps, Michael Benedetti, Amanda L. Foster, Maithili Vantipalli, James N. Sampayo,<br />

Matthew S. Gill, Anders Olsen, Gord<strong>on</strong> J. Lithgow<br />

Buck Institute, 8001 Redwood Blvd, 94945 Novato, USA, e-mail: odescamp@buckinstitute.org<br />

Aging is the most important risk factor in developed countries. C<strong>on</strong>sequently, there is c<strong>on</strong>siderable interest in<br />

the pharmacological manipulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> aging as a way to uncover new avenues <str<strong>on</strong>g>for</str<strong>on</strong>g> development <str<strong>on</strong>g>of</str<strong>on</strong>g> therapeutics.<br />

Genetic alterati<strong>on</strong>s can extend the lifespan <str<strong>on</strong>g>of</str<strong>on</strong>g> the nematode C. elegans as much as five-fold and in principle<br />

chemical compounds could do the same. Based <strong>on</strong> the mechanistic relati<strong>on</strong>ship between aging and stress<br />

resp<strong>on</strong>se, we have screened <str<strong>on</strong>g>for</str<strong>on</strong>g> compounds that enhance stress resistance. Lipoic acid (LA), a c<str<strong>on</strong>g>of</str<strong>on</strong>g>actor <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

Krebs cycle enzymes pyruvate and glyceraldehyde dehydrogenases, increases respirati<strong>on</strong> and lifespan in the<br />

adult nematode worm C. elegans. This seems somewhat paradoxical in the c<strong>on</strong>text <str<strong>on</strong>g>of</str<strong>on</strong>g> the rate <str<strong>on</strong>g>of</str<strong>on</strong>g> living theory<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> aging in which lifespan is inversely proporti<strong>on</strong>al to the rates <str<strong>on</strong>g>of</str<strong>on</strong>g> metabolic processes. We investigated the<br />

effect <str<strong>on</strong>g>of</str<strong>on</strong>g> the LA treatment <strong>on</strong> the mitoch<strong>on</strong>drial physiology <str<strong>on</strong>g>of</str<strong>on</strong>g> C. elegans. We find that lipoic acid decreases<br />

the mitoch<strong>on</strong>drial membrane potential and see some evidence <str<strong>on</strong>g>for</str<strong>on</strong>g> an increase in the amount <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>dria.<br />

This in turn results in lower oxygen radical producti<strong>on</strong> (steady state superoxide ani<strong>on</strong> levels), which may<br />

explain the lifespan extensi<strong>on</strong>. The LA-treated worms are not overtly hyperactive and have the same levels <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

ATP and fat c<strong>on</strong>tent as untreated c<strong>on</strong>trols. Our working hypothesis is that stimulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the Krebs cycle,<br />

without changing the other metabolic parameters, increases the number <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>dria and in c<strong>on</strong>sequence<br />

lowers the burden <str<strong>on</strong>g>of</str<strong>on</strong>g> activity in each <str<strong>on</strong>g>of</str<strong>on</strong>g> the mitoch<strong>on</strong>dria, resulting in less stress and l<strong>on</strong>ger lifespan.<br />

240


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5A_04_P<br />

(poster secti<strong>on</strong> B1, poster board #184, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EARLY STRESS RESPONSE IN FXTAS AN INHERITED NEURODEGENERATIVE<br />

DISORDER<br />

D. Garcia-Arocena 1 , C. K. Iwahashi 1 , F. Tass<strong>on</strong>e 1 , J. B. Brower 2 , E. M. Berry-Kravis 3 ,<br />

R. Willemsen 2 , P. J. Hagerman 1<br />

1 Dept. Biochemistry and Molecular Medicine, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Cali<str<strong>on</strong>g>for</str<strong>on</strong>g>nia Davis, School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, One Shields Ave,<br />

Davis, 95616, USA<br />

2 Dept. Clinical Genetics, Erasmus University, Rotterdam, The Netherlands.<br />

3 Neurology and Biochemistry, Rush University Medical Center, Chicago, Illinois, USA<br />

Fragile X-associated tremor/ataxia syndrome (FXTAS) is a late-<strong>on</strong>set neurodegenerative disorder that<br />

appears to affect carriers <str<strong>on</strong>g>of</str<strong>on</strong>g> premutati<strong>on</strong> alleles (55 to 200 CGG repeats) <str<strong>on</strong>g>of</str<strong>on</strong>g> the FMR1 gene. The<br />

pathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> FXTAS involves the direct toxic effect <str<strong>on</strong>g>of</str<strong>on</strong>g> the elevated levels <str<strong>on</strong>g>of</str<strong>on</strong>g> expanded CGG repeat FMR1<br />

mRNA. We have recently described neural cell models that are capable <str<strong>on</strong>g>of</str<strong>on</strong>g> recapitulating the <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

intranuclear inclusi<strong>on</strong>s, the pathologic hallmark <str<strong>on</strong>g>of</str<strong>on</strong>g> FXTAS. We have discovered that several proteins<br />

including lamin A/C and the stress resp<strong>on</strong>se proteins αB-crystallin and Hsp27 genes are dysregulated in<br />

resp<strong>on</strong>se to expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the expanded CGG repeat. To quantify these observati<strong>on</strong>s we analyzed the<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> FMR1, αB-crystallin, Hsp27, and lamin A/C by real time quantitative PCR in cultured human<br />

cells derived from subjects with severe FXTAS and age-matched c<strong>on</strong>trols. We also studied cultured cells<br />

derived from mice with expanded CGG repeats and wt c<strong>on</strong>trols. Our results show that subjects with<br />

premutati<strong>on</strong>s have elevated levels <str<strong>on</strong>g>of</str<strong>on</strong>g> expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> stress resp<strong>on</strong>se genes and cellular redistributi<strong>on</strong> in cultured<br />

cells. Immun<str<strong>on</strong>g>of</str<strong>on</strong>g>luorescent studies <str<strong>on</strong>g>of</str<strong>on</strong>g> additi<strong>on</strong>al proteins and flow cytometry DNA analysis indicate cell cycle<br />

dysregulati<strong>on</strong> in cells with expanded CGG repeats c<strong>on</strong>sistent with accelerated aging in both mouse and<br />

human models. We discuss the implicati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> these results <strong>on</strong> neural plasticity.<br />

5A_05_P<br />

(poster secti<strong>on</strong> B1, poster board #185, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DIURNAL AND AGE CHANGES IN STRESS RESPONSIVENESS OF THE<br />

HYPOTHALAMIC-PITUITARY-ADRENAL (HPA) AXIS AND ERYTHROCYTE<br />

ANTIOXIDANT ENZYMES<br />

N. D. G<strong>on</strong>charova 1 , T. N. Bogatyrenko 2<br />

1Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Primatology, Sochi-Adler, Veseloye 1, 354376, Russia; 2 Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Problems <str<strong>on</strong>g>of</str<strong>on</strong>g> Chemical<br />

Physics, Chernogolovka, Russia<br />

The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> the study was to examine chr<strong>on</strong>obiological characteristics <str<strong>on</strong>g>of</str<strong>on</strong>g> the HPA axis functi<strong>on</strong>ing and the<br />

antioxidant enzyme activities in stress and aging. Female Macaca mulatta <str<strong>on</strong>g>of</str<strong>on</strong>g> 6-8 years (young mature) and 20-27<br />

years (old) were subjected to acute psycho-emoti<strong>on</strong>al stress (two hours immobilizati<strong>on</strong>) at 9.00 or 15.00 h.<br />

Levels <str<strong>on</strong>g>of</str<strong>on</strong>g> ACTH, cortisol (F), dehydroepiandroster<strong>on</strong>e sulfate (DHEAS), and testoster<strong>on</strong>e (T) in peripheral<br />

blood plasma were measured be<str<strong>on</strong>g>for</str<strong>on</strong>g>e the stress and 5, 15, 30, 60, 120, 240 min and 24 h after the challenge. In<br />

parallel, activities <str<strong>on</strong>g>of</str<strong>on</strong>g> superoxide dismutase (SOD), glutathi<strong>on</strong>e peroxidase, glutathi<strong>on</strong>e reductase (GR),<br />

glutathi<strong>on</strong>e-S-transferase, and lipid peroxidati<strong>on</strong> (LPO) were measured in hemolyzed erythrocytes. Young<br />

m<strong>on</strong>keys dem<strong>on</strong>strated much higher increase <str<strong>on</strong>g>of</str<strong>on</strong>g> ACTH, F, T, DHEAS levels and GR activity in resp<strong>on</strong>se to<br />

241


23-26 August 2007,<br />

Budapest, Hungary<br />

the stress imposed at 15.00 than to identical stress imposed at 9.00. However, no such circadian differences in<br />

dynamics <str<strong>on</strong>g>of</str<strong>on</strong>g> the horm<strong>on</strong>al secreti<strong>on</strong> and GR activity after the stress were found <str<strong>on</strong>g>for</str<strong>on</strong>g> old animals. Young<br />

m<strong>on</strong>keys dem<strong>on</strong>strated also much higher accreti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ACTH, F, T, DHEAS levels and GR activity in<br />

comparis<strong>on</strong> with old m<strong>on</strong>keys in resp<strong>on</strong>se to the afterno<strong>on</strong> stress. The changes in GR activity with stress and<br />

aging correlated well with the changes in the F, DHEAS, and T level. SOD activity in old m<strong>on</strong>keys was lower<br />

than in young <strong>on</strong>es be<str<strong>on</strong>g>for</str<strong>on</strong>g>e the stress and increased in resp<strong>on</strong>se to the stress that was accompanied by light<br />

decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> LPO. In c<strong>on</strong>trast to old m<strong>on</strong>keys, young <strong>on</strong>es dem<strong>on</strong>strated decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> SOD activity and some<br />

increase <str<strong>on</strong>g>of</str<strong>on</strong>g> LPO with stress. There was a high level <str<strong>on</strong>g>of</str<strong>on</strong>g> correlati<strong>on</strong> between the stress changes in SOD activity<br />

and the changes in the F level in old m<strong>on</strong>keys but there was no correlati<strong>on</strong> in young m<strong>on</strong>keys. The lack <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

correlati<strong>on</strong> between stress dynamics <str<strong>on</strong>g>of</str<strong>on</strong>g> F level and SOD activity may be caused by marked activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

adrenal T secreti<strong>on</strong> in young m<strong>on</strong>keys. These results suggest that the HPA axis plays an important role in<br />

regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> antioxidant enzymes defense in stress c<strong>on</strong>diti<strong>on</strong>s and that this regulati<strong>on</strong> shows age differences.<br />

5A_06_P<br />

(poster secti<strong>on</strong> B1, poster board #186, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

POTENTIAL USE OF PLANT ADAPTOGENS IN AGE-RELATED DISORDERS<br />

1V. Makarov, M. N. Makarova, N. V. Stoloschyck, 2 G. Wikman, A. Panossian<br />

1Interregi<strong>on</strong>al Center “Adaptogen”, Piskarevsky ave., 47/5, St-Petersburg, 195067, Russia<br />

2Swedish Herbal Institute, Prinsgatan 12 5tr, SE-413 05 GÖTEBORG, Sweden<br />

In traditi<strong>on</strong>al medicinal systems, members <str<strong>on</strong>g>of</str<strong>on</strong>g> a small group <str<strong>on</strong>g>of</str<strong>on</strong>g> higher plants were c<strong>on</strong>sidered to possess<br />

restorative properties and were used as general t<strong>on</strong>ics in the treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> disease and in c<strong>on</strong>valescence.<br />

Through a series <str<strong>on</strong>g>of</str<strong>on</strong>g> studies c<strong>on</strong>ducted in the early 1950s, Soviet scientists first established that several <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

these plants also possessed the capacity to enhance the “state <str<strong>on</strong>g>of</str<strong>on</strong>g> n<strong>on</strong>-specific resistance” (SNSR) <str<strong>on</strong>g>of</str<strong>on</strong>g> an<br />

organism to stress. Plants that exhibit this “adaptogenic effect” are currently used to increase mental and<br />

physical work capacity and per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance against a background <str<strong>on</strong>g>of</str<strong>on</strong>g> fatigue and stress, and also to improve the<br />

quality <str<strong>on</strong>g>of</str<strong>on</strong>g> life. Additi<strong>on</strong>ally, the efficacy <str<strong>on</strong>g>of</str<strong>on</strong>g> plant adaptogens in the relief <str<strong>on</strong>g>of</str<strong>on</strong>g> mild and moderate depressi<strong>on</strong> has<br />

recently been dem<strong>on</strong>strated. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> the present study was to investigate the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> adaptogens,<br />

administered in the <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> standardised extracts <str<strong>on</strong>g>of</str<strong>on</strong>g> Rhodiola rosea, Schisandra chinensis and Eleutherococcus<br />

senticosus in their fixed combinati<strong>on</strong>s (ADAPT-232, ADAPT-Plus and ADAPT-Extra-Plus), <strong>on</strong> the age-related<br />

deteriorati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the innate defence, cardiovascular and central nervous systems in 2 year old rats.<br />

Drugs and placebo were administered daily over a period <str<strong>on</strong>g>of</str<strong>on</strong>g> 4 m<strong>on</strong>ths, and alterati<strong>on</strong>s in adrenal weight,<br />

ECG parameters, motor activity, learning ability, durati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> hexanal-induced sleep, <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

immobilisati<strong>on</strong> stress-induced stomach ulcers, sp<strong>on</strong>taneous promoti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> tumours, apoptosis <str<strong>on</strong>g>of</str<strong>on</strong>g> spleen<br />

lymphocytes, and levels <str<strong>on</strong>g>of</str<strong>on</strong>g> 17- hydroxy-corticosteroids in the urine, and cholesterol, albumins and total<br />

proteins in the blood, were m<strong>on</strong>itored. The results showed that repeated administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> adaptogens can<br />

diminish or prevent a range <str<strong>on</strong>g>of</str<strong>on</strong>g> age-related disorders including reduced liver detoxifying functi<strong>on</strong>, malfuncti<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the central nervous system (i.e. loss <str<strong>on</strong>g>of</str<strong>on</strong>g> memory and learning ability), development and progressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

cardiac insufficiency and hypercholesterolemia, impaired protein synthesis, reduced activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the horm<strong>on</strong>al<br />

system, increased sensitivity to stress (hypodynamia-induced damage to the stomach and adrenals), impaired<br />

apoptosis, and sp<strong>on</strong>taneous promoti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> tumours. It is c<strong>on</strong>cluded that adaptogens have potential value in<br />

the treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> age-related disorders <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress system in the elderly and may be effective in maintaining<br />

the health status <str<strong>on</strong>g>of</str<strong>on</strong>g> such individuals at the normal level.<br />

242


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5A_07_P<br />

(poster secti<strong>on</strong> B1, poster board #187, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

TISSUE AND CELLULAR RESPONSES TO STRESSES OF HYPOXIA AND<br />

HYPEROXIA OF IMPORTANCE IN TISSUE ENGINEERING<br />

George K. B. Sándor, Riitta Suur<strong>on</strong>en<br />

Regea Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Regenerative Medicine, Tampere University, Tampere, Finland and The Hospital <str<strong>on</strong>g>for</str<strong>on</strong>g> Sick<br />

Children, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Tor<strong>on</strong>to, Tor<strong>on</strong>to, Canada<br />

e-mail: george.sandor@utor<strong>on</strong>to.ca<br />

Future rec<strong>on</strong>structive surgical procedures will have the possibility <str<strong>on</strong>g>of</str<strong>on</strong>g> using tissue engineered c<strong>on</strong>structs to<br />

rebuild missing or defective body parts in humans and in veterinary medicine. Such c<strong>on</strong>structs require three<br />

essential comp<strong>on</strong>ents: cells, a scaffold and signalling molecules. A great deal <str<strong>on</strong>g>of</str<strong>on</strong>g> attenti<strong>on</strong> has been paid to the<br />

harvesting <str<strong>on</strong>g>of</str<strong>on</strong>g> stem-cells both <str<strong>on</strong>g>of</str<strong>on</strong>g> fetal and autogenous adult origins which are to be expanded in tissue culture.<br />

Such expanded stem-cell derrived populati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> cells are used to populate resorbable scaffolds. Much work<br />

has been d<strong>on</strong>e in understanding the potential sources and roles <str<strong>on</strong>g>of</str<strong>on</strong>g> key bioactive cell signalling molecules such<br />

as B<strong>on</strong>e Morphogenetic Protein (BMP). Cells that exist in isolati<strong>on</strong> or as parts <str<strong>on</strong>g>of</str<strong>on</strong>g> complex organs and tissues<br />

have known resp<strong>on</strong>ses to certain stressors. Cells and tissues that are grown ex vivo in extra corporeal<br />

bioreactors must be provided with envir<strong>on</strong>ments that minimize cellular stress and promote maximal growth<br />

when cellular populati<strong>on</strong>s are to be expanded. These cellular c<strong>on</strong>structs are sensitive to varying<br />

c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> ascorbic acid, dexamethas<strong>on</strong>e and β-glycerophosphate. The reacti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> cells and tissues to<br />

hypoxic and hyperoxic envir<strong>on</strong>ments are also important. At the molecular level these reacti<strong>on</strong>s occur in part<br />

due to superoxide radicals while at the tissue level they are mediated by bioactive signalling proteins such as<br />

Vascular Endothelial Growth Factor (VEGF). There is evidence that both hypoxia and hyperoxic states may<br />

promote angiogenesis and further tissue growth. This presentati<strong>on</strong> will review our understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> these<br />

processes and mechanisms in the c<strong>on</strong>text <str<strong>on</strong>g>of</str<strong>on</strong>g> tissue engineering.<br />

5A_08_P<br />

(poster secti<strong>on</strong> B1, poster board #188, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CREB-INDUCING ACTION OF HYPERICUM PERFORATUM MAY BE<br />

RESPONSIBLE FOR ITS PREVENTIVE EFFECTS ON STRESS- AND AGING-<br />

RELATED MEMORY IMPAIRMENT<br />

Emil Tr<str<strong>on</strong>g>of</str<strong>on</strong>g>imiuk, Adam Hołownia, Jan J. Braszko<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Clinical Pharmacology, Medical University <str<strong>on</strong>g>of</str<strong>on</strong>g> Bialystok,<br />

Waszyngt<strong>on</strong>a 15A, 15-274 Bialystok, Poland<br />

Chr<strong>on</strong>ic stress impairs a number <str<strong>on</strong>g>of</str<strong>on</strong>g> aspects <str<strong>on</strong>g>of</str<strong>on</strong>g> cogniti<strong>on</strong> such as acquisiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> memory, its c<strong>on</strong>solidati<strong>on</strong> and<br />

recall. Similar alterati<strong>on</strong>s were observed in aged rats. It is well known that stress causes accelerati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

brain aging. In stressed as well as in aged rats disturbances <str<strong>on</strong>g>of</str<strong>on</strong>g> hypothalamo-pituitary-adrenal axis and<br />

c<strong>on</strong>secutive hypercortisolemia are seen. Excessive cortisolemia leads to a number <str<strong>on</strong>g>of</str<strong>on</strong>g> neurochemical and<br />

neuroanatomical changes in brain, especially in the hippocampus, which is particularly vulnerable because <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

high density <str<strong>on</strong>g>of</str<strong>on</strong>g> glucocorticoid receptors. As a result neurogenesis is disturbed in the hippocampal areas CA1<br />

and CA3 and the neur<strong>on</strong>al atrophy occurs leading to the decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> its volume and lowering total number <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

neur<strong>on</strong>s and their ramificati<strong>on</strong>s.<br />

243


23-26 August 2007,<br />

Budapest, Hungary<br />

We recently found, that dried crude herb <str<strong>on</strong>g>of</str<strong>on</strong>g> H. per<str<strong>on</strong>g>for</str<strong>on</strong>g>atum (350 mg kg -1 <str<strong>on</strong>g>for</str<strong>on</strong>g> 21 days orally) reversed negative<br />

effects <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic stress <strong>on</strong> cogniti<strong>on</strong> (spatial reference and working memory, recogniti<strong>on</strong> memory,<br />

c<strong>on</strong>diti<strong>on</strong>ed behaviour, learning, acquisiti<strong>on</strong> and recall <str<strong>on</strong>g>of</str<strong>on</strong>g> the avoidance behaviour), but still our knowledge<br />

about the mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> this improvement is poor. It is known that H. per<str<strong>on</strong>g>for</str<strong>on</strong>g>atum normalizes the<br />

dopaminergic and noradrenergic transmissi<strong>on</strong> in medial prefr<strong>on</strong>tal cortex, normalizes decreased by stress 5-<br />

HT1A and 5-HT2 receptors, balance exaggerated NMDA receptor functi<strong>on</strong>, and also restores normal<br />

cortisolemia.<br />

This study assessed an associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the H. per<str<strong>on</strong>g>for</str<strong>on</strong>g>atum improvement <str<strong>on</strong>g>of</str<strong>on</strong>g> age-related memory impairments with<br />

the increase <str<strong>on</strong>g>of</str<strong>on</strong>g> CREB and phosphorylated CREB in hippocampus. Middle-aged rats (78-weeks old) displayed<br />

clear-out decline in the acquisiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> spatial working memory in the Morris water maze similar that in the<br />

young rats (8-weeks old) after 21-day restraint stress. Chr<strong>on</strong>ic administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> H. per<str<strong>on</strong>g>for</str<strong>on</strong>g>atum effectively<br />

prevented these negative effects <str<strong>on</strong>g>of</str<strong>on</strong>g> aging and stress (p


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5A_10_P<br />

(poster secti<strong>on</strong> B1, poster board #190, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SPECIFIC DOWNREGULATION OF HSP72 OR HSP27 IN CANCER CELLS<br />

SUPPRESSES THE DNA DAMAGE RESPONSE<br />

Julia Yaglom<br />

Assistant Pr<str<strong>on</strong>g>of</str<strong>on</strong>g>essor, Bost<strong>on</strong> University Medical Center, 715 Albany St., Bost<strong>on</strong>, MA, 02118<br />

e-mail: Yaglom@biochem.bumc.bu.edu<br />

Various cancer cell lines and a large fracti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> biopsies are characterized by high expressi<strong>on</strong> levels <str<strong>on</strong>g>of</str<strong>on</strong>g> heat<br />

shock proteins, Hsp72 and Hsp27. Furthermore, high expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp72 and Hsp27 in human tumors<br />

correlates with invasiveness, metastasis, resistance to chemotherapy, and poor prognosis <str<strong>on</strong>g>of</str<strong>on</strong>g> the disease.<br />

Specific down regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp72 or Hsp27 using retrovirus-based delivery <str<strong>on</strong>g>of</str<strong>on</strong>g> shRNA, results in cell’s<br />

transiti<strong>on</strong> into a state where ~30%-40% <str<strong>on</strong>g>of</str<strong>on</strong>g> cells became senescent while the rest <str<strong>on</strong>g>of</str<strong>on</strong>g> the populati<strong>on</strong> c<strong>on</strong>tinued<br />

to divide, though with significantly reduced rates (pre-senescent cells).<br />

Noteworthy, pre-senescent cells became extremely sensitive to DNA damaging drugs and stresses. Further<br />

investigati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> this phenomen<strong>on</strong> led us to an unexpected discovery that inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> pre-senescent state was<br />

accompanied by severe defect in cellular DNA damage resp<strong>on</strong>se (DDR). Accordingly, Hsp72 or Hsp27-<br />

depleted cells when challenged with DNA damaging stress, like UVC irradiati<strong>on</strong>, failed to phosphorylate<br />

hist<strong>on</strong>e H2A.X, which is an essential step required <str<strong>on</strong>g>for</str<strong>on</strong>g> an efficient DNA repair. Moreover, activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a<br />

checkpoint kinase Chk1 was also compromised.<br />

These results suggest that triggering pre-senescence by Hsps downregulati<strong>on</strong> may become a novel interesting<br />

approach towards sensitizing cancer cells to chemotherapy and overcoming drug resistance.<br />

245


23-26 August 2007,<br />

Budapest, Hungary<br />

246<br />

5B. HSP-S IN CEREBRAL PROTECTION<br />

(IAN R. BROWN)<br />

5B_01_S<br />

HEAT SHOCK PROTEINS AND PROTECTIVE EFFECTS IN THE NERVOUS<br />

SYSTEM<br />

Ian R. Brown, Sheng Chen, Ari Chow<br />

Center <str<strong>on</strong>g>for</str<strong>on</strong>g> the Neurobiology <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Tor<strong>on</strong>to at Scarborough<br />

Tor<strong>on</strong>to, Ontario, Canada M1C1A4<br />

Hsp70 is a multi-gene family composed <str<strong>on</strong>g>of</str<strong>on</strong>g> stress-inducible members (Hsp70) and other members that are<br />

c<strong>on</strong>stitutively expressed (Hsc70). The heat shock proteins Hsp70 and Hsc70 exhibit similar molecular<br />

structure and biochemical functi<strong>on</strong>s. C<strong>on</strong>stitutively expressed Hsc70 is enriched in the mammalian nervous<br />

system compared with n<strong>on</strong>-neural tissues and present at high levels in neur<strong>on</strong>al cell bodies. After thermal<br />

stress, Hsc70 is translocated to synapse-enriched areas <str<strong>on</strong>g>of</str<strong>on</strong>g> the cerebral cortex where it associates with Hsp40<br />

to <str<strong>on</strong>g>for</str<strong>on</strong>g>m a complex that can refold denaturated proteins. These results suggest that the heat shock resp<strong>on</strong>se in<br />

the nervous system involves not <strong>on</strong>ly the synthesis <str<strong>on</strong>g>of</str<strong>on</strong>g> stress-inducible Hsps but also translocati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

c<strong>on</strong>stitutively expressed Hsc70 to synapse-enriched areas where it could participate in neuroprotective<br />

mechanism that preserve synaptic functi<strong>on</strong> during times <str<strong>on</strong>g>of</str<strong>on</strong>g> stress. Neurodegenerative disorders such as<br />

Alzheimer's disease, Parkins<strong>on</strong>'s disease and ALS have been termed 'protein misfolding disorders'. These<br />

diseases differ widely in frequency and impact different classes <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>s. Hsps provide a line <str<strong>on</strong>g>of</str<strong>on</strong>g> defense<br />

against misfolded, aggregati<strong>on</strong>-pr<strong>on</strong>e proteins and are am<strong>on</strong>g the most potent suppressors <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

neurodegenerati<strong>on</strong> in animal models. Analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> c<strong>on</strong>stitutively expressed heat shock proteins revealed<br />

variable levels <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsc70 and Hsp27 in different classes <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>s in the adult brain. The differing levels <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

these c<strong>on</strong>stitutively expressed heat shock proteins in neur<strong>on</strong>al cell populati<strong>on</strong>s may c<strong>on</strong>fer a variable<br />

buffering capacity against protein misfolding disorders that correlates with the relative frequencies <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

previously menti<strong>on</strong>ed neurodegenerative diseases. Upregulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsps could <str<strong>on</strong>g>of</str<strong>on</strong>g>fer a therapeutic strategy to<br />

counter c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>al changes in neur<strong>on</strong>al proteins that trigger pathogenic cascades resulting in<br />

neurodegenerative diseases. Differentiated neur<strong>on</strong>s in both in vivo and in vitro systems have been reported to<br />

be refractory to Hsp inducti<strong>on</strong> by c<strong>on</strong>venti<strong>on</strong>al heat shock. We will report <strong>on</strong> a compound that induces Hsps<br />

in differentiated neur<strong>on</strong>s with the interesting feature that it induces a wider set <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsps in differentiated<br />

human neur<strong>on</strong>s compared to differentiated rodent neur<strong>on</strong>s.<br />

5B_02_S<br />

MOLECULAR CHAPERONES AND PROTECTION FROM CEREBRAL ISCHEMIA<br />

R<strong>on</strong>a G. Giffard, Yibing Ouyang, LiJun Xu, Midori Yenari<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Anesthesia, Stan<str<strong>on</strong>g>for</str<strong>on</strong>g>d University, Stan<str<strong>on</strong>g>for</str<strong>on</strong>g>d, CA USA<br />

Stroke is the third leading cause <str<strong>on</strong>g>of</str<strong>on</strong>g> death in many countries. Data from both animal stroke models and<br />

primary cultures have dem<strong>on</strong>strated that Hsp70 overexpressi<strong>on</strong> can reduce cerebral ischemic injury. We have<br />

used viral vectors and DNA transfecti<strong>on</strong> to overexpress Hsp70 in the brain and in brain cells. We have<br />

previously reported that overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 and Hsp40 reduce ischemic injury in in vitro models <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

ischemia, and Hsp70 overexpressi<strong>on</strong> is effective in rodent models <str<strong>on</strong>g>of</str<strong>on</strong>g> stroke. To delineate which domains<br />

within Hsp70 are important <str<strong>on</strong>g>for</str<strong>on</strong>g> protecti<strong>on</strong> from ischemia in the brain we compared two mutants to full


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

length wild type Hsp70. One was a point mutant in the ATPase domain- K71E, the sec<strong>on</strong>d was a deleti<strong>on</strong><br />

mutant encoding the carboxy terminal domain, amino acids 381-640. The c<strong>on</strong>structs were injected<br />

intracerebroventricularly, resulting in transfecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> both neur<strong>on</strong>s and astrocytes. We observed that both the<br />

ATPase deficient mutant and the carboxyterminal domain al<strong>on</strong>e were similar to the wild type HSP70 in ability<br />

to significantly reduce focal ischemic injury in the rat. Additi<strong>on</strong>al effects <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 that could c<strong>on</strong>tribute to<br />

protecti<strong>on</strong> were studied in Hsp70 overexpressing mice subjected to focal ischemia. Here we found evidence<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> inflammati<strong>on</strong>- reduced activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> NFkB, and decreased expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> iNOS, in the Hsp70<br />

overexpressing mice. We also found previously that Hsp70 overexpressi<strong>on</strong> was associated with higher levels<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the antiapoptotic protein Bcl-2. We have thus found at least three mechanisms c<strong>on</strong>tributing to protecti<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the brain from ischemia by Hsp70. These are binding <str<strong>on</strong>g>of</str<strong>on</strong>g> unfolded proteins, inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> inflammati<strong>on</strong> and<br />

inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis. Hsp70 is a multifaceted protein, and at least several <str<strong>on</strong>g>of</str<strong>on</strong>g> its effects are involved in<br />

protecting the brain from ischemia.<br />

5B_03_S<br />

MANIPULATING THE HEAT SHOCK RESPONSE<br />

AS A THERAPEUTIC STRATEGY IN ALS<br />

Bernadett Kalmar, Linda Greensmith<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Neurology, UCL, UK, e-mail: l.greensmith@i<strong>on</strong>.ucl.ac.uk<br />

Heat shock proteins (hsps) are know to play a role in neurodegenerative diseases, including Amyotrophic<br />

Lateral Sclerosis (ALS), in which mot<strong>on</strong>eur<strong>on</strong>s degenerate, resulting in muscle weakness, paralysis and finally<br />

death, usually within 2-5 years <str<strong>on</strong>g>of</str<strong>on</strong>g> diagnosis. Evidence suggests a relati<strong>on</strong>ship between the level <str<strong>on</strong>g>of</str<strong>on</strong>g> the heat<br />

shock resp<strong>on</strong>se (HSR) and the specific vulnerability <str<strong>on</strong>g>of</str<strong>on</strong>g> mot<strong>on</strong>eur<strong>on</strong>s to degenerati<strong>on</strong> in ALS. For example,<br />

mot<strong>on</strong>eur<strong>on</strong>s have an unusually high threshold <str<strong>on</strong>g>for</str<strong>on</strong>g> the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the HSR which may result in abnormal<br />

protein folding and trafficking and an increased susceptibility to apoptotic insults. We have recently upregulated<br />

the HSR in a mouse model <str<strong>on</strong>g>of</str<strong>on</strong>g> ALS. Treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> SOD1 G93A mice with arimoclomol, induces the<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp70 and hsp90, resulting in a delay in disease progressi<strong>on</strong> and a significant increase in<br />

survival. However, the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> arimoclomol are unlikely to result <strong>on</strong>ly by increasing hsp levels in<br />

mot<strong>on</strong>eur<strong>on</strong>s since genetic manipulati<strong>on</strong>s that increase hsp70 in SOD1 G93A mice fail to alter disease<br />

progressi<strong>on</strong>. It is not <strong>on</strong>ly the mechanism, but the site <str<strong>on</strong>g>of</str<strong>on</strong>g> acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> arimoclomol and hsps in SOD1 G93A mice<br />

that remains unclear. We are characterising the HSR in this model <str<strong>on</strong>g>of</str<strong>on</strong>g> ALS as a functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> disease<br />

progressi<strong>on</strong> and testing the differential neuroprotective effects <str<strong>on</strong>g>of</str<strong>on</strong>g> a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> manipulati<strong>on</strong>s that up-regulate<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> hsps. Our results show that not all agents that activate the HSR will necessarily have beneficial<br />

neuroprotective effects <strong>on</strong> mot<strong>on</strong>eur<strong>on</strong>s. Moreover, activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the HSR in n<strong>on</strong>-neur<strong>on</strong>al cells may have<br />

greater neuroprotective effects than targeting the HSR within mot<strong>on</strong>eur<strong>on</strong>s themselves. We hope that a<br />

better understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> the role <str<strong>on</strong>g>of</str<strong>on</strong>g> the hsps in ALS will help to optimize targeting <str<strong>on</strong>g>of</str<strong>on</strong>g> the HSR as an effective<br />

therapeutic strategy <str<strong>on</strong>g>for</str<strong>on</strong>g> this fatal disease.<br />

247


23-26 August 2007,<br />

Budapest, Hungary<br />

5B_04_S<br />

NEUROPROTECTIVE EFFECTS OF EXTRACELLULAR HSP70<br />

Michael Tytell<br />

Neurobiology & Anatomy, Wake Forest Univ. School <str<strong>on</strong>g>of</str<strong>on</strong>g> Med., Winst<strong>on</strong>-Salem, NC 27157<br />

e-mail: tytellm@wfu.edu<br />

When the cytoprotective functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsps was a relatively new c<strong>on</strong>cept in the 1980s, we and Hightower<br />

respectively observed that Hsp70 was passed from <strong>on</strong>e cell to another and released into the extracellular fluid<br />

(Tytell & Lasek, Brain Res 324:223, 1984; Hightower & Guid<strong>on</strong>, J Cell Physiol 138:257, 1989). These were<br />

unexpected results because the Hsp70 family was thought to be exclusively intracellular proteins. In the two<br />

decades since, extracellular Hsp70 has become well-documented. Our 1984 observati<strong>on</strong> that glial Hsp70 was<br />

transferred into the ax<strong>on</strong> led to the novel hypothesis that, in the nervous system at least, the stress tolerance<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>s was not solely dependent <strong>on</strong> their own Hsp70, but could be supplemented by additi<strong>on</strong>al Hsp70<br />

from adjacent glia. For neur<strong>on</strong>s, this hypothesis has major implicati<strong>on</strong>s because those with l<strong>on</strong>g ax<strong>on</strong>s (mm<br />

to meters in length) require newly synthesized proteins to be transported from the neur<strong>on</strong>al cell body to<br />

remain functi<strong>on</strong>al. If stress or injury occurs in ax<strong>on</strong>s at some distance from their cell bodies, hours to days are<br />

required <str<strong>on</strong>g>for</str<strong>on</strong>g> newly synthesized Hsp70 to reach the injury site, which is too l<strong>on</strong>g to be <str<strong>on</strong>g>of</str<strong>on</strong>g> use. Thus, supplying<br />

Hsp70 at the injury site would be more effective in maintaining neur<strong>on</strong>al and ax<strong>on</strong>al functi<strong>on</strong>. We have tested<br />

this idea in a number <str<strong>on</strong>g>of</str<strong>on</strong>g> models. . In the injured sciatic nerve, local applicati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 improved sensory<br />

and motor neur<strong>on</strong> survival. Similarly, in the light-damaged rat eye, Hsp70 injected into the vitreous chamber<br />

promoted survival <str<strong>on</strong>g>of</str<strong>on</strong>g> photoreceptors. In cultured glia and neur<strong>on</strong>s, we found that glia released Hsp70 and<br />

that neur<strong>on</strong>s took up Hsp70 from the medium, leading to greater resistance to apoptosis induced chemically<br />

or by lack <str<strong>on</strong>g>of</str<strong>on</strong>g> trophic factor. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, Hsp70 has potential as a therapeutic agent in acute nervous system<br />

injury.<br />

248


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5B_02_P<br />

(poster secti<strong>on</strong> B1, poster board #191, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ELECTRO-ACUPUNCTURE AND BRAIN PROTECTION FROM CEREBRAL<br />

ISCHEMIA: DIFFERENTIAL ROLES OF ACUPOINTS<br />

F. Zhou 1,2 , J. C. Guo 2,1 , J. S. Cheng 2,1 , G. C. Wu 2,1 , Y. Xia 3,1<br />

1Shanghai <str<strong>on</strong>g>Research</str<strong>on</strong>g> Center <str<strong>on</strong>g>for</str<strong>on</strong>g> Acupuncture and Moxibusti<strong>on</strong>, Shanghai, China;<br />

2Fudan University Shanghai Medical College, Shanghai, China;<br />

3Yale University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, New Haven, CT, USA<br />

e-mail: Ying.Xia@Yale.Edu<br />

We have shown that electro-acupuncture (EA) significantly reduces ischemic infarcti<strong>on</strong> in the rat model <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

cerebral ischemia (right middle cerebral artery occlusi<strong>on</strong>, MCAO). Since there are multiple acupoints that may<br />

cause different effects <strong>on</strong> the body, we asked, in this work, whether the EA protecti<strong>on</strong> from cerebral ischemia<br />

varies with the acupoints stimulated. We observed that 1-hour MCAO greatly reduced cerebral blood flow<br />

and caused the brain infarcti<strong>on</strong> and EA with sparse-dense wave (5Hz/4s-20Hz/4s) at 1.0mA <str<strong>on</strong>g>for</str<strong>on</strong>g> 30 minutes<br />

differentially attenuated the ischemic infarcti<strong>on</strong> depending <strong>on</strong> the acupoints used. In the group <str<strong>on</strong>g>of</str<strong>on</strong>g> head<br />

acupoints, i.e., “Shuigou” (Du 26) and “Baihui” (Du 20), the cerebral infarcti<strong>on</strong> was greatly reduced from<br />

33.4%±3.1% to 4.9%±1.2% <str<strong>on</strong>g>of</str<strong>on</strong>g> the brain (n=30, P


23-26 August 2007,<br />

Budapest, Hungary<br />

1.0mA±0.1mA, starting from 5 mins after the <strong>on</strong>set <str<strong>on</strong>g>of</str<strong>on</strong>g> MCAO. The results showed that EA induced a<br />

significant increase in cerebral blood flow in the ischemic cortex. EA <str<strong>on</strong>g>for</str<strong>on</strong>g> 5-30 mins significantly reduced<br />

ischemic infarct volume and relieved neurological deficits. This benefit effect increased with EA length<br />

between 5 and 30 min with the infarct volume being reduced by >80% in the 30-min group (n=60, P


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5C. HEAVY METAL STRESS<br />

(LAWRENCE E. HIGHTOWER)<br />

5C_01_S<br />

METALLOTHIONEIN-MEDIATED IMMUNOMODULATION; NEW ROLES FOR<br />

AN OLD STRESS RESPONSE<br />

* M. A. Lynes, * X. Yin, * D. W. Unfricht, * G. Marusov, ** B. L. Ly<strong>on</strong>s, † R. T. Emeny,<br />

†D. A. Lawrence<br />

*University <str<strong>on</strong>g>of</str<strong>on</strong>g> C<strong>on</strong>necticut, Storrs, CT, USA<br />

**Jacks<strong>on</strong> Laboratory, Bar Harbor, ME, USA<br />

†Wadsworth Center, NYSDH, Albany, NY, USA<br />

Metallothi<strong>on</strong>ein (MT) is an unusual stress resp<strong>on</strong>se protein with a novel and intriguing structure and a set <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

distinct biological functi<strong>on</strong>s. The protein is remarkably cysteine rich, and many <str<strong>on</strong>g>of</str<strong>on</strong>g> its functi<strong>on</strong>s revolve<br />

around cysteine-associated thiols. This highly c<strong>on</strong>served protein is known to be a reservoir <str<strong>on</strong>g>of</str<strong>on</strong>g> essential<br />

divalent heavy metals, it sequesters toxicants and scavenges free radicals, and it can act to regulate<br />

transcripti<strong>on</strong> factor activity. In each <str<strong>on</strong>g>of</str<strong>on</strong>g> these instances, the principal site <str<strong>on</strong>g>of</str<strong>on</strong>g> acti<strong>on</strong> is thought to be in the<br />

nuclear and cytosolic compartments, and this perspective is c<strong>on</strong>sistent with the observati<strong>on</strong> that MT does not<br />

have a signal sequence <str<strong>on</strong>g>for</str<strong>on</strong>g> export through the endoplasmic reticulum and golgi apparatus. Despite this signal<br />

sequence absence, there is str<strong>on</strong>g evidence that MT can be released to extracellular spaces in significant<br />

amounts under stressful circumstances. We have found that MT in the extracellular envir<strong>on</strong>ment can act as a<br />

chemotactic agent, and that this resp<strong>on</strong>se may be mediated by interacti<strong>on</strong>s with G-coupled protein receptors.<br />

Our recent studies have focused <strong>on</strong> mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> cadmium-mediated immunomodulati<strong>on</strong>, and the effect <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

manipulati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> MT gene dose <strong>on</strong> the progressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> humoral immunity against T-dependent antigens<br />

during low dose cadmium or zinc exposure. At doses <str<strong>on</strong>g>of</str<strong>on</strong>g> metal that do not influence the humoral resp<strong>on</strong>se in<br />

wild type mice, mice that carry a targeted disrupti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the Mt1 and Mt2 genes show significant<br />

immunosuppressi<strong>on</strong> in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> cadmium, but not zinc. In c<strong>on</strong>trast, both cadmium and zinc suppress<br />

the humoral resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> mice that carry the Mt1 transgene c<strong>on</strong>struct that drives over-expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

metallothi<strong>on</strong>ein. This suppressi<strong>on</strong> is associated with changes in the cytokine pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ile produced by stimulated<br />

splenocytes. Both cadmium and zinc are inducers <str<strong>on</strong>g>of</str<strong>on</strong>g> MT mRNA and protein expressi<strong>on</strong>, but at sub-toxic<br />

c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> these metals <strong>on</strong>ly cadmium provokes a selective release <str<strong>on</strong>g>of</str<strong>on</strong>g> MT to the extracellular<br />

envir<strong>on</strong>ment. We have also explored the impact <str<strong>on</strong>g>of</str<strong>on</strong>g> changes to the MT gene dose in mice infected with<br />

Listeria m<strong>on</strong>ocytogenes. Clearance <str<strong>on</strong>g>of</str<strong>on</strong>g> L.m. was accelerated in mice that carry both the targeted disrupti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> Mt1<br />

and Mt2, as well as in mice that carry the Mt1 transgene c<strong>on</strong>struct when compared to c<strong>on</strong>genic wild type<br />

c<strong>on</strong>trols. We hypothesize that MT may represent an central regulator <str<strong>on</strong>g>of</str<strong>on</strong>g> immune functi<strong>on</strong>s that occur under<br />

stressful circumstances, and that manipulati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> MT levels in the extracellular envir<strong>on</strong>ment may provide an<br />

avenue <str<strong>on</strong>g>for</str<strong>on</strong>g> the management <str<strong>on</strong>g>of</str<strong>on</strong>g> inflammatory, infectious, autoimmune and neoplastic disease processes.<br />

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23-26 August 2007,<br />

Budapest, Hungary<br />

252<br />

5C_02_S<br />

COPPER METABOLIC DISORDER IN MYOCARDIAL PATHOGENESIS<br />

Y. James Kang<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine and Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmacology and Toxicology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Louisville<br />

School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Louisville, KY, USA<br />

The associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> copper (Cu) with cardiomyopathy has been recognized <str<strong>on</strong>g>for</str<strong>on</strong>g> a l<strong>on</strong>g time, but its clinical<br />

significance has not been explored until recently. In diabetic patients and rat model, Cu chelati<strong>on</strong> therapy<br />

using a Cu chelator trientine has been shown to ameliorate cardiac pathological changes and improve heart<br />

functi<strong>on</strong>. In c<strong>on</strong>trary, we have observed that dietary supplementati<strong>on</strong> with physiologically relevant levels <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Cu can reverse established hypertrophic cardiomyopathy in aortic banding mouse model even in the presence<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> pressure overload. Both results have indicati<strong>on</strong>s <str<strong>on</strong>g>for</str<strong>on</strong>g> Cu metabolic disorders in cardiomyopathy in humans.<br />

However, it is important to distinguish the fundamental differences in Cu metabolic disorder between<br />

diabetic and pressure overload cardiomyopathy. Both diabetes and pressure overload cause cardiac oxidative<br />

stress, mitoch<strong>on</strong>drial damage, hypertrophy and dysfuncti<strong>on</strong>. However, we have observed that Cu levels<br />

slightly increase in the heart, but significantly decrease in the liver <str<strong>on</strong>g>of</str<strong>on</strong>g> streptozotocin (STZ)-induced diabetic<br />

mice. In c<strong>on</strong>trary, Cu c<strong>on</strong>centrati<strong>on</strong>s decrease in the heart, but do not change in the liver <str<strong>on</strong>g>of</str<strong>on</strong>g> pressure overload<br />

mice. It is possible that diabetic complicati<strong>on</strong>s lead to systemic disorder <str<strong>on</strong>g>of</str<strong>on</strong>g> Cu metabolism and disrupti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Cu detoxificati<strong>on</strong> in the liver, thus increasing the risk <str<strong>on</strong>g>of</str<strong>on</strong>g> Cu toxicity to organs including the heart, but in<br />

pressure overload the heart is the primary organ <str<strong>on</strong>g>of</str<strong>on</strong>g> Cu metabolic disorder leading to cardiac Cu deficiency but<br />

maintaining liver detoxificati<strong>on</strong> functi<strong>on</strong>. In this c<strong>on</strong>text, Cu chelati<strong>on</strong> would be beneficial to patients with<br />

diabetic cardiomyopathy, but Cu supplementati<strong>on</strong> would improve the c<strong>on</strong>diti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> pressure overload<br />

cardiomyopathy. Both cases have been dem<strong>on</strong>strated in animal studies and it is important to apply the<br />

in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> generated from animal studies to human cardiomyopathy patients. These studies were supported<br />

in part by US-NIH grants HL59225 and HL63760.<br />

5C_03_S<br />

BRAIN INJURY AND OXIDATIVE STRESS: THINK ZINC!<br />

Stefano Sensi 1,2<br />

Depts. <str<strong>on</strong>g>of</str<strong>on</strong>g> Neurology, and Basic and Applied Medical Sciences, 1Center <str<strong>on</strong>g>of</str<strong>on</strong>g> Excellence <strong>on</strong> Aging / University<br />

“G. d’Annunzio”, Chieti-Pescara, Italy, 2University <str<strong>on</strong>g>of</str<strong>on</strong>g> Cali<str<strong>on</strong>g>for</str<strong>on</strong>g>nia-Irivine, Irvine., USA,<br />

e-mail: ssensi@uci.edu<br />

Zn2+ is a potent mediator <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>al injury both “in vitro”and “in vivo”, and trans-synaptic movement <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the cati<strong>on</strong> from pre- to post-synaptic neur<strong>on</strong>s has been shown to greatly c<strong>on</strong>tribute to a variety <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

neurological c<strong>on</strong>diti<strong>on</strong>s including cerebral ischemia, brain trauma and epilepsy. Zn2+ can enter neur<strong>on</strong>s<br />

through glutamate receptor-associated channels [NMDA and AMPA/kainate channels (Ca-A/K channels),<br />

voltage-sensitive calcium channels (VSCC), or Zn2+-sensitive membrane transporters (like the Na+/Ca2+ and<br />

Na+/Zn2+ exchangers). Mechanisms by which Zn2+ exerts its potent neurotoxic effects include both<br />

mitoch<strong>on</strong>drial and extra-mitoch<strong>on</strong>drial pathways. Experiments in cortical neur<strong>on</strong>s have shown that<br />

mitoch<strong>on</strong>dria play an important role in restoring Zn2+ i homeostasis but this Zn2+ uptake leads also to<br />

prol<strong>on</strong>ged mitoch<strong>on</strong>drial depolarizati<strong>on</strong> and free radicals generati<strong>on</strong>. In additi<strong>on</strong> to roles in acute injury, Zn2+<br />

might play roles in the selective neurodegenerati<strong>on</strong> associated with aging and Alzheimer’s disease (AD).<br />

Indeed, cumulative effects <str<strong>on</strong>g>of</str<strong>on</strong>g> repeated Zn2+ exposures could c<strong>on</strong>tribute to the oxidative damage and


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

mitoch<strong>on</strong>drial dysfuncti<strong>on</strong> seen in AD. Interestingly, Zn2+ homeostasis is affected by oxidative stress, as<br />

reactive oxygen species are potent triggers <str<strong>on</strong>g>for</str<strong>on</strong>g> injurious cati<strong>on</strong> release from Zn2+ binding proteins<br />

(metallothi<strong>on</strong>eins). In this talk, we examine how Zn2+ dyshomeostasis and oxidative stress might act<br />

synergistically to promote degenerati<strong>on</strong> in the c<strong>on</strong>text <str<strong>on</strong>g>of</str<strong>on</strong>g> several neurological c<strong>on</strong>diti<strong>on</strong>s.<br />

5C_04_S<br />

CYTOPROTECTIVE METAL IONS<br />

George A. Perdrizet a , Lawrence E. Hightower b<br />

a Dept. Trauma, Hart<str<strong>on</strong>g>for</str<strong>on</strong>g>d Hospital and Univ. C<strong>on</strong>necticut, Hart<str<strong>on</strong>g>for</str<strong>on</strong>g>d, CT 06102<br />

b Dept Cell and Molecular Biology, Univ. C<strong>on</strong>necticut, Storrs, 06269<br />

The cellular stress resp<strong>on</strong>se and stress protein expressi<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g>m the biologic basis <str<strong>on</strong>g>of</str<strong>on</strong>g> stress c<strong>on</strong>diti<strong>on</strong>ing<br />

protocols. Currently most in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <strong>on</strong> inducible cytoprotecti<strong>on</strong> has been focused <strong>on</strong> the use <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

hyperthermia as the inducing agent. Heat shock prec<strong>on</strong>diti<strong>on</strong>ing has amply dem<strong>on</strong>strated the pro<str<strong>on</strong>g>of</str<strong>on</strong>g> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

c<strong>on</strong>cept that stress c<strong>on</strong>diti<strong>on</strong>ing protocols have the potential to provide beneficial cytoprotecti<strong>on</strong> in the<br />

setting <str<strong>on</strong>g>of</str<strong>on</strong>g> elective invasive medical and surgical procedures. While whole-body hyperthermia is a very<br />

effective agent, a pharmacologic agent would be more attractive as a clinical method to achieve a<br />

cytoprotected state in diseased humans. Metal i<strong>on</strong>s have been shown to be potent inducers <str<strong>on</strong>g>of</str<strong>on</strong>g> the cellular<br />

stress resp<strong>on</strong>se, are anti-inflammatory and capable <str<strong>on</strong>g>of</str<strong>on</strong>g> providing clinically relevant cytoprotecti<strong>on</strong>. We have<br />

successfully used the chloride salts <str<strong>on</strong>g>of</str<strong>on</strong>g> tin and zinc to provide in vivo cytoprotecti<strong>on</strong> in animal models <str<strong>on</strong>g>of</str<strong>on</strong>g>: 1.<br />

Rodent renal artery occlusi<strong>on</strong>, 2. rabbit spinal cord ischemia 3. acute pulm<strong>on</strong>ary inflammati<strong>on</strong>, rabbit fat<br />

embolism syndrome (intravenous administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> oleic acid), 4. acute inflammati<strong>on</strong> within the rodent<br />

mesenteric blood vessels. In these models, evidence <str<strong>on</strong>g>for</str<strong>on</strong>g> inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> stress proteins and inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> acute<br />

inflammati<strong>on</strong> will be presented as potential mechanisms <str<strong>on</strong>g>for</str<strong>on</strong>g> the observed cytoprotecti<strong>on</strong>. The nature and<br />

activities <str<strong>on</strong>g>of</str<strong>on</strong>g> tin and zinc metals will be compared and c<strong>on</strong>trasted.<br />

5C_05_S<br />

(poster secti<strong>on</strong> B1, poster board #194, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

INFLUENCE OF MERCURY ON RAT RENAL GLUCOCORTICOID RECEPTOR<br />

ASSOCIATION WITH HSP90 AND HSP70<br />

Jadranka Dundjerski, Jelena Brkljačić, Tatjana Perišić, Gordana Matić<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Biological <str<strong>on</strong>g>Research</str<strong>on</strong>g> »Siniša Stanković«, Belgrade, Serbia<br />

e-mail: jadund@ibiss.bg.ac.yu<br />

Unliganded glucocorticoid receptor (GR) is located in cytoplasm in <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> multiprotein heterocomplex with<br />

various proteins including Hsp90 and Hsp70. The structure, compositi<strong>on</strong>, assembly and functi<strong>on</strong>al<br />

significance <str<strong>on</strong>g>of</str<strong>on</strong>g> this complex depends <strong>on</strong> cell type, physiological demands and envir<strong>on</strong>mental c<strong>on</strong>diti<strong>on</strong>s. It is<br />

known that toxic effects <str<strong>on</strong>g>of</str<strong>on</strong>g> mercury could be reflected <strong>on</strong> structure and functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> transcripti<strong>on</strong>al factors<br />

such as GR. The influence <str<strong>on</strong>g>of</str<strong>on</strong>g> mercury <strong>on</strong> associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> rat kidney GR with heat shock proteins Hsp90 and<br />

Hsp70 and <strong>on</strong> cytosolic levels <str<strong>on</strong>g>of</str<strong>on</strong>g> these Hsps was investigated. The GR heterocomplexes with Hsp90 and<br />

Hsp70 were immunopurified from renal cytosol <str<strong>on</strong>g>of</str<strong>on</strong>g> rats administered with different doses <str<strong>on</strong>g>of</str<strong>on</strong>g> mercury (1, 2<br />

and 3 mg Hg/kg b.w.). A quantitative immunoblotting procedure was applied to determine the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> GR,<br />

Hsp90 and two Hsp70 is<str<strong>on</strong>g>of</str<strong>on</strong>g>orms (c<strong>on</strong>stitutive Hsp73 and inducible Hsp72) in the renal cytosol, as well as the<br />

amounts <str<strong>on</strong>g>of</str<strong>on</strong>g> these proteins within GR heterocomplexes immunoprecipitated by anti-GR antibody. Mercury<br />

253


23-26 August 2007,<br />

Budapest, Hungary<br />

was found to stimulate GR associati<strong>on</strong> with all examined Hsps. The most prominent effect <str<strong>on</strong>g>of</str<strong>on</strong>g> the metal was<br />

stimulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp72 interacti<strong>on</strong> with GR. On the other hand, the metal administrati<strong>on</strong> led to an increase <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Hsp90 level in the cytosol, while the cytosolic levels <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 is<str<strong>on</strong>g>of</str<strong>on</strong>g>orms remained unaltered. These findings<br />

suggest that associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsps, at least Hsp70, with the GR might be ascribed to changes in the affinity <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

their interacti<strong>on</strong> rather than to changes in Hsps availability in the cytosol. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, GR heterocomplex<br />

assembly seems to be a c<strong>on</strong>trolled process enabling chaper<strong>on</strong>ing and functi<strong>on</strong>ing <str<strong>on</strong>g>of</str<strong>on</strong>g> the GR in c<strong>on</strong>cert with<br />

physiological demands.<br />

5C_06_S<br />

(poster secti<strong>on</strong> B1, poster board #195, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DIFFERENTIAL STRESS PROTEINS EXPRESSION IN NRK-52E CELLS EXPOSED<br />

TO HG(II) OR PB(II)<br />

§ A. Stacchiotti, * F. Morandini, * F. Bett<strong>on</strong>i, § E. Foglio, ° M. Cadei, § L. F. Rodella, ° P. Grigolato,<br />

* M. F. Aleo<br />

§Human Anatomy, °2nd Pathology and *Biochemistry Units, School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Brescia, Viale<br />

Europa 11, 25123-Brescia, Italy<br />

e-mail: aleo@med.unibs.it<br />

Mercury Hg(II) and lead Pb(II), are two hazardous envir<strong>on</strong>mental c<strong>on</strong>taminants having nephrotoxic effects<br />

and whose toxic acti<strong>on</strong> appear to be associated to the cellular increase <str<strong>on</strong>g>of</str<strong>on</strong>g> reactive oxygen species (ROS). Cells<br />

resp<strong>on</strong>d to oxidative damage by synthesizing highly c<strong>on</strong>served stress proteins, heat shock (HSP) and glucose<br />

related proteins (GRP). These molecular chaper<strong>on</strong>es protect other cellular proteins and organules from<br />

injuries induced by a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> stressors, including heavy metals. So, the present in vitro study is undertaken to<br />

compare the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> five stress proteins in rat proximal tubular cells exposed to subcytotoxic<br />

c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> Hg(II) or Pb(II) salts. Proliferating NRK-52E cells received <str<strong>on</strong>g>for</str<strong>on</strong>g> 24h culture media<br />

c<strong>on</strong>taining 20µM HgCl 2 or 60µM PbCl 2 then the presence and abundance <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP25, HSP60, HSP70,<br />

GRP75, GRP78 were analysed by immunohistochemistry and Western blotting. C<strong>on</strong>comitantly, in order to<br />

check if stress proteins resp<strong>on</strong>se might be really related to a renal oxidative damage, ROS and glutathi<strong>on</strong>e<br />

(GSH) levels were measured by flow cytometry and spettrophotometric analysis. Our data proved that in<br />

NRK-52E both HgCl 2 and PbCl 2 treatments enhance the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> c<strong>on</strong>stitutive chaper<strong>on</strong>es (HSP25,<br />

GRP75 and GRP78) and the cell GSH c<strong>on</strong>tent, even if at different grade. Interestingly, <strong>on</strong>ly Hg(II) i<strong>on</strong>s<br />

stimulate either the ROS <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> or the inducible HSP70 protein. These results suggest that, in our in vitro<br />

system, Hg(II)-induced ROS producti<strong>on</strong> and expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> peculiar stress proteins could be a c<strong>on</strong>sequence <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

a different mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> this metal with respect to Pb(II).<br />

254


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5C_02_P<br />

(poster secti<strong>on</strong> B1, poster board #196, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

OXIDATIVE STRESS PRODUCED BY ENVIRONMENTAL AGENTS ON<br />

AMPHIBIAN EMBRYOS<br />

Jorge Herkovits 1 , Cristina S. Pérez-Coll 1 , Jose L. D´Eramo 1 , Luis A. Castañaga 1 ,<br />

Carolina Ar<strong>on</strong>z<strong>on</strong> 1 , Abelardo A. Sztrum 1 , Juan C. Stockert 1,2<br />

Instituto de Ciencias Ambientales y Salud, Fundación PROSAMA, Buenos Aires, Argentina<br />

e-mail: herkovit@mail.retina.ar<br />

1Instituto de Ciencias Ambientales y Salud<br />

2Departamento de Biología, Facultad de Ciencias,<br />

Universidad Autónoma de Madrid, Cantoblanco, Madrid, España<br />

The decline <str<strong>on</strong>g>of</str<strong>on</strong>g> amphibian populati<strong>on</strong>s could be related to physico chemical agents producing oxidative stress.<br />

Although Reactive Oxygen Species (ROS) participate in metabolic signalling pathways that regulate cell<br />

survival, proliferati<strong>on</strong> and apoptosis, an excess <str<strong>on</strong>g>of</str<strong>on</strong>g> ROS induce oxidative stress and could result in damage and<br />

death. In this study c<strong>on</strong>ducted with Chaunus arenarum embryos we report the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> different experimental<br />

c<strong>on</strong>diti<strong>on</strong>s that show the possibility (i) to prevent, or at least, to mitigate the adverse effects <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidative<br />

stress, and (ii) to exacerbate the toxicity by means <str<strong>on</strong>g>of</str<strong>on</strong>g> the synergic effect <str<strong>on</strong>g>of</str<strong>on</strong>g> agents inducing oxidative stress. In<br />

the first case the lethality induced by ultraviolet B light (UV-B) was prevented with Zn and Se in single or<br />

combined pre-treatments. Zn also protected against the lethality produced by Cu, Al, Cd, Ni, and Pb.<br />

Synergic effects were obtained by simultaneous treatments <str<strong>on</strong>g>of</str<strong>on</strong>g> photodynamic toxicity plus Zn, UV-B plus Ni<br />

and 2,4-D plus Cu. The results point out the complexity <str<strong>on</strong>g>of</str<strong>on</strong>g> the envir<strong>on</strong>mental scenarios related to oxidative<br />

stress effects <strong>on</strong> living organisms which could be <str<strong>on</strong>g>of</str<strong>on</strong>g> major relevance in the case <str<strong>on</strong>g>of</str<strong>on</strong>g> endangered species.<br />

5C_03_P<br />

(poster secti<strong>on</strong> B1, poster board #197, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STRESS, HOST DEFENSE AND HEALING<br />

Istvan Berczi 1 , Andres Quintanar Stephano 2 , Kalman Kovacs 3<br />

1 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Immunology, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, the University <str<strong>on</strong>g>of</str<strong>on</strong>g> Manitoba,<br />

Winnipeg, MB, Canada, R3E 0W3<br />

2 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Physiology, Free University <str<strong>on</strong>g>of</str<strong>on</strong>g> Augascaliantes, Augascaliantes, Mexico<br />

3 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Pathology, St. Michael’s Hospital, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Tor<strong>on</strong>to, Tor<strong>on</strong>to ON Canada<br />

Hans Selye c<strong>on</strong>cluded after 10 years <str<strong>on</strong>g>of</str<strong>on</strong>g> experimentati<strong>on</strong> that stressed animals are resistant to their stressors.<br />

Stressed animals also showed resistance to various other noxious agents. Selye called this phenomen<strong>on</strong> “the<br />

general adaptati<strong>on</strong> syndrome” (GAS). Resistant animals <str<strong>on</strong>g>of</str<strong>on</strong>g>ten survived, when compared with n<strong>on</strong>-stressed<br />

animals, which died after exposure to severe stress [1].<br />

The beneficial effects <str<strong>on</strong>g>of</str<strong>on</strong>g> stress are <str<strong>on</strong>g>of</str<strong>on</strong>g>ten overlooked today. It is comm<strong>on</strong> belief that stress is harmful and<br />

dangerous. Indeed, stress has been shown repeatedly to be immunosuppressive (<str<strong>on</strong>g>for</str<strong>on</strong>g> adaptive immunity) and<br />

also to weaken the animal’s resistance to various pathogenic challenges. Some observati<strong>on</strong>s in man also<br />

support the c<strong>on</strong>clusi<strong>on</strong> that stress is harmful.<br />

The stress syndrome, as described by Selye, is analogous to the Acute Phase Resp<strong>on</strong>se (APR, or febrile illness) as<br />

we know it today [1,2]. APR is an emergency defense reacti<strong>on</strong>, whereby the adaptive (thymus dependent) immune<br />

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23-26 August 2007,<br />

Budapest, Hungary<br />

resp<strong>on</strong>se is suppressed and the innate or natural immune functi<strong>on</strong> is dramatically amplified. Natural immune<br />

reacti<strong>on</strong>s are poly-specific, but were viewed earlier as “n<strong>on</strong>specific “. We suffer from febrile illness <strong>on</strong><br />

numerous occasi<strong>on</strong>s in our lifetime and in most instances recover completely, which is pro<str<strong>on</strong>g>of</str<strong>on</strong>g> <str<strong>on</strong>g>of</str<strong>on</strong>g> the efficacy<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> innate immunity. Sometimes, however, APR is unable to eliminate the problem and prol<strong>on</strong>ged disease or<br />

death may follow.<br />

Infecti<strong>on</strong> and various <str<strong>on</strong>g>for</str<strong>on</strong>g>ms <str<strong>on</strong>g>of</str<strong>on</strong>g> injury, including emoti<strong>on</strong>al trauma, induce APR. Cytokines (primarily IL-1-<br />

beta, TNF-alpha and IL-6) stimulate corticotropin releasing horm<strong>on</strong>e (CRH) and vasopressin (VP) secreti<strong>on</strong> and<br />

cause a “sympathetic outflow”. CRH is a powerful activator <str<strong>on</strong>g>of</str<strong>on</strong>g> the hypothalamic-pituitary adrenal (HPA) axis,<br />

which play a fundamental role in febrile illness, al<strong>on</strong>g with catecholamines. VP also stimulates the HPA axis.<br />

However VP remains active and elevated <str<strong>on</strong>g>for</str<strong>on</strong>g> a l<strong>on</strong>ger period than is CRH. VP, but not CRH, is elevated<br />

during chr<strong>on</strong>ic inflammatory diseases [2].<br />

We observed recently that VP c<strong>on</strong>trols adaptive immune functi<strong>on</strong> in rats. It is known from the literature, that<br />

VP stimulates the secreti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> both prolactin and growth horm<strong>on</strong>e, which support the functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the thymus and<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the adaptive immune system [2,3].<br />

It seems apparent from the evidence available to date that CRH and catecholamines boost the innate immune<br />

system, which are greatly amplified during APR and increase host resistance poly-specifically. This is the<br />

phenomen<strong>on</strong>, what was coined by Selye as GAS. On the other hand we propose that VP is the hypothalamic<br />

coordinator <str<strong>on</strong>g>of</str<strong>on</strong>g> healing and recovery. VP is capable <str<strong>on</strong>g>of</str<strong>on</strong>g> restoring homeostatic c<strong>on</strong>diti<strong>on</strong>s after acute illness and<br />

thereby bringing back also adaptive immunocompetence. Stress is harmful <strong>on</strong>ly when things get out <str<strong>on</strong>g>of</str<strong>on</strong>g> hand.<br />

256


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5D. STRESS AND THE IMMUNE RESPONSE<br />

(TAMAS BARTFAI)<br />

5D_01_S<br />

MOLECULAR UNDERSTANDING OF HPA AXIS INVOLVEMENT IN<br />

STRESS/IMMUNE CIRCUITS<br />

Eduardo Arzt<br />

Laboratorio de Fisiologia y Biologia Molecular, Universtity <str<strong>on</strong>g>of</str<strong>on</strong>g> Buenos Aires, and CONICET, Argentina<br />

The immune-neuroendocrine systems have an intimate cross communicati<strong>on</strong> making possible a satisfactory<br />

resp<strong>on</strong>se to envir<strong>on</strong>mental changes and stress. The hypothalamic-pituitary-adrenal axis (HPA) has a key role<br />

in the interacti<strong>on</strong> between the immune and neuroendocrine systems and in the stress resp<strong>on</strong>se. Cytokines<br />

activate the HPA axis and induce a rise in glucocorticoid levels, which are instrumental in order to c<strong>on</strong>trol<br />

immune-cytokine overreacti<strong>on</strong>s, acting <strong>on</strong> T lymphocytes and other ctyokine/target cells. The specificity <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

horm<strong>on</strong>al and cytokine signals <strong>on</strong> their target cells is crucial to provide specificity to their acti<strong>on</strong>s. The crosstalk<br />

between cytokine-induced transcripti<strong>on</strong> factors such as Tbet, GATA-3, NFκB, and AP-1 and steroid (i.e.<br />

glucocorticoid) receptors involve both genomic and n<strong>on</strong>-genomic acti<strong>on</strong>s, and c<strong>on</strong>stitutes the mechanism <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

fine tuning both horm<strong>on</strong>e and cytokine resp<strong>on</strong>ses. Corticotrophin releasing horm<strong>on</strong>e (CRH) is the key<br />

mediator <str<strong>on</strong>g>of</str<strong>on</strong>g> the HPA axis <str<strong>on</strong>g>of</str<strong>on</strong>g> the central nervous system resp<strong>on</strong>se needed to adapt to stressful c<strong>on</strong>diti<strong>on</strong>s. The<br />

final outcome <str<strong>on</strong>g>of</str<strong>on</strong>g> CRH/CRH1 signaling depends <strong>on</strong> the c<strong>on</strong>text <str<strong>on</strong>g>of</str<strong>on</strong>g> specific cells and ligands, cross-talk <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

signaling pathways and the effector acti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> the pathways <strong>on</strong>ce they are activated. This specificity bears<br />

c<strong>on</strong>sequences at the CNS level where CRH activates through the same receptor (CRHR1) different signaling<br />

pathways depending <strong>on</strong> neuroanatomical c<strong>on</strong>text. All these molecular interacti<strong>on</strong>s represent a key step <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

understanding, at the cellular and genetic level, the specificity and ultimate resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> physiological<br />

neuroendocrine-immune interacti<strong>on</strong>s during stress.<br />

5D_02_S<br />

INTERLEUKIN-1 SIGNALING AT THE INTERSECTION OF NEURONAL,<br />

IMMUNE AND METABOLIC STRESS<br />

Tamas Bartfai<br />

The Harold L Dorris Neurological research Institute. Molecular and Integrative Neurosciences Department,<br />

The Scripps <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute , La Jolla Ca 92037 USA<br />

Macrophages, adipocytes and macrophages in white adipose tissue, microglia and neur<strong>on</strong>s can synthesize the<br />

proinflammatory cytokine and endogenous pyrogen; IL-1 β in resp<strong>on</strong>se to acute immune, mechanic,<br />

oxidative and hypoxic insult, and to excitatory neurotoxic events. IL-1 synthesis is also induced by chr<strong>on</strong>ic<br />

elevati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> misfolded proteins or metabolic/endocrine stressors like elevated leptin levels. The IL-1 levels<br />

being highly inducible within short time serve as a comm<strong>on</strong> signal in neur<strong>on</strong>al immune and endocrine<br />

signaling.<br />

The wide spread expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> IL-1type 1 receptor <strong>on</strong> neur<strong>on</strong>s, pancreatic-β cells and macrophages, T and B<br />

cells accounts <str<strong>on</strong>g>for</str<strong>on</strong>g> the global inflammatory effects f IL-1.<br />

The acute stress leading to rapid activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the HPA axis, lowering seizure threshold and fever resp<strong>on</strong>se by<br />

IL-1 utilizes the rapid transcripti<strong>on</strong> independent neur<strong>on</strong>al effects <str<strong>on</strong>g>of</str<strong>on</strong>g> IL-1. The molecular steps <str<strong>on</strong>g>of</str<strong>on</strong>g> this<br />

signaling have been delineated in molecular detail showing that IL-1R1-MyD88-Neutral Spingomyelinase-<br />

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23-26 August 2007,<br />

Budapest, Hungary<br />

ceramide cSrc – phosphoprotein steps mediate the early IL-1 resp<strong>on</strong>se. After 45-60 min the classical Toll<br />

signaling pathway involving the NFKB mediated inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> COX2 and the subsequent producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

inflammatory mediator PGE2.<br />

The chr<strong>on</strong>ic inflammatory stress presented by obesity and the role <str<strong>on</strong>g>of</str<strong>on</strong>g> IL-1 in the c<strong>on</strong>versi<strong>on</strong> from insulin<br />

resistance to type 2 diabetes will also be discussed.<br />

258<br />

5D_03_S<br />

ROLE OF LIPIDS IN THE MODULATION OF HUMAN T CELL ACTIVATION<br />

T. Fulop 1 , P. Brassard 2 , A. Larbi 1 , F. Frisch 2 , C. Fortin 1 , A. Carpentier 2<br />

1Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Geriatrics and the Program <str<strong>on</strong>g>of</str<strong>on</strong>g> Immunology, 2 Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Endocrinology, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine,<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Sherbrooke, Sherbrooke, Qc, Canada<br />

Studies have shown suppressive effects <str<strong>on</strong>g>of</str<strong>on</strong>g> PUFA <strong>on</strong> T cell functi<strong>on</strong>s suggesting that lipids can have potent<br />

immunomodulatory effects. It is now accepted that the membrane <str<strong>on</strong>g>of</str<strong>on</strong>g> T cells is heterogeneous and c<strong>on</strong>tains<br />

microdomains called lipid rafts (LR) playing an important role in TCR signalling. Thus, variati<strong>on</strong>s in lipid<br />

levels and compositi<strong>on</strong> may determine their effects <strong>on</strong> immune resp<strong>on</strong>se. Each time when lipids are ingested<br />

the immune system is submitted to a lipid stress. The precise mechanism <str<strong>on</strong>g>for</str<strong>on</strong>g> this effect has not been fully<br />

investigated in vivo in humans. Our aim was to determine whether there are differences in T cell functi<strong>on</strong>s and<br />

signalling depending <strong>on</strong> the way <str<strong>on</strong>g>of</str<strong>on</strong>g> lipid administrati<strong>on</strong> and compositi<strong>on</strong>. Peripheral T cells were isolated<br />

from healthy subjects be<str<strong>on</strong>g>for</str<strong>on</strong>g>e and after 2-hours <str<strong>on</strong>g>of</str<strong>on</strong>g> an intravenous infusi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> heparin + Intralipid (HI) during<br />

a euglycemic hyperinsulinemic clamp to induce a 2.5-fold elevati<strong>on</strong> in plasma linoleic acid c<strong>on</strong>centrati<strong>on</strong>.<br />

Similar experimental setting was designed after an oral meal (OM).. HI and OM reduced peripheral T cell<br />

membrane fluidity and altered lipid raft organisati<strong>on</strong>. Both associated with reduced T cell proliferati<strong>on</strong> up<strong>on</strong><br />

CD3 + CD28 co-stimulati<strong>on</strong>. Tyrosine phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> LAT and activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Akt in T cells were also<br />

impaired without a reducti<strong>on</strong> in T cell receptor expressi<strong>on</strong>. The LR polarizati<strong>on</strong> was also altered. A selective<br />

increase in plasma linoleic acid c<strong>on</strong>centrati<strong>on</strong> and in intravascular lipolysis there<str<strong>on</strong>g>for</str<strong>on</strong>g>e have a suppressive effect<br />

<strong>on</strong> peripheral T cell CD28-dependent activati<strong>on</strong> and this effect associates with changes in plasma membrane<br />

properties. The lipid compositi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> nutriti<strong>on</strong>al therapy in patients at high risk <str<strong>on</strong>g>of</str<strong>on</strong>g> septic complicati<strong>on</strong>s may be<br />

crucial and may also be <str<strong>on</strong>g>of</str<strong>on</strong>g> relevance <str<strong>on</strong>g>for</str<strong>on</strong>g> type 2 diabetes. Furthermore, oral nutriti<strong>on</strong> rich in lipids c<strong>on</strong>stituting<br />

a chr<strong>on</strong>ic lipid stress, at l<strong>on</strong>g term, could c<strong>on</strong>tribute to the observed immunosenescence.<br />

5D_04_S<br />

NEUROENDOCRINE RESPONSE IN SUSCEPTIBILITY TO INFLAMMATORY,<br />

AUTOIMMUNE AND INFECTIOUS DISEASES<br />

Esther M. Sternberg<br />

NIMH/NIH, Bethesda, MD, USA<br />

The central nervous system (CNS) plays an important role in regulating immunity and in susceptibility and<br />

resistance to autoimmune, inflammatory and infectious diseases. Cytokines released during inflammati<strong>on</strong><br />

mediate changes in brain functi<strong>on</strong>, inducing sickness behaviors, sleep, fever and activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the horm<strong>on</strong>al<br />

stress resp<strong>on</strong>se (hypothalamic-pituitary-adrenal axis, HPA axis). In turn neural and neuroendocrine resp<strong>on</strong>ses,<br />

including the HPA axis, sympathetic and parasympathetic resp<strong>on</strong>ses, regulate immune resp<strong>on</strong>ses, thus<br />

providing important extra-immune system feedback c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> immunity. HPA axis activati<strong>on</strong> inhibits<br />

inflammati<strong>on</strong> through the generally anti-inflammatory acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the glucocorticoids. However, physiologic


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> glucocorticoids are immunomodulatory, shifting cytokine producti<strong>on</strong> from a TH1 to a TH2<br />

resp<strong>on</strong>se and enhancing delayed type hypersensitivity. Interrupti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> the HPA axis at any level and through<br />

multiple mechanisms, whether genetic, surgical or pharmacological, render inflammatory resistant hosts<br />

susceptible to inflammatory disease. In c<strong>on</strong>trast, over-activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> this axis, as in chr<strong>on</strong>ic stress, enhances<br />

severity <str<strong>on</strong>g>of</str<strong>on</strong>g> infecti<strong>on</strong>s, through the immunosuppressive effects <str<strong>on</strong>g>of</str<strong>on</strong>g> the glucocorticoids. The associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a<br />

blunted HPA axis with autoimmune/inflammatory disease occurs across species, strains and diseases, and in a<br />

variety <str<strong>on</strong>g>of</str<strong>on</strong>g> human autoimmune/inflammatory illnesses. Tissue resistance to glucocorticoids resulting from<br />

polymorphisms, mutati<strong>on</strong>s or dysfuncti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the glucocorticoid receptor (GR) is also associated with<br />

enhanced autoimmune/inflammatory disease expressi<strong>on</strong>. We have described a new mechanism <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

glucocorticoid resistance related to bacterial toxin repressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the GR and other nuclear horm<strong>on</strong>e receptors.<br />

This suggests a potential new mechanism <str<strong>on</strong>g>for</str<strong>on</strong>g> shock and inflammatory sequelae <str<strong>on</strong>g>of</str<strong>on</strong>g> bacterial infecti<strong>on</strong>s, and<br />

potential new approaches to developing therapies <str<strong>on</strong>g>for</str<strong>on</strong>g> these c<strong>on</strong>diti<strong>on</strong>s.<br />

5D_05_S<br />

(poster secti<strong>on</strong> B1, poster board #198, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ADRENOCORTICAL HYPERRESPONSIVENESS DURING POSTNATAL<br />

DEVELOPMENT OF STRESS SYSTEM IN MICE<br />

A. Yu. Shevchenko, T. V. Yakovleva, E. N. Makarova, N. M. Bazhan<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Cytology and Genetics, Siberian Branch <str<strong>on</strong>g>of</str<strong>on</strong>g> the Russian Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences,<br />

Lavrenteva 10, Novosibirsk, 630090 Russia<br />

The stress system in the mouse from postnatal days 1-12 is characterized by very low basal corticoster<strong>on</strong>e<br />

levels and an inability <str<strong>on</strong>g>of</str<strong>on</strong>g> mild stressors to induce an enhanced corticoster<strong>on</strong>e release. This period known as<br />

the stress-hyporesp<strong>on</strong>sive period. We previously dem<strong>on</strong>strated that in 3-week-old mice basal plasma<br />

corticoster<strong>on</strong>e levels and adrenal corticoster<strong>on</strong>e producti<strong>on</strong> were higher than both in 2-week-old mice and in<br />

adult mice. It is unknown whether stress-resp<strong>on</strong>se is increased and what changes in regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> adrenal<br />

steroidogenesis occur in 3-week-old mice. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to compare stress-resp<strong>on</strong>se, as well as<br />

activity <str<strong>on</strong>g>of</str<strong>on</strong>g> adenylate cyclase and steroidogenic enzymes in adrenal cells in 3-week-old mice with that in adult<br />

15-week-old mice. Stress-induced plasma corticoster<strong>on</strong>e level in 3-week-old mice was greater and developed<br />

faster than in adult mice. Corticoster<strong>on</strong>e producti<strong>on</strong> stimulated by ACTH, exogenous cAMP and<br />

progester<strong>on</strong>e, and ACTH- and <str<strong>on</strong>g>for</str<strong>on</strong>g>skolin-induced cAMP accumulati<strong>on</strong> in vitro were higher in 3-week-old mice<br />

than in adult mice. Thus, our data dem<strong>on</strong>strate that in the mouse the stress-hyperresp<strong>on</strong>sive period occurs<br />

during the third week after birth. This period is characterized by very high basal and stress induced plasma<br />

corticoster<strong>on</strong>e levels, which seems to be related to adrenal ACTH-resp<strong>on</strong>siveness. Adrenocortical<br />

hyperresp<strong>on</strong>siveness during the third week is caused by high adenylate cyclase activity and high intracellular<br />

steroidogenic enzyme activity.<br />

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23-26 August 2007,<br />

Budapest, Hungary<br />

5D_06_S<br />

(poster secti<strong>on</strong> B1, poster board #199, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STRESS-INDUCED CORTICOSTERONE ENHANCE BRAIN SURVEILLANCE BY T<br />

CELLS<br />

Gil. M Lewitus 1 , Hagit Cohen 2 , Michal Schwartz 1<br />

1The Weizmann Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Science, Rehovot, Israel<br />

2Ben-Guri<strong>on</strong> University <str<strong>on</strong>g>of</str<strong>on</strong>g> the Negev, Be’er Sheva, Israel<br />

Acute psychological stress was shown to act as an endogenous adjuvant, c<strong>on</strong>sequently enhancing cellmediated<br />

immunity. Independent work carried out by our research group, has dem<strong>on</strong>strated that whilst T<br />

cell-mediated immunity, recognizes antigens residing in the CNS (autoimmune cells), and well-regulated with<br />

respect to activity and length <str<strong>on</strong>g>of</str<strong>on</strong>g> time can fight <str<strong>on</strong>g>of</str<strong>on</strong>g>f abnormal behavior resulting from the stress. We examined<br />

the mechanism underlying the stress-induced T cell-mediated protective autoimmunity. We found that short<br />

exposure <str<strong>on</strong>g>of</str<strong>on</strong>g> mice to stressor (predator odor), especially in the hippocampus and the paraventricular nucleus,<br />

enhanced T-cell infiltrati<strong>on</strong> to the CNS. The peak <str<strong>on</strong>g>of</str<strong>on</strong>g> infiltrati<strong>on</strong> was 72 hr following mice exposure to the<br />

stressor. The T cells trafficking to the CNS was associated with ICAM-1 expressi<strong>on</strong>. This peak was preceded<br />

by a transient elevati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> corticoster<strong>on</strong>e. Assessment <str<strong>on</strong>g>of</str<strong>on</strong>g> the relati<strong>on</strong>ship between the stress-induced increase<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> corticoster<strong>on</strong>e and infiltrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> T cells revealed; while corticoster<strong>on</strong>e induces T cells mobilizati<strong>on</strong> to the<br />

CNS by inducing the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ICAM-1, the T cells are necessary <str<strong>on</strong>g>for</str<strong>on</strong>g> the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> corticoster<strong>on</strong>e<br />

following exposure to acute stress. Moreover, the beneficial effect <str<strong>on</strong>g>of</str<strong>on</strong>g> the elevati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> corticoster<strong>on</strong>e after the<br />

stress is partly mediated by the T cells. These results identify a new relati<strong>on</strong>ship between acute stress,<br />

corticoster<strong>on</strong>e, and adaptive immunity and support the argument that the immune system is needed to<br />

maintain normal brain homeostasis. They may also point the way to the development <str<strong>on</strong>g>of</str<strong>on</strong>g> immune-based<br />

therapies <str<strong>on</strong>g>for</str<strong>on</strong>g> post traumatic stress disorder.<br />

260


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5D_01_P<br />

(poster secti<strong>on</strong> B1, poster board #200, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HEAT SHOCK PROTEINS ARE RELEASED BY AN INTESTINAL EPITHELIAL<br />

CELL LINE INFECTED WITH ROTAVIRUS<br />

A. Barreto 1,2 , L. S. Rodriguez 1 , J. Angel 1 , M. Franco 1<br />

1Instituto de Genética Humana, Facultad de Medicina;<br />

2Departamento de Microbiología, Facultad de Ciencias, P<strong>on</strong>tificia Universidad Javeriana<br />

Carrera 7 No. 43-82, Bogotá-Colombia, e-mail: alf<strong>on</strong>so.barreto@javeriana.edu.co<br />

HSPs modulate the immune resp<strong>on</strong>se by signaling throughout receptors and transporting antigenic peptides<br />

to antigen presenting cells. Here we have studied if an intestinal epithelial cell line infected with rotavirus<br />

(RV) release c<strong>on</strong>stitutive and inducible heat shock proteins (Hsc70 and Hsp70, respectively). Caco-2 cells (a<br />

human col<strong>on</strong> adenocarcinoma cell line) were grown 14-21 days in a transwell system until they achieved<br />

polarizati<strong>on</strong>. Cells were c<strong>on</strong>sidered to be polarized when the transepithelial cell resistance (TER) was above<br />

300 Ohms/cm 2 . Caco-2 cells were infected with the RV strain RRV with a multiplicity <str<strong>on</strong>g>of</str<strong>on</strong>g> infecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 5, and<br />

the release <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsc70 and Hsp70 was evaluated at different time points after infecti<strong>on</strong> in both the apical and<br />

basal chamber by western blot and ELISA, respectively. 24 h post-infecti<strong>on</strong> both HSP were found in the<br />

apical chamber <str<strong>on</strong>g>of</str<strong>on</strong>g> infected, but not in c<strong>on</strong>trol treated cells. At this time the TER was still above 300<br />

Ohms/cm2, and we detected a higher number <str<strong>on</strong>g>of</str<strong>on</strong>g> infectious RRV particles in the apical side <str<strong>on</strong>g>of</str<strong>on</strong>g> the wells than<br />

in the basal side. At 48 h, we evidenced a fall <str<strong>on</strong>g>of</str<strong>on</strong>g> the TER due to m<strong>on</strong>olayer disrupti<strong>on</strong>, and we could detect<br />

both infectious virus and HSPs in the apical and basal side <str<strong>on</strong>g>of</str<strong>on</strong>g> the cultures. We also determined that released<br />

HSPs are mainly in a soluble <str<strong>on</strong>g>for</str<strong>on</strong>g>m, not precipitable by ultracentrifugati<strong>on</strong>. The simultaneous release <str<strong>on</strong>g>of</str<strong>on</strong>g> HSPs<br />

and RV by infected intestinal epithelial cell lines suggest that HSPs may modulate the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

mucosal immune resp<strong>on</strong>se against this pathogen.<br />

5D_02_P<br />

(poster secti<strong>on</strong> B1, poster board #201, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PERIPHERAL LEUKOCYTE APOPTOSIS IN ACUTE CORONARY SYNDROME<br />

M. Boutsikou 1 , C. Aggeli 2 , C. Tentolouris 2 , V. Bistola 1 , V. Ec<strong>on</strong>omou 1 , P. Doumba 1 ,<br />

E. Vavuranakis 2 , C. Stefanadis 2 , M. M. K<strong>on</strong>stadoulakis 1<br />

1Lab <str<strong>on</strong>g>of</str<strong>on</strong>g> Surgical <str<strong>on</strong>g>Research</str<strong>on</strong>g>, 1st Dept <str<strong>on</strong>g>of</str<strong>on</strong>g> Propaedeutic Surgery, Athens University<br />

21st Dept <str<strong>on</strong>g>of</str<strong>on</strong>g> Cardiology, Athens University, 114 V. S<str<strong>on</strong>g>of</str<strong>on</strong>g>ias Ave, 115 27 Athens<br />

e-mail: labsures@med.uoa.gr<br />

Purpose: To evaluate leukocyte apoptosis in patients with acute myocardial infarcti<strong>on</strong> (AMI) and with<br />

unstable angina pectoris (UAP). Methods: The study comprised <str<strong>on</strong>g>of</str<strong>on</strong>g> 41 c<strong>on</strong>secutive patients with AMI<br />

(mean±SD age:57±11) and 29 with UAP (61±10), all admitted in the Cor<strong>on</strong>ary Care Unit (CCU) within 6<br />

hours from the <strong>on</strong>set <str<strong>on</strong>g>of</str<strong>on</strong>g> symptoms. Ten patients without acute heart disease hospitalized in the cardiology<br />

department, served as C<strong>on</strong>trol group (C) (56±10). Blood samples were obtained <strong>on</strong> admissi<strong>on</strong> (0h), 24h, 48h<br />

and 72h after enrollment. Apoptosis in peripheral leukocytes was determined <strong>on</strong>ce in 19 healthy volunteers<br />

(H) (52±9). Leukocyte apoptosis was evaluated by flow cytometry. Early apoptotic intact cells (FITC + /PI - )<br />

were discriminated from late apoptotic cells lacking membrane integrity (FITC + /PI + ) by double staining with<br />

Annexin V-FITC and Propidium Iodine (PI). Results: Early apoptosis <str<strong>on</strong>g>of</str<strong>on</strong>g> lymphocytes was c<strong>on</strong>siderably<br />

decreased at 0h in AMI when compared to C and H group ([mean % ±SD] 6.3±5.1 vs 11.4±8.3, p=0.048<br />

261


23-26 August 2007,<br />

Budapest, Hungary<br />

and 6.4±3.7 vs 10.2±4.8, p=0.042 respectively). Similarly, early apoptosis <str<strong>on</strong>g>of</str<strong>on</strong>g> m<strong>on</strong>ocytes was decreased at 0h<br />

in AMI and UAP when compared to H group ([mean % ±SD] 42.3±26.4 vs 63.2±24.5, p=0.016 and<br />

42.9±20.3 vs 63.2±24.5, p=0.033 respectively). In c<strong>on</strong>trast, early apoptotic neutrophils were notably<br />

increased at 0h in AMI compared to UAP group (6.2±7.8 vs 3.1±3.1, p=0.042). Leukocyte apoptotic<br />

expressi<strong>on</strong> patterns were m<strong>on</strong>itored during the first 72 hours after admissi<strong>on</strong>. Late apoptotic lymphocytes<br />

increased at 72 hours after admissi<strong>on</strong> in AMI and UAP group. In c<strong>on</strong>trast, early apoptotic neutrophils were<br />

gradually decreased from admissi<strong>on</strong> (0h) in AMI by 3.39% (p=0.001) at 24h, 3.38% (p=0.026) at 48h and<br />

4.43% (p


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5D_04_P<br />

(poster secti<strong>on</strong> B1, poster board #203, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STUDYING QUINAZOLINONES DERIVATIVES TREATMENTS AS KIDNEY<br />

CELLS STRESS IN BALB/C NEW BORN MICE<br />

Samar Etemad, Maryam Shams Lahijani<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Science, Shahid Behehsti University<br />

Water insoluble hetrocylcic compounds, such as quinazolin<strong>on</strong>es are reported to having pharmacological<br />

effects such as anti-inflammati<strong>on</strong>, sedati<strong>on</strong>, anti-depressi<strong>on</strong>, anti-bacterial, anti-allergic, anti- blood pressur<br />

and aspects <str<strong>on</strong>g>of</str<strong>on</strong>g> their properties <strong>on</strong> reducing blood fat, inhibiting some proteins and specifially preventing cell<br />

movements, are the newest ways <str<strong>on</strong>g>of</str<strong>on</strong>g> treating cancer and HIV disease.<br />

Previous studies at the Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Science, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Shahid Behehsti, showed<br />

significant teratogenic effects <str<strong>on</strong>g>of</str<strong>on</strong>g> two new derivatives <str<strong>on</strong>g>of</str<strong>on</strong>g> quinazl<strong>on</strong>ines <strong>on</strong> skeletal and morphological<br />

structures (exencephaly, exophthalmia, microphthalmia, anophthalmia, disturbance in polarity <str<strong>on</strong>g>of</str<strong>on</strong>g> limbs<br />

and…). So, finding any effects at histological level would have been quite interesting results, regarding not<br />

observing any obvious disturbences in organs such as kindneys.Pregnant Balb/C mice(n=20) were divided<br />

into 3 groups <str<strong>on</strong>g>of</str<strong>on</strong>g>: c<strong>on</strong>trol, receiving <strong>on</strong>ly distilled water, sham, receiving 0.5% (methylcelloluse 9the solvent)<br />

and experimental groups, receiving 100mg/kg/ body weight <str<strong>on</strong>g>of</str<strong>on</strong>g> 4(3H) quinazolin<strong>on</strong>e-2-propyl-2-phenylethyl)<br />

and 4(3H)quinazl<strong>on</strong><strong>on</strong>es-2-ethyl-2-phenylethyl, by IP injecti<strong>on</strong>s, <strong>on</strong> day 8 th <str<strong>on</strong>g>of</str<strong>on</strong>g> gestati<strong>on</strong>. Kidneys were fixed<br />

in <str<strong>on</strong>g>for</str<strong>on</strong>g>maldehyde after birth and stained with H & E. C<strong>on</strong>firming the pathological results, <strong>on</strong>e- way ANOVA<br />

and Chi-Square tests were applied <str<strong>on</strong>g>for</str<strong>on</strong>g> quantitative and qualititative data (P


23-26 August 2007,<br />

Budapest, Hungary<br />

amphibians, but has been dem<strong>on</strong>strated in fish, reptiles, and mammals (including laboratory rodents and<br />

humans cell lines) as well. In additi<strong>on</strong>, several independent studies in amphibians and laboratory rodents have<br />

dem<strong>on</strong>strated now that atrazine also increases stress horm<strong>on</strong>e (glucocorticoid) synthesis and secreti<strong>on</strong> as well.<br />

Furthermore, when combined with other comm<strong>on</strong>ly applied pesticides, the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> atrazine are magnified.<br />

Exposed larvae suffer from retarded growth and development and immuno-suppressi<strong>on</strong> resulting in<br />

increased disease c<strong>on</strong>tracti<strong>on</strong> and increased mortality. These effects have been dem<strong>on</strong>strated in both the<br />

laboratory and in the field: Tadpoles downstream <str<strong>on</strong>g>of</str<strong>on</strong>g> agriculture show retarded growth and development and<br />

suffer high mortality rates in resp<strong>on</strong>se to pathogens relative to tadpoles upstream <str<strong>on</strong>g>of</str<strong>on</strong>g> agriculture. These effects<br />

indicate that pesticide c<strong>on</strong>taminati<strong>on</strong> may play a critical role in amphibian declines even in localities and<br />

incidences where populati<strong>on</strong> declines appear to be due to other causes such as disease.<br />

5D_06_P<br />

(poster secti<strong>on</strong> B1, poster board #205, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

OPPOSING EFFECTS OF β-ENDORPHIN AND RESTRAINT STRESS ON PAW<br />

INFLAMMATION REFLECT CHANGES IN THE FUNCTIONS OF<br />

INFLAMMATORY CELLS<br />

N. Kuštrimović, K. Mitić, S. Stanojević, M. Dimitrijević<br />

Immunology <str<strong>on</strong>g>Research</str<strong>on</strong>g> Center “Branislav Janković” at Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Virology,<br />

Vaccines and Sera “Torlak”, Belgrade, Serbia<br />

It is well established that stress- induced release <str<strong>on</strong>g>of</str<strong>on</strong>g> opioid peptides could affect development <str<strong>on</strong>g>of</str<strong>on</strong>g> inflammati<strong>on</strong><br />

via mechanisms involving changes in the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> inflammatory cells both locally and systemically. To<br />

compare the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> restraint stress (RS) and local injecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> opioid peptide <strong>on</strong> C<strong>on</strong>canavalin A-induced<br />

paw inflammati<strong>on</strong>, male Dark Agouti (DA) rats were either exposed to RS <str<strong>on</strong>g>for</str<strong>on</strong>g> 2h or intraplantarly treated<br />

with β-endorphin (β -End, 0.01-10 µg). While 0.01 µg <str<strong>on</strong>g>of</str<strong>on</strong>g> β -End increased paw edema, exposure <str<strong>on</strong>g>of</str<strong>on</strong>g> rats to 2h<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> RS significantly diminished it. In additi<strong>on</strong>, perit<strong>on</strong>eal macrophages from intact and previously stressed rats<br />

were tested <str<strong>on</strong>g>for</str<strong>on</strong>g> adherence and producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> reactive oxygen species (ROS) in the presence or absence <str<strong>on</strong>g>of</str<strong>on</strong>g> 10 -<br />

12 -10 -8 M <str<strong>on</strong>g>of</str<strong>on</strong>g> β-End in vitro. Results showed that β-End suppressed macrophages adherence and ROI<br />

producti<strong>on</strong>, in c<strong>on</strong>trast to RS that increased both macrophage functi<strong>on</strong>s. However, previous exposure to RS<br />

counteracted suppressive influence <str<strong>on</strong>g>of</str<strong>on</strong>g> β-End <strong>on</strong> macrophage adherence but did not change the suppressive<br />

effect <str<strong>on</strong>g>of</str<strong>on</strong>g> β-End <strong>on</strong> macrophage ROI producti<strong>on</strong>. Our results suggest two main c<strong>on</strong>clusi<strong>on</strong>s. First, ROI<br />

produced by inflammatory cells more likely suppress development <str<strong>on</strong>g>of</str<strong>on</strong>g> tissue inflammati<strong>on</strong> than promote tissue<br />

destructi<strong>on</strong> and swelling. Sec<strong>on</strong>d, exposure to RS can functi<strong>on</strong>ally alter macrophages and make them less<br />

sensitive to the in vitro treatment with β-End.<br />

264


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5D_07_P<br />

(poster secti<strong>on</strong> B1, poster board #206, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

LASER-INDUCED AND EXOGENOUSLY INTRODUCED HSP70 SERVE AS THE<br />

EFFICIENT ADJUVANTS FOR INFLUENZA VACCINE<br />

Sergey Onikienko 1 , Irina Guzhova 2 , Boris Margulis 2 , Alexander Zemlyanoi 3<br />

1 Military Medical Academy, St Petersburg, Russia, e-mail: <strong>on</strong>ik@nwgsm.ru<br />

2 Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Cytology RAS, St. Petersburg, Russia<br />

3 Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Military Medicine, St. Petersburg, Russia<br />

Hsp70 chaper<strong>on</strong>e is known to possess high adjuvant activity; <str<strong>on</strong>g>for</str<strong>on</strong>g> instance Hsp70 <str<strong>on</strong>g>of</str<strong>on</strong>g> Mycobacterium<br />

tuberculosis c<strong>on</strong>stitutes the active part <str<strong>on</strong>g>of</str<strong>on</strong>g> BCG vaccine. In order to assess the adjuvant ability <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 we<br />

used laser irradiati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> skin <str<strong>on</strong>g>of</str<strong>on</strong>g> internal surface <str<strong>on</strong>g>of</str<strong>on</strong>g> mouse ear followed by the injecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> anti-influenza<br />

vaccine Vaxigrip into the same site. According to Western blotting data Hsp70 almost completely<br />

disappeared from the area <str<strong>on</strong>g>of</str<strong>on</strong>g> the exposure to laser in comparis<strong>on</strong> with n<strong>on</strong>-treated site; 48 hours later the<br />

Hsp70 c<strong>on</strong>tent returned back to basal value. We suggest that the chaper<strong>on</strong>e can be released from laser-treated<br />

cells as shown earlier <str<strong>on</strong>g>for</str<strong>on</strong>g> other Hsp70-inducing factors. In another experimental setting pure Hsp70/Hsc70<br />

preparati<strong>on</strong> from bovine muscle was injected al<strong>on</strong>g with influenza vaccine. In both cases, stimulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

endogenous Hsp70 release or exogenous chaper<strong>on</strong>e delivery was sufficient to induce stable cellular and<br />

humoral resp<strong>on</strong>se to vaccine antigen as proved by in vitro and in vivo tests. Additi<strong>on</strong>ally, bacterial<br />

lipopolysaccharide (LPS) from Serratia marcescens (prodigiosin) modified by electr<strong>on</strong> beam or laser irradiati<strong>on</strong><br />

was also found to possess adjuvant activity to Vaxigrip vaccine, when employed al<strong>on</strong>e or with the exogenous<br />

Hsp70. The adjuvant effect <str<strong>on</strong>g>of</str<strong>on</strong>g> prodigiosin was due to activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> TLR2/4 and trans-activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> receptors<br />

to alpha-fetoprotein, GM-CSF and IL-2. In summary these data prove that the combinatory applicati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

chaper<strong>on</strong>es with modified LPS can be promising in the novel vaccine development.<br />

5D_08_P<br />

(poster secti<strong>on</strong> B1, poster board #207, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE ROLE OF APOPTOSIS IN IMMUNE SYSTEM RESPONSES TO ACUTE STRESS:<br />

IMPACT OF GENDER AND MENSTRUAL CYCLE<br />

Bilge Pehlivanoglu, Sibel Bayrak, Esin Ileri, Dicle Z. Balkanci<br />

Hacettepe University, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Physiology Ankara Turkey<br />

e-mail: pbilge@hacettepe.edu.tr<br />

It is evident that there are differences in immune variables under acute stress between genders and menstrual<br />

cycle phases. Marked impact <str<strong>on</strong>g>of</str<strong>on</strong>g> gender causes women to be more pr<strong>on</strong>e to stress-induced diseases.<br />

Involvement <str<strong>on</strong>g>of</str<strong>on</strong>g> the apoptosis in stress resp<strong>on</strong>se has been studied recently and has a potential to be <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

mechanisms in varying immune resp<strong>on</strong>se. We aimed to investigate the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> acute stress <strong>on</strong> apoptosis <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

immune cells in men and women and at different phases <str<strong>on</strong>g>of</str<strong>on</strong>g> the cycle. Healthy men (n=17) and women<br />

(n=16) volunteers (age 18-25) were subjected to Stroop color-word interference and cold pressor tests.<br />

Women tested both at follicular and luteal phases. Pre and post-test lymphocyte subsets and apoptotic<br />

lymphocytes were determined flowcytometrically. Menstrual phase was assured by plasma estrogen and<br />

progester<strong>on</strong>e levels. Stress resp<strong>on</strong>se was evaluated by blood pressure and heart rate measurements and<br />

plasma cortisol levels. Nitric oxide (NO) was measured by chemiluminescence. All the data was evaluated<br />

statistically. Acute stress decreased CD4+ cells in all groups, CD4+/CD8+ ratio in men and women at<br />

265


23-26 August 2007,<br />

Budapest, Hungary<br />

follicular phase. CD19+ cells were reduced in women at follicular phase and increased in men, whereas<br />

CD56+ cells were increased in women and decreased in men after the stress. Annexin V+ helper T cell ratio<br />

was higher in all groups in post-test samples. Acute stress resulted in increased NO levels in men. Stressrelated<br />

apoptosis <str<strong>on</strong>g>of</str<strong>on</strong>g> T cells can explain the depressed cellular immunity, diverse immune resp<strong>on</strong>ses in men<br />

and women at follicular and luteal phases which may be modulated by NO. Further studies can enlighten the<br />

diversity <str<strong>on</strong>g>of</str<strong>on</strong>g> immune resp<strong>on</strong>ses to stress between men and women via apoptosis.<br />

266<br />

5D_09_P<br />

(poster secti<strong>on</strong> B1, poster board #208, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

GALECTIN-1 REGULATES RADIATION-INDUCED FIBROBLAST<br />

DIFFERENTIATION<br />

Ji-Ye<strong>on</strong> Ahn, Sarah Park, Youngsoo Han, Ji-Young Shim, Ye<strong>on</strong>-Sook Yun, Jie-Young S<strong>on</strong>g<br />

Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Radiati<strong>on</strong> Immunology, Korea Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Radiological and Medical Sciences, 215-4 g<strong>on</strong>gneung-d<strong>on</strong>g,<br />

now<strong>on</strong>-ku, Seoul 139-706, Korea, (Corresp<strong>on</strong>dence; Jie-Young S<strong>on</strong>g, immu@kcch.re.kr)<br />

Radiati<strong>on</strong>-induced lung fibrosis is <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> severe side effects <str<strong>on</strong>g>for</str<strong>on</strong>g> radiotherapy in lung or breast cancer patients.<br />

We previously reported that Galectin-1 was significantly increased in radiati<strong>on</strong>-induced fibrotic lungs by<br />

proteomic analysis in vivo. In this study, we show that Galectin-1 mediates radiati<strong>on</strong>-induced differentiati<strong>on</strong><br />

from fibroblast into my<str<strong>on</strong>g>of</str<strong>on</strong>g>ibroblast characterized by accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> smooth muscle alpha actin (α-SMA) and<br />

collagen type I. Morphological changes <str<strong>on</strong>g>of</str<strong>on</strong>g> WI38 human lung fibroblast cells were observed from day 2 after<br />

4Gy irradiati<strong>on</strong> and sustained up to 9 days. Galectin-1 expressi<strong>on</strong> was up-regulated by radiati<strong>on</strong>, and<br />

knockdown <str<strong>on</strong>g>of</str<strong>on</strong>g> endogenous Galectin-1 by specific short hairpin RNA (shRNA) significantly reduced α-SMA<br />

synthesis as well as switched the phenotype <str<strong>on</strong>g>of</str<strong>on</strong>g> my<str<strong>on</strong>g>of</str<strong>on</strong>g>ibroblast. Exposure to radiati<strong>on</strong> <strong>on</strong> WI38 cells also<br />

activated TGF-β secreti<strong>on</strong>, which is a major pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ibrotic cytokine and a crucial mediator <str<strong>on</strong>g>of</str<strong>on</strong>g> fibrosis in many<br />

different tissues, and stimulated its downstream signaling molecule Smad-2 and -3 activati<strong>on</strong>. However,<br />

knockdown <str<strong>on</strong>g>of</str<strong>on</strong>g> Galectin-1 did not affect the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> Smad-2, -3 or inhibitory Smad-7 expressi<strong>on</strong>s. These data<br />

taken together suggest that galectin-1 could regulate radiati<strong>on</strong>-induced fibroblast differentiati<strong>on</strong> via Smadindependent<br />

pathway and may provide an alternative approach to the treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> lung fibrosis.<br />

5D_10_P<br />

(poster secti<strong>on</strong> B1, poster board #209, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PILOTING ANTIGEN TO CROSS-PRESENTATION PATHWAY BY HEAT SHOCK<br />

PROTEIN<br />

Yasuaki Tamura, Goro Kutomi, Kenjirou Kamiguchi, Toshihiko Torigoe, Noriyuki Sato<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Pathology, Sapporo Medical University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine<br />

Recent evidences have been indicating that Heat shock proteins (HSPs) play an important role as a “danger<br />

signal” in the extracellular milieu <strong>on</strong> behalf <str<strong>on</strong>g>of</str<strong>on</strong>g> immune surveillance. Above all, Hsp70 and Hsp90 elicit<br />

intriguing efficient CTL resp<strong>on</strong>ses by so called “cross-presentati<strong>on</strong>” with yet entirely unknown mechanism.<br />

Here, we discuss that the immunologic roles <str<strong>on</strong>g>of</str<strong>on</strong>g> HSPs, particularly Hsp90, in the MHC class I-restricted crosspresentati<strong>on</strong><br />

by b<strong>on</strong>e marrow-derived dendritic cells (DCs). We show that Hsp90-peptide complex enter the<br />

endocytic pathway via putative Hsp90 receptor and associated peptide might be transferred <strong>on</strong>to endosomal<br />

MHC class I molecules. Moreover, we show that immunizati<strong>on</strong> with Hsp90-peptide complex efficiently elicits


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

CTL resp<strong>on</strong>ses and antitumor effect. Interestingly, this presentati<strong>on</strong> is TAP-independent, but rather follows<br />

endocytic pathway. Meanwhile, when Hsp90-whole protein (OVA) antigen complex were pulsed to DCs, this<br />

protein antigen could enter at least in part via TAP-dependent pathway to the ER, and finally was presented<br />

to MHC class I molecules. However, OVA al<strong>on</strong>e without Hsp90 could not enter into this pathway, but rather<br />

into MHC class II pathway.<br />

Here we discuss novel insights into the immunologic role <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90 in cross-presentati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> antigens,<br />

efficient inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> MHC class I-restricted CTL resp<strong>on</strong>ses, and applicati<strong>on</strong> to peptide/protein antigenbased<br />

immunotherapy <str<strong>on</strong>g>of</str<strong>on</strong>g> cancers.<br />

5D_11_P<br />

(poster secti<strong>on</strong> B1, poster board #210, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STUDY ON THE ACTION MECHANISM OF “SHUGAN LIFEI” THERAPY ON<br />

EXPERIMENTAL RATS WITH ASTHMA AND UNDER STRESS<br />

T<strong>on</strong>g Y. 1* , Zhang N. X. 2 , Liu Q. 1<br />

School <str<strong>on</strong>g>of</str<strong>on</strong>g> Chinese Medicine, the University <str<strong>on</strong>g>of</str<strong>on</strong>g> H<strong>on</strong>g K<strong>on</strong>g<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Rehabilitati<strong>on</strong>, Pu Tuo Hospital,<br />

Shanghai University <str<strong>on</strong>g>of</str<strong>on</strong>g> Traditi<strong>on</strong>al Chinese Medicine<br />

*e-mail: t<strong>on</strong>gyao@hku.hk<br />

Traditi<strong>on</strong>al Chinese medicine (TCM) has l<strong>on</strong>g realized that an unpleasant envir<strong>on</strong>ment or mental stimulati<strong>on</strong><br />

is closely related to the occurrence and progressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> disease. In TCM, there<str<strong>on</strong>g>for</str<strong>on</strong>g>e, it is viewed that “Gan” can<br />

regulate psychological status. In additi<strong>on</strong>, numerous studies believe that the functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the “Gan” in TCM is<br />

closely related to the neuroendocrine-immune network <str<strong>on</strong>g>of</str<strong>on</strong>g> modern medicine. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, this study discusses<br />

the influence <str<strong>on</strong>g>of</str<strong>on</strong>g> psychological factors <strong>on</strong> the occurrence <str<strong>on</strong>g>of</str<strong>on</strong>g> asthma and the acti<strong>on</strong> mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> “ShuGan”<br />

therapy in regulating stress to prevent and treat asthma. This is d<strong>on</strong>e through the use <str<strong>on</strong>g>of</str<strong>on</strong>g> “ShuGan” and<br />

“LiFei” (lung-regulating) therapy to treat rats with asthma and under stress.<br />

Objective: To discuss the influence <str<strong>on</strong>g>of</str<strong>on</strong>g> psychological factors <strong>on</strong> asthma occurrence and the acti<strong>on</strong> mechanism<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> “ShuGan LiFei” prescripti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> TCM in regulating stress to prevent and treat asthma.<br />

Results: “ShuGan LiFei” therapy could reduce plasma corticoster<strong>on</strong>e, decrease CRH positive neur<strong>on</strong>s in the<br />

paraventricular nucleus <str<strong>on</strong>g>of</str<strong>on</strong>g> hypothalamus, up-regulate the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> GRmRNA in hippocampus CA3, and<br />

alleviate hippocampal neur<strong>on</strong>al lesi<strong>on</strong>. Moreover, the therapy could significantly alleviate pulm<strong>on</strong>ary<br />

pathological changes in rats with asthma under the c<strong>on</strong>diti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> restraint stress through increasing CD 4 + ,<br />

CD 4+ /CD 8<br />

+<br />

in peripheral blood and plasma IFN-γ, and decreasing plasma IL-4.<br />

C<strong>on</strong>clusi<strong>on</strong>s: “ShuGan LiFei” therapy may regulate the hyperfuncti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HPA axis and the immune<br />

dysfuncti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> asthma in rats under the c<strong>on</strong>diti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> restraint stress.<br />

267


23-26 August 2007,<br />

Budapest, Hungary<br />

268<br />

5D_12_P<br />

(poster secti<strong>on</strong> B1, poster board #211, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HSP70 IS AN ANTIGENIC CHAPERONE SERVING FOR BOTH INTRACELLULAR<br />

AND EXTRACELLULAR ANTIGEN PRESENTATION<br />

Toshihiko Torigoe, Yasuaki Tamura, Kenjiro Kamiguchi, Atsushi Sugawara, Shin Ohshima,<br />

Takehiro Kurotaki, Noriyuki Sato<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Pathology, Sapporo Medical University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, S-1 W-17 Chuo-ku 060-8556, Japan<br />

E- mail: torigoe@sapmed.ac.jp<br />

It is speculated that Hsp70 serves as an antigenic chaper<strong>on</strong>e in the cytosol <str<strong>on</strong>g>for</str<strong>on</strong>g> MHC class I antigen<br />

presentati<strong>on</strong>. However, there have been <strong>on</strong>ly a few evidences that support the hypothesis. We have found<br />

that HSP70 was associated with TAP in cells. In the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> ATP, HSP70 was dissociated from TAP,<br />

indicating that peptide-binding <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 might be preferentially bound to TAP and peptide-free <str<strong>on</strong>g>for</str<strong>on</strong>g>m<br />

might be detached from TAP. In additi<strong>on</strong>, we found that Hsp70-binding polyamine compound<br />

deoxyspergualine (DSG) could inhibit the associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 with TAP. TAP-mediated transfer <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

cytosolic peptides was inhibited in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> DSG and cell surface level <str<strong>on</strong>g>of</str<strong>on</strong>g> MHC class I molecules was<br />

decreased to almost the equal level <str<strong>on</strong>g>of</str<strong>on</strong>g> those <strong>on</strong> TAP-deficient cells. By using radio-labeled antigenic peptides,<br />

it was shown that cytosolic peptides with high affinity to Hsp70 could be transported into the ER by TAP<br />

more efficiently than those with low affinity. Taken together, our data str<strong>on</strong>gly suggest that cytosolic Hsp70<br />

serves as a chaper<strong>on</strong>e <str<strong>on</strong>g>for</str<strong>on</strong>g> cytosolic peptides and facilitate the antigen presentati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the client peptides.<br />

Stimulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> dendritic cells (DC) with Hsp70 induced activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> DC, leading to TNF release. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e,<br />

extracellular Hsp70 has a cytokine-like functi<strong>on</strong> that activates innate immunity. When mice were immunized<br />

with Hsp70-binding peptides in combinati<strong>on</strong> with Hsp70, antigen-specific cytotoxic T cells (CTL) was<br />

efficiently induced. In c<strong>on</strong>trast, immunizati<strong>on</strong> with n<strong>on</strong>-binding peptides in combinati<strong>on</strong> with Hsp70 failed<br />

to elicit the CTL resp<strong>on</strong>se. These data indicate that extracellular Hsp70 might serve as a chaper<strong>on</strong>e <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

extracellular antigens. Hsp70 might facilitate the cross-presentati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> client peptides to MHC class I<br />

molecules in DC, thus activating adaptive immunity as well as innate immunity. Interestingly, the binding <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Hsp70 to the cell surface Hsp-receptors <strong>on</strong> DC was inhibited in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> DSG. It is there<str<strong>on</strong>g>for</str<strong>on</strong>g>e<br />

suggested that immunosuppressive and anti-inflammatory acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> DSG may be explained at least in part by<br />

the suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> both intracellular and extracellular antigenic chaper<strong>on</strong>e functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70. Important<br />

roles <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 as an antigenic chaper<strong>on</strong>e in the immune system and host defense system would be<br />

highlighted in this presentati<strong>on</strong>.<br />

5D_13_P<br />

(poster secti<strong>on</strong> B1, poster board #212, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HSF2 AS A REGULATOR OF CYTOKINE EXPRESSION IN SPERMATOGENESIS<br />

Anniina Vihervaara, Malin Åkerfelt, Eva Henrikss<strong>on</strong>, Lea Sist<strong>on</strong>en<br />

Turku <str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g> Biotechnology and Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, Åbo Akademi University,<br />

BioCity, Tykistökatu 6, 20520 Turku, Finland<br />

e-mail: aviherva@btk.fi<br />

Heat shock factor 2 (HSF2) bel<strong>on</strong>gs to the family <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock transcripti<strong>on</strong> factors (HSFs), the members <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

which orchestrate cellular stress resp<strong>on</strong>ses and participate in processes <str<strong>on</strong>g>of</str<strong>on</strong>g> development and differentiati<strong>on</strong>.<br />

While HSF1 is the major executor <str<strong>on</strong>g>of</str<strong>on</strong>g> the heat shock resp<strong>on</strong>se, HSF2 is involved in germ cell differentiati<strong>on</strong>


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

and in cortical development. To investigate the targets <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF2 in mouse spermatogenesis, a ChIP-<strong>on</strong>-chip<br />

screen <strong>on</strong> whole testis was carried out. Our results reveal HSF2 binding <strong>on</strong> various cytokine genes, and show<br />

the corresp<strong>on</strong>ding proteins to be differentially expressed in wt and Hsf2 -/- mice. Since HSF2 cooperates with<br />

and is regulated by HSF1 during heat shock, we investigated the possible influence <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF1 <strong>on</strong> the novel<br />

HSF2 targets. Whereas certain cytokine genes are occupied by both factors, we, interestingly, also discovered<br />

HSF2 targets that are not bound by HSF1. Accordingly, similar protein levels <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF2-specific target genes<br />

were detected in wt and Hsf1 -/- mice, which indicates gene-specific effects <str<strong>on</strong>g>for</str<strong>on</strong>g> HSF1 and HSF2. Diverse<br />

functi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF1 and HSF2 are further supported by their different expressi<strong>on</strong> patterns in seminiferous<br />

tubules where HSF2 is expressed in a stage-dependent manner and the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF1 remain relatively<br />

c<strong>on</strong>stant throughout spermatogenesis. In testis, cytokines are important <str<strong>on</strong>g>for</str<strong>on</strong>g> germ cell migrati<strong>on</strong> and <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

creating and maintaining the site <str<strong>on</strong>g>of</str<strong>on</strong>g> immune privilege. Furthermore, the detected cytokine envir<strong>on</strong>ment in<br />

Hsf2 -/- mice is associated with testicular inflammati<strong>on</strong> and infertility. Thus, HSF2-regulated cytokine<br />

expressi<strong>on</strong> elucidates novel testicular and immunological functi<strong>on</strong>s, emphasizing the multifaceted character <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

HSF2.<br />

5D_14_P<br />

(poster secti<strong>on</strong> B1, poster board #213, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EUKARYOTICALLY EXPRESSED, ENDOTOXIN-FREE RECOMBINANT HUMAN<br />

HSP70 ENHANCES UPTAKE OF ANTIGENIC PEPTIDES IN MACROPHAGES AND<br />

PRIMARY HUMAN EPITHELIAL CELLS<br />

D<strong>on</strong>g Wang#, Ralf Dressel, Rainer Blasczyk*, Thomas Werfel#, Britta Eiz-Vesper*,<br />

Miriam Wittmann#<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Dermatology# and Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Transfusi<strong>on</strong> Medicine*, Hannover Medical School, Germany<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Cellular and Molecular Immunology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Goettingen, Germany<br />

Stress-inducible heat shock protein 70 (HSP70/HSP72) has been shown to enhance or enable inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

antigen-specific T cell resp<strong>on</strong>ses. Recently, the ability <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70/peptide complexes to elicit CTL resp<strong>on</strong>ses<br />

by cross-presentati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> exogenous antigen via HLA class I has been coming into focus <str<strong>on</strong>g>of</str<strong>on</strong>g> immunotherapy.<br />

Little is known about differences in the functi<strong>on</strong> between pro- and eukaryotically expressed HSP70 used <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

the in vitro inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> specific CTLs. When prokaryotically expressed HSP70 is used, endotoxin<br />

c<strong>on</strong>taminati<strong>on</strong> is always a major c<strong>on</strong>cern. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, we expressed HSP70 eukaryotically to analyse the role <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

endotoxin-free HSP70 in the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> specific CTLs. FITC-labelled HSP70 could be shown to be rapidly<br />

taken up by m<strong>on</strong>ocytes and macrophages <str<strong>on</strong>g>of</str<strong>on</strong>g> the peripheral blood as well as by primary human keratinocytes.<br />

In our endotoxin-free system uptake <str<strong>on</strong>g>of</str<strong>on</strong>g> Tamra-labelled n<strong>on</strong>apeptides <str<strong>on</strong>g>of</str<strong>on</strong>g> different specificity was significantly<br />

enhanced in peripheral blood m<strong>on</strong>ocytes as well as in keratinocytes in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> rhHSP70. rhHSP70<br />

could be shown to enhance cross-presentati<strong>on</strong> to CD8 T-cells in different experimental approaches.<br />

Preliminary results support the predominant role <str<strong>on</strong>g>of</str<strong>on</strong>g> CD91 as HSP70 receptor. Our results indicate that<br />

eukaryotically expressed HSP70/peptide complexes could be a useful tool to generate antigen specific CD8 T<br />

cell resp<strong>on</strong>ses.<br />

269


23-26 August 2007,<br />

Budapest, Hungary<br />

5D_16_P<br />

(poster secti<strong>on</strong> B1, poster board #214, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HSP70 POLYMORPHISMS ARE GENETICALLY ASSOCIATED WITH SYSTEMIC<br />

LUPUS ERYTHEMATOSOUS<br />

Barbara G. Fürnrohr, Sven Wach, Rüdiger Müller, Christian Stach, Thomas Winkler,<br />

Reinhard E. Voll<br />

IZKF <str<strong>on</strong>g>Research</str<strong>on</strong>g> Group 2, Nikolaus-Fiebiger-Center, Glückstrasse 6, 91054 Erlangen, Germany, Ph<strong>on</strong>e: +49-<br />

9131-85-39303, FAX: +49-9131-85-39311, Corresp<strong>on</strong>ding author: Barbara Fürnrohr<br />

e-mail: bfuernro@molmed.uni-erlangen.de<br />

Systemic lupus erythematosous (SLE) is an autoimmune disease <str<strong>on</strong>g>of</str<strong>on</strong>g> unknown origin characterised by defecitve<br />

immune regulatory mechanisms. Complicated interacti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> various genetic and envir<strong>on</strong>mental factors are<br />

discussed to be relevant <str<strong>on</strong>g>for</str<strong>on</strong>g> the manifestati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the disease. Since heat shock proteins (Hsps) are thought to<br />

play a role in multifactorial diseases such as SLE, we investigated the three cytosolic Hsp70 genes. Genetic<br />

variants <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70-Hom, Hsp70A1A and Hsp70A1B were analysed, which are localised in the SLEsusceptibility<br />

locus <strong>on</strong> chromosome 6p21.3. In a case-c<strong>on</strong>trol study using a haplotype-tagging SNP approach,<br />

we identified a polymorphism in the regi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70A1B, which was significantly associated (p = 0.0086)<br />

with SLE. Evaluati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> clinical data has shown that patients exhibiting the significantly associated haplotype<br />

c<strong>on</strong>tain higher titers <str<strong>on</strong>g>of</str<strong>on</strong>g> autoantibodies to dsDNA, Ro, and La. Our results may help to gain new insights into<br />

the pathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> SLE and, possibly, to develop novel therapeutic strategies.<br />

270


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5E. ISN SYMPOSIUM ON STRESS ENDOCRINOLOGY<br />

(FRANCOIS TRONCHE)<br />

5E_01_S<br />

THE MECHANISMS BEHIND THE BRAIN’S RESPONSE TO STRESS<br />

Marian Joëls<br />

SILS-CNS, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Amsterdam, The Netherlands<br />

When an organism experiences a stressful situati<strong>on</strong>, this is perceived through the brain and thus gives rise to<br />

the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the sympathetic nervous system and the hypothalamo-pituitary-adrenal axis. As a result brain<br />

cells, including those involved in the initial appreciati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the stressor, are exposed to elevated levels <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

m<strong>on</strong>oamines like noradrenaline, to specific neuropeptides and to corticosteroids. We have examined the<br />

effects <str<strong>on</strong>g>of</str<strong>on</strong>g> these horm<strong>on</strong>es <strong>on</strong> neur<strong>on</strong>al functi<strong>on</strong> in limbic regi<strong>on</strong>s, particularly the hippocampal CA1 area. In<br />

the initial phase after stress (


23-26 August 2007,<br />

Budapest, Hungary<br />

localized receptor in isolated NPC from the adult hippocampus. Treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> NPC grown in vitro with GCs<br />

induces decreased proliferati<strong>on</strong> index, and a down-regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Nestin, a protein marker that is down<br />

regulated as NPC stop dividing and differentiate. This resp<strong>on</strong>se is blocked using the GR-specific antag<strong>on</strong>ist<br />

indicating that the GCs resp<strong>on</strong>se is mediated by the glucocorticoid receptor. We dem<strong>on</strong>strate decreased<br />

proliferati<strong>on</strong> and a shift in cell fate choice in NPC following GCs treatment, which is blocked by the anti-<br />

GCs genetic manipulati<strong>on</strong>. The apparent resp<strong>on</strong>siveness <str<strong>on</strong>g>of</str<strong>on</strong>g> NPC to GCs suggests that neurogenesis is directly<br />

modulated via GR signalling pathways.<br />

5E_03_S<br />

STRESS AND TRANSCRIPTION: SPECIFIC INACTIVATION OF THE<br />

GLUCOCORTICOID RECEPTOR GENE IN THE DOPAMINERGIC SYSTEM: NEW<br />

INSIGHTS ON DRUG ADDICTION<br />

Francois Tr<strong>on</strong>che<br />

CNRS UMR7148 « Molecular Genetics, Neurophysiology and Behavior », Collège de France, 75005 Paris France<br />

e-mail: francois.tr<strong>on</strong>che@gmail.com<br />

A dysfuncti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress resp<strong>on</strong>se is suspected in the etiology <str<strong>on</strong>g>of</str<strong>on</strong>g> metabolic and behavioral disorders (ie<br />

anxiety, depressi<strong>on</strong>, drug dependence). A major comp<strong>on</strong>ent <str<strong>on</strong>g>of</str<strong>on</strong>g> this resp<strong>on</strong>se involves the release <str<strong>on</strong>g>of</str<strong>on</strong>g> adrenal<br />

glucocorticoids (GCs) and the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the glucocorticoid receptor (GR), a transcripti<strong>on</strong> factor.<br />

To study GR functi<strong>on</strong> in vivo, we develop and analyze animal models lacking GR in given brain cell<br />

populati<strong>on</strong>s. Molecular, physiological, electrophysiological and behavioral studies <str<strong>on</strong>g>of</str<strong>on</strong>g> these animals should<br />

allow us to locate the cell types involved and to better define the mechanisms underlying GR functi<strong>on</strong> in<br />

brain physiology and in pathological situati<strong>on</strong>s. We previously showed that the selective inactivati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

glucocorticoid receptor (GR) gene in mice brains (GRNesCre) pr<str<strong>on</strong>g>of</str<strong>on</strong>g>oundly reduces motivati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> cocaine.<br />

More recently, we showed that these behavioural effects are associated with a change in the impulse activity<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> midbrain dopamine neur<strong>on</strong>es. To determine in which cell type the functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> GR is required to modulate<br />

motivati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> cocaine, we generated animal models in which GR is selectively inactivated in either presynaptic<br />

dopamine neur<strong>on</strong>s (GRDATCre) or post-synaptic cells (GRD1Cre). Characterizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these<br />

models will be presented. To address the questi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the interacti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> GCs and serot<strong>on</strong>ergic pathway, we<br />

generated a mouse transgenic line that allow Cre recombinati<strong>on</strong> in all 5-HT1A neur<strong>on</strong>es and obtained<br />

c<strong>on</strong>diti<strong>on</strong>al GR inactivati<strong>on</strong>. Analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> this animal model will be presented.<br />

5E_04_S<br />

GLUCOCORTICOIDS AND PANCREAS DEVELOPMENT: STUDIES IN RODENTS<br />

AND HUMAN<br />

Bernadette Breant<br />

<str<strong>on</strong>g>Centre</str<strong>on</strong>g> de Recherche des Cordeliers, UMRS 872 - Université Paris VI, INSERM U872, Paris, France<br />

Low birth weight is str<strong>on</strong>gly predictive <str<strong>on</strong>g>of</str<strong>on</strong>g> hypertensi<strong>on</strong>, cardiovascular diseases, obesity, insulin resistance and<br />

diabetes. The mechanisms by which fetal undernutriti<strong>on</strong> or low birth weight, increases the risk <str<strong>on</strong>g>of</str<strong>on</strong>g> developing<br />

these diseases are unclear. In a rat model <str<strong>on</strong>g>of</str<strong>on</strong>g> undernutriti<strong>on</strong>, involving an overall reducti<strong>on</strong> in maternal food<br />

intake, we investigated the hypothesis <str<strong>on</strong>g>of</str<strong>on</strong>g> a primary defect in beta-cell development. In this model, fetuses<br />

with intrauterine growth retardati<strong>on</strong> show decreased beta-cell mass, which persists into adulthood and<br />

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3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

ultimately causes glucose intolerance, thereby mimicking features <str<strong>on</strong>g>of</str<strong>on</strong>g> the metabolic syndrome. Maternal<br />

undernutriti<strong>on</strong> caused glucocorticoid elevati<strong>on</strong>, which, in turn, causes a reducti<strong>on</strong> in beta-cell mass in the<br />

fetuses. Our data also suggest a key role <str<strong>on</strong>g>of</str<strong>on</strong>g> glucocorticoids under normal nutriti<strong>on</strong>al supply. By combining invitro<br />

studies with in-vivo investigati<strong>on</strong>s in mice lacking the glucocorticoid receptor (GR) in the whole<br />

organism or in specific pancreatic cell populati<strong>on</strong>s, we show that the GR is critical <str<strong>on</strong>g>for</str<strong>on</strong>g> pancreatic architecture<br />

and survival, as well as <str<strong>on</strong>g>for</str<strong>on</strong>g> beta-cell mass expansi<strong>on</strong> during a critical developmental window. Glucocorticoids<br />

act <strong>on</strong> precursor cells be<str<strong>on</strong>g>for</str<strong>on</strong>g>e the <strong>on</strong>set <str<strong>on</strong>g>of</str<strong>on</strong>g> horm<strong>on</strong>e gene expressi<strong>on</strong> and are likely to programme beta-cell<br />

differentiati<strong>on</strong> by modifying the balance <str<strong>on</strong>g>of</str<strong>on</strong>g> specific transcripti<strong>on</strong> factors. To investigate whether this would<br />

also apply to human pancreatic development, we study the expressi<strong>on</strong> and localizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the GR and that <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

various transcripti<strong>on</strong> factors and pancreatic horm<strong>on</strong>es <strong>on</strong> fetal pancreatic specimens. The results are<br />

c<strong>on</strong>sistent with a possible role <str<strong>on</strong>g>for</str<strong>on</strong>g> glucocorticoids during human pancreatic development.<br />

5E_05_S<br />

(poster secti<strong>on</strong> B1, poster board #242, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

INTERPLAY OF STRESS-RELATED HORMONES IN THE CONTROL OF<br />

INSECT FITNESS<br />

N. E. Gruntenko, N. A. Chentsova, N. V. Ad<strong>on</strong>yeva, E. K. Karpova, E. V. Bogomolova,<br />

I. Yu. Rauschenbach<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Cytology and Genetics SD RAS, Novosibirsk 630090, Russia<br />

e-mail: nataly@bi<strong>on</strong>et.nsc.ru<br />

In insects, biogenic amines, dopamine (DA) and octopamine (OA), and g<strong>on</strong>adotropins, juvenile horm<strong>on</strong>e<br />

(JH) and 20-hydroxyecdys<strong>on</strong>e (20E), are the main comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g> neurohorm<strong>on</strong>al stress-reacti<strong>on</strong>. For the<br />

progress <str<strong>on</strong>g>of</str<strong>on</strong>g> oogenesis in Drosophila under normal and stress c<strong>on</strong>diti<strong>on</strong>s a proper balance between JH and 20E<br />

is <str<strong>on</strong>g>of</str<strong>on</strong>g> a paramount importance: imbalance (a shift <str<strong>on</strong>g>of</str<strong>on</strong>g> the balance in the directi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> JH or 20E) leads to<br />

dramatic changes in oogenesis and fecundity (Soller et al., 1999; Gruntenko et al., 2003, 2005; Rauschenbach<br />

et al., 2004). Here we dem<strong>on</strong>strate that (I) D. virilis and D. melanogaster females possess a mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

reciprocal regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> JH and 20E which is essential to maintain the JH/20E balance: (a) a rise in JH titre<br />

leads to a rise in the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> ecdys<strong>on</strong>e 20-m<strong>on</strong>ooxygenase (which c<strong>on</strong>verts ecdys<strong>on</strong>e to 20E) and their 20E<br />

titre; (b) a rise in 20E titre results in a dose-dependent rise in JH levels; (c) 20E regulates JH indirectly via DA<br />

metabolic system - a rise in 20E titre increases DA c<strong>on</strong>tent in young females and decreases it in mature<br />

females, thus leading to a rise in JH levels in both; (d) there is a feedback in the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> JH by DA - a<br />

rise in JH titre leads to a decrease in DA in young females and its rise in mature females. (II) Under normal<br />

c<strong>on</strong>diti<strong>on</strong>s, reproducti<strong>on</strong> is regulated by genes that c<strong>on</strong>trol DA metabolic pathways (indirectly via JH); under<br />

unfavorable c<strong>on</strong>diti<strong>on</strong>s, reproducti<strong>on</strong> is regulated by genes that c<strong>on</strong>trol OA metabolism. (III) Individual<br />

stress-resistance depends <strong>on</strong> expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> genes c<strong>on</strong>trolling the background level <str<strong>on</strong>g>of</str<strong>on</strong>g> DA.<br />

273


23-26 August 2007,<br />

Budapest, Hungary<br />

5E_06_S<br />

(poster secti<strong>on</strong> B1, poster board #243, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DISSECTING CRHR1-MEDIATED PATHWAYS VIA MICROARRAY TECHNOLOGY<br />

C. Graf, P. Weber, B. Pütz, F. Holsboer, W. Wurst, J. M. Deussing<br />

Max-Planck-Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychiatry, Kraepelinstr. 2, 80804 München<br />

e-mail: cgraf@mpipsykl.mpg.de<br />

The corticotropin releasing horm<strong>on</strong>e (CRH) system is involved in endocrine, aut<strong>on</strong>omic and behavioural<br />

resp<strong>on</strong>ses to stress. The biological acti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> CRH-like neuropeptides are mediated by G-protein-coupled<br />

receptors, CRH receptor 1 (CRHR1) and CRHR2. CRHR1 is widely expressed in the mammalian brain (e.g.<br />

cerebral cortex, cerebellum, amygdala, hippocampus) and in the pituitary gland. Mice deficient <str<strong>on</strong>g>for</str<strong>on</strong>g> CRHR1<br />

display decreased anxiety-like behaviour and dysregulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the hypothalamic-pituitary-adrenal (HPA) axis.<br />

Ligand binding increases the affinity <str<strong>on</strong>g>of</str<strong>on</strong>g> the CRHR to G-proteins. Binding <str<strong>on</strong>g>of</str<strong>on</strong>g> a Gαs-protein will activate<br />

adenylate cyclase (AC) and protein kinase A (PKA) as well as other cyclic adenosine m<strong>on</strong>ophoshpate (cAMP)<br />

dependent pathways. The reported coupling <str<strong>on</strong>g>of</str<strong>on</strong>g> additi<strong>on</strong>al G-proteins to CRHR1 suggests that also other<br />

sec<strong>on</strong>d messengers are involved in CRHR-signaling. In order to identify specific target genes <str<strong>on</strong>g>of</str<strong>on</strong>g> CRHR1-<br />

mediated signaling pathways we applied cDNA and olig<strong>on</strong>ucleotide microarray technology using a mouse<br />

corticotroph cell line (AtT20) and pituitaries <str<strong>on</strong>g>of</str<strong>on</strong>g> CRHR1-deficient mice. 102 genes in vitro and more than 400<br />

genes in vivo were found stress- and/or CRH- and CRHR1-dependently regulated. A subset <str<strong>on</strong>g>of</str<strong>on</strong>g> candidate<br />

genes was validated in independent material by quantitative real time PCR (qRT-PCR). These candidates are<br />

involved e.g. in cAMP, mitogen activated protein kinase (MAPK) or epidermal growth factor receptor<br />

(EGFR) signaling. In order to examine their functi<strong>on</strong>al role the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> these genes <strong>on</strong> different known<br />

target genes downstream <str<strong>on</strong>g>of</str<strong>on</strong>g> CRHR1 signaling is analyzed using reporter assays.<br />

274


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5E_02_P<br />

(poster secti<strong>on</strong> B1, poster board #244, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

FIRING INCREASES IN AUTONOMIC PERIPHERAL NERVES AND C-FOS<br />

EXPRESSION IN HYPOTHALAMUS AND MEDULLA DURING SEIZURES IN RATS<br />

Kiyomi Koizumi, Rena Orman, Mark Stewart<br />

SUNY Downstate Medical Center, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Physiology & Pharmacology, Brooklyn, NY, USA<br />

Aut<strong>on</strong>omic nervous system c<strong>on</strong>sequences <str<strong>on</strong>g>of</str<strong>on</strong>g> seizures range from mild (e.g. sweating, piloerecti<strong>on</strong>) to severe<br />

(Sudden Unexplained Death in EPilepsy, SUDEP), but the severity <str<strong>on</strong>g>of</str<strong>on</strong>g> aut<strong>on</strong>omic disturbances are poorly<br />

correlated with seizure severity. We used urethane anesthetized rats to study the spread <str<strong>on</strong>g>of</str<strong>on</strong>g> kainic acidinduced<br />

limbic seizure activity into hypothalamic, medullary, and peripheral aut<strong>on</strong>omic relays. Peripheral<br />

nerve activity was recorded with a linear array electrode. ECG and blood pressure were c<strong>on</strong>tinuously<br />

m<strong>on</strong>itored.<br />

Peripheral multi- and single-unit activity, recorded from right or left vagus nerve, cervical sympathetic<br />

gangli<strong>on</strong>, and renal sympathetic nerve was elevated during each recurrent seizure. Activity recorded from the<br />

splanchnic nerve was not changed substantially. Increases in firing were sustained during seizures <str<strong>on</strong>g>for</str<strong>on</strong>g> vagus<br />

nerve in particular, whereas short episodes <str<strong>on</strong>g>of</str<strong>on</strong>g> increased activity were most frequently observed in renal<br />

sympathetic nerve recordings. Baroreceptor reflex resp<strong>on</strong>ses to phenylephrine or nitroprusside were in the<br />

right directi<strong>on</strong>, but prol<strong>on</strong>ged.<br />

Increased expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> c-fos was detected in aut<strong>on</strong>omic and neuroendocrine parts <str<strong>on</strong>g>of</str<strong>on</strong>g> paraventricular nucleus<br />

and in the supraoptic nucleus. Increased expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> c-fos was also detected in the rostral ventrolateral<br />

medulla and dorsal motor nucleus <str<strong>on</strong>g>of</str<strong>on</strong>g> the vagus. Increases in expressi<strong>on</strong> were comparable in the right and left<br />

hemispheres.<br />

We c<strong>on</strong>clude that aut<strong>on</strong>omic disturbances that occur during seizures will be “balanced” if seizure activity is<br />

bilateral. We suggest that the most sever disrupti<strong>on</strong>s will occur when the seizure activity, or its pattern <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

spread, results in a transient unilateral over-activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> hypothalamic or medullary brain regi<strong>on</strong>s. On the<br />

other hand, we suggest that the neuroendocrine c<strong>on</strong>sequences <str<strong>on</strong>g>of</str<strong>on</strong>g> seizures will be correlated with seizure<br />

severity, and will likely result in significant gender differences in seizure patients.<br />

5E_03_P<br />

(poster secti<strong>on</strong> B1, poster board #245, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ENDOCRINE STRESS RESPONSE INDUCED BY HYPOBARIC-HYPOXIA<br />

S. Sime<strong>on</strong>i, P. M<strong>on</strong>nazzi, R. Biselli * , F. R. Patacchioli<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Human Physiology and Pharmacology “V. Erspamer”, University “La Sapienza”<br />

* Experimental Flight Center, Dpt. <str<strong>on</strong>g>of</str<strong>on</strong>g> Aviati<strong>on</strong> and Space Medicine, Pratica di Mare, Rome, Italy<br />

e-mail: francesca.patacchioli@uniroma1.it<br />

The measurement <str<strong>on</strong>g>of</str<strong>on</strong>g> horm<strong>on</strong>es in saliva provides an interesting and reliable tool to assess the resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

neuroendocrine system to different stressors. Hypoxia familiarizati<strong>on</strong> is an integral part <str<strong>on</strong>g>of</str<strong>on</strong>g> the aviators'<br />

educati<strong>on</strong>. For many aviators, hypoxia preventi<strong>on</strong> begins in the hypobaric (altitude) chamber. In order to<br />

reach the experience <str<strong>on</strong>g>of</str<strong>on</strong>g> typical signs and symptoms <str<strong>on</strong>g>of</str<strong>on</strong>g> hypoxic hypoxia, twelve pr<str<strong>on</strong>g>of</str<strong>on</strong>g>icient paratroops were<br />

exposed to high altitude in a hypobaric chamber.<br />

275


23-26 August 2007,<br />

Budapest, Hungary<br />

We m<strong>on</strong>itored the hypothalamic-pituitary-adrenal axis resp<strong>on</strong>se to hypobaric hypoxia collecting salivary<br />

samples each two hours in the day <str<strong>on</strong>g>of</str<strong>on</strong>g> the simulati<strong>on</strong> in the hypobaric chamber, and also the day be<str<strong>on</strong>g>for</str<strong>on</strong>g>e and<br />

the day after.<br />

Altitude-induced hypoxia represents a str<strong>on</strong>g envir<strong>on</strong>mental stressor as shown by the significant increase <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

cortisol released during the “hypobaric chamber day”. Moreover, dhea-s levels were significantly increased in<br />

the day <str<strong>on</strong>g>of</str<strong>on</strong>g> the strenuous hypobaric chamber training: this result is c<strong>on</strong>sistent with the more recent literature<br />

suggesting a stress-buffering role <str<strong>on</strong>g>of</str<strong>on</strong>g> dhea-s. A comparis<strong>on</strong> most <str<strong>on</strong>g>of</str<strong>on</strong>g> interest was that c<strong>on</strong>trasting the cortisol to<br />

dhea-s ratio measured in the day <str<strong>on</strong>g>of</str<strong>on</strong>g> stress exposure with that <strong>on</strong>e measured the day be<str<strong>on</strong>g>for</str<strong>on</strong>g>e: no difference was<br />

found between the two measures. What can be drawn from this result, together with the observati<strong>on</strong> that this<br />

very stressful experience appears not to be associated with an impairment <str<strong>on</strong>g>of</str<strong>on</strong>g> cortisol and dheas circadian<br />

fluctuati<strong>on</strong>, is that the group <str<strong>on</strong>g>of</str<strong>on</strong>g> paratroops we have studied presented a good level <str<strong>on</strong>g>of</str<strong>on</strong>g> stress resilience.<br />

276


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5F. EXERCISE STRESS<br />

5F_01_P<br />

(poster secti<strong>on</strong> B1, poster board #215, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EXERCISE TRAINING MODIFIES INTESTINAL CELLULAR STRESS RESPONSE IN<br />

YOUNG MICE<br />

I. Brenner, K. Bret<strong>on</strong>-H<strong>on</strong>eyman<br />

Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, Trent University, 1600 West Bank Dr. Peterborough, ON, Canada, K9J 7B8.<br />

e-mail: ibrenner @trentu.ca<br />

Regular exercise reduces the incidence <str<strong>on</strong>g>of</str<strong>on</strong>g> col<strong>on</strong> cancer and possibly inflammatory disorders <str<strong>on</strong>g>of</str<strong>on</strong>g> the intestine,<br />

however, the mechanisms underlying these exercise-induced adaptati<strong>on</strong>s are not well understood. This study<br />

examined the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> an exhaustive bout <str<strong>on</strong>g>of</str<strong>on</strong>g> exercise <strong>on</strong> cellular stress proteins. Fifteen young (4 weeks)<br />

C57BL/6 mice were randomly assigned to <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> three experimental groups: (1) sedentary (S), (2) treadmill<br />

training (TT) and (3) wheel running (WR). Sedentary mice maintained normal cage activity, TT mice ran 5<br />

days per week, at approximately 75% VO & 2 max, <strong>on</strong> a motorized treadmill and WR mice had free access to<br />

running wheels. Following 12-14 weeks <str<strong>on</strong>g>of</str<strong>on</strong>g> training, all mice underwent an exhaustive bout <str<strong>on</strong>g>of</str<strong>on</strong>g> exercise <strong>on</strong> the<br />

treadmill and were euthanized 2 hours afterwards by cervical dislocati<strong>on</strong>. Blood samples were obtained by<br />

cardiac puncture and the intestines were removed <str<strong>on</strong>g>for</str<strong>on</strong>g> processing. Total HSP 70 expressi<strong>on</strong>, in segments <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the small and large intestine, and plasma IL-6 c<strong>on</strong>centrati<strong>on</strong> were quantified by ELISAs. Levels <str<strong>on</strong>g>of</str<strong>on</strong>g> JNK and<br />

ERK½ in the small and large intestine were measured by Western Blot techniques. Both modes <str<strong>on</strong>g>of</str<strong>on</strong>g> exercise<br />

training (TT and WR) significantly increased HSP 70 expressi<strong>on</strong> to exhaustive exercise in the large intestine,<br />

whereas in the small intestine, HSP70 expressi<strong>on</strong> was significantly increased by exhaustive exercise in the TT<br />

mice <strong>on</strong>ly. Preliminary results indicate that JNK and ERK1/2 expressi<strong>on</strong> are down-regulated in the TT mice.<br />

There were no significant differences in plasma IL-6 levels between the groups. The results <str<strong>on</strong>g>of</str<strong>on</strong>g> this study are<br />

useful <str<strong>on</strong>g>for</str<strong>on</strong>g> identifying the intestinal molecular pathways that may be modified by different <str<strong>on</strong>g>for</str<strong>on</strong>g>ms <str<strong>on</strong>g>of</str<strong>on</strong>g> exercise<br />

training and by the stress <str<strong>on</strong>g>of</str<strong>on</strong>g> exhaustive exercise.<br />

5F_02_P<br />

(poster secti<strong>on</strong> B1, poster board #216, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE RESPONSE OF INTERLEUKIN-6 AND ITS SOLUBLE RECEPTOR TO<br />

REPEATED EXERCISE AND HEAT STRESS<br />

Stuart R Gray, Helen C Marshall, Myra A Nimmo<br />

SIPBS, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Strathclyde, Glasgow, UK<br />

e-mail: m.a.nimmo@strath.ac.uk<br />

Interleukin-6 (IL-6) is a pleiotropic cytokine that increases up to 100 fold with exercise (Ostrowski et al,<br />

1998). It has also been linked to immunosupressi<strong>on</strong> (Smith, 2003) and underper<str<strong>on</strong>g>for</str<strong>on</strong>g>mance syndrome (UPS)<br />

(Robs<strong>on</strong>, 2003). The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> the present study was to investigate the IL-6 resp<strong>on</strong>se when subjects are under<br />

c<strong>on</strong>siderable stress from the combinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> exercise and heat. This exposure regimen is similar to that <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

athletes acclimating to the heat. Seven male subjects (age 30 ± 4 yr, height 1.80 ± 0.08 m, body mass 73.8 ±<br />

8.5 kg; means ± SD) completed 2 hr cycle exercise (44% O 2peak ) in a hot, humid envir<strong>on</strong>ment (38 ºC, 60%<br />

relative humidity) <strong>on</strong> 3 c<strong>on</strong>secutive days. Blood samples, obtained at rest and immediately post exercise <strong>on</strong><br />

days 1 and 3, were analysed <str<strong>on</strong>g>for</str<strong>on</strong>g> plasma IL-6 and soluble IL-6 receptor (sIL-6R) via ELISA. Statistical analyses<br />

277


23-26 August 2007,<br />

Budapest, Hungary<br />

were per<str<strong>on</strong>g>for</str<strong>on</strong>g>med using two-way repeated measures ANOVA and post-hoc B<strong>on</strong>ferr<strong>on</strong>i corrected paired t-tests<br />

where appropriate.<br />

There were no differences in resting IL-6 or sIL-6R c<strong>on</strong>centrati<strong>on</strong> between days. Exercise heat stress resulted<br />

in an increase (P


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5F_04_P<br />

(poster secti<strong>on</strong> B1, poster board #218, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HORMONAL STRESS RESPONSES TO AN IMMERSION IN A SIPHON PLACED ON<br />

THE BOTTOM OF AN ALPINE CAVE OF 700M DEPTH<br />

E. Stenner°, A. Bussani^, B. Biasioli*, W. Micheli*, G. Delbello°, E. Gianoli*, C. Piccinini<br />

°School <str<strong>on</strong>g>of</str<strong>on</strong>g> Sports Medicine, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine and Surgery, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Trieste, Italy<br />

^Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biological Oceanography, Nati<strong>on</strong>al Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Oceanography and Applied Geophisics<br />

*Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Laboratory Medicine, A.O.U, Ospedali Riuniti di Trieste, Italy<br />

e-mail: elisabetta.stenner@libero.it<br />

We studied the hypothalamus pituitary adrenocortical (cortisol) and the hypothalamus pituitary (growth<br />

horm<strong>on</strong>e GH) system resp<strong>on</strong>ses to an immersi<strong>on</strong> d<strong>on</strong>e in an unexplored siph<strong>on</strong> placed in a cave, 700m under<br />

the surface. The combinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> heavy exercise in a demanding envir<strong>on</strong>ment be<str<strong>on</strong>g>for</str<strong>on</strong>g>e diving, the absolute<br />

darkness, the c<strong>on</strong>finement, the cold temperatures and the emoti<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> the explorati<strong>on</strong> as well as the<br />

awareness that if an accident happened the situati<strong>on</strong> became drastically very critical represent a unique<br />

multiple stress model that may be helpful in understanding the endocrine expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> exercise and<br />

psychological stress. Owing to the high skill <str<strong>on</strong>g>of</str<strong>on</strong>g> the per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance <strong>on</strong>ly <strong>on</strong>e cave-diver was tested; however this<br />

subject repeats the same immersi<strong>on</strong> twice. Two blood drawings were per<str<strong>on</strong>g>for</str<strong>on</strong>g>med: (1) at the bottom <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

cave, pre siph<strong>on</strong>, (2) at the end <str<strong>on</strong>g>of</str<strong>on</strong>g> the explorati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the siph<strong>on</strong> lasted about 45 minutes. Two blood<br />

drawings, as c<strong>on</strong>trols, were per<str<strong>on</strong>g>for</str<strong>on</strong>g>med <strong>on</strong> the same potholers, at the same resting time envisaged <str<strong>on</strong>g>for</str<strong>on</strong>g> the day<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the experiment to minimize the specimen processing time influences and any circadian fluctuati<strong>on</strong>. Serum<br />

GH and cortisol were measured with a chemiluminescence assay (DXI 800 Beckman Coulter, Fullert<strong>on</strong>,<br />

Cali<str<strong>on</strong>g>for</str<strong>on</strong>g>nia, 2004) within 24 hours. The marked rise <str<strong>on</strong>g>of</str<strong>on</strong>g> GH values during diving underlines the great intensity<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> cave diving ef<str<strong>on</strong>g>for</str<strong>on</strong>g>t probably due to the difficult route that the cave diver covered and the bulky technical<br />

equipment dressed while the rise <str<strong>on</strong>g>of</str<strong>on</strong>g> cortisol is likely due to the combinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> exercise, emoti<strong>on</strong>al stress, cold<br />

temperature and darkness.<br />

5F_05_P<br />

(poster secti<strong>on</strong> B1, poster board #219, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

WOULD LITERATURE ON TAI CHI FOR STRESS BE SUFFICIENT FOR<br />

A META-ANALYSIS<br />

Chenchen Wang, Timothy McAlind<strong>on</strong><br />

New England Medical Center, USA<br />

Purpose: To systematically review world literature <str<strong>on</strong>g>for</str<strong>on</strong>g> the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> Tai Chi (TC) exercise <strong>on</strong> psychological<br />

stress and determine the sufficiency <str<strong>on</strong>g>of</str<strong>on</strong>g> data <str<strong>on</strong>g>for</str<strong>on</strong>g> a meta-analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> their outcomes.<br />

Methods: We searched Medline, PsycINFO, Sportdiscus, Social Sciences, Health Star, EMBASE, China<br />

Hospital Knowledge, China Nati<strong>on</strong>al Knowledge Infrastructure, Traditi<strong>on</strong>al Chinese Medicine Databases and<br />

article references until February 2007. Inclusi<strong>on</strong> criteria were all types <str<strong>on</strong>g>of</str<strong>on</strong>g> human clinical studies that reported<br />

original data written in English or Chinese and which included at least <strong>on</strong>e clinical outcome relevant to<br />

psychological health. We extracted data <strong>on</strong> the populati<strong>on</strong> characteristics, study setting, type and durati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

TC, study design, psychological outcome, and primary results.<br />

Results: We screened 1172 abstracts and identified 23 studies (10 randomized c<strong>on</strong>trolled trials, 9<br />

n<strong>on</strong>randomized c<strong>on</strong>trolled trials, 4 observati<strong>on</strong>al studies). These eligible studies were c<strong>on</strong>ducted in 4<br />

279


23-26 August 2007,<br />

Budapest, Hungary<br />

countries (USA, Canada, China, Australia) involving a total <str<strong>on</strong>g>of</str<strong>on</strong>g> 2335 participants (age range is from 10 to 92).<br />

Of the 23 studies, 19 clinical trials with a total <str<strong>on</strong>g>of</str<strong>on</strong>g> 1153 subjects c<strong>on</strong>sistently reported that 4 weeks to 46<br />

m<strong>on</strong>ths <str<strong>on</strong>g>of</str<strong>on</strong>g> TC training resulted in higher levels <str<strong>on</strong>g>of</str<strong>on</strong>g> several indices <str<strong>on</strong>g>of</str<strong>on</strong>g> psychological well-being, including<br />

reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> stress, anxiety and depressi<strong>on</strong>, and improved life satisfacti<strong>on</strong> compared with the c<strong>on</strong>trols. Four<br />

observati<strong>on</strong>al studies further dem<strong>on</strong>strated that 6 m<strong>on</strong>ths to over 20 years TC practice improved mood, and<br />

reduced stress and depressi<strong>on</strong>.<br />

C<strong>on</strong>clusi<strong>on</strong>s: The heterogeneity <str<strong>on</strong>g>of</str<strong>on</strong>g> the study designs, settings, methods <str<strong>on</strong>g>of</str<strong>on</strong>g> comparis<strong>on</strong>s, exercise background,<br />

TC style, and durati<strong>on</strong>, precluded a <str<strong>on</strong>g>for</str<strong>on</strong>g>mal meta-analysis. However, TC appears to be associated with<br />

improvements in psychological stress and well-being in both eastern and western populati<strong>on</strong>s.<br />

5F_06_P<br />

(poster secti<strong>on</strong> B1, poster board #220, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ALPHA-TOCOPHEROL SUPPLEMENTATION PROTECTS SERUM PARAOXONASE<br />

1/ARYLESTERASE ACTIVITIES FROM EXERCISE-INDUCED REDUCTION<br />

Stylianos Tsakiris 1 , George A. Karikas 2 , Theodore Parthimos 1 , Theodore Tsakiris 1 ,<br />

C<strong>on</strong>stantinos Bakogiannis 1 , Kleopatra H. Schulpis 3<br />

1 Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Exp. Physiology, Athens University Medical School, PO Box 65257, GR-15401 Athens, Greece<br />

2 Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Laboratories, Technological Educati<strong>on</strong>al Instituti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Athens, Greece<br />

3 Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Child Health, “Aghia Sophia” Children’s Hospital, Athens, Greece<br />

Aim: To investigate the closely related to oxidati<strong>on</strong> and atherogenesis Paraox<strong>on</strong>ase 1/Arylesterase (PON<br />

1/Aryl) activities in basketball players with or without alpha-Tocopherol (alpha-T) supplementati<strong>on</strong> pre- and<br />

post-training. Subject and Methods: Blood was obtained from 10 players pre- (group A), post- exercise<br />

(group B) and after 1 m<strong>on</strong>th <strong>on</strong> alpha-T (200 mg/24 h orally) supplementati<strong>on</strong> pre- (group C) and postexercise<br />

(group D). Lactate, pyruvate, muscle enzyme activities (CK, LDH) and total antioxidant status (TAS)<br />

were measured with commercial kits. Catecholamines and alpha-T were determined with HPLC and PON<br />

1/Aryl activities spectrophotometrically. Results: Lactate, pyruvate, muscle enzyme activities and<br />

catecholamines were increased (p


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5F_07_P<br />

(poster secti<strong>on</strong> B1, poster board #221, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECTS OF PHYSICAL EXERCISE ON MR AND GR EXPRESSION IN THE<br />

HIPPOCAMPAL FORMATION OF THE ADULT RAT<br />

I. H. J<strong>on</strong>sdottir 1, 2 , A. S. Naylor 2 , T. Thorlin 2 , S.-A. Bergström 3 , I. Söderström 3 , T. Olss<strong>on</strong> 3<br />

1The Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress Medicine,<br />

2Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> neuroscience and physiology, Göteborg University<br />

3The Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Umeå University, Sweden<br />

Email: inga.j<strong>on</strong>sdottir@stressmedicin.com<br />

We have previously shown that short-term voluntary running <str<strong>on</strong>g>for</str<strong>on</strong>g> 9 days results in a fivefold increase in newly<br />

divided progenitors in the hippocampus, whereas l<strong>on</strong>g-term voluntary running <str<strong>on</strong>g>for</str<strong>on</strong>g> 24 days paradoxically<br />

decreases by half dividing progenitors in the adult rat brain. In additi<strong>on</strong>, a c<strong>on</strong>comitant increase in stress<br />

resp<strong>on</strong>ses (increased plasma corticoster<strong>on</strong>e levels and adrenal gland weight with decreased thymus weight)<br />

was seen after l<strong>on</strong>g-term running <strong>on</strong>ly. We have now analysed the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> different voluntary running<br />

durati<strong>on</strong> <strong>on</strong> expressi<strong>on</strong> <strong>on</strong> mineralocorticoid (MR) and glucocorticoid (GR) receptors in hippocampal<br />

subregi<strong>on</strong>s (dentate gyrus (DG) and CA1 and CA3) within the adult rodent brain (40 female rats,<br />

n=20/group, single-housed with/without a running wheel). Nine days <str<strong>on</strong>g>of</str<strong>on</strong>g> voluntary running induced a<br />

significant increase in MR expressi<strong>on</strong> in the DG (p=0.008) and CA1 (p=0.006) subregi<strong>on</strong>s without effects in<br />

CA3. No effects were seen in GR expressi<strong>on</strong> after 9 or 24 days <str<strong>on</strong>g>of</str<strong>on</strong>g> running. However, there was a significant<br />

decrease in GR expressi<strong>on</strong> between 9 and 24 days in the n<strong>on</strong>-running group. This decrease was not seen in<br />

the running group. We c<strong>on</strong>clude that short-term running results in upregulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> MR expressi<strong>on</strong> in the<br />

hippocampus. This may be a key in the previously suggested beneficial effects <str<strong>on</strong>g>of</str<strong>on</strong>g> exercise <strong>on</strong> neur<strong>on</strong>al<br />

plasticity. L<strong>on</strong>g-term running abolishes this effect. A decrease in GR expressi<strong>on</strong> in n<strong>on</strong>-runners may be<br />

sec<strong>on</strong>dary to social isolati<strong>on</strong>. Running seems to protect against this negative effect. Effects <str<strong>on</strong>g>of</str<strong>on</strong>g> MR and GR<br />

modulati<strong>on</strong> in this model <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>al plasticity should be <str<strong>on</strong>g>of</str<strong>on</strong>g> major interest.<br />

281


23-26 August 2007,<br />

Budapest, Hungary<br />

282<br />

5G. STRESS AND NUTRITION<br />

5G_01_P<br />

(poster secti<strong>on</strong> B1, poster board #222, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

QUERCETIN, A DIETARY-DERIVED FLAVONOID, ACTS AS A FREE RADICAL<br />

SCAVENGER IN SWINE GRANULOSA CELL<br />

Giuseppina Basini, Sujen Ele<strong>on</strong>ora Santini, Francesca Grasselli<br />

Dipartimento di Produzi<strong>on</strong>i Animali, Biotecnologie Veterinarie, Qualità e Sicurezza degli Alimenti, Sezi<strong>on</strong>e di<br />

Fisiologia Veterinaria, Università di Parma, 43100 Parma, Italy<br />

e-mail: basini@unipr.it<br />

Quercetin (3,3’,4’,5,7-penthydroxy flav<strong>on</strong>e) is an important c<strong>on</strong>stituent <str<strong>on</strong>g>of</str<strong>on</strong>g> the flav<strong>on</strong>oid family present in<br />

many fruits and vegetables. Various pharmacological activities <str<strong>on</strong>g>of</str<strong>on</strong>g> quercetin have been dem<strong>on</strong>strated including<br />

suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> tumor growth by inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> neovascularizati<strong>on</strong>. However, its mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> acti<strong>on</strong> has not<br />

been clarified yet. Ovarian follicle is an excellent model to study angiogenesis since it is an unique excepti<strong>on</strong><br />

to the general quiescence <str<strong>on</strong>g>of</str<strong>on</strong>g> the adult vasculature. In previous studies, we dem<strong>on</strong>strated an involvement <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

reactive oxygen species (ROS) in angiogenesis signalling. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, the aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this research was to evaluate if<br />

the treatment with 5 or 50 µg/ml <str<strong>on</strong>g>of</str<strong>on</strong>g> quercetin modulates superoxide ani<strong>on</strong> (O 2- ) generati<strong>on</strong> and superoxide<br />

dismutase (SOD) activity in granulosa cells collected from swine follicles. Our results evidenced that the<br />

-<br />

highest c<strong>on</strong>centrati<strong>on</strong> af quercetin was able to significantly (p


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

(61.79+/- 14.9) experienced the highest stress level (F=2.43, p=0.04). NES was present in 7.7 % <str<strong>on</strong>g>of</str<strong>on</strong>g> students<br />

(4 males, 3 females), while evening hyperphagia was presentin 72.8% students (20 males, 47 females). Evening<br />

hyperphagia correlated with anxiety (r= -0.24, p=0.020). CONCLUSIONS: 1) Senior college students<br />

experienced significantly more stress than juniors. 2) Anxiety was more prevalent in female students. 3)<br />

Evening hyperphagia was comm<strong>on</strong> and was associated with anxiety. Stress in college students is prevalent and<br />

may lead to abnormal eating patterns, which may partially explain the c<strong>on</strong>tributi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> stress to development<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> obesity.<br />

5G_03_P<br />

(poster secti<strong>on</strong> B1, poster board #224, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

AMBULATORY ACTIVITY AND THE ACCUMBENS C-FOS EXPRESSION WERE<br />

DECREASED BY REPEATED ORAL CAPSAICIN IN RATS<br />

Xue Feng Gu 1 , J<strong>on</strong>g-Ho Lee 1 , Vitaly Ryu 1 , Sang Bae Yoo 1 , Young Wha Mo<strong>on</strong> 2 , Je<strong>on</strong>g W<strong>on</strong> Jahng 1<br />

1. Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Oral and Maxill<str<strong>on</strong>g>of</str<strong>on</strong>g>acial Surgery, Dental research institute, Seoul Nati<strong>on</strong>al University School <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Dentistry, Seoul, 110-744, Korea; 2. Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, The Catholic University <str<strong>on</strong>g>of</str<strong>on</strong>g> Korea College <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine,<br />

Seoul, 137-701, Korea<br />

Capsaicin (8-methyl-N-vanillyl-6-n<strong>on</strong>enamide), the active comp<strong>on</strong>ent <str<strong>on</strong>g>of</str<strong>on</strong>g> chili peppers, is widely believed to<br />

stimulate appetite. The "capsaicin high" is a euphoric sensati<strong>on</strong> caused by the c<strong>on</strong>sumpti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> large quantities<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> capsaicin from capsaicin-laden foods. We have previously found that gene expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a sweet taste<br />

receptor T1R3 as well as capsaicin receptor VR1 was decreased in the circumvallate papilla <str<strong>on</strong>g>of</str<strong>on</strong>g> rats that had<br />

repeated exposure to oral capsaicin, and they showed preferences <strong>on</strong> sweet soluti<strong>on</strong>s, revealing a molecular<br />

basis underlying the hed<strong>on</strong>ic effect <str<strong>on</strong>g>of</str<strong>on</strong>g> oral capsaicin. In this study, we examined ambulatory activity and c-Fos<br />

expressi<strong>on</strong> in the nucleus accumbens (Accb) in rats with repeated oral capsaicin. Male SD rats received daily<br />

1 ml <str<strong>on</strong>g>of</str<strong>on</strong>g> 0.02 % capsaicin or distilled water in their oral cavity with ad libitum access to rodent chow and water.<br />

On days 1, 5 and 10, rats were placed in the activity chamber 30 min after capsaicin exposure, and then the<br />

ambulatory activity was recorded <str<strong>on</strong>g>for</str<strong>on</strong>g> 30 min. Two days after the behavioral sessi<strong>on</strong> without capsaicin, rats<br />

were transcardially perfused with 4 % PFA and the brain tissues were processed <str<strong>on</strong>g>for</str<strong>on</strong>g> c-Fos<br />

immunohistochemistry. Ambulatory counts decreased significantly in the capsaicin group compared with the<br />

c<strong>on</strong>trol group <strong>on</strong> each test day. The ambulatory counts <str<strong>on</strong>g>of</str<strong>on</strong>g> capsaicin rats, but not <str<strong>on</strong>g>of</str<strong>on</strong>g> c<strong>on</strong>trol rats, further<br />

decreased <strong>on</strong> the sec<strong>on</strong>d test day compared to the first test day. The number <str<strong>on</strong>g>of</str<strong>on</strong>g> c-Fos-ir nuclei significantly<br />

decreased in the Accb and the cingulate cortex <str<strong>on</strong>g>of</str<strong>on</strong>g> capsaicin rats compared to the c<strong>on</strong>trol group. Together<br />

with our previous finding, this result suggests that hed<strong>on</strong>ic properties <str<strong>on</strong>g>of</str<strong>on</strong>g> oral capsaicin may be mediated by<br />

decreased c-Fos expressi<strong>on</strong> in the brain reward pathway. Supported by KMOST (JWJ).<br />

283


23-26 August 2007,<br />

Budapest, Hungary<br />

284<br />

5G_04_P<br />

(poster secti<strong>on</strong> B1, poster board #225, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

OXIDATIVE STRESS IN HUMAN BODY AND PROBIOTIC APPLICATION<br />

Tiiu Kullisaar, Tiiu Vihalemm, Marika Mikelsaar, Mihkel Zilmer<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry University <str<strong>on</strong>g>of</str<strong>on</strong>g> Tartu, Tartu, Est<strong>on</strong>ia<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Microbiology University <str<strong>on</strong>g>of</str<strong>on</strong>g> Tartu, Tartu, Est<strong>on</strong>ia<br />

Human beings living in oxygen envir<strong>on</strong>ment and using oxygen <str<strong>on</strong>g>for</str<strong>on</strong>g> their aerobic metabolisms have to deal<br />

with free radicals, which are normally, at very low levels, produced by several metabolic processes. When<br />

antioxidants/oxidants ratio is balanced, nothing will happen, but when this balance is disturbed the oxidative<br />

stress (OxS) will appear. High-grade OxS has an impact <strong>on</strong> development <str<strong>on</strong>g>of</str<strong>on</strong>g> inflammatory diseases and any<br />

kinds <str<strong>on</strong>g>of</str<strong>on</strong>g> approach, which have beneficial effects, are welcome.<br />

Lactic acid bacteria (LAB) <str<strong>on</strong>g>of</str<strong>on</strong>g> our intestinal micr<str<strong>on</strong>g>of</str<strong>on</strong>g>lora play a significant role in the gut ecosystem having<br />

beneficial cross- talk with human organism. LAB <str<strong>on</strong>g>of</str<strong>on</strong>g> human origin helps to restore normal microbial functi<strong>on</strong>,<br />

stimulating immune system, and some <str<strong>on</strong>g>of</str<strong>on</strong>g> them carry remarkable antioxidative, anti-atherogenic and anticancerogenic<br />

properties. Viable LAB c<strong>on</strong>sidered as good tools <str<strong>on</strong>g>for</str<strong>on</strong>g> designing health supporting functi<strong>on</strong>al<br />

food.<br />

Our previous studies have shown that antioxidative probiotic L. fermentum ME-3 possesses substantial<br />

antioxidative and antimicrobial activity and has beneficial effects <strong>on</strong> human well being. C<strong>on</strong>sumpti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

probiotic ME-3 correct OxS status in clinically n<strong>on</strong>-problematic subjects, suppresses high-grade OxS in<br />

diseased pers<strong>on</strong>s in the associati<strong>on</strong> with clinical outcome, which points to the possibility to gain the better<br />

quality <str<strong>on</strong>g>of</str<strong>on</strong>g> life as adjunct therapy.<br />

Regular intake <str<strong>on</strong>g>of</str<strong>on</strong>g> dietary products, enriched with special probiotics, without necessity to change typical eating<br />

habits <str<strong>on</strong>g>of</str<strong>on</strong>g> pers<strong>on</strong> (patient) might be used easily accessible low cost large scale tool <str<strong>on</strong>g>for</str<strong>on</strong>g> populati<strong>on</strong>.<br />

5G_05_P<br />

(poster secti<strong>on</strong> B1, poster board #226, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SAGE DRINKING IMPROVES PLASMA LIPID PROFILE, ERYTHROCYTE<br />

ANTIOXIDANT DEFENCES AND INCREASES LYMPHOCYTE HSP70<br />

P. C. Piairo, C. M. Sa, A. Abreu-Ramos, C. F. Lima*, M. F. Azevedo, M. J. Almeida,<br />

O. P. Coutinho, C. Pereira-Wils<strong>on</strong><br />

<str<strong>on</strong>g>Centre</str<strong>on</strong>g>/Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, School <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Minho, Braga, Portugal<br />

e-mail: lima@bio.uminho.pt<br />

Salvia <str<strong>on</strong>g>of</str<strong>on</strong>g>ficinalis (comm<strong>on</strong> sage) is a medicinal plant to which antioxidant, anti-inflammatory and antimutagenic<br />

properties have been attributed. Recent results from our laboratory showed cellular and in vivo antioxidant<br />

effects <str<strong>on</strong>g>of</str<strong>on</strong>g> sage as well as met<str<strong>on</strong>g>for</str<strong>on</strong>g>min-like effects at the rat liver level, suggesting an antidiabetic potential <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

sage. In order to test these effects in humans, we per<str<strong>on</strong>g>for</str<strong>on</strong>g>med a pilot trial with six healthy female volunteers.<br />

The trial was carried out in three phases, which includes two weeks <str<strong>on</strong>g>of</str<strong>on</strong>g> baseline, four weeks <str<strong>on</strong>g>of</str<strong>on</strong>g> sage treatment<br />

(drinking <str<strong>on</strong>g>of</str<strong>on</strong>g> a sage infusi<strong>on</strong> twice a day) and two weeks <str<strong>on</strong>g>of</str<strong>on</strong>g> wash-out. Sage treatment positively affected the<br />

erythrocyte antioxidant status as shown by increased SOD and CAT activities. Cholesterol and LDL levels<br />

significantly decreased and HDL levels significantly increased after treatment, indicating benefits also in lipid<br />

metabolism. However, no changes in glucose clearance were observed in the oral glucose tolerance tests at<br />

the end <str<strong>on</strong>g>of</str<strong>on</strong>g> treatment period. In additi<strong>on</strong>, a reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> in vitro lymphocyte DNA damage induced by H 2 O 2


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

was observed during the treatment period, which was maintained through the wash-out period. During the<br />

S. <str<strong>on</strong>g>of</str<strong>on</strong>g>ficinalis drinking period, lymphocyte Hsp70 protein expressi<strong>on</strong> was significantly increased (about 2.25<br />

times) and decreased to baseline following the wash-out period. Overall these results c<strong>on</strong>firm the health<br />

improving potential <str<strong>on</strong>g>of</str<strong>on</strong>g> sage infusi<strong>on</strong> drinking.<br />

5G_06_P<br />

(poster secti<strong>on</strong> B1, poster board #227, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

BIOACTIVE PEPTIDES FROM BUFFALO MOZZARELLA-CHEESE WASTE WHEY:<br />

ANTIOXIDANT AND DIFFERENTIATION EFFECTS ON COLON<br />

ADENOCARCINOMA CACO-2 CELLS<br />

P. Ferranti, D. Cassese, G. Picariello, A. Dicitore, C. De Sim<strong>on</strong>e, G. Giuberti, A. Abbruzzese,<br />

P. Stiuso, Paola Stiuso<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry and Biophysics, Sec<strong>on</strong>da Università degli Studi <str<strong>on</strong>g>of</str<strong>on</strong>g> Naples, Italy<br />

e-mail: paola.stiuso@unina2.it<br />

The gastrointestinal mucosa is c<strong>on</strong>stantly exposed to luminal oxidants from ingested foods as reactive oxygen<br />

species (ROS) that elicit membrane destabilizati<strong>on</strong>, alterati<strong>on</strong>s in DNA, inactivati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proteolytic enzymes .<br />

The ingesti<strong>on</strong> and/or occurrence <str<strong>on</strong>g>of</str<strong>on</strong>g> these reactive species, particularly in the l<strong>on</strong>g term, are resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

injury to the intestinal mucosa in several disease states, as well as inflammatory bowel disease and intestinal<br />

cancer. Bovine milk proteins are a source <str<strong>on</strong>g>of</str<strong>on</strong>g> biologically active peptides that can be released during<br />

gastrointestinal digesti<strong>on</strong> or food processing. They may act as regulatory compounds which exhibit a wide<br />

range <str<strong>on</strong>g>of</str<strong>on</strong>g> biological functi<strong>on</strong>s such as antimicrobial, antihypertensive, antioxidant and opioid activities.<br />

However no study have been carried out <strong>on</strong> peptides in buffalo milk and/or <strong>on</strong> peptides released in whey<br />

during technological trans<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> buffalo milk. Our study focuses <strong>on</strong> characterizing peptide fracti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

buffalo mozzarella-cheese waste whey by complementary mass spectrometric techniques. Moreover we have<br />

investigated a cytomodulatory and antioxidant effect <str<strong>on</strong>g>of</str<strong>on</strong>g> peptide fracti<strong>on</strong>s from buffalo mozzarella-cheese<br />

whey <strong>on</strong> hydrogen peroxide-induced oxidative damage in CaCo-2 cell lines. We have dem<strong>on</strong>strated that a<br />

specific peptide fracti<strong>on</strong> (F3) after 12h <str<strong>on</strong>g>of</str<strong>on</strong>g> treatment, decreased ROS producti<strong>on</strong> (-42%O 2- ) and enhanced cell<br />

differentiati<strong>on</strong> (27%Alkaline Phosphatase activity/min). Our results showed that peptide fracti<strong>on</strong> (F3)<br />

derived from buffalo mozzarella-cheese waste whey is potential health enhancing nutriaceutical <str<strong>on</strong>g>for</str<strong>on</strong>g> food and<br />

pharmaceutical applicati<strong>on</strong>s.<br />

5G_07_P<br />

(poster secti<strong>on</strong> B1, poster board #228, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECT OF IRRIGATION INTERVALS AND POTASSIUM APPLICATION ON SOME<br />

BIOCHEMICAL INDICES OF PISTACHIO<br />

A. Tajabadi Pour<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Soil Science, College <str<strong>on</strong>g>of</str<strong>on</strong>g> Agriculture, Rafsanjan University, Rafsanjan, Iran<br />

e-mail: ahtajabadi@yahoo.com.au<br />

Water deficit <str<strong>on</strong>g>of</str<strong>on</strong>g> soils is the principle factor limiting crop yield in large areas <str<strong>on</strong>g>of</str<strong>on</strong>g> world. Plants undergo<br />

significant morphological and metabolic changes in resp<strong>on</strong>se to drought. Many <str<strong>on</strong>g>of</str<strong>on</strong>g> these changes are believed<br />

to adaptive resp<strong>on</strong>ses by which plants cope with water stress. It is generally accepted that the first result <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

285


23-26 August 2007,<br />

Budapest, Hungary<br />

stress is an alterati<strong>on</strong> in the structure and functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cell membranes. There are more than 420000 ha <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

n<strong>on</strong>bearing and bearing pistachio trees in Iran. Despite the ec<strong>on</strong>omic importance <str<strong>on</strong>g>of</str<strong>on</strong>g> this crop, very little<br />

in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> is available <strong>on</strong> its moisture needs and nutriti<strong>on</strong>al requirements. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> three<br />

irrigati<strong>on</strong> frequencies (1, 3, and 7 days) and five potassium (K) levels (0, 75, 150, 225, and 300 mg K kg -1 soil<br />

as K 2 SO 4 ) <strong>on</strong> the proline and reducing sugars c<strong>on</strong>tent and activity <str<strong>on</strong>g>of</str<strong>on</strong>g> catalase and peroxidase <str<strong>on</strong>g>of</str<strong>on</strong>g> pistachio<br />

were studied in a glasshouse experiment. Water frequency decreased all <str<strong>on</strong>g>of</str<strong>on</strong>g> growth parameters (shoot and root<br />

dry weight, stem length and leaf area) but potassium applicati<strong>on</strong> had no significant effects <strong>on</strong> the growth<br />

parameters. As the irrigati<strong>on</strong> intervals and K applicati<strong>on</strong> increased, the proline c<strong>on</strong>tent significantly increased.<br />

In c<strong>on</strong>trast to proline, with increasing irrigati<strong>on</strong> frequencies and K applicati<strong>on</strong> the reducing sugars<br />

c<strong>on</strong>centrati<strong>on</strong> significantly decreased. A significant decrease in peroxidaes and catalase activity was observed<br />

with increased K levels. However, with increasing irrigati<strong>on</strong> intervals, activity <str<strong>on</strong>g>of</str<strong>on</strong>g> these enzymes increased. In<br />

c<strong>on</strong>clusi<strong>on</strong>, the results <str<strong>on</strong>g>of</str<strong>on</strong>g> this study clearly dem<strong>on</strong>strate that although K applicati<strong>on</strong> had no significant effects<br />

<strong>on</strong> some growth parameters, it significantly modified the biochemical indices and somewhat overcame the<br />

depressing effects <str<strong>on</strong>g>of</str<strong>on</strong>g> water stress.<br />

286


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5H. STRESS OF DOMESTIC ANIMALS AND FISH<br />

5H_01_P<br />

(poster secti<strong>on</strong> B1, poster board #229, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CASPASE-3-MEDIATED PROAPOPTOTIC SIGNALING IN INTERSEXUAL GONAD<br />

OF ZEBRAFISH<br />

Nanami Mizusawa, Michiaki Yamashita<br />

Nati<strong>on</strong>al <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Fisheries Science, Stress-induced apoptosis in g<strong>on</strong>adal masculinizati<strong>on</strong><br />

e-mail: nkawa@fra.affrc.go.jp<br />

Phenotypic sex in fish is influenced by envir<strong>on</strong>mental c<strong>on</strong>diti<strong>on</strong>s in additi<strong>on</strong> to genotype. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, various<br />

stressors, such as temperature, hypoxia and polluti<strong>on</strong> at sublethal levels, are assumed to influence the sex<br />

ratio. In order to evaluate the hypothesis, we designed an experiment to assess the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock <strong>on</strong><br />

sex ratio using zebrafish as the model. As the time <str<strong>on</strong>g>of</str<strong>on</strong>g> heat stress during development is critical, temperature<br />

sensitive stage was selected <str<strong>on</strong>g>for</str<strong>on</strong>g> the study. The phenotypic sex ratio was examined until 40 days, using<br />

histological observati<strong>on</strong>. The results were compared with a c<strong>on</strong>trol without heat shock. Results indicated<br />

that in c<strong>on</strong>trol a 50/50 male to female ratio was maintained. However, a significantly higher increase in male<br />

phenotype was observed. Our study also indicates that 14 to 16 days post fertilizati<strong>on</strong> is critical <str<strong>on</strong>g>for</str<strong>on</strong>g> sex<br />

differentiati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> primitive g<strong>on</strong>ad. TUNEL staining and immunohistochemical observati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the g<strong>on</strong>ads<br />

showed the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> caspase-3 mediated proapoptotic signaling in the g<strong>on</strong>adal cells after the heat shock.<br />

There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, proapoptotic signaling mechanism may regulate the sex differentiati<strong>on</strong> in the g<strong>on</strong>ads at this stage<br />

and may be markedly induced by stress c<strong>on</strong>diti<strong>on</strong>s. We report a model <str<strong>on</strong>g>of</str<strong>on</strong>g> the evaluati<strong>on</strong> effects <strong>on</strong> sex<br />

differentiati<strong>on</strong> using zebrafish. We anticipate that this model will facilitate further study <str<strong>on</strong>g>of</str<strong>on</strong>g> the exposure to<br />

stressors by m<strong>on</strong>itoring their sex ratio and gene expressi<strong>on</strong> patterns in the g<strong>on</strong>ad.<br />

5H_02_P<br />

(poster secti<strong>on</strong> B1, poster board #230, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

VALIDATION OF A FECAL GLUCOCORTICOID ASSAY FOR THE SYRIAN<br />

HAMSTER (MESOCRICETUS AURATUS)<br />

Marie-Odile M. Chelini 1 , Priscila F. Viau 2 , Cláudio A. de Oliveira 2 , Emma Otta 1<br />

1Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychology, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Experimental Psychology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> São Paulo, São Paulo, Brazil<br />

2 Horm<strong>on</strong>e Quantificati<strong>on</strong> Laboratory, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Animal Reproducti<strong>on</strong>, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Veterinary Medicine,<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> SãoPaulo, São Paulo, Brazil<br />

The measurement <str<strong>on</strong>g>of</str<strong>on</strong>g> fecal steroid metabolites has proved to be a reliable technique <str<strong>on</strong>g>for</str<strong>on</strong>g> m<strong>on</strong>itoring endocrine<br />

status in a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> mammalian species. We verified the biological relevance <str<strong>on</strong>g>of</str<strong>on</strong>g> quantifying the c<strong>on</strong>centrati<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> cortisol fecal metabolites to assess the physiologic stress resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> the Syrian hamster. Two experiments<br />

were per<str<strong>on</strong>g>for</str<strong>on</strong>g>med using a total <str<strong>on</strong>g>of</str<strong>on</strong>g> 10 adult female hamsters. To suppress adrenocortical activity, five females<br />

were injected with 200 µg/100 g body weight <str<strong>on</strong>g>of</str<strong>on</strong>g> dexamethas<strong>on</strong>e, dissolved in 1 ml <str<strong>on</strong>g>of</str<strong>on</strong>g> sterile isot<strong>on</strong>ic saline<br />

soluti<strong>on</strong>. Dexamethas<strong>on</strong>e is an artificial steroid that mimics endogenous glucocorticoids and reduces<br />

circulating corticosteroid levels via the negative-feedback mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> the HPA axis. To investigate the<br />

effects <str<strong>on</strong>g>of</str<strong>on</strong>g> the injecti<strong>on</strong> procedure itself <strong>on</strong> the pattern <str<strong>on</strong>g>of</str<strong>on</strong>g> excreted fecal steroid metabolites, five females<br />

received an injecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 1 ml <str<strong>on</strong>g>of</str<strong>on</strong>g> sterile isot<strong>on</strong>ic saline soluti<strong>on</strong>. All fecal samples voided by each group were<br />

collected and homogenized <strong>on</strong> a daily basis <str<strong>on</strong>g>for</str<strong>on</strong>g> 5 days be<str<strong>on</strong>g>for</str<strong>on</strong>g>e (to assess basal c<strong>on</strong>centrati<strong>on</strong>) and thereafter <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

287


23-26 August 2007,<br />

Budapest, Hungary<br />

4 days after the injecti<strong>on</strong>s. Cortisol fecal metabolites were extracted with ethanol and quantified by<br />

radioimmunoassay. The basal c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cortisol fecal metabolites was 3.95 ± 1.25 µg/g <str<strong>on</strong>g>of</str<strong>on</strong>g> feces in the<br />

c<strong>on</strong>trol group and 5.78 ± 0.62 µg/g <str<strong>on</strong>g>of</str<strong>on</strong>g> feces in the group injected with dexamethas<strong>on</strong>e. On the first day after<br />

injecti<strong>on</strong>, whereas a slight increase in cortisol c<strong>on</strong>centrati<strong>on</strong> in the c<strong>on</strong>trol group (4.81 µg/g <str<strong>on</strong>g>of</str<strong>on</strong>g> feces)<br />

(p


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5H_04_P<br />

(poster secti<strong>on</strong> B1, poster board #232, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECTS OF SEX AND GENETICS ON BEHAVIOR AND STRESS RESPONSE OF<br />

TURKEYS<br />

G. R. Huff, W. E. Huff, N. C. Rath, A. M. D<strong>on</strong>oghue, N. B. Anth<strong>on</strong>y, K. E. Nestor<br />

USDA, Agricultural <str<strong>on</strong>g>Research</str<strong>on</strong>g> Service, Fayetteville, AR USA 72701<br />

e-mail: grhuff@uark.edu<br />

Stress can lead to changes in the immune resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> turkeys resulting in decreased resistance to<br />

opportunistic bacterial pathogens. Three lines <str<strong>on</strong>g>of</str<strong>on</strong>g> turkeys were tested <str<strong>on</strong>g>for</str<strong>on</strong>g> resp<strong>on</strong>se in T-maze and open field<br />

tests during the first 8 days after hatch and behavior was observed after catching, moving, and transport.<br />

They were also compared <str<strong>on</strong>g>for</str<strong>on</strong>g> corticoster<strong>on</strong>e (Cort) levels and heterophil/lymphocyte ratios (H/L) at 15 wk<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> age, in resp<strong>on</strong>se to an Escherichia coli challenge followed by transport stress. Large commercial line birds<br />

(Comm) were faster and more active in the T maze at day 2 than smaller Egg line birds. Male Comm line<br />

birds were faster than male Egg line birds when tested in an open field at day 8. Egg line birds had more<br />

sleeping behavior after moving to a new floor pen as compared to both an intermediate-sized line (F line) and<br />

the Comm line. Transport stress increased Cort levels in all 3 lines and the increase was greater in males<br />

compared to females. The Egg line had higher basal Cort levels (P = 0.03) and higher levels after transport<br />

(P< 0.0001). The H/L ratios were affected by both transport stress and line but not by sex. The H/L ratio<br />

was lower in the Egg line as compared to both the F line and the Comm line (P< 0.0001), with the Comm<br />

line having the greatest increase in resp<strong>on</strong>se to transport. Previous studies determined that Egg line birds<br />

were more resistant to the deleterious effects <str<strong>on</strong>g>of</str<strong>on</strong>g> challenge and the Comm line displayed the most adverse<br />

effects. These data suggest that differences in activity <str<strong>on</strong>g>of</str<strong>on</strong>g> fast-growing turkeys may be used to select birds that<br />

are less susceptible to inflammatory bacterial disease and that the H/L ratio may be more useful than serum<br />

Cort in evaluating the deleterious effects <str<strong>on</strong>g>of</str<strong>on</strong>g> stress.<br />

5H_05_P<br />

(poster secti<strong>on</strong> B1, poster board #233, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DIFFERENTIAL REGULATION OF GLUCOCORTICOID AND<br />

MINERALOCORTICOID RECEPTOR AND 11ß-HYDROXYSTEROID<br />

DEHYDROGENASE 1 AND 2 MRNAS BY ACUTE PSYCHOSOCIAL STRESS IN<br />

PORCINE BRAIN<br />

E. Kanitz, M. Tuchscherer, B. Puppe, T. Viergutz, A. Tuchscherer<br />

<str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> the Biology <str<strong>on</strong>g>of</str<strong>on</strong>g> Farm Animals, <str<strong>on</strong>g>Research</str<strong>on</strong>g> Unit Behavioural Physiology, Dummerstorf, Germany<br />

e-mail: ekanitz@fbn-dummerstorf.de<br />

Early life events can have short and l<strong>on</strong>g-term effects <strong>on</strong> neuroendocrine, aut<strong>on</strong>omic, and behavioural<br />

resp<strong>on</strong>ses to stress and appear to play an important role in the etiology <str<strong>on</strong>g>of</str<strong>on</strong>g> stress-related disorders.<br />

Glucocorticoids are key mediators <str<strong>on</strong>g>of</str<strong>on</strong>g> these brain-endocrine-immune c<strong>on</strong>necti<strong>on</strong>s. To gain in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <strong>on</strong><br />

molecular modificati<strong>on</strong>s caused by psychosocial stress in coincidence with weaning <str<strong>on</strong>g>of</str<strong>on</strong>g> piglets, we investigated<br />

the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> social isolati<strong>on</strong> <strong>on</strong> days 7, 21 and 35 <str<strong>on</strong>g>of</str<strong>on</strong>g> age <strong>on</strong> the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> genes regulating glucocorticoid<br />

resp<strong>on</strong>se in stress-related brain regi<strong>on</strong>s. Evaluati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> glucocorticoid receptor (GR), mineralocorticoid<br />

receptor (MR) and 11ß-hydroxysteroid dehydrogenase 1 and 2 (11ß-HSD1 and 11ß-HSD2) mRNA<br />

expressi<strong>on</strong> by real-time RT-PCR revealed significant differences in quantity <str<strong>on</strong>g>of</str<strong>on</strong>g> these genes in hypothalamus,<br />

289


23-26 August 2007,<br />

Budapest, Hungary<br />

hippocampus and amygdala. A single isolati<strong>on</strong> from mother and siblings <str<strong>on</strong>g>for</str<strong>on</strong>g> 4 hours caused a significant<br />

increase <str<strong>on</strong>g>of</str<strong>on</strong>g> ACTH and cortisol c<strong>on</strong>centrati<strong>on</strong>s. The hypothalamic GR, MR and 11ß-HSD1 mRNA levels<br />

significantly increased in piglets exposed to isolati<strong>on</strong> stress, whereas in the amygdala the MR mRNA<br />

expressi<strong>on</strong> significantly decreased. The 11ß-HSD2 mRNA levels were not influenced by social isolati<strong>on</strong> in the<br />

brain regi<strong>on</strong>s. In c<strong>on</strong>clusi<strong>on</strong>s, psychosocial stress in <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> a short-term maternal deprivati<strong>on</strong> and social<br />

isolati<strong>on</strong> in piglets caused age-dependent and regi<strong>on</strong>-specific modificati<strong>on</strong>s in mRNA levels <str<strong>on</strong>g>of</str<strong>on</strong>g> stress-related<br />

genes in the brain. Furthermore, the results emphasize that these glucocorticoid regulating genes are involved<br />

in mediating emoti<strong>on</strong>al experience in pigs.<br />

290<br />

5H_06_P<br />

(poster secti<strong>on</strong> B1, poster board #234, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

INFLUENCE OF ACUTE AND LONG-TERM STRESS ON SIGA- AND CORTISOL-<br />

CONCENTRATIONS IN SALIVA OF WORKING DOGS<br />

A. Scheideler, M. Schneider, M. H. Erhard, S. Wilhelm, C. Waldhauser, S. Pauly, L. Slotta-Bachmayr,<br />

F. Ahrens<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Animal Welfare, Ethology and Animal Hygiene, Ludwig-Maximilians-University Munich<br />

In humans salivary secretory immunoglobulin A (sIgA) has been proved an objective and sensitive marker <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

stress. In animals, however, stress is hard to determine, and reliable, n<strong>on</strong>-invasive parameters are needed <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

the assessment <str<strong>on</strong>g>of</str<strong>on</strong>g> strain. Salivary cortisol is comm<strong>on</strong>ly used to detect stress resp<strong>on</strong>ses in animals but<br />

interpretati<strong>on</strong> is not always easy <str<strong>on</strong>g>for</str<strong>on</strong>g> various reas<strong>on</strong>s. In the present work sIgA is established as a dependable<br />

indicator <str<strong>on</strong>g>for</str<strong>on</strong>g> both physical and psychological stress in working dogs. Hence, the results <str<strong>on</strong>g>of</str<strong>on</strong>g> three different<br />

studies will be presented: 1) 19 search-and-rescue-dogs underwent search exercises <strong>on</strong> five different days,<br />

with 2 searches per day and a 20 min break in between; 2) 20 search-and-rescue-dogs took part in a 3-dayslasting<br />

program with 4 rubble searches per day, strains had durati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 20 min each and a 60 min break in<br />

between; 3) 26 police dogs were examined during 4 weeks <str<strong>on</strong>g>of</str<strong>on</strong>g> basic training with protecti<strong>on</strong> exercise units in<br />

weeks 1 and 4, each included a strain phase <str<strong>on</strong>g>of</str<strong>on</strong>g> 3-minute protecti<strong>on</strong> exercise. Salivary samples were collected<br />

in intervals be<str<strong>on</strong>g>for</str<strong>on</strong>g>e and after the strain periods. Salivary sIgA and cortisol were analyzed by ELISA. Resulting<br />

data from the three studies showed significantly, that acute stress leads to a decrease in sIgA from mean<br />

c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 1 g/L down to 0.2 g/L, whereas salivary cortisol c<strong>on</strong>centrati<strong>on</strong> increased. L<strong>on</strong>g-term strain<br />

resulted in an increase in sIgA in search-and-rescue-dogs during the 3-days-lasting program, while police dogs<br />

showed a reducti<strong>on</strong> after their 4-week-training.<br />

5H_07_P<br />

(poster secti<strong>on</strong> B1, poster board #235, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ENDOCRINE, HEMATOLOGICAL AND BEHAVIORAL STRESS RESPONSES TO<br />

TAIL DOCKING IN PIGS<br />

M. A. Sutherland * , P. Bryer, N. Krebs, J. J. McGl<strong>on</strong>e<br />

Pork Industry Institute, Texas Tech University, Lubbock, TX, USA<br />

*e-mail: mhairi.sutherland@ttu.edu<br />

Tail docking <str<strong>on</strong>g>of</str<strong>on</strong>g> piglets is a routine procedure <strong>on</strong> farms to prevent tail biting behavior, however docking<br />

causes an acute stress resp<strong>on</strong>se. The objectives <str<strong>on</strong>g>of</str<strong>on</strong>g> this research were to determine stress resp<strong>on</strong>ses to tail


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

docking in piglets and to compare two methods <str<strong>on</strong>g>of</str<strong>on</strong>g> tail docking; cautery ir<strong>on</strong> (CAUT) and the more<br />

comm<strong>on</strong>ly used blunt trauma cutters (BT). At approximately 6 days <str<strong>on</strong>g>of</str<strong>on</strong>g> age, piglets were tail docked using<br />

CAUT (n = 20), BT (n = 20), or sham tail docked, but their tails were left intact (CON; n = 40). Blood<br />

samples were taken prior to tail docking and at 30, 60 and 90 min after tail docking. Behavior and<br />

vocalizati<strong>on</strong>s were recorded during the tail docking procedure to measure the immediate reacti<strong>on</strong> to tail<br />

docking and in the home pen be<str<strong>on</strong>g>for</str<strong>on</strong>g>e and after tail docking. Behaviors measured included tail movement, butt<br />

movement, leg kicking and overall activity. Hematological measures did not differ (P > 0.05) am<strong>on</strong>g<br />

treatments. Cortisol c<strong>on</strong>centrati<strong>on</strong>s were higher (P < 0.05) am<strong>on</strong>g BT compared with CON piglets 60 min<br />

after tail docking, however, cortisol c<strong>on</strong>centrati<strong>on</strong>s did not differ (P > 0.05) am<strong>on</strong>g CAUT and CON piglets<br />

at this time. Furthermore, cortisol c<strong>on</strong>centrati<strong>on</strong>s were higher (P < 0.01) am<strong>on</strong>g BT than CAUT piglets 60<br />

min after tail docking. Behavioral activity and vocalizati<strong>on</strong>s were greater (P < 0.01) in docked piglets<br />

compared with CON, regardless <str<strong>on</strong>g>of</str<strong>on</strong>g> treatment. Furthermore, butt movement was greater (P < 0.005) in BT<br />

compared with CAUT piglets. The sharp pain <str<strong>on</strong>g>of</str<strong>on</strong>g> BT caused both a rise in cortisol c<strong>on</strong>centrati<strong>on</strong>s and more<br />

behavioral activity compared with CON or CAUT piglets. Combining behavioral and physiological measures<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> stress provided a more complete picture <str<strong>on</strong>g>of</str<strong>on</strong>g> piglet welfare in a challenging farm situati<strong>on</strong>.<br />

5H_08_P<br />

(poster secti<strong>on</strong> B1, poster board #236, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ANTIOXIDATIVE ACTIVITY OF PARAOXONASE-1 (PON1) AND<br />

MALONDIALDEHYDE (MDA) LEVEL THROUGHOUT PREGNANCY AND<br />

POSTPARTUM PERIOD IN DAIRY COWS<br />

Romana Turk a *, Dubravka Juretić b , Darko Gereš a , Nenad Turk a Zlata Flegar-Meštrić c , Ante Svetina a<br />

aFaculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Veterinary Medicine University <str<strong>on</strong>g>of</str<strong>on</strong>g> Zagreb, Croatia;<br />

bFaculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmacy and Biochemistry, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Zagreb, Croatia; cInstitute <str<strong>on</strong>g>of</str<strong>on</strong>g> Clinical Chemistry, Clinical<br />

Hospital “Merkur”, Zagreb, Croatia;<br />

*e-mail: rturk@vef.hr<br />

Serum PON1 is HDL-associated enzyme that hydrolyzes oxidized phospholipids generated <strong>on</strong> LDL and<br />

HDL during oxidative stress and it is c<strong>on</strong>sidered as an anti-oxidative/anti-inflammatory comp<strong>on</strong>ent <str<strong>on</strong>g>of</str<strong>on</strong>g> HDL.<br />

The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to investigate serum PON1 activity and MDA level as well as the relati<strong>on</strong>ship<br />

between PON1 activity and total cholesterol and HDL-C c<strong>on</strong>centrati<strong>on</strong>s in dairy cows during pregnancy and<br />

postpartum period. A total <str<strong>on</strong>g>of</str<strong>on</strong>g> 133 dairy cows were distributed into six groups: pregnant cows in 1 st trimester<br />

(P1, n=21), 2 nd trimester (P2, n=17) and 3 rd trimester <str<strong>on</strong>g>of</str<strong>on</strong>g> pregnancy (P3, n=10); lactating cows in the early<br />

puerperium (L1, 1-15 days postpartum, n=23), late puerperium (L2, 16-30 days postpartum, n=27) and midlactati<strong>on</strong><br />

(L3, 40-60 days postpartum, n=11). The median PON1 activity was significantly lower (P


23-26 August 2007,<br />

Budapest, Hungary<br />

5H_09_P<br />

(poster secti<strong>on</strong> B1, poster board #237, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

INDIVIDUAL DIFFERENCES IN STRESS COPING STYLES IN POLICE DOGS<br />

EXPOSED TO THREATENING SITUATION<br />

Zsuzsánna Horváth a,* , Bot<strong>on</strong>d-Zoltán Igyártó b , Attila Magyar b , Ádám Miklósi a<br />

aDepartment <str<strong>on</strong>g>of</str<strong>on</strong>g> Ethology, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, Eötvös Loránd University Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Science, Budapest, Hungary<br />

bDepartment <str<strong>on</strong>g>of</str<strong>on</strong>g> Human Morphology and Developmental Biology, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Semmelweis University,<br />

Budapest, Hungary, e-mail: horvath.zsuzsanna@gmail.com<br />

According to some researchers animals use certain coping strategies to deal with stressful situati<strong>on</strong>s. In the<br />

case <str<strong>on</strong>g>of</str<strong>on</strong>g> social carnivores social stress is a substantial part <str<strong>on</strong>g>of</str<strong>on</strong>g> the overall stress load. Previous research has<br />

established two extreme (proactive and reactive) coping styles in several animal species, but means <str<strong>on</strong>g>of</str<strong>on</strong>g> coping<br />

with social stress has not yet been investigated in the case <str<strong>on</strong>g>of</str<strong>on</strong>g> dogs. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to develop a<br />

testing procedure, where police dogs had to deal with social stress caused by a strange human approaching<br />

threateningly in the absence <str<strong>on</strong>g>of</str<strong>on</strong>g> their handler. We simultaneously measured changes <str<strong>on</strong>g>of</str<strong>on</strong>g> cortisol levels and<br />

described the behavior displayed in the course <str<strong>on</strong>g>of</str<strong>on</strong>g> the test. A factor analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> the behavioral variables<br />

discriminated three factors (‘shyness’, ‘boldness’, ‘ambivalence’). Based <strong>on</strong> these three factors as sec<strong>on</strong>dary<br />

behavioral variables a combinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> an cluster and a discriminant analysis established that in case <str<strong>on</strong>g>of</str<strong>on</strong>g> police<br />

dogs three types <str<strong>on</strong>g>of</str<strong>on</strong>g> coping strategies can be identified, namely ‘proactive’, ‘reactive’ and ‘passive’. Individuals<br />

in our ‘Proactive’ group are characterized by low HPA-axis reactivity, high level <str<strong>on</strong>g>of</str<strong>on</strong>g> activity and the short<br />

attack latency, while subjects in our ‘Reactive’ group show high HPA-axis reactivity and l<strong>on</strong>g attack latency.<br />

Our ‘Passive’ group c<strong>on</strong>sists <str<strong>on</strong>g>of</str<strong>on</strong>g> the least active individuals, they failed to show attack behavior and reacted to<br />

approaching human with passivity and submissi<strong>on</strong>, while their HPA-axis reactivity has been normal.<br />

292<br />

5H_10_P<br />

(poster secti<strong>on</strong> B1, poster board #238, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

NEW POTENTIALS FOR THE DETECTION OF LONG-TERM STRESS IN FRESH-<br />

WATER FISH SPECIES<br />

Árpád Hegyi, Katalin Kinga Lefler, Zsolt Csenki, Ákos Horváth, Béla Urbányi<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Fish Culture, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Agricultural and Envir<strong>on</strong>mental Sciences, Szent István University,<br />

Gödöllő, H-2103 Páter Károly u. 1., Hungary<br />

e-mail: Hegyi.Arpad@mkk.szie.hu<br />

In<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> about l<strong>on</strong>g-term stress is gaining more importance in fish cultures. As a result <str<strong>on</strong>g>of</str<strong>on</strong>g> unfavorable<br />

weather changes and more intensive p<strong>on</strong>d fish producti<strong>on</strong> many different stressors can have an effect <strong>on</strong> fish<br />

and induce essential metabolic disturbances. Most important stressors are significant changes in water<br />

temperature (Lyytikainen et al. 2002), lasting lack <str<strong>on</strong>g>of</str<strong>on</strong>g> oxygen (Henrique et al. 2002), increasing stocking density<br />

(Bart<strong>on</strong> et al. 1985) and handling <str<strong>on</strong>g>of</str<strong>on</strong>g> fish (Hoeger et al. 2004). Under the influence <str<strong>on</strong>g>of</str<strong>on</strong>g> these stressors<br />

disturbance appears in carbohydrate, protein and lipid metabolism and processes <str<strong>on</strong>g>of</str<strong>on</strong>g> electrolyte transport can<br />

also be damaged.<br />

Measurement <str<strong>on</strong>g>of</str<strong>on</strong>g> serum/plasma fructosamine (SeFa) applied routinely in human diabetes (diabetes mellitus) was<br />

worked out <strong>on</strong> fresh-water fish species which is exceedingly suitable <str<strong>on</strong>g>for</str<strong>on</strong>g> the detecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> l<strong>on</strong>g-term stress.<br />

Effect <str<strong>on</strong>g>of</str<strong>on</strong>g> many stressors was already examined <strong>on</strong> Prussian carp (Carassius auratus gibelio BLOCH, 1783) and<br />

comm<strong>on</strong> carp (Cyprinus carpio L., 1758) (Hegyi et al. 2006), by which the utility <str<strong>on</strong>g>of</str<strong>on</strong>g> SeFa was proved.


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Blood plasma cortisol, glucose and SeFa levels were determined <str<strong>on</strong>g>for</str<strong>on</strong>g> grass carp (Ctenopharyngod<strong>on</strong> idella<br />

VALENCIENNES, 1844) kept in an oxygen deficient envir<strong>on</strong>ment <str<strong>on</strong>g>for</str<strong>on</strong>g> four weeks. Changes in the quantity <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

blood plasma comp<strong>on</strong>ents were followed up by cupping <strong>on</strong>ce in every week. As an effect <str<strong>on</strong>g>of</str<strong>on</strong>g> corticotropreleasing<br />

factor (CRF) by the end <str<strong>on</strong>g>of</str<strong>on</strong>g> the experiment the amount <str<strong>on</strong>g>of</str<strong>on</strong>g> cortisol significantly decreased (P


23-26 August 2007,<br />

Budapest, Hungary<br />

294<br />

5I. STRESS AND NO<br />

5I_01_P<br />

(poster secti<strong>on</strong> B1, poster board #239, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

WITHANIA SOMNIFERA INDUCES THE STRESS INDUCED MODULATION OF<br />

NEUROBEHAVIORAL PATTERNS VIA NITRIC OXIDE SYNTHASE<br />

Zaved Ahmed Khan 1 , Iftikhar Aslam Tayubi 2<br />

1,2School <str<strong>on</strong>g>of</str<strong>on</strong>g> Biotechnology , Chemical and Biomedical Engineering,Vellore Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Technology University, Vellore<br />

TN-632014, India<br />

Withania somnifera (ashwagandha, Indian ginseng) is an immunostimulant herbal medicine used to improve<br />

overall health and prevent diseases, particularly in the elderly. We had found to be an effective stress<br />

modulator herbal medicine. However, the mechanisms underlying its anxiolytics effect is poorly understood.<br />

To elucidate the mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> Withania somnifera, we investigated the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> a methanolic extract from<br />

the root and stem <str<strong>on</strong>g>of</str<strong>on</strong>g> Withania somnifera (WS) <strong>on</strong> nitric oxide (NO) producti<strong>on</strong> in wistar rats.We found that<br />

WS (1–256 µg/ml) produced a significant and c<strong>on</strong>centrati<strong>on</strong>-dependent increase in NO producti<strong>on</strong>, an effect<br />

which was abolished by N-nitro-L-arginine methyl ester (L-NAME, 3–300 µM), a n<strong>on</strong>selective inhibitor <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

NO synthase (NOS).As it has been already shown about the involvement <str<strong>on</strong>g>of</str<strong>on</strong>g> nitric oxide (NO) in stressinduced<br />

neurobehavioral changes in rats .Moreover, elevated plus maze (EPM) and open field test (OFT)<br />

analysis showed that WS increased, in a c<strong>on</strong>centrati<strong>on</strong>-dependent fashi<strong>on</strong> modulates stress induced<br />

neurobehavioral patterns. Activity <str<strong>on</strong>g>of</str<strong>on</strong>g> NOS was checked using NOS detecti<strong>on</strong> system(Sigma FCANOS ) in<br />

brain sample. These results dem<strong>on</strong>strate that WS may induce the synthesis <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>al NOS expressi<strong>on</strong> likely<br />

by acting at transcripti<strong>on</strong>al level. nNOS level is checked by western blot analysis. The increased NO<br />

producti<strong>on</strong> by brain could account, at least in part, <str<strong>on</strong>g>for</str<strong>on</strong>g> the anxiolytics properties <str<strong>on</strong>g>of</str<strong>on</strong>g> Withania somnifera.<br />

5I_02_P<br />

(poster secti<strong>on</strong> B1, poster board #240, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ROOT DEVELOPMENT UNDER OSMOTIC STRESS AND IN THE PRESENCE OF<br />

EXOGENOUS AUXIN IN PISUM SATIVUM L.: THE ROLE OF NITRIC OXIDE<br />

Zs. Kolbert, B. Bartha, L. Erdei<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Physiology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Szeged, P.O.B. 654., 6701 Szeged, Hungary<br />

e-mail: kolzsu@bio.u-szeged.hu; erdei@bio.u-szeged.hu<br />

During this work the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> exogenous auxin (indolebutyric acid, IBA) and osmotic stress <strong>on</strong> root<br />

morphology and nitric oxide (NO) generati<strong>on</strong> in roots were compared in pea plants. Five-day old plants were<br />

treated with IBA at a c<strong>on</strong>centrati<strong>on</strong> range <str<strong>on</strong>g>of</str<strong>on</strong>g> 10 -9 - 10 -3 M and osmotic stress was induced by 50-400 mOsm<br />

polyethylene glycol (PEG 6000) <str<strong>on</strong>g>for</str<strong>on</strong>g> another 5 days. NO generati<strong>on</strong> was examined by in situ and in vivo<br />

fluorescence method. Both root characteristics and NO intensities were followed as the functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> time. The<br />

increasing c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> IBA as well as PEG resulted in shortening <str<strong>on</strong>g>of</str<strong>on</strong>g> primary roots (PRs), enhancement<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> lateral root (LR) number and significant increase <str<strong>on</strong>g>of</str<strong>on</strong>g> NO generati<strong>on</strong>. By virtual secti<strong>on</strong>ing <str<strong>on</strong>g>of</str<strong>on</strong>g> PRs, it was<br />

determined that the accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> NO occurred at the basis <str<strong>on</strong>g>of</str<strong>on</strong>g> LRs during both treatments. Timedependence<br />

investigati<strong>on</strong>s revealed that in the case <str<strong>on</strong>g>of</str<strong>on</strong>g> IBA treatments, the LR number increased in parallel<br />

with an intensified NO generati<strong>on</strong>, while el<strong>on</strong>gati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> PRs was not followed by changes in NO levels.<br />

During osmotic stress, the time curve <str<strong>on</strong>g>of</str<strong>on</strong>g> LR development was similar to that <str<strong>on</strong>g>of</str<strong>on</strong>g> IBA-treated roots, but,


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

significantly, it was preceded by a burst <str<strong>on</strong>g>of</str<strong>on</strong>g> NO. This early phase <str<strong>on</strong>g>of</str<strong>on</strong>g> NO generati<strong>on</strong> under osmotic stress was<br />

clearly distinguishable from that which accompanies LR initiati<strong>on</strong>. It is c<strong>on</strong>cluded that osmotic stress and the<br />

presence <str<strong>on</strong>g>of</str<strong>on</strong>g> exogenous auxin resulted in partly similar root architecture but different time courses <str<strong>on</strong>g>of</str<strong>on</strong>g> NO<br />

synthesis. We suppose that the early phase <str<strong>on</strong>g>of</str<strong>on</strong>g> NO generati<strong>on</strong> may fulfil a role in the osmotic stress-induced<br />

signalizati<strong>on</strong> process leading to the modificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> root morphology.<br />

5I_03_P<br />

(poster secti<strong>on</strong> B1, poster board #241, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EVALUATION OF IN VITRO LDL OXIDATION MEDIATED BY CUPPER AND<br />

NITRIC OXIDE PRODUCTION IN HUVEC IN PRESENCE OF HUMAN<br />

DYSLIPIDEMIC PLASMAS<br />

L. Lamperti, P. Binder, D. Nova, I. G<strong>on</strong>zález, A. Searle, R. Aguilar, C. Aguayo<br />

Clinic Biochemistry and Immunology Department, Pharmacy Faculty, POX-237, University <str<strong>on</strong>g>of</str<strong>on</strong>g> C<strong>on</strong>cepción, Chile<br />

e-mail: llampert@udec.cl<br />

Low-density lipoprotein (LDL) is involved in the pathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> atherosclerotic lesi<strong>on</strong>s, through modifying<br />

processes such as oxidati<strong>on</strong>. We examined the in vitro susceptibility to oxidati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> LDL isolated from normal<br />

individuals in presence <str<strong>on</strong>g>of</str<strong>on</strong>g> hypercholesterolemic human plasma and we assayed the same plasmas to evaluate<br />

the effects <strong>on</strong> the nitric oxide (NO) producti<strong>on</strong> in human fetal endothelial cells (HUVEC). Plasma from 20<br />

dyslipidemic patients and 10 normal individuals were analyzed <str<strong>on</strong>g>for</str<strong>on</strong>g> total cholesterol (TC), high density<br />

lipoprotein (HDL), LDL, triglycerides and the ratio TC/HDL was calculated. The LDL susceptibility to<br />

oxidati<strong>on</strong> was measured and expressed in sec<strong>on</strong>ds (lag phase and maximal rate time (MRT) <str<strong>on</strong>g>for</str<strong>on</strong>g> c<strong>on</strong>jugated<br />

dienes <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>). The NO producti<strong>on</strong> was evidenced by florescence microscopy and flow cytometry (FC)<br />

using 4,5-Diamin<str<strong>on</strong>g>of</str<strong>on</strong>g>luorescein Diacetate (DAF-2 DA). The kinetic curve <str<strong>on</strong>g>for</str<strong>on</strong>g> isolated normal c<strong>on</strong>trol LDL (50<br />

ug/mL, 3 uM Cu) shown a lag phase <str<strong>on</strong>g>of</str<strong>on</strong>g> 3145s ±43 and MRT <str<strong>on</strong>g>of</str<strong>on</strong>g> 5030s ±80. Plasma <str<strong>on</strong>g>of</str<strong>on</strong>g> patients with high<br />

cholesterol (>200 mg/dl) exhibited a shorter MRT <str<strong>on</strong>g>for</str<strong>on</strong>g> c<strong>on</strong>jugated dienes (-15% ±4 vs c<strong>on</strong>trol MRT) when<br />

the ratio TC/HDL is > 5.0, but MRT is l<strong>on</strong>ger (+25% ±11 vs c<strong>on</strong>trol MRT) when the hypercholesterolemic<br />

patients ratio TC/HDL is 5 (62.2% ±3.8 vs 49.2% ±6.2 normal cholesterol plasma). The<br />

mayor susceptibility to LDL oxidati<strong>on</strong> showed a correlati<strong>on</strong> with a minor disp<strong>on</strong>ibility <str<strong>on</strong>g>of</str<strong>on</strong>g> NO in<br />

hypercholesterolemic patients.<br />

5I_04_P<br />

(poster secti<strong>on</strong> B1, poster board #242, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

UNCOUPLING ENDOTHELIAL NITRIC OXIDE SYNTHASE CAUSES<br />

ENDOTHELIAL CELL STRESS BY SUPEROXIDE FORMATION DUE TO<br />

DIMINISHED SUPPLY OF THE COFACTOR TETRAHYDROBIOPTERIN<br />

Pavel Martasek † , Jeannette Vasquez-Vivar § , B. Kalyanaraman ‡<br />

†Charles University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine I and Univ. General Hospital, Dpt <str<strong>on</strong>g>of</str<strong>on</strong>g> Pediatrics, 128 08 Prague,<br />

Czech Republic, e-mail: pavel.martasek@gmail.com<br />

§Dpt <str<strong>on</strong>g>of</str<strong>on</strong>g> Biophysics, Medical College <str<strong>on</strong>g>of</str<strong>on</strong>g> Wisc<strong>on</strong>sin, 8701 Watertown Plank Road, Milwaukee, WI 53226, USA<br />

295


23-26 August 2007,<br />

Budapest, Hungary<br />

In blood vessels, superoxide ani<strong>on</strong> radical (O •− 2 ) <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> has been ascribed to different enzymes, am<strong>on</strong>g<br />

them endothelial nitric oxide synthase (eNOS). The improved methods <str<strong>on</strong>g>for</str<strong>on</strong>g> the detecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> O •− 2 has greatly<br />

c<strong>on</strong>tributed to the identificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the mechanisms involved in the shift <str<strong>on</strong>g>of</str<strong>on</strong>g> eNOS activity from a NO to a<br />

O •−<br />

2 synthase. This is <str<strong>on</strong>g>of</str<strong>on</strong>g> high importance because this shift in activity plays a significant role in the<br />

pathophysiological mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> vascular disease. In this presentati<strong>on</strong>, we show evidence <str<strong>on</strong>g>for</str<strong>on</strong>g> the role <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

c<str<strong>on</strong>g>of</str<strong>on</strong>g>actor tetrahydrobiopterin (BH4) as the molecular switch that c<strong>on</strong>trols O •− 2 release from eNOS. BH4<br />

effects were shown to be dependent <strong>on</strong> both its redox state and c<strong>on</strong>centrati<strong>on</strong>. For the enzyme reacti<strong>on</strong> to be<br />

coupled and <str<strong>on</strong>g>for</str<strong>on</strong>g>m exclusively NO, however, requires optimal BH4 supply. In the absence <str<strong>on</strong>g>of</str<strong>on</strong>g> BH4, eNOS<br />

generates superoxide from the breakdown <str<strong>on</strong>g>of</str<strong>on</strong>g> the heme–dioxygen complex at the oxygenase domain <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

enzyme. Asymmetrical-dimethyl-L-argine (ADMA) or L-NMMA did not alter O •− 2 release, and L-NAME<br />

caused a partial inhibiti<strong>on</strong>. Examining the role <str<strong>on</strong>g>of</str<strong>on</strong>g> eNOS protein interacti<strong>on</strong> revealed that caveolin-1-<br />

scaffolding peptide inhibits O •− 2 producti<strong>on</strong> in a dose dependent manner. Overall, the data presented indicate<br />

that O •− 2 producti<strong>on</strong> from eNOS is increased by the diminished availability <str<strong>on</strong>g>of</str<strong>on</strong>g> the reduced BH4 c<str<strong>on</strong>g>of</str<strong>on</strong>g>actor but<br />

not by deficient levels <str<strong>on</strong>g>of</str<strong>on</strong>g> L-arginine or accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> methylated L-arginine in the endothelium.<br />

5I_05_P<br />

(poster secti<strong>on</strong> B1, poster board #243, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE EFFECT OF BERBERIS VULGARIS ON INFLAMMATION-INDUCED STRESS<br />

Elena Parvu Alina 1 , M. Parvu 2 , Cristina Mogosan 3 , M. Tamas 4<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine and Pharmacy Iuliu Hatieganu Cluj-Napoca –<br />

1Pathophisiology,<br />

3Pharmacology,<br />

4Pharmaceutical Botany;<br />

2University Babes-Bolyai Cluj-Napoca - Biology Faculty<br />

1 Adress: Emil Isac Street 13, 400020 Cluj-Napoca, Romania,<br />

e-mail: parvualinaelena@yahoo.com<br />

Excessive nitric oxide (NO) synthesis by nitric oxide synthase 2 (NOS2) has been implicated in the<br />

pathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> inflammatory illness, including inflammati<strong>on</strong>-induced stress. Novel therapeutic approaches<br />

have changed the trend towards pharmacologic modulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> exaggerated host resp<strong>on</strong>se, namely host<br />

modulatory therapy. That is why the present study evaluated Berberis vulgaris extract effect <strong>on</strong> NO synthesis in<br />

experimental inflammati<strong>on</strong> models. There have been used 10 groups <str<strong>on</strong>g>of</str<strong>on</strong>g> Wistar-Bratislawa male rats: 2 groups<br />

with experimental inflammati<strong>on</strong> (turpentine-induced acute inflamati<strong>on</strong>, Freund’s Adjuvant-induced arthritis),<br />

4 groups with experimental inflammati<strong>on</strong> treated with Berberis vulgaris extract (administrated i.p. in two<br />

diluti<strong>on</strong>s: 0,2 %, respectively 0,02% expressed in berberine), 2 groups with experimental inflammati<strong>on</strong> treated<br />

with a n<strong>on</strong>selective NOS inhibitor (L-NAME), and 2 groups with experimental inflammati<strong>on</strong> treated with<br />

ibupr<str<strong>on</strong>g>of</str<strong>on</strong>g>en, as an NOS2 inhibitor. After 24 hours from turpentine administrati<strong>on</strong>, respectivelly after 4 weeks<br />

from Freund’s Adjuvant administrati<strong>on</strong>, NO synthesis was appreciated in all groups by measuring serum<br />

nitrite/nitrates (Griess) and citrulline c<strong>on</strong>centrati<strong>on</strong>s. C<strong>on</strong>clusi<strong>on</strong>s: 1. Berberis vulgaris extract significantly<br />

reduced NO synthesis in experimental inflammati<strong>on</strong> models; 2. The higer diluti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Berberis vulgaris extract<br />

had a str<strong>on</strong>ger inhibitory effect <strong>on</strong> NO synthesis; 3. The effect <str<strong>on</strong>g>of</str<strong>on</strong>g> Berberis vulgaris extract was smaller than that<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> L-NAME and ibupr<str<strong>on</strong>g>of</str<strong>on</strong>g>en.<br />

296


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5I_06_P<br />

(poster secti<strong>on</strong> B1, poster board #244, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PRELIMINARY APPLICATION OF HIGH SPEED NO-METABOLITE-ASSAY IN<br />

HUMAN SALIVA FOR EXERCISE STRESS USING LAB-ON-A-CHIP TECHNOLOGY<br />

Shin-ichi Wakida, Takashi Miyado, Yoshihide Tanaka, Hidenori Nagai, Yasukazu Yoshida,<br />

Etsuo Niki<br />

Human Stress Signal <str<strong>on</strong>g>Research</str<strong>on</strong>g> Center, Nati<strong>on</strong>al Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Advanced Industrial Science and Technology (AIST),<br />

Japan, Ikeda, Osaka 563-8577, Japan<br />

e-mail: s.wakida@aist.go.jp<br />

Nitric oxide (NO) has been identified as a mediator in many physiological functi<strong>on</strong>s. Several research<br />

scientists have reported the possibility that acute exercise induced endothelium-dependent vasodilati<strong>on</strong> is due<br />

mainly to an increase in NO release and also discussed NO metabolites change in saliva. In general, NO<br />

metabolites assay is used by the Griess reacti<strong>on</strong>, however, the assay is time-c<strong>on</strong>suming. We have studied high<br />

speed NO assay in human saliva using Lab-<strong>on</strong>-a-Chip integrated with analytical procedures, such as <strong>on</strong>-chip<br />

pre-c<strong>on</strong>centrati<strong>on</strong>, separati<strong>on</strong> and simultaneous detecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> NO metabolites, NO2- and NO3- within 1<br />

minutes including <str<strong>on</strong>g>of</str<strong>on</strong>g>f-line sample pre-treatment with 10-fold pure water diluti<strong>on</strong>. In this paper, we will<br />

introduce the high speed Lab-<strong>on</strong>-a-Chip assay based <strong>on</strong> capillary z<strong>on</strong>e electrophoresis with UV detecti<strong>on</strong><br />

method using a novel running buffer and an advanced micr<str<strong>on</strong>g>of</str<strong>on</strong>g>luidic c<strong>on</strong>trol. We achieved complete separati<strong>on</strong><br />

assay <str<strong>on</strong>g>of</str<strong>on</strong>g> NO metabolites in 10-fold diluted saliva within 15 sec<strong>on</strong>ds. And we will also introduce preliminary<br />

applicati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> exercise tolerance test using bicycle ergometer. The exercise tolerance level was changed<br />

stepwisely from 0 to 183 W <str<strong>on</strong>g>for</str<strong>on</strong>g> 15 minutes. Human saliva samples were collected be<str<strong>on</strong>g>for</str<strong>on</strong>g>e and after the test in<br />

high exercise intensity group, 6 healthy volunteer (5 female and 1 male aged from 40 to 66 years) and low<br />

exercise intensity group, 4 healthy volunteer (1 female and 3 male aged from 21 to 47 years). The preliminary<br />

results <str<strong>on</strong>g>of</str<strong>on</strong>g> the change <str<strong>on</strong>g>of</str<strong>on</strong>g> NO metabolites in saliva will be introduced and discussed in the c<strong>on</strong>gress.<br />

297


23-26 August 2007,<br />

Budapest, Hungary<br />

298<br />

5J. OTHER STRESS TOPICS AT LEVEL OF THE ORGANISM (GRAVITY STRESS,<br />

STRESS OF VIRAL INFECTIONS, ACUTE PHASE RESPONSE,<br />

HSP-S IN DEVELOPMENT, RADIATION STRESS – ELECTROSMOG)<br />

5J_01_P<br />

(poster secti<strong>on</strong> B1, poster board #245, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HYPERGRAVITY STRESS RESPONSE IN RAT HYPOTHALAMUS AND<br />

MODULATION BY VESTIBULAR BLOCKADE<br />

Yasuhiro Kumei 1 *, Jorge Zeredo 2 , Mari Kimoto 3 , Tohru Ikeda 2 , Kazuo Toda 2<br />

1Graduate School <str<strong>on</strong>g>of</str<strong>on</strong>g> Tokyo Medical and Dental University, Tokyo 113-8549<br />

2Graduate School <str<strong>on</strong>g>of</str<strong>on</strong>g> Nagasaki University, 852-8588 Nagasaki<br />

3Japan Women’s University, Tokyo 112-8681, Japan<br />

*e-mail: kumei.bch@tmd.ac.jp<br />

Spaceflight influences vestibular functi<strong>on</strong> and causes moti<strong>on</strong> sickness in astr<strong>on</strong>auts. Stress resp<strong>on</strong>se is<br />

induced by microgravity and other spaceflight c<strong>on</strong>diti<strong>on</strong>s. However, little is known about the relati<strong>on</strong>ship<br />

between stress resp<strong>on</strong>se and vestibular functi<strong>on</strong> in gravitati<strong>on</strong>al alterati<strong>on</strong>. The purpose <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to<br />

examine the hypergravity effects <strong>on</strong> the neur<strong>on</strong>al activities <str<strong>on</strong>g>of</str<strong>on</strong>g> hypothalamic arcuate nucleus (ARC) in the<br />

presence and absence <str<strong>on</strong>g>of</str<strong>on</strong>g> vestibular functi<strong>on</strong> in rats. Young male Wistar rats were exposed to gradually<br />

increasing gravity-<str<strong>on</strong>g>for</str<strong>on</strong>g>ce in a light-blocked dark centrifuge. Electrophysiological data were obtained through<br />

chr<strong>on</strong>ically implanted electrodes by telemetrical recording <str<strong>on</strong>g>of</str<strong>on</strong>g> the unit activities at ARC with or without<br />

injecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> xylocaine into both inner ears. The firing frequency in the ARC started to increase when the<br />

centrifugal <str<strong>on</strong>g>for</str<strong>on</strong>g>ce reached to 2.0G, 40 sec<strong>on</strong>ds after initiating centrifugati<strong>on</strong>. By c<strong>on</strong>trast, the firing frequency<br />

in the ARC <str<strong>on</strong>g>of</str<strong>on</strong>g> xylocaine-injected rat started to increase when the centrifugal <str<strong>on</strong>g>for</str<strong>on</strong>g>ce reached 1.4G, 15 sec<strong>on</strong>ds<br />

after starting centrifugati<strong>on</strong>. Thus, the latent period <str<strong>on</strong>g>for</str<strong>on</strong>g> hypergravity-resp<strong>on</strong>sive ARC activity was significantly<br />

shortened by vestibular blockade. On the following day when xylocaine effects were expired, the ARC<br />

resp<strong>on</strong>siveness to hypergravity resumed the basal level. Data suggest that the vestibular functi<strong>on</strong> may<br />

modulate the gravity-stress resp<strong>on</strong>se in rat hypothalamus. Supported by Grant-in-aid from the Ministry <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Educati<strong>on</strong>, Science, and Culture, Japan.<br />

5J_02_P<br />

(poster secti<strong>on</strong> B1, poster board #246, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

GRAVITATIONAL STRESS ON SYMPATHETIC NERVE ACTIVITY IN HUMANS<br />

Tadaaki Mano, Satoshi Iwase<br />

Gifu University <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Science, Seki, Gifu 501-3892, Japan and Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Physiology, Aichi Medical<br />

University, Nagakute, Aichi 480-1195, Japan, e-mail: tadaaki.mano@nifty.com<br />

Gravity is an important envir<strong>on</strong>mental stress <str<strong>on</strong>g>for</str<strong>on</strong>g> human being who maintains upright posture. Neural and<br />

humoral mechanisms operate to maintain upright posture in humans against gravitati<strong>on</strong>al stress. One <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

mechanisms is sympathetic neural c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> blood pressure. Sympathetic neural traffic leading to skeletal<br />

muscles called muscle sympathetic nerve activity (MSNA) has been proved to play an essential role to<br />

maintain blood pressure homeostasis through baroreflexes. The present study aimed to clarify how<br />

gravitati<strong>on</strong>al stress influences MSNA in humans. Subjects were 20 healthy male subjects ranging in age<br />

between 18 and 24 years. MSNA was recorded and identified from the tibial nerve using micr<strong>on</strong>eurography


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

(1), simultaneously with hemodynamic functi<strong>on</strong>s. In 10 subjects, gravitati<strong>on</strong>al stress was loaded by head-up<br />

tilt changing posture from supine to standing using a tilt table. We analyzed how gravitati<strong>on</strong>al stress from the<br />

head to the leg (+Gz) expressed as sine functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the tilt angle influences MSNA and cardiovascular<br />

functi<strong>on</strong>s. We also analyzed MSNA resp<strong>on</strong>ses in subjects who complained <str<strong>on</strong>g>of</str<strong>on</strong>g> orthostatic hypotensi<strong>on</strong> during<br />

head-up tilt. In another 10 subjects, MSNA was measured during exposure to short- and l<strong>on</strong>g-term<br />

microgravity induced by parabolic flight and simulated by head-down bed rest <str<strong>on</strong>g>for</str<strong>on</strong>g> 14 days, respectively. We<br />

analyzed the influence <str<strong>on</strong>g>of</str<strong>on</strong>g> microgravity <strong>on</strong> MSNA and compared the results with changes in the same activity<br />

reported in microgravity in space. We c<strong>on</strong>clude that +Gz stress is a str<strong>on</strong>g activator <str<strong>on</strong>g>of</str<strong>on</strong>g> sympathetic neural<br />

traffic leading to skeletal muscles in humans to maintain blood pressure homeostasis during standing.<br />

Excessively low and high sympathetic resp<strong>on</strong>ders to gravitati<strong>on</strong>al stress can cause orthostatic hypotensi<strong>on</strong>.<br />

Short-term microgravity suppresses sympathetic nerve activity to muscles but l<strong>on</strong>g-term exposure to<br />

simulated microgravity rather enhances the same activity as reported in spaceflight depending <strong>on</strong> complex<br />

mechanisms including plasma volume reducti<strong>on</strong>.<br />

5J_03_P<br />

(poster secti<strong>on</strong> B1, poster board #247, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HSP72 DETERMINES THE OUTCOME OF VIRAL INFECTION IN BRAIN<br />

Michael Oglesbee, Thomas Carsillo<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Veterinary Biosciences, The Ohio State University, Columbus, OH;<br />

e-mail: oglesbee.1@osu.edu<br />

Fever is a potent inducer <str<strong>on</strong>g>of</str<strong>on</strong>g> 70 kDa heat shock proteins, particularly hsp72, and in vitro studies show that<br />

hsp72 supports replicati<strong>on</strong> and gene expressi<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> multiple viral families. Despite the frequent occurrence <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

fever in viral infecti<strong>on</strong>, the biological (i.e., in vivo) significance <str<strong>on</strong>g>of</str<strong>on</strong>g> virus-hsp72 interacti<strong>on</strong> is poorly defined.<br />

The present work addresses the in vivo significance <str<strong>on</strong>g>of</str<strong>on</strong>g> virus-hsp72 interacti<strong>on</strong> using a mouse model <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

measles virus (MV) encephalitis. Hsp72 stimulates MV transcripti<strong>on</strong>, a resp<strong>on</strong>se that is disrupted in an N<br />

protein amino acid substituti<strong>on</strong> mutant (N522D). Ne<strong>on</strong>atal mice receiving an intracranial inoculati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> MV<br />

are afebrile and exhibit H-2 restricted differences in susceptibility to infecti<strong>on</strong>; H-2 d mice are resistant to<br />

infecti<strong>on</strong> due to robust adaptive antiviral immune resp<strong>on</strong>ses whereas H-2 b are susceptible due to deficiencies<br />

in this resp<strong>on</strong>se. Here we show that transgenic overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp72 increases MV transcripti<strong>on</strong> within<br />

infected brains <str<strong>on</strong>g>of</str<strong>on</strong>g> H-2 b (susceptible) C57BL/6 mice, resulting in increased viral protein expressi<strong>on</strong>, CPE and<br />

virus-induced mortality. In c<strong>on</strong>trast, overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp72 in c<strong>on</strong>genic C57BL/10 having the H-2 d<br />

resistance phenotype are completely protected against MV challenge, compared to > 30% mortality in n<strong>on</strong>transgenic<br />

mice. Protecti<strong>on</strong> reflects a viral transcripti<strong>on</strong>al resp<strong>on</strong>se to hsp72 based up<strong>on</strong> no significant<br />

differences in infecti<strong>on</strong>s by the N522D variant in transgenic versus n<strong>on</strong>-transgenic mice. Collectively, these<br />

findings suggest that hsp72-mediated stimulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> viral transcripti<strong>on</strong> (and thus antigenic burden) can<br />

facilitate adaptive immune resp<strong>on</strong>se leading to viral clearance, but if the host is immune compromised, this<br />

host protective strategy back-fires, leading to enhanced viral virulence.<br />

299


23-26 August 2007,<br />

Budapest, Hungary<br />

300<br />

5J_04_P<br />

(poster secti<strong>on</strong> B1, poster board #248, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CHRONIC STRESS AFFECTS THE EXPRESSION OF NEUROTROPHINS IN THE<br />

RAT SALIVARY GLANDS<br />

Juri Saruta 1* , Ko Nakajima 1 , Lee Tae Ki 1 , Masayoshi Shirasu 1 , Takeshi Takahashi 1 , Chikatoshi Sato 1 ,<br />

Keiichi Tsukinoki 2 , Sadao Sato 1<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> 1 Crani<str<strong>on</strong>g>of</str<strong>on</strong>g>acial Growth and Development Dentistry, Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Orthod<strong>on</strong>tics, 2 Maxill<str<strong>on</strong>g>of</str<strong>on</strong>g>acial Diagnostic<br />

Science, Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Pathology, Kanagawa Dental College, Yokosuka, Japan;<br />

*e-mail: sarujuri@kdcnet.ac.jp<br />

Purpose <str<strong>on</strong>g>of</str<strong>on</strong>g> this experiment; In the previous study, we reported that BDNF was produced from salivary gland<br />

in the c<strong>on</strong>diti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> acute immobilizati<strong>on</strong> stress. In additi<strong>on</strong>, salivary glands were origin <str<strong>on</strong>g>of</str<strong>on</strong>g> plasma BDNF in<br />

that c<strong>on</strong>diti<strong>on</strong>. Next, in chr<strong>on</strong>ic immobilizati<strong>on</strong> stress, we thought to examine whether there is a relati<strong>on</strong>ship<br />

between salivary gland and neurotrophins expressi<strong>on</strong>.<br />

Materials & Methods; Chr<strong>on</strong>ic stress group was also per<str<strong>on</strong>g>for</str<strong>on</strong>g>med against rats which were resected the bilateral<br />

major salivary glands (sialoadenoectomy). Chr<strong>on</strong>ic stress was induced by enclosing each animal in flexible<br />

wire mesh shaped to fit its body.<br />

Results; Increased neurotrophins mRNA and protein expressi<strong>on</strong> were observed in duct cells as a result <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

chr<strong>on</strong>ic stress, as dem<strong>on</strong>strated by real-time PCR, immunohistochemistry and ELISA. Chr<strong>on</strong>ic stress<br />

significantly increased the level <str<strong>on</strong>g>of</str<strong>on</strong>g> plasma neurotrophins. There were significant differences between n<strong>on</strong>stress<br />

and chr<strong>on</strong>ic stress in the plasma neurotrophins level (p>0.05). Rat submandibular gland was identified<br />

as an organ which expresses neurotrophins. Furthermore, the results <str<strong>on</strong>g>of</str<strong>on</strong>g> this study suggest that increased<br />

salivary neurotrophins expressi<strong>on</strong> occurs following chr<strong>on</strong>ic stress. There were no significant differences<br />

between stressed rats and stressed sialoadenoectomy rats in the plasma neurotrophins level.<br />

C<strong>on</strong>clusi<strong>on</strong>; Our data, there<str<strong>on</strong>g>for</str<strong>on</strong>g>e, suggest an existence <str<strong>on</strong>g>of</str<strong>on</strong>g> a different between acute and chr<strong>on</strong>ic stress was<br />

suggested in source <str<strong>on</strong>g>of</str<strong>on</strong>g> plasma BDNF and NT-3. Future studies are needed to examine the origin <str<strong>on</strong>g>of</str<strong>on</strong>g> plasma<br />

BDNF and NT-3 in chr<strong>on</strong>ic stress.<br />

5J_05_P<br />

(poster secti<strong>on</strong> B1, poster board #249, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ACUTE IMMOBILIZATION STRESS AFFECTS THE EXPRESSION OF BDNF IN<br />

THE RAT SALIVARY GLANDS<br />

Keiichi Tsukinoki 1 , Juri Saruta 2 , Lee Tae Ki 2 , Sadao Sato 2<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> 1 Maxill<str<strong>on</strong>g>of</str<strong>on</strong>g>acial Diagnostic Science, Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Pathology,<br />

2Crani<str<strong>on</strong>g>of</str<strong>on</strong>g>acial Growth and Development Dentistry, Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Orthod<strong>on</strong>tics,<br />

Kanagawa Dental College, Yokosuka, Japan;<br />

*e-mail: ktsukino@kdcnet.ac.jp<br />

Purpose <str<strong>on</strong>g>of</str<strong>on</strong>g> this experiment: The present study examined the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> immobilizati<strong>on</strong> stress <strong>on</strong> BDNF and<br />

TrkB expressi<strong>on</strong> in male rat submandibular glands.<br />

Materials & Methods; Male Sprague-Dawley rats, 9 weeks <str<strong>on</strong>g>of</str<strong>on</strong>g> age were used in this study. All experiments<br />

were per<str<strong>on</strong>g>for</str<strong>on</strong>g>med using four rats per group. The rats were exposed to immobilizati<strong>on</strong> stress <str<strong>on</strong>g>for</str<strong>on</strong>g> 30 min, 60<br />

min, or 180 min.


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Results; Increased BDNF mRNA and protein expressi<strong>on</strong> were observed in duct cells as a result <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

immobilizati<strong>on</strong> stress, as dem<strong>on</strong>strated by real-time PCR, western blot, immunohistochemistry, and analysis<br />

by microdissecti<strong>on</strong>. TrkB mRNA was not detected in salivary gland tissue, or oral or esophageal mucosa, by<br />

RT-PCR. Rat submandibular gland was thus identified as an organ which expresses BDNF. Acute<br />

immobilizati<strong>on</strong> stress <str<strong>on</strong>g>for</str<strong>on</strong>g> 60 min significantly increased the level <str<strong>on</strong>g>of</str<strong>on</strong>g> plasma BDNF. However, plasma BDNF<br />

elevati<strong>on</strong> was markedly suppressed in bilaterally sialoadenectomized rats. There were no significant<br />

differences between stressed (60 min) and n<strong>on</strong>-stressed rats with respect to the BDNF mRNA expressi<strong>on</strong> in<br />

the hippocampus, heart, lung, liver, pancreas, or spleen, as determined by real-time PCR.<br />

C<strong>on</strong>clusi<strong>on</strong>; Under acute immobilizati<strong>on</strong> stress, the rat submandibular gland has been shown to increase<br />

producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> BDNF, thereby c<strong>on</strong>tributing to the elevati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> plasma BDNF levels. We believe that salivary<br />

gland can be influenced the health <str<strong>on</strong>g>of</str<strong>on</strong>g> distant organs. It is necessary to determine the target organs <str<strong>on</strong>g>of</str<strong>on</strong>g> plasma<br />

BDNF in this stress model.<br />

5J_06_P<br />

(poster secti<strong>on</strong> B1, poster board #250, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DEVELOPMENTAL EXPRESSION OF HEAT SHOCK PROTEIN 70 IN RAT KIDNEY<br />

Chul-Ho Lee 1 , Ji-Hyun S<strong>on</strong>g 2 , Y<strong>on</strong>g-Hwan Kim 2 , Si-Yun Ryu 2 , Jin Kim 3 , Ju-Young Jung 2<br />

1Genetic Resources Center, Korea <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Bioscience and Biotechnology (KRIBB), Daeje<strong>on</strong>, Korea;<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Veterinary Medicine & Institutes <str<strong>on</strong>g>of</str<strong>on</strong>g> Veterinary Science, Chungnam Nati<strong>on</strong>al University, Daeje<strong>on</strong><br />

305764, Korea; 3Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Anatomy, College <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, The Catholic University <str<strong>on</strong>g>of</str<strong>on</strong>g> Korea, Seoul 137701,<br />

Korea<br />

Heat shock protein 70 (HSP70) is a major molecular chaper<strong>on</strong>e and plays an important role in protecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

cells in renal medulla against high osmolality. As the urinary c<strong>on</strong>centrating mechanism is being built up after<br />

birth, urine osmolality increases gradually <str<strong>on</strong>g>for</str<strong>on</strong>g>m 300 mOsm/kg to 2,000 mOsm/kgH 2 O; accordingly, cells in<br />

renal medulla are exposed to an elevated and variable interstitial osmolality. To understand the possible roles<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 in the developing kidney, expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 and the changes <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 expressi<strong>on</strong> in the loop<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> Henle following infusi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> furosemide were investigated. Immunohistochemistry analysis revealed that<br />

little immunoreactivity <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 was detected in embry<strong>on</strong>ic kidneys. On postnatal day 1, HSP70 was first<br />

detected in the inner medullary collecting duct (IMCD) <str<strong>on</strong>g>of</str<strong>on</strong>g> the papillary tip. The intensity <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70<br />

immunostaining in the IMCD steeply increases ascending to the border between outer and inner medulla<br />

during the first three week after birth. At birth, all loop <str<strong>on</strong>g>of</str<strong>on</strong>g> Henle in rat renal papilla have the c<strong>on</strong>figurati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

short loop and there is no ascending thin limb. During the first two week <str<strong>on</strong>g>of</str<strong>on</strong>g> life the cuboidal epithelium <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the thick ascending limb is gradually trans<str<strong>on</strong>g>for</str<strong>on</strong>g>med into the ascending thin limb by a process that start just<br />

be<str<strong>on</strong>g>for</str<strong>on</strong>g>e the bend <str<strong>on</strong>g>of</str<strong>on</strong>g> the loop and proceeds toward the outer medulla. From 4 to 14 days <str<strong>on</strong>g>of</str<strong>on</strong>g> age, HSP70 was<br />

detected in the cells after trans<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> in the ascending thin limb, beginning at the papillary tip and<br />

ascending to the border between the outer medulla and the inner medulla. Since then expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70<br />

gradually increased, and by 21 days after birth, HSP70 was detected in IMCD and ATL in most part <str<strong>on</strong>g>of</str<strong>on</strong>g> inner<br />

medulla. In adult rat kidney, expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 was highest in IMCD. The gradual increase in HSP70 is<br />

associated with an increase in its mRNA abundance. However, furosemide infusi<strong>on</strong> resulted the significantly<br />

decrease HSP70 expressi<strong>on</strong> in the renal papilla. These data dem<strong>on</strong>strated that expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 was<br />

closely correlated with the changes in interstitial osmolality during the development <str<strong>on</strong>g>of</str<strong>on</strong>g> kidney. We suggest<br />

that HSP70 protects tubular epithelial cells in inner medulla from the stress <str<strong>on</strong>g>of</str<strong>on</strong>g> high osmolality during the<br />

development <str<strong>on</strong>g>of</str<strong>on</strong>g> kidney.<br />

5J_07_P<br />

301


23-26 August 2007,<br />

Budapest, Hungary<br />

302<br />

(poster secti<strong>on</strong> B1, poster board #251, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PHYSIOLOGICAL AND MOLECULAR RESPONSES TO HEAT STRESS DURING<br />

AVIAN ONTOGENY: HOW DO THEY REFLECT EVOLUTIONARY CHANGES IN<br />

THERMAL RESISTANCE<br />

Ariel Shabtay 1 , Zeev Arad 2<br />

1 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Cattle and Genetic Sciences, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Animal Science, ARO, Newe Ya’ar <str<strong>on</strong>g>Research</str<strong>on</strong>g> Center,<br />

P. O. Box 1021, Ramat Yishay 30095, Israel<br />

2 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, Techni<strong>on</strong> – Israel Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Technology, Haifa 32000, Israel<br />

Ectothermy and endothermy c<strong>on</strong>stitute the major thermoregulatory strategies in living organisms. While<br />

ectotherms can be pr<strong>on</strong>e to changes in body temperature (T b ) in correlati<strong>on</strong> with ambient temperature (T a )<br />

variati<strong>on</strong>s, the internal physiological milieu <str<strong>on</strong>g>of</str<strong>on</strong>g> endotherms remains relatively stable despite external thermal<br />

fluctuati<strong>on</strong>s. The chicken is ideal <str<strong>on</strong>g>for</str<strong>on</strong>g> comparing ectothermic and endothermic ‘thermal states’, as during<br />

<strong>on</strong>togeny its embryo undergoes a transiti<strong>on</strong> from ectothermy to endothermy. By using avian development as<br />

a model system <str<strong>on</strong>g>for</str<strong>on</strong>g> transiti<strong>on</strong> from ectothermy to endothermy, we show that, in c<strong>on</strong>trast to the ectothermic<br />

state, in the endothermic state the organism is more resistant to heat but relies less <strong>on</strong> HSPs as a first-line<br />

thermoprotective mechanism. Moreover, intraspecific, real-time, in vivo measurements in genetically diverse<br />

fowl strains relate improvement <str<strong>on</strong>g>of</str<strong>on</strong>g> thermoresistance in endotherms to improved T b regulati<strong>on</strong>, with a<br />

c<strong>on</strong>comitant delay in HSF activati<strong>on</strong> and HSPs expressi<strong>on</strong>. The time course <str<strong>on</strong>g>of</str<strong>on</strong>g> this delay and the T b at which<br />

it occurs imply that the <strong>on</strong>togenetic and evoluti<strong>on</strong>ary pathways leading to improved thermoresistance may<br />

have followed two, apparently n<strong>on</strong>-related, parallel routes: a cellular <strong>on</strong>e, in which the acquisiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

thermoresistance is not HSP-dependent and could result from altered mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> thermal sensati<strong>on</strong>, and<br />

a peripheral <strong>on</strong>e, characterized by altered homeostatic mechanisms that lead to differential patterns <str<strong>on</strong>g>of</str<strong>on</strong>g> T b<br />

regulati<strong>on</strong>.<br />

5J_08_P<br />

(poster secti<strong>on</strong> B1, poster board #252, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ELECTROSTRESS AS A RISK FACTOR OF THE RAT POSTNATAL<br />

NEUROGENESIS: IMMUNOHISTOCHEMICAL AND QUANTITATIVE STUDY<br />

J. Orendáčová 1 , M. Orendáč 2 , J. Labun 3 , E. Račeková 1 , K. Saganová 1 , M. Mojžiš 2 , H. Abdiová 4<br />

1Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Neurobiology, CE, SAS, Šoltésovej 4, Košice 040 01, Slovakia; e-mail: orendac@saske.sk<br />

2Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Electrical Engineering and In<str<strong>on</strong>g>for</str<strong>on</strong>g>matics, TU Košice; 3 Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Aer<strong>on</strong>autics, TU Košice; 4 Sec<strong>on</strong>dary<br />

School <str<strong>on</strong>g>for</str<strong>on</strong>g> Medical Assistants, Košice, Slovakia<br />

Findings <str<strong>on</strong>g>of</str<strong>on</strong>g> the last years put evidence about persisted neurogenesis during the whole postnatal age and not<br />

<strong>on</strong>ly during prenatal period. One <str<strong>on</strong>g>of</str<strong>on</strong>g> the most studied regi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> postnatal neurogenesis is the subventricular<br />

z<strong>on</strong>e, which harbours steadily dividing stem cells and progenitors, which migrate al<strong>on</strong>g a distinct pathway the<br />

rostral migratory stream (RMS), to populate the olfactory bulb and establish c<strong>on</strong>necti<strong>on</strong>s with their neur<strong>on</strong>al<br />

targets. This system represents a simple well-defined model allowing experimental observati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> postnatal<br />

neurogenesis. We used BrdU immunohistochemistry to study the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> EMR <strong>on</strong> postnatal neurogenesis<br />

within the RMS in the newborn, adult and aging rats. Quantitative analysis showed significant differences <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

postirradiati<strong>on</strong> changes in dependency to the rat age, the EMR durati<strong>on</strong> and postirradiati<strong>on</strong> survival <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

animals. The EMR <str<strong>on</strong>g>of</str<strong>on</strong>g> 2-6.7 mW/cm 2 induced significant changes <str<strong>on</strong>g>of</str<strong>on</strong>g> proliferating cells number within the<br />

RMS and disrupts typical double peeks <str<strong>on</strong>g>of</str<strong>on</strong>g> proliferating cells number, characteristic <str<strong>on</strong>g>for</str<strong>on</strong>g> the first postnatal


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

m<strong>on</strong>th. Three sets <str<strong>on</strong>g>of</str<strong>on</strong>g> the menti<strong>on</strong>ed EMR doses (5 hr/day) induced significant decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> proliferating cell<br />

number within the RMS <str<strong>on</strong>g>of</str<strong>on</strong>g> rats irradiated at P8 regardless postirradiati<strong>on</strong> survival period. On the other hand<br />

the same doses <str<strong>on</strong>g>of</str<strong>on</strong>g> the EMR applied in two sets induced increase <str<strong>on</strong>g>of</str<strong>on</strong>g> proliferating cells numbers <strong>on</strong>ly in the<br />

acute postirradiati<strong>on</strong> period. In the aging rats the same doses <str<strong>on</strong>g>of</str<strong>on</strong>g> EMR induced decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> proliferati<strong>on</strong>g cell<br />

number regardless postirradiati<strong>on</strong> survival period.<br />

5J_09_P<br />

(poster secti<strong>on</strong> B1, poster board #253, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

IMPULSE ACTIVITY OF SUPRAOPTIC CELLS OF HYPOTHALAMUS AFTER LONG-<br />

LASTING INFLUENCE OF LOW-INTENSITY EXTREMELY HIGH FREQUENCY<br />

RADIATION<br />

G. Yu. Grigoryan, S. M. Minassian, S. G. Sahakyan<br />

Yerevan State University, Alek Manoogyan Str. 1, 375025 Yerevan, Armenia<br />

The influence <str<strong>on</strong>g>of</str<strong>on</strong>g> envir<strong>on</strong>mental factors was sharply increased in existing ecological c<strong>on</strong>diti<strong>on</strong>. In particular,<br />

l<strong>on</strong>g-lasting influence <str<strong>on</strong>g>of</str<strong>on</strong>g> electromagnetic radiati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> different ranges results in a stress-related strained state<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the organism. Our investigati<strong>on</strong> was aimed at the revealing <str<strong>on</strong>g>of</str<strong>on</strong>g> effects <str<strong>on</strong>g>of</str<strong>on</strong>g> low-intensity extremely high<br />

frequency radiati<strong>on</strong> <strong>on</strong> the sp<strong>on</strong>taneous activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the rats’ supraoptic nucleus neur<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> hypothalamus<br />

within different periods <str<strong>on</strong>g>of</str<strong>on</strong>g> influence (up to 15 days). In acute experiments <strong>on</strong> nembutal-anesthetized rats, the<br />

extracellular recording and analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> impulse activity <str<strong>on</strong>g>of</str<strong>on</strong>g> supraoptic cells in norm and after exposure were<br />

carried out. The distributi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>s by the degree <str<strong>on</strong>g>of</str<strong>on</strong>g> regularity and dynamics <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>al current flows<br />

as well as the average frequency <str<strong>on</strong>g>of</str<strong>on</strong>g> impulsati<strong>on</strong> and the interspike intervals' coefficient <str<strong>on</strong>g>of</str<strong>on</strong>g> variati<strong>on</strong> were<br />

evaluated. The shifts <str<strong>on</strong>g>of</str<strong>on</strong>g> the impulse activity <str<strong>on</strong>g>of</str<strong>on</strong>g> supraoptic cells, c<strong>on</strong>cerning both <str<strong>on</strong>g>of</str<strong>on</strong>g> internal structure and <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

statistic parameters <str<strong>on</strong>g>of</str<strong>on</strong>g> the impulse streams, under the 15-day exposure were revealed. The presence <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

statistically significant maximal expressiveness <str<strong>on</strong>g>of</str<strong>on</strong>g> the investigated parameters shifts <strong>on</strong> the 10th day and the<br />

tendency <str<strong>on</strong>g>of</str<strong>on</strong>g> their similarity with the initial data <strong>on</strong> the 15th day <str<strong>on</strong>g>of</str<strong>on</strong>g> the electromagnetic factor influence was<br />

find out. We have observed increase <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>s’ regularity grade that is correlated with some decrease in<br />

interspike intervals' coefficient <str<strong>on</strong>g>of</str<strong>on</strong>g> variati<strong>on</strong>. The significant reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> quantity <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>s with group<br />

activity was revealed as well. Phase character <str<strong>on</strong>g>of</str<strong>on</strong>g> changes <str<strong>on</strong>g>of</str<strong>on</strong>g> supraoptic cells’ average frequency in dynamics <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

15-day radiati<strong>on</strong> was observed also. It allowed us to assume that the influence <str<strong>on</strong>g>of</str<strong>on</strong>g> electromagnetic radiati<strong>on</strong><br />

could lead to the modulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>s excitability.<br />

303


23-26 August 2007,<br />

Budapest, Hungary<br />

5J_10_P<br />

(poster secti<strong>on</strong> B1, poster board #254, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CLONING AND TISSUE DISTRIBUTION OF CHICKEN UROCORTIN 3<br />

B. Leen, V. M. Darras, B. De Groef<br />

Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> comparative endocrinology KULeuven, Naamsetraat 61 (B)3000 Leuven,<br />

e-mail: bert.leen@bio.kuleuven.be<br />

Urocortins are neuropeptides bel<strong>on</strong>ging to the corticotropin-releasing horm<strong>on</strong>e family. They are known to<br />

cause stress-coping resp<strong>on</strong>ses in the brain and they have peripheral effects <strong>on</strong> the cardiovascular and immune<br />

system. To come to a better general understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress resp<strong>on</strong>se in chicken it is important to identify<br />

and localize these UCN’s in order to know their role. We were able to cl<strong>on</strong>e the cDNA encoding <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

chicken UCN3 by RT-PCR based <strong>on</strong> sequence similarity with the Xenopus UCN3. After cl<strong>on</strong>ing the chicken<br />

UCN3 sequence we further investigated the tissue distributi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> UCN3 using RT-PCR. A more detailed<br />

localisati<strong>on</strong> study in the chicken brain was per<str<strong>on</strong>g>for</str<strong>on</strong>g>med by in situ hybridisati<strong>on</strong> and by immmunohistochemistry<br />

using a heterologous antibody. UCN3 shows a widespread expressi<strong>on</strong> throughout the chicken body including<br />

not <strong>on</strong>ly various brain areas, but also most <str<strong>on</strong>g>of</str<strong>on</strong>g> the peripheral tissues tested. A closer look at the distributi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

UCN3 mRNA in the chicken brain showed mainly expressi<strong>on</strong> in the outer layer <str<strong>on</strong>g>of</str<strong>on</strong>g> the telencephal<strong>on</strong>. We also<br />

observed mRNA expressi<strong>on</strong> in the Edinger-Westphal nucleus, which is known to be a source <str<strong>on</strong>g>of</str<strong>on</strong>g> urocortin in<br />

the rodent brain. At the protein level we saw a corresp<strong>on</strong>ding colouring <str<strong>on</strong>g>of</str<strong>on</strong>g> the telencephal<strong>on</strong> and we could<br />

also detect a signal in the area <str<strong>on</strong>g>of</str<strong>on</strong>g> the Edinger-Westphal nucleus and in the locus coeruleus, which is<br />

resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> physiological resp<strong>on</strong>ses to stress. The tissue distributi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> UCN3 together with its colocalizati<strong>on</strong><br />

with the corticotropin-releasing horm<strong>on</strong>e receptor 2 (CRHR2) shows that UCN3 is a possibly<br />

important factor in the stress resp<strong>on</strong>se in chicken.<br />

304


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

MODULE 6<br />

STRESS IN MEDICINE<br />

305


23-26 August 2007,<br />

Budapest, Hungary<br />

SYMPOSIA<br />

• 6A. Sick chaper<strong>on</strong>es and chaper<strong>on</strong>opathies (August 25th Saturday afterno<strong>on</strong>)<br />

(Organizer and chair: Alberto J. L. Macario)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> B2, Building B<br />

• 6B. Chaper<strong>on</strong>es and cancer (August 24th Friday afterno<strong>on</strong>)<br />

(Organizer and chair: Gabriele Multh<str<strong>on</strong>g>of</str<strong>on</strong>g>f, and Michael Sherman)<br />

Poster: August 24 th Friday, Poster secti<strong>on</strong> B1, Building B<br />

• 6C. Hsp90 inhibitors and their clinical applicati<strong>on</strong>s (August 26th Sunday morning)<br />

(Organizers and Speakers: Francis Burrows, Len Neckers, Neal Rosen, Paul Workman)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> B2, Building B<br />

• 6D. Stress, Hsps and autoimmunity (August 26th Sunday morning)<br />

(Organizer and chair: Willem van Eden)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> B2, Building B<br />

• 6E. Stress and the metabolic syndrome: hypertensi<strong>on</strong>, atherosclerosis, obesity and diabetes<br />

(August 24th Friday afterno<strong>on</strong>)<br />

(Organizers and chairs: Jan Erikss<strong>on</strong> and R. Paul Roberts<strong>on</strong>)<br />

Poster: August 24 th Friday, Poster secti<strong>on</strong> B1, Building B<br />

• 6F. Sepsis, endotoxins, stress and organ dysfuncti<strong>on</strong> (August 25th Saturday morning)<br />

(Organizer and chairs: Robert S. Mun<str<strong>on</strong>g>for</str<strong>on</strong>g>d and Jean-Marc Cavaill<strong>on</strong>)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> B2, Building B<br />

• 6G. Protein misfolding, neurodegenerati<strong>on</strong> and disease (August 26th Sunday morning)<br />

(Organizer and chair: William E. Balch)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> B2, Building B<br />

• 6H. Stress in the development <str<strong>on</strong>g>of</str<strong>on</strong>g> schizophrenia (August 26th Sunday morning)<br />

(Organizer and chair: Kunio Yui and Michio Suzuki)<br />

Poster: August 24 th Friday, Poster secti<strong>on</strong> B1, Building B<br />

• 6I. Modern molecular mechanisms and therapeutics <str<strong>on</strong>g>of</str<strong>on</strong>g> stress-related gastroinestinal<br />

disorders (August 25th Saturday afterno<strong>on</strong>)<br />

(Organizers and chairs: Sándor Szabó and Yvette Taché)<br />

Poster: August 25 th Saturday, Poster secti<strong>on</strong> B2, Building B<br />

POSTER SYMPOSIA<br />

• 6J. Stress, Hsps and inflammati<strong>on</strong><br />

August 24 th Friday, Poster secti<strong>on</strong> B1, Building B<br />

• 6K. Maternal, fetal and ne<strong>on</strong>atal stress<br />

August 24 th Friday, Poster secti<strong>on</strong> B1, Building B<br />

• 6L. Molecular mechanisms and c<strong>on</strong>sequences <str<strong>on</strong>g>of</str<strong>on</strong>g> substance abuse:<br />

smoking, alcoholism, drugs<br />

August 24 th Friday, Poster secti<strong>on</strong> B1, Building B<br />

• 6M. Other stress topics in medicine (Gender differences in pathophysiological stress, Hsps<br />

and wound healing, Stress and myocardial protecti<strong>on</strong>, Pre- and postoperative stress)<br />

Poster: August 24 th Friday, Poster secti<strong>on</strong> B1, Building B<br />

306


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

6A. SICK CHAPERONES AND CHAPERONOPATHIES<br />

(ALBERTO J. L. MACARIO)<br />

6A_01_S<br />

CHAPERONOPATHIES BY DEFECT, EXCESS, OR MISTAKE<br />

Alberto J. L. Macario, Everly C<strong>on</strong>way De Macario<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Maryland Biotechnology Institute, Center <str<strong>on</strong>g>of</str<strong>on</strong>g> Marine Biotechnology, Baltimore, Maryland 21202, USA<br />

The stress resp<strong>on</strong>se, stress proteins, heat-shock genes and proteins, molecular chaper<strong>on</strong>e genes and proteins,<br />

and a number <str<strong>on</strong>g>of</str<strong>on</strong>g> closely related molecules and cellular processes have been studied over the last few decades.<br />

A huge amount <str<strong>on</strong>g>of</str<strong>on</strong>g> in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> has accumulated that is scattered in printed and electr<strong>on</strong>ic literature and data<br />

bases. Most <str<strong>on</strong>g>of</str<strong>on</strong>g> this in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> c<strong>on</strong>stitutes the subject matter <str<strong>on</strong>g>of</str<strong>on</strong>g> the science <str<strong>on</strong>g>of</str<strong>on</strong>g> chaper<strong>on</strong>ology. More recently,<br />

the c<strong>on</strong>cept <str<strong>on</strong>g>of</str<strong>on</strong>g> chaper<strong>on</strong>e pathology, sick chaper<strong>on</strong>es, has evolved since various pathological c<strong>on</strong>diti<strong>on</strong>s have<br />

been identified in which defective chaper<strong>on</strong>es play an etiologic role. These c<strong>on</strong>diti<strong>on</strong>s are the<br />

chaper<strong>on</strong>opathies. Recent findings <strong>on</strong> chaper<strong>on</strong>opathies are briefly discussed in this article. Chaper<strong>on</strong>opathies<br />

occur at all ages; as a rule the genetic cases have an early clinical <strong>on</strong>set while the acquired chaper<strong>on</strong>opathies<br />

become manifest in the elderly and/or in associati<strong>on</strong> with other diseases. Other fields <str<strong>on</strong>g>of</str<strong>on</strong>g> chaper<strong>on</strong>ology,<br />

which will most likely be expanded in the near future, are the study <str<strong>on</strong>g>of</str<strong>on</strong>g> extracellular chaper<strong>on</strong>es, chaper<strong>on</strong>e<br />

networks, the therapeutic use <str<strong>on</strong>g>of</str<strong>on</strong>g> chaper<strong>on</strong>es (i.e, chaper<strong>on</strong>otherapy) to manage chaper<strong>on</strong>opathies and to<br />

improve cell per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance in the face <str<strong>on</strong>g>of</str<strong>on</strong>g> stress, the evaluati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> chaper<strong>on</strong>es as diagnostic markers and as<br />

prognostic indicators, and the development <str<strong>on</strong>g>of</str<strong>on</strong>g> anti-chaper<strong>on</strong>e agents to suppress chaper<strong>on</strong>e-gene expressi<strong>on</strong><br />

or inhibit chaper<strong>on</strong>e functi<strong>on</strong> when chaper<strong>on</strong>es c<strong>on</strong>tribute to disease rather than the opposite.<br />

6A_02_S<br />

MOLECULAR GENETICS AND BIOLOGY OF SMALL HEAT SHOCK PROTEINS<br />

CAUSING INHERITED PERIPHERAL NEUROPATHY<br />

Joy Irobi 1 , Ines Dierick 1 , Berlinda Vanloo 2 , Jan Gettemans 2 , Ludo Van Den Bosch 3 ,<br />

Wim Robberecht 3 , Jean-Pierre Timmermans 5 , Peter De J<strong>on</strong>ghe 1,4 , Vincent Timmerman 1<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular Genetics, VIB, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Antwerp, Antwerpen, Belgium<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Protein <str<strong>on</strong>g>Research</str<strong>on</strong>g>, VIB, Ghent University, Gent, Belgium<br />

Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Neurobiology, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Experimental Neurology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Leuven, Leuven, Belgium<br />

Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Neurology, University Hospital Antwerp, Antwerpen, Belgium<br />

Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Cell Biology & Histology, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Veterinary Sciences, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Antwerp,<br />

Antwerpen, Belgium<br />

Distal hereditary motor neuropathies (distal HMN) are characterized by a selective degenerati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the ax<strong>on</strong>s<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> motor neur<strong>on</strong>s, while the sensory neur<strong>on</strong>s are spared. The biological process <str<strong>on</strong>g>of</str<strong>on</strong>g> this selective degenerati<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> motor neur<strong>on</strong>s is still unknown. In 2 distal HMN type II pedigrees linked to chromosome 12q24.3, we<br />

identified the same mutati<strong>on</strong> (K141N) in the small heat shock 22kDa protein 8 (HSPB8). A sec<strong>on</strong>d mutati<strong>on</strong><br />

(K141E) was found in 2 smaller families. Co-immunoprecipitati<strong>on</strong> showed an increased binding <str<strong>on</strong>g>of</str<strong>on</strong>g> both<br />

HSPB8 mutants to the interacting partner heat shock protein27 (HSPB1). We previously reported a Russian<br />

family with autosomal dominant ax<strong>on</strong>al Charcot-Marie-Tooth disease (CMT) and assigned the locus<br />

(CMT2F) to chromosome 7q11-q21. This locus c<strong>on</strong>tained HSPB1 as <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the candidate genes. Mutati<strong>on</strong><br />

analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> HSPB1 revealed a (S135F) missense mutati<strong>on</strong> segregating in the CMT2F family. Screening <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

307


23-26 August 2007,<br />

Budapest, Hungary<br />

HSPB1 mutati<strong>on</strong>s in a large cohort <str<strong>on</strong>g>of</str<strong>on</strong>g> CMT2/distal HMN patients identified additi<strong>on</strong>al mutati<strong>on</strong>s.<br />

Expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> mutant HSPB8 in COS-1 and N2a cells promoted <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> intracellular aggregates and a<br />

reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>al cell survival. In vitro chaper<strong>on</strong>e activity assay showed a reducti<strong>on</strong> <strong>on</strong> the cytoprotective<br />

functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> mutant proteins. Early passages <str<strong>on</strong>g>of</str<strong>on</strong>g> the primary fibroblast cultures from the distal HMN patient’s<br />

skin biopsy showed the <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> aggregate/aggresome complex, which sequestered several molecules<br />

including mitoch<strong>on</strong>dria. Measurement <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>dria transmembrane potentials in these primary fibroblast<br />

cells evidenced that patients but not c<strong>on</strong>trols pers<strong>on</strong>s exhibited a depolarized mitoch<strong>on</strong>drial potential.<br />

308<br />

6A_04_S<br />

DEFECTIVE CHAPERONE NETWORKS<br />

Péter Csermely, Tamás Korcsmáros, István A. Kovács, Máté S. Szalay<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Chemistry, Semmelweis University, Puskin street 9., H-1088 Budapest, Hungary<br />

e-mail: csermely@puskin.sote.hu<br />

Molecular chaper<strong>on</strong>es are not <strong>on</strong>ly fascinating molecular machines, but have a number <str<strong>on</strong>g>of</str<strong>on</strong>g> functi<strong>on</strong>s, which<br />

can be understood <strong>on</strong>ly by c<strong>on</strong>sidering the emergent properties <str<strong>on</strong>g>of</str<strong>on</strong>g> protein-protein interacti<strong>on</strong>, signalling and<br />

organelle networks – and that <str<strong>on</strong>g>of</str<strong>on</strong>g> chaper<strong>on</strong>es as special c<strong>on</strong>stituents <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular networks. Moreover,<br />

chaper<strong>on</strong>es themselves are networks <str<strong>on</strong>g>of</str<strong>on</strong>g> amino acid side chains <str<strong>on</strong>g>of</str<strong>on</strong>g>fering vulnerable points <str<strong>on</strong>g>for</str<strong>on</strong>g> damage. Why<br />

are chaper<strong>on</strong>es special in the c<strong>on</strong>text <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular networks Chaper<strong>on</strong>es (1) have weak links, i.e. low affinity,<br />

transient interacti<strong>on</strong>s with most <str<strong>on</strong>g>of</str<strong>on</strong>g> their partners; (2) c<strong>on</strong>nect hubs, i.e. act as ‘masterminds’ <str<strong>on</strong>g>of</str<strong>on</strong>g> the cell being<br />

close to several centre proteins with a lot <str<strong>on</strong>g>of</str<strong>on</strong>g> neighbours; (3) are in the overlaps <str<strong>on</strong>g>of</str<strong>on</strong>g> network modules, which<br />

c<strong>on</strong>fers them a special regulatory role. Chaper<strong>on</strong>es may be inhibited by (1) mutati<strong>on</strong>s; (2) their damage e.g.<br />

after oxidati<strong>on</strong>; (3) their overload, i.e. a growth in the need <str<strong>on</strong>g>of</str<strong>on</strong>g> chaper<strong>on</strong>es and/or a decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> their<br />

availability or (4) by pharmacological inhibitors. Inhibitory modes (1) through (3) may occur in various<br />

diseases and during the aging process. Chaper<strong>on</strong>e inhibitors are efficient multi-target drugs in several diseases<br />

such as cancer. Defective chaper<strong>on</strong>es may uncouple or quarantine modules <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular networks, which<br />

increase protecti<strong>on</strong> and efficiency <str<strong>on</strong>g>of</str<strong>on</strong>g> the cell during stress. Moreover, after stress chaper<strong>on</strong>es are essential to<br />

re-build inter-modular c<strong>on</strong>tacts by their low affinity, ‘quasi-random’ sampling <str<strong>on</strong>g>of</str<strong>on</strong>g> the potential interacti<strong>on</strong><br />

partners in different cellular modules. This opens the way to the chaper<strong>on</strong>e-regulated disassembly, reassembly,<br />

adaptati<strong>on</strong> and modular evoluti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular networks, and helps us to design novel therapeutic<br />

and anti-aging strategies.<br />

6A_05_S<br />

PROTEIN AGGREGATION MECHANISMS: NEW LESSONS FROM HUMAN<br />

ALPHAB-CRYSTALLIN CHAPERONAPATHY IN MICE<br />

Ivor J. Benjamin<br />

Salt Lake City, USA<br />

Protein aggregati<strong>on</strong> skeletal and cardiac diseases are caused by mutati<strong>on</strong>s in αB-crystallin (CryAB, HSPB5) or<br />

desmin and exhibit characteristic hallmarks <str<strong>on</strong>g>of</str<strong>on</strong>g> protein misfolding and large cytoplasmic aggregates. The<br />

mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> protein misfolding diseases remain poorly understood but defining their pathogenesis might<br />

uncover new pathways as potential targets <str<strong>on</strong>g>for</str<strong>on</strong>g> therapeutic interventi<strong>on</strong>s. Our recent genetic studies in mice<br />

have shown that selective hR120GCryAB expressi<strong>on</strong> in the heart induces a novel toxic gain-<str<strong>on</strong>g>of</str<strong>on</strong>g>-functi<strong>on</strong>


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

mechanism mimicking reductive stress. Reductive stress refers to an abnormal increase in the amounts <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

reducing equivalents (e.g., glutathi<strong>on</strong>e, NADPH), which has been dem<strong>on</strong>strated in lower eukaryotes but is<br />

uncomm<strong>on</strong> in the mammals and/or in disease states. This work has broad implicati<strong>on</strong>s <str<strong>on</strong>g>for</str<strong>on</strong>g> the treatment <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

chaper<strong>on</strong>opathies and neurodegenerative disorders in which small MW Hsps participate as key players and<br />

not innocent bystanders in disease pathogenesis involving novel mechanisms.<br />

309


23-26 August 2007,<br />

Budapest, Hungary<br />

310<br />

6A_01_P<br />

(poster secti<strong>on</strong> B2, poster board #259, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

GENETIC VARIANT IN THE HSPB1 PROMOTER REGION IMPAIRS THE HSP27<br />

STRESS RESPONSE<br />

Ines Dierick 1,4,5 , Joy Irobi 1,4,5 , Sophie Janssens 6 , Jessie Theuns 2,4,5 , Ludo Van Den Bosch 7 ,<br />

Wim Robberecht 7 , Peter De J<strong>on</strong>ghe 3,4,5,8 , Christine Van Broeckhoven 2,4,5 , Vincent Timmerman 1,4,5<br />

1Peripheral Neuropathy Group, 2 Neurodegenerative Brain Diseases Group and 3 Neurogenetics Group, Department <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Molecular Genetics, VIB; 4 Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Neurogenetics, Institute Born-Bunge; 5 University <str<strong>on</strong>g>of</str<strong>on</strong>g> Antwerp; Antwerpen.<br />

6Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular Biomedical <str<strong>on</strong>g>Research</str<strong>on</strong>g>, VIB, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Ghent, Gent. 7 Neurobiology, Experimental<br />

Neurology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Leuven, Leuven. 8 Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Neurology, University Hospital <str<strong>on</strong>g>of</str<strong>on</strong>g> Antwerp,<br />

Antwerpen, Belgium<br />

The 27kDa heat shock protein 1 (HSP27) is a member <str<strong>on</strong>g>of</str<strong>on</strong>g> the ubiquitously expressed small heat shock protein<br />

family and has pleiotropic cytoprotective functi<strong>on</strong>s. Since HSP27 may act as a motor neur<strong>on</strong> survival factor,<br />

we analyzed the genetic c<strong>on</strong>tributi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the human HSPB1 gene (HSPB1) to the etiology <str<strong>on</strong>g>of</str<strong>on</strong>g> amyotrophic<br />

lateral sclerosis (ALS). In a cohort <str<strong>on</strong>g>of</str<strong>on</strong>g> sporadic ALS patients, we identified three rare genetic variati<strong>on</strong>s and<br />

<strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> which (c.-217T>C) targeted a c<strong>on</strong>served nucleotide <str<strong>on</strong>g>of</str<strong>on</strong>g> the Heat Shock Element (HSE) in the HSPB1<br />

promoter. Since binding <str<strong>on</strong>g>of</str<strong>on</strong>g> Heat Shock Factor 1 (HSF1) to this HSE is essential <str<strong>on</strong>g>for</str<strong>on</strong>g> stress-induced<br />

transcripti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSPB1, we examined the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> the c.-217C allele <strong>on</strong> transcripti<strong>on</strong>al activity and HSF<br />

binding. The promoter activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the HSPB1 c.-217C mutant allele decreased significantly to that <str<strong>on</strong>g>of</str<strong>on</strong>g> the wildtype<br />

promoter under basal and heat shock c<strong>on</strong>diti<strong>on</strong>s in neur<strong>on</strong>al and n<strong>on</strong>-neur<strong>on</strong>al cells. This impaired<br />

transcripti<strong>on</strong>al activity was mirrored by a drastic reducti<strong>on</strong> in HSF binding. In c<strong>on</strong>clusi<strong>on</strong>, our study<br />

underscores the importance <str<strong>on</strong>g>of</str<strong>on</strong>g> the c.-217T nucleotide <str<strong>on</strong>g>for</str<strong>on</strong>g> HSF binding and heat inducibility <str<strong>on</strong>g>of</str<strong>on</strong>g> HSPB1. It is<br />

c<strong>on</strong>ceivable that a weakened heat-shock mediated protecti<strong>on</strong> mechanism could affect age-related motor<br />

neur<strong>on</strong> integrity and there<str<strong>on</strong>g>for</str<strong>on</strong>g>e may c<strong>on</strong>tribute to the pathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> motor neur<strong>on</strong> disease.<br />

6A_02_P<br />

(poster secti<strong>on</strong> B2, poster board #260, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ABNORMAL INTERACTION OF MUTANT HSP22 (HSPB8) WITH THE RNA<br />

HELICASE DDX20 (GEMIN3, DP103)<br />

Xiankui Sun 1 , Jean-M. F<strong>on</strong>taine 1 , Adam D. Hoppe 2 , Serena Carra 3 , Cheryl DeGuzman 1 ,<br />

Jody L. Martin 4 , Stephanie Sim<strong>on</strong> 5 , Patrick Vicart 5 , Michael J. Welsh 1 , Jacques Landry 3 ,<br />

Rainer Benndorf 1<br />

Departments <str<strong>on</strong>g>of</str<strong>on</strong>g> 1 Cell and Developmental Biology and 2 Microbiology and Immunology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Michigan Medical<br />

School, Ann Arbor, MI, USA; 3 <str<strong>on</strong>g>Centre</str<strong>on</strong>g> de recherche en cancérologie de l'Université Laval, L'Hôtel-Dieu de Québec,<br />

Québec, Canada; 4 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Cardiovascular Institute, Loyola University Chicago, Maywood, IL,<br />

USA; and 5 EA 300 Stress et pathologies du cytosquelette, Université Paris 7, France<br />

e-mail: rbenndo@umich.edu<br />

Eight mutati<strong>on</strong>s in the small heat shock proteins (sHSP) Hsp22 and Hsp27 have been associated with the<br />

motor neur<strong>on</strong> diseases (MND) distal hereditary motor neuropathy and Charcot-Marie-Tooth disease. Hsp22<br />

and Hsp27 interact with each other, suggesting that these two etiologic factors may act in the same pathway.<br />

In an ef<str<strong>on</strong>g>for</str<strong>on</strong>g>t to learn about the role <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp22 in MND, we screened a human cDNA library by the yeast two-


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

hybrid method <str<strong>on</strong>g>for</str<strong>on</strong>g> potential binding proteins. One identified protein was the RNA helicase Ddx20, a core<br />

comp<strong>on</strong>ent <str<strong>on</strong>g>of</str<strong>on</strong>g> the survival-<str<strong>on</strong>g>of</str<strong>on</strong>g>-motor neur<strong>on</strong> (SMN) complexes. This interacti<strong>on</strong> was verified by independent<br />

methods including FRET. Both mutant Hsp22 <str<strong>on</strong>g>for</str<strong>on</strong>g>ms showed abnormally increased binding to Ddx20.<br />

Interestingly, Ddx20 itself binds to the SMN protein, and mutati<strong>on</strong>s in the SMN1 gene cause spinal muscular<br />

atrophy, another MND. Thus, these protein interacti<strong>on</strong> data have linked the etiologic factors Hsp22, Hsp27,<br />

and SMN, and mutati<strong>on</strong>s in any <str<strong>on</strong>g>of</str<strong>on</strong>g> these genes cause the various <str<strong>on</strong>g>for</str<strong>on</strong>g>ms <str<strong>on</strong>g>of</str<strong>on</strong>g> MND. SMN complexes are<br />

involved in RNP processing. The mutant Hsp22/Ddx20 interacti<strong>on</strong> was sensitive to treatment with RNase<br />

suggesting involvement <str<strong>on</strong>g>of</str<strong>on</strong>g> RNA in this interacti<strong>on</strong> and a potential role <str<strong>on</strong>g>of</str<strong>on</strong>g> sHSPs in RNP processing.<br />

6A_03_P<br />

(poster secti<strong>on</strong> B2, poster board #261, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE EXTENDED CCT-HSP60 GENE FAMILY IN THE HUMAN GENOME<br />

Dennis L. Maeder 1 , Luciano Brocchieri 2 , Alberto J. L. Macario 1 , Everly C<strong>on</strong>way de Macario 1<br />

1Center <str<strong>on</strong>g>of</str<strong>on</strong>g> Marine Biotechnology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Maryland Biotechnology Institute, Baltimore, MD, USA<br />

2University <str<strong>on</strong>g>of</str<strong>on</strong>g> Florida, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular Genetics and Microbiology and UF Genetics Institute,<br />

Gainesville, FL, USA<br />

The chaper<strong>on</strong>ins, a subset <str<strong>on</strong>g>of</str<strong>on</strong>g> molecular chaper<strong>on</strong>es have been classified into groups I and II, and we are<br />

investigating their occurrence in the human genome. Here we report <strong>on</strong> human CCT, which bel<strong>on</strong>gs to group<br />

II. Extensive searches <str<strong>on</strong>g>of</str<strong>on</strong>g> databases and published literature were per<str<strong>on</strong>g>for</str<strong>on</strong>g>med using bioin<str<strong>on</strong>g>for</str<strong>on</strong>g>matics and<br />

complementary methods. The nine can<strong>on</strong>ical cct subunit genes (cct1-cct8, cct6A and B included) were<br />

characterized. In additi<strong>on</strong>, at least 15 cct-related pseudogenes and 3 cct-related protein-encoding genes were<br />

identified. All 9 cct genes are multiex<strong>on</strong>ic and several have more than <strong>on</strong>e mRNA variant with multiple<br />

protein is<str<strong>on</strong>g>of</str<strong>on</strong>g>orms. The proteins encoded by these genes with their mRNA variants so far identified have 339<br />

(cct7, is<str<strong>on</strong>g>of</str<strong>on</strong>g>orm “b”) to 556 amino acids (cct1, is<str<strong>on</strong>g>of</str<strong>on</strong>g>orm “a”), with the great majority having over 530 amino acids.<br />

Most <str<strong>on</strong>g>of</str<strong>on</strong>g> the cct genes have related protein-encoding genes and/or pseudogenes. Our analysis thus far revealed<br />

that <str<strong>on</strong>g>of</str<strong>on</strong>g> the 15 pseudogenes three are related to cct1, <strong>on</strong>e to cct3, three to cct4, two to cct5, three to cct6A, two to<br />

cct7, and <strong>on</strong>e to cct8. Several chromosomes do not harbor cct genes or pseudogenes but others have them in<br />

various numbers, e.g., chromosome 7 has <strong>on</strong>e cct subunit gene and six pseudogenes; chromosome 5 has <strong>on</strong>e<br />

cct subunit gene, and four pseudogenes; other chromosomes have between <strong>on</strong>e and three genes and/or<br />

pseudogenes. We investigate the evoluti<strong>on</strong>ary relati<strong>on</strong>ships <str<strong>on</strong>g>of</str<strong>on</strong>g> cct, hsp60, and related protein-encoding genes<br />

and pseudogenes, and the diversity <str<strong>on</strong>g>of</str<strong>on</strong>g> their proteins (origin, structure, distributi<strong>on</strong>, functi<strong>on</strong>, pathology).<br />

311


23-26 August 2007,<br />

Budapest, Hungary<br />

6B. CHAPERONES AND CANCER<br />

(GABRIELE MULTHOFF, MICHAEL SHERMAN)<br />

6B_01_S<br />

LEDGF IS AN HSP70-2 REGULATED GUARDIAN OF LYSOSOMAL STABILITY IN<br />

HUMAN CANCER<br />

Mads Daugaard, Thomas Kirkegaard-Sørensen, Mikkel Rohde, Marja Jäättelä<br />

Apoptosis Department and <str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g> Genotoxic Stress, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Cancer Biology, Danish Cancer Society,<br />

Strandboulevarden 49, DK-2100 Copenhagen, Denmark<br />

e-mail: mj@cancer.dk<br />

Heat shock protein 70-2 (Hsp70-2) is a chaper<strong>on</strong>e protein essential <str<strong>on</strong>g>for</str<strong>on</strong>g> the growth <str<strong>on</strong>g>of</str<strong>on</strong>g> spermatocytes and<br />

cancer cells. Here we show that Hsp70-2 depleti<strong>on</strong> triggers lysosomal membrane permeabilizati<strong>on</strong> and<br />

cathepsin-dependent cell death and identify lens epithelial derived growth factor (LEDGF) as an Hsp70-2-<br />

regulated guardian <str<strong>on</strong>g>of</str<strong>on</strong>g> lysosomal stability in human cancer. Knockdown <str<strong>on</strong>g>of</str<strong>on</strong>g> LEDGF in cancer cells induces<br />

destabilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> lysosomal membranes followed by caspase-independent and Bcl-2 resistant cell death.<br />

Accordingly, ectopic LEDGF stabilizes lysosomes and protects cancer cells against cytotoxicity induced by<br />

anti-cancer agents that trigger the lysosomal cell death pathway. Remarkably, ectopic LEDGF also increases<br />

the tumorigenic potential <str<strong>on</strong>g>of</str<strong>on</strong>g> human cancer cells in immunodeficient mice and LEDGF expressi<strong>on</strong> is<br />

increased in human breast and bladder carcinomas correlating with that <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70-2 in invasive bladder<br />

cancer. Taken together, these data reveal LEDGF as an <strong>on</strong>cogenic protein that c<strong>on</strong>trols a caspaseindependent<br />

lysosomal cell death pathway.<br />

6B_02_S<br />

αB-CRYSTALLIN: A NEW PLAYER IN CANCER<br />

Vincent Cryns<br />

Cell Death Regulati<strong>on</strong> Lab, Feinberg School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Northwestern University, Chicago, IL USA<br />

e-mail: v-cryns@northwestern.edu<br />

αB-crystallin, a member <str<strong>on</strong>g>of</str<strong>on</strong>g> the small heat shock protein family, is induced by cellular stress and functi<strong>on</strong>s to<br />

limit stress-induced damage by suppressing protein aggregati<strong>on</strong>. We have dem<strong>on</strong>strated that αB-crystallin<br />

inhibits apoptosis induced many stimuli, at least in part, by disrupting activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the cell death protease<br />

caspase-3. We have also shown that αB-crystallin is comm<strong>on</strong>ly expressed in poor prognosis basal-like breast<br />

cancer and likely c<strong>on</strong>tributes to the aggressive behavior <str<strong>on</strong>g>of</str<strong>on</strong>g> these tumors. This presentati<strong>on</strong> will focus <strong>on</strong> new<br />

insights into the mechanisms by which αB-crystallin inhibits apoptosis and promotes tumor progressi<strong>on</strong> and<br />

its integral role in the cellular stress resp<strong>on</strong>se.<br />

312


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

6B_03_S<br />

IMMUNOLOGICAL ROLE OF MEMBRANE-BOUND AND EXPORTED HEAT<br />

SHOCK PROTEIN 70 (HSP70)<br />

Gabriele Multh<str<strong>on</strong>g>of</str<strong>on</strong>g>f 1 , Mathias Gehrmann 1 , Jürgen Rad<strong>on</strong>s 2<br />

1Dpt. <str<strong>on</strong>g>of</str<strong>on</strong>g> Radiotherapy and Radio<strong>on</strong>cology, Technical University Munich, and GSF – Inst. <str<strong>on</strong>g>of</str<strong>on</strong>g> Pathology,<br />

Munich, Germany<br />

2Inst. <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Biochemistry and Molecular Biology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Greifswald, Germany<br />

Stress or heat shock proteins (HSPs) are remarkably c<strong>on</strong>served in all living organisms. Their synthesis is<br />

induced in resp<strong>on</strong>se to a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> physiological and envir<strong>on</strong>mental insults. In the cytosol HSPs play an<br />

essential role as molecular chaper<strong>on</strong>es by assisting the correct folding <str<strong>on</strong>g>of</str<strong>on</strong>g> nascent and stress-accumulated<br />

misfolded proteins, preventing protein aggregati<strong>on</strong>, transport <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins, and supporting antigen processing<br />

and presentati<strong>on</strong>. On the plasma membrane and in the extracellular milieu they act as danger signals <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

adaptive and innate immune system. Either they act as carriers <str<strong>on</strong>g>for</str<strong>on</strong>g> immunogenic peptides, induce cytokine<br />

release or provide recogniti<strong>on</strong> sites <str<strong>on</strong>g>for</str<strong>on</strong>g> activated natural killer (NK) cells. Here we will discuss the problem<br />

why Hsp70, the major stress-inducible, cytosolic member <str<strong>on</strong>g>of</str<strong>on</strong>g> the HSP70 family is selectively found in the<br />

plasma membrane <str<strong>on</strong>g>of</str<strong>on</strong>g> tumor cells but not <strong>on</strong> normal cells and elucidate the immunological c<strong>on</strong>sequences:<br />

6B_04_S<br />

HSP72 AND HSP27 REGULATE THE P53 PATHWAY AND SUPPRESS THE<br />

SENESCENCE PROGRAM IN CANCER CELLS<br />

J. Yaglom, V. L Gabai, C. O’Callaghan, M. Y. Sherman<br />

Bost<strong>on</strong> University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine<br />

Many tumors have high levels <str<strong>on</strong>g>of</str<strong>on</strong>g> the major heat shock proteins Hsp72 and/or Hsp27, which correlates with<br />

aggressiveness <str<strong>on</strong>g>of</str<strong>on</strong>g> tumors, resistance to chemotherapy and poor prognosis. Originally, it was suggested that<br />

these Hsps facilitate tumorigenesis because they can suppress apoptosis <str<strong>on</strong>g>of</str<strong>on</strong>g> cancer cells caused by activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

<strong>on</strong>cogenes, like myc, or by the adverse factors <str<strong>on</strong>g>of</str<strong>on</strong>g> tumor microenvir<strong>on</strong>ment. Here we dem<strong>on</strong>strate that both<br />

Hsp72 and Hsp27 can suppress the p53 pathway and prevent senescence, another major break <strong>on</strong> cancer<br />

development. For example, over-expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp27 inhibited activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the p53 pathway by<br />

doxorubicin, nutlin-3 or TGF-β, and suppressed development <str<strong>on</strong>g>of</str<strong>on</strong>g> senescence in resp<strong>on</strong>se to these stimuli. In<br />

c<strong>on</strong>trast, specific depleti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp27 or Hsp72 in a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> cancer cell lines led to inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hdm2,<br />

activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> p53 and inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> p21. As a result, about 30-40% <str<strong>on</strong>g>of</str<strong>on</strong>g> cell populati<strong>on</strong> became growth arrested<br />

and developed features <str<strong>on</strong>g>of</str<strong>on</strong>g> cell senescence, while the rest <str<strong>on</strong>g>of</str<strong>on</strong>g> the populati<strong>on</strong> also having activated p53<br />

c<strong>on</strong>tinued to divide slowly, but became sensitive to radiati<strong>on</strong>, doxorubicin and other drugs. These data<br />

indicate that high levels <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp72 and/or Hsp27 allow cancer cells to avoid activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> an intrinsic<br />

senescence program by suppressing p53. At least in certain cancer lines, the intrinsic senescence program was<br />

associated with activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> <strong>on</strong>cogenes. In fact, in MCF-7 cells, which have c<strong>on</strong>stitutively active mutant <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

PI3 kinase (PIK3CA <strong>on</strong>cogene), depleti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp72 or Hsp27 led to activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> p53 and senescence.<br />

However, inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> PIK3CA that down-regulates PIP3 prevented activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> p53 and development <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

senescence up<strong>on</strong> depleti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsps. Similar results were seen with down-regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> PIP3 in cell lines<br />

trans<str<strong>on</strong>g>for</str<strong>on</strong>g>med by PTEN <strong>on</strong>cogenic mutati<strong>on</strong>s. These data indicate that overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsps plays a crucial<br />

role in supporting divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cells at early stage <str<strong>on</strong>g>of</str<strong>on</strong>g> trans<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> up<strong>on</strong> activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> <strong>on</strong>cogenes that c<strong>on</strong>trol<br />

PIP3 levels.<br />

313


23-26 August 2007,<br />

Budapest, Hungary<br />

6B_05_S<br />

HSP70 INTERACTS WITH THE PLASMA MEMBRANE PHOSPHOLIPID<br />

PHOSPHATIDYL SERINE<br />

Mathias Gehrmann 1 , Daniela Schilling 1 , Claudia Steinem 2 , A. Graham Pockley 1,3 ,Gabriele Multh<str<strong>on</strong>g>of</str<strong>on</strong>g>f 1<br />

1Technische Universität München and GSF - Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Pathology, Munich, Germany<br />

2University <str<strong>on</strong>g>of</str<strong>on</strong>g> Göttingen, Göttingen, Germany, 3 University <str<strong>on</strong>g>of</str<strong>on</strong>g> Sheffield, Sheffield, UK<br />

We have previously dem<strong>on</strong>strated a tumour-specific, plasma membrane localisati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 and shown that<br />

membrane bound Hsp70 acts as a target recogniti<strong>on</strong> structure <str<strong>on</strong>g>for</str<strong>on</strong>g> NK cell-mediated cytolysis. The membrane<br />

associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 remains poorly understood and this study used artificial unilamellar liposomes having<br />

different phosphatidylserine (PS), phosphatidylcholine (PC) and phosphatidylglycerol (PG) compositi<strong>on</strong>s to<br />

investigate the interacti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 with plasma membrane phospholipids. For the assay, Hsp70 (5µg) was<br />

incubated with liposomes (10µl) <str<strong>on</strong>g>for</str<strong>on</strong>g> 30 min at room temperature, after which the mixture was centrifuged <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

2 h at 200,000g. The presence <str<strong>on</strong>g>of</str<strong>on</strong>g> protein in the pelleted fracti<strong>on</strong>, which is indicative <str<strong>on</strong>g>of</str<strong>on</strong>g> protein/lipid<br />

interacti<strong>on</strong>s, was determined by Western blot analysis. Hsp70, but not Hsp60 or Hsp90, bound to unilamellar<br />

PC/PS (8/2) liposomes under physiological c<strong>on</strong>diti<strong>on</strong>s (pH 7.4), even in the absence <str<strong>on</strong>g>of</str<strong>on</strong>g> CaCl 2 or MgCl 2 . The<br />

amount <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 in the pellet increased as the amount <str<strong>on</strong>g>of</str<strong>on</strong>g> PS in the liposomes increased (PC/PS, 9/1, 8/2,<br />

7/3, 2/8, 1/9). Hsp70 did not bind to PC/PG (8/2) liposomes, thereby c<strong>on</strong>firming that the interacti<strong>on</strong>s are<br />

not charge-related. Hsp70 bound to PS-c<strong>on</strong>taining liposomes was recognised by commercially available anti-<br />

Hsp70 antibodies, whereas membrane-bound Hsp70 is <strong>on</strong>ly detectable using the cmHsp70.1 antibody<br />

(multimmune GmbH, Munich). If PS is indeed a natural binding partner <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 in vivo, then this might<br />

have important physiological implicati<strong>on</strong>s. It might also be that the plasma membrane PS c<strong>on</strong>tent <str<strong>on</strong>g>of</str<strong>on</strong>g> tumour<br />

sublines exhibiting distinct membrane Hsp70 expressi<strong>on</strong> patterns differs.<br />

314<br />

6B_06_S<br />

(poster secti<strong>on</strong> B1, poster board #179, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EXTRACELLULAR MTHSP70 AND IMMUNE RESISTANCE<br />

Zvi Fishels<strong>on</strong>, Oren Moskovich, David Pilzer<br />

Tel Aviv University, Dept. Cell & Developmental Biology, Sackler School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Tel Aviv, Israel<br />

e-mail: lifish@post.tau.ac.il<br />

Mitoch<strong>on</strong>drial hsp70 is involved in mitoch<strong>on</strong>drial import/export <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins, protecti<strong>on</strong> from stress and<br />

apoptosis, senescence and cancer. The level <str<strong>on</strong>g>of</str<strong>on</strong>g> mtHsp70 is elevated in many tumor types. Our findings (Pilzer<br />

and Fishels<strong>on</strong>, Int. Immunol. 17: 1239, 2005) suggested a role <str<strong>on</strong>g>for</str<strong>on</strong>g> mtHsp70 in protecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cancer cells from<br />

immune death. Following complement attack, the complement membrane attack complexes (MAC) and<br />

mtHsp70 were both found <strong>on</strong> membrane vesicles released from the cell surface. Vesicle release and<br />

accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> extracellular mtHsp70 could be inhibited by exogenous applicati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> anti-mtHsp70<br />

antibodies. Silencing <str<strong>on</strong>g>of</str<strong>on</strong>g> mtHsp70 with siRNA reduced MAC eliminati<strong>on</strong> by vesiculati<strong>on</strong> and enhanced<br />

sensitivity to MAC-induced cell death. MKT-077 is a cati<strong>on</strong>ic rhodacyanine dye that is highly toxic selectively<br />

to cancer cells. MKT-077 binds to mtHsp70 and dissociates it from p53, <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> its client proteins. Our results<br />

dem<strong>on</strong>strated that MKT-077 similarly prevented binding <str<strong>on</strong>g>of</str<strong>on</strong>g> mtHsp70 to purified complement C9, the main<br />

MAC comp<strong>on</strong>ent. Treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> human erythroleukemia K562 cells with MKT-077 sensitized them to MACmediated<br />

lysis but not to lysis induced by another pore-<str<strong>on</strong>g>for</str<strong>on</strong>g>mer, streptolysin O. Pre-treatment with MKT-077<br />

also reduced the extent <str<strong>on</strong>g>of</str<strong>on</strong>g> MAC-mtHsp70 vesiculati<strong>on</strong> after sublytic complement attack and, thus, enhanced


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

the amount <str<strong>on</strong>g>of</str<strong>on</strong>g> MAC remaining <strong>on</strong> the cells. Experiments with human colorectal carcinoma HCT116 p53 +/+<br />

and p53 -/- cells excluded the involvement <str<strong>on</strong>g>of</str<strong>on</strong>g> p53 in the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> MKT-077 <strong>on</strong> complement-mediated lysis.<br />

Our results suggest that during an immune attack, mtHsp70 binds to the MAC at the plasma membrane plane<br />

and plays a role in its clearance from the cell surface by vesiculati<strong>on</strong> and thus, supports cell resistance to<br />

complement-mediated lysis.<br />

6B_07_S<br />

(poster secti<strong>on</strong> B1, poster board #180, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ANTAGONISM BETWEEN MAJOR STRESS PROTEIN HSP70 AND MYC<br />

ONCOPROTEINS IN THE EXECUTION OF APOPTOTIC PROGRAM<br />

Irina Guzhova 1 , Elena Afanasyeva 1 , Elena Komarova 1 , Lars-Gunnar Larss<strong>on</strong> 2 , Boris Margulis 1<br />

1Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Cytology RAS, St Petersburg, Russia<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Biology and Forest Genetics, Swedish University <str<strong>on</strong>g>of</str<strong>on</strong>g> Agricultural Sciences, Uppsala, Sweden<br />

e-mail: guzhova@mail.cytspb.rssi.ru<br />

Two proteins c<strong>on</strong>tribute to the targeting <str<strong>on</strong>g>of</str<strong>on</strong>g> a cancer cell populati<strong>on</strong> towards apoptosis or survival, Myc<br />

<strong>on</strong>cogene known mainly as proapoptotic factor and Hsp70 stress protein possessing the ubiquitous protective<br />

activity. Based <strong>on</strong> our data indicating that Hsp70 can have especially high protective activity in heat shocktreated<br />

U-937-v-Myc cells we suggested that the key functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the <str<strong>on</strong>g>for</str<strong>on</strong>g>mer is the abrogati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis<br />

specifically mediated by Myc. To check this we used two models, in <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> whose over-expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Myc<br />

<strong>on</strong>cogene was c<strong>on</strong>tinuous (v-Myc in U-937 cells) and in another Myc expressi<strong>on</strong> was c<strong>on</strong>trolled by estradiol<br />

(c-Myc in Rat1MycER cells). It was found that the stable expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 rendered U-937-v-Myc cells<br />

more resistant than the original U-937 cell cl<strong>on</strong>es when both having been induced to apoptosis with<br />

etoposide and camptothecin. Using anti-Hsp70 siRNA we also dem<strong>on</strong>strated that resistance <str<strong>on</strong>g>of</str<strong>on</strong>g> U-937-v-Myc<br />

cells to the cytotoxic effect <str<strong>on</strong>g>of</str<strong>on</strong>g> above anti-cancer drugs was fully related to Hsp70 level, whereas in U-937 cells<br />

such effect <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 depleti<strong>on</strong> was not observed. In order to prove the data we employed another model<br />

system, Rat1MycERcells, with inducible expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 and <str<strong>on</strong>g>of</str<strong>on</strong>g> c-Myc. It was found that apoptosis<br />

induced by camptothecin and sodium butyrate and mediated by active MycER was efficiently and dosedependently<br />

suppressed by Hsp70. Taken together, our results led us to c<strong>on</strong>clusi<strong>on</strong> that Hsp70 might act as a<br />

complement to Myc-driven <strong>on</strong>cogenesis.<br />

315


23-26 August 2007,<br />

Budapest, Hungary<br />

316<br />

6B_01_P<br />

(poster secti<strong>on</strong> B1, poster board #181, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SURFACE AND INTRACELLULAR HSP70 EXPRESSION IN MALIGNANT<br />

BLOOD DISEASES<br />

F. Le<strong>on</strong>i, N. C. Dempsey 1 , C. Hoyle 2 , C. Hunter-Lavin 1 , J. H. H. Williams 1<br />

1University <str<strong>on</strong>g>of</str<strong>on</strong>g> Chester, UK, 2 Glan Clwyd Hospital. Bodelwyddan,UK<br />

e-mail: n.dempsey@chester.ac.uk<br />

The expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 has been extensively studied in many cancer types, however there is limited<br />

literature <strong>on</strong> Hsp70 expressi<strong>on</strong> in Myelodysplasic Syndrome (MDS). MDS is a cl<strong>on</strong>al disorder <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

haematopoietic stem cells characterised by apoptosis <str<strong>on</strong>g>of</str<strong>on</strong>g> progenitor stem cells leading to dysplastic<br />

haematopoiesis and variable cytopenias. Certain subtypes are associated with a high risk <str<strong>on</strong>g>of</str<strong>on</strong>g> progressi<strong>on</strong> to<br />

Acute Myeloid Leukemia (AML). Chr<strong>on</strong>ic Lymphocytic Leukemia (CLL) is a haematological malignancy<br />

which may progress from a stable <str<strong>on</strong>g>for</str<strong>on</strong>g>m to acute disease requiring treatment. We compared blood samples<br />

from patients with MDS, AML, CLL and normal age matched c<strong>on</strong>trols by flow cytometry <str<strong>on</strong>g>for</str<strong>on</strong>g> intracellular<br />

and extracellular expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70. CD3 (T cells), CD14 (M<strong>on</strong>ocytes) and CD15 (Neutrophils) were used<br />

to detect differences in Hsp70 expressi<strong>on</strong> between the main leucocyte populati<strong>on</strong>s. CD34+ was used as a<br />

marker <str<strong>on</strong>g>for</str<strong>on</strong>g> AML blasts and CD5+/CD19+ as a marker <str<strong>on</strong>g>for</str<strong>on</strong>g> CLL cells. Preliminary data show different<br />

patterns <str<strong>on</strong>g>of</str<strong>on</strong>g> expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> surface and intracellular Hsp70 with different subtypes <str<strong>on</strong>g>of</str<strong>on</strong>g> MDS. In some patients it<br />

is possible to show two distinct sub-populati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> neutrophils <strong>on</strong>e expressing more surface Hsp70. The<br />

AML data show populati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> blast cells which are both intracellular and surface Hsp70+ and this<br />

expressi<strong>on</strong> is higher than in the normal cells. These preliminary data would suggest correlati<strong>on</strong> with disease<br />

severity. CLL samples show Hsp70 expressi<strong>on</strong> <strong>on</strong> CD5+/CD19+ cells. We are currently increasing sample<br />

numbers <str<strong>on</strong>g>for</str<strong>on</strong>g> each stages <str<strong>on</strong>g>of</str<strong>on</strong>g> the individual diseases. We will use these results to determine the involvement <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Hsp70 in the progressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the diseases and whether manipulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 in these cells would be<br />

beneficial.<br />

6B_02_P<br />

(poster secti<strong>on</strong> B1, poster board #182, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE STRESS-INDUCIBLE IMMUNOLOGICAL DANGER SIGNALS HSP70 AND<br />

MICA/B JOINTLY AUGMENT THE CYTOTOXIC ACTIVITY OF HUMAN NK CELLS<br />

AGAINST TUMOR CELLS<br />

Leslie Elsner 1 , Perris F. Flügge 1 , Vijayakumar Muppala 1 , Jingky Lozano 2 , Dörthe Malzahn 2 ,<br />

Heike Bickeböller 2 , Edgar Brunner 3 , Gabriele Multh<str<strong>on</strong>g>of</str<strong>on</strong>g>f 4 , Lutz Walter 5 , Ralf Dressel 1*<br />

1Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Cellular and Molecular Immunology, 2 Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Genetic Epidemiology, 3 Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Statistics,<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Göttingen, 37099 Göttingen, Germany<br />

4Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Radiotherapy and Radio<strong>on</strong>cology, Technical University Munich, 81675 Munich, Germany<br />

5Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Primate Genetics, German Primate Center, 37077 Göttingen, Germany<br />

e-mail: rdresse@gwdg.de<br />

The stress-inducible heat shock protein (HSP) 70 is known to functi<strong>on</strong> as an endogenous danger signal which<br />

can increase the immunogenicity <str<strong>on</strong>g>of</str<strong>on</strong>g> tumors and induce cytotoxic T lymphocyte resp<strong>on</strong>ses. We show here that<br />

HSP70 also activates human natural killer (NK) cells which recognize the stress-inducible MHC class I chain-


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

related (MIC) A and B molecules <strong>on</strong> tumor cells. We observed that the stress-inducible HSP70, in c<strong>on</strong>trast to<br />

the c<strong>on</strong>stitutively expressed HSC70, activated human peripheral blood m<strong>on</strong><strong>on</strong>uclear cells in vitro to kill<br />

MICA-transfected target cells. The HSP70-derived peptide TKD (Multh<str<strong>on</strong>g>of</str<strong>on</strong>g>f et al., Cell Stress Chaper<strong>on</strong> 6:337,<br />

2001) was able to replace the full-length HSP70 in these assays. Cell separati<strong>on</strong> experiments identified NK<br />

cells as the cytotoxic effector cells which were activated by HSP70 or TKD. When MICA/B expressi<strong>on</strong> was<br />

induced <strong>on</strong> human melanoma cells by pharmacological means and NK cells were activated by HSP70 or<br />

TKD, both treatments jointly improved the killing <str<strong>on</strong>g>of</str<strong>on</strong>g> the tumor cells. Thus, the synergistic activity <str<strong>on</strong>g>of</str<strong>on</strong>g> two<br />

stress-inducible immunological danger signals, HSP70 and MICA/B, leads to enhanced cytotoxicity <str<strong>on</strong>g>of</str<strong>on</strong>g> NK<br />

cells. This work was supported by grants from the Deutsche Forschungsgemeinschaft (DR 394/2, GRK 289,<br />

GRK 1034) and the European Uni<strong>on</strong> (MRTN-CT-2004-512253; TRANS-NET).<br />

6B_03_P<br />

(poster secti<strong>on</strong> B1, poster board #183, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

RESISTANCE TO HSP90-TARGETED THERAPY IS MEDIATED THROUGH THE<br />

HEAT SHOCK RESPONSE<br />

A. McCollum, C. Ten Eyck, B. Stensgard, D. T<str<strong>on</strong>g>of</str<strong>on</strong>g>t, C. Erlichman<br />

Mayo Clinic College <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Rochester MN 55905<br />

e-mail: erlichman.charles@mayo.edu<br />

HSP90 is critical in the proper folding, stabilizati<strong>on</strong>, and trafficking <str<strong>on</strong>g>of</str<strong>on</strong>g> many client proteins that have a major<br />

impact <strong>on</strong> proliferati<strong>on</strong> and antiapoptotic signaling necessary <str<strong>on</strong>g>for</str<strong>on</strong>g> the malignant process. HSP90-targeted<br />

therapy has been introduced into clinical testing with the expectati<strong>on</strong> that with degradati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> many client<br />

proteins, a significant antitumor resp<strong>on</strong>se would be observed. However, clinical trials to date have shown<br />

<strong>on</strong>ly limited and anecdotal activity <str<strong>on</strong>g>of</str<strong>on</strong>g> this multitargeted kinase approach, even though stress resp<strong>on</strong>se<br />

proteins such as Hsp27 and Hsp70 are induced in patients after treatment with Hsp90-directed agents. We<br />

have undertaken studies to delineate factors that may c<strong>on</strong>tribute to the apparent resistance that has been<br />

observed in patients. A549 cells have been selected <str<strong>on</strong>g>for</str<strong>on</strong>g> resistance to geldanamycin (GA) after exposure to<br />

increasing c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> GA in a stepwise fashi<strong>on</strong>. The cells are resistant to not <strong>on</strong>ly GA but also to the<br />

clinically relevant compounds 17-allylaminogeldanamycin (17AAG) and (dimethylaminoethylamino)-17-<br />

demethoxygeldanamycin (DMAG). These cells overexpress the ABC transporter p-glycoprotein (pgp) which<br />

has been implicated in GA resistance previously. Treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> these cells with verapamil or GF120918,<br />

inhibitors <str<strong>on</strong>g>of</str<strong>on</strong>g> pgp, does not reverse the resistance to GA. However, knockdown <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP27 or HSP70 with<br />

siRNA reverses the resistance <str<strong>on</strong>g>of</str<strong>on</strong>g> the cells resulting in an IC 50 similar to that <str<strong>on</strong>g>of</str<strong>on</strong>g> the parent cell line. These<br />

results suggest that selective targeting <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP90 al<strong>on</strong>e will have limited clinical efficacy. On the other hand, a<br />

combined approach that affects the stress resp<strong>on</strong>se by targeting other stress-related proteins in additi<strong>on</strong> to<br />

HSP90 may overcome this self-induced resistance.<br />

317


23-26 August 2007,<br />

Budapest, Hungary<br />

318<br />

6B_05_P<br />

(poster secti<strong>on</strong> B1, poster board #184, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE LEVEL HSP72 AND HSP27 EXPRESSION AND APOPTOSIS INDUCTION IN<br />

NEUROBLASTOMA CELLS AND NEURONS AFTER QUERCETIN TREATMENT<br />

Joanna Jakubowicz-Gil 1 , Wojciech Rzeski 2 , Barbara Zdzisinska 2 , Katarzyna Wejksza 2 ,<br />

Tomasz Piersiak 1 , Ant<strong>on</strong>i Gawr<strong>on</strong> 1<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Comparative Anatomy & Anthropology and 2 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Virology & Immunology,<br />

Maria Curie-Sklodowska University, Akademicka 19, 20-033 Lublin, Poland<br />

e-mail: asiajgil@o2.pl<br />

We studied the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> quercetin, natural flav<strong>on</strong>oid, <strong>on</strong> apoptosis and necrosis inducti<strong>on</strong> in neuroblastoma<br />

cell line and the culture <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>s. Quercetin induced apoptosis and necrosis in both studied cell lines,<br />

whereas neur<strong>on</strong>s were much more sensitive to cell death up<strong>on</strong> quercetin treatment. The number <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptotic,<br />

as well as necrotic cells was 3-6 times higher in neur<strong>on</strong>s than in neuroblastoma cells.<br />

We also investigated the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> quercetin <strong>on</strong> the level <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp72 and Hsp27 expressi<strong>on</strong> in neuroblastoma<br />

cells and neur<strong>on</strong>s. It is worth to note, that no Hsp72 expressi<strong>on</strong> was observed in neur<strong>on</strong>s. Quercetin<br />

appeared to be a good inhibitor <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp27 expressi<strong>on</strong> in neuroblastoma cells and in neur<strong>on</strong>s and Hsp72 in<br />

cancer cells. Inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp72 and Hsp27 expressi<strong>on</strong> by specific antisense olig<strong>on</strong>ucleotides in<br />

neuroblastoma cells made the cells more vulnerable <str<strong>on</strong>g>for</str<strong>on</strong>g> apoptosis inducti<strong>on</strong> after quercetin treatment. The<br />

number <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptotic cells was comparable with results observed in neur<strong>on</strong>s after incubati<strong>on</strong> with flav<strong>on</strong>oid.<br />

Quercetin changed the localizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp27 in cancer cells. Direct correlati<strong>on</strong> between drug c<strong>on</strong>centrati<strong>on</strong><br />

and gradual migrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> protein from cytoplasm to the nucleus was observed. Quercetin had no effect <strong>on</strong><br />

localizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp72 in neuroblastoma and Hsp27 in neur<strong>on</strong>s, where proteins were observed in nuclei.<br />

Our results indicate that sensitivity <str<strong>on</strong>g>of</str<strong>on</strong>g> neuroblastoma cells and neur<strong>on</strong>s to quercetin induced apoptosis<br />

depends <strong>on</strong> the level <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp72 and Hsp27 expressi<strong>on</strong>.<br />

6B_06_P<br />

(poster secti<strong>on</strong> B1, poster board #185, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CASPASES-7 IS SPECIFICALLY ACTIVATED BY TAXOL IN OVARIAN CARCINOMA<br />

CELLS<br />

Anna Luria, Marina Wolfs<strong>on</strong><br />

Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular Biology <str<strong>on</strong>g>of</str<strong>on</strong>g> the Cell, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Microbiology and Immunology,<br />

Ben Guri<strong>on</strong> University <str<strong>on</strong>g>of</str<strong>on</strong>g> the Negev, Beer Sheva 84105, Israel<br />

e-mail: wolfs<strong>on</strong>@bgumail.bgu.ac.il<br />

Despite years <str<strong>on</strong>g>of</str<strong>on</strong>g> study, ovarian cancer remains <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the leading causes <str<strong>on</strong>g>of</str<strong>on</strong>g> cancer death in women. Primary<br />

ovarian carcinomas exhibit mitoch<strong>on</strong>drial dysfuncti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis in ~70% cases. Taxol, the first-line<br />

ovarian therapy drug, induces apoptosis in SKOV3 cell line and primary human ovarian carcinoma cells. We<br />

reported previously that taxol-induced apoptosis in these cells does not include activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> caspase-9 and<br />

caspase-3 (Ofir et al, 2002.Cell Death Diff 9,636-642). Recently, we showed that taxol-induced apoptosis is<br />

associated with procaspase-7 processing in ovarian cancer cells. Molecular chaper<strong>on</strong>e Hsp90 could directly<br />

inhibit apoptosome <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> and caspase-9 and -3 activati<strong>on</strong>, and its elevated expressi<strong>on</strong> was found in<br />

ovarian cancers. Using the specific Hsp90 inhibitor Novobiocin in combinati<strong>on</strong> with Taxol <strong>on</strong> SKOV3 cells,<br />

we observed the processing <str<strong>on</strong>g>of</str<strong>on</strong>g> caspase-3, however, synergistic effect <str<strong>on</strong>g>of</str<strong>on</strong>g> drug combinati<strong>on</strong> <strong>on</strong> apoptosis rate


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

was insignificant. Primary ovarian carcinoma cells isolated from ascites <str<strong>on</strong>g>of</str<strong>on</strong>g> patients exhibited different<br />

sensitivity to taxol. Processing <str<strong>on</strong>g>of</str<strong>on</strong>g> caspase-7 was observed in “taxol-sensitive” primary cultures, whereas no<br />

processed caspase-7 has been found in “taxol-resistant” primary cultures. In accordance to the lack <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

caspase-7 cleavage, viability <str<strong>on</strong>g>of</str<strong>on</strong>g> "resistant" cells was not affected by taxol. However, processing <str<strong>on</strong>g>of</str<strong>on</strong>g> caspase-7<br />

and decreased cell viability were observed under Novobiocin treatment in “taxol-resistant” primary cultures.<br />

Altogether, our results dem<strong>on</strong>strate that an inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90 sensitizes ovarian cancer cells to drug<br />

treatment, and that the processing <str<strong>on</strong>g>of</str<strong>on</strong>g> caspase-7 could be a marker <str<strong>on</strong>g>for</str<strong>on</strong>g> in vitro testing <str<strong>on</strong>g>of</str<strong>on</strong>g> susceptibility to<br />

chemotherapy.<br />

6B_07_P<br />

(poster secti<strong>on</strong> B1, poster board #186, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PHARMACOLOGICAL INHIBITION OF STRESS-ACTIVATED P38 AND<br />

EXTRACELLULAR-REGULATED P44/42 PROTEIN KINASES STIMULATE P-<br />

GLYCOPROTEIN AND DECREASE APOPTOSIS IN DOXORUBICIN-EXPOSED<br />

HEP3B CELLS<br />

Irena Manov, Yulia Bashenko, Theodore C. Iancu<br />

Pediatric <str<strong>on</strong>g>Research</str<strong>on</strong>g> and Electr<strong>on</strong> Microscopy Unit, Ruth and Bruce Rappaport Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine,<br />

Techni<strong>on</strong>-Israel Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Technology, Haifa, Israel<br />

e-mail: irmanov@tx.techni<strong>on</strong>.ac.il<br />

Doxorubicin (DOX) is the anthracycline with the widest spectrum <str<strong>on</strong>g>of</str<strong>on</strong>g> antitumor activity, but its clinical<br />

effectiveness is limited by the development <str<strong>on</strong>g>of</str<strong>on</strong>g> multidrug resistance (MDR). The phenomen<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> MDR in<br />

cancer cells is mainly associated with overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> P-glycoprotein (P-gp) leading to increased efflux <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

anticancer drugs and down-regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis. The p38 and p44/42 signaling pathways were shown to<br />

play pro-apoptotic roles in DOX-induced apoptosis. The cross-talk between MAPK and P-gp pathways in<br />

cancer cells is still poorly understood. We aimed to investigate whether the MAPK pathways are involved in<br />

P-gp activati<strong>on</strong> and whether their modificati<strong>on</strong> can regulate P-gp and DOX-induced apoptosis in Hep3B<br />

cells. The p38 and p44/42 cascades in cells exposed or not to DOX (5 mM) were selectively inhibited by<br />

SB202190 (SB, 0.1-30 µM) and PD98059 (PD, 10 and 30 µM) respectively. Viability, cell cycle distributi<strong>on</strong>, P-<br />

gp activity and c<strong>on</strong>tent were studied. Both SB and PD str<strong>on</strong>gly stimulated P-gp functi<strong>on</strong> as was dem<strong>on</strong>strated<br />

by the Rhodamine-123 efflux assay. Western blot revealed the increase <str<strong>on</strong>g>of</str<strong>on</strong>g> P-gp expressi<strong>on</strong> in cells treated<br />

either p38 or p44/42 inhibitors. Combining SB or PD with DOX inhibited apoptosis and increased cell<br />

viability. In c<strong>on</strong>clusi<strong>on</strong>, the selective inhibitors SB202190 and PD98059 activate P-gp in Hep3B cells. MAPK<br />

pathways p38 and p44/42 can regulate P-gp transporter in cancer cells and modify the cellular resp<strong>on</strong>se to<br />

chemotherapy.<br />

319


23-26 August 2007,<br />

Budapest, Hungary<br />

320<br />

6B_08_P<br />

(poster secti<strong>on</strong> B1, poster board #187, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EXPRESSION OF HUMAN HTRA1, HTRA2 AND HTRA3 GENES IN OVARIAN AND<br />

ENDOMETRIAL CANCERS<br />

Joanna Narkiewicz 1 *, Dagmara Klasa-Mazurkiewcz 2 , Sylwia Lapinska-Szumczyk 2 , Janusz Emerich 2 ,<br />

Barbara Lipinska 1<br />

1University <str<strong>on</strong>g>of</str<strong>on</strong>g> Gdansk, Gdansk, Poland<br />

2Medical University <str<strong>on</strong>g>of</str<strong>on</strong>g> Gdansk, Gdansk, Poland<br />

e-mail: j<strong>on</strong>ark@biotech.univ.gda.pl<br />

The HtrA family <str<strong>on</strong>g>of</str<strong>on</strong>g> serine proteases takes part in cellular stress resp<strong>on</strong>se including heat shock, inflammati<strong>on</strong><br />

and cancer. Down-regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> human HtrA1 and HtrA3 genes has been reported in some cancers,<br />

suggesting tumor-suppressor role <str<strong>on</strong>g>for</str<strong>on</strong>g> both genes. An evidence exists showing that both HtrA1 and HtrA3<br />

regulate biological processes by modulating TGFβ signaling. HtrA2 is a unique HtrA family member playing<br />

proapoptotic functi<strong>on</strong>s. In the present study expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> human HtrA1, HtrA2, HtrA3 and TGF-β1 genes<br />

in ovarian and endometrial cancers was examined by semi-quantitative RT-PCR and Western blotting<br />

methods. Analyses were carried out <strong>on</strong> 98 ovarian and 123 endometrial tissue specimens, including tumors<br />

and healthy c<strong>on</strong>trols. Our results showed statistically significant decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> HtrA1 and HtrA3 expressi<strong>on</strong> in<br />

ovarian and endometrial tumors comparing to normal tissues. For the ovarian tumors, we have found<br />

decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the HtrA1 and HtrA3 genes in all tested tumor groups including the benign,<br />

borderline, malignant and Krukenberg tumors. In the case <str<strong>on</strong>g>of</str<strong>on</strong>g> HtrA3, the decrease was dramatic both at the<br />

mRNA and protein levels; we did not find detectable HtrA3 protein in about 38% <str<strong>on</strong>g>of</str<strong>on</strong>g> ovarian and 20% <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

endometrial tumors. Our results showed a slight decrease in HtrA2 expressi<strong>on</strong> in the examined tumor tissues.<br />

Moreover, our results showed significant negative correlati<strong>on</strong> between HtrA1 and HtrA3, and TGF-β1<br />

relative protein levels in endometrial tissues, suggesting inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> TGF-β1 signaling by HtrAs in<br />

endometrial cancer. Our results are in agreement with previous reports showing downregulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HtrA1<br />

and HtrA3 genes’ expressi<strong>on</strong> in cancers and show additi<strong>on</strong>al data <strong>on</strong> correlati<strong>on</strong> between tumor type and<br />

HtrA expressi<strong>on</strong>. This is the first report showing correlati<strong>on</strong> between TGF-β1 and HtrA proteins’ levels in<br />

endometrial cancer.<br />

6B_09_P<br />

(poster secti<strong>on</strong> B1, poster board #188, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HSF-1 IN BREAST CANCER GROWTH, MIGRATION AND METASTASIS<br />

M. Kouspou 1,2 , K. Waldeck 1,2 , J. Vieusseux 1 , J. Harris 1 , J. Ojaimi 1 , J. Price 1<br />

1 Cancer Biology and Metastasis Lab, M<strong>on</strong>ash University, Clayt<strong>on</strong><br />

2Melbourne University, Melbourne VIC, Australia<br />

e-mail: John.Price@med.m<strong>on</strong>ash.edu.au<br />

Breast cancer metastasis is the major cause <str<strong>on</strong>g>of</str<strong>on</strong>g> morbidity and mortality am<strong>on</strong>gst patients. To identify novel<br />

mediators <str<strong>on</strong>g>of</str<strong>on</strong>g> breast cancer metastasis we have identified cl<strong>on</strong>es <str<strong>on</strong>g>of</str<strong>on</strong>g> the human breast cancer cell line, MDA-<br />

MB-231, that have differential migratory and metastatic propensities. Gene array analysis identified an<br />

increased expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> several HSF-1 target genes in cl<strong>on</strong>es possessing a high migratory and metastatic<br />

ability. Gene expressi<strong>on</strong> data mining <str<strong>on</strong>g>of</str<strong>on</strong>g> cohorts <str<strong>on</strong>g>of</str<strong>on</strong>g> breast cancer patient specimens identified HSF-1 to be


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

positively correlated to increased metastatic burden and decreased survival. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, to investigate the role<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> HSF-1 in breast cancer we stably transfected MDA-MB-231 cells with wild-type HSF-1 (HSF-1 WT ) and a<br />

dominant negative mutant <str<strong>on</strong>g>for</str<strong>on</strong>g>m (HSF-1 DN ). Characterizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these cells revealed that inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF-1<br />

functi<strong>on</strong> rather than its expressi<strong>on</strong> per se substantially inhibited anchorage-independent growth. In additi<strong>on</strong><br />

to this growth effect, HSF-1 was also identified to be fundamental in the migratory resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> the cells<br />

towards a number <str<strong>on</strong>g>of</str<strong>on</strong>g> growth factors, including EGF and IGF-I. Moreover, use <str<strong>on</strong>g>of</str<strong>on</strong>g> known HSF-1 inhibitors,<br />

KNK437 and quercetin, resulted in a potent inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> migrati<strong>on</strong>. Examinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cell signalling pathways<br />

previously described as vital to the migratory behaviour <str<strong>on</strong>g>of</str<strong>on</strong>g> these cells identified PLCγ1 as being<br />

downregulated by HSF-1 inhibiti<strong>on</strong>, suggesting an important interplay between these molecules. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e,<br />

these findings describe <str<strong>on</strong>g>for</str<strong>on</strong>g> the first time a direct role <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF-1 in tumour cell migrati<strong>on</strong> and identify a novel<br />

role <str<strong>on</strong>g>for</str<strong>on</strong>g> HSF-1 in the metastatic cascade. As such, HSF-1 may represent a potential therapeutic target in<br />

breast cancer metastasis.<br />

6B_10_P<br />

(poster secti<strong>on</strong> B1, poster board #189, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

UPR ACTIVATION IN THE HUMAN GASTRIC ADENOCARCINOMA CELL LINE<br />

AGS ENHANCES RESISTANCE TO THE COMMONLY USED ANTI CANCER<br />

AGENTS CISPLATIN, DOXORUBICIN AND 5-FLUROURACIL<br />

P. Scriven, J. Ritchie, L. Wyld<br />

Academic Surgical Oncology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Sheffield, Royal Hallamshire Hospital, Sheffield, United Kingdom<br />

e-mail: p.scriven@sheffield.ac.uk<br />

Physiological stress such as glucose deprivati<strong>on</strong> adversely impacts <strong>on</strong> protein folding within the ER and via<br />

recruitment <str<strong>on</strong>g>of</str<strong>on</strong>g> chaper<strong>on</strong>e proteins such as GRP78 activates a signal cascade known as the Unfolded Protein<br />

Resp<strong>on</strong>se (UPR). This mechanism is cytoprotective if the stressor is mild or <str<strong>on</strong>g>of</str<strong>on</strong>g> short durati<strong>on</strong>, if stress is<br />

severe or prol<strong>on</strong>ged then the UPR can initiate apoptosis. The UPR is activated in many solid tumours and<br />

has been shown to affect chemosensitivity, specifically increasing resistance to topoisomerase II inhibitors<br />

such as doxorubicin whilst possibly enhancing susceptibility to DNA cross linking agents such as cisplatin.<br />

By depriving the AGS cell line <str<strong>on</strong>g>of</str<strong>on</strong>g> glucose our group was able to induce UPR activati<strong>on</strong> as dem<strong>on</strong>strated by a<br />

rise in GRP78 levels <strong>on</strong> Western Blotting after 24hours <str<strong>on</strong>g>of</str<strong>on</strong>g> culture in media c<strong>on</strong>taining 0.5mmol and 0.2mmol<br />

glucose (compared to 11mmol in standard culture). The sensitivity <str<strong>on</strong>g>of</str<strong>on</strong>g> UPR activated AGS was compared<br />

against a standard c<strong>on</strong>trol <str<strong>on</strong>g>for</str<strong>on</strong>g> predetermined IC50 doses <str<strong>on</strong>g>of</str<strong>on</strong>g> the comm<strong>on</strong>ly utilised anti cancer agents<br />

Cisplatin, Doxorubicin and 5-Flurouracil. Toxicity was determined by MTT assay relative to an untreated<br />

c<strong>on</strong>trol group <str<strong>on</strong>g>for</str<strong>on</strong>g> each glucose c<strong>on</strong>centrati<strong>on</strong>. Relative to standard 11mmol glucose c<strong>on</strong>diti<strong>on</strong>s AGS cells<br />

cultured in 0.5 and 0.2mmol showed a statistically significant increase in survival (p


23-26 August 2007,<br />

Budapest, Hungary<br />

322<br />

6B_11_P<br />

(poster secti<strong>on</strong> B1, poster board #190, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

REGULATION OF TUMOR GROWTH BY HSPBP1, AN HSP70 COCHAPERONE<br />

Vince Guerriero, Roger Miesfeld, Randy Burd, Shamarie Black, Deborah A. Raynes<br />

Depts. <str<strong>on</strong>g>of</str<strong>on</strong>g> Animal Sciences, Molecular and Cellular Biology, Biochemistry and Molecular Biophysics and Nutriti<strong>on</strong>al<br />

Sciences, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Ariz<strong>on</strong>a, Tucs<strong>on</strong>, AZ 85721<br />

e-mail: Guerrier@email.ariz<strong>on</strong>a.edu<br />

The Hsp70 family <str<strong>on</strong>g>of</str<strong>on</strong>g> heat stress proteins can be regulated by a group <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins collectively known as<br />

cochaper<strong>on</strong>es. HspBP1 is a cochaper<strong>on</strong>e with nucleotide exchange activity that binds to and regulates Hsp70.<br />

Hsp70 is elevated in numerous types <str<strong>on</strong>g>of</str<strong>on</strong>g> cancer cells and has anti-apoptotic activity. Data from this laboratory<br />

has shown that HspBP1 levels are also elevated in a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> cancer types and there<str<strong>on</strong>g>for</str<strong>on</strong>g>e the HspBP1 to<br />

Hsp70 ratio remained c<strong>on</strong>stant. HspBP1 and HspBP1 mutants were over expressed in a cancer cell line to<br />

determine if increasing the HspBP1 to Hsp70 ratio would alter cell growth and tumor <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>. Increased<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HspBP1 (approximately 3-4 fold) did not alter cell proliferati<strong>on</strong>, cell growth at low densities,<br />

cell cycle parameters, apoptosis, or cell migrati<strong>on</strong>. However, injecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these cells into SCID mice resulted<br />

in a lag period be<str<strong>on</strong>g>for</str<strong>on</strong>g>e tumor growth and smaller tumors compared to c<strong>on</strong>trols. These results suggest that<br />

HspBP1 can alter the tumor host interacti<strong>on</strong>. One possible alterati<strong>on</strong> is a suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> vascular endothelial<br />

growth factor (VEGF) secreti<strong>on</strong> by the tumor cells. Measurement <str<strong>on</strong>g>of</str<strong>on</strong>g> VEGF secreti<strong>on</strong> revealed that over<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HspBP1 did not alter secreti<strong>on</strong> rates. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, HspBP1 can alter tumor growth in mice but<br />

the mechanism is unknown. Understanding how HspBP1 alters tumor cell growth will provide new<br />

in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> that can be used to develop novel therapies <str<strong>on</strong>g>for</str<strong>on</strong>g> tumor treatment.<br />

6B_12_P<br />

(poster secti<strong>on</strong> B1, poster board #191, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PROTEIN KINASE Cα ACTIVATION BY LEAD ACETATE LINKS TO THE<br />

EGFR/SRC/RAS/RAF/ERK SIGNALING AND PREVENTS CYTOTOXICITY AND<br />

MUTAGENICITY IN HUMAN LUNG CANCER CELLS<br />

Chun-Yu Wang, Yun-Wei Lin, Jia-Ling Yang 1<br />

Molecular Carcinogenesis Laboratory, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Biotechnology & Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Life Sciences,<br />

Nati<strong>on</strong>al Tsing Hua University, Hsinchu 30013, Taiwan<br />

e-mail: jlyang@life.nthu.edu.tw<br />

Lead acetate [Pb(II)] exhibits weak genotoxicity in mammalian cells due in part to the enhanced nucleotide<br />

excisi<strong>on</strong> repair mediated through ERK activati<strong>on</strong>. However, the ERK upstream signaling linking to Pb(II)<br />

weak genotoxicity remains unknown. Here we report the essential signaling transducers <str<strong>on</strong>g>for</str<strong>on</strong>g> ERK activati<strong>on</strong><br />

that prevent Pb(II) cytotoxicity and mutagenicity in human n<strong>on</strong>-small cell lung adenocarcinoma CL3 cells.<br />

Pb(II) stimulated the membrane localizati<strong>on</strong> associated PKCα activati<strong>on</strong>, the dissociati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Raf kinase<br />

inhibitory protein from Raf-1, and the phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Raf-1 <strong>on</strong> S338. The Pb(II)-induced phospho-Raf-<br />

1(S338) were markedly decreased by <str<strong>on</strong>g>for</str<strong>on</strong>g>ced expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a dominant-negative Ras. Pre-treatment with<br />

Gö6976 (a c<strong>on</strong>venti<strong>on</strong>al PKC inhibitor) or introducing PKCα small interfering RNA (siPKCα) blocked the<br />

Raf/ERK activati<strong>on</strong> and significantly increased the cytotoxicity and the hprt mutati<strong>on</strong> frequency in Pb(II)-<br />

exposed cells. PD153035 (an EGFR inhibitor) or SU6656 (a Src family inhibitor) but not AG1296 (a PDGFR


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

inhibitor) suppressed the Pb(II)-induced activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> tyrosine kinases and PKCα, and phospho-Raf-1(S338)<br />

levels. Pb(II) sequentially induced the autophosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> EGFR <strong>on</strong> Y1173 and Src <strong>on</strong> Y419. Thus, the<br />

PKCα participates in the EGFR/Src/Ras/Raf/ERK signaling cascade and avoids cell death and genomic<br />

alterati<strong>on</strong> during Pb(II) exposure.<br />

6B_13_P<br />

(poster secti<strong>on</strong> B1, poster board #192, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DHA INDUCES DISTURBANCE IN CHOLESTEROL HOMEOSTASIS, ER STRESS<br />

AND CELL CYCLE ARREST IN COLON CANCER CELLS<br />

G. L. Størvold, C. H. Jakobsen, H. Bremseth, S. A. Schønberg<br />

Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Laboratory Medicine, Children’s and Women’s Health,<br />

NTNU Erling Skjalgss<strong>on</strong>s gt 1, 7006 Tr<strong>on</strong>dheim, Norway<br />

e-mail: gro.l.storvold@ntnu.no<br />

Epidemiological, animal and cell culture studies indicate an inverse associati<strong>on</strong> between l<strong>on</strong>g chain omega-3<br />

fatty acids and growth <str<strong>on</strong>g>of</str<strong>on</strong>g> cancers such as breast, prostate and col<strong>on</strong>. We have previously shown that<br />

docosahexaenoic acid (DHA) induces growth arrest in different col<strong>on</strong> cancer cell lines, but little in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong><br />

regarding the molecular mechanisms exists. Microarray analysis has revealed that DHA induces cellular stress,<br />

and that activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the endoplasmic reticulum (ER) stress pathway may be <strong>on</strong>e mechanism leading to<br />

growth arrest. The main functi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> ER are regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> calcium homeostasis, protein folding and<br />

processing, and lipid synthesis. Disturbances in any <str<strong>on</strong>g>of</str<strong>on</strong>g> these functi<strong>on</strong>s may lead to ER stress. We have<br />

observed an accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cholesteryl esters in our cell lines after DHA treatment, but <strong>on</strong>ly slight increases<br />

in total cholesterol levels. In additi<strong>on</strong> we observe increased protein levels <str<strong>on</strong>g>of</str<strong>on</strong>g> HMG-CoA reductase, the rate<br />

limiting enzyme in cholesterol biosynthesis, and nSREBP2, the main regulator <str<strong>on</strong>g>of</str<strong>on</strong>g> genes involved in<br />

cholesterol synthesis and uptake. This indicates a cellular demand <str<strong>on</strong>g>for</str<strong>on</strong>g> cholesterol. Previously we have<br />

observed that treatment with DHA leads to an accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cells in G2/M phase <str<strong>on</strong>g>of</str<strong>on</strong>g> the cell cycle.<br />

Progressi<strong>on</strong> through G2/M phase is cholesterol-dependent and cholesterol depleti<strong>on</strong> will lead to inactivati<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> CDK1, downregulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cyclin B1, and G2/M arrest. In additi<strong>on</strong> activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ER stress may lead to cell<br />

cycle arrest by depleti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cyclin D1. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this study is to investigate the mechanisms by which DHA<br />

affects cholesterol homeostasis, induces cell cycle arrest and ER stress, and the possible link between these<br />

events.<br />

6B_14_P<br />

(poster secti<strong>on</strong> B1, poster board #193, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECTS OF HSC70/HSP70 ON IMMUNOGENICITY OF MITOMYCIN C-TREATED<br />

LEWIS LUNG CARCINOMA CELLS AND ON THE NEUTROPHIL ROS<br />

PRODUCTION<br />

V. N. Maltseva, V. V. Vrublevskaya, O. S. Morenkov, V. G. Safr<strong>on</strong>ova<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Cell Biophysics, Russian Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences, Pushchino, Moscow regi<strong>on</strong>, 142290 Russia<br />

e-mail: mvn3@mail.ru<br />

Heat shock proteins (hsp) play an important role in activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> innate immune system, antigen presenting<br />

and are used in specific antitumor immunotherapy. It was shown that co-injecti<strong>on</strong> in mice apoptotic<br />

323


23-26 August 2007,<br />

Budapest, Hungary<br />

mitomycin C-treated Lewis lung carcinoma (Mit-C-treated LLC) cells with hsc70/hsp70 preparati<strong>on</strong>s derived<br />

from mouse brain resulted in inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> specific anti-tumor immunity in comparis<strong>on</strong> with mice injected<br />

with Mit-C-treated LLC cells al<strong>on</strong>e. The decelerati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> tumor growth after live LLC cells inoculati<strong>on</strong>, the<br />

suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> metastatic tumor progressi<strong>on</strong> and delay <str<strong>on</strong>g>of</str<strong>on</strong>g> animal death was observed. Additi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> exogenous<br />

hsc70/hsp70 preparati<strong>on</strong>s in c<strong>on</strong>centrati<strong>on</strong> 16 µ/ml to perit<strong>on</strong>eal evoked neutrophils increased sp<strong>on</strong>taneous<br />

and induced reactive oxygen species (ROS) producti<strong>on</strong> while low c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> hsc70/hsp70 (1 µ/ml)<br />

suppressed them. These findings show that hsc70/hsp70 can enhance the immunogenicity <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptotic<br />

tumor cells and may acts as activati<strong>on</strong> signal <str<strong>on</strong>g>for</str<strong>on</strong>g> the cells <str<strong>on</strong>g>of</str<strong>on</strong>g> innate immune system.<br />

6B_15_P<br />

(poster secti<strong>on</strong> B1, poster board #194, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HSP70-2 PROMOTES CANCER CELL SURVIVAL BY INHIBITING A LYSOSOMAL<br />

DEATH PATHWAY THAT DEPENDS ON THE CELLULAR ABLATION OF LENS<br />

EPITHELIUM DERIVED GROWTH FACTOR (LEDGF/P75)<br />

Mads Daugaard 1,* , Thomas Kirkegaard 1 , Marie Stampe Ostenfeld 2 , Mads Aaboe 2 ,<br />

Torben F. Ørnt<str<strong>on</strong>g>of</str<strong>on</strong>g>t 2 , Mikkel Rohde 1 , Marja Jäättelä 1<br />

1Apoptosis Laboratory, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Cancer Biology, The Danish Cancer Society, Strandboulevarden 49, 2100<br />

Copenhagen East, DK-Denmark<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Clinical Biochemistry, Aarhus University Hospital, Skejby, DK-8200 Aarhus N, Denmark<br />

e-mail: mdj@cancer.dk<br />

Heat Shock Protein 70-2 (Hsp70-2) is a c<strong>on</strong>stitutively expressed member <str<strong>on</strong>g>of</str<strong>on</strong>g> the human Hsp70 chaper<strong>on</strong>e<br />

family and required <str<strong>on</strong>g>for</str<strong>on</strong>g> growth and survival <str<strong>on</strong>g>of</str<strong>on</strong>g> spermatocytes and cancer cells. Target specific knockdown <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Hsp70-2 drives cancer cells into a G 1 -phase cell cycle arrest that is mediated by the p53 tumor suppressor and<br />

the TGF-β family-member Macrophage inhibitor cytokine 1 (MIC-1). Here we show that cancer cells<br />

depleted <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70-2 also engage a delayed cell death program that is independent <str<strong>on</strong>g>of</str<strong>on</strong>g> MIC-1 but relies <strong>on</strong> the<br />

cellular ablati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Lens epithelium derived growth factor (LEDGF). Direct knockdown <str<strong>on</strong>g>of</str<strong>on</strong>g> LEDGF induces<br />

cancer cell specific destabilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> lysosomal membranes followed by caspase-independent and Bcl-2<br />

resistant cell death. Accordingly, over-expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the protein protects cancer cell lines against osmotic<br />

stress-induced lysosomal rupture and cytotoxic agents known to compromise lysosome integrity. Cell lines<br />

over-expressing LEDGF produce large and fast growing tumors in immune-deficient mice and LEDGF is<br />

up-regulated in clinical samples <str<strong>on</strong>g>of</str<strong>on</strong>g> invasive bladder and breast cancer. These data identify LEDGF as an<br />

Hsp70-2-regulated cancer cell survival factor and a putative target <str<strong>on</strong>g>for</str<strong>on</strong>g> cancer therapy.<br />

324


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

6B_16_P<br />

(poster secti<strong>on</strong> B1, poster board #195, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

MOLECULAR AND FUNCTIONAL ALTERATIONS OF ORGANELLE CROSSTALK<br />

DURING CELLULAR STRESS AND TUMORIGENESIS<br />

G. Szabadkai 1,2* , K. Bianchi 1,2 , D. De Stefani 2 , M. Chami 1 , B. Oules 1 , R. Rizzuto 2 ,<br />

P. Paterlini-Brechot 1<br />

1INSERM U 807, Paris, France<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Experimental and Diagnostic Medicine, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Ferrara, Italy<br />

e-mail: gszabadkai@necker.fr<br />

We have recently shown that protein complexes at the ER-mitoch<strong>on</strong>dria interface, involving Ca 2+ channels<br />

(VDAC and IP 3 R) and molecular chaper<strong>on</strong>es (grp75), c<strong>on</strong>tribute to Ca 2+ signal transmissi<strong>on</strong> between the<br />

organelles, and that ER stress induces mitoch<strong>on</strong>dria-mediated cell death through ER-mitoch<strong>on</strong>drial crosstalk.<br />

At the initial phases <str<strong>on</strong>g>of</str<strong>on</strong>g> tumorigenesis, solid tumors were shown to undergo ER stress due to limited nutrient<br />

and oxygen availability. Still, trans<str<strong>on</strong>g>for</str<strong>on</strong>g>med cells survive, suggesting that cells gain resistance against the ER<br />

stress induced cell death. In our present work we challenged two issues <str<strong>on</strong>g>of</str<strong>on</strong>g> the process: (i) We showed that ER<br />

stress inducti<strong>on</strong> leads to depleti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the ER Ca 2+ store by inducing a truncated spliced variant <str<strong>on</strong>g>of</str<strong>on</strong>g> the SERCA<br />

pump (S1T) devoid <str<strong>on</strong>g>of</str<strong>on</strong>g> the ATPase domain, introducing Ca 2+ leak into the ER membrane. The elevati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the [Ca 2+ ] in the ER associated microdomains immobilized mitoch<strong>on</strong>dria at the ER surface. This<br />

morphological rearrangement amplifies ER-mitoch<strong>on</strong>drial Ca 2+ transfer, resulting in mitoch<strong>on</strong>drial Ca 2+<br />

overload and inducti<strong>on</strong> the mitoch<strong>on</strong>drial apoptotis. (ii) We applied a chemically induced hepatocellular<br />

carcinoma model to study the stress resp<strong>on</strong>se, ER-mitoch<strong>on</strong>drial interacti<strong>on</strong>s and followed up the relative<br />

expressi<strong>on</strong> changes and the reorganizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> grp75 interactome during different phases <str<strong>on</strong>g>of</str<strong>on</strong>g> tumorigenesis by<br />

proteomic approaches. The observed overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> grp75 in the early phase <str<strong>on</strong>g>of</str<strong>on</strong>g> tumor development<br />

underlie the role <str<strong>on</strong>g>of</str<strong>on</strong>g> grp75 in maintaining the communicati<strong>on</strong> between the ER and mitoch<strong>on</strong>dria, while<br />

changes in the interactome might reflect a protective effect against stress induced death.<br />

325


23-26 August 2007,<br />

Budapest, Hungary<br />

326<br />

6C. HSP90 INHIBITORS AND THEIR CLINICAL APPLICATIONS<br />

(FRANCIS BURROWS, LEN NECKERS, NEAL ROSEN, PAUL WORKMAN)<br />

6C_01_S<br />

DIVERGENT OUTCOMES OF HSP90 INHIBITION –<br />

PROBLEM OR OPPORTUNITY<br />

Francis Burrows<br />

Biogen IDEC, San Diego, USA<br />

Hsp90 is a ubiquitous molecular chaper<strong>on</strong>e implicated in the pathophysiology <str<strong>on</strong>g>of</str<strong>on</strong>g> a range <str<strong>on</strong>g>of</str<strong>on</strong>g> diseases and so is<br />

a target <str<strong>on</strong>g>of</str<strong>on</strong>g> high interest <str<strong>on</strong>g>for</str<strong>on</strong>g> therapeutic interventi<strong>on</strong>. Most drug targets eventually become resistant, e.g. by<br />

mutati<strong>on</strong>, overexpressi<strong>on</strong> or anergy, but Hsp90 is uncomm<strong>on</strong> in that it directly c<strong>on</strong>trols both cytotoxic and<br />

cytoprotective pathways through it’s key role in regulating mitogenic and survival signaling as well as<br />

antiapoptotic stress resp<strong>on</strong>ses. Recent work has indicated that the HSF-1-dependent heat shock resp<strong>on</strong>se<br />

(HSR) decreases the cytotoxic potency <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90 inhibitors in cancer cells in vitro by several mechanisms, but,<br />

since normal cells are thought to mount a more robust HSR than their trans<str<strong>on</strong>g>for</str<strong>on</strong>g>med counterparts, it is unclear<br />

what impact inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the HSR might have <strong>on</strong> the therapeutic index <str<strong>on</strong>g>of</str<strong>on</strong>g> these drugs in vivo. On the other<br />

hand, c<strong>on</strong>diti<strong>on</strong>s characterized by excessive cell death, such as neurodegenerative diseases, stroke and<br />

peripheral neuropathy, could potentially be treated by pharmacological inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the HSR in neur<strong>on</strong>s.<br />

Biogen IDEC has identified potent Hsp90 modulators that induce client protein degradati<strong>on</strong> and the HSR<br />

with equal potency, which are in clinical development <str<strong>on</strong>g>for</str<strong>on</strong>g> cancer. In additi<strong>on</strong>, we are also exploring the<br />

potential <str<strong>on</strong>g>of</str<strong>on</strong>g> compounds that selectively induce client protein degradati<strong>on</strong> or favour cytoprotective pathways<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> use in Alzheimer’s Disease and Huntingt<strong>on</strong>’s Disease in additi<strong>on</strong> to <strong>on</strong>cology.<br />

6C_02_S<br />

ACETYLATION AS A DYNAMIC REGULATOR OF HSP90 FUNCTION:<br />

IMPLICATIONS FOR FURTHER DEVELOPMENT OF PHARMACOLOGIC HSP90<br />

INHIBITORS<br />

L. Neckers<br />

Urologic Oncology Branch, Nati<strong>on</strong>al Cancer Institute, Bethesda, MD, USA<br />

Heat shock protein 90 (Hsp90) chaper<strong>on</strong>es a key subset <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular signaling proteins and is necessary <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

malignant trans<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>. However, many aspects <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90 regulati<strong>on</strong> remain unresolved. Hsp90 is subject<br />

to an array <str<strong>on</strong>g>of</str<strong>on</strong>g> post-translati<strong>on</strong>al modificati<strong>on</strong>s which affect its functi<strong>on</strong>, including acetylati<strong>on</strong>. In the last few<br />

years, several investigators have reported that hist<strong>on</strong>e deacetylase (HDAC) inhibitors and knock-down <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

HDAC6 induce Hsp90 acetylati<strong>on</strong> and inhibit its activity. While pharmacologic inhibiti<strong>on</strong> and RNA<br />

knockdown <str<strong>on</strong>g>of</str<strong>on</strong>g> HDACs have been very useful in identifying reversible acetylati<strong>on</strong> as a potential regulator <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Hsp90 activity, it is unclear how this process functi<strong>on</strong>s at the molecular level. Use <str<strong>on</strong>g>of</str<strong>on</strong>g> HDACi and/or HDAC<br />

knockdown techniques allow study <str<strong>on</strong>g>of</str<strong>on</strong>g> <strong>on</strong>ly the hyperacetylated (but not hypoacetylated) chaper<strong>on</strong>e, and the<br />

importance <str<strong>on</strong>g>of</str<strong>on</strong>g> the acetylati<strong>on</strong> state <str<strong>on</strong>g>of</str<strong>on</strong>g> individual residues <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90 in the c<strong>on</strong>text <str<strong>on</strong>g>of</str<strong>on</strong>g> hyperacetylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

proteome cannot be queried. Furthermore, c<strong>on</strong>tributory effects due to hist<strong>on</strong>e hyperacetylati<strong>on</strong> cannot be<br />

discounted. Direct determinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the functi<strong>on</strong>al c<strong>on</strong>sequences <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90 acetylati<strong>on</strong> has awaited mapping<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> specific sites. These sites are now being identified. K294 (Hsp90alpha) was the first amino acid in Hsp90<br />

to be <str<strong>on</strong>g>for</str<strong>on</strong>g>mally identified as an acetylati<strong>on</strong> site and this residue is highly c<strong>on</strong>served in eukaryotic Hsp90.


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

C<strong>on</strong>servative mutati<strong>on</strong>al analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> K294 revealed its acetylati<strong>on</strong> status to be a str<strong>on</strong>g determinant <str<strong>on</strong>g>of</str<strong>on</strong>g> client<br />

protein and co-chaper<strong>on</strong>e binding to Hsp90 in mammalian cells. Interestingly, although acetylati<strong>on</strong> status <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

K294 affects Hsp90 ATPase activity, ATP binding is not altered. In yeast, human Hsp90 mutants that cannot<br />

be acetylated at K294 have reduced ability to support viability, while an acetylati<strong>on</strong>-mimicking mutati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

K294 possesses the opposite property. Further, acetylati<strong>on</strong> status <str<strong>on</strong>g>of</str<strong>on</strong>g> K294 may determine Hsp90 sensitivity<br />

to N-terminal pharmacologic inhibitors. These data suggest that c<strong>on</strong>trolled acetylati<strong>on</strong>/deacetylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> K294<br />

plays an important role in regulating the Hsp90 chaper<strong>on</strong>e cycle.<br />

6C_04_S<br />

DRUGGING THE CANCER CHAPERONE: PRECLINICAL DISCOVERY AND<br />

CLINICAL DEVELOPMENT OF HSP90 INHIBITORS<br />

Paul Workman<br />

Cancer <str<strong>on</strong>g>Research</str<strong>on</strong>g> UK <str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g> Cancer Therapeutics, The Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Cancer <str<strong>on</strong>g>Research</str<strong>on</strong>g>, Haddow Laboratories,<br />

Sutt<strong>on</strong>, Surrey SM2 5NG<br />

We are in a very exciting era <str<strong>on</strong>g>of</str<strong>on</strong>g> cancer drug discovery in which effective mechanism-based therapies are being<br />

designed and developed that act <strong>on</strong> the <strong>on</strong>cogenic proteins and pathways that are hijacked by pathological<br />

genetic and epigenetic changes to bring about the initiati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cancer and subsequent malignant progressi<strong>on</strong>.<br />

Therapeutic selectivity <str<strong>on</strong>g>for</str<strong>on</strong>g> tumor versus normal cells is achieved by taking advantage <str<strong>on</strong>g>of</str<strong>on</strong>g> various<br />

‘dependencies’ that develop during the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cancers. Two major <str<strong>on</strong>g>for</str<strong>on</strong>g>ms <str<strong>on</strong>g>of</str<strong>on</strong>g> cancer dependency are<br />

related to the molecular chaper<strong>on</strong>e and heat shock protein 90 (Hsp90). Firstly, many mutant and<br />

overexpressed <strong>on</strong>coproteins that are involved in <strong>on</strong>cogene dependence or addicti<strong>on</strong> require Hsp90 <str<strong>on</strong>g>for</str<strong>on</strong>g> their<br />

stability and functi<strong>on</strong>. Sec<strong>on</strong>dly, cancer cells also require Hsp90 and other stress resp<strong>on</strong>se proteins to help<br />

protect them against the adverse envir<strong>on</strong>mental c<strong>on</strong>diti<strong>on</strong>s present in solid tumors. Moreover, since Hsp90<br />

inhibiti<strong>on</strong> causes combinatorial degradati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> many cancer-causing proteins, Hsp90 inhibitors are able to<br />

attack all <str<strong>on</strong>g>of</str<strong>on</strong>g> the hallmark phenotypic traits <str<strong>on</strong>g>of</str<strong>on</strong>g> cancer cells and should have a reduced liability to the<br />

development <str<strong>on</strong>g>of</str<strong>on</strong>g> resistance compared to more c<strong>on</strong>venti<strong>on</strong>al drugs. In this presentati<strong>on</strong>, I will provide an<br />

update <strong>on</strong> the preclinical discovery and clinical development <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90 inhibitors. In particular, I will<br />

describe what we have learned from the clinical experience with the first-in-class Hsp90 inhibitor 17-AAG.<br />

This is a derivative <str<strong>on</strong>g>of</str<strong>on</strong>g> the natural product geldanamycin which has been a pathfinder Hsp90 drug,<br />

dem<strong>on</strong>strating evidence <str<strong>on</strong>g>of</str<strong>on</strong>g> target inhibiti<strong>on</strong> and therapeutic activity in various cancers, including melanoma,<br />

breast, prostate and multiple myeloma. I will also describe our discovery <str<strong>on</strong>g>of</str<strong>on</strong>g> the new synthetic small molecule<br />

class <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90 inhibitors based <strong>on</strong> the resorcinylic pyrazole and isoxazole scaffold Hsp90 and discuss their<br />

optimizati<strong>on</strong> by structure-based design in collaborati<strong>on</strong> with Vernalis Limited. Finally, I will describe our<br />

recent work identifying genes and proteins that are involved in sensitivity to Hsp90 inhibitors and that have<br />

potential to act as biomarkers <str<strong>on</strong>g>for</str<strong>on</strong>g> clinical use.<br />

327


23-26 August 2007,<br />

Budapest, Hungary<br />

328<br />

6C_05_S<br />

(poster secti<strong>on</strong> B2, poster board #262, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ALCOHOL EXPOSURE REGULATES HSF-1 AND HEAT SHOCK PROTEINS 70 AND<br />

90 IN MURINE MACROPHAGES: IMPLICATION IN TNFα PRODUCTION<br />

P. Mandrekar, D. Catalano, V. Jeliazkova, G. Szabo<br />

Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Univ. Mass. Med. Ctr, Worcester, MA, USA<br />

e-mail: pranoti.mandrekar@umassmed.edu<br />

Alcohol use affects innate immune resp<strong>on</strong>ses, particularly TNFα, and results in alcoholic liver disease. Heat<br />

shock proteins, mediators <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress resp<strong>on</strong>ses influence TNFα producti<strong>on</strong>. Here we hypothesized that<br />

alcohol exposure regulates TNFα producti<strong>on</strong> and NFκB activati<strong>on</strong> via modulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the heat shock<br />

transcripti<strong>on</strong> factor (HSF), hsp70 and hsp90. Murine RAW 264.7 macrophages were exposed to 25mM,<br />

50mm and 75mM c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> alcohol <str<strong>on</strong>g>for</str<strong>on</strong>g> 24h, 48h and 72h followed by lipopolysaccharide (LPS) <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

15 minutes (<str<strong>on</strong>g>for</str<strong>on</strong>g> HSF and NFκB binding) or 16h (<str<strong>on</strong>g>for</str<strong>on</strong>g> hsp70, hsp90) to study the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> alcohol. At the end<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the stimulati<strong>on</strong>, nuclear extracts prepared and subjected to HSF and NFκB binding assays whereas hsp70<br />

and hsp90 in whole cell lysates were determined by Western blotting. Our findings dem<strong>on</strong>strate that HSF<br />

binding was increased after alcohol exposure. Supershift analysis showed the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF-1 proteins and<br />

not HSF-2. Interestingly, hsp70 was increased by short-term alcohol but prol<strong>on</strong>ged alcohol exposure<br />

decreased hsp70. C<strong>on</strong>versely, hsp90 levels were decreased after short-term alcohol treatment and increased by<br />

prol<strong>on</strong>ged alcohol treatment in macrophages. Immunoprecipitati<strong>on</strong> experiments <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp90, a chaper<strong>on</strong>e <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

IKK, showed decreased hsp90-IKKβ complexes after short-term alcohol whereas prol<strong>on</strong>ged alcohol revealed<br />

presence <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp90-IKKβ and increased IKK kinase activity. Geldanamycin, an hsp90 inhibitor, blocked<br />

alcohol-induced increase in TNFα producti<strong>on</strong> suggesting an important role <str<strong>on</strong>g>for</str<strong>on</strong>g> hsp90 in alcohol-induced<br />

inflammati<strong>on</strong>. Collectively, our results suggest that alcohol exposure regulates HSF-1 binding, hsp70 and<br />

hsp90 and thus could c<strong>on</strong>tribute to elevated TNFα and alcohol-induced liver injury.<br />

6C_06_S<br />

(poster secti<strong>on</strong> B2, poster board #263, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HSP90 AND ERBB2 IN THE CARDIAC RESPONSE TO DOXORUBICIN INJURY<br />

Kathleen Gabriels<strong>on</strong>, Djahida Bedja, Scott Pin, Allis<strong>on</strong> Tsao, Lucio Gama, Bibo Yuan,<br />

Nicole Muratore, Yi Xu, Nazareno Paolocci<br />

Johns Hopkins University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, 733 N Broadway Street,<br />

21205 Baltimore, USA,<br />

e-mail: kgabriel@jhmi.edu<br />

A major drawback to chemotherapeutic agent doxorubicin is cardiac toxicity. To understand the signal<br />

transducti<strong>on</strong> pathways activated in doxorubicin cardiac toxicity, and the potent synergic effect seen when<br />

doxorubicin is combined with anti-ErbB2 (trastuzumab), we developed an in vivo rat model that exhibits<br />

progressive dose-dependent cardiac damage and loss <str<strong>on</strong>g>of</str<strong>on</strong>g> cardiac functi<strong>on</strong> after doxorubicin treatment. The<br />

hearts <str<strong>on</strong>g>of</str<strong>on</strong>g> these animals resp<strong>on</strong>d to doxorubicin damage by increasing levels <str<strong>on</strong>g>of</str<strong>on</strong>g> ErbB2 and the ErbB family<br />

ligand, NRG1β, and by activating the downstream Akt signaling pathway. These increases in ErbB2 protein<br />

levels are not due to increased ErbB2 mRNA, however, suggesting post-transcripti<strong>on</strong>al mechanisms <str<strong>on</strong>g>for</str<strong>on</strong>g>


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

regulating this protein in the heart. Accordingly, levels <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP90, a known ErbB2 protein stabilizer and<br />

chaper<strong>on</strong>e, are increased by doxorubicin treatment and co-immunoprecipitati<strong>on</strong> reveals binding <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP90 to<br />

ErbB2. Isolated cardiomyocytes are more susceptible to doxorubicin after treatment with HSP90 inhibitor,<br />

17AAG, suggesting that the HSP90 is protective during doxorubicin treatment. Thus, our results provide <strong>on</strong>e<br />

plausible mechanism <str<strong>on</strong>g>for</str<strong>on</strong>g> the susceptibility <str<strong>on</strong>g>of</str<strong>on</strong>g> the heart to anti-ErbB2 therapy post-doxorubicin therapy in<br />

subclinical and clinical c<strong>on</strong>diti<strong>on</strong>s. Additi<strong>on</strong>ally, these results have lead us to further investigate the biology <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

HSP90 inhibiti<strong>on</strong> in the heart under various c<strong>on</strong>diti<strong>on</strong>s. These isolated heart studies will also be included in<br />

this presentati<strong>on</strong>.<br />

329


23-26 August 2007,<br />

Budapest, Hungary<br />

6D. STRESS, HSPS AND AUTOIMMUNITY<br />

(WILLEM VAN EDEN)<br />

6D_01_S<br />

STRESS PROTEINS ARE INDUCERS OF ANTI-INFLAMMATORY<br />

REGULATORY T CELLS<br />

Willem van Eden<br />

Dept <str<strong>on</strong>g>of</str<strong>on</strong>g> Infectious Diseases and Immunology, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Veterinary Medicine, Yalelaan 1, 3584 CL Utrecht,<br />

The Netherlands<br />

e-mail: w.vaneden@uu.nl<br />

Especially since the (re-)discovery <str<strong>on</strong>g>of</str<strong>on</strong>g> T cell subpopulati<strong>on</strong>s with specialized regulatory activities, mechanisms<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> anti-inflammatory T cell regulati<strong>on</strong> are studied very actively and are expected to lead to the development <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

novel immunotherapeutic approaches, especially in chr<strong>on</strong>ic inflammatory diseases. HSP are possible targets<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> regulatory T cells due to their enhanced expressi<strong>on</strong> in inflamed (stressed) tissues and the evidence that<br />

HSP induce anti-inflammatory immuno-regulatory T cell resp<strong>on</strong>ses.<br />

Initial evidence <str<strong>on</strong>g>for</str<strong>on</strong>g> an immuno-regulatory role <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock proteins (HSP) in chr<strong>on</strong>ic inflammati<strong>on</strong> was<br />

obtained through analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> T cell resp<strong>on</strong>ses in the rat model <str<strong>on</strong>g>of</str<strong>on</strong>g> adjuvant arthritis and the findings that HSP<br />

immunisati<strong>on</strong>s protected against the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> various <str<strong>on</strong>g>for</str<strong>on</strong>g>ms <str<strong>on</strong>g>of</str<strong>on</strong>g> autoimmune arthritis in rat and mouse<br />

models. Since then, immune reactivity to HSP was found to result from inflammati<strong>on</strong> in various disease<br />

models and human inflammatory c<strong>on</strong>diti<strong>on</strong>s, such as RA, diabetes type 1 and atherosclerosis.<br />

Now, also in the light <str<strong>on</strong>g>of</str<strong>on</strong>g> a growing interest in T cell regulati<strong>on</strong>, it is <str<strong>on</strong>g>of</str<strong>on</strong>g> interest to further explore the<br />

mechanisms through which HSP can be utilised to trigger immuno-regulatory pathways, capable <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

suppressing such a wide and diversified spectrum <str<strong>on</strong>g>of</str<strong>on</strong>g> inflammatory diseases.<br />

6D_02_S<br />

HEAT SHOCK PROTEINS AS IMMUNOMODULATORS<br />

Salvo Albani<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Cali<str<strong>on</strong>g>for</str<strong>on</strong>g>nia, San Diego, 9500 Gilman Drive, 92093-0377 La Jolla, USA<br />

e-mail: salbani@ucsd.edu<br />

The focus in clinical research in rheumatoid arthritis is increasingly shifting toward early aggressive<br />

interventi<strong>on</strong> to achieve remissi<strong>on</strong> or a state <str<strong>on</strong>g>of</str<strong>on</strong>g> low disease and to prevent the development <str<strong>on</strong>g>of</str<strong>on</strong>g> erosi<strong>on</strong>s and<br />

functi<strong>on</strong>al limitati<strong>on</strong>s. These objectives have been achieved in general by c<strong>on</strong>tinuing therapies that are costly<br />

and have l<strong>on</strong>g-term side effects. The questi<strong>on</strong> remains <strong>on</strong> whether we can induce lasting immune tolerance<br />

and, if so, what mechanisms should we target Are the drugs currently available sufficient to meet this goal<br />

and are we using them properly The fact that DMARD withdrawal leads to relapses <str<strong>on</strong>g>of</str<strong>on</strong>g> active diseases in<br />

many patients indicates that complementary approaches aimed at maintaining tolerance are needed. A<br />

translati<strong>on</strong>al itinerary from idea to end <str<strong>on</strong>g>of</str<strong>on</strong>g> Phase II trial epitope specific T cell immune therapy will be<br />

discussed as a potential complementary therapeutic tool to currently existing therapies <str<strong>on</strong>g>for</str<strong>on</strong>g> RA. This approach<br />

relies <strong>on</strong> a mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> modulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> autoimmune inflammati<strong>on</strong> based <strong>on</strong> immune recogniti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> heat<br />

shock protein derived epitopes.<br />

330


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

6D_03_S<br />

WHAT MAKES ARTERIAL ENDOTHELIAL CELLS A TARGET FOR THE<br />

AUTOIMMUNE ATTACK IN THE EARLIEST STAGES OF ATHEROSCLEROSIS<br />

Georg Wick<br />

Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Experimental Pathophysiology and Immunology, Biocenter, Innsbruck Medical University<br />

Fritz-Pregl-Str. 3/IV, 6020 Innsbruck, Austria<br />

The autoimmune hypothesis <str<strong>on</strong>g>of</str<strong>on</strong>g> atherogenesis postulates that preexisting cellular and humoral immunity to<br />

either microbial heat shock protein 60 (HSP60) or b<strong>on</strong>a fide autoimmunity to biochemically altered<br />

autologous HSP60 leads to an attack <strong>on</strong> stressed arterial endothelial cells (ECs). We have shown in various<br />

animal models with sp<strong>on</strong>taneously occurring autoimmune diseases that two essential sets <str<strong>on</strong>g>of</str<strong>on</strong>g> genes have to be<br />

present <str<strong>on</strong>g>for</str<strong>on</strong>g> an autoimmune disease to develop, i.e. genes that code <str<strong>on</strong>g>for</str<strong>on</strong>g> autoimmune reactivity <str<strong>on</strong>g>of</str<strong>on</strong>g> the immune<br />

system and genes that are resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> target organ susceptibility. In the case <str<strong>on</strong>g>of</str<strong>on</strong>g> atherosclerosis, the arterial<br />

ECs express HSP60 that is also transported to the cell surface after being subjected to classical atherosclerosis<br />

risk-factors. We have shown the HSP60-inducing effect <str<strong>on</strong>g>of</str<strong>on</strong>g> most <str<strong>on</strong>g>of</str<strong>on</strong>g> these risk-factors, including mechanical<br />

stress (hypertensi<strong>on</strong>), oxygen radicals, oxidized low-density lipoproteins (oxLDL), proinflammatory cytokines<br />

(TNFα), and cigarette smoke c<strong>on</strong>stituents. Exposure to these classical atherosclerosis risk-factors entail the<br />

simultaneous expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP60 and various adhesi<strong>on</strong> molecules (ICAM-1, VCAM-1, P-selectin). Due to<br />

their lifel<strong>on</strong>g mechanical pre-stress by the arterial blood pressure, arterial ECs have a lower threshold <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

HSP60 inducing effect <str<strong>on</strong>g>of</str<strong>on</strong>g> atherosclerosis risk-factors as compared to venous ECs. However, when veins are<br />

subjected to arterial blood flow c<strong>on</strong>diti<strong>on</strong>s, e.g. after arterial-venous bypass operati<strong>on</strong>s, HSP60 expressi<strong>on</strong><br />

and intimal infiltrati<strong>on</strong> with m<strong>on</strong><strong>on</strong>uclear cells with subsequent restenosis occurs similar to the pathogenetic<br />

events in classical atherosclerosis.<br />

6D_04_S<br />

SIX WAYS THE IMMUNE SYSTEM RECOGNIZES HSP60, A DOMINANT SELF-<br />

ANTIGEN<br />

Irun R. Cohen<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Immunology, The Weizmann Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Science, Rehovot 76100, Israel<br />

e-mail: Irun.cohen@weizmann.ac.il<br />

The immune system, using a combinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> innate and adaptive receptors, maintains the body by initiating<br />

and regulating the inflammatory resp<strong>on</strong>ses required <str<strong>on</strong>g>for</str<strong>on</strong>g> healing and protecting the body. The immune system<br />

resp<strong>on</strong>ds to a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> stresses that visit up<strong>on</strong> the body. To know when and where to heal wounds,<br />

stimulate or arrest blood-vessel growth, kill cells or stimulate their growth, dispose <str<strong>on</strong>g>of</str<strong>on</strong>g> molecular and cellular<br />

waste, and carry out the all the other immune maintenance functi<strong>on</strong>, the immune system receives vital<br />

in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> from key body molecules that serve as immune biomarkers. One such in<str<strong>on</strong>g>for</str<strong>on</strong>g>mative immune<br />

biomarker is HSP60, an infallible sign <str<strong>on</strong>g>for</str<strong>on</strong>g> regulating inflammati<strong>on</strong>. The state <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP60 reflects the state <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the cell. The immune system has at least six ways to read HSP60 signals: via innate TLR4 signaling <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

dendritic cells and macrophages; via innate TLR4 signaling <str<strong>on</strong>g>of</str<strong>on</strong>g> B cells; via innate TLR2 signaling <str<strong>on</strong>g>of</str<strong>on</strong>g> CD4CD25<br />

Tregs; via adaptive TCR signaling <str<strong>on</strong>g>of</str<strong>on</strong>g> T effector cl<strong>on</strong>es and anti-ergotypic T regulator cl<strong>on</strong>es; via adaptive<br />

BCR signaling <str<strong>on</strong>g>of</str<strong>on</strong>g> B cell cl<strong>on</strong>es; and via natural autoantibodies. Each type <str<strong>on</strong>g>of</str<strong>on</strong>g> recogniti<strong>on</strong> has its own features<br />

and its own type <str<strong>on</strong>g>of</str<strong>on</strong>g> biological output. Integrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these HSP60 signals helps the immune system reach the<br />

right decisi<strong>on</strong>s – most <str<strong>on</strong>g>of</str<strong>on</strong>g> the time. Wr<strong>on</strong>g decisi<strong>on</strong>s can lead to inflammatory disease.<br />

331


23-26 August 2007,<br />

Budapest, Hungary<br />

332<br />

6D_05_S<br />

(poster secti<strong>on</strong> B2, poster board #264, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HEAT SHOCK MODULATES INFLAMMATORY ACTIVATION OF HUMAN<br />

INTESTINAL MICROVASCULAR ENDOTHELIAL CELLS (HIMEC)<br />

Parvaneh Rafiee, Mary F. Otters<strong>on</strong>, David G. Bini<strong>on</strong><br />

Medical College <str<strong>on</strong>g>of</str<strong>on</strong>g> Wisc<strong>on</strong>sin, Milwaukee, WI<br />

Background and Aims: The heat shock resp<strong>on</strong>se is an evoluti<strong>on</strong>arily c<strong>on</strong>served mechanism <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

maintenance <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular homeostasis following sublethal noxious stimuli, where heat shock proteins (HSPs)<br />

induced by stress, chaper<strong>on</strong>e intracellular proteins that might otherwise be denatured. HSPs are now<br />

appreciated to modulate signaling cascades during periods <str<strong>on</strong>g>of</str<strong>on</strong>g> repeated stress, including chr<strong>on</strong>ic inflammati<strong>on</strong>.<br />

Although the microvascular endothelium plays a critical role in chr<strong>on</strong>ic intestinal inflammati<strong>on</strong> in IBD, little<br />

is known about the role <str<strong>on</strong>g>of</str<strong>on</strong>g> HSPs in the inflammatory activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HIMEC, the gut specific microvascular<br />

populati<strong>on</strong>.<br />

Methods: HIMECs from small and large intestine between passages 8 to12 were used. HIMECs were<br />

activated with thermal stress as well as TNF-α and LPS. HSPs and iNOS expressi<strong>on</strong> were characterized using<br />

RT-PCR and Western blot analysis. HIMEC activati<strong>on</strong> was assessed using whole cell ELISA <str<strong>on</strong>g>for</str<strong>on</strong>g> detecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

cell adhesi<strong>on</strong> molecules, and leukocyte binding under low shear stress flow adhesi<strong>on</strong>.<br />

Results: HIMECs exposed to thermal stress (42C, 1h) with recovery times <str<strong>on</strong>g>of</str<strong>on</strong>g> 1, 6 and 20h dem<strong>on</strong>strated<br />

inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP family members including HSP32 (HO-1), HSP60, HSP70, HSP90. TNF-α/LPS activati<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> HIMEC maintained at 37C also dem<strong>on</strong>strated induced expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP60 and HSP70 at the mRNA and<br />

protein levels. To investigate the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP inducti<strong>on</strong> <strong>on</strong> inflammatory activati<strong>on</strong>, HIMEC<br />

prec<strong>on</strong>diti<strong>on</strong>ed with thermal stress were assessed with and without TNF-α/LPS activati<strong>on</strong>. Heat shock al<strong>on</strong>e<br />

induced significant increases in ICAM 1. TNF-α/LPS activati<strong>on</strong> following heat shock, resulted in the highest<br />

levels <str<strong>on</strong>g>of</str<strong>on</strong>g> ICAM 1 and E selectin expressi<strong>on</strong> detected in these cells, which corresp<strong>on</strong>ded to significantly<br />

increased leukocyte adhesi<strong>on</strong>. Heat shock with and without TNF-α/LPS failed to induce increased iNOS<br />

expressi<strong>on</strong>, which normally functi<strong>on</strong>s to downregulate inflammatory activati<strong>on</strong> in HIMEC.<br />

C<strong>on</strong>clusi<strong>on</strong>s: Heat shock significantly enhanced TNF-α/LPS induced activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HIMEC. The<br />

mechanisms involved increased pro-inflammatory ICAM 1 and E selectin expressi<strong>on</strong>, which was not<br />

accompanied by an increase in the downregulatory expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> iNOS. Further defining the heat shock<br />

resp<strong>on</strong>se may yield insight into mechanisms which enhance inflammatory activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HIMEC in the setting<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic inflammati<strong>on</strong> in IBD.<br />

6D_06_S<br />

(poster secti<strong>on</strong> B2, poster board #265, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

A NOVEL ROLE OF HSP70 AS A MODULATORY AGENT FOR DENDRITIC CELLS<br />

PHENOTYPE AND FUNCTION<br />

P. Stocki, U. Holtick, N. J. Morris, A.M. Dickins<strong>on</strong><br />

Haematological Sciences, Newcastle University, UK<br />

e-mail: pawel.stocki@ncl.ac.uk<br />

Heat shock protein 70 (HSP70) is a chaper<strong>on</strong>e proteins which was reported to be implicated in an activati<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> innate immunity. However, recent data str<strong>on</strong>gly opposes the proposed functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 as a


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

proinflammatory cytokine highlighting the problem <str<strong>on</strong>g>of</str<strong>on</strong>g> a bacterial c<strong>on</strong>taminati<strong>on</strong> in a purified recombinant<br />

human HSP70 (rhHSP70). As it was shown, endotoxin free rhHSP70 is incapable to launch immune<br />

activati<strong>on</strong> raising the rhHSP70-endotoxin debate and suggesting that the real role <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 in an immune<br />

system, if any, hasn’t been reviled yet. To eliminate the problem <str<strong>on</strong>g>of</str<strong>on</strong>g> endotoxin c<strong>on</strong>taminati<strong>on</strong> we purified<br />

HSP70 from two different leukemic cell lines which were growing in the endotoxin free envir<strong>on</strong>ment. In<br />

additi<strong>on</strong>, the purificati<strong>on</strong> products were tested <str<strong>on</strong>g>for</str<strong>on</strong>g> LPS c<strong>on</strong>tent giving negative result. We examined the<br />

HSP70 role <strong>on</strong> m<strong>on</strong>ocyte derived dendritic cells (DC) presenting its modulatory effect <strong>on</strong> a DC activati<strong>on</strong><br />

state. 24h incubati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> DC with HSP70 was effective in down regulating surface expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HLA-DR as<br />

well as CD80/86/83. As a c<strong>on</strong>trol we used an endotoxin-c<strong>on</strong>taminated rhHSP70 (erhHSP70) which in<br />

c<strong>on</strong>trast to HSP70 was highly potent DC maturati<strong>on</strong> molecule. We further examined the functi<strong>on</strong>al effect <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

HSP70 treated DC <strong>on</strong> proliferati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> allogeneic T cells, where HSP70 was resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> its down<br />

regulati<strong>on</strong>. The modulatory effect <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 <strong>on</strong> DC was also investigated <strong>on</strong> DC cytokine level after CD40L<br />

activati<strong>on</strong>, presenting decreased level <str<strong>on</strong>g>of</str<strong>on</strong>g> IL-4/6/10 and TNFα c<strong>on</strong>firming the observed phenomen<strong>on</strong>.<br />

Although our results which clearly show that HSP70 has an immunological potential as an anti inflammatory<br />

agent are c<strong>on</strong>tradicting most <str<strong>on</strong>g>of</str<strong>on</strong>g> the published data they are supported by the most recent publicati<strong>on</strong> <strong>on</strong> the<br />

field <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 c<strong>on</strong>scious about possible endotoxin c<strong>on</strong>taminati<strong>on</strong>.<br />

333


23-26 August 2007,<br />

Budapest, Hungary<br />

334<br />

6D_01_P<br />

(poster secti<strong>on</strong> B2, poster board #266, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

BRIEF ACADEMIC STRESS AFFECTS ANTI HSP70 ANTIBODIES, CORTISOL AND<br />

PSYCHOLOGICAL FACTORS<br />

Orly Sarid 1 , Ofra Ans<strong>on</strong> 2 , Arieh Yaari 2<br />

1Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Humanities & Social Sciences, 2 Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Health Sciences; Ben-Guri<strong>on</strong> University <str<strong>on</strong>g>of</str<strong>on</strong>g> the Negev<br />

Beer-Sheva, Israel<br />

The objectives <str<strong>on</strong>g>of</str<strong>on</strong>g> the study were to explore the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> brief academic stress <strong>on</strong> anti Hsp70 and cortisol in<br />

serum and saliva and <strong>on</strong> psychological indicators; and to examine the associati<strong>on</strong>s between psycho-biological<br />

indicators. A brief academic stress <str<strong>on</strong>g>of</str<strong>on</strong>g> venopuncture and blood withdrawal was chosen as a model <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

measuring transient mood, endocrine and cell chaper<strong>on</strong>e changes. The study populati<strong>on</strong> c<strong>on</strong>sisted <str<strong>on</strong>g>of</str<strong>on</strong>g> 28<br />

healthy students, practicing venopuncture as part <str<strong>on</strong>g>of</str<strong>on</strong>g> their curricular activity. Sera and saliva were collected<br />

and analyzed <str<strong>on</strong>g>for</str<strong>on</strong>g> the existence <str<strong>on</strong>g>of</str<strong>on</strong>g> anti Hsp70 and cortisol using an anti-human Hsp70 (IgG/A/M) and<br />

cortisol ELISA Kits and following the manufacture’s instructi<strong>on</strong>s. Pencil and paper questi<strong>on</strong>naires were<br />

administered be<str<strong>on</strong>g>for</str<strong>on</strong>g>e (t1) and after (t2) venopuncture and saliva d<strong>on</strong>ati<strong>on</strong>. Cortisol and anti Hsp70 antibodies<br />

were detected in serum and saliva am<strong>on</strong>g all participants. Additi<strong>on</strong>ally, a change in some psychological<br />

indicators between t1 and t2 took place. Anti Hsp70 in serum was positively correlated with anxiety (t2)<br />

(r=0.47, p≤0.05) and with anger at t2 ((r=0.54, p≤0.05). Cortisol in serum was negatively correlated with<br />

vigor (t1: r=-0.44, t2: r=-0.48, p≤0.05).<br />

In c<strong>on</strong>clusi<strong>on</strong>, a brief stress activates endocrine and cell chaper<strong>on</strong>e system in serum and saliva, which is<br />

rarely studied in Hsps c<strong>on</strong>text. However associati<strong>on</strong>s with psychological variables were detected <strong>on</strong>ly in<br />

the serum.<br />

6D_02_P<br />

(poster secti<strong>on</strong> B2, poster board #267, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE ROLE OF BACTERIAL DNAJ PROTEIN AND ITS HUMAN HOMOLOG HDJ-2<br />

IN PATHOGENESIS OF RHEUMATOID ARTHRITIS<br />

Stefan Tukaj 1 , Agnieszka Kotlarz 1 , Jolanta Tyborowska 2 , Barbara Lipinska 1<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Gdansk, Kladki 24, Gdansk, Poland<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular Virology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Gdansk, Kladki 24, Poland<br />

e-mail: tukaj@biotech.univ.gda.pl<br />

Escherichia coli DnaJ (Hsp40) heat shock protein is suspected to participate in pathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> rheumatoid<br />

arthritis (RA) in humans by an autoimmune process. We have found previously, using ELISA assay, that<br />

reacti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the RA patients sera with DnaJ protein and with its human homolog HDJ-2 (produced in<br />

bacteria) was at least 2-fold and 5-fold str<strong>on</strong>ger, respectively, than the reacti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the c<strong>on</strong>trol group, and there<br />

was a str<strong>on</strong>g positive correlati<strong>on</strong> between the immune resp<strong>on</strong>se to DnaJ protein and HDJ-2 in RA.<br />

There<str<strong>on</strong>g>for</str<strong>on</strong>g>e it is possible that in RA the immune resp<strong>on</strong>se directed against the bacterial protein could crossreact<br />

with the human homologous protein. In this study, similar results were obtained when the HDJ-2<br />

protein produced in insect cells (and there<str<strong>on</strong>g>for</str<strong>on</strong>g>e modified by farnezylati<strong>on</strong> at the C-terminal end) was used as<br />

an antigen. These results c<strong>on</strong>firmed previous findings and indicated also that antigenic properties <str<strong>on</strong>g>of</str<strong>on</strong>g> HDJ-2<br />

did not change significantly up<strong>on</strong> farnezylati<strong>on</strong>. We have assayed antibody levels against the N-terminal<br />

fragment <str<strong>on</strong>g>of</str<strong>on</strong>g> HDJ-2 encompassing the “J” domain, and against its C-terminal domain. The humoral resp<strong>on</strong>se


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

against the C-terminal domain was significantly increased in the RA patients compared to c<strong>on</strong>trols, and this<br />

resp<strong>on</strong>se correlated very well with the anti-DnaJ resp<strong>on</strong>se. The anti-DnaJ m<strong>on</strong>ocl<strong>on</strong>al antibodies recognizing<br />

the C-terminal domain <str<strong>on</strong>g>of</str<strong>on</strong>g> DnaJ reacted very efficiently with the human HDJ-2, indicating that the C-terminal<br />

part <str<strong>on</strong>g>of</str<strong>on</strong>g> HDJ-2 is immunologically similar to its counterpart <str<strong>on</strong>g>of</str<strong>on</strong>g> DnaJ. These results support a hypothesis that<br />

human HDJ-2 may functi<strong>on</strong> as an autoantigen in RA and suggest that the C-terminal part <str<strong>on</strong>g>of</str<strong>on</strong>g> HDJ-2 plays an<br />

important role in this process.<br />

6D_03_P<br />

(poster secti<strong>on</strong> B2, poster board #268, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

NASAL ADMINISTRATION OF MYCOBACTERIAL HSP70 OR DERIVED PEPTIDES<br />

SUPPRESS PROTEOGLYCAN INDUCED ARTHRITIS<br />

Lotte Wieten, Ruurd van der Zee, Elles Klein Koerkamp, Jose Wagenaar-Hilbers, Willem van Eden,<br />

Femke Hauet-Broere<br />

Dept. Inf. Dis. & Immunol., FacVetMed,Utrecht University, The Netherlands<br />

e-mail: r.vanderzee@vet.uu.nl<br />

T cells specific <str<strong>on</strong>g>for</str<strong>on</strong>g> bacterial Hsp’s and cross-reactive with self are involved in regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic<br />

inflammatory diseases. In the present study we investigated the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> nasal administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

mycobacterial Hsp70 and cross-reactive peptides <strong>on</strong> proteoglycan induced arthritis, a mouse model <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

rheumatoid arthritis. Arthritis was induced in Balb/c mice by i.p. immunisati<strong>on</strong> with human proteoglycan<br />

(hPG), <strong>on</strong> day 0 and 21. Nasal treatment with mycob. Hsp70 or peptides was d<strong>on</strong>e <strong>on</strong> day -7, -5 and -3. Mice<br />

were sacrificed 3.5 weeks after development <str<strong>on</strong>g>of</str<strong>on</strong>g> arthritis and splenocytes were stimulated with Hsp70 and PG,<br />

and proliferati<strong>on</strong>, cytokine producti<strong>on</strong> and mRNA expressi<strong>on</strong> were analysed. Intranasal administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

mycobacterial HspP70 led to a delayed <strong>on</strong>set <str<strong>on</strong>g>of</str<strong>on</strong>g> arthritis after which the mice developed less severe arthritis<br />

than c<strong>on</strong>trol mice receiving PBS intranasally. The maximum mean arthritis score was significantly (p=0.026)<br />

lower in Hsp70 treated mice (3.8±3.3) compared to c<strong>on</strong>trol mice (9.4±2.6). RT- PCR analysis showed that<br />

intranasal administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 led to reduced pro-inflammatory cytokine mRNA expressi<strong>on</strong> and<br />

augmented IL-10 mRNA expressi<strong>on</strong>. CD4 mRNA expressi<strong>on</strong> was decreased in the joints <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 treated<br />

mice, indicating a reduced (pathogenic) CD4 + T-cell influx in the joint. Similar treatments with mycob. Hsp70<br />

peptides comprising cross-reactive T cell inducing epitopes also ameliorated disease development. Thus,<br />

intranasal administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> mycob. Hsp70 induces a protective immune resp<strong>on</strong>se in the model <str<strong>on</strong>g>of</str<strong>on</strong>g> PGIA. In<br />

additi<strong>on</strong>, we identified Hsp70 peptides capable <str<strong>on</strong>g>of</str<strong>on</strong>g> inducing regulatory T cells that can inhibit experimentally<br />

induced arthritis.<br />

335


23-26 August 2007,<br />

Budapest, Hungary<br />

336<br />

6E. STRESS AND THE METABOLIC SYNDROME:<br />

HYPERTENSION, ATHEROSCLEROSIS, OBESITY AND DIABETES<br />

(JAN ERIKSSON, R. PAUL ROBERTSON)<br />

6E_01_S<br />

PSYCHOSOCIAL FACTORS IN THE METABOLIC SYNDROME<br />

Eric Brunner<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Epidemiology and Public Health, University College L<strong>on</strong>d<strong>on</strong>, WC1E 6BT, UK<br />

e-mail: e.brunner@ucl.ac.uk<br />

The cluster <str<strong>on</strong>g>of</str<strong>on</strong>g> cardiovascular risk factors labeled the metabolic syndrome is linked with low social status.<br />

Prevalence studies show a stepwise relati<strong>on</strong>ship - the lower the social positi<strong>on</strong>, the greater the probability the<br />

syndrome will be present. Systematic differences in diet and physical activity c<strong>on</strong>tribute to social patterning <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the syndrome. In additi<strong>on</strong>, psychosocial factors including chr<strong>on</strong>ic work stress are linked with its development.<br />

The Whitehall II study, set up in 1985 to study the psychosocial, behavioral and biological causes <str<strong>on</strong>g>of</str<strong>on</strong>g> health<br />

inequalities in an <str<strong>on</strong>g>of</str<strong>on</strong>g>fice-based cohort <str<strong>on</strong>g>of</str<strong>on</strong>g> 10,308 adults, obtained repeat measures <str<strong>on</strong>g>of</str<strong>on</strong>g> job stress using a standard<br />

self-report questi<strong>on</strong>naire (Karasek-Theorell job strain questi<strong>on</strong>naire). Comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g> the metabolic syndrome<br />

(waist circumference, BP, fasting glucose, serum HDL-cholesterol and triglycerides) were measured after 14<br />

years <str<strong>on</strong>g>of</str<strong>on</strong>g> follow-up. A dose-resp<strong>on</strong>se relati<strong>on</strong> was found between exposure to job stress and risk <str<strong>on</strong>g>of</str<strong>on</strong>g> ATPIII<br />

metabolic syndrome which remained after adjustment <str<strong>on</strong>g>for</str<strong>on</strong>g> age and socioec<strong>on</strong>omic status (OR=2.25 (95% CI<br />

1.3-3.9)). A sec<strong>on</strong>d prospective study showed a similar dose-resp<strong>on</strong>se relati<strong>on</strong> with general (BMI) obesity and<br />

central obesity. These findings add to other evidence that psychosocial stressors from everyday life are linked<br />

to cor<strong>on</strong>ary risk. Cross-secti<strong>on</strong>al studies link the metabolic syndrome to adverse neuroendocrine and<br />

aut<strong>on</strong>omic activity (increased urinary cortisol and noradrenaline metabolite outputs, and decreased heart rate<br />

variability). The metabolic syndrome is a valuable research c<strong>on</strong>cept <str<strong>on</strong>g>for</str<strong>on</strong>g> studying populati<strong>on</strong> health and socialbiological<br />

translati<strong>on</strong>.<br />

6E_02_S<br />

NEUROENDOCRINE MECHANISMS IN THE METABOLIC SYNDROME<br />

Jan W. Erikss<strong>on</strong><br />

Sahlgrenska University Hospital, Gothenburg and AstraZeneca R&D, Molndal, Sweden<br />

e-mail address: jan.erikss<strong>on</strong>@medic.gu.se, jan.erikss<strong>on</strong>@astrazeneca.com<br />

Psychosocial stress is now c<strong>on</strong>sidered to be an important risk factor <str<strong>on</strong>g>for</str<strong>on</strong>g> metabolic syndrome and type 2<br />

diabetes, and development <str<strong>on</strong>g>of</str<strong>on</strong>g> insulin resistance can be a comm<strong>on</strong> denominator. Insulin resistance can be<br />

described as an impaired insulin acti<strong>on</strong> in target tissues, i.e. muscle, fat and liver. In muscle, insulin-stimulated<br />

transmembrane glucose transport and the first step in intracellar glucose metabolism, i.e. glucose-6-<br />

phosphorylati<strong>on</strong>, appear to be rate-limiting defects. In adipose tissue, insulin resistance is manifested as<br />

impaired glucose uptake but also as an impaired suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> lipolysis and, in additi<strong>on</strong>, there may<br />

potentially be dysregulated producti<strong>on</strong> and secreti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> adipokines and other adipose-derived biomolecules.<br />

In the liver, there is attenuated insulin acti<strong>on</strong> with respect to glucose uptake and storage as well as<br />

suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> glucose producti<strong>on</strong>.<br />

Type 2 diabetes is in most cases caused by a combinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> beta cell dysfuncti<strong>on</strong> and insulin resistance.<br />

Physical inactivity, adiposity due to overeating, stress and smoking are risk factors that interact with


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

susceptibility genes in the development <str<strong>on</strong>g>of</str<strong>on</strong>g> the disease. The metabolic syndrome is <str<strong>on</strong>g>of</str<strong>on</strong>g>ten used to define a<br />

cluster <str<strong>on</strong>g>of</str<strong>on</strong>g> risk markers that predict cardiovascular disease but also type 2 diabetes.<br />

There is no c<strong>on</strong>sensus about a unifying underlying mechanism in insulin resistance. However, it is now<br />

generally accepted that central adiposity plays a key role in the metabolic syndrome and it is linked to insulin<br />

resistance and risk <str<strong>on</strong>g>for</str<strong>on</strong>g> type 2 diabetes as well as cardiovascular disease. Dysregulated neuroendocrine<br />

signalling in adipose tissue can c<strong>on</strong>tribute to the development <str<strong>on</strong>g>of</str<strong>on</strong>g> insulin resistance and metabolic syndrome,<br />

and such pathways can mediate the metabolic effect <str<strong>on</strong>g>of</str<strong>on</strong>g> stress resp<strong>on</strong>ses that are evoked in the CNS. An<br />

overview <str<strong>on</strong>g>of</str<strong>on</strong>g> the field will be given and recent results will be presented. Topics will include the links between<br />

neuroendocrine dysregulati<strong>on</strong> and visceral fat accumulati<strong>on</strong>, fat cell metabolism, alterati<strong>on</strong>s in fat cell<br />

recruitment, differentiati<strong>on</strong> and growth as well as adipokine producti<strong>on</strong>.<br />

Recent data will be presented with focus <strong>on</strong> the the interplay between different neuroendocrine sysems - the<br />

cortisol axis, sex horm<strong>on</strong>es, the aut<strong>on</strong>omic nervous system, catecholamines and the renin-angiotensin system.<br />

A hypothesis will be presented about a comm<strong>on</strong> ‘final pathway’ that might merge the insulin-antag<strong>on</strong>istic<br />

acti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> horm<strong>on</strong>es, inflammati<strong>on</strong>, glucose and lipids, namely the generati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> excess reactive oxygen<br />

species (ROS). Potential pharmacological interventi<strong>on</strong>s targeting the different signalling pathways, including<br />

oxidative stress, will also be discussed.<br />

6E_03_S<br />

ROLE OF STRESS RESPONSE GENES, INCLUDING HO-1, IN TYPE 1 DIABETES<br />

AND DIABETIC COMPLICATIONS<br />

Nader G. Abraham<br />

Stem Cells and Gene Therapy, New York Medical College, Valhalla, New York 10595<br />

Hyperglycemic episodes, which cause vascular complicati<strong>on</strong>s as a result <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidative stress and complicate<br />

even well c<strong>on</strong>trolled cases <str<strong>on</strong>g>of</str<strong>on</strong>g> diabetes, are closely associated with the development <str<strong>on</strong>g>of</str<strong>on</strong>g> vascular disease. Several<br />

antioxidants and stress resp<strong>on</strong>se genes have been shown to partially protect the vascular system. However,<br />

the c<strong>on</strong>tinuous generati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> O 2- via heme-dependent enzymes, including NADPH oxidase and mitoch<strong>on</strong>drial<br />

cytochrome, has shown that such a strategy is difficult to achieve and problematic in diabetes. Heme<br />

oxygenase (HO-1) is the sole enzyme that degrades heme and subsequently regulates NADPH oxidase and<br />

the cytochrome oxidase system and O 2- <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> via increases in bilirubin generati<strong>on</strong> and ferritin synthesis,<br />

which are anti-oxidants, and CO, which is anti-apoptotic. In an ef<str<strong>on</strong>g>for</str<strong>on</strong>g>t to overcome this impasse, we examined<br />

molecular (gene transfer) and pharmacological (cobalt protoporphyrin, CoPP) approaches to increased HO-<br />

1/HO-2 expressi<strong>on</strong> in providing cardiovascular protecti<strong>on</strong> in Type I diabetic animal models. Upregulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

HO-1 expressi<strong>on</strong> attenuates hyperglycemia-mediated increases in vascular dysfuncti<strong>on</strong>, circulating endothelial<br />

cells and fragmentati<strong>on</strong>, decreases O 2- , heme levels and iNOS, but increases EC-SOD. Translocati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HO-<br />

1 to the mitoch<strong>on</strong>drial compartments enhances cytochrome oxidase and anti-apoptotic molecules, and<br />

decreases cytochrome C release, suggesting that HO-1 may regulate mitoch<strong>on</strong>drial pro-apoptotic and antiapoptotic<br />

proteins. Using the loss-<str<strong>on</strong>g>of</str<strong>on</strong>g>-functi<strong>on</strong>, HO-2(-/-) and gain-<str<strong>on</strong>g>of</str<strong>on</strong>g>-functi<strong>on</strong> strategy, we will present data<br />

to document that HO-2 siRNA treatment decreased basal levels <str<strong>on</strong>g>of</str<strong>on</strong>g> EC-SOD and phosphorylated Akt<br />

proteins (serine-473 and thre<strong>on</strong>ine-308), although no change in Akt protein expressi<strong>on</strong> was observed. HO-2<br />

siRNA was also associated with an increase in 3-nitrotyrosine (3-NT) and apoptotic signaling kinase-1 (ASK-<br />

1). Additi<strong>on</strong>ally, we will present data to show that an increase in HO-1/HO-2 levels, i.e., CO and bilirubin,<br />

has a salutary effect, modulating the vascular phenotype, as reflected by the increases in anti-apoptotic<br />

proteins, thus rendering endothelial cells resistant to oxidant stress hence preventing vascular dysfuncti<strong>on</strong> and<br />

the development <str<strong>on</strong>g>of</str<strong>on</strong>g> Type 1 diabetes.<br />

337


23-26 August 2007,<br />

Budapest, Hungary<br />

338<br />

6E_04_S<br />

CHRONIC OXIDATIVE STRESS AS A MECHANISM FOR GLUCOSE TOXICITY OF<br />

THE BETA CELL<br />

Paul Roberts<strong>on</strong><br />

Pacific Northwest <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute, 98122, 720 Broadway, Seattle, WA 98122, USA<br />

e-mail: rpr@pnri.org<br />

In studies that used the beta cell line HIT-T15, we made the observati<strong>on</strong> that over many passages chr<strong>on</strong>ic<br />

exposure <str<strong>on</strong>g>of</str<strong>on</strong>g> beta cells leads to decreased insulin gene expressi<strong>on</strong>, insulin stores, and glucose-induced insulin<br />

secreti<strong>on</strong>. We next identified the molecular defects <str<strong>on</strong>g>for</str<strong>on</strong>g> decreased insulin mRNA levels. Protein levels <str<strong>on</strong>g>of</str<strong>on</strong>g> two<br />

critically important transcripti<strong>on</strong> factors, PDX-1 and MafA, were low to n<strong>on</strong>-detectable after prol<strong>on</strong>ged<br />

culturing <str<strong>on</strong>g>of</str<strong>on</strong>g> HIT-T15 cells in media c<strong>on</strong>taining supraphysiologic c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> glucose. We was<br />

determined that the molecular mechanism <str<strong>on</strong>g>for</str<strong>on</strong>g> decreased PDX-1 levels is post-transcripti<strong>on</strong>al, whereas as the<br />

mechanism <str<strong>on</strong>g>for</str<strong>on</strong>g> decreased MafA is post-translati<strong>on</strong>al. The transcripti<strong>on</strong>al machinery needed <str<strong>on</strong>g>for</str<strong>on</strong>g> gene<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> insulin, PDX-1, and MafA was not abnormal. Attenti<strong>on</strong> was focused <strong>on</strong> the insulin promoter<br />

regi<strong>on</strong> through a series <str<strong>on</strong>g>of</str<strong>on</strong>g> studies involving mutati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the PDX-1 and MafA DNA binding sites and<br />

rec<strong>on</strong>stituti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> glucotoxic beta cells with these two proteins. Mutati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the insulin promoter binding sites<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> either protein in n<strong>on</strong>-glucose toxic beta cells led to marked decreased in promoter activity. Rec<strong>on</strong>stituti<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> glucotoxic beta cells by transient transfecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> either PDX-1 or MafA lead to improved promoter<br />

activity. Most recently, it was shown that adenoviral rec<strong>on</strong>stituti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HIT-T15 cells with both PDX-1 and<br />

MafA fully restored insulin promoter reporter activity and partially normalized levels <str<strong>on</strong>g>of</str<strong>on</strong>g> endogenous insulin<br />

mRNA. As these experiments were evolving, it became evident from the c<strong>on</strong>cepts put <str<strong>on</strong>g>for</str<strong>on</strong>g>ward by Wolff and<br />

colleagues that the mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> glucose toxicity might involve generati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ROS, specifically from glucose<br />

autoxidati<strong>on</strong>. This line <str<strong>on</strong>g>of</str<strong>on</strong>g> thinking was instrumental in early work assessing whether the defects in insulin<br />

gene expressi<strong>on</strong> and abnormal insulin secreti<strong>on</strong> associated with exposure to high glucose c<strong>on</strong>centrati<strong>on</strong>s<br />

could be ameliorated by antioxidants. This led to reports <str<strong>on</strong>g>of</str<strong>on</strong>g> preventive effects <str<strong>on</strong>g>of</str<strong>on</strong>g> antioxidants against glucose<br />

toxicity in vitro in beta cell lines and in vivo in animal models <str<strong>on</strong>g>of</str<strong>on</strong>g> type 2 diabetes. Inclusi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> NAC or<br />

aminoguanidine in media c<strong>on</strong>taining supraphysiologic c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> glucose protected HIT-T15 cells<br />

against the loss <str<strong>on</strong>g>of</str<strong>on</strong>g> insulin gene expressi<strong>on</strong>. NAC preserved insulin gene expressi<strong>on</strong> and beta cell functi<strong>on</strong> in<br />

Zucker Diabetic Fatty rats and db/db mice. These lines <str<strong>on</strong>g>of</str<strong>on</strong>g> research str<strong>on</strong>gly suggest that glucose toxicity <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the beta cell is mediated through oxidative stress, which in turn causes defects in synthesis <str<strong>on</strong>g>of</str<strong>on</strong>g> PDX-1 and<br />

MafA.<br />

6E_05_S<br />

HSP72 PROTECTS AGAINST OBESITY-INDUCED INSULIN RESISTANCE<br />

Mark A. Febbraio<br />

Baker Heart <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute, Melbourne, Australia<br />

e-mail: mark.febbraio@baker.edu.au<br />

Patients with type 2 diabetes have reduced gene expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Heat Shock Protein (HSP) 72 which correlates<br />

with reduced insulin sensitivity. Heat therapy, which activates HSP72, improves clinical parameters in these<br />

patients. Activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> several inflammatory signaling proteins such as c-jun amino terminal kinase (JNK),<br />

inhibitor <str<strong>on</strong>g>of</str<strong>on</strong>g> κB kinase and tumor necrosis factor-α can induce insulin resistance, but HSP 72 can block the<br />

inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these molecules in vitro. Accordingly, we examined whether activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP72 can protect


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

against the development <str<strong>on</strong>g>of</str<strong>on</strong>g> insulin resistance. We first show that obese, insulin resistant humans have<br />

reduced HSP72 protein expressi<strong>on</strong> and increased JNK phosphorylati<strong>on</strong> in skeletal muscle. We next utilized<br />

heat shock therapy, transgenic overexpressi<strong>on</strong>, and pharmacologic means to overexpress HSP72 either<br />

specifically in skeletal muscle or globally in mice. Herein we show that regardless <str<strong>on</strong>g>of</str<strong>on</strong>g> the means used to<br />

achieve an elevati<strong>on</strong> in HSP72 protein, protecti<strong>on</strong> against diet or obesity-induced hyperglycemia,<br />

hyperinsulinemia, glucose intolerance and insulin resistance was observed. This protecti<strong>on</strong> was tightly<br />

associated with the preventi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> JNK phosphorylati<strong>on</strong>. These findings identify an essential role <str<strong>on</strong>g>for</str<strong>on</strong>g> HSP72<br />

in blocking inflammati<strong>on</strong> and preventing insulin resistance in the c<strong>on</strong>text <str<strong>on</strong>g>of</str<strong>on</strong>g> genetic obesity or high fat<br />

feeding.<br />

6E_06_S<br />

(poster secti<strong>on</strong> B1, poster board #196, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HEART RATE VARIABILITY AND GLYCATED HEMOGLOBIN<br />

Nanna Hurwitz Eller<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Occupati<strong>on</strong>al Medicine, Hilleroed Hospital, 3400 Hilleroed, Denmark<br />

Introducti<strong>on</strong>: By the means <str<strong>on</strong>g>of</str<strong>on</strong>g> spectral analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> heart rate, the activity in the aut<strong>on</strong>omous nervous system<br />

can be expressed as Heart Rate Variability, HRV. Low Total Power, TP (total variance in HRV, ms 2 ) and low High<br />

Frequency power, HF (0,18-0,4Hz), the latter known to mirror vagal t<strong>on</strong>e, is associated with higher<br />

cardiovascular morbidity.<br />

Aim: To analyse associati<strong>on</strong>s between TP, HF and HbA1c.<br />

Methods: The study included seventy-four healthy n<strong>on</strong>-diabetic participants (50 women, 24 men, mean age<br />

49 years). Levels <str<strong>on</strong>g>of</str<strong>on</strong>g> HbA1c were obtained in 1998 and 2002. In 2002 ECG during the 45 minutes clinical<br />

examinati<strong>on</strong> were analysed and spectral analysis carried out. LogTP, logHF and logHF /TP were used as<br />

dependent variables. The analyses were carried out by means <str<strong>on</strong>g>of</str<strong>on</strong>g> general linear models, repeated<br />

measurements (9 levels <str<strong>on</strong>g>of</str<strong>on</strong>g> 5 minutes), separately <str<strong>on</strong>g>for</str<strong>on</strong>g> each gender using HbA1c in 1998, 2002 and expressed as<br />

change between 1998 and 2002 as the independent variable, <strong>on</strong>e at a time. Results were adjusted <str<strong>on</strong>g>for</str<strong>on</strong>g> age and<br />

time <str<strong>on</strong>g>of</str<strong>on</strong>g> examinati<strong>on</strong>.<br />

Results: HbA1c in 1998 was <strong>on</strong>ly weakly associated with HRV in 2002. HbA1c in 2002 and expressed as<br />

change (2002-1998) were significantly associated with HRV, i.e. high HbA1c and high change were associated<br />

with low HRV, most pr<strong>on</strong>ounced in men.<br />

C<strong>on</strong>clusi<strong>on</strong>s: Increase in HbA1c and actual level <str<strong>on</strong>g>of</str<strong>on</strong>g> HbA1c was str<strong>on</strong>gly associated with HRV, in n<strong>on</strong>diabetic<br />

women and men. This indicates that the increased cardiovascular risk accompanying insulin<br />

resistance may be caused by low HRV.<br />

339


23-26 August 2007,<br />

Budapest, Hungary<br />

340<br />

6E_02_P<br />

(poster secti<strong>on</strong> B1, poster board #197, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EPIDEMIOLOGICAL STUDY OF POLYMORPHISMS OF HSP72 PROMOTER GENE<br />

AND ITS TRANSLATION PRODUCT<br />

M. C. Guisasola, E. Dulin*, M. M. Desco, M. J. Sanchez, P. García-Barreno<br />

Experimental Med & Surgical Unit.*Clinical Biochemistry Dep. Hosp Gen Univ Gregorio Maran<strong>on</strong>.<br />

Dr Esquerdo 46. 28007.Madrid, Spain<br />

e-mail: mguisasola@mce.hggm.es<br />

Atheroesclerosis is a chr<strong>on</strong>ic inflammatory disease: cells involved are m<strong>on</strong>ocytes, macrophages, and T<br />

lymphocytes. Neutrophil polymorph<strong>on</strong>uclear leukocytes (PMNs) are also pathogenic releasing<br />

myeloperoxidase tightly linked to atherosclerosis and cardiovascular disease. Candidate<br />

molecules/autoantigens proposed include members <str<strong>on</strong>g>of</str<strong>on</strong>g> the Heat Shock Proteins (HSPs). Polymorphisms <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the hsp72 gene regulator regi<strong>on</strong>s, could account <str<strong>on</strong>g>for</str<strong>on</strong>g> different sensitivities <str<strong>on</strong>g>for</str<strong>on</strong>g> the development <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

cardiovascular disease. Epidemiological study <str<strong>on</strong>g>of</str<strong>on</strong>g> these polymorphisms and its possible correlati<strong>on</strong> with<br />

morbi-mortality <str<strong>on</strong>g>of</str<strong>on</strong>g> populati<strong>on</strong>, may lead to identify efficient tools to be used as biomarkers. 92 ♀ (age 49,01<br />

± 0.68) and 106 ♂ (48,05 ± 0,66) were included randomly. Molecular study <str<strong>on</strong>g>of</str<strong>on</strong>g> promoter and flanking<br />

sequence <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp72 gene (NT_ 007592) in PMNs, PMNs[HSP72] and circulating HSP72 were per<str<strong>on</strong>g>for</str<strong>on</strong>g>med.<br />

Statistical analysis: Mann-Whitney’s U-test, and ANOVA <strong>on</strong>e-way. Results: Subjects were divided into 3<br />

groups: G0 without vascular risk factors [n=113 (54 men, 59 women)]; G1 with moderate risk factors (10%),<br />

without illness [n=55 (32 men, 23 women)]; and G2 with evident AT disease [n= 30 (19 men, 11<br />

women)].Three SNPs were detected: -325A→C,-95T→C and -27G→C. SNP -325: 71 subjects (35,9%) were<br />

wild type (AA), 99 heterozygotes (46,5%) (AC), and 35 (17,7%) homozygotes (CC). Homozygotes had the<br />

highest [HSP72]i in G2 (p= 0,038 Anova). SNP -27 was co-expressed with -325 in 98,49% <str<strong>on</strong>g>of</str<strong>on</strong>g> total cases (73<br />

WT, 93 HT, 32 HM).In the same way with SNP –325, CC phenotypes presents significant highest [HSP72]i<br />

in G2. SNP -95 was <strong>on</strong>ly detected in heterozygosis in 20 subjects (10.1%) and it was related to lowest<br />

[HSP72]i in AT (p=0,052, Anova), and to maximal levels in patients with vascular risk factors. With respect<br />

to circulating HSP72, the lowest levels bel<strong>on</strong>ged to G2 (p= 0,94), independently <str<strong>on</strong>g>of</str<strong>on</strong>g> the phenotype <str<strong>on</strong>g>for</str<strong>on</strong>g> any <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the SNPs. C<strong>on</strong>clusi<strong>on</strong>s: in our study, SNPs -325 and –27 were related in their homozygote <str<strong>on</strong>g>for</str<strong>on</strong>g>m, with<br />

highest [HSP7] i , which has been proved previously to be antinmflamatory, and in c<strong>on</strong>sequence, protective <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

AT development and progressi<strong>on</strong>. Heterozygosis <str<strong>on</strong>g>of</str<strong>on</strong>g> –95 may be cardiovascular protective, but when AT is<br />

yet established, results in an additi<strong>on</strong>al vascular risk factor <str<strong>on</strong>g>of</str<strong>on</strong>g> AT disease. Grants from FIS 03/1308 and<br />

FMM.<br />

6E_03_P<br />

(poster secti<strong>on</strong> B1, poster board #198, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

MODULATION OF ANTIOXIDANT STATUS IN A MACROPHAGE CELL LINE<br />

DURING CHRONIC EXPOSURE TO GLYCATED SERUM<br />

Anna Bassi<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Genoa, Experimental Medicine (DiMeS), General Pathology Secti<strong>on</strong><br />

16132 Via L.B. Alberti 2, Genoa, Italy<br />

e-mail: ambassi@medicina.unige.it<br />

Advanced glycati<strong>on</strong> end-products (AGEs) are involved in the pathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> aging and correlated<br />

degenerative pathologies, such as diabetic and uremic complicati<strong>on</strong>s, atherosclerosis and amyloidosis. The


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

aim <str<strong>on</strong>g>of</str<strong>on</strong>g> the present study was to reproduce those physiopathological c<strong>on</strong>diti<strong>on</strong>s that precede the clinical<br />

aspects <str<strong>on</strong>g>of</str<strong>on</strong>g> diabetes or other aging-related diseases and resulted <strong>on</strong>ly in mild symptoms such as less<br />

appreciable levels <str<strong>on</strong>g>of</str<strong>on</strong>g> AGE products (i.e. pentosidine). Oxidative stress-related parameters were nalysed in<br />

J774A.1 murine macrophage cells during chr<strong>on</strong>ic exposure (7 days) to subtoxic c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> AGE (5%<br />

ribose-glycated serum, GS) and subsequently <str<strong>on</strong>g>for</str<strong>on</strong>g> 48 hours to higher dose (10%GS). To assess the effects <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

exposure under the different experimental c<strong>on</strong>diti<strong>on</strong>s, we analyzed both the cytotoxic and<br />

oxidative/glycoxidative effects <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular damage.<br />

No effects <strong>on</strong> cell viability was evidenced in either experimental c<strong>on</strong>diti<strong>on</strong>. During chr<strong>on</strong>ic treatment,<br />

glycative markers (free and bound pentosidine) increased significantly in intra- and extra-cellular<br />

envir<strong>on</strong>ments, but the producti<strong>on</strong> and release <str<strong>on</strong>g>of</str<strong>on</strong>g> thiobarbituric acid reactive substances (TBARs), as index <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

lipid peroxidati<strong>on</strong>, underwent a time-dependent decrease. Exposure to 10% GS, evidenced that glycative<br />

markers rose further while TBARs elicited a cellular defence against oxidative stress. N<strong>on</strong>-adapted cultures<br />

showed an accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> AGEs, a marked oxidative stress and a loss <str<strong>on</strong>g>of</str<strong>on</strong>g> viability. During 10% GS<br />

exposure, GSH levels in adapted cultures remained c<strong>on</strong>stant, as did GSSG/GSH ratio, while n<strong>on</strong>-adapted<br />

cells showed a markedly increased redox ratio. A c<strong>on</strong>stant increase <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 mRNA was observed in all<br />

experimental c<strong>on</strong>diti<strong>on</strong>s. On the c<strong>on</strong>trary HSP70 protein expressi<strong>on</strong> became undetectable <str<strong>on</strong>g>for</str<strong>on</strong>g> l<strong>on</strong>ger<br />

exposure time; this could be due to the direct involvement <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 in the refolding <str<strong>on</strong>g>of</str<strong>on</strong>g> damaged proteins.<br />

The increased levels <str<strong>on</strong>g>of</str<strong>on</strong>g> Cu/Zn SOD protein and mRNA expressi<strong>on</strong>s observed during chr<strong>on</strong>ic exposure, was<br />

followed by a marked decrease after subsequent 48 hours 10%GS exposure. MnSOD protein and mRNA<br />

expressi<strong>on</strong>s resulted not affected in all experimental c<strong>on</strong>diti<strong>on</strong>s. Our findings suggest an adaptive resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

macrophages to subtoxic doses <str<strong>on</strong>g>of</str<strong>on</strong>g> AGE, which could c<strong>on</strong>stitute an important factor in the spread <str<strong>on</strong>g>of</str<strong>on</strong>g> damage<br />

to other cellular types during aging.<br />

6E_04_P<br />

(poster secti<strong>on</strong> B1, poster board #199, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

METABOLIC AND CARDIOVASCULAR ALTERATIONS INDUCED BY CHRONIC<br />

SOCIAL STRESS IN NORMOTENSIVE AND HYPERTENSIVE RATS<br />

I. Bernatova 1 , A. Puzserova 1 , M. Barancik 2 , M. Ivanova 2 , Z. Csizmadiova 1,3 , M. Zeman 3<br />

1Inst Norm Pathol Physiol, 2 Inst Heart Res, Slovak Acad Sci; 3 Dept <str<strong>on</strong>g>of</str<strong>on</strong>g> Animal Physiol and Ethology,<br />

Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Nat Sci, Comenius Univ, Bratislava, Slovak Republic<br />

e-mail: Iveta.Bernatova@savba.sk<br />

This study investigated the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic crowding stress <strong>on</strong> metabolism and cardiovascular functi<strong>on</strong>s in<br />

normotensive WKY and sp<strong>on</strong>taneously hypertensive (SHR) rats. Adult males were exposed to 8-week<br />

crowding (5 rats/cage, 200 cm 2 /rat). C<strong>on</strong>trols were kept 4 rats/cage (480 cm 2 /rat). Blood pressure (BP) was<br />

determined by tail-cuff method. Basal BP <str<strong>on</strong>g>of</str<strong>on</strong>g> WKY and SHR was 110+/-2 and 180+/-5 mm Hg, respectively<br />

and crowding significantly elevated BP <strong>on</strong>ly in SHR. No phenotype-related alterati<strong>on</strong>s in plasma<br />

corticoster<strong>on</strong>e (pCort) were observed in c<strong>on</strong>trols. Crowding significantly elevated pCort and reduced blood<br />

glucose levels, similarly in both phenotypes. Crowding increased activati<strong>on</strong> (Thr202/Tyr204 phosphorylati<strong>on</strong>)<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> extracellular-signal regulated protein kinases in the heart <str<strong>on</strong>g>of</str<strong>on</strong>g> both WKY and SHR, suggesting involvement<br />

and similar role <str<strong>on</strong>g>of</str<strong>on</strong>g> this pathway in resp<strong>on</strong>ses to crowding stress in both phenotypes. However, phenotyperelated<br />

differences were observed in cardiovascular resp<strong>on</strong>ses to stress. Crowding reduced NO producti<strong>on</strong> in<br />

the heart and aorta <str<strong>on</strong>g>of</str<strong>on</strong>g> SHR and reduced acetylcholine-induced vasorelaxati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the femoral artery. On the<br />

other hand, crowding elevated NO producti<strong>on</strong> in the aorta <str<strong>on</strong>g>of</str<strong>on</strong>g> WKY, without changes in the heart, and<br />

improved vasorelaxati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the femoral artery. In c<strong>on</strong>clusi<strong>on</strong>, chr<strong>on</strong>ic crowding stress evoked similar<br />

metabolic changes in both phenotypes <str<strong>on</strong>g>of</str<strong>on</strong>g> rats while NO producti<strong>on</strong> and endothelium-dependent<br />

341


23-26 August 2007,<br />

Budapest, Hungary<br />

vasorelaxati<strong>on</strong> were affected differently in WKY and SHR. This suggest impaired cardiovascular adaptati<strong>on</strong><br />

to stress in SHR.<br />

6E_05_P<br />

(poster secti<strong>on</strong> B1, poster board #200, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECTS OF CHRONIC EXERCISE AND ACUTE STRESS ON CARDIOVASCULAR<br />

PERFORMANCE IN RATS<br />

Chauying J. Jen*, Yuan-Chang Hsu, Hsiun-ing Chen<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Physiology and Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Basic Medical Sciences, College <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine,<br />

Nati<strong>on</strong>al Cheng Kung University, Tainan 70101, Taiwan<br />

e-mail: jen@mail.ncku.edu.tw<br />

It is well accepted that the cardiovascular per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance is improved by chr<strong>on</strong>ic exercise and hampered by<br />

stress. However, the detailed mechanisms remain unclear. Wistar rats were divided into sedentary c<strong>on</strong>trol and<br />

chr<strong>on</strong>ic exercise groups; the latter were trained <strong>on</strong> a treadmill <str<strong>on</strong>g>for</str<strong>on</strong>g> 8 weeks. Rats from either group were<br />

exposed to restraint stress and their heart rate (HR), mean blood pressure (mBP) and pulse pressure (PP)<br />

were m<strong>on</strong>itored by telemetric measurements under c<strong>on</strong>scious c<strong>on</strong>diti<strong>on</strong>s. Moreover, we quantified the<br />

acetylcholine-evoked endothelial calcium signaling and the heat shock protein 72 (HSP72) expressi<strong>on</strong> in<br />

isolated aortic segments. In general, acute stress induced rapid elevati<strong>on</strong>s in HR, mBP and PP, which were<br />

partially recovered during 1 hr <str<strong>on</strong>g>of</str<strong>on</strong>g> stress period. However, these parameters did not fully return to resting<br />

levels even after 1 hr <str<strong>on</strong>g>of</str<strong>on</strong>g> post-stress resting. In additi<strong>on</strong>, stress also suppressed endothelial calcium signaling<br />

and caused elevated HSP72 expressi<strong>on</strong> in aortas. Comparing with sedentary c<strong>on</strong>trols, exercise-trained rats<br />

showed i) enhanced PP al<strong>on</strong>g with reduced HR and mBP at rest and during stress, ii) faster recovery <str<strong>on</strong>g>of</str<strong>on</strong>g> these<br />

parameters during post-stress period, iii) faster adaptati<strong>on</strong> due to repeated stress, iv) minimal stresssuppressed<br />

endothelial calcium signaling, and v) augmented stress-induced HSP72 expressi<strong>on</strong>. Taken<br />

together, exercise training made the cardiovascular system more resistant to stress partially by reducing the<br />

stress-induced adverse effects and enhancing the stress-induced protective effects in vessels.<br />

6E_06_P<br />

(poster secti<strong>on</strong> B1, poster board #201, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECTS OF ALFA1-ACID GLYCOPROTEIN ON FATTY ACIDS OVERLOAD IN<br />

PERIPHERAL BLOOD MONONUCLEAR CELLS FROM DAIRY COWS<br />

N. Lacetera 1 , F. Ceciliani 2 , G. Kuzminsky 1 , C. Lecchi 2 , U. Bernabucci 1 , A. Nard<strong>on</strong>e 1 , P. Sartorelli 2<br />

1 Dipartimento di Produzi<strong>on</strong>i Animali, Università della Tuscia, Viterbo, Italy<br />

2 Dipartimento di Patologia Animale, Igiene e Sanità Pubblica Veterinaria, Università di Milano, Milano, Italy<br />

Fatty acids (FA) overload can alter leukocyte functi<strong>on</strong>s in dairy ruminants. Alfa1-acid glycoprotein (AGP)<br />

may functi<strong>on</strong> as a binding protein <str<strong>on</strong>g>for</str<strong>on</strong>g> hydrophobic molecules. Periparturient cows <str<strong>on</strong>g>of</str<strong>on</strong>g>ten experience a<br />

c<strong>on</strong>comitant increase <str<strong>on</strong>g>of</str<strong>on</strong>g> plasma FA and AGP. This study was aimed to assess proliferative resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

bovine peripheral blood m<strong>on</strong><strong>on</strong>uclear cells (PBMC) cultured in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> various c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> FA<br />

and AGP. Nine pregnant Holstein heifers were utilized as blood d<strong>on</strong>ors. PBMC were incubated with a<br />

mixture <str<strong>on</strong>g>of</str<strong>on</strong>g> FA reflecting compositi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> bovine plasma FA, and at c<strong>on</strong>centrati<strong>on</strong>s mimicking those <str<strong>on</strong>g>of</str<strong>on</strong>g> cows<br />

342


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

undergoing various degree <str<strong>on</strong>g>of</str<strong>on</strong>g> lipomobilizati<strong>on</strong> (500, 250, 125 e 62.5 µmol/l). AGP was purified from bovine<br />

plasma and added to PBMC culture at c<strong>on</strong>centrati<strong>on</strong>s, which reflected those found in physiological<br />

c<strong>on</strong>diti<strong>on</strong>s or during the systemic reacti<strong>on</strong> to inflammati<strong>on</strong> (0.3 and 1.5 mg/ml, respectively). C<strong>on</strong>canavalin A<br />

and pokeweed mitogen (PWM) were utilized as mitogens. PBMC proliferati<strong>on</strong> was assessed by measuring the<br />

5-bromo-2'-deoxyuridine incorporated during DNA synthesis. The highest c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> FA significantly<br />

impaired proliferative resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> PBMC to both mitogens. In PWM-stimulated PBMC <strong>on</strong>ly, the additi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

1.5 mg/ml AGP attenuated to a significant extent the negative effects <str<strong>on</strong>g>of</str<strong>on</strong>g> FA overload <strong>on</strong> PBMC<br />

proliferati<strong>on</strong>. Present results suggest that under c<strong>on</strong>diti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> intense lipomobilizati<strong>on</strong>, levels <str<strong>on</strong>g>of</str<strong>on</strong>g> AGP<br />

mimicking those observed during the systemic reacti<strong>on</strong> to inflammati<strong>on</strong> may at least partially attenuate the<br />

negative effects <str<strong>on</strong>g>of</str<strong>on</strong>g> FA overload <strong>on</strong> leukocyte functi<strong>on</strong>s.<br />

6E_07_P<br />

(poster secti<strong>on</strong> B1, poster board #202, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

TREATMENT WITH SODIUM PENTOSAN POLIYSULFATE AMELIORATES<br />

MORPHOLOGICAL RENAL ALTERATION AND ALBUMINURIA IN<br />

DIABETIC RATS<br />

1Y. Mathis<strong>on</strong>, 2 H. J. Finol, 1 Z. Quero, 2 S. Mayora<br />

1Schools <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine J.M. Vargas;<br />

2Center <str<strong>on</strong>g>of</str<strong>on</strong>g> Electr<strong>on</strong> Microscopy, School <str<strong>on</strong>g>of</str<strong>on</strong>g> Science, Central University <str<strong>on</strong>g>of</str<strong>on</strong>g> Venezuela<br />

e-mail: yaira57@yahoo.com<br />

Decrease levels <str<strong>on</strong>g>of</str<strong>on</strong>g> glycosaminoglycans (GAGs) have been observed in kidney and other organs, in human<br />

and animal models <str<strong>on</strong>g>of</str<strong>on</strong>g> diabetes. L<strong>on</strong>g term administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> heparins and other glycosam<strong>on</strong>oglycans have<br />

dem<strong>on</strong>strated a beneficial effect <strong>on</strong> morphological and functi<strong>on</strong>al renal abnormalities in diabetic rats. We<br />

assessed the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> sodium pentosan polysulfate (SPP), a semi-synthetic orally active glycosaminoglycan<br />

with low anticoagulant activity, <strong>on</strong> renal involvement in streptozotocin diabetic rats. Diabetes was induced in<br />

male Sprague-Dawley rats by i.v. administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> streptozotocin (STZ). Animals were randomly allocated in<br />

three groups: C=c<strong>on</strong>trol, STZ and STZ+SPP= pretreated with SPP (15mg/kg, s.c.). After three m<strong>on</strong>ths <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

follow-up, blood and 24h-urine samples were obtained and than the animals were sacrificed and the kidney<br />

microdissected <str<strong>on</strong>g>for</str<strong>on</strong>g> morphometric analysis. Urinary albumin excreti<strong>on</strong> was markedly increased in untreated<br />

diabetic rats (C= 0,26± 0,03 vs. STZ= 7,75±1,8 mg/24h) and SPP treatment partially prevented the albumin<br />

rise (3,7±0,7 mg/24h), without affecting the metabolic c<strong>on</strong>trol HbA1c (C=3,6±1,7; STZ=8,82±0,47;<br />

STZ+PPSNa= 8,63 ± 0,54). Electr<strong>on</strong> microscope observati<strong>on</strong> revealed typical renal lesi<strong>on</strong>s described in<br />

experimental diabetes (STZ group). SPP administrati<strong>on</strong> prevent the glomerular and tubular basement<br />

membrane thickening and the lost <str<strong>on</strong>g>of</str<strong>on</strong>g> cytoarchitecture induced by experimental diabetes. Our data<br />

dem<strong>on</strong>strated that l<strong>on</strong>g-term administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> SPP have a favorable effect <strong>on</strong> morphological and functi<strong>on</strong>al<br />

abnormalities in kidney <str<strong>on</strong>g>of</str<strong>on</strong>g> diabetic rats and suggests a potential therapeutic use <str<strong>on</strong>g>of</str<strong>on</strong>g> this compound.<br />

343


23-26 August 2007,<br />

Budapest, Hungary<br />

344<br />

6E_09_P<br />

(poster secti<strong>on</strong> B1, poster board #203, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECTS OF FAT LOSS STRESS IN FAT CELLS<br />

K. T. Tan, Y. H. Lee<br />

Academia Sinica, Inst. Mol. Biol., Taipei 115, Taiwan<br />

Recent years, obesity and the ensuing metabolic disorders such as hypertensi<strong>on</strong>, type 2 diabetes mellitus have<br />

become the severe healthcare problem in industrial nati<strong>on</strong>s. To cope with this problem, ef<str<strong>on</strong>g>for</str<strong>on</strong>g>ts such as<br />

behavior therapy, surgical interventi<strong>on</strong> and pharmacologic treatment have been applied. Although weighreducing<br />

drugs, due to their fast effect, tend to be the most popular choice <str<strong>on</strong>g>for</str<strong>on</strong>g> people wishing to shed a few<br />

pounds <str<strong>on</strong>g>of</str<strong>on</strong>g> their body weight, it was found that there is a high frequency <str<strong>on</strong>g>of</str<strong>on</strong>g> weight regain and/or the loss <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

their effects <strong>on</strong> reducing weight. In fact, weight recurrence occurs frequently with all kinds <str<strong>on</strong>g>of</str<strong>on</strong>g> weightreducti<strong>on</strong><br />

programs, especially with those that produce quick effects. Our evidences show that this<br />

phenomen<strong>on</strong> may be due to the cellular and physiological rescuing-resp<strong>on</strong>ses triggered by stress from the<br />

sudden loss <str<strong>on</strong>g>of</str<strong>on</strong>g> body weight/fat. We have used both cell and animal systems to study the slim-fast stress and<br />

its effects, and our findings indicate that the stressed fat cells are more active than those unstressed in gene<br />

expressi<strong>on</strong>, glucose uptake, protein synthesis and energy oxidati<strong>on</strong>, etc. In simple words, the stressed cells<br />

have become more viable. Accordingly, this increased viability might <str<strong>on</strong>g>for</str<strong>on</strong>g>m a driving <str<strong>on</strong>g>for</str<strong>on</strong>g>ce <str<strong>on</strong>g>for</str<strong>on</strong>g> cells to recover,<br />

even to exceed, rapidly their status be<str<strong>on</strong>g>for</str<strong>on</strong>g>e stress. The molecular mechanism underlying this rapid recovery<br />

was investigated, and the results will be discussed.<br />

6E_10_P<br />

(poster secti<strong>on</strong> B1, poster board #204, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SPOTLIGHT ON ETHNOMEDICINE: USABILITY OF GYMNEMA MONTANUM IN<br />

THE TREATMENT OF DIABETES<br />

Kunga Mohan Ramkumar*, Palanisamy Rajaguru<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biotechnology, Bharathidasan Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Technology<br />

Bharathidasan University, Tiruchirappalli – 620 024, Tamilnadu, India<br />

Oxidative stress caused by free radicals has become an area <str<strong>on</strong>g>of</str<strong>on</strong>g> interest in understanding the process <str<strong>on</strong>g>of</str<strong>on</strong>g> human<br />

diseases. Diabetes mellitus is <strong>on</strong>e such disease and it is estimated that the number <str<strong>on</strong>g>of</str<strong>on</strong>g> diabetic patients will<br />

c<strong>on</strong>tinue to increase in the future. The elevated levels <str<strong>on</strong>g>of</str<strong>on</strong>g> blood glucose in diabetes produce oxygen free<br />

radicals that cause membrane damage due to peroxidati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> membrane lipids and protein glycati<strong>on</strong>.<br />

Indigenous natural resources such as plants/plant products are the potential sources <str<strong>on</strong>g>of</str<strong>on</strong>g> chemicals <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

effective medicati<strong>on</strong>. The antioxidant treatment suppressed the apoptosis in β-cells <str<strong>on</strong>g>of</str<strong>on</strong>g> pancreas without<br />

changing the rate <str<strong>on</strong>g>of</str<strong>on</strong>g> β-cells proliferati<strong>on</strong>. Recently, an intensive search <str<strong>on</strong>g>for</str<strong>on</strong>g> novel types <str<strong>on</strong>g>of</str<strong>on</strong>g> antioxidants has<br />

been carried out from numerous plant materials. Gymnema m<strong>on</strong>tanum Hook, bel<strong>on</strong>ging to the family<br />

Asclepiadaceae, is an endemic plant species <str<strong>on</strong>g>of</str<strong>on</strong>g> India found mainly in Western Ghats. The present<br />

investigati<strong>on</strong> was carried out to evaluate the protective effect <str<strong>on</strong>g>of</str<strong>on</strong>g> Gymnema m<strong>on</strong>tanum leaves <strong>on</strong> red blood cells<br />

(RBC) membrane lipid peroxidati<strong>on</strong> in diabetic rats. Ethanol extract <str<strong>on</strong>g>of</str<strong>on</strong>g> G. m<strong>on</strong>tanum leaves (GLEt) was<br />

administered orally to alloxan induced diabetic rats and the effects <strong>on</strong> blood glucose, plasma Insulin, lipid<br />

peroxidati<strong>on</strong> markers TBARS, hydroperoxides in plasma and enzymic antioxidants Superoxide dismutase<br />

(SOD), catalase (CAT) and glutathi<strong>on</strong>e peroxidase (GPx) activities in erythrocytes were studied.<br />

Administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> GLEt at doses <str<strong>on</strong>g>of</str<strong>on</strong>g> 50, 100 and 200 mg/kg body weight <str<strong>on</strong>g>for</str<strong>on</strong>g> three weeks suppressed the


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

elevated blood glucose and lipid peroxidati<strong>on</strong> in diabetic rats. GLEt at 200 mg/kg body weight was found to<br />

be comparable to glibenclamide, a reference drug. Further, erythrocytes from the extract treated group rats<br />

were found to be more resistant to H 2 O 2 induced peroxidati<strong>on</strong> than the diabetic animals. In additi<strong>on</strong>, we<br />

studied the protective effect <str<strong>on</strong>g>of</str<strong>on</strong>g> GLEt against oxidative DNA damage caused by H 2 O 2 in human peripheral<br />

blood lymphocytes in vitro using the single cell gel electrophoresis. In order to throw light <strong>on</strong> the chemical<br />

nature <str<strong>on</strong>g>of</str<strong>on</strong>g> the active comp<strong>on</strong>ents, the extract was subjected to GC-MS analysis. We suggest that G. m<strong>on</strong>tanum<br />

may be useful <str<strong>on</strong>g>for</str<strong>on</strong>g> the c<strong>on</strong>trol, management and preventi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> oxidative stress associated with diabetes.<br />

6E_11_P<br />

(poster secti<strong>on</strong> B1, poster board #205, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE ROLE OF A SYMPATHETIC NEUROTRANSMITTER, NEUROPEPTIDE Y, IN<br />

STRESS AND STRESS-RELATED DISORDERS<br />

Joanna Kittinska<br />

Georgetown University, 3900 Reservoir Rd. NW, 20057, Washingt<strong>on</strong>, D.C., USA<br />

e-mail: zzukow01@georgetown.edu<br />

Stress has many different meanings in various fields <str<strong>on</strong>g>of</str<strong>on</strong>g> science. It is c<strong>on</strong>sidered a matter <str<strong>on</strong>g>of</str<strong>on</strong>g> the mind, the<br />

place where percepti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ‘disturbance <str<strong>on</strong>g>of</str<strong>on</strong>g> homeostasis’ is born, leading to activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> hypothalamicpituitary-adrenocortical<br />

axis and sympathetic nerves. However, the patterns <str<strong>on</strong>g>of</str<strong>on</strong>g> stress-induced resp<strong>on</strong>ses are<br />

highly variable, and depend also <strong>on</strong> factors outside the brain. Cardiovascular and metabolic changes during<br />

stress vary with durati<strong>on</strong> and intensity <str<strong>on</strong>g>of</str<strong>on</strong>g> stress, and this is reflected in a differential pattern <str<strong>on</strong>g>of</str<strong>on</strong>g> the release <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

sympathetic neurotransmitters, norepinephrine (NE), purines and neuropeptide Y (NPY). Mild and acute<br />

stress releases NE, whereas more intense and/or prol<strong>on</strong>ged stress is required <str<strong>on</strong>g>for</str<strong>on</strong>g> NPY. Prol<strong>on</strong>ged stressinduced<br />

sympathetic activati<strong>on</strong> also raises platelet NPY levels; hence platelet NPY may be a particularly good<br />

index <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic stress. NPY’s acti<strong>on</strong>s are both similar and dissimilar to those <str<strong>on</strong>g>of</str<strong>on</strong>g> NE. Like NE, NPY is a<br />

vasoc<strong>on</strong>strictor acting directly via its Y1 receptor (R), and indirectly, by potentiating NE-induced<br />

c<strong>on</strong>stricti<strong>on</strong>. However, unlike NE, NPY also stimulates growth <str<strong>on</strong>g>of</str<strong>on</strong>g> variety <str<strong>on</strong>g>of</str<strong>on</strong>g> cells: neur<strong>on</strong>al cell precursors<br />

via Y1Rs, vascular smooth muscle cells via Y1/Y5 Rs, endothelial cells (angiogenesis) via Y2/Y5R and<br />

predipocytes (adipogenesis) via Y2Rs. In a setting <str<strong>on</strong>g>of</str<strong>on</strong>g> vascular injury, stress up-regulates NPY-Y1R and<br />

augments atherosclerosis. C<strong>on</strong>versely, stress appears to be inhibitory <str<strong>on</strong>g>for</str<strong>on</strong>g> the NPY-Y2R angiogenic system in<br />

ischemic tissues but stimulatory <str<strong>on</strong>g>for</str<strong>on</strong>g> fat tissue angiogenesis and adipogenesis leading to obesity. Thus, stressinduced<br />

high NPY levels are risk factors <str<strong>on</strong>g>for</str<strong>on</strong>g> cardiovascular and metabolic disorders and any other<br />

angiogenesis-dependent disorders such as tumors and retinopathy.<br />

345


23-26 August 2007,<br />

Budapest, Hungary<br />

346<br />

6F. SEPSIS, ENDOTOXINS, STRESS AND ORGAN DYSFUNCTION<br />

(ROBERT S. MUNFORD, JEAN-MARC CAVAILLON)<br />

6F_01_S<br />

LEUKOCYTE REPROGRAMMING AFTER ENDOTOXIN ENCOUNTER<br />

Jean-Marc Cavaill<strong>on</strong><br />

Unit Cytokines & Inflammati<strong>on</strong>, Institut Pasteur, Paris<br />

e-mail: jmcavail@pasteur.fr<br />

Stressfull situati<strong>on</strong> associated with sepsis and n<strong>on</strong>-infectious systemic inflammatory resp<strong>on</strong>se syndrome<br />

(SIRS) such as trauma, surgery, haemorrhage, or ischemia/reperfusi<strong>on</strong>, is associated with a pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ound<br />

alterati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> immune status that could explain the increased sensisitivity <str<strong>on</strong>g>of</str<strong>on</strong>g> the patients to nosocomial<br />

infecti<strong>on</strong>s. Endotoxin (lipopolysaccharide, LPS) is <str<strong>on</strong>g>of</str<strong>on</strong>g>ten found in plasma <str<strong>on</strong>g>of</str<strong>on</strong>g> sepsis patients, and endotoxin<br />

translocati<strong>on</strong> from the gut frequently occurs in SIRS patients. During sepsis and SIRS, pro-inflammatory<br />

cytokines and inflammatory mediators c<strong>on</strong>tribute in synergy to tissue injury, organ dysfuncti<strong>on</strong>, and possibly<br />

to lethality. To dampen this overzealous process, a counter regulatory loop exists, involving antiinflammatory<br />

cytokines and neuromediators that can also alter the immune status. In sepsis and SIRS<br />

patients, a reduced ex vivo pro-inflammatory cytokine producti<strong>on</strong>, particularly tumor necrosis factor (TNF), in<br />

resp<strong>on</strong>se to LPS and other Toll-like receptor (TLR) ag<strong>on</strong>ists has been reported. However, cells remain<br />

resp<strong>on</strong>sive to whole bacteria, and their capacity to produce anti-inflammatory cytokine is maintained or even<br />

enhanced. Thus the terms "leukocyte reprogramming" well define the process. Modificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> intracellular<br />

signaling are presently deciphered in patients' leukocytes and some aspects recall what is known about<br />

"endotoxin tolerance". The reduced capacity <str<strong>on</strong>g>of</str<strong>on</strong>g> circulating m<strong>on</strong>ocytes to produce TNF and other cytokines<br />

can be mimicked in vitro by a pre-treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> m<strong>on</strong>ocytes or macrophages with lipolysaccharide (LPS). This is<br />

not a specific phenomen<strong>on</strong> and it can be induced by other agents or events. Cross-tolerance between LPS,<br />

TLR2, 4, 5, or 9 specific ligands, and TNF has been reported. It is possible that cross-tolerance is induced by<br />

microbial and endogenous (alarmins) signals <str<strong>on</strong>g>of</str<strong>on</strong>g> danger.<br />

6F_02_S<br />

ACYLOXYACYL HYDROLASE SHORTENS THE DURATION OF POST-INFECTION<br />

IMMUNOSUPPRESSION<br />

Mingfang Lu, Robert S. Mun<str<strong>on</strong>g>for</str<strong>on</strong>g>d<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Texas Southwestern Medical Center, Dallas, Texas, USA 75390-9113<br />

e-mail: Mingfang.Lu@utsouthwestern.edu, Robert.mun<str<strong>on</strong>g>for</str<strong>on</strong>g>d@utsouthwestern.edu<br />

Serious tissue infecti<strong>on</strong>s may be followed by a period during which the host animal is much more susceptible<br />

to infecti<strong>on</strong> with the same or other microbes. One experimental model <str<strong>on</strong>g>for</str<strong>on</strong>g> this clinical phenomen<strong>on</strong> is<br />

endotoxin tolerance or reprogramming. For several days after receiving a low parenteral dose <str<strong>on</strong>g>of</str<strong>on</strong>g> Gramnegative<br />

bacteria or their cell wall lipopolysaccharide (LPS, endotoxin), many <str<strong>on</strong>g>of</str<strong>on</strong>g> an animals’ reacti<strong>on</strong>s to LPS<br />

and other microbial ag<strong>on</strong>ists are “reprogrammed” so that proinflammatory resp<strong>on</strong>ses are reduced while antiinflammatory<br />

<strong>on</strong>es are maintained or increased. Whereas the adaptive functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> this stress reacti<strong>on</strong> may be<br />

to prevent harmful inflammati<strong>on</strong>-induced damage to the host, it is <str<strong>on</strong>g>of</str<strong>on</strong>g>ten also immunosuppressive. We found<br />

that deacylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> LPS by a host lipase, acyloxyacyl hydrolase (AOAH), is required to terminate LPS-induced<br />

immunosuppressi<strong>on</strong> in mice. Whereas wildtype mice recover within 10 days after receiving 10 µg LPS


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

intraperit<strong>on</strong>eally, mice that lack AOAH remain immunosuppressed <str<strong>on</strong>g>for</str<strong>on</strong>g> at least <strong>on</strong>e m<strong>on</strong>th. The perit<strong>on</strong>eal<br />

cells in Aoah -/- mice retain intact LPS <str<strong>on</strong>g>for</str<strong>on</strong>g> much <str<strong>on</strong>g>of</str<strong>on</strong>g> this time, their proinflammatory resp<strong>on</strong>ses to LPS (a TLR4<br />

ag<strong>on</strong>ist) and Micrococcus luteus (a TLR2 ag<strong>on</strong>ist) are blunted, and their ability to kill Gram-negative bacteria is<br />

impaired. By extinguishing the LPS signal, AOAH seems to help the body “re-load” its defensive<br />

armamentarium to fight another infecti<strong>on</strong>.<br />

6F_03_S<br />

REPROGRAMMING OF MACROPHAGES BY ENDOTOXIN TOLERANCE<br />

F. Ulrich Schade, Katja Butterbach, Daniela Plitzko<br />

Surgical <str<strong>on</strong>g>Research</str<strong>on</strong>g> - Trauma Surgery, University Hospital Essen, D-45122 Essen, Germany<br />

e-mail: ulrich.schade@uni-due.de<br />

The term "Endotoxin Tolerance" (ET) has been introduced <str<strong>on</strong>g>for</str<strong>on</strong>g> a state <str<strong>on</strong>g>of</str<strong>on</strong>g> low resp<strong>on</strong>siveness <str<strong>on</strong>g>of</str<strong>on</strong>g> experimental<br />

animals or humans treated with a low dose <str<strong>on</strong>g>of</str<strong>on</strong>g> lipopolysaccharide (LPS, endotoxin). ET has attracted a lot <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

attenti<strong>on</strong> since it was shown that cross tolerance exists to other bacterial products such as lipoteichoic acid or<br />

lipopeptides. Furthermore, it turned out that ET protected against bacterial infecti<strong>on</strong>s - Gram-negative, as<br />

well as Gram-positive - and ischemia-reperfusi<strong>on</strong> injury. It, there<str<strong>on</strong>g>for</str<strong>on</strong>g>e, is <str<strong>on</strong>g>of</str<strong>on</strong>g> potential clinical interest to get<br />

insight into the cellular and molecular basis <str<strong>on</strong>g>of</str<strong>on</strong>g> ET. To this end, we studied different cellular comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the perit<strong>on</strong>eal cell populati<strong>on</strong>s and Kupffer cells <str<strong>on</strong>g>of</str<strong>on</strong>g> endotoxin tolerant and normal mice with regard to gene<br />

expressi<strong>on</strong>, intracellular signal transducti<strong>on</strong> and regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cytokine producti<strong>on</strong>. Corresp<strong>on</strong>ding results will<br />

be presented. HSP 70 is am<strong>on</strong>g the genes that are significantly upregulated in perit<strong>on</strong>eal cells <str<strong>on</strong>g>of</str<strong>on</strong>g> ET mice<br />

compared to normal mice.<br />

6F_04_S<br />

IMMUNO-NEURO-ENDOCRINE ADAPTATION IN CRITICAL ILLNESS<br />

I.Vermes, A. Beishuizen, A. B. J.Groeneveld<br />

Medical Spectrum Twente, Enschede & Free University Medical <str<strong>on</strong>g>Centre</str<strong>on</strong>g> Amsterdam, The Netherlands<br />

e-mail: i.vermes@home.nl<br />

Critical illness is a severe stress stimulus that disturbs the milieu interior and induces homeostatic resp<strong>on</strong>ses<br />

specific to the stimulus and generalized resp<strong>on</strong>ses when the disturbances are prol<strong>on</strong>ged and severe. The<br />

immune-neuroendocrine resp<strong>on</strong>se during critical illness is a dynamic process, differing between the acute and<br />

chr<strong>on</strong>ic phases. This acute phase lasts typically from about a few hours to several days, depending <strong>on</strong> the<br />

severity <str<strong>on</strong>g>of</str<strong>on</strong>g> the illness and is characterized by an appropriate horm<strong>on</strong>al reacti<strong>on</strong>: the mobilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> fuel stores<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the organism, together with apparent restrains <strong>on</strong> their utilizati<strong>on</strong>. During the acute phase <str<strong>on</strong>g>of</str<strong>on</strong>g> critical illness<br />

the secretory activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the pituitary gland is stimulated whereas anabolic target organ horm<strong>on</strong>es are<br />

inactivated. Due to the development <str<strong>on</strong>g>of</str<strong>on</strong>g> intensive care medicine patients who would previously have died<br />

during the acute phase nowadays survive and enter the chr<strong>on</strong>ic or prol<strong>on</strong>ged phase <str<strong>on</strong>g>of</str<strong>on</strong>g> the critical illness. This<br />

prol<strong>on</strong>ged phase lasts <str<strong>on</strong>g>for</str<strong>on</strong>g> days and is characterized by an inappropriate horm<strong>on</strong>al resp<strong>on</strong>se, resulting in a<br />

chr<strong>on</strong>ic increase in metabolic rate and a breakdown <str<strong>on</strong>g>of</str<strong>on</strong>g> body tissue. The secretory activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the pituitary<br />

gland is uni<str<strong>on</strong>g>for</str<strong>on</strong>g>mly inhibited in relati<strong>on</strong> to reduced levels <str<strong>on</strong>g>of</str<strong>on</strong>g> target organ horm<strong>on</strong>es. These horm<strong>on</strong>al changes<br />

suggest an imbalance between immunosuppressive and immunostimulatory horm<strong>on</strong>es which might be the<br />

cause <str<strong>on</strong>g>of</str<strong>on</strong>g> the increased susceptibility to infectious complicati<strong>on</strong>s during the chr<strong>on</strong>ic phase <str<strong>on</strong>g>of</str<strong>on</strong>g> severe illness.<br />

The suppressed pituitary activity allows the respective target organ horm<strong>on</strong>es to decrease, resulting in a<br />

347


23-26 August 2007,<br />

Budapest, Hungary<br />

restored balance between the catabolic and anabolic horm<strong>on</strong>al resp<strong>on</strong>ses. This recovery phase is<br />

characterized by restored sensitivity <str<strong>on</strong>g>of</str<strong>on</strong>g> the pituitary gland to reduced feedback c<strong>on</strong>trol. The distincti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

acute and prol<strong>on</strong>ged critical illness as different entities with regard to the immuno-neuroendocrine adaptati<strong>on</strong><br />

may be helpful in further understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> the pathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> critical illness and the targeting <str<strong>on</strong>g>of</str<strong>on</strong>g> therapeutic<br />

interventi<strong>on</strong>.<br />

6F_05_S<br />

IN VIVO IMAGING OF THE EFFECT OF BACTERIAL ENDOTOXINS ON<br />

ARTERIAL ENDOTHELIAL CELLS: MOLECULAR IMAGING OF HEAT SHOCK<br />

PROTEIN 60 EXPRESSION<br />

Marius C. Wick 1 , Christina Mayerl 2 , Georg Dobrozemsky 3 , Roland Haubner 3 , Hermann Dietrich 4 ,<br />

Werner Jaschke 1 , Georg Wick 2 .<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Radiology, 2 Divisi<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> Experimental Pathophysiology and Immunology, Biocenter,<br />

3Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Nuclear Medicine, 4 Central Animal Laboratory Facilities,<br />

Innsbruck Medical University, Austria<br />

Background. Bacterial endotoxins (LPS) are known to act as stress factors <str<strong>on</strong>g>for</str<strong>on</strong>g> endothelial cells.<br />

Aims. As a pro<str<strong>on</strong>g>of</str<strong>on</strong>g> <str<strong>on</strong>g>of</str<strong>on</strong>g> principle, the present project aimed at developing a novel radiotracer-based n<strong>on</strong>-invasive<br />

molecular imaging method <str<strong>on</strong>g>for</str<strong>on</strong>g> in vivo visualizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> early aortal HSP60-expressi<strong>on</strong> in a<br />

normocholesterolemic rabbit model after inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> endothelial stress.<br />

Material and Methods.<br />

In 14 rabbits, endothelial stress was induced by i.v. injecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> bacterial endotoxin (lipopolysaccharide, LPS<br />

at 10µg/kg) while 8 animals were untreated c<strong>on</strong>trols. N<strong>on</strong>-invasive in vivo molecular imaging was per<str<strong>on</strong>g>for</str<strong>on</strong>g>med<br />

24 hours after intravenous injecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> I-124 radiolabelled m<strong>on</strong>ocl<strong>on</strong>al anti-HSP60 antibodies using<br />

computertomography (CT) and positr<strong>on</strong>-emissi<strong>on</strong>-tomography (PET). In vitro correlati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> in vivo imaging<br />

was d<strong>on</strong>e by en face immunohistochemistry and autoradiography <str<strong>on</strong>g>of</str<strong>on</strong>g> the aorta.<br />

Results.<br />

Compared to c<strong>on</strong>trol animals, quantitative analyses <str<strong>on</strong>g>of</str<strong>on</strong>g> in vivo n<strong>on</strong>-invasive molecular PET-CT images <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

rabbit aortae revealed a significantly increased endothelial binding <str<strong>on</strong>g>of</str<strong>on</strong>g> I-124 anti-HSP60 antibodies up<strong>on</strong><br />

applicati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> LPS as an endothelial stressor, especially at sites <str<strong>on</strong>g>of</str<strong>on</strong>g> aortal branching. This was c<strong>on</strong>firmed by in<br />

vitro anti-HSP60 immunohistochemistry and autoradiography data.<br />

C<strong>on</strong>clusi<strong>on</strong>.<br />

Our first results showed, as a pro<str<strong>on</strong>g>of</str<strong>on</strong>g> <str<strong>on</strong>g>of</str<strong>on</strong>g> principle, that HSP60-expressi<strong>on</strong> in normocholesterolemic rabbits is<br />

significantly increased after the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> endothelial stress and n<strong>on</strong>-invasive in vivo molecular imaging <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

early aortal HSP60-expressi<strong>on</strong> using radiotracer labelled anti-HSP60 antibodies is possible.<br />

348


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

6F_01_P<br />

(poster secti<strong>on</strong> B2, poster board #269, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

VASCULAR HYPOREACTIVITY IS NOT ASSOCIATED WITH MORTALITY IN RATS<br />

WITH ENDOTOXIN STRESS<br />

S. J. Chen 1 , P. L. Chiang 2 , H. L. Liu 2 , R. L. Chen 2 , M. H. Liao 3 , C. C. Wu 2<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Nursing, Kang-Ning Junior College <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Care and Management, Taipei; 2 Institute <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Pharmacology and 3 Graduate Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Sciences, Nati<strong>on</strong>al Defense Medical Center, Taipei; Taiwan<br />

e-mail: sjchen@knjc.edu.tw<br />

Sepsis is the systemic inflammatory stress induced by infecti<strong>on</strong>. In previous studies, the administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

lipopolysaccharide (LPS) to animals produces a sepsis-like syndrome stress, characterized by low blood<br />

pressure and vascular hyporeactivity to vasoc<strong>on</strong>strictor agents (norepinephrine, NE). The vascular<br />

hyporeactivity to vasoc<strong>on</strong>strictor agents, an important characterizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> circulatory failure in septic shock,<br />

occurred at the late phase <str<strong>on</strong>g>of</str<strong>on</strong>g> sepsis. The present study was to compare vascular reactivity between survival<br />

and n<strong>on</strong>-survival rats with endotoxic shock. Endotoxemic stress was induced by intravenous administrati<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> LPS in male Wister rats. We measured resting membrane potential (RMP) and NE-induced tensi<strong>on</strong> in<br />

isolated thoracic aortas by electrophysiology and tensi<strong>on</strong> recording experiments. Our results revealed that<br />

hypotensi<strong>on</strong>, bradycardia, hypoglycemia and hypothermia were found in n<strong>on</strong>-survival septic rats. However,<br />

the RMP and vascular reactivity to NE in aorta were not significant different between n<strong>on</strong>-survival and<br />

survival septic rats. Thus, our results suggest that the mortality in LPS-induced stress was not associated with<br />

vascular hyporeactivity. Indeed, other mechanisms c<strong>on</strong>tributed to the mortality induced by LPS should be<br />

elucidated in future studies.<br />

6F_02_P<br />

(poster secti<strong>on</strong> B2, poster board #270, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SESAMOL PROTECTS AGAINST FERRIC NITRILOTRIACETATE-INDUCED<br />

OXIDATIVE STRESS IN ACUTE RENAL INJURY IN MICE<br />

Bu-Miin Huang 1 , Dur-Z<strong>on</strong>g Hsu 2 , Chang-Hsin Wan 2 , Ming-Yie Liu 2<br />

Departments <str<strong>on</strong>g>of</str<strong>on</strong>g> Cell Biology and Anatomy 1 and Envir<strong>on</strong>mental and Occupati<strong>on</strong>al Health 2 , Nati<strong>on</strong>al Cheng Kung<br />

University Medical College, Tainan, Taiwan<br />

Ferric-nitrilotriacetate (Fe-NTA) has been reported to cause acute nephro-toxicity in animals and in humans.<br />

The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to examine the protective effects <str<strong>on</strong>g>of</str<strong>on</strong>g> sesamol <strong>on</strong> acute renal damage induced by Fe-<br />

NTA in mice. Acute renal injury was induced by giving Fe-NTA to mice <str<strong>on</strong>g>for</str<strong>on</strong>g> 24 h. Renal functi<strong>on</strong> was<br />

assessed by blood biochemistry, creatinine clearance, and histological examinati<strong>on</strong>. Free radicals were<br />

determined using a high-per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance chemiluminescence analyzer. Fe-NTA (4 mg/kg) increased renal lipid<br />

peroxidati<strong>on</strong> and induced acute renal injury. Sesamol (30 mg/kg) attenuated Fe-NTA-induced acute renal<br />

injury, decreased renal lipid peroxidati<strong>on</strong>, reduced the generati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> renal hydroxyl radical and superoxide<br />

ani<strong>on</strong>, and inhibited the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> xanthine oxidase in mice. Thus, sesamol might ameliorate oxidative-stressassociated-acute<br />

renal injury by inhibiting xanthine-oxidase-initiated superoxide ani<strong>on</strong> generati<strong>on</strong>, thereby<br />

reducing hydroxyl radical producti<strong>on</strong>, at least partially, in acutely ir<strong>on</strong>-intoxicated mice.<br />

349


23-26 August 2007,<br />

Budapest, Hungary<br />

350<br />

6F_03_P<br />

(poster secti<strong>on</strong> B2, poster board #271, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

UNDETOXIFIED ENDOTOXINS CAUSING PERSISTENT SEPSIS WITH<br />

PROGRESSIVE PARKINSONISM (A CASE REPORT)<br />

I. Niehaus<br />

Bad Segeberg, Germany, e-mail: Ines_Niehaus@gmx.de<br />

A 22-year old female was c<strong>on</strong>taminated with 10 microgram Salm<strong>on</strong>ella minnesota smooth lipopolysaccharides<br />

(LPS or endotoxin) by an accident in lab in 1995. 21 days later parkins<strong>on</strong>ism started after acute sepsis-like<br />

symptoms (high fever, chills, myalgia, flu-like symptoms). The LPS-induced neur<strong>on</strong>al inflammati<strong>on</strong> (6.6 ng<br />

LPS/ml cerebrospinal fluid measured in 2001) causes polyneuropathy, myocl<strong>on</strong>i, and reduced metabolism <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

dopamine and glucose shown by positr<strong>on</strong> emissi<strong>on</strong> tomographies <str<strong>on</strong>g>of</str<strong>on</strong>g> the brain. The chr<strong>on</strong>ic inflammatory<br />

LPS-induced stress also leads to a multiple chemical sensitivity because <str<strong>on</strong>g>of</str<strong>on</strong>g> the reduced capacity <str<strong>on</strong>g>of</str<strong>on</strong>g> the liver,<br />

which had up taken a part <str<strong>on</strong>g>of</str<strong>on</strong>g> the LPS, as a filter organ. The liver was unable to eliminate the still circulating<br />

LPS in the blood causing a chr<strong>on</strong>ically sub acute sepsis with e. g. tachycardia, orthostatic hypotensi<strong>on</strong>.<br />

Treatment with endotoxin-binding agents like the antibiotic drug <str<strong>on</strong>g>of</str<strong>on</strong>g>loxacin is improving all symptoms<br />

including parkins<strong>on</strong>ism but is unable to cure. After stop taking it the inflammatory reacti<strong>on</strong>s are increasing<br />

again in the whole body because <str<strong>on</strong>g>of</str<strong>on</strong>g> the re-binding <str<strong>on</strong>g>of</str<strong>on</strong>g> LPS to the cells. Acute feverish phases <str<strong>on</strong>g>of</str<strong>on</strong>g> inflammati<strong>on</strong><br />

provoked by all kinds <str<strong>on</strong>g>of</str<strong>on</strong>g> stress (e. g. physical or mental exhausting, sec<strong>on</strong>dary viral or bacterial infecti<strong>on</strong>s) are<br />

marked by a rapid deteriorati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> parkins<strong>on</strong>ism with akinetic crises (muteness, inability to walk),<br />

disorientati<strong>on</strong>, difficulties in hearing and seeing. Glucocorticoids, which are not effective in idiopathic<br />

Parkins<strong>on</strong> ’s disease, are able to stop these crises, which is comparable with some patients suffering from post<br />

encephalitic parkins<strong>on</strong>ism. In rats <strong>on</strong>e single i. p. injecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> LPS is causing a progressive parkins<strong>on</strong>ism<br />

similar to this case report.<br />

6F_04_P<br />

(poster secti<strong>on</strong> B2, poster board #272, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

NON-TOXIC LIPOPOLYSACCHARIDE DERIVATIVES AND EXOGENOUS HSP70<br />

PROTECT AGAINST LETHAL SEPTIC SHOCK AND IMMUNO-METHABOLIC<br />

INJURIES INDUCED BY BACTERIAL ENDOTOXIN AND XENOBIOTICS<br />

Sergey Onikienko 1 , Alexander Zemlyanoi 3 , Irina Guzhova 2 , Boris Margulis 2 , Maxim Vinokurov 4<br />

1 Military Medical Academy, St Petersburg, Russia, <strong>on</strong>ik@nwgsm.ru<br />

2 Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Cytology RAS, St Petersburg, Russia<br />

3 Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Military Medicine, St Petersburg, Russia<br />

4 Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Cell Biophysics RAS, Pushino, Russia<br />

Serratia marcescens endotoxin (LPS) modificati<strong>on</strong> by electr<strong>on</strong>ic beam and CO 2 -laser radiati<strong>on</strong> resulted to<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> its n<strong>on</strong>-toxic derivatives. The parent LPS were modified to 1,5-5,0 kDa oligomers with reduced<br />

toxicity and enhanced immunogenicity. Treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> the mice with n<strong>on</strong>-toxic LPS derivatives significantly<br />

attenuated the lethality, deleterious immune and metabolic injuries induced by parent LPS (1-7LD 50 ,i.p). The<br />

protective effect str<strong>on</strong>gly correlated with the degree <str<strong>on</strong>g>of</str<strong>on</strong>g> LPS molecular structure modificati<strong>on</strong> and was<br />

significantly enhanced by the combined impact <str<strong>on</strong>g>of</str<strong>on</strong>g> detoxified LPS and extracellular Hsp70. Detoxified LPS<br />

activated antiendotoxin immune resp<strong>on</strong>se and reduced LPS-induced inflammatory cytokine producti<strong>on</strong>. Also<br />

it was effective in the treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> endotoxin-related injuries, inhibited the deleterious effects <str<strong>on</strong>g>of</str<strong>on</strong>g> ecological


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

and occupati<strong>on</strong>al toxicants. Significant exacerbati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> whole blood Hsp70 c<strong>on</strong>tent after its incubati<strong>on</strong> with<br />

modified LPS (in vitro) or in resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> the patient to high elevati<strong>on</strong> (5000 m) hypoxia and total body<br />

hyperthermia reveals the necessity <str<strong>on</strong>g>for</str<strong>on</strong>g> n<strong>on</strong>toxic LPS applicati<strong>on</strong>. N<strong>on</strong>resp<strong>on</strong>ders need the applicati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

exogenous chaper<strong>on</strong>s. The activati<strong>on</strong> product <str<strong>on</strong>g>of</str<strong>on</strong>g> yeasts culture Saccharomyces cerevisiae may be a promising<br />

source <str<strong>on</strong>g>of</str<strong>on</strong>g> these cytokines. The yeasts were activated by laser radiati<strong>on</strong>, electr<strong>on</strong>ic beam and cavitati<strong>on</strong>. In<br />

summary the data prove that applicati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> modified LPS and exogenous chaper<strong>on</strong>es can be promising in<br />

treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> toxin-related injuries.<br />

6F_08_P<br />

(poster secti<strong>on</strong> B2, poster board #273, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

COMPARISON OF VASCULAR REACTIVITY BETWEEN SURVIVAL AND NON-<br />

SURVIVAL RATS WITH SEPTIC STRESS INDUCED BY PERITONITIS<br />

C. C. Wu 1 , S. J. Chen 2 , P. L. Chiang 1 , Z. F. Chen 1 , C. T. Chiu 1 , W. J. Liaw 3<br />

Departments <str<strong>on</strong>g>of</str<strong>on</strong>g> 1 Pharmacology and 3 Anesthiology, Nati<strong>on</strong>al Defense Medical Center, Taipei; Department <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Nursing, Kang-Ning Junior College <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Care and Management, Taipei; Taiwan. ccwu@ndmctsgh.edu.tw<br />

The sepsis-induced metabolism has two stress phases: hyper-metabolism followed by hypo-metabolism. The<br />

vascular hyporeactivity to vasoc<strong>on</strong>strictor agents, an important characterizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> circulatory failure in septic<br />

stress, occurred at the late phase <str<strong>on</strong>g>of</str<strong>on</strong>g> sepsis. This study was to compare vascular reactivity between survival and<br />

n<strong>on</strong>-survival rats with perit<strong>on</strong>itis-induced septic stress and to evaluate whether this is associated with<br />

physiological metabolism. Sepsis stress was induced by a surgery <str<strong>on</strong>g>of</str<strong>on</strong>g> cecal ligati<strong>on</strong> and puncture (CLP) in male<br />

Wister rats. We measured resting membrane potential (RMP) and norepinephrine (NE)-induced tensi<strong>on</strong> in<br />

isolated thoracic aortas by electrophysiology and tensi<strong>on</strong> recording experiments. Our results dem<strong>on</strong>strated<br />

that hypotensi<strong>on</strong>, hypoglycemia and multiple organ dysfuncti<strong>on</strong> syndrome (MODS) occurred in n<strong>on</strong>-survival<br />

rats. In additi<strong>on</strong>, not <strong>on</strong>ly the RMP was more hyperpolarized in aorta from n<strong>on</strong>-survival rats, but also the<br />

vascular reactivity to NE was reduced in n<strong>on</strong>-survival rats than that in survival rats. However, the RMP and<br />

vascular reactivity to NE were not significant different in n<strong>on</strong>-survival groups whether they died at early or<br />

late phase. Our results suggest that n<strong>on</strong>-survival rats after CLP were associated with vascular hyporeactivity<br />

which may lead to MODS.<br />

6F_09_P<br />

(poster secti<strong>on</strong> B2, poster board #274, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SESAME OIL PROTECTS AGAINST HYDROXYL-RADICAL-ASSOCIATED LIPID<br />

PEROXIDATION IN LIVER AFTER CECAL LIGATION AND PUNCTURE IN RATS<br />

Ming-Yie Liu a , Ya-Hui Li a , Se-Ping Chien b , Dur-Z<strong>on</strong>g Hsu a<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Envir<strong>on</strong>mental and Occupati<strong>on</strong>al Health,<br />

Nati<strong>on</strong>al Cheng Kung University, College <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine<br />

Address: 138 Sheng-Li Road, Tainan, Taiwan 704.<br />

e-mail: myliu@mail.ncku.edu.tw<br />

Sesame oil protects against oxidative-stress-associated hepatic injury in experimental sepsis. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this<br />

study was to explore the effects underlying the inhibitory effect <str<strong>on</strong>g>of</str<strong>on</strong>g> sesame oil <strong>on</strong> hepatic lipid peroxidati<strong>on</strong> in<br />

septic rats. After <strong>on</strong>e-week daily supplement <str<strong>on</strong>g>of</str<strong>on</strong>g> sesame oil (4 mL/kg/day, orally) to rats, hepatic injury was<br />

351


23-26 August 2007,<br />

Budapest, Hungary<br />

induced by cecal ligati<strong>on</strong> and puncture. Hepatic oxidative stress was assessed by determining the levels <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

hepatic lipid peroxidati<strong>on</strong>, hydroxyl radical, superoxide ani<strong>on</strong>, and nitric oxide 12 h after cecal ligati<strong>on</strong> and<br />

puncture in rats. In additi<strong>on</strong>, the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> xanthine oxidase and the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> inducible nitric oxide<br />

synthase were also determined. Hepatic lipid peroxidati<strong>on</strong>, hydroxyl radical, superoxide ani<strong>on</strong>, and nitrite<br />

levels were significantly lower in sesame-oil-treated septic rats. Furthermore, sesame oil decreased the activity<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> xanthine oxidase and the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> inducible nitric oxide synthase in septic rats. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, sesame oil<br />

might reduce hydroxyl-radical-associated hepatic lipid peroxidati<strong>on</strong> by inhibiting superoxide ani<strong>on</strong> and nitric<br />

oxide, at least partially, in septic rats.<br />

352


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

6G. PROTEIN MISFOLDING, NEURODEGENERATION AND DISEASE<br />

(WILLIAM E. BALCH)<br />

6G_01_S<br />

STRESS AND PRIONS: LESSONS FROM THE YEAST MODEL<br />

Yury O. Chern<str<strong>on</strong>g>of</str<strong>on</strong>g>f<br />

School <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology and Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Bioengineering and Bioscience, Georgia Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Technology,<br />

Altlanta, Georgia 30332-0230, USA<br />

e-mail: yury.chern<str<strong>on</strong>g>of</str<strong>on</strong>g>f@biology.gatech.edu<br />

Amyloids are fiber-like ordered aggregates generated via intermolecular cross-ß interacti<strong>on</strong>s. In vivo amyloid<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> is a widespread phenomen<strong>on</strong> in eukaryotes. Self-perpetuating amyloids provide a basis <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

transmissible protein is<str<strong>on</strong>g>of</str<strong>on</strong>g>orms (pri<strong>on</strong>s) that cause infectious neurodegenerative diseases in mammals<br />

(including humans) and manifest themselves as n<strong>on</strong>-Mendelian heritable elements in yeast and other fungi.<br />

Propagati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the pri<strong>on</strong> state in yeast is c<strong>on</strong>trolled by the c<strong>on</strong>certed acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> chaper<strong>on</strong>e proteins. A crucial<br />

role in this process is played by the chaper<strong>on</strong>e protein Hsp104, which promotes breakage <str<strong>on</strong>g>of</str<strong>on</strong>g> amyloids into<br />

smaller oligomeric seeds initiating new rounds <str<strong>on</strong>g>of</str<strong>on</strong>g> pri<strong>on</strong> proliferati<strong>on</strong>. Pri<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> and propagati<strong>on</strong> are<br />

also influenced by other stress-related chaper<strong>on</strong>es (Hsp70 and Hsp40), and by alterati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> the protein<br />

trafficking and degradati<strong>on</strong> networks. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, pri<strong>on</strong> propagati<strong>on</strong> employs enzymatic machinery which is<br />

normally supposed to protect cells from envir<strong>on</strong>mental stresses. Some stresses induce pri<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> or<br />

loss. It is possible that pri<strong>on</strong>s arise as by-products <str<strong>on</strong>g>of</str<strong>on</strong>g> the reversible assembly <str<strong>on</strong>g>of</str<strong>on</strong>g> highly ordered complexes,<br />

protecting certain proteins from destructi<strong>on</strong> during unfavorable c<strong>on</strong>diti<strong>on</strong>s.<br />

6G_02_S<br />

INTRACELLULAR FATE OF MISFOLDED PROTEINS ASSOCIATED WITH<br />

RETINAL DEGENERATION<br />

Shalesh Kaushal, Syed M. Noorwez<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Florida, Departments <str<strong>on</strong>g>of</str<strong>on</strong>g> Ophthalmology and Cell Biology, Gainesville, Florida, USA<br />

e-mail: skaushal@ufl.edu<br />

Misfolded proteins are found in many inherited diseases, which collectively are called protein c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>al<br />

disorders. These misfolded proteins, typically c<strong>on</strong>taining <strong>on</strong>e or more mutati<strong>on</strong>s, are retained intracellularly in<br />

the endoplasmic reticulum or in cytoplasmic aggregates. Our previous studies clearly dem<strong>on</strong>strated that the<br />

clinically comm<strong>on</strong> P23H opsin mutant associated with autosomal dominant retinitis pigmentosa is misfolded<br />

and retained in the cell. Pharmacological inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the heat shock resp<strong>on</strong>se has emerged as an attractive<br />

strategy <str<strong>on</strong>g>for</str<strong>on</strong>g> modulating the cellular folding envir<strong>on</strong>ment and treating protein c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>al diseases. The<br />

induced ensemble <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock proteins act as molecular chaper<strong>on</strong>es in protein folding and protect against<br />

the <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> misfolded protein aggregates. Using a tetracycline-inducible HEK293 cell lines, we have<br />

studied the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> the heat shock resp<strong>on</strong>se <strong>on</strong> P23H opsin and folded rhodopsin levels in the cell. The<br />

known chemical inducers <str<strong>on</strong>g>of</str<strong>on</strong>g> the heat shock resp<strong>on</strong>se, geldanamycin (GA) and celastrol, increase the total<br />

opsin levels by 1.8- and 1.6-fold (n=3), respectively. Further, GA and celastrol reproducibly increase folded<br />

rhodopsin levels by 1.7- and 1.4-fold (n=3). The level <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp70 in the treated cells was elevated by 4 fold by<br />

GA and 1.2-fold with celastrol. Both treatments appear to induce the heat shock resp<strong>on</strong>se and stabilize the<br />

folded <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> opsin i.e. the <str<strong>on</strong>g>for</str<strong>on</strong>g>m capable <str<strong>on</strong>g>of</str<strong>on</strong>g> binding retinal. Further, the extent <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp70 increase correlates<br />

353


23-26 August 2007,<br />

Budapest, Hungary<br />

with te yields <str<strong>on</strong>g>of</str<strong>on</strong>g> folded P23H opsin. Thus, heat shock inducti<strong>on</strong> clearly alters the cellular stability and fate <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

P23H opsin. Both compounds are attractive candidates <str<strong>on</strong>g>for</str<strong>on</strong>g> treating P23H opsin associated retinal<br />

degenerati<strong>on</strong>.<br />

354<br />

6G_03_S<br />

MISFOLDED PROTEINS IN AGING AND NEURODEGENERATIVE DISEASE<br />

Cindy Voisine, Michael Schieber, Daniel Czyz, Richard I. Morimoto<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, Molecular Biology and Cell Biology, Northwestern University, Evanst<strong>on</strong>, IL 60208<br />

e-mail: c-voisine@northwestern.edu<br />

Misfolded and damaged proteins challenge homeostasis and the capacity <str<strong>on</strong>g>of</str<strong>on</strong>g> molecular chaper<strong>on</strong>es and<br />

clearance machineries to restore the health <str<strong>on</strong>g>of</str<strong>on</strong>g> the cell. Whereas acute exposures to heat shock and other<br />

envir<strong>on</strong>ment stress leads to the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cytoprotective resp<strong>on</strong>ses, the chr<strong>on</strong>ic expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> misfolded<br />

and mutant proteins can result in irreversible toxicity as occurs in human neurodegenerative disease. We have<br />

established C. elegans models to identify the genes that regulate protein homeostasis in resp<strong>on</strong>se to chr<strong>on</strong>ic<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> aggregati<strong>on</strong>-pr<strong>on</strong>e proteins associated with neurodegenerative disease. These studies have<br />

identified an important molecular link between the insulin-signaling pathway that regulates l<strong>on</strong>gevity and<br />

HSF-1, the master regulator <str<strong>on</strong>g>of</str<strong>on</strong>g> protein folding quality c<strong>on</strong>trol and the heat shock resp<strong>on</strong>se, revealing that<br />

l<strong>on</strong>gevity is closely linked to the cellular folding capacity. Using <str<strong>on</strong>g>for</str<strong>on</strong>g>ward genetics, genome-wide RNAi screens<br />

and a candidate gene approach, we have identified a functi<strong>on</strong>ally diverse set <str<strong>on</strong>g>of</str<strong>on</strong>g> genes that sort into six<br />

networks involved in RNA metabolism, protein synthesis, protein folding, protein degradati<strong>on</strong>, protein<br />

trafficking, and mitoch<strong>on</strong>drial functi<strong>on</strong> and energy metabolism that influence protein homeostasis. Many <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

these genes are also important <str<strong>on</strong>g>for</str<strong>on</strong>g> the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock gene expressi<strong>on</strong> which suggests that genetic<br />

modifiers <str<strong>on</strong>g>for</str<strong>on</strong>g> protein quality c<strong>on</strong>trol defines a protein-folding proteome that also functi<strong>on</strong>s to sense an<br />

imbalance in protein homeostasis and regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> chaper<strong>on</strong>e expressi<strong>on</strong>. Using our C. elegans models, we are<br />

characterizing Hsp70 chaper<strong>on</strong>e networks that coordinately functi<strong>on</strong> to protect cells in resp<strong>on</strong>se to misfolded<br />

disease proteins.<br />

6G_04_S<br />

CHAPEROME SYSTEMS AND PROTEIN MISFOLDING STRESS IN<br />

TRAFFICKING DISEASE<br />

William E. Balch 1,2,6 , Atanas Koulov 1 , Darren Hutt 1 , Bogdan Tanasa 1 , Wendy Kellner 1 ,<br />

Helen Plutner 1 , Paul Lapointe 1 , Scott Stagg 1 , Clint S. Potter 1,5 , Bridget Carragher 1,5 ,<br />

Jeffery W. Kelly 3,4 , John R. Yates III 1<br />

1The Scripps <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute, Departments <str<strong>on</strong>g>of</str<strong>on</strong>g> Cell and 2 Molecular Biology and 3 Chemistry,<br />

4The Skaggs Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Chemical Biology, 5 The Nati<strong>on</strong>al Resource <str<strong>on</strong>g>for</str<strong>on</strong>g>Automated Molecular Microscopy,<br />

6The Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Childhood and Neglected Diseases,<br />

10550 North Torrey Pines Road, La Jolla, CA 92037<br />

Protein folding in the eukaryotic cell is highly sensitive to the local envir<strong>on</strong>ment <str<strong>on</strong>g>of</str<strong>on</strong>g> the cytoplasm or<br />

membrane trafficking compartments, and requires the assistance <str<strong>on</strong>g>of</str<strong>on</strong>g> multiple chaper<strong>on</strong>es that define the<br />

folding buffer <str<strong>on</strong>g>of</str<strong>on</strong>g> the cell- the chaperome (Wang et al. (2006) Cell, 127:803). Different cell types exploit the<br />

variable compositi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the chaperome envir<strong>on</strong>ment to maintain protein folding homeostasis- reflecting the


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

kinetic and thermodynamic properties <str<strong>on</strong>g>of</str<strong>on</strong>g> the protein fold. Perturbati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> this relati<strong>on</strong>ship, as occurs in<br />

misfolding diseases such as cystic fibrosis (CFTR), Gaucher's (glucocerebrosidase), childhood emphysema<br />

(alpha-1-antitrypsin), type II diabetes and numerous amyloid diseases (Alzheimers (APP), Parkins<strong>on</strong>s (alphasynuclein)),<br />

results in an imbalance between the protein fold and the folding envir<strong>on</strong>ment leading to<br />

disrupti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> normal physiology. It is becoming increasingly apparent that folding pathways in aging and<br />

disease can be altered by the cell through stress sensitive pathways to maintain or reestablish the proper<br />

chaperome envir<strong>on</strong>ment to maintain functi<strong>on</strong>ality. Emerging mechanisms by which these activities work<br />

provides insight into the pathways that can be manipulated through biological and chemical approaches to<br />

adjust the flow <str<strong>on</strong>g>of</str<strong>on</strong>g> folded and functi<strong>on</strong>al protein through the exocytic and endocytic pathways.<br />

6G_05_S<br />

(poster secti<strong>on</strong> B2, poster board #275, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

A REVERSE GENETIC OVEREXPRESSION SCREEN REVEALS NON-CANONICAL<br />

CHAPERONES AS POTENT SUPPRESSORS OF POLYGLUTAMINE PROTEINS<br />

Harm H. Kampinga, J. Hageman, M. Rujano, M. Vos<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Radiati<strong>on</strong> and Stress Cell Biology, University Medical Center Gr<strong>on</strong>ingen, Univ. <str<strong>on</strong>g>of</str<strong>on</strong>g> Gr<strong>on</strong>ingen,<br />

The Netherlands<br />

e-mail: h.h.kampinga@med.umcg.nl<br />

As they are able to bind misfolded proteins, Heat Shock Proteins (HSP) may prevent accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> toxic<br />

poly-Q protein aggregates and as such inhibit disease-associated pathogenesis. Several in vitro reports have<br />

pointed to the classical Hsp70 chaper<strong>on</strong>e machines and their c<str<strong>on</strong>g>of</str<strong>on</strong>g>actors as suppressors <str<strong>on</strong>g>of</str<strong>on</strong>g> polyglutaminerelated<br />

protein aggregati<strong>on</strong>. Yet, the (limited) in vivo work with mouse models has yielded disappointing<br />

results from minor delays in disease progressi<strong>on</strong> to no effect at all. It has become clear recently that the many<br />

different Hsp subfamily members may have different and substrate specific chaper<strong>on</strong>e-like functi<strong>on</strong>s. To seek<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> chaper<strong>on</strong>es that may be particularly effective in dealing with polyglutamine proteins, we c<strong>on</strong>ducted a<br />

reverse genetic overexpressi<strong>on</strong> screen <str<strong>on</strong>g>of</str<strong>on</strong>g> the human chaper<strong>on</strong>ome. We identified a number <str<strong>on</strong>g>of</str<strong>on</strong>g> n<strong>on</strong>-can<strong>on</strong>ical<br />

Hsp40 family members as well as a new member <str<strong>on</strong>g>of</str<strong>on</strong>g> the small Hsp family as superior inhibitors <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

polyglutamine aggregati<strong>on</strong>. Unlike the can<strong>on</strong>ical DnaJB1 (that has mild suppressive activity in vitro), the<br />

suppressi<strong>on</strong> by the n<strong>on</strong>-can<strong>on</strong>ical Hsp40’s was not annihilated by mutating the HPD motif in their J-domain,<br />

normally required <str<strong>on</strong>g>for</str<strong>on</strong>g> interacti<strong>on</strong> with Hsp70 family members. At suboptimal c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> the n<strong>on</strong>can<strong>on</strong>ical<br />

Hsp40’s, overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> n<strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the Hsp70 family members was able to further reduce the<br />

polyglutamine aggregati<strong>on</strong>. All these data suggest that they act independently <str<strong>on</strong>g>of</str<strong>on</strong>g> the Hsp70 machine. The<br />

acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the small Hsp was independent <strong>on</strong> its N-termimal domain and as such seems to be executed by the<br />

crystallin domain.<br />

355


23-26 August 2007,<br />

Budapest, Hungary<br />

6G_06_S<br />

(poster secti<strong>on</strong> B2, poster board #276, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE HSP60-(P.VAL98ILE) VARIANT PROTEIN ASSOCIATED WITH AUTOSOMAL<br />

DOMINANT SPASTIC PARAPLEGIA SPG13 DISPLAYS SUBTLE EFFECTS WHEN<br />

CO-EXPRESSED WITH WILD TYPE HSP60.<br />

Peter Bross 1 , Marit N. Nielsen 1 , Jane H. Christensen 1 , Jakob Hansen 1 ,Niels Gregersen 1 ,<br />

Debbie Ang 2 , Costa Georgopoulos 2<br />

1<str<strong>on</strong>g>Research</str<strong>on</strong>g> Unit <str<strong>on</strong>g>for</str<strong>on</strong>g> Molecular Medicine, Aarhus University Hospital and Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Health Sciences,<br />

Århus, Denmark<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Microbiology and Molecular Medicine, <str<strong>on</strong>g>Centre</str<strong>on</strong>g> Médical Universitaire, Geneva, Switzerland<br />

e-mail: Peter.Bross@ki.au.dk<br />

We have earlier reported that a mutati<strong>on</strong> (c.292G>A/p.Val98Ile) in the human HSPD1 gene that encodes the<br />

mitoch<strong>on</strong>drial Hsp60 chaper<strong>on</strong>in is associated with dominantly inherited hereditary spastic paraplegia SPG13<br />

[1]. To mimic heterozygosity <str<strong>on</strong>g>for</str<strong>on</strong>g> the Hsp60-(p.Val98Ile) mutati<strong>on</strong> and to assess a potential dominant negative<br />

effect <str<strong>on</strong>g>of</str<strong>on</strong>g> co-expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a wild type and the mutant variant <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp60 <strong>on</strong> the functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the 7-meric<br />

chaper<strong>on</strong>in complex, we used a flexible bacterial model system. These cells lack the endogenous chaper<strong>on</strong>in<br />

genes and are maintained alive by expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a plasmid-encoded Hsp60/Hsp10 oper<strong>on</strong>. The cells harbour<br />

a sec<strong>on</strong>d plasmid with a Hsp60/Hsp10 oper<strong>on</strong> encoding a mutant Hsp60. The two oper<strong>on</strong>s can be turned <strong>on</strong><br />

and <str<strong>on</strong>g>of</str<strong>on</strong>g>f separately. We compared the behaviour <str<strong>on</strong>g>of</str<strong>on</strong>g> bacterial cells co-expressing either wild type Hsp60 and<br />

Hsp60-(p.Val98Ile) or wild type Hsp60 and an artificially c<strong>on</strong>structed Hsp60 ATPase mutant. Inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

co-expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the Hsp60 ATPase mutant severely inhibited bacterial growth, whereas co-expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

Hsp60-(p.Val98Ile) mutant variant had <strong>on</strong>ly subtle yet specific effects. Additi<strong>on</strong>al experiments indicate that<br />

both mutant Hsp60 variants <str<strong>on</strong>g>for</str<strong>on</strong>g>m heterologous complexes with wild type subunits. By varying temperature<br />

and relative amounts <str<strong>on</strong>g>of</str<strong>on</strong>g> the wild type and Hsp60-(p.Val98Ile) mutant variant, we have found c<strong>on</strong>diti<strong>on</strong>s<br />

under which the mutant exerts a clear effect. We propose that the major in vivo c<strong>on</strong>sequences <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

heterozygosity <str<strong>on</strong>g>for</str<strong>on</strong>g> the Hsp60-(p.Val98Ile) variati<strong>on</strong> are due to subtle qualitative and quantitative effects<br />

remaining to be discovered.<br />

356


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

6G_01_P<br />

(poster secti<strong>on</strong> B2, poster board #277, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

INACTIVATION OF THE HEREDITARY SPASTIC PARAPLEGIA-ASSOCIATED<br />

HSPD1 GENE RESULTS IN A RECESSIVE EMBRYONIC LETHAL<br />

PHENOTYPE IN MICE<br />

Jane H. Christensen 1 , Marit N. Nielsen 1 , Ernst-Martin Füchtbauer 2 , Thomas J. Coryd<strong>on</strong> 3 ,<br />

Jakob Hansen 1 , Niels Gregersen 1 , Peter G. Bross 1<br />

1) <str<strong>on</strong>g>Research</str<strong>on</strong>g> Unit <str<strong>on</strong>g>for</str<strong>on</strong>g> Molecular Medicine, Aarhus University Hospital, Skejby; 2) Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular Biology,<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Aarhus; 3) Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Human Genetics, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Aarhus, Denmark<br />

e-mail: jane.hvarregaard@ki.au.dk<br />

To establish an animal model <str<strong>on</strong>g>for</str<strong>on</strong>g> the neurodegenerative disease, hereditary spastic paraplegia, we have<br />

generated a mouse which is heterozygous <str<strong>on</strong>g>for</str<strong>on</strong>g> an inactivating inserti<strong>on</strong> in the hspd1 gene. Hspd1 encodes the<br />

mitoch<strong>on</strong>drial chaper<strong>on</strong>in Hsp60. Missense mutati<strong>on</strong>s in <strong>on</strong>e allele <str<strong>on</strong>g>of</str<strong>on</strong>g> the gene have been associated with<br />

hereditary spastic paraplegia in humans. Heterozygous mice were born at the expected frequency compared<br />

to wild type and displayed no obvious severe phenotype. By quantitative RT-PCR we found decreased levels<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> hspd1 transcript in all tissues examined, dem<strong>on</strong>strating that the inactivati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the hspd1 gene is efficient. By<br />

Western blotting analysis, we found that the amount <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp60 protein compared to either cytosolic tubulin<br />

or mitoch<strong>on</strong>drial VDAC/porin was decreased as well. Breeding <str<strong>on</strong>g>of</str<strong>on</strong>g> heterozygous animals resulted in reduced<br />

litter sizes and <str<strong>on</strong>g>of</str<strong>on</strong>g>fspring homozygous <str<strong>on</strong>g>for</str<strong>on</strong>g> the inactivated allele were missing. Timed mating revealed a<br />

significant proporti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> degenerated or growth retarded embryos at 8.5 and 10.5 dpc and normal developed<br />

embryos were either wild type or heterozygous. In c<strong>on</strong>clusi<strong>on</strong>, the inactivati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the hspd1 gene is efficient at<br />

the molecular level and it is associated with a recessive peri-implantati<strong>on</strong>al embry<strong>on</strong>ic lethal phenotype in<br />

mice. We expect that heterozygous mice will develop hereditary spastic paraplegia due to shortage <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp60<br />

chaper<strong>on</strong>in activity, protein quality c<strong>on</strong>trol failure, and mitoch<strong>on</strong>drial dysfuncti<strong>on</strong> in motor neur<strong>on</strong>s with very<br />

l<strong>on</strong>g ax<strong>on</strong>s.<br />

6G_02_P<br />

(poster secti<strong>on</strong> B2, poster board #278, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SMALL HEAT SHOCK PROTEIN HSPB8 AND PROTEIN DEGRADATION IN<br />

MOTONEURON DISEASES<br />

V. Crippa 1 , F. Sim<strong>on</strong>ini 1 , S. Carra 2 , E. Bolz<strong>on</strong>i 1 , E. Onesto 1 , P. Rusmini 1 , D. Sau 1 , A. Poletti 1<br />

1Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Endocrinology, CEND, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Milan. Via Balzaretti 9, 20133 Milan, ITALY<br />

2<str<strong>on</strong>g>Centre</str<strong>on</strong>g> de recherche en cancerologie de l'Universite Laval, CANADA<br />

e-mail: angelo.poletti@unimi.it<br />

In mammals, the small Heat Shock Proteins (HSP) family comprises 10 members called HSPB proteins<br />

(HSPB1-10), with chaper<strong>on</strong>e activity. They are upregulated in neurodegenerative disorders exerting a<br />

neuroprotective role. Mutati<strong>on</strong>s in HSPB1 and HSPB8 have been linked to peripheral neuropathies. We<br />

investigated the role <str<strong>on</strong>g>of</str<strong>on</strong>g> HSPB8 in two mot<strong>on</strong>eur<strong>on</strong>al diseases: spinal and bulbar muscular atrophy (SBMA)<br />

and familial amyotrophic lateral sclerosis (fALS). SBMA is caused by polyglutamine tract expansi<strong>on</strong> (polyQ)<br />

in the Androgen Receptor (AR), while fALS is <str<strong>on</strong>g>of</str<strong>on</strong>g>ten associated to mutati<strong>on</strong>s in Superoxide Dismutase 1<br />

(SOD1). Mutant AR and SOD1 do not share structural or functi<strong>on</strong>al domains, but are unstable and tend to<br />

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23-26 August 2007,<br />

Budapest, Hungary<br />

aggregate. Immortalized mot<strong>on</strong>eur<strong>on</strong>, NSC34, expressing mutant <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> SOD1 (SOD1G93A) or ARpolyQ<br />

(ARQ46) c<strong>on</strong>tain intracellular aggregates <str<strong>on</strong>g>of</str<strong>on</strong>g> the mutant proteins and have a reduced proteasome activity,<br />

measured with accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the proteasome reporter YFPu. HSPB8 overexpressi<strong>on</strong> decreased the levels<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> both mutant proteins and reduced YFPu levels, suggesting a desaturati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the proteasome system. Thus,<br />

HSPB8 exerts chaper<strong>on</strong>e activity towards both mutated AR and SOD1. By immunoprecipitati<strong>on</strong>, we showed<br />

that HSPB8 does not need a protein-protein interacti<strong>on</strong> to reduce mutant AR and SOD1 levels. Moreover,<br />

proteasome inhibiti<strong>on</strong> with MG132 did not block HSPB8 chaper<strong>on</strong>e activity, suggesting that HSPB8 could<br />

activate other degradative pathways, such as the autophagy. Grants Teleth<strong>on</strong> - Italy (#GGP06063), MIUR-<br />

FIRB (#RBAU01NXFP), MIUR-C<str<strong>on</strong>g>of</str<strong>on</strong>g>in (2005057598_002), University <str<strong>on</strong>g>of</str<strong>on</strong>g> Milan-FIRST, FONDAZIONE<br />

CARIPLO.<br />

6G_03_P<br />

(poster secti<strong>on</strong> B2, poster board #279, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ACTIVATION OF STRESS RESPONSE PATHWAYS IN MOTOR NEURON DISEASES:<br />

IMPLICATIONS FOR PATHOGENESIS AND THERAPY<br />

Heather D. Durham, Benoit Gentil, Miranda L. Tradewell, David M. Taylor, Walter E. Mushynski,<br />

Laura Senese, Sandra Minotti<br />

M<strong>on</strong>treal Neurological Institute, McGill University, M<strong>on</strong>treal QC, Canada<br />

e-mail: heather.durham@mcgill.ca<br />

Neural cells differ in their ability to induce heat shock genes in resp<strong>on</strong>se to thermal or disease-related stresses.<br />

Motor neur<strong>on</strong>s have a high threshold <str<strong>on</strong>g>for</str<strong>on</strong>g> up-regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> stress-inducible Hsp70 [Manzerra and Brown,<br />

1992; Batulan et al, 2003]. However, increasing levels <str<strong>on</strong>g>of</str<strong>on</strong>g> multiple chaper<strong>on</strong>es is protective in culture and<br />

mouse models <str<strong>on</strong>g>of</str<strong>on</strong>g> motor neur<strong>on</strong> diseases including amyotrophic lateral sclerosis [Bruening et al, 1999; Kieran<br />

et al, 2004; Batulan et al, 2006], spinal bulbar muscular atrophy [Katsuno et al, 2005], and Charcot-Marie-<br />

Tooth (CMT) disease type 1A due to missense mutati<strong>on</strong> in PMP22 [Fortun et al, 2007]. We have<br />

dem<strong>on</strong>strated preventi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the disease-related phenotype in a primary culture model <str<strong>on</strong>g>of</str<strong>on</strong>g> CMT2 caused by<br />

mutati<strong>on</strong>s in nefl encoding the neur<str<strong>on</strong>g>of</str<strong>on</strong>g>ilament light protein (NFL), as well as <str<strong>on</strong>g>of</str<strong>on</strong>g> ALS due to sod1 mutati<strong>on</strong>.<br />

Increasing levels <str<strong>on</strong>g>of</str<strong>on</strong>g> HSPs, including Hsp70 and Hsp40, by over-expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> c<strong>on</strong>stitutively active Hsf1<br />

prevented bundling and fragmenting <str<strong>on</strong>g>of</str<strong>on</strong>g> the neur<str<strong>on</strong>g>of</str<strong>on</strong>g>ilament network and aggregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> CMT-related mutant<br />

NFL in motor neur<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> dissociated spinal cord cultures. Upregulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> chaper<strong>on</strong>es, including Hsp27,<br />

facilitated degradati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> NFL mutants in SW13 vim- cells. Given the potential <str<strong>on</strong>g>for</str<strong>on</strong>g> chaper<strong>on</strong>e-based therapies<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> neurological disorders, our group is studying the mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp regulati<strong>on</strong> in motor neur<strong>on</strong>s and is<br />

using primary culture models <str<strong>on</strong>g>of</str<strong>on</strong>g> motor neur<strong>on</strong> diseases to screen compounds <str<strong>on</strong>g>for</str<strong>on</strong>g> their ability to increase<br />

neur<strong>on</strong>al HSP levels and <str<strong>on</strong>g>for</str<strong>on</strong>g> neuroprotecti<strong>on</strong>. Differences identified in mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> activating hsp<br />

expressi<strong>on</strong> in neur<strong>on</strong>s included a CaMKIV-dependent pathway <str<strong>on</strong>g>for</str<strong>on</strong>g> upregulating Hsp70 in motor neur<strong>on</strong>s, but<br />

not in fibroblasts.<br />

358


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

6G_04_P<br />

(poster secti<strong>on</strong> B2, poster board #280, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DECREASED EXPRESSION OF THE MITOCHONDRIAL MATRIX PROTEASES<br />

CLPP AND LON IN CELLS FROM PATIENT WITH HEREDITARY SPASTIC<br />

PARAPLEGIA (SPG13)<br />

Jakob Hansen 1 , Thomas J. Coryd<strong>on</strong> 2 , Alexandra Dürr 3,4 , Marit N. Nielsen 1 , Jane H. Christensen 1 ,<br />

Niels Gregersen 1 , Bertrand F<strong>on</strong>taine 5 , Peter Bross 1<br />

1<str<strong>on</strong>g>Research</str<strong>on</strong>g> Unit <str<strong>on</strong>g>for</str<strong>on</strong>g> Molecular Medicine, Aarhus University Hospital, Skejby, Denmark<br />

2Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Human Genetics, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Aarhus, Aarhus, Denmark<br />

3INSERM U679, 4 Departement de genetique, 5 INSERM U546, Hôpital de la Salpêtrière, Paris, France<br />

e-mail: Jakob.hansen@ki.au.dk<br />

The mitoch<strong>on</strong>drial Hsp60 chaper<strong>on</strong>e promotes folding <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins in the mitoch<strong>on</strong>drial matrix space, and<br />

plays a crucial role in protein quality c<strong>on</strong>trol. A mutati<strong>on</strong> in the HSPD1 (SPG13) gene encoding the mutant<br />

Hsp60-(p.Val98Ile) protein has been associated with a dominantly inherited <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> spastic paraplegia. The<br />

Hsp60-(p.Val98Ile) protein is functi<strong>on</strong>ally impaired and displays a reduced efficiency in mediating folding <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the malate dehydrogenase substrate protein possibly related to a reduced ATP-ase activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the chaper<strong>on</strong>in<br />

complex, but the molecular defect involved in ax<strong>on</strong>al degenerati<strong>on</strong> in spastic paraplegia is unknown. We have<br />

investigated mitoch<strong>on</strong>drial functi<strong>on</strong> and gene expressi<strong>on</strong> levels <str<strong>on</strong>g>of</str<strong>on</strong>g> key mitoch<strong>on</strong>drial chaper<strong>on</strong>es and<br />

proteases in cultured lymphoblastoid and fibroblast cells from SPG13 patient cells. We found that impaired<br />

Hsp60-(p.Val98Ile) functi<strong>on</strong> is not related to a severe mitoch<strong>on</strong>drial dysfuncti<strong>on</strong> phenotype as indicated by<br />

assessment <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>drial membrane potential, cell vitality, and sensitivity towards oxidative stress insults.<br />

However, a decreased expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> protein quality c<strong>on</strong>trol proteases L<strong>on</strong> and ClpP in SPG13 patient cells<br />

was dem<strong>on</strong>strated. We propose that decreased protease levels may represent an adaptive change <str<strong>on</strong>g>of</str<strong>on</strong>g> protein<br />

quality c<strong>on</strong>trol giving more time to the folding <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins whose folding is impaired due to a reduced activity<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp60-(p.Val98Ile).<br />

6G_05_P<br />

(poster secti<strong>on</strong> B2, poster board #281, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

A NOVEL NUCLEAR DNAJ PROTEIN, DNAJC8, CAN SUPPRESS THE ATAXIN-3-<br />

POLYQ AGGREGATION AND CELL DEATH IN A J-DOMAIN INDEPENDENT<br />

MANNER<br />

Norie Ito 1, 3 , Kenjiro Kamiguchi 1 , Toshihiko Torigoe 1* , Katsuya Nakanishi 1 , Alisa Sokolovskya 1 ,<br />

Susumu Chiba 3 , Shun Shimohama 2 , Noriyuki Sato 1<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Pathology 1 and Neurology 2 , Sapporo Medical University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine<br />

South-1, West-17, Chuo-ku, Sapporo 060-8556, Japan, Sapporo Yamano-ue Hospital<br />

e-mail: torigoe@sapmed.ac.jp *<br />

Machado-Joseph disease (MJD), also termed spinocerebellar ataxia type3, is a neurodegenerative disorder<br />

characterized by abnormal movement coordinati<strong>on</strong>. The disease is fatal and is inherited in a dominant<br />

manner. MJD is caused by a pathogenic ataxin-3 protein with an expanded polyglutamine (polyQ) stretch.<br />

The neurotoxicity <str<strong>on</strong>g>of</str<strong>on</strong>g> the expanded polyQ-c<strong>on</strong>taining ataxin-3 is closely associated with its aggregate<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>. We report here that a novel J-protein, DNAJC8 (JC8) suppresses the ataxin-3-polyQ aggregati<strong>on</strong><br />

359


23-26 August 2007,<br />

Budapest, Hungary<br />

in a cellular model <str<strong>on</strong>g>of</str<strong>on</strong>g> MJD. JC8 was identified by the human EST database search <str<strong>on</strong>g>for</str<strong>on</strong>g> nuclear J-proteins,<br />

since ataxin-3 protein with expanded polyQ stretch <str<strong>on</strong>g>for</str<strong>on</strong>g>ms aggregati<strong>on</strong> in the nucleus <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>al cells.<br />

Overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> JC8 in SH-SY5Y neuroblastoma cells significantly suppressed the polyQ aggregati<strong>on</strong> and<br />

cell death. JC8 was co-localized with the polyQ-c<strong>on</strong>taining protein in the nucleus. To identify a resp<strong>on</strong>sible<br />

regi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> JC8 affecting the polyQ aggregati<strong>on</strong>, a series <str<strong>on</strong>g>of</str<strong>on</strong>g> JC8 deleti<strong>on</strong> mutants were examined <str<strong>on</strong>g>for</str<strong>on</strong>g> their ability<br />

to suppress the aggregati<strong>on</strong>. Interestingly, C-terminal domain <str<strong>on</strong>g>of</str<strong>on</strong>g> JC8 was essential <str<strong>on</strong>g>for</str<strong>on</strong>g> the suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

polyQ aggregati<strong>on</strong>, whereas J-domain was dispensable. These results indicate that JC8 might suppress the<br />

polyQ aggregati<strong>on</strong> by a distinct mechanism independent <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70-based chaper<strong>on</strong>e machinery and have a<br />

unique protective role against the aggregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> expanded polyQ-c<strong>on</strong>taining proteins such as a pathogenic<br />

ataxin-3 protein.<br />

6G_06_P<br />

(poster secti<strong>on</strong> B2, poster board #282, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

OVEREXPRESSION OF HSP27 HAS NO EFFECT ON SURVIVAL IN A MOUSE<br />

MODEL OF AMYOTROPHIC LATERAL SCLEROSIS<br />

J. Krishnan, K. Vannuvel, D. Kiraly, W. Robberecht, L. Van Den Bosch<br />

Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Neurobiology, K. U. Leuven, B-3000 Leuven, Belgium<br />

e-mail: Jyothsna.Krishnan@med.kuleuven.be<br />

Amyotrophic Lateral Sclerosis (ALS) is a chr<strong>on</strong>ic neurodegenerative disorder characterized by the selective<br />

loss <str<strong>on</strong>g>of</str<strong>on</strong>g> upper and lower motor neur<strong>on</strong>s. This disease results in gradual atrophy <str<strong>on</strong>g>of</str<strong>on</strong>g> muscles leading to paralysis<br />

and death about 3-5 years after diagnosis. Familial ALS (fALS) is resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> 10% <str<strong>on</strong>g>of</str<strong>on</strong>g> cases and mutati<strong>on</strong>s<br />

in the superoxide dismutase 1 (SOD1) gene cause 2% <str<strong>on</strong>g>of</str<strong>on</strong>g> fALS. The exact pathogenic mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> mutant<br />

SOD1 induced motor neur<strong>on</strong> death is still elusive and is thought to involve oxidative stress and protein<br />

aggregati<strong>on</strong>. These two phenomena are known to induce heat shock proteins (Hsp’s) which protect cells<br />

through their chaper<strong>on</strong>ing and anti-apoptotic activity. Loss <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp27 from motor neur<strong>on</strong>s preceding disease<br />

has been suggested to c<strong>on</strong>tribute to their degenerati<strong>on</strong> in ALS mice. Moreover, several Hsp’s, notably Hsp27,<br />

have been shown to be protective in a number <str<strong>on</strong>g>of</str<strong>on</strong>g> in vitro ALS models. However, we recently showed a lack<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> protective effect <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp27 against mutant SOD1-dependent cell death in N2a cells. In this study, we show<br />

corroborative results using the ALS mice. Mice that overexpress the human G93A SOD1 mutant developing<br />

ALS-like symptoms were crossed with ubiquitous Hsp27 overexpressors and the resulting double transgenic<br />

mice (Dtg) were tested. The Dtg mice did not live l<strong>on</strong>ger, nor had significant delayed <strong>on</strong>set <str<strong>on</strong>g>of</str<strong>on</strong>g> disease<br />

compared to their G93A SOD1 littermates. No evidence that motor neur<strong>on</strong>s were protected in these mice<br />

was found histologically. Also no difference in the activati<strong>on</strong>, nor in the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> members <str<strong>on</strong>g>of</str<strong>on</strong>g> the apoptotic<br />

machinery was found in these mice. In c<strong>on</strong>clusi<strong>on</strong>, Hsp27 al<strong>on</strong>e does not seem to be sufficient to protect<br />

against the devastating effects <str<strong>on</strong>g>of</str<strong>on</strong>g> mutant SOD1.<br />

360


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

6G_07_P<br />

(poster secti<strong>on</strong> B2, poster board #283, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EXTREMOLYTES: STRESS PROTECTIVE LOW-MOLECULAR-WEIGHT<br />

COMPOUNDS AS INHIBITORS OF AMYLOID FORMATION<br />

Jungki Ryu 1 , Chan Beum Park 1 , Georg Lentzen 2<br />

1 Korea Advanced Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Science and Technology (KAIST), Daeje<strong>on</strong>, Republic <str<strong>on</strong>g>of</str<strong>on</strong>g> Korea<br />

2 Bitop AG, Stockumer Str. 28, 58453 Witten, Germany<br />

Extremolytes are low-molecular weight osmolytes accumulated by extremophilic microorganisms as a<br />

resp<strong>on</strong>se to osmotic and temperature stress and are known to act as chemical chaper<strong>on</strong>es by stabilizing<br />

cellular structures (1). Extremolytes have been shown to prevent the misfolding <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins and to maintain<br />

their stability and can reach high intracellular c<strong>on</strong>centrati<strong>on</strong>s without interfering with metabolism.<br />

β-Amyloid peptide (Aβ) is the major c<strong>on</strong>stituent <str<strong>on</strong>g>of</str<strong>on</strong>g> senile plaques, the key pathological feature <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Alzheimer´s disease. Aβ is physiologically produced as a soluble <str<strong>on</strong>g>for</str<strong>on</strong>g>m, but the aggregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Aβ m<strong>on</strong>omers<br />

causes neurotoxicity. Ectoine, an extremolyte widespread in extremophilic (halophilic) bacteria has been<br />

shown to interfere with the <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> amyloid aggregates in vitro and amyloid-induced cytotoxicity (2,3).<br />

Using atomic <str<strong>on</strong>g>for</str<strong>on</strong>g>ce microscopy and an assay based <strong>on</strong> thi<str<strong>on</strong>g>of</str<strong>on</strong>g>lavin T fluorescence, we have now tested<br />

synthetic ectoine analogs and other osmolytes <str<strong>on</strong>g>for</str<strong>on</strong>g> their ability to interfere with Alzheimer peptide Aß amyloid<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> in vitro. We show that a synthetic analog <str<strong>on</strong>g>of</str<strong>on</strong>g> ectoine with widened ring size interferes effectively<br />

with amyloid fibril <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>. The results indicate the possibility <str<strong>on</strong>g>of</str<strong>on</strong>g> designing synthetic compounds with<br />

chemical chaper<strong>on</strong>e properties as potential inhibitors <str<strong>on</strong>g>of</str<strong>on</strong>g> amyloid <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> associated with neurodegenerative<br />

diseases.<br />

6G_08_P<br />

(poster secti<strong>on</strong> B2, poster board #284, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HSP90Α AS PART OF THE POLYGENIC RESPONSE TO SCRAPIE DEVELOPMENT<br />

IN SHEEP<br />

A. Marcos-Carcavilla 1 , J. H. Calvo 2 , C. G<strong>on</strong>zález 1 , K. Moazami-Goudarzi 3 , P. Laurent 3 , M. Bertaud 3 ,<br />

H. Hayes 3 , A. E. Beattie 4 , C. Serrano 5 , J. Lyahyai 5 , I. Martín-Burriel 5 , M. Serrano 1<br />

1INIA, 28040, Madrid, Spain, 2 CITA, 50059, Zaragoza, Spain<br />

3INRA, F-78350 Jouy-en-Josas, France<br />

4Ag<str<strong>on</strong>g>Research</str<strong>on</strong>g>, Invermay Agricultural <str<strong>on</strong>g>Centre</str<strong>on</strong>g>, Private Bag 50034, Mosgiel, New Zealand<br />

5Universidad de Zaragoza, 50013, Zaragoza, Spain<br />

e-mail: amarcos@inia.es<br />

Pri<strong>on</strong> diseases, including ovine scrapie, are fatal neurodegenerative diseases characterized by the c<strong>on</strong>versi<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the host encoded PrP c into its aberrant and aggregate-pr<strong>on</strong>e counterpart PrP sc . Molecular chaper<strong>on</strong>es<br />

provide the first line <str<strong>on</strong>g>of</str<strong>on</strong>g> defence against misfolded proteins and probably functi<strong>on</strong> at the earliest stages <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

pri<strong>on</strong> pathogenesis. In the present work, the ovine gene encoding the Hsp90α (HSPCAA1), including the<br />

promoter and other regulatory regi<strong>on</strong>s, has been isolated and characterized. Several sequence polymorphisms<br />

have also been identified. Genetic and cytogenetic mapping localized the ovine HSPCAA1 gene <strong>on</strong> a<br />

chromosome regi<strong>on</strong> previously described as a QTL interval <str<strong>on</strong>g>for</str<strong>on</strong>g> scrapie incubati<strong>on</strong> period in sheep<br />

(OAR19q24dist). Quantitative PCR results revealed no changes at HSPCAA1 mRNA level as c<strong>on</strong>sequence<br />

361


23-26 August 2007,<br />

Budapest, Hungary<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the scrapie infecti<strong>on</strong>. Nevertheless, associati<strong>on</strong> analyses revealed that several polymorphisms in the 5’<br />

flanking regi<strong>on</strong> and intr<strong>on</strong> 10 <str<strong>on</strong>g>of</str<strong>on</strong>g> the ovine HSPCAA1 gene were differentially distributed between sheep with<br />

different resp<strong>on</strong>ses to scrapie infecti<strong>on</strong>. Taking into account the implicati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90 family in triggering<br />

stress resp<strong>on</strong>se under several envir<strong>on</strong>mental insults, and in protein degradati<strong>on</strong>, results presented here point<br />

to HSPCAA1 as a good positi<strong>on</strong>al and functi<strong>on</strong>al candidate gene modulating the resp<strong>on</strong>se to scrapie in<br />

sheep. Its possible importance in other amyloidosis modulati<strong>on</strong> shouldn’t be ruled out.<br />

6G_09_P<br />

(poster secti<strong>on</strong> B2, poster board #285, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ONE OF THE IMPLICATIONS OF HSP70 CHAPERONE ANTI-AGGREGATE<br />

ACTIVITY IN A CELL MODEL OF HUNTINGTON DISEASE<br />

Irina Guzhova, Anastasia Kaznacheeva, Maria Ippolitova, Alexander Kinev, Boris Margulis<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Cytology RAS, Tikhoretsky, 4, StPetersburg, Russia<br />

e-mail: margulis@mail.cytspb.rssi.ru<br />

Misfolded proteins and their aggregates is the cause <str<strong>on</strong>g>of</str<strong>on</strong>g> a great variety <str<strong>on</strong>g>of</str<strong>on</strong>g> pathologies including polyglutaminelinked<br />

diseases, like Huntingt<strong>on</strong>’s disease (HD). Since most <str<strong>on</strong>g>of</str<strong>on</strong>g> these diseases can be successfully treated by<br />

inducing Hsp70 chaper<strong>on</strong>e expressi<strong>on</strong> we have used the intrinsic property <str<strong>on</strong>g>of</str<strong>on</strong>g> the <str<strong>on</strong>g>for</str<strong>on</strong>g>mer to penetrate inside<br />

living cells in a cellular model <str<strong>on</strong>g>of</str<strong>on</strong>g> HD. We found that the incubati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> SK-N-SH human neuroblastoma cells<br />

overexpressing 103-glutamine expansi<strong>on</strong> with pure Hsp70 substantially lowered the amount <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptotic<br />

cells. This protecti<strong>on</strong> was attributed to the anti-aggregate effect <str<strong>on</strong>g>of</str<strong>on</strong>g> the chaper<strong>on</strong>e that was proved by reduced<br />

number and size <str<strong>on</strong>g>of</str<strong>on</strong>g> aggresomes, especially when the chaper<strong>on</strong>e has been applied prior to the aggregate<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>. At the initial stage aggregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> polyglutamine repeats is thought to be accompanied with the<br />

latter cross-linking to lysines <str<strong>on</strong>g>of</str<strong>on</strong>g> certain cellular molecules in a process catalyzed by tissue transglutaminse<br />

(tTG). One <str<strong>on</strong>g>of</str<strong>on</strong>g> the d<strong>on</strong>ors <str<strong>on</strong>g>of</str<strong>on</strong>g> such lysines is glyceraldehydephosphate dehydrogenase (GAPDH), and since<br />

the enzyme was shown to specifically bind Hsp70, we suggested that the chaper<strong>on</strong>e can prevent aggregati<strong>on</strong><br />

by depriving tTG <str<strong>on</strong>g>of</str<strong>on</strong>g> the substrate. Using the same cell model <str<strong>on</strong>g>of</str<strong>on</strong>g> HD we found that Hsp70 and GAPDH are<br />

co-localized with polyglutamine aggregates. Furthermore, the chaper<strong>on</strong>e was shown to physically interact with<br />

GAPDH in Hsp70-overexpressing cells. It was found <str<strong>on</strong>g>for</str<strong>on</strong>g> the first time that the up-regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70<br />

c<strong>on</strong>tent was accompanied with the reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> GAPDH quantity to be sequestered by SDS-insoluble<br />

aggresomes. In c<strong>on</strong>clusi<strong>on</strong> exogenously administered Hsp70 can prevent aggregate <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> in cells<br />

overexpressing l<strong>on</strong>g polyglutamine tracts and there<str<strong>on</strong>g>for</str<strong>on</strong>g>e the chaper<strong>on</strong>e itself serves as a potent antineurodegenerative<br />

drug.<br />

362


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

6G_10_P<br />

(poster secti<strong>on</strong> B2, poster board #286, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

FORMATION OF HIGHLY TOXIC AMYLOID BETA OLIGOMERS IS MEDIATED BY<br />

MOLECULAR CHAPERONES<br />

Masafumi Sak<strong>on</strong>o 1,2 *, Tamotsu Zako 1 , Hiroshi Ueda 3 , Masafumi Yohda 4 , Mizuo Maeda 1<br />

1 RIKEN, 2 JST, 3 Univ. Tokyo, 4 Tokyo Univ. Agric. Tech.<br />

*RIKEN Bioengineering Lab., 2-1, Hirosawa, Wako, Saitama 351-0198, Japan<br />

e-mail: msak<strong>on</strong>o@riken.jp<br />

Since the amyloid beta (Aβ) aggregates are the cause <str<strong>on</strong>g>of</str<strong>on</strong>g> Alzheimer's disease, studies <strong>on</strong> its <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong><br />

mechanism is important to develop the methods <str<strong>on</strong>g>for</str<strong>on</strong>g> preventi<strong>on</strong> and treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> Alzheimer's disease. In this<br />

report, we studied the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> a molecular chaper<strong>on</strong>e, prefoldin (PFD) from Pyrococcus horikoshii OT3 <strong>on</strong><br />

Aβ aggregati<strong>on</strong>. Lyophilized Aβ peptide (2 mg/ml) was dissolved in HFIP. The solvent was then dried using<br />

a spin-vacuum system, and stored in -80 °C. The HFIP-treated peptide was dissolved in distilled water by<br />

vortex. After s<strong>on</strong>icati<strong>on</strong>, 1 mM peptide soluti<strong>on</strong> was diluted to 50 µM in PBS buffer with or without 50 µM<br />

PFD, and incubated at 50 °C. Transmissi<strong>on</strong> electr<strong>on</strong> microscope (TEM) observati<strong>on</strong> and Western blotting<br />

analysis showed that the soluble oligomeric (3-50mer) Aβ particles (typically about 10 nm in diameter) were<br />

produced by incubati<strong>on</strong> with PFD, while fibrillar aggregates were produced in the absence <str<strong>on</strong>g>of</str<strong>on</strong>g> PFD. The<br />

cytotoxicity <str<strong>on</strong>g>of</str<strong>on</strong>g> Aβ aggregates against PC12 cells (rat pheochromocytoma cells) was measured by MTT<br />

method and terminal deoxynucletidyl transferase-mediated biotin-dUTP nick end labeling (TUNEL) method.<br />

The Aβ soluble oligomer <str<strong>on</strong>g>for</str<strong>on</strong>g>med by the incubati<strong>on</strong> with PFD exhibited higher toxicity than the fibrillar<br />

aggregates without PFD. This finding is c<strong>on</strong>sistent with the recent results that soluble oligomers, the<br />

intermediates <str<strong>on</strong>g>of</str<strong>on</strong>g> amyloid fibrillati<strong>on</strong>, have higher toxicity than amyloid fibril. We also c<strong>on</strong>firmed that the<br />

soluble β soluble oligomer induced apoptosis. Our data suggest that PFD recognizes these toxic soluble<br />

oligomers and prevents further fibrillati<strong>on</strong>.<br />

6G_11_P<br />

(poster secti<strong>on</strong> B2, poster board #287, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

IMPAIRMENT OF THE UBIQUITIN-PROTEASOME SYSTEM ASSOCIATED WITH<br />

EXTRACELLULAR TRANSTHYRETIN AGGREGATES IN FAMILIAL<br />

AMYLOIDOTIC POLYNEUROPATHY<br />

S<str<strong>on</strong>g>of</str<strong>on</strong>g>ia Duque Santos<br />

Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Molecular and Cell Biology, Porto, Portugal<br />

e-mail: sasantos@ibmc.up.pt<br />

The mechanisms developed by cells to acquire protecti<strong>on</strong> from the deleterious effects <str<strong>on</strong>g>of</str<strong>on</strong>g> misfolded proteins<br />

are currently well characterized. One essential mechanism is the ubiquitin-proteasome system (UPS) which<br />

has been associated with neurodegenerative disorders <str<strong>on</strong>g>of</str<strong>on</strong>g> intracellular protein aggregati<strong>on</strong>.<br />

We have studied the UPS in familial amyloidotic polyneuropathy (FAP), a neurodegenerative disorder caused<br />

by extracellular depositi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> mutant transthyretin (TTR). The studies were c<strong>on</strong>ducted in TTR synthesizing<br />

and n<strong>on</strong>-synthesizing tissues from affected individuals, in transgenic mice models <str<strong>on</strong>g>for</str<strong>on</strong>g> FAP and in<br />

neuroblastoma or Schwannoma cell lines cultured with TTR oligomers. We observed that in human FAP<br />

tissues, presenting extracellular TTR aggregates ubiquitin protein c<strong>on</strong>jugates were up-regulated, the<br />

proteasome levels were decreased and parkin and alpha-synuclein expressi<strong>on</strong>s were both decreased. On the<br />

363


23-26 August 2007,<br />

Budapest, Hungary<br />

other hand, the liver, that normally synthesizes variant TTR V30M, did not show this resp<strong>on</strong>se. A similar<br />

resp<strong>on</strong>se was detected in mice models <str<strong>on</strong>g>for</str<strong>on</strong>g> TTR V30M or TTR L55P. Furthermore, transgenic mice<br />

immunized to decrease TTR depositi<strong>on</strong> showed a significant decrease in ubiquitin levels and an increase in<br />

parkin and alpha-synuclein levels in comparis<strong>on</strong> to c<strong>on</strong>trol mice. Studies per<str<strong>on</strong>g>for</str<strong>on</strong>g>med in cell lines with<br />

aggregates in medium resulted in increased ubiquitin levels.<br />

The overall results are indicative <str<strong>on</strong>g>of</str<strong>on</strong>g> TTR depositi<strong>on</strong> as an external stimulus to an intracellular UPS resp<strong>on</strong>se<br />

in FAP.<br />

364<br />

6G_12_P<br />

(poster secti<strong>on</strong> B2, poster board #288, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ENDOPLASMIC RETICULUM STRESS IN ALZHEIMER’S DISEASE<br />

W. Scheper, J. J. M. Hoozemans, S. M. Chafekar, A. van Aken, M. López-Cavanillas, R. Zwart,<br />

P. Eikelenboom, F. Baas<br />

Neurogenetics Laboratory and Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Neurology, Academic Medical Center, 1100 DE Amsterdam,<br />

The Netherlands<br />

e-mail: w.scheper@amc.uva.nl<br />

The unfolded protein resp<strong>on</strong>se (UPR) is a protein quality c<strong>on</strong>trol mechanism that initially protects the cells<br />

against endoplasmic reticulum (ER) stress caused by the toxicity <str<strong>on</strong>g>of</str<strong>on</strong>g> unfolded proteins. We have previously<br />

shown that the UPR is activated in neur<strong>on</strong>s in Alzheimer’s disease (AD) brain. In additi<strong>on</strong> we find<br />

upregulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> the trafficking protein Rab6, which may also be involved in protein quality<br />

c<strong>on</strong>trol. The levels <str<strong>on</strong>g>of</str<strong>on</strong>g> Rab6 show a very str<strong>on</strong>g correlati<strong>on</strong> with levels <str<strong>on</strong>g>of</str<strong>on</strong>g> the ER chaper<strong>on</strong>e BiP in AD brain.<br />

We find ER stress as well as high Rab6 levels in n<strong>on</strong>-tangle bearing neur<strong>on</strong>s early in AD pathogenesis,<br />

suggesting that it precedes the late Aβ (plaques) and tau pathology (tangle <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>). Because ER stress is a<br />

very early event in AD pathogenesis it is a potential therapeutic target.<br />

Accumulati<strong>on</strong> and aggregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> unfolded proteins is a major neuropathological characteristic <str<strong>on</strong>g>of</str<strong>on</strong>g> AD,<br />

however, no aggregates are found in the ER itself. This suggests that other processes, <str<strong>on</strong>g>for</str<strong>on</strong>g> example disturbed<br />

protein trafficking out <str<strong>on</strong>g>of</str<strong>on</strong>g> the ER are involved. We use cellular models <str<strong>on</strong>g>for</str<strong>on</strong>g> early Aβ and tau pathology to<br />

delineate mechanistic interacti<strong>on</strong>s with the ER stress resp<strong>on</strong>se, in order to prevent neurotoxicity. We find that<br />

early intermediates in Aβ aggregati<strong>on</strong> can induce a mild ER stress resp<strong>on</strong>se. In additi<strong>on</strong>, preliminary data<br />

indicate that increased expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> tau interacts with the ER stress resp<strong>on</strong>se as well. Our ultimate aim is to<br />

modulate the ER stress resp<strong>on</strong>se in AD to prevent or limit neur<strong>on</strong>al loss.<br />

6G_13_P<br />

(poster secti<strong>on</strong> B2, poster board #289, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

A DISEASE-CAUSING VARIANT OF SHORT-CHAIN ACYL-COA DEHYDROGENASE<br />

PROMOTES OXIDATIVE STRESS<br />

S. P. Schmidt 1 , T. J. Coryd<strong>on</strong> 2 , N. Gregersen 1<br />

1<str<strong>on</strong>g>Research</str<strong>on</strong>g> Unit <str<strong>on</strong>g>for</str<strong>on</strong>g> Molecular Medicine, Skejby Sygehus, Brendstrupgaardsvej 8200 Aarhus N, Denmark<br />

2Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Human Genetics, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Aarhus, Bartholin Building, 800 Aarhus C, Denmark<br />

e-mail: stinne@ki.au.dk<br />

Short-chain acyl-CoA dehydrogenase deficiency (SCADD) is a rare recessively inherited metabolic disorder,<br />

affecting the mitoch<strong>on</strong>drial β-oxidati<strong>on</strong>. Patients are usually presenting neuromuscular features such as<br />

developmental delay, hypot<strong>on</strong>ia, seizures, as well as a general failure to thrive. To study the pathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g>


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

the disease, transduced astrocytic cells stably expressing five different disease-associated SCAD protein<br />

variants were established. In the cl<strong>on</strong>ing process, the viability <str<strong>on</strong>g>of</str<strong>on</strong>g> cells expressing each <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> two severe<br />

SCAD variant proteins was severely reduced, compared with cells expressing the SCAD wild-type (wt)<br />

protein. One <str<strong>on</strong>g>of</str<strong>on</strong>g> these was the rare variati<strong>on</strong> 319C>T (Arg83Cys), which is unable to assemble into<br />

catalytically active SCAD tetramers, as well as having aggregati<strong>on</strong>al tendencies in vitro. Six out <str<strong>on</strong>g>of</str<strong>on</strong>g> six SCAD wt<br />

cell col<strong>on</strong>ies survived, whereas <strong>on</strong>ly four out <str<strong>on</strong>g>of</str<strong>on</strong>g> seven SCAD Arg83Cys col<strong>on</strong>ies survived col<strong>on</strong>y transfer. To<br />

investigate whether this variati<strong>on</strong> inflicts with the ability <str<strong>on</strong>g>of</str<strong>on</strong>g> the cell to overcome a stress-full situati<strong>on</strong>, cells<br />

expressing the wt or the 319C>T variati<strong>on</strong> was subjected to heat stress <str<strong>on</strong>g>of</str<strong>on</strong>g> 40°C, and the stress-resp<strong>on</strong>se was<br />

followed over a time period <str<strong>on</strong>g>of</str<strong>on</strong>g> 24 hours, m<strong>on</strong>itored by selected stress resp<strong>on</strong>se genes (Hsp70, Hsp60 and<br />

HO-1 (Hemeoxygenase-1)). The cell line expressing the SCAD Arg83Cys variant protein revealed an elevated<br />

producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HO-1 compared with the SCAD wt cells, indicating oxidative stress, elicited by the misfolded<br />

mitoch<strong>on</strong>drial SCAD variant protein.<br />

6G_14_P<br />

(poster secti<strong>on</strong> B2, poster board #290, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CHARACTERIZING THE STRESS RESPONSE TO DIFFERENT MODELS OF<br />

PROTEIN MISFOLDING<br />

Mehmet Alper Arslan, Balázs Dancsó, Christopher D. Link, Péter Csermely, Csaba Sőti<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Chemistry, Semmelweis University, Budapest, POBox 260, Hungary<br />

e-mail: alpera@alumni.bilkent.edu.tr<br />

Ageing is characterized by a progressive accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cytotoxic and proteotoxic damage caused by<br />

envir<strong>on</strong>mental stress. Molecular chaper<strong>on</strong>es are ubiquitous, highly c<strong>on</strong>served proteins resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

maintenance <str<strong>on</strong>g>of</str<strong>on</strong>g> protein folding homeostasis in cells. Damaged, misfolding proteins are readily recognized by<br />

the complex chaper<strong>on</strong>e system in cells and are either refolded/repaired at the level <str<strong>on</strong>g>of</str<strong>on</strong>g> c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> or<br />

targeted predominantly <str<strong>on</strong>g>for</str<strong>on</strong>g> the proteasomal degradati<strong>on</strong> if the damage is irreparable. Damaged proteins<br />

accumulate in postmitotic cells, and in cells harboring <str<strong>on</strong>g>of</str<strong>on</strong>g> severe inheritable mutati<strong>on</strong>s (such as polyQ<br />

proteins) and may have pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ound cytotoxic effects, including transcripti<strong>on</strong> factor deactivati<strong>on</strong> and<br />

cytoskeletal derangements, apoptosis inducti<strong>on</strong>. These pathologies have wide clinical implicati<strong>on</strong>s from<br />

neurodegenerati<strong>on</strong> to aging.<br />

To better understand the role <str<strong>on</strong>g>of</str<strong>on</strong>g> protein structure and the role <str<strong>on</strong>g>of</str<strong>on</strong>g> molecular chaper<strong>on</strong>es in these processes,<br />

we have overexpressed several protein-misfolding models in Cos-7 cells and currently study there aggregati<strong>on</strong><br />

properties and the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress resp<strong>on</strong>se by sedimentati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> detergent insoluble fracti<strong>on</strong>s, western<br />

blotting and immun<str<strong>on</strong>g>of</str<strong>on</strong>g>luorescence analysis. Of these, GFP::degr<strong>on</strong>, a C-terminal fusi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a 16-residue<br />

“degr<strong>on</strong>” peptide to GFP, was found to <str<strong>on</strong>g>for</str<strong>on</strong>g>m iuxtanuclear aggregates resembling to aggresomes and<br />

markedly induced the major stress protein Hsp70. Results <str<strong>on</strong>g>of</str<strong>on</strong>g> our further <strong>on</strong>going studies will be presented.<br />

365


23-26 August 2007,<br />

Budapest, Hungary<br />

366<br />

6H. STRESS IN THE DEVELOPMENT OF SCHIZOPHRENIA<br />

(KUNIO YUI, MICHIO SUZUKI)<br />

6H_01_S<br />

THE NEURAL BASES OF NEGATIVE AFFECT AND STRESS RESPONSES<br />

IN SCHIZOPHRENIA<br />

Ute Habel<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychiatry and Psychotherapy, RWTH Aachen University<br />

e-mail: uhabel@ukaachen.de<br />

Stress has a major impact <strong>on</strong> the course and psychopathology <str<strong>on</strong>g>of</str<strong>on</strong>g> schizophrenia, having significant influences<br />

especially <strong>on</strong> emoti<strong>on</strong> but also cognitive processes. In schizophrenia, emoti<strong>on</strong> dysfuncti<strong>on</strong>s are a hallmark<br />

with a special role <str<strong>on</strong>g>of</str<strong>on</strong>g> negative affect and reduced capacities to manage stressful situati<strong>on</strong>s and stimulati<strong>on</strong>s.<br />

Data acquired from different samples <str<strong>on</strong>g>of</str<strong>on</strong>g> healthy subjects and schizophrenia patients will be presented to<br />

clarify the neural basis <str<strong>on</strong>g>of</str<strong>on</strong>g> dysfuncti<strong>on</strong>al emoti<strong>on</strong> processes applying fMRI. Emoti<strong>on</strong> and cogniti<strong>on</strong> are mostly<br />

investigated as different entities, while practically both functi<strong>on</strong>s are inseparable and are interacting with each<br />

other c<strong>on</strong>tinuously. However, interacti<strong>on</strong>s between emoti<strong>on</strong> and cogniti<strong>on</strong> have not been investigated in<br />

greater detail in patients. Similarly, in healthy pers<strong>on</strong>s there are <strong>on</strong>ly a few and rather c<strong>on</strong>tradictory studies<br />

exploring the general effects <str<strong>on</strong>g>of</str<strong>on</strong>g> emoti<strong>on</strong> <strong>on</strong> cognitive functi<strong>on</strong>. We developed and validated tasks<br />

investigating the interplay between emoti<strong>on</strong>s and cogniti<strong>on</strong>. Stress was induced in patients and comparis<strong>on</strong><br />

subjects by means <str<strong>on</strong>g>of</str<strong>on</strong>g> negative olfactory stimulati<strong>on</strong> while subjects had to per<str<strong>on</strong>g>for</str<strong>on</strong>g>m a combined 0-back/2-back<br />

task. A c<strong>on</strong>trol c<strong>on</strong>diti<strong>on</strong> with neutral room air stimulati<strong>on</strong> was also applied. According to subjective ratings<br />

mood inducti<strong>on</strong> was successful. The impairing effect <str<strong>on</strong>g>of</str<strong>on</strong>g> the negative mood inducti<strong>on</strong> was visible during<br />

working memory per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance (2-back) <strong>on</strong>ly, in which the mean number <str<strong>on</strong>g>of</str<strong>on</strong>g> correct target reacti<strong>on</strong>s was<br />

significantly lower during negative olfactory stimulati<strong>on</strong> as compared to the neutral c<strong>on</strong>diti<strong>on</strong>. The<br />

neuroimaging data reveal a complex dysfuncti<strong>on</strong>al interacti<strong>on</strong> in patients, where especially fr<strong>on</strong>tal and<br />

cingular regi<strong>on</strong>s show aberrant activati<strong>on</strong> patterns. Hence, regi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> major importance in emoti<strong>on</strong> regulati<strong>on</strong><br />

and integrati<strong>on</strong> are affected.<br />

6H_02_S<br />

STRESS, DOPAMINE, BRAIN TISSUE VOLUMES AND VULNERABILITY<br />

TO PSYCHOSIS<br />

Machteld Marcelis, E. Cavalier, John Suckling, Paul H<str<strong>on</strong>g>of</str<strong>on</strong>g>man, Philippe Delespaul, Jim van Os<br />

Dept <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychiatry & Neuropsychology, POBox 616, 6200 MD, Maastricht University, The Netherlands<br />

e-mail: m.marcelis@sp.unimaas.nl<br />

The present study investigated whether individuals with schizophrenia, and those with an elevated genetic risk<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> schizophrenia, display an altered plasma HVA-resp<strong>on</strong>se to metabolic stress. Besides, associati<strong>on</strong>s between<br />

the metabolic stress resp<strong>on</strong>se and brain tissue volumes were examined. Patients with psychosis (n=50), n<strong>on</strong>psychotic<br />

first-degree relatives <str<strong>on</strong>g>of</str<strong>on</strong>g> patients with psychosis (n=51) and c<strong>on</strong>trols (n=50) underwent, in<br />

randomised order, double-blind administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> placebo and the glucose analog 2-deoxy-D-glucose (2DG),<br />

which induces a mild, transient clinical state <str<strong>on</strong>g>of</str<strong>on</strong>g> glucoprivati<strong>on</strong>. Plasma HVA was assessed twice be<str<strong>on</strong>g>for</str<strong>on</strong>g>e the<br />

start <str<strong>on</strong>g>of</str<strong>on</strong>g> the 2DG/placebo infusi<strong>on</strong> (baseline), as well as four times post infusi<strong>on</strong>. MRI cerebral tissue volumes<br />

were derived from automated segmentati<strong>on</strong> procedures. During the stress c<strong>on</strong>diti<strong>on</strong>, significant increases in


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

plasma HVA were found. The increase in plasma HVA level during the stress c<strong>on</strong>diti<strong>on</strong> was significantly<br />

str<strong>on</strong>ger in patients than in c<strong>on</strong>trols, whereas this was not the case in relatives v. c<strong>on</strong>trols. The HVA level<br />

increase in the stress c<strong>on</strong>diti<strong>on</strong> was str<strong>on</strong>ger in patients with lower grey and white matter volumes. In<br />

c<strong>on</strong>clusi<strong>on</strong>, patients with psychosis, but not their n<strong>on</strong>-psychotic relatives, show an altered<br />

dopaminergic/noradrenergic mediated stress resp<strong>on</strong>se, possibly reflecting acquired sensitizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

catecholamine systems by repeated envir<strong>on</strong>mental stressors or repeated stimulati<strong>on</strong> with ag<strong>on</strong>istic drugs.<br />

HVA level increases were str<strong>on</strong>ger in patients with lower grey and white matter volumes, supporting the<br />

hypothesis that alterati<strong>on</strong>s in cortico-subcortical c<strong>on</strong>necti<strong>on</strong>s affect psychosis susceptibility through an altered<br />

stress resp<strong>on</strong>se.<br />

6H_03_S<br />

STRUCTURAL BRAIN CHANGES UNDERLYING VULNERABILITY<br />

TO SCHIZOPHRENIA<br />

Michio Suzuki 1,2 , Tsutomu Takahashi 1,2 , Shi-Yu Zhou 1,2,3 , Yasuhiro Kawasaki 1,2 ,<br />

Masayoshi Kurachi 1,2<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Neuropsychiatry, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Toyama, 2630 Sugitani, Toyama 930-0194, Japan,<br />

2CREST, JST, Tokyo, Japan, 3 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychiatry and Medical Psychology, Dalian Medical University,<br />

Dalian, China<br />

Morphologic changes in schizophrenia are thought to represent a complex and dynamic process in which<br />

multiple brain regi<strong>on</strong>s are differentially affected. Comm<strong>on</strong> neurobiological abnormalities am<strong>on</strong>g the<br />

schizophrenia-spectrum may be essential <str<strong>on</strong>g>for</str<strong>on</strong>g> the pathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> schizophrenia, but some additi<strong>on</strong>al<br />

pathological changes may also be required <str<strong>on</strong>g>for</str<strong>on</strong>g> the development <str<strong>on</strong>g>of</str<strong>on</strong>g> full-blown schizophrenia. Clarifying the<br />

neurobiological similarities and differences between established schizophrenia and schizotypal (pers<strong>on</strong>ality)<br />

disorder, a schizophrenia-spectrum disorder without manifestati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> overt and sustained psychosis, would<br />

potentially discriminate the pathophysiological mechanisms underlying the vulnerability to schizophrenia<br />

from those associated with overt psychosis. Detailed volumetric comparis<strong>on</strong>s using magnetic res<strong>on</strong>ance<br />

imaging (MRI) revealed differential morphologic alterati<strong>on</strong>s in the brain between the patients with schizotypal<br />

disorder and those with schizophrenia. Volume reducti<strong>on</strong>s in the amygdala, hippocampus, superior temporal<br />

gyrus, and anterior parietal cortex comm<strong>on</strong> to both patient groups may represent the vulnerability to<br />

schizophrenia. Volume loss <str<strong>on</strong>g>of</str<strong>on</strong>g> the prefr<strong>on</strong>tal cortex, posterior parietal cortex, cingulate, insula, and fusi<str<strong>on</strong>g>for</str<strong>on</strong>g>m<br />

cortex preferentially observed in schizophrenia may be critical <str<strong>on</strong>g>for</str<strong>on</strong>g> overt manifestati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> psychosis. On the<br />

other hand, preserved volume in these regi<strong>on</strong>s might have relevance to the protecti<strong>on</strong> factors from overt<br />

psychosis in schizotypal disorder.<br />

6H_04_S<br />

EARLY EMOTIONAL STRESS BY ANALYSIS OF STRESS LINE IN MOLAR AND<br />

SUSCEPTIBILITY TO SCHIZOPHRENIA<br />

Kunio Yui 1, 2 , Koichi Watanabe 3 , Masayoshi Kobayashi 3 , Koichi Nishijima 4<br />

1Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Human Science, Kansai University <str<strong>on</strong>g>of</str<strong>on</strong>g> Internati<strong>on</strong>al Studies<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychiatry and Neurology, Kobe University, Graduate School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine<br />

3X-ray Microanalyzer secti<strong>on</strong>, Center <str<strong>on</strong>g>for</str<strong>on</strong>g> Instrumental Analysis, Niigata University<br />

4Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychiatry, Jichi Medical School<br />

367


23-26 August 2007,<br />

Budapest, Hungary<br />

[Introducti<strong>on</strong>]: It is well known that early life events or urbanicity, that interacts with multiple genes, induces<br />

persistent sensitizati<strong>on</strong> to stress possibly through an imbalance in interacti<strong>on</strong>s between dopaminergic and<br />

glutamatergic systems. This stress sensitizati<strong>on</strong> may be critical in the development or relapse <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

schizophrenia. Laboratory method <str<strong>on</strong>g>for</str<strong>on</strong>g> estimating early stress is there<str<strong>on</strong>g>for</str<strong>on</strong>g>e needed. Ameloblast activity in<br />

human molar’s enamel is slowed during 1-2 days extreme stress, and the segment <str<strong>on</strong>g>of</str<strong>on</strong>g> enamel rods is smaller<br />

and <str<strong>on</strong>g>of</str<strong>on</strong>g>ten misshapen, making a particular dark line seen by microscopy, which we called Pathological Stress<br />

Line (PSL). We studied the nature <str<strong>on</strong>g>of</str<strong>on</strong>g> stress sensitizati<strong>on</strong> by analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> PSL. According to animal and clinical<br />

studies, severe emoti<strong>on</strong>al stress induce changes <str<strong>on</strong>g>of</str<strong>on</strong>g> mineral density in b<strong>on</strong>e. Thus, to clarify the type <str<strong>on</strong>g>of</str<strong>on</strong>g> stress<br />

related to PSL, we examine mineral changes in PSL porti<strong>on</strong>s.<br />

[Methods]: We examined PSL in third molar in 35 chr<strong>on</strong>ic paranoid schizophrenics - 25 males and 10<br />

females, 41.9±13.5 years old - and 32 normal c<strong>on</strong>trols (5 males and 27 female, 28.3±9.1 years old). Changes<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> density in potassium (P), calcium (Ca) and magnesium (Mg) in PSL porti<strong>on</strong> were examined by Scanning<br />

Microscope and Electr<strong>on</strong> Probe Microanalyser (EPMA). Since the rate <str<strong>on</strong>g>of</str<strong>on</strong>g> enamel el<strong>on</strong>gate is well known, PSL<br />

was assessed due to its length and definiti<strong>on</strong> at half yearly between 9 to 13 years old: 0, n<strong>on</strong>e; 1, slight; 2, mild;<br />

3, moderate; 4, severe. Mineral changes were assessed in PSL porti<strong>on</strong>s rated as 2-4 due to extent <str<strong>on</strong>g>of</str<strong>on</strong>g> changes<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> mineral density in <strong>on</strong>e area (1.5 X 1.5 mm): 0, n<strong>on</strong>e; 1, 1/4; 2, 1/3; 3, 1/2, 4, 2/3.<br />

[Results]: PSL scores in the 35 schizophrenics were significantly higher than the 32 normal c<strong>on</strong>trols (4.8±5.1<br />

vs. 2.0±2.3, P


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Brattleboro rats (e.g. oxytocin, CRH). Moreover both the adrenomedullary system and the sympathetic nerves<br />

are more active in vasopressin deficient rats. So our c<strong>on</strong>clusi<strong>on</strong> is that vasopressin may act <strong>on</strong> the<br />

development <str<strong>on</strong>g>of</str<strong>on</strong>g> schizophrenia through influencing other neurotransmitters in brain and not the HPA axis.<br />

Chr<strong>on</strong>ic stress may exacerbate the schizophrenia not through HPA axis changes but e.g. glucocorticoid<br />

toxicity in hippocampus.<br />

6H_06_S<br />

(poster secti<strong>on</strong> B1, poster board #206, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

NUTRITIONAL METABOLIC STRESS AND EPIGENETIC MECHANISM OF<br />

SCHIZOPHRENIA<br />

Sahebarao Mahadik<br />

Medical College <str<strong>on</strong>g>of</str<strong>on</strong>g> Georgia, Medical <str<strong>on</strong>g>Research</str<strong>on</strong>g>/Psychiatry and Health Behavior<br />

1 Freedom Way, 30904 Augusta, USA<br />

e-mail: smahadik@mail.mcg.edu<br />

Stress, both envir<strong>on</strong>mental (nutriti<strong>on</strong>, toxins, hypoxia) and emoti<strong>on</strong>al is ubiquitous and can c<strong>on</strong>tribute to<br />

epigenetic molecular mechanisms <str<strong>on</strong>g>for</str<strong>on</strong>g> a wide variety <str<strong>on</strong>g>of</str<strong>on</strong>g> n<strong>on</strong>-communicable diseases from birth to aging.<br />

Maternal as well as fetal nutriti<strong>on</strong>al stress (deficiency <str<strong>on</strong>g>of</str<strong>on</strong>g> micr<strong>on</strong>utrients or oxidative cell damage due to very<br />

high caloric intake) has been shown to imprint the risk <str<strong>on</strong>g>for</str<strong>on</strong>g> adolescent and adult n<strong>on</strong>-communicable disorders<br />

such as brain disorders, diabetes and cardiovascular disease. Am<strong>on</strong>g these, maternal folic acid and vitamin<br />

B12 deficiency has been c<strong>on</strong>sidered to c<strong>on</strong>tribute to the altered gene expressi<strong>on</strong> by altered <strong>on</strong>e carb<strong>on</strong><br />

metabolism and thereby methylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proteins, DNA and phospholipids, particularly c<strong>on</strong>taining omega-3<br />

fatty acids. Role <str<strong>on</strong>g>of</str<strong>on</strong>g> maternal nutriti<strong>on</strong> has been c<strong>on</strong>sidered in schizophrenia but evidence is based <strong>on</strong><br />

epidemiological data. We studied the plasma levels <str<strong>on</strong>g>of</str<strong>on</strong>g> folic acid, vitamin B12, cortisol and homocysteine, <strong>on</strong>e<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the most important factor in oxidative stress-mediated cellular dysfuncti<strong>on</strong>, particularly neural cell functi<strong>on</strong><br />

during brain development. The drug-naïve patients at the early <strong>on</strong>set <str<strong>on</strong>g>of</str<strong>on</strong>g> psychosis (N=28) and matched<br />

healthy c<strong>on</strong>trols (N=34) were enrolled in a study. All the study protocols were properly approved by the<br />

Instituti<strong>on</strong>al Ethical Committee and each study subject signed the c<strong>on</strong>sent to participate in the study.<br />

Compared to normal c<strong>on</strong>trols, patients had significantly (p


23-26 August 2007,<br />

Budapest, Hungary<br />

6H_01_P<br />

(poster secti<strong>on</strong> B1, poster board #207, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE EFFECT OF SUBCHRONIC KETAMINE TREATMENT AND SOCIAL<br />

ISOLATION ON PAIN SENSITIVITY<br />

G. Tuboly, G. Horvath, G. Benedek<br />

Dept. Physiol., Fac. Med., Univ. Szeged, Dom ter 10. Hungary<br />

e-mail: tuboly@phys.szote.u-szeged.hu<br />

Chr<strong>on</strong>ic exposure to stressful events precipitates or exacerbates many neuropsychiatric disorders, including<br />

schizophrenia, resulting in the stress hypothesis <str<strong>on</strong>g>of</str<strong>on</strong>g> schizophrenia. Clinical studies have proven that<br />

schizophrenia is accompanied by hypoalgesia. Relevant animal models are <str<strong>on</strong>g>of</str<strong>on</strong>g> decisive importance in the<br />

study <str<strong>on</strong>g>of</str<strong>on</strong>g> psychiatric diseases. It is well-known that subchr<strong>on</strong>ic treatment with ketamine or social isolati<strong>on</strong><br />

induces schizophrenia-related alterati<strong>on</strong>s, which are ameliorated by clinically used neuroleptics. The purpose<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to test nociceptive resp<strong>on</strong>ses in singly housed and/or subchr<strong>on</strong>ic ketamine treatment.<br />

After weaning <strong>on</strong> postantal day 21, male Wistar rat pups were either group housed (5-6; GR. I and II) or<br />

isolated (GR. III and IV) <str<strong>on</strong>g>for</str<strong>on</strong>g> 20 days. 6 days later (30 days old rats), animals (Gr. II and IV) were injected<br />

with 30 mg/kg ketamine daily <str<strong>on</strong>g>for</str<strong>on</strong>g> 14 days. C<strong>on</strong>trol rats (Gr. I. and III) received saline. The tail-flick latency<br />

tests were per<str<strong>on</strong>g>for</str<strong>on</strong>g>med at 41. day at 48 and 52 o C.<br />

At 48 o C we found a significant effect <str<strong>on</strong>g>of</str<strong>on</strong>g> housing c<strong>on</strong>diti<strong>on</strong>s, whereas ketamine treatment had no effect <strong>on</strong><br />

reacti<strong>on</strong> time. Both single housed groups <str<strong>on</strong>g>of</str<strong>on</strong>g> animals had significantly higher reacti<strong>on</strong> times compared with<br />

group-housed rats irrespective <str<strong>on</strong>g>of</str<strong>on</strong>g> ketamin treatment. In c<strong>on</strong>trast, at 52 o C there were no significant<br />

differences between the four groups.<br />

Our study suggests that isolati<strong>on</strong> has effect <strong>on</strong> acute heat pain sensitivity. Since low temperature activates<br />

mainly the C-fibers, while the high temperature the Aδ-fibers, we suppose that this type <str<strong>on</strong>g>of</str<strong>on</strong>g> stress disturbed<br />

primarily the C-fiber linked pain pathways.<br />

370


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

6I. MODERN MOLECULAR MECHANISMS AND THERAPEUTICS<br />

OF STRESS-RELATED GASTROINESTINAL DISORDERS<br />

(SÁNDOR SZABÓ, YVETTE TACHÉ)<br />

6I_01_S<br />

STRESS AND THE GUT: ROLE OF CORTICOTROPIN RELEASING FACTOR (CRF)<br />

SIGNALING PATHWAYS<br />

Y. Taché<br />

Center <str<strong>on</strong>g>for</str<strong>on</strong>g> Neurovisceral Sciences, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Cali<str<strong>on</strong>g>for</str<strong>on</strong>g>nia, Los Angeles, USA<br />

The characterizati<strong>on</strong> and distributi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> corticotropin-releasing factor (CRF) family <str<strong>on</strong>g>of</str<strong>on</strong>g> peptides, CRF,<br />

urocortin 1, urocortin 2 and urocortin 3, and the two G-protein coupled receptors, CRF 1 and CRF 2 , as well<br />

as the development <str<strong>on</strong>g>of</str<strong>on</strong>g> selective CRF 1 and CRF 2 receptor antag<strong>on</strong>ists provided novel means to understand<br />

mechanisms involve in the stress resp<strong>on</strong>se. The activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> brain CRF 1 signaling pathway plays a primary<br />

role in the endocrine (activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> pituitary adrenal axis), behavioral (anxiety, depressi<strong>on</strong>), aut<strong>on</strong>omic<br />

(sympathetic activati<strong>on</strong>), and decrease immune resp<strong>on</strong>ses to stress. Combined anatomical, pharmacologic and<br />

molecular approaches support a role <str<strong>on</strong>g>of</str<strong>on</strong>g> CRF receptor activati<strong>on</strong> in both the brain and the gut as part <str<strong>on</strong>g>of</str<strong>on</strong>g> key<br />

mechanisms involved in stress-related alterati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> gut propulsive functi<strong>on</strong>. Inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> gastric emptying<br />

and stimulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> col<strong>on</strong>ic motor functi<strong>on</strong> are the comm<strong>on</strong>ly encountered patterns resulting from exposure<br />

to various stressors in experimental animals and humans. Activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> brain and peripheral CRF 2 receptors<br />

mediates stress-related inhibiti<strong>on</strong> gastric motor functi<strong>on</strong> while that <str<strong>on</strong>g>of</str<strong>on</strong>g> CRF 1 receptors are involved in the<br />

stimulati<strong>on</strong> col<strong>on</strong>ic secretory and motor functi<strong>on</strong>s. Clinical investigati<strong>on</strong>s also support the noti<strong>on</strong> that stress<br />

c<strong>on</strong>tributes to visceral hypersensitivity <str<strong>on</strong>g>of</str<strong>on</strong>g> the gut observed in patients with irritable bowel syndrome (IBS) as<br />

established by their lowered threshold <str<strong>on</strong>g>of</str<strong>on</strong>g> pain to colorectal distensi<strong>on</strong> (CRD). Experimental models have<br />

been developed that recapture clinical features <str<strong>on</strong>g>of</str<strong>on</strong>g> IBS with regard to stress-related hyperalgesia to CRD,<br />

gender differences, comorbidity with anxiety/depressi<strong>on</strong> and altered bowel habit. Data obtained using<br />

pharmacologic approaches in rodents subjected to stress indicate that the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> CRF 1 receptor<br />

c<strong>on</strong>tributes to the development <str<strong>on</strong>g>of</str<strong>on</strong>g> hyperalgesia to CRD while CRF 2 receptors display opposite effects. The<br />

mechanisms through which CRF 1 antag<strong>on</strong>ists alleviate col<strong>on</strong>ic resp<strong>on</strong>ses involved the preventi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> stressrelated<br />

activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> sacral parasympathetic ouflow, col<strong>on</strong>ic enteric cholinergic neur<strong>on</strong>s and mast cells. The<br />

pre-clinical and clinical phase I data support that targeting <str<strong>on</strong>g>of</str<strong>on</strong>g> CRF 1 receptors may open new therapeutic<br />

venues <str<strong>on</strong>g>for</str<strong>on</strong>g> stress-related functi<strong>on</strong>al gastrointestinal disorders. Supported by NIHDDK and VA Merit grants.<br />

6I_02_S<br />

EFFECT OF CAPSAICIN ON THE ESOPHAGEAL MOTILITY OF PATIENTS WITH<br />

GASTRO-ESOPHAGEAL REFLUX DISEASE (GERD)<br />

Agnes Király<br />

3rd Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Pécs, Hungary<br />

e-mail: agnes.kiraly@aok.pte.hu<br />

Capsaicin-sensitive afferents play a role in the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> esophageal motility. Vanilloid receptor is activated<br />

by acidic pH which triggers pain and motor resp<strong>on</strong>ses leading to esophageal emptying . The aim was to<br />

investigate the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> topical capsaicin (Tabasco) suspensi<strong>on</strong> <strong>on</strong> esophageal sensati<strong>on</strong> and motility in<br />

healthy c<strong>on</strong>trols (N=10), patients with n<strong>on</strong>-erosive GERD (NERD) (N=10), erosive esophagitis (ERD)<br />

371


23-26 August 2007,<br />

Budapest, Hungary<br />

(N=10) and with Barrett’s metaplasia (BM) (N=10). Visual analog scale was used to determine sensati<strong>on</strong>, the<br />

esophageal body and lower esophageal sphincter (LES) resp<strong>on</strong>se were analyzed. Topical 1.5 x 10 -4 M<br />

capsaicin significantly increased the amplitudes (68 +/- 3 to 88.5 +/- 4.7 ; 66+/-2 to 80+/-3; 70+/-1 to<br />

96+/-4 mm Hg, P < 0.05) and propagati<strong>on</strong> velocity (2.7 +/- 0.3 to 4.3 +/- 0.3 ; 3.1+/-0.3 to 4.2+/-0.1;<br />

2.5+/-0.1 to 3.7+/-0.2 cm/s, P < 0.05) <str<strong>on</strong>g>of</str<strong>on</strong>g> esophageal pressure waves and LES pressure (15 +/- 1.4 to 27<br />

+/- 2.6; 13+/-1 to 20+/-1; 14+/-1 to 29+/-3 mm Hg, P < 0.05) in c<strong>on</strong>trols, NERD and ERD patients<br />

respectively. N<strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the above resp<strong>on</strong>ses were present in BM patients. Capsaicin enhanced esophageal bolus<br />

transit (3.3+/-0.1 to 2.7+/-0.1; 3.2+/- 0.1 to 2.5 +/- 0.1 s, P < 0.05) in c<strong>on</strong>trols and NERD patients.<br />

Unchanged bolus clearance was found in ERD and BM patients (4.0+/-0.1 vs. 4.1+/-0.1; 4.2+/- 0.2 vs. 3.9<br />

+/-0.1 s NS). Significantly increased percepti<strong>on</strong> to capsaicin was found in NERD (66+/-7) and ERD<br />

(78+/-9) patients vs. to healthy c<strong>on</strong>trols (47+/-3) mm. Esophageal capsaicin induced a pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ound increase in<br />

emptying which could improve clearance <str<strong>on</strong>g>of</str<strong>on</strong>g> the esophagus. Chr<strong>on</strong>ic acid exposure produces allodynia in<br />

NERD and ERD patients which is related to their symptom development. Capsaicin-mediated esophageal<br />

clearance is disturbed in ERD and BM patients.<br />

6I_03_S<br />

STRESS AT THE CELLULAR LEVEL – NOVEL ROLES OF HEAT SHOCK<br />

PROTEINS IN GASTRIC MUCOSAL PROTECTION AND HEALING<br />

A. S. Tarnawski, H. Gergely<br />

VALBHS & Univ <str<strong>on</strong>g>of</str<strong>on</strong>g> Cali<str<strong>on</strong>g>for</str<strong>on</strong>g>nia, Irvine, CA, USA<br />

e-mail: atarnawski@yahoo.com<br />

Cells resp<strong>on</strong>d to stress by activating heat shock protein (HSP) resp<strong>on</strong>se, which c<strong>on</strong>stitutes an universal<br />

defense system, essential <str<strong>on</strong>g>for</str<strong>on</strong>g> maintaining cell and tissue homeostasis by stabilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> denatured proteins.<br />

In gastric mucosa, the surface epithelial cells are directly exposed to temperatures ranging from 4°C to 75°C,<br />

there<str<strong>on</strong>g>for</str<strong>on</strong>g>e HSP activati<strong>on</strong> is very relevant. We examined expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> c<strong>on</strong>stitutive and inducible HSP70<br />

(c and iHSP70) in gastric mucosa <str<strong>on</strong>g>of</str<strong>on</strong>g> rats at baseline and following treatment with cytoprotective drugs:<br />

sucralfate, rebamipide, and talcid. In normal gastric mucosa cHSP70 is expressed mainly in the surface<br />

epithelium. Cytoprotective drugs enhanced expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cHSP70 in surface epithelial and progenitor cells<br />

induced iHSP70 in the same areas and significantly reduced ethanol-induced gastric mucosal injury. Ethanolinduced<br />

gastric injury is significantly increased in mice with KO gene <str<strong>on</strong>g>for</str<strong>on</strong>g> heat shock factor 1 (HSF-1) a<br />

transcripti<strong>on</strong> factor <str<strong>on</strong>g>for</str<strong>on</strong>g> HSP genes. This is due to increased apoptosis in resp<strong>on</strong>se to ethanol injury in HSF-1<br />

null mice. This study provides evidence that HSPs after their HSF-1-dependent upregulati<strong>on</strong> protect gastric<br />

mucosa against injury. Recent data indicate that HSPs are involved in healing <str<strong>on</strong>g>of</str<strong>on</strong>g> experimental gastric ulcers.<br />

HSP32 are elevated during early stage <str<strong>on</strong>g>of</str<strong>on</strong>g> healing; HSP47 are increased in granulati<strong>on</strong> tissue and HSP70 is<br />

increased in the epithelial cell <str<strong>on</strong>g>of</str<strong>on</strong>g> the ulcer margin and progenitor cells where it co localized to EGF, IGF-1<br />

and Cox2. C<strong>on</strong>clusi<strong>on</strong>s: 1) Gastric mucosa expressed HSP at baseline and their expresses is activated by<br />

cytoprotective drugs and in resp<strong>on</strong>se to injury. 2) HSPs also play important roles in ulcer healing by local<br />

interacti<strong>on</strong>s with growth factors and by protecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> regenerating cells.<br />

372


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

6I_04_S<br />

INCREASED EXPRESSION OF TRANSCRIPTION FACTOR EGR-1 IN STRESS-<br />

INDUCED GASTRIC ULCERATION<br />

T. Beregova 1 , V. Kucharskyij 1 , O. Drobinska 1 , S. Szabo 2<br />

1Taras Shevchenko Kyiv Nati<strong>on</strong>al University, Ukraine; 2 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Phatology and Pharmacology,<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Cali<str<strong>on</strong>g>for</str<strong>on</strong>g>nia, Irvine, VA L<strong>on</strong>g Beach Healthcare System, CA, USA<br />

Revutskogo str., 44, apt.86, Kyiv, Ukraine 02140,<br />

e-mail: tberegova@univ.kiev.ua<br />

The transcripti<strong>on</strong> factor early growth resp<strong>on</strong>se-1 (Egr-1) is activated by many envir<strong>on</strong>mental signals. Recently<br />

it was dem<strong>on</strong>strated the enhanced duodenal Egr-1 transcripti<strong>on</strong> activity and followed increased expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

target genes such as bFGF, PDGF and VEGF in cysteamine-induced duodenal ulcerati<strong>on</strong> in rats. However,<br />

the involvement <str<strong>on</strong>g>of</str<strong>on</strong>g> Egr-1 expressi<strong>on</strong> in stress-induced gastric ulcerati<strong>on</strong> was not determined. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> our<br />

study was to investigate the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> stress <strong>on</strong> Egr-1 expressi<strong>on</strong> in rat gastric mucosa (GM) at the different<br />

time point. The rats were immobilized <str<strong>on</strong>g>for</str<strong>on</strong>g> 6, 12 and 24 hr. Egr-1 expressi<strong>on</strong> was determined by<br />

immunoblotting. It was established that 6 hr <str<strong>on</strong>g>of</str<strong>on</strong>g> stress exposure induced <strong>on</strong>ly single punctated hemorragies in<br />

GM. 12 hr <str<strong>on</strong>g>of</str<strong>on</strong>g> stress produced the multiply punctated hemorragies. 24 hr <str<strong>on</strong>g>of</str<strong>on</strong>g> stress exposure developed ulcers,<br />

errosi<strong>on</strong>s and massive hemorragies in rat stomach. The gastric injuries were 23,01±4,07 mm 2 . Egr-1<br />

expressi<strong>on</strong> was barely detectable in GM <str<strong>on</strong>g>of</str<strong>on</strong>g> untreated rats. 6 hours <str<strong>on</strong>g>of</str<strong>on</strong>g> stress increased the gastric Egr-1<br />

expressi<strong>on</strong> by 8 fold (p


23-26 August 2007,<br />

Budapest, Hungary<br />

Thus, the initial injury to endothelial & epithelial cells in DU seems to be aggravated (& not initiated) by HCl<br />

& proteolytic enzymes. The resulting mucosal necrosis does not rapidly heal because <str<strong>on</strong>g>of</str<strong>on</strong>g> the imbalance <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

VEGF & angiostatin/endostatin, hence duodenal ulcers develop. The experimental ulcers Selye described<br />

morphologically are now characterized at the molecular & genome level, involving unexpected mediators like<br />

ET-1, egr-1 & angiogenesis-related molecules.<br />

6I_06_S<br />

NEW MOLECULAR MECHANISMS OF ULCERATIVE COLITIS/IBD: DOES<br />

STRESS MATTER<br />

Zs. Sandor, G. Tolstanova, X. M. Deng, L. Chen, T. Khomenko, S. Szabo<br />

VA Med. Cent. & Univ. <str<strong>on</strong>g>of</str<strong>on</strong>g> Cali<str<strong>on</strong>g>for</str<strong>on</strong>g>nia-Irvine, Sch. <str<strong>on</strong>g>of</str<strong>on</strong>g> Med., L<strong>on</strong>g Beach, CA, USA<br />

Recent experimental & clinical studies indicate that adverse life events, chr<strong>on</strong>ic stress, & depressi<strong>on</strong> increase<br />

the likelihood <str<strong>on</strong>g>of</str<strong>on</strong>g> relapse in patients with quiescent IBD. Phsycologic stress increases plasma corticoster<strong>on</strong>e,<br />

mast cells degranulati<strong>on</strong>, reduces col<strong>on</strong>ic mucin secreti<strong>on</strong> and enhances intestinal mucosal permeability,<br />

facilitating bacterial adherence & uptake, followed by the sensitizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> T cells and initiati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

inflammatory/immune reacti<strong>on</strong>. Here we review our recent findings <strong>on</strong> the new molecular mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

experimental IBD induced by chemicals in rats & IL-10-KO mice. Namely, we recently found a<br />

c<strong>on</strong>comitantly increased expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the angiogenic (VEGF/VPF) and anti-angiogenic (angiostatin,<br />

endostatin) factors in the col<strong>on</strong>ic mucosa be<str<strong>on</strong>g>for</str<strong>on</strong>g>e the morphologic appearance <str<strong>on</strong>g>of</str<strong>on</strong>g> chemically-induced col<strong>on</strong>ic<br />

lesi<strong>on</strong>s in rats. Furthermore, the elevated levels <str<strong>on</strong>g>of</str<strong>on</strong>g> endostatin correlated with the severity <str<strong>on</strong>g>of</str<strong>on</strong>g> chemicallyinduced<br />

ulcerative colitis (UC) and disease progressi<strong>on</strong> in IL-10-KO mice. This probably results in<br />

insufficient angiogenesis & poor healing (by interfering with the proliferative acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> VEGF), while letting<br />

the VPF acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> VEGF/VPF be manifested in the early stages <str<strong>on</strong>g>of</str<strong>on</strong>g> IBD. This new mechanism is probably<br />

supported by another recent finding dem<strong>on</strong>strating a beneficial effect <str<strong>on</strong>g>of</str<strong>on</strong>g> VEGF neutralizati<strong>on</strong> <strong>on</strong> the severity<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> chemically-induced UC in rats that was associated with attenuati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> VEGF, PDGF & bFGF expressi<strong>on</strong>.<br />

Moreover, we found rapidly increased phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Erk1/2 and expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Egr-1, its DNA binding<br />

and interacti<strong>on</strong> with other transcripti<strong>on</strong> factors (AP-2, PPAR, NF-kβ) in col<strong>on</strong> during chemically-induced<br />

UC. VEGF neutralizati<strong>on</strong> significantly inhibited expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> pErk1/2 and Egr-1. C<strong>on</strong>clusi<strong>on</strong>s: 1) The<br />

unexpected parallel increased expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> both VEGF/VPF & angiostatin/endostatin in UC may explain<br />

the poor healing & sustained inflammati<strong>on</strong>. 2) Transcripti<strong>on</strong> factor Egr-1 may play a pivotal role in the<br />

VEGF acti<strong>on</strong>s in the pathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> experimental UC. 3) Stress may play a role in the exacerbati<strong>on</strong> but<br />

apparently not in the initiati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> IBD.<br />

374


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

6I_01_P<br />

(poster secti<strong>on</strong> B2, poster board #291, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STRESS-RELATED GASTRIC ULCERATION: THE GASTROPROTECTIVE ROLE OF<br />

GLUCOCORTICOID HORMONES<br />

Ludmila Filaretova 1 , Tatiana Bagaeva 1 , Koji Takeuchi 2 , Gabor Makara 3<br />

1Pavlov Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Physiology, nab. Makarova 6, St.Petersburg, Russia<br />

2Kyoto Pharmaceutical University, Japan; 3 Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Experimental Medicine, Budapest, Hungary<br />

e-mail: filaretova@pavlov.infran.ru<br />

Various stressors stimulate the producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> glucocorticoids, and severe stressors may also induce gastric<br />

erosi<strong>on</strong>, “stress ulcer”. From the very outset, researchers have focused <strong>on</strong> the point <str<strong>on</strong>g>of</str<strong>on</strong>g> view that stressgenerated<br />

glucocorticoids are causally related with gastric ulcerogenesis. In our studies we have focused <strong>on</strong><br />

the idea that glucocorticoids released during acute stress have an adaptive effect <strong>on</strong> the stomach and,<br />

there<str<strong>on</strong>g>for</str<strong>on</strong>g>e, are gastroprotective rather than ulcerogenic. To test this idea, we examined the effect <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

glucocorticoid deficiency (created by different approaches and followed by corticoster<strong>on</strong>e replacement) or the<br />

glucocorticoid receptor antag<strong>on</strong>ist RU-38486 <strong>on</strong> stress-induced gastric erosi<strong>on</strong> in rats. The data obtained<br />

show that the reducti<strong>on</strong> in the stress-induced corticoster<strong>on</strong>e release, or its acti<strong>on</strong>s, aggravates stress-caused<br />

gastric erosi<strong>on</strong>. It is suggested that an acute increase in corticoster<strong>on</strong>e during stress protects the stomach<br />

against injury caused by this stress. Extending this idea, we also hypothesized that glucocorticoids may play a<br />

role in the gastroprotective effect <str<strong>on</strong>g>of</str<strong>on</strong>g> prec<strong>on</strong>diti<strong>on</strong>ing stress. It is known that mild, prec<strong>on</strong>diti<strong>on</strong>ing, stress may<br />

attenuate stress-induced gastric ulcerati<strong>on</strong> and this effect is mediated by prostaglandins. We c<strong>on</strong>firmed that<br />

mild, prec<strong>on</strong>diti<strong>on</strong>ing, stress decrease the gastric ulcerati<strong>on</strong> caused by severe stress and dem<strong>on</strong>strated <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

first time that this effect is also provided by glucocorticoids released in resp<strong>on</strong>se to prec<strong>on</strong>diti<strong>on</strong>ing stress.<br />

The results support gastroprotective role glucocorticoids released during stress.<br />

375


23-26 August 2007,<br />

Budapest, Hungary<br />

6J. STRESS, HSPS AND INFLAMMATION<br />

6J_01_P<br />

(poster secti<strong>on</strong> B1, poster board #208, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECTS OF ANTI-INFLAMMATORY DRUG SUPPLEMENTATION ON HSP70<br />

EXPRESSION AND RELEASE DURING ANAEROBIC EXERCISE<br />

F. Le<strong>on</strong>i, T. Couls<strong>on</strong>, G. J<strong>on</strong>es, H. E. Ireland, C. Hunter-Lavin, J. H. H. Williams<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Chester, UK<br />

e-mail: f.le<strong>on</strong>i@chester.ac.uk<br />

Hsp70 expressi<strong>on</strong> and release increases in resp<strong>on</strong>se to exercise. To determine whether the inflammatory<br />

resp<strong>on</strong>se is involved in the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70, we analysed the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> Aspirin and Ibupr<str<strong>on</strong>g>of</str<strong>on</strong>g>en<br />

supplementati<strong>on</strong> and the release <str<strong>on</strong>g>of</str<strong>on</strong>g> pro and anti-inflammatory cytokines <strong>on</strong> Hsp70 after an intensive<br />

anaerobic exercise (30 sec<strong>on</strong>d maximal intensity Wingate test). A group <str<strong>on</strong>g>of</str<strong>on</strong>g> trained males (N=12) were<br />

administered Ibupr<str<strong>on</strong>g>of</str<strong>on</strong>g>en (Ibu, 1200mg) or Aspirin (Asp, 2700mg) and compared to a c<strong>on</strong>trol group (C<strong>on</strong>).<br />

Blood samples were taken be<str<strong>on</strong>g>for</str<strong>on</strong>g>e the exercise, 10 minutes after, and 1 hour after the sec<strong>on</strong>d blood taking.<br />

Hsp70 mRNA, intracellular protein expressi<strong>on</strong>, release from leucocytes and target cytokine release (IL-6,<br />

TNFα, IL-10) were measured. Flow Cytometry data showed a decrease <strong>on</strong> general Hsp70+ lymphocyte<br />

populati<strong>on</strong> after the exercise in C<strong>on</strong> and Asp but an increase in Ibu group; in particular the B cell expressing<br />

Hsp70 shows a decrease in Asp but an increase after 1h after in Ibu. Hsp70+ T helper lymphocytes showed<br />

no change in C<strong>on</strong>, a decrease in Asp and an increase in Ibu 1h after the exercise. Hsp70+ NK populati<strong>on</strong><br />

decreased in Asp which was not evident in C<strong>on</strong> and increased in Ibu group 1h after the exercise. Hsp70+<br />

m<strong>on</strong>ocytes decreased straight after the exercise, then returned to normal levels in C<strong>on</strong> and Asp. An increase<br />

in Hsp70+ m<strong>on</strong>ocytes was shown after the exercise in Ibu group. No change in Hsp70+ neutrophils was<br />

observed. We also showed a slight increase in Hsp70 gene expressi<strong>on</strong> straight after the exercise in C<strong>on</strong> but<br />

not in Asp or Ibu.<br />

376


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

6K. MATERNAL, FETAL AND NEONATAL STRESS<br />

6K_01_P<br />

(poster secti<strong>on</strong> B1, poster board #209, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

C-FOS EXPRESSION IN MEDIODORSAL THALAMIC AREA DURING THE<br />

NEONATAL PERIOD: INFLUENCE OF THE MATERNAL DEPRIVATION AND<br />

INJECTION-STRESS<br />

Katsumasa Muneoka 1 , Tetsuo Ogawa 1 , Makiko Kuwagata 1, 2 , Seiji Shioda 1<br />

1 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Anatomy I, Showa University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Tokyo, Japan<br />

2 Lab.<str<strong>on</strong>g>of</str<strong>on</strong>g> Pathology, Toxicology divisi<strong>on</strong>, Hatano <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institure, FDSC, Kanagawa, Japan<br />

e-mail: kmuneoka@med.showa-u.ac.jp<br />

Mediodorsal thalamic area (MDT) is a brain area innervating the prefr<strong>on</strong>tal cortex (PFC) and implicated as a<br />

site <str<strong>on</strong>g>of</str<strong>on</strong>g> neuropathology in schizophrenia. We investigated the functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> MDT in ne<strong>on</strong>atal rats as well as<br />

influence <str<strong>on</strong>g>of</str<strong>on</strong>g> stressful c<strong>on</strong>diti<strong>on</strong>s via examining c-Fos expressi<strong>on</strong> at postnatal day (PND) 11. We tested three<br />

experimental groups that are the home cage- (H), maternal deprivati<strong>on</strong>- (M) and injecti<strong>on</strong>-group (I). Rats in<br />

H-group were perfused transcranially immediately (within 5 mins) after picking up from their home cages.<br />

Rats in M-group were deprived from their mother <str<strong>on</strong>g>for</str<strong>on</strong>g> 60 mins in a novel cage and thereafter perfused. Rats in<br />

I-group were perfused 60 mins after subcutaneous injecti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> 100µl saline. Removed brains were cut with<br />

vibratome and c-Fos- immunoreactivity (ir)-positive cells were visualized with immunohistochemical<br />

reacti<strong>on</strong>s. Images were digitally taken and the numbers <str<strong>on</strong>g>of</str<strong>on</strong>g> c-Fos-ir-positive cells in the MDT were counted.<br />

Although c-Fos-ir-positive cells were detected in all three groups, the total number was significantly increased<br />

in I-group compared with H or M groups. These results suggest that the MDT-PFC neur<strong>on</strong>al system during<br />

the ne<strong>on</strong>atal period is active even under undisturbed envir<strong>on</strong>ment, and that injecti<strong>on</strong> stress but not maternal<br />

deprivati<strong>on</strong> stimulate the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the MDT. The present findings support the involvement <str<strong>on</strong>g>of</str<strong>on</strong>g> stress during<br />

the development in the pathophysiology <str<strong>on</strong>g>of</str<strong>on</strong>g> schizophrenia.<br />

6K_02_P<br />

(poster secti<strong>on</strong> B1, poster board #210, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PRENATALLY INCREASED MATERNAL CORTISOL: EFFECTS ON HPA AXIS,<br />

BRAIN NEUROTRANSMITTER SYSTEMS, IMMUNE RESPONSES AND<br />

BEHAVIOUR IN THE OFFSPRING OF PIGS<br />

W. Otten, E. Kanitz, M. Tuchscherer, B. Puppe<br />

<str<strong>on</strong>g>Research</str<strong>on</strong>g> Unit Behavioural Physiology, <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> the Biology <str<strong>on</strong>g>of</str<strong>on</strong>g> Farm Animals<br />

18196- Dummerstorf, Germany<br />

e-mail: otten@fbn-dummerstorf.de<br />

Envir<strong>on</strong>mental factors acting prenatally <strong>on</strong> the developing foetus are important determinants <str<strong>on</strong>g>for</str<strong>on</strong>g> disorders<br />

later in life. Maternal glucocorticoids are c<strong>on</strong>sidered as major candidates <str<strong>on</strong>g>for</str<strong>on</strong>g> the mediati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> stressful<br />

prenatal events to the <str<strong>on</strong>g>of</str<strong>on</strong>g>fspring. We investigated the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> increased maternal cortisol levels in pregnant<br />

sows <strong>on</strong> central and peripheral alterati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> the HPA axis, brain neurotransmitter pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iles, immune<br />

resp<strong>on</strong>ses and open-field behaviour <str<strong>on</strong>g>of</str<strong>on</strong>g> their piglets. Increased endogenous cortisol release was induced in<br />

pregnant sows by repeated intramuscular administrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> ACTH every sec<strong>on</strong>d day during late gestati<strong>on</strong>,<br />

whereas c<strong>on</strong>trol sows received injecti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> saline. The ACTH treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> sows significantly increased the<br />

377


23-26 August 2007,<br />

Budapest, Hungary<br />

birth weight <str<strong>on</strong>g>of</str<strong>on</strong>g> piglets without affecting gestati<strong>on</strong> length, number <str<strong>on</strong>g>of</str<strong>on</strong>g> total born piglets or frequency <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

stillborn piglets. After birth, piglets from ACTH-treated sows showed a suppressed cell-mediated immunity.<br />

Decreased corticosteroid-binding globulin levels indicate a higher amount <str<strong>on</strong>g>of</str<strong>on</strong>g> biologically available free<br />

cortisol and elevated noradrenaline levels dem<strong>on</strong>strate an enhanced activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the SAM system in the<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g>fspring from ACTH sows. Hypothalamic HPA axis feedback seems to be attenuated after birth in ACTH<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g>fspring as shown by a slightly reduced glucocorticoid receptor binding. Together with a significantly<br />

decreased serot<strong>on</strong>ergic activity in the locus coeruleus regi<strong>on</strong> these alterati<strong>on</strong>s may account <str<strong>on</strong>g>for</str<strong>on</strong>g> the increased<br />

resp<strong>on</strong>ses <str<strong>on</strong>g>of</str<strong>on</strong>g> the HPA and SAM system after an acute restraint stress and the increased emoti<strong>on</strong>al reactivity<br />

during an open-field test.<br />

6K_03_P<br />

(poster secti<strong>on</strong> B1, poster board #211, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE EFFECTS OF MATERNAL SEPARATION ON NEUROGENESIS IN THE RAT<br />

ROSTRAL MIGRATORY STREAM<br />

1 E. Račeková, 1 J. Orendáčová, 2 Ľ. Raček,<br />

1Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Neurobiology, CE, SAS; 2 Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Animal Physiology, SAS, Košice, Slovak Republic<br />

e-mail: racekova@saske.sk<br />

The olfactory bulb is the first relay <strong>on</strong> the olfactory sensory pathway and the target <str<strong>on</strong>g>for</str<strong>on</strong>g> neural progenitors<br />

generated in the subventricular z<strong>on</strong>e (SVZ) and which migrate al<strong>on</strong>g a well-demarcated pathway - the rostral<br />

migratory stream (RMS). Although neurogenesis in the SVZ/RMS occurs c<strong>on</strong>tinuously throughout<br />

adulthood, the rate <str<strong>on</strong>g>of</str<strong>on</strong>g> proliferati<strong>on</strong> and the fate <str<strong>on</strong>g>of</str<strong>on</strong>g> new cells may be affected by exogenous factors. Maternal<br />

separati<strong>on</strong> (MS) is a well-described model <str<strong>on</strong>g>of</str<strong>on</strong>g> envir<strong>on</strong>mental influences <strong>on</strong> development and subsequent<br />

nervous system functi<strong>on</strong>. In our study MS has been used as a model <str<strong>on</strong>g>of</str<strong>on</strong>g> early olfactory deprivati<strong>on</strong> to<br />

investigate its effect <strong>on</strong> neurogenesis in the rat RMS. Pups from postnatal day 1 (P1) to P21 were separated<br />

from the dam <str<strong>on</strong>g>for</str<strong>on</strong>g> 3 hrs daily and compared to pups that remained with the dam. At the end <str<strong>on</strong>g>of</str<strong>on</strong>g> MS, resp. <strong>on</strong>e<br />

week or three m<strong>on</strong>ths later the rats were injected with BrdU to determine the number <str<strong>on</strong>g>of</str<strong>on</strong>g> dividing cells in the<br />

RMS. Dying cells were visualized by Fluoro Jade-B histochemistry. The RMS <strong>on</strong> hematoxylin stained secti<strong>on</strong>s<br />

after all survival times was composed <str<strong>on</strong>g>of</str<strong>on</strong>g> tightly packed cells as seen in c<strong>on</strong>trol animals. However, cells<br />

proliferati<strong>on</strong> was specifically affected immediately after the finishing <str<strong>on</strong>g>of</str<strong>on</strong>g> MS resp. after short-term survival. At<br />

P21 there was almost complete depleti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> dividing cells in the RMS vertical arm. With reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> dividing<br />

cells, the number <str<strong>on</strong>g>of</str<strong>on</strong>g> dying cells noticeably increased in this part <str<strong>on</strong>g>of</str<strong>on</strong>g> the RMS. Three m<strong>on</strong>ths after MS, BrdUpositive<br />

cells displayed the similar pattern <str<strong>on</strong>g>of</str<strong>on</strong>g> density as seen in c<strong>on</strong>trol animals. These data indicate that<br />

adverse experience in life may induce acute site-specific changes in the RMS neurogenesis.<br />

378


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

6K_04_P<br />

(poster secti<strong>on</strong> B1, poster board #212, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

A FLATTENED DIURNAL CORTISOL PROFILE AND DEPRESSION IN<br />

ADOLESCENCE ARE RELATED TO ANTENATAL MATERNAL ANXIETY<br />

Bea R. H. Van den Bergh 1 , Ben Van Calster 2 , Tim Smits 3 , Lieven Lagae 4 , Sabine Van Huffel 2<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychology, K.U.Leuven, Leuven and Flemish Community Brussels, Belgium<br />

e-mail: bea.vandenbergh@psy.kuleuven.be<br />

In a prospective-l<strong>on</strong>gitudinal study maternal anxiety was measured at 12-22, 23-32 and 32-40 weeks <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

pregnancy (wp) with the State Trait Anxiety Inventory. 68 firstborns <str<strong>on</strong>g>of</str<strong>on</strong>g> 86 mothers, recruited at the<br />

Obstetrical and Gynecological Board c<strong>on</strong>sultati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> a University Hospital were followed up to age 15.<br />

HPAA functi<strong>on</strong>ing was measured through establishing a saliva day-time cortisol pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ile (shortened versi<strong>on</strong>);<br />

during a week-end day 3 saliva samples were provided, namely immediately after awakening, at no<strong>on</strong> and in<br />

the evening. Severity <str<strong>on</strong>g>of</str<strong>on</strong>g> depressive symptoms was measured with the Children’s Depressi<strong>on</strong> Inventory (CDI).<br />

We tested the following two hypotheses: (a) that maternal anxiety during pregnancy predicts altered HPAA<br />

functi<strong>on</strong> (e.g., a flattened cortisol pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ile) in adolescent <str<strong>on</strong>g>of</str<strong>on</strong>g>fspring; (b) that altered HPAA functi<strong>on</strong> mediates<br />

the associati<strong>on</strong> between prenatal maternal anxiety and depressi<strong>on</strong> in these adolescents. Repeated<br />

measurements regressi<strong>on</strong> analysis and ordinary least-squares regressi<strong>on</strong> analyses indicated that maternal<br />

anxiety at 12-22 wp was in female and male <str<strong>on</strong>g>of</str<strong>on</strong>g>fspring (all having reached Tanner puberty phase IV),<br />

associated with flattening <str<strong>on</strong>g>of</str<strong>on</strong>g> the diurnal salivary cortisol curve (P =.0463). Moreover in female adolescents<br />

this flatter cortisol curve was associated with depressi<strong>on</strong> (P = .0077). Effects remained when c<strong>on</strong>trolling <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

covariates such as postnatal maternal anxiety, smoking during pregnancy, birth weight and obstetrical<br />

optimality. Our results indicate that maternal anxiety during pregnancy enhances neurobiological vulnerability<br />

to depressi<strong>on</strong> - possibly by altering (or programming) foetal physiology - and dem<strong>on</strong>strate the mediating role<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> HPAA-dysregulati<strong>on</strong> in linking an adverse foetal envir<strong>on</strong>ment to depressi<strong>on</strong>. These results may lead to a<br />

re-orientati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the target <str<strong>on</strong>g>of</str<strong>on</strong>g> primary preventi<strong>on</strong> and treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> depressi<strong>on</strong>.<br />

6K_05_P<br />

(poster secti<strong>on</strong> B1, poster board #213, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

MATERNAL DEPRIVATION AND STRESS IN ADULT RATS: IMPORTANCE OF<br />

GENDER AND ESTROUS CYCLE STATE<br />

M. Crumeyrolle-Arias, V. Mourl<strong>on</strong>, B. Giros, V. Daugé<br />

INSERM U513, Créteil, France<br />

e-mail: crumeyrolle@creteil.inserm.fr<br />

Early adverse experiences can trigger l<strong>on</strong>g-lasting molecular and cellular modificati<strong>on</strong>s in brain and facilitate<br />

the appearance <str<strong>on</strong>g>of</str<strong>on</strong>g> pathological behaviors. This study analyzed the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> maternal deprivati<strong>on</strong> <strong>on</strong> the<br />

reactivity <str<strong>on</strong>g>of</str<strong>on</strong>g> L<strong>on</strong>g-Evans rats to stress according to gender and state <str<strong>on</strong>g>of</str<strong>on</strong>g> estrous cycle. Rat pups were separated<br />

from their mother and isolated <str<strong>on</strong>g>for</str<strong>on</strong>g> 3h/day during 14 days. At adulthood, different stressful situati<strong>on</strong>s were<br />

applied: change <str<strong>on</strong>g>of</str<strong>on</strong>g> animal housing, food deprivati<strong>on</strong> (18h), <str<strong>on</strong>g>for</str<strong>on</strong>g>ced swimming stress (5 min) or restraint stress<br />

(20 min). Serum corticoster<strong>on</strong>e level was quantified and the dexamethas<strong>on</strong>e sensitivity after stress tested.<br />

N<strong>on</strong>-deprived (ND) females presented a higher proporti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 5-day than 4-day cycle lengths. This difference<br />

was reduced <str<strong>on</strong>g>for</str<strong>on</strong>g> deprived (D) females. After the change <str<strong>on</strong>g>of</str<strong>on</strong>g> animal housing, 4- and 5-day cycles proporti<strong>on</strong><br />

was the same in both group and irregular cycles increased also similarly. Maternal deprivati<strong>on</strong> had no effect<br />

379


23-26 August 2007,<br />

Budapest, Hungary<br />

<strong>on</strong> corticoster<strong>on</strong>e level in both genders after the <str<strong>on</strong>g>for</str<strong>on</strong>g>ced swimming stress. D females in diestrus had higher<br />

level <str<strong>on</strong>g>of</str<strong>on</strong>g> corticoster<strong>on</strong>e than males and D proestrus females. Both ND and D proestrus females showed a<br />

higher resp<strong>on</strong>se to food deprivati<strong>on</strong> than males. Food deprived D females in diestrus presented a loss <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

sensitivity <str<strong>on</strong>g>of</str<strong>on</strong>g> the HPA axis to the negative feedback <str<strong>on</strong>g>of</str<strong>on</strong>g> dexamethas<strong>on</strong>e. These results indicate that maternal<br />

deprivati<strong>on</strong> had l<strong>on</strong>g-lasting effects <strong>on</strong> the estrous cycle and the reactivity to stress in female rats, depending<br />

<strong>on</strong> the stress applied and the state <str<strong>on</strong>g>of</str<strong>on</strong>g> the estrous cycle. Maternal deprivati<strong>on</strong> in female rats c<strong>on</strong>stitutes a new<br />

model to study the neurobiological basis <str<strong>on</strong>g>of</str<strong>on</strong>g> the vulnerability to psychiatric disorders, especially those in which<br />

a vulnerability <str<strong>on</strong>g>of</str<strong>on</strong>g> women has been shown such as depressi<strong>on</strong>.<br />

6K_06_P<br />

(poster secti<strong>on</strong> B1, poster board #214, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

NITRATED PLASMA ALBUMIN: A POTENTIAL MARKER OF NITRATIVE STRESS<br />

LINKED TO ASPHYXIA AND HYPOGLYCEMIA IN NEWBORNS<br />

J.- L. Wayenberg 1 , C. Caved<strong>on</strong> 1 , V. Ransy 1 , D. Vermeylen, E. Damis, Serge P. Bottari 2<br />

1 Depts. <str<strong>on</strong>g>of</str<strong>on</strong>g> Paediatrics and Ne<strong>on</strong>atology, ULB and CHIREC, Brussels, Belgium<br />

2 CHU & INSERM U884, Grenoble, France<br />

Several stresses induce an increased generati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> peroxynitrite which nitrates specific proteins <strong>on</strong> tyrosines<br />

am<strong>on</strong>g which albumin. A recent study showed a str<strong>on</strong>g increase <str<strong>on</strong>g>of</str<strong>on</strong>g> nitrotyrosine in the brains <str<strong>on</strong>g>of</str<strong>on</strong>g> ne<strong>on</strong>ates<br />

deceased from perinatal asphyxia, indicating the occurrence <str<strong>on</strong>g>of</str<strong>on</strong>g> an important nitrative stress. Similarly,<br />

ne<strong>on</strong>atal hypoglycemia has been shown to induce nitrotyrosine generati<strong>on</strong> in the brain <str<strong>on</strong>g>of</str<strong>on</strong>g> animal models. To<br />

investigate nitrative stress in clinical practice, we developed a novel ELISA allowing the quantitative<br />

determinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> nitrated albumin (Alb-NO 2 ) in plasma as a marker <str<strong>on</strong>g>of</str<strong>on</strong>g> peroxynitrite generati<strong>on</strong>.<br />

In an <strong>on</strong>going study <strong>on</strong> nitrative stress in human newborns (n = 194), we investigated the plasma Alb-NO 2<br />

c<strong>on</strong>centrati<strong>on</strong>s in ne<strong>on</strong>ates who suffered perpartal asphyxia or ne<strong>on</strong>atal hypoglycemia.<br />

We found a significant increase (p < 0.05) in Alb-NO 2 c<strong>on</strong>centrati<strong>on</strong> at D1 in children who developed<br />

moderate or severe ne<strong>on</strong>atal post-asphyxial encephalopathy (NE). Newborns who had a normal evoluti<strong>on</strong> or<br />

who developed mild NE had Alb-NO 2 levels similar to those <str<strong>on</strong>g>of</str<strong>on</strong>g> matched c<strong>on</strong>trols. Alb-NO 2 levels were<br />

normalized at day 4.<br />

There also was a significant (p = 0.05) inverse correlati<strong>on</strong> between glycemia and Alb-NO 2 levels in preterm<br />

ne<strong>on</strong>ates (n = 51). After exclusi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> c<strong>on</strong>current pathologies or c<strong>on</strong>founding factors significance increased to<br />

p < 0.005. This correlati<strong>on</strong> was found both at H1 and D1, indicating that nitrative stress is an early but<br />

sustained resp<strong>on</strong>se to hypoglycemia. Levels were normalized at D4.<br />

Our data indicate the occurrence <str<strong>on</strong>g>of</str<strong>on</strong>g> nitrative stress in newborns exposed to perpartal asphyxia and preterm<br />

ne<strong>on</strong>ates suffering from moderate to severe hypoglycemia in agreement with the increased tyrosine nitrati<strong>on</strong><br />

observed in brains in post-mortem studies. Our findings indicate that plasma Alb-NO 2 c<strong>on</strong>centrati<strong>on</strong> is a<br />

reliable marker <str<strong>on</strong>g>of</str<strong>on</strong>g> nitrative stress in clinical practice and might there<str<strong>on</strong>g>for</str<strong>on</strong>g>e have a prognostic value <str<strong>on</strong>g>for</str<strong>on</strong>g> the l<strong>on</strong>g<br />

term outcome <str<strong>on</strong>g>of</str<strong>on</strong>g> asphyxiated and preterm hypoglycaemic newborns.<br />

380


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

6L. MOLECULAR MECHANISMS AND CONSEQUENCES OF SUBSTANCE ABUSE:<br />

SMOKING, ALCOHOLISM, DRUGS<br />

6L_01_P<br />

(poster secti<strong>on</strong> B1, poster board #215, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CHRONIC ETHANOL INTAKE IMPAIRS BRAIN RENIN-ANGIOTENSIN SYSTEM<br />

FUNCTION. ROLE OF CALCIUM<br />

J. M. Martínez-Martos, M. D. Mayas, M. P. Cobo*, P. Cortes-Denia, S. De la Chica-Rodriguez,<br />

M. J. Ramírez-Exposito<br />

Experimental and Clinical Physiopathology <str<strong>on</strong>g>Research</str<strong>on</strong>g> Group CVI296, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Health Sciences,<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Jaén, Spain, *Servicio de Neurología, Complejo Hospitalario de Jaén, Spain<br />

Aspartyl aminopeptidase (AspAP) and glutamyl aminopeptidase (GluAP), named together as aminopeptidase<br />

A activity (APA) exert angiotensinase activity due to their relati<strong>on</strong> to the metabolism <str<strong>on</strong>g>of</str<strong>on</strong>g> angiotensins in the<br />

regi<strong>on</strong>al brain renin-angiotensin-system (RAS). APA c<strong>on</strong>verts angiotensin II (AngII) to angiotensin III<br />

(AngIII). AngIII possesses most <str<strong>on</strong>g>of</str<strong>on</strong>g> the properties <str<strong>on</strong>g>of</str<strong>on</strong>g> AngII and shares the same receptors. Both angiotensins<br />

participate not <strong>on</strong>ly in the central regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> blood pressure, but also participate as neurotransmitters/<br />

neuromodulators. Ethanol (EtOH) is a drug <str<strong>on</strong>g>of</str<strong>on</strong>g> abuse that produces hypertensi<strong>on</strong>, and the brain is specially<br />

sensitive to hypertensi<strong>on</strong>, which causes a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> vascular and cellular changes. Previous in vitro studies<br />

dem<strong>on</strong>strated that EtOH inhibits APA activity in basal c<strong>on</strong>diti<strong>on</strong>s (independently <str<strong>on</strong>g>of</str<strong>on</strong>g> Ca2+) and also under<br />

depolarisati<strong>on</strong> (dependently <str<strong>on</strong>g>of</str<strong>on</strong>g> Ca2+). In this work, we analyse the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic EtOH intake <strong>on</strong> APA<br />

activity under basal/resting and K+-stimulated c<strong>on</strong>diti<strong>on</strong>s in mouse fr<strong>on</strong>tal cortex synaptosomes in a Ca2+c<strong>on</strong>taining<br />

or Ca2+-free artificial cerebrospinal fluid (aCSF). The results show that EtOH does not modify<br />

APA activity under resting c<strong>on</strong>diti<strong>on</strong>s in presence <str<strong>on</strong>g>of</str<strong>on</strong>g> Ca2+, but is modified in absence <str<strong>on</strong>g>of</str<strong>on</strong>g> Ca2+. However,<br />

EtOH inhibits the K+-stimulated increase in APA activity in presence <str<strong>on</strong>g>of</str<strong>on</strong>g> Ca2+. In absence <str<strong>on</strong>g>of</str<strong>on</strong>g> Ca2+, this<br />

effect is not observed. Due to brain RAS is related with the pathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> hypertensi<strong>on</strong> at different levels,<br />

the inhibitory effects <str<strong>on</strong>g>of</str<strong>on</strong>g> EtOH <strong>on</strong> angiotensinase activity may explain the changes observed in blood pressure<br />

regulati<strong>on</strong>, local blood flow and/or fluid and electrolyte homeostasis, related to alcohol abuse.<br />

6L_02_P<br />

(poster secti<strong>on</strong> B1, poster board #216, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CHRONIC ETHANOL INTAKE AND OXIDATIVE STRESS IN MOUSE BRAIN: A<br />

CALCIUM-DEPENDENT MECHANISM<br />

M. J. Ramírez-Expósito, M. D. Mayas, M.P. Cobo *, S. De la Chica-Rodríguez, P. Cortes-Denia,<br />

J. M. Martínez-Martos<br />

Experimental and Clinical Physiopathology <str<strong>on</strong>g>Research</str<strong>on</strong>g> Group CVI296, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Health Sciences,<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Jaén, Spain. *Servicio de Neurología, Complejo Hospitalario de Jaén, Spain<br />

Ethanol (EtOH) is the most widely and frequently used psychoactive drug in modern society. Chr<strong>on</strong>ic<br />

alcoholism is associated with numerous degenerative and inflammatory disorders <str<strong>on</strong>g>of</str<strong>on</strong>g> the central nervous<br />

system. The purpose <str<strong>on</strong>g>of</str<strong>on</strong>g> the present work was to study the influence <str<strong>on</strong>g>of</str<strong>on</strong>g> EtOH in mouse fr<strong>on</strong>tal cortex<br />

synaptosomes after chr<strong>on</strong>ic EtOH intake (15% in drinking water during 30 days), under resting and K + -<br />

stimulated c<strong>on</strong>diti<strong>on</strong>s. The neurotoxic effects were analysed by free radicals generati<strong>on</strong> (luminol- or lucigenin-<br />

381


23-26 August 2007,<br />

Budapest, Hungary<br />

enhanced chemiluminiscence), lipid peroxidati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the membrane lipids (thiobarbituric acid reactive<br />

substances -TBARS-) and the oxidati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> synaptosomal proteins (carb<strong>on</strong>yl groups c<strong>on</strong>tent). In additi<strong>on</strong>, the<br />

bioenergetic behaviour <str<strong>on</strong>g>of</str<strong>on</strong>g> synaptosomes was analysed under the different experimental protocols. Oxidative<br />

stress parameters were modified in a different degree after EtOH intake. Thus, under resting c<strong>on</strong>diti<strong>on</strong>s,<br />

chr<strong>on</strong>ic EtOH intake did not modify lucigenin-enhanced chemiluminescence, but increased significantly<br />

(P


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

6L_04_P<br />

(poster secti<strong>on</strong> B1, poster board #218, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

IMPACT OF A CHRONIC ETHANOL TREATMENT IN PRENATALLY STRESSED<br />

MALE RATS ON THE ETHANOL APPETENCE AND ON LEVELS OF DELTAFOSB<br />

IN THE NUCLEUS ACCUMBENS<br />

Vincent Van Waes 1 , Mihaela Enache 1 , Stéfania Maccari 1,2 , Muriel Darnaudéry 1<br />

1 Unité de Neurosciences & Physiologie Adaptatives EA 4052, Equipe Stress Périnatal , Université de Lille 1,<br />

Villeneuve d'Ascq, France<br />

2 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Experimental Medicine & Pathology, Università di Roma “La Sapienza”, Roma, Italy<br />

In rats, prenatal stress affects the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the hypothalamic-pituitary-adrenocortical axis and increases<br />

vulnerability to several drugs <str<strong>on</strong>g>of</str<strong>on</strong>g> abuse (amphetamine, 3,4-methylenedioxymethamphetamine and nicotine).<br />

Repeated exposure to drugs <str<strong>on</strong>g>of</str<strong>on</strong>g> abuse persistently induces the transcripti<strong>on</strong> factor deltaFosB in the nucleus<br />

accumbens, effect hypothesized to c<strong>on</strong>tribute to neuroadaptati<strong>on</strong>s in dopamine-regulated signalling. The aim<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the present work was 1) to determine whether prenatal stress have an impact <strong>on</strong> ethanol appetence in rats;<br />

2) to assess deltaFosB levels in the nucleus accumbens after a chr<strong>on</strong>ic ethanol treatment in prenatally stressed<br />

and c<strong>on</strong>trol rats.<br />

Preference <str<strong>on</strong>g>for</str<strong>on</strong>g> ethanol was measured in a two-bottle choice paradigm (water versus ethanol 2.5%, 5% or<br />

10%) in adolescent (postnatal days 28-38) naive rats and in adult rats after a chr<strong>on</strong>ic oral treatment (ethanol<br />

2.5%, 5% or 10%). The level <str<strong>on</strong>g>of</str<strong>on</strong>g> motivati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> ethanol was determined in adult animals under challenge<br />

situati<strong>on</strong>s (reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ethanol c<strong>on</strong>centrati<strong>on</strong>, ethanol withdrawal, c<strong>on</strong>current choice with sucrose). After 9<br />

m<strong>on</strong>ths <str<strong>on</strong>g>of</str<strong>on</strong>g> ethanol oral treatment, animal were killed, nuclei accumbens dissected and deltaFosB levels<br />

assessed by western blot.<br />

Our results indicated that prenatal stress did not modify ethanol preference, neither during adolescence, nor<br />

after the <str<strong>on</strong>g>for</str<strong>on</strong>g>ced ethanol drinking period in adulthood. Moreover, animals’ motivati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> ethanol was weak,<br />

in spite <str<strong>on</strong>g>of</str<strong>on</strong>g> the l<strong>on</strong>g-lasting ethanol exposure. In c<strong>on</strong>trast, the chr<strong>on</strong>ic oral treatment with ethanol 10%<br />

significantly increased deltaFosB levels in the nucleus accumbens. This increase was more important in<br />

prenatally stressed than in c<strong>on</strong>trol animals. The dissociati<strong>on</strong> between the impact <str<strong>on</strong>g>of</str<strong>on</strong>g> prenatal stress <strong>on</strong> brain<br />

deltaFosB and <strong>on</strong> ethanol c<strong>on</strong>sumpti<strong>on</strong> suggests that it would be important to evaluate more finely the<br />

motivati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> ethanol in our populati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> animals, in particular by modulating the palatability <str<strong>on</strong>g>of</str<strong>on</strong>g> ethanol<br />

and/or by c<strong>on</strong>sidering interindividual differences.<br />

383


23-26 August 2007,<br />

Budapest, Hungary<br />

384<br />

6M. OTHER STRESS TOPICS IN MEDICINE (GENDER DIFFERENCES IN<br />

PATHOPHYSIOLOGICAL STRESS, HSPS AND WOUND HEALING, STRESS AND<br />

MYOCARDIAL PROTECTION, PRE- AND POSTOPERATIVE STRESS)<br />

6M_01_P<br />

(poster secti<strong>on</strong> B1, poster board #219, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

GENDER-RELATED DIFFERENCES IN RESPONSE OF RAT BRAIN<br />

CORTICOSTEROID RECEPTORS AND HEAT SHOCK PROTEINS TO<br />

ANTIDEPRESSANT IMIPRAMINE<br />

Gordana Matić, Ivana Elaković, Jelena Brkljačić<br />

Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Biological <str<strong>on</strong>g>Research</str<strong>on</strong>g>, Despot Stefan Blvd. 142, 11000 Belgrade, Serbia<br />

e-mail: gormatic@ibiss.bg.ac.yu<br />

Sexual dimorphism in the prevalence <str<strong>on</strong>g>of</str<strong>on</strong>g> major depressi<strong>on</strong> 1 has been c<strong>on</strong>sidered a c<strong>on</strong>sequence <str<strong>on</strong>g>of</str<strong>on</strong>g> sex<br />

differences in the HPA axis functi<strong>on</strong>ing 2,3 . Antidepressant drugs enhance glucocorticoid signaling and<br />

normalize HPA axis reactivity 4 , representing useful tools <str<strong>on</strong>g>for</str<strong>on</strong>g> animal studies <strong>on</strong> the pathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> this<br />

disorder. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> the present study was to examine gender-related differences in the resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> pituitary<br />

and brain corticosteroid receptors (CRs) to a typical tricyclic antidepressant drug, imipramine. To that end,<br />

the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> a l<strong>on</strong>g-term imipramine treatment <strong>on</strong> the protein levels <str<strong>on</strong>g>of</str<strong>on</strong>g> glucocorticoid receptor (GR), and<br />

heat shock proteins Hsp90 and Hsp70, as well as <strong>on</strong> dexamethas<strong>on</strong>e binding to both GR and<br />

mineralocorticoid receptor (MR) in the pituitary, hypothalamus, hippocampus and brain cortex <str<strong>on</strong>g>of</str<strong>on</strong>g> n<strong>on</strong>depressed<br />

rats were studied. Differences in the GR protein level in the tissues <str<strong>on</strong>g>of</str<strong>on</strong>g> untreated female vs. male<br />

animals were not noticed. However, imipramine led to opposite changes in the cellular level <str<strong>on</strong>g>of</str<strong>on</strong>g> GR protein in<br />

the brain <str<strong>on</strong>g>of</str<strong>on</strong>g> female and male rats, as well as to gender- and tissue-specific changes in in vitro dexamethas<strong>on</strong>e<br />

binding to GR and MR in the hippocampus and brain cortex. Gender-related differences in the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Hsp90 and Hsp70 were noticed mainly in the hippocampus, <strong>on</strong>ly after imipramine treatment. The results<br />

suggest that this antidepressant may affect both the GR level and the mechanisms regulating its binding<br />

ability in a gender-related manner.<br />

6M_02_P<br />

(poster secti<strong>on</strong> B1, poster board #220, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HSP27 IS INVOLVED IN ACTIN BASED CELL MOVEMENT DURING<br />

WOUND HEALING<br />

Bindi Doshi, Lawrence Hightower, Juliet Lee<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> C<strong>on</strong>necticut, 91 N. Eagleville Rd, U3125, Storrs, CT 06269, USA<br />

e-mail: Bindi.Doshi@uc<strong>on</strong>n.edu<br />

Heat inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP27 in vertebrate cells causes micr<str<strong>on</strong>g>of</str<strong>on</strong>g>ilament stabilizati<strong>on</strong>, increased pinocytosis and cell<br />

migrati<strong>on</strong>. HSP27 plays an important role in cytoprotecti<strong>on</strong>, has actin capping activity and inhibits actin<br />

polymerizati<strong>on</strong> in vitro. The amount <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP27 expressed is regulated by gene activati<strong>on</strong> and post<br />

transcripti<strong>on</strong>ally whereas activity is regulated by phosphorylati<strong>on</strong> in resp<strong>on</strong>se to stress. Neither<br />

phosphorylated nor unphosphorylated multimers alter actin dynamics whereas unphosphorylated m<strong>on</strong>omers<br />

inhibit actin polymerizati<strong>on</strong> in vitro. C<strong>on</strong>sequently, HSP27 has been proposed as a regulator <str<strong>on</strong>g>of</str<strong>on</strong>g> cell invasi<strong>on</strong>;<br />

however, the direct role <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP27 in cell motility is unclear. In the present study, we c<strong>on</strong>structed a


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

eukaryotic expressi<strong>on</strong> vector using fish DNA to investigate the role <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP27 in cell motility. Heat shock<br />

treatment caused relocati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP27GFP from diffuse cytosolic distributi<strong>on</strong>s to c<strong>on</strong>centrati<strong>on</strong> in<br />

perinuclear regi<strong>on</strong>s, suggesting a cytoprotective role involving protein aggregati<strong>on</strong> in resp<strong>on</strong>se to heat.<br />

CRL1807, a human col<strong>on</strong> epithelial cell line, was transfected with HSP27GFP driven by a CMV c<strong>on</strong>stitutive<br />

promoter to over-express HSP27. In vitro wound assays showed that cells over-expressing HSP27 had a<br />

faster wound closure rate compared to c<strong>on</strong>trol cells. HSP27 levels were knocked down in CRL1807 cells<br />

using siRNA. Cells with decreased HSP27 levels showed slower wound closure rates compared to c<strong>on</strong>trol<br />

cells implying a role in regulating motility. Rapidly moving fish epithelial keratocytes were also used as a<br />

model cell motility system. Keratocytes over-expressing HSP27GFP had greater cell speed than c<strong>on</strong>trol cells.<br />

Western blotting, immunostaining and co-localizati<strong>on</strong> with the actin cytoskelet<strong>on</strong> will be presented.<br />

6M_03_P<br />

(poster secti<strong>on</strong> B1, poster board #221, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PROHIBITIN IS REQUIRED FOR SURVIVAL OF CARDIOMYOCYTES AGAINST<br />

STRESS INDUCED DAMAGE<br />

Lingjia Qian*, Xiaohua Liu, Zhen Ren, Zhihua Yang<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Health and Envir<strong>on</strong>mental Medicine, China<br />

e-mail: newjia@vip.sina.com<br />

Stress is c<strong>on</strong>sidered as <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> main causes inducing myocardial injury. Mitoch<strong>on</strong>dria play a key role in<br />

triggering the necrosis and apoptosis pathway <str<strong>on</strong>g>of</str<strong>on</strong>g> cardiomyocytes under stress. Prohibitin has been shown to<br />

stabilize and regulate the respiratory chain enzymes in mitoch<strong>on</strong>dria. The present research was designed to<br />

investigate the role <str<strong>on</strong>g>of</str<strong>on</strong>g> prohibitin in mitoch<strong>on</strong>dria <str<strong>on</strong>g>for</str<strong>on</strong>g> cardioprotecti<strong>on</strong>. Stress had no influence <strong>on</strong> prohibitin<br />

gene expressi<strong>on</strong> in cardiomyocytes, but time-and dose-dependently increased the prohibitin c<strong>on</strong>tent in<br />

mitoch<strong>on</strong>dria. The overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> prohibitin in cardiomyocytes by transfecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> prohibitin gene resulted<br />

in an increase <str<strong>on</strong>g>of</str<strong>on</strong>g> prohibitin in mitoch<strong>on</strong>dria. Compared with the n<strong>on</strong>-transfected cardiomyocytes, prohibitin<br />

overexpressi<strong>on</strong> could protect the mitoch<strong>on</strong>dria from stress-induced injury. C<strong>on</strong>sistently, the mitoch<strong>on</strong>driamediated<br />

apoptosis pathway was suppressed in prohibitin overexpressed cardiomyocytes after stress<br />

treatment, including the reduced change <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>drial membrane permeability transiti<strong>on</strong> and inhibited<br />

release <str<strong>on</strong>g>of</str<strong>on</strong>g> CytC from mitoch<strong>on</strong>dria to cytoplasma. As a result, stress induced cardiomyocyte apoptosis was<br />

inhinbited. Moreover, in stressed cardiomyocytes the interacti<strong>on</strong> between prohibitin and HSP70 was found,<br />

and the increase <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 expressi<strong>on</strong> promoted the translocati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> prohibitin to mitoch<strong>on</strong>dria and<br />

improved the survival <str<strong>on</strong>g>of</str<strong>on</strong>g> cardiomyocytes against stress injury. These indicated that prohibitin protected the<br />

cardiomyocytes from stress induced damage, increasing prohibitin c<strong>on</strong>tent in mitoch<strong>on</strong>dria via HSP70 may<br />

c<strong>on</strong>stitute a new therapeutic target <str<strong>on</strong>g>for</str<strong>on</strong>g> myocardium injury.<br />

385


23-26 August 2007,<br />

Budapest, Hungary<br />

6M_04_P<br />

(poster secti<strong>on</strong> B1, poster board #222, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ELEVATED HEAT SHOCK PROTEINS 70 AND 27 ARE CORRELATED WITH CNS<br />

INJURY IN SURGICAL PATIENTS UNDERGOING DEEP HYPOTHERMIA<br />

CIRCULATORY ARREST AND DESCENDING THORACIC AORTIC ANEURYSM<br />

RESECTION<br />

James Hecker<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Pennsylvania, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Anesthesia, 305 Morgan, 3620 Hamilt<strong>on</strong> Walk, 19104-6112,<br />

Philadelphia, USA<br />

e-mail: heckerj@uphs.upenn.edu<br />

Introducti<strong>on</strong>. The Heat Shock Proteins (HSPs) are members <str<strong>on</strong>g>of</str<strong>on</strong>g> a highly c<strong>on</strong>served family <str<strong>on</strong>g>of</str<strong>on</strong>g> molecular<br />

chaper<strong>on</strong>es, some <str<strong>on</strong>g>of</str<strong>on</strong>g> which are rapidly induced by severe stress. The inducible <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 is normally<br />

near the lower limit <str<strong>on</strong>g>of</str<strong>on</strong>g> detecti<strong>on</strong> by ELISA in the cerebral spinal fluid (CSF) <str<strong>on</strong>g>of</str<strong>on</strong>g> humans, and remains so<br />

despite exercise to exhausti<strong>on</strong> Dalsgaard et al (1). The inducible members <str<strong>on</strong>g>of</str<strong>on</strong>g> the Hsp70 and Hsp27 families<br />

are associated with cellular protecti<strong>on</strong> and recovery after a near lethal stress. Whole animals, isolated organs<br />

and cells subjected to heat shock are protected against a subsequent near lethal ischemic or hypoxic event.<br />

Inducible HSPs, in particular Hsp70 and 27, have also been used as markers <str<strong>on</strong>g>for</str<strong>on</strong>g> cells and tissues that have<br />

been exposed to a near-lethal stress. Although the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> protective intracellular resp<strong>on</strong>ses by heat<br />

shock is not clinically useful, an understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> the time course and correlati<strong>on</strong> with injury <str<strong>on</strong>g>of</str<strong>on</strong>g> HSPs<br />

released during brain and/or spinal cord cellular stress (ischemia) is critical in understanding the role <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

HSPs in cellular survival, and is potentially very useful as a biomarker <str<strong>on</strong>g>of</str<strong>on</strong>g> severe cellular stress.<br />

Methods. As a part <str<strong>on</strong>g>of</str<strong>on</strong>g> the c<strong>on</strong>tinuing ef<str<strong>on</strong>g>for</str<strong>on</strong>g>ts to improve peri-operative management by the Hospital <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Pennsylvania Aortic Surgery <str<strong>on</strong>g>Research</str<strong>on</strong>g> Group, serial human CSF measurements <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 and<br />

27 during Deep Hypothermic Circulatory Arrest (DHCA) and Descending Thoracic Aorta repair procedures<br />

were collected. These patients are at high risk <str<strong>on</strong>g>for</str<strong>on</strong>g> spinal cord ischemia with surgical resecti<strong>on</strong> due to the<br />

locati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the procedure, and routine invasive m<strong>on</strong>itors there<str<strong>on</strong>g>for</str<strong>on</strong>g>e includes a lumbar intrathecal (‘spinal’)<br />

drain. In this IRB Approved protocol, CSF samples were collected at the following time points: lumbar drain<br />

placement immediately following inducti<strong>on</strong>; at time <str<strong>on</strong>g>of</str<strong>on</strong>g> aortic cross clamp; <strong>on</strong>e, two, and twelve hours after<br />

crossclamp; and at regular intervals following, or if signs/symptoms <str<strong>on</strong>g>of</str<strong>on</strong>g> paraparesis developed postoperatively.<br />

Samples were immediately spun and frozen <str<strong>on</strong>g>for</str<strong>on</strong>g> simultaneous analysis by ELISA. Preliminary<br />

results show marked increases in these Heat Shock Proteins.<br />

Results. Of the first 23 DHCA and thoracoabdominal aneurysm patients (not c<strong>on</strong>secutive), nine have large<br />

increases in Hsp70 and 27 by 2 hours after crossclamp, nine more have large increases by 48 hours after<br />

crossclamp, and <str<strong>on</strong>g>of</str<strong>on</strong>g> these 18, eight have post-operative paraplegia. All patients with transient or permanent<br />

paraparesis had large increases in Hsp70 and Hsp27 at multiple time points in the peri-operative period. One<br />

patient had high levels <str<strong>on</strong>g>of</str<strong>on</strong>g> these HSPs at the initial time point just after inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> anesthesia, and he had<br />

had a carotid endarterectomy procedure the preceding day. Reported another way, <str<strong>on</strong>g>of</str<strong>on</strong>g> the total <str<strong>on</strong>g>of</str<strong>on</strong>g> 23 patients<br />

examined, all 8 patients with post-operative paraplegia had multiple elevated HSPs over the first 48 hours,<br />

four patients had multiple elevated HSPs without paraplegia, and seven patients had <strong>on</strong>ly a few mildly<br />

elevated time points with no post-op paraplegia.<br />

C<strong>on</strong>clusi<strong>on</strong>s. These preliminary results are supportive <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 and Hsp27 as biomarkers <str<strong>on</strong>g>for</str<strong>on</strong>g> significant<br />

spinal cord and perhaps brain ischemia.<br />

386


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

6M_05_P<br />

(poster secti<strong>on</strong> B1, poster board #223, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

N-TERMINAL MUTATIONS IN CRYAB ARE ASSOCIATED WITH HYPERTROPHIC<br />

CARDIOMYOPATHY<br />

B. Stoevring 1,2,* , O. Vang 1 , W. J. Mckenna 3 , M. Christiansen 2<br />

1Roskilde University, Roskilde Denmark, 2 Statens Serum Institut, Copenhagen Denmark<br />

3University College <str<strong>on</strong>g>of</str<strong>on</strong>g> L<strong>on</strong>d<strong>on</strong>, L<strong>on</strong>d<strong>on</strong>, UK<br />

e-mail: biv@ssi.dk<br />

Introducti<strong>on</strong>: Hypertrophic cardiomyopathy (HCM) is a comm<strong>on</strong> genetic disease characterised by the<br />

presence <str<strong>on</strong>g>of</str<strong>on</strong>g> unexplained cardiac left ventricular hypertrophy. αB-crystallin is a small heat shock protein<br />

expressed at a high level in the human heart and skeletal muscles. During cardiac ischemia αB-crystallin<br />

translocates from the cytosol to the my<str<strong>on</strong>g>of</str<strong>on</strong>g>ibrils where it protects my<str<strong>on</strong>g>of</str<strong>on</strong>g>ibrillar proteins from unfolding or<br />

aggregati<strong>on</strong>. Mutati<strong>on</strong>s in the C-terminus <str<strong>on</strong>g>of</str<strong>on</strong>g> the αB-crystallin gene (CRYAB) have previously been associated<br />

with My<str<strong>on</strong>g>of</str<strong>on</strong>g>ibrillar Myopathy, cataract and recently with Dilated Cardiomyopathy (DCM). Methods: We<br />

analyzed 339 patients clinically diagnosed with HCM and 190 patients with DCM <str<strong>on</strong>g>for</str<strong>on</strong>g> mutati<strong>on</strong>s in CRYAB<br />

by high-resoluti<strong>on</strong> melting curve analysis and subsequent DNA sequencing. Circular dichroism (CD)<br />

spectroscopy was used to assess changes in sec<strong>on</strong>dary structure in synthetic peptides corresp<strong>on</strong>ding to the<br />

αB-crystallin-N-terminal (AA 1-60) with and without mutati<strong>on</strong>s. Results: Four HCM probands were<br />

heterozygous carriers <str<strong>on</strong>g>of</str<strong>on</strong>g> the N-terminal mutati<strong>on</strong>s Q26X, P39L, M1I and R50Q, respectively. The P39L<br />

segregated with family disease and CD analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> WT and P39L peptides showed a 50 % increase in helix<br />

c<strong>on</strong>tent as a c<strong>on</strong>sequence <str<strong>on</strong>g>of</str<strong>on</strong>g> the P39L mutati<strong>on</strong>. In the DCM group, <strong>on</strong>e proband was heterozygous <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

previously described DCM associated C-terminal mutati<strong>on</strong> G154S changing a fully c<strong>on</strong>served glycine into a<br />

serine and most likely associated with decreased chaper<strong>on</strong>e activity. Discussi<strong>on</strong>: This is the first study to<br />

identify N-terminal mutati<strong>on</strong>s in αB-crystallin and an associati<strong>on</strong> with HCM, indicati<strong>on</strong> genotype phenotype<br />

differences depending <strong>on</strong> the mutated domain <str<strong>on</strong>g>of</str<strong>on</strong>g> αB-crystallin and thus different pathogenic mechanisms.<br />

We show that the P39L mutati<strong>on</strong> cause severe sec<strong>on</strong>dary structure changes and suggest that this may result in<br />

changed intermolecular interacti<strong>on</strong>s compromising the functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> αB-crystallin.<br />

6M_06_P<br />

(poster secti<strong>on</strong> B1, poster board #224, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PECULIARITIES OF HSP60 AND HSP70 EXPRESSION IN CARDIOVASCULAR<br />

DISEASES CAUSED BY CHRONIC STRESS<br />

L. Kapustian, V. Bobyk, O. Rozhko, I. Kroupskaya, D. Ryabenko*, V. Gurtovyy**,<br />

Yu. Khozhaenko**, V. Usenko**, L. Sidorik<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular Biology and Genetics NAS <str<strong>on</strong>g>of</str<strong>on</strong>g> Ukraine, Kyiv, Ukraine<br />

*Strazhesko Cardiology Institute MAS <str<strong>on</strong>g>of</str<strong>on</strong>g> Ukraine<br />

**Morphological Laboratory BIONTEK, Dnipropetrovsk, Ukraine<br />

Investigati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the role <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock proteins in pathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> cardiovascular diseases is <str<strong>on</strong>g>of</str<strong>on</strong>g> great scientific<br />

and practical importance. Overexpressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these proteins has significant cytoprotective effect in<br />

cardiovascular system. This phenomen<strong>on</strong> was dem<strong>on</strong>strated in culture <str<strong>on</strong>g>of</str<strong>on</strong>g> cardiomyocytes, transgenic animals<br />

and intact hearts. However, the majority <str<strong>on</strong>g>of</str<strong>on</strong>g> these investigati<strong>on</strong>s were carried out in the acute stress<br />

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Budapest, Hungary<br />

c<strong>on</strong>diti<strong>on</strong>s, and our knowledge about peculiarities <str<strong>on</strong>g>of</str<strong>on</strong>g> such proteins under chr<strong>on</strong>ic heart stress, including<br />

cardiomyopathies, is restricted. Dilated cardiomyopathy (DCM) is a severe autoimmune cardiospecific disease<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> unknown origin characterized by the dilatati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> left and/or right ventricular, mitoch<strong>on</strong>drial dysfuncti<strong>on</strong><br />

and massive apoptotic cardiomyocytes death with collagenosis/fibrosis <str<strong>on</strong>g>of</str<strong>on</strong>g> heart muscle. We studied the<br />

expressi<strong>on</strong> and localizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> abundant cytoplasmic (Hsp70) and mitoch<strong>on</strong>drial (Hsp60) chaper<strong>on</strong>s at DCM<br />

progressi<strong>on</strong> at the end stage <str<strong>on</strong>g>of</str<strong>on</strong>g> heart failure (secti<strong>on</strong>al pathomorphologic samples <str<strong>on</strong>g>of</str<strong>on</strong>g> the human myocardium)<br />

and in dynamics (using the mouse model <str<strong>on</strong>g>of</str<strong>on</strong>g> myosin-induced autoimmune myocardial damage like human<br />

DCM). The decreased level <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp70 and increased level <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp60 have been observed in DCM affected<br />

hearts by Western-blot analysis. These changes were accompanied by significant increase <str<strong>on</strong>g>of</str<strong>on</strong>g> anti-myosin<br />

autoantibodies level in DCM patients sera and the accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>drial proteins precursors in<br />

DCM cardiomyocytes. Moreover, we revealed the decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp60 in the cytoplasmic fracti<strong>on</strong> and<br />

significant increase in mitoch<strong>on</strong>drial fracti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> DCM-affected hearts by Western blotting. The data <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

immunohystochemical analysis have shown changes <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp60 and Hsp70 expressi<strong>on</strong> in myocardium <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

model animals at different stages <str<strong>on</strong>g>of</str<strong>on</strong>g> the model disease progressi<strong>on</strong>. Our results taken together with recently<br />

published data c<strong>on</strong>cerning antiapoptotic role <str<strong>on</strong>g>of</str<strong>on</strong>g> cytosolic Hsp60 in cultured embry<strong>on</strong>ic cardiomyocytes allow<br />

to propose that the changes in the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp60 and Hsp70 cytosolic fracti<strong>on</strong>s may play an important role<br />

in the apoptotic events in cardiomyocytes at DCM.<br />

388


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

MODULE 7<br />

PSYCHOSOCIAL STRESS<br />

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23-26 August 2007,<br />

Budapest, Hungary<br />

SYMPOSIA<br />

• 7A. Stress, cognitive functi<strong>on</strong> and behavior (August 24th Friday afterno<strong>on</strong>)<br />

(Organizers and chairs: József Haller and Jeansok Kim)<br />

Poster: August 24 th Friday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

• 7B. Psychosocial stress and disease (August 24th Friday morning)<br />

(Organizer and chair: Holger Ursin)<br />

Poster: August 24 th Friday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

• 7C. Psycho-neuro-immunology (August 24th Friday morning)<br />

(Organizer and chair: Judit Szelényi)<br />

Poster: August 24 th Friday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

• 7D. Cultural differences, cultural changes and stress (August 24th Friday afterno<strong>on</strong>)<br />

(Organizer and chair: Mária Kopp)<br />

Poster: August 24 th Friday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

• 7E. Dental stress (August 24th Friday afterno<strong>on</strong>)<br />

(Organizer and chair: Tibor Károly Fábián)<br />

Poster: August 24 th Friday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

• 7F. Post traumatic stress disorder (August 24th Friday afterno<strong>on</strong>)<br />

(Organizer and chair: Marcus Ising, and Christoph Czermak)<br />

Poster: August 24 th Friday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

• 7G. Stress and reproducti<strong>on</strong>: From sexual dysfuncti<strong>on</strong> to sp<strong>on</strong>taneous aborti<strong>on</strong> (August 24th<br />

Friday morning)<br />

(Organizer and chair: Pablo Nepomnaschy)<br />

Poster: August 24 th Friday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

POSTER SYMPOSIA<br />

• 7I. Work stress<br />

August 24 th Friday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

• 7J. Stress and collective behavior: Outbreaks <str<strong>on</strong>g>of</str<strong>on</strong>g> panic, war, market crash<br />

August 24 th Friday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

• 7K. Stress and gender<br />

August 24 th Friday, Poster secti<strong>on</strong> A2, Building A 1st floor<br />

390


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7A. STRESS, COGNITIVE FUNCTION AND BEHAVIOR<br />

(JÓZSEF HALLER, JEANSOK KIM)<br />

7A_01_S<br />

PREDATOR STRESS, MEMORY AND BRAIN PLASTICITY<br />

David M. Diam<strong>on</strong>d<br />

Departments <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychology, Molecular Pharmacology and Physiology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> South Florida, Tampa, Florida<br />

33620 USA<br />

e-mail: ddiam<strong>on</strong>d@mail.usf.edu<br />

It is well-known that the hippocampus is critically involved in the <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> new memories. However,<br />

extensive research has provided str<strong>on</strong>g support <str<strong>on</strong>g>for</str<strong>on</strong>g> the view that hippocampal functi<strong>on</strong>ing is impaired in<br />

times <str<strong>on</strong>g>of</str<strong>on</strong>g> str<strong>on</strong>g emoti<strong>on</strong>ality. For example, studies have shown that stress suppresses the inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> l<strong>on</strong>gterm<br />

potentiati<strong>on</strong> (LTP), a well-accepted physiological model <str<strong>on</strong>g>of</str<strong>on</strong>g> memory, in rats. Moreover, research <strong>on</strong><br />

people indicates that in times <str<strong>on</strong>g>of</str<strong>on</strong>g> str<strong>on</strong>g emoti<strong>on</strong>ality there is a suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> hippocampal functi<strong>on</strong>ing. Thus,<br />

despite the importance <str<strong>on</strong>g>of</str<strong>on</strong>g> the hippocampus in processes involved in memory <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>, in times <str<strong>on</strong>g>of</str<strong>on</strong>g> stress the<br />

hippocampus appears to be inhibited from participating in the <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> emoti<strong>on</strong>al memories, including<br />

flashbulb and traumatic memories. In this talk I will incorporate rodent and human studies to provide an<br />

alternative interpretati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the findings. I will suggest that c<strong>on</strong>diti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> str<strong>on</strong>g emoti<strong>on</strong>ality actually produce<br />

enhanced neuroplasticity in the hippocampus (and amygdala), which underlies the declarative comp<strong>on</strong>ent <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

emoti<strong>on</strong>al memories. The hyperactivati<strong>on</strong> phase <str<strong>on</strong>g>of</str<strong>on</strong>g> plasticity, which is then followed by an inhibitory phase <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

hippocampal plasticity, underlies well-described psychological phenomena in emoti<strong>on</strong>al memory processing,<br />

such as the great durability <str<strong>on</strong>g>of</str<strong>on</strong>g> flashbulb and traumatic memories, as well as stress-induced amnesia.<br />

7A_02_S<br />

HYPER- AND HYPO-AROUSAL IN THE CONTROL OF AGGRESSIVENESS: THE<br />

ROLE OF GLUCOCORTICOIDS<br />

J. Haller, J. Halasz, E. Mikics, M. Toth, B. Barsy<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Experimental Medicine, Budapest, Hungary<br />

We will show that - bey<strong>on</strong>d chr<strong>on</strong>ic HPA-axis hyperfuncti<strong>on</strong> -, decreased glucocorticoid producti<strong>on</strong> also leads<br />

to psychopathologies. This point will be exemplified by studies <strong>on</strong> aggressive behavior. Psychopathologyassociated<br />

human aggressi<strong>on</strong> types are induced by a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> c<strong>on</strong>diti<strong>on</strong>s, are behaviorally variable, and show<br />

a differential pharmacological resp<strong>on</strong>siveness. Thus, there are several types <str<strong>on</strong>g>of</str<strong>on</strong>g> abnormal human aggressi<strong>on</strong>.<br />

This diversity was not reflected by c<strong>on</strong>venti<strong>on</strong>al laboratory approaches that focused <strong>on</strong> the quantitative<br />

aspects <str<strong>on</strong>g>of</str<strong>on</strong>g> aggressive behavior. Recently, several laboratory models <str<strong>on</strong>g>of</str<strong>on</strong>g> abnormal aggressi<strong>on</strong> were proposed,<br />

which mainly model hyperarousal-driven aggressiveness (characteristic to intermittent explosive disorder,<br />

posttraumatic stress disorder, depressi<strong>on</strong>, chr<strong>on</strong>ic burnout, etc.) and hypoarousal-driven aggressiveness<br />

(characteristic mainly to antisocial pers<strong>on</strong>ality disorder and its childhood antecedent c<strong>on</strong>duct disorder).<br />

Findings obtained with these models suggest that hyperarousal-driven aggressiveness has at its roots an<br />

excessive acute glucocorticoid stress resp<strong>on</strong>se (and probably an exaggerated resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> other stress-related<br />

systems), whereas chr<strong>on</strong>ic hypoarousal-associated aggressiveness is due to glucocorticoid deficits that affect<br />

brain functi<strong>on</strong> <strong>on</strong> the l<strong>on</strong>g term. In hypoarousal-driven aggressiveness, serot<strong>on</strong>ergic neurotransmissi<strong>on</strong><br />

appears to lose its impact <strong>on</strong> aggressi<strong>on</strong> (which it has in normal aggressi<strong>on</strong>), certain prefr<strong>on</strong>tal neur<strong>on</strong>s are<br />

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23-26 August 2007,<br />

Budapest, Hungary<br />

weakly activated, whereas the central amygdala (no, or weakly involved in the c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> normal aggressi<strong>on</strong>)<br />

acquires important roles. We suggest that the specific study <str<strong>on</strong>g>of</str<strong>on</strong>g> abnormal aspects <str<strong>on</strong>g>of</str<strong>on</strong>g> aggressive behavior would<br />

lead to important developments in understanding the specific mechanisms underlying different <str<strong>on</strong>g>for</str<strong>on</strong>g>ms <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

aggressi<strong>on</strong>, and may ultimately lead to the development <str<strong>on</strong>g>of</str<strong>on</strong>g> better treatment approaches.<br />

7A_04_S<br />

A SYSTEMS-LEVEL MODEL OF STRESS EFFECTS ON COGNITION<br />

Jeansok J. Kim, Taejib Yo<strong>on</strong><br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychology and Program in Neurobiology & Behavior, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Washingt<strong>on</strong>, Seattle,<br />

Washingt<strong>on</strong>, USA<br />

Stress is a biologically significant social-envir<strong>on</strong>mental factor that plays a pervasive role in our lives, from<br />

impacting our daily behaviors to producing and exacerbating myriad physical and mental illness. An<br />

accumulating body <str<strong>on</strong>g>of</str<strong>on</strong>g> evidence from human and animal studies reveals that while the acute resp<strong>on</strong>se to stress<br />

(i.e., heightened cogniti<strong>on</strong>) is an adaptive mechanism, exposures to unc<strong>on</strong>trollable (unpredictable and<br />

inescapable) stress can subsequently produce detrimental neurocognitive effects, particularly in the<br />

hippocampus. Rodent studies further indicate that stress impairs l<strong>on</strong>g-term potentiati<strong>on</strong> (LTP), a leading<br />

candidate cellular mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> storage, in the hippocampus. We have recently discovered that<br />

amygdalar lesi<strong>on</strong>s/inactivati<strong>on</strong> and prefr<strong>on</strong>tal cortex lesi<strong>on</strong>s block and exacerbate, respectively, stress-induced<br />

impairments in hippocampal LTP and spatial memory. Moreover, single unit recording data indicate that<br />

stress alters the firing rate <str<strong>on</strong>g>of</str<strong>on</strong>g> place cells recorded from dorsal hippocampus, providing an empirical bridge<br />

between stress effects <strong>on</strong> synaptic plasticity and spatial memory. Based <strong>on</strong> these findings, we will present a<br />

c<strong>on</strong>ceptual model <str<strong>on</strong>g>of</str<strong>on</strong>g> the central stress mechanism (a neural-endocrine network comprising <str<strong>on</strong>g>of</str<strong>on</strong>g> amygdala,<br />

prefr<strong>on</strong>tal cortex and glucocorticoids) regulating hippocampal functi<strong>on</strong>ing.<br />

392<br />

7A_05_S<br />

(poster secti<strong>on</strong> A2, poster board #75, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STRESS COPING STRATEGIES IN A 3D ESCAPE MAZE REVEAL TWO DISTINCT<br />

TYPES OF SOCIAL FEAR LEARNING<br />

Cliff H. Summers, Russ E. Carpenter<br />

Biology, Neuroscience Group, University <str<strong>on</strong>g>of</str<strong>on</strong>g> South Dakota, Vermilli<strong>on</strong>, SD 57069 USA, e-mail: cliff@usd.edu<br />

Rainbow trout interact aggressively, and <str<strong>on</strong>g>for</str<strong>on</strong>g>m distinct social hierarchies. We hypothesized that social<br />

interacti<strong>on</strong> against a larger opp<strong>on</strong>ent provides the impetus and in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> necessary to stimulate goaloriented<br />

learning in subordinate individuals. A small (~100 g) juvenile rainbow trout in <strong>on</strong>e tank<br />

compartment was adjacent to a large fish (~300 g; US) in another. A small (5 cm) hole leading to an empty<br />

compartment, large enough <strong>on</strong>ly <str<strong>on</strong>g>for</str<strong>on</strong>g> the smaller fish to pass was <strong>on</strong>ly available when the larger fish was<br />

present. Once a day the water inflow was turned <str<strong>on</strong>g>of</str<strong>on</strong>g>f (CS); 15s later dividers were removed and fish interacted<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> 15 min. Larger fish were aggressive, and the learning curve <str<strong>on</strong>g>for</str<strong>on</strong>g> subordinate fish was dramatic (600%)<br />

over seven days. Escape time improved daily until fish escaped in approximately <strong>on</strong>e minute. Plasma was<br />

taken 3 days be<str<strong>on</strong>g>for</str<strong>on</strong>g>e and 1 day after social interacti<strong>on</strong> and learning trials. For samples taken after the trials,<br />

fish were presented with inflow water <str<strong>on</strong>g>of</str<strong>on</strong>g>f but no large fish challenge. Fish learning to escape showed no<br />

change in plasma cortisol. However, fish that did not learn to escape exhibited a four-fold increase in<br />

cortisol, although no large fish was presented as a social challenge. Turning <str<strong>on</strong>g>of</str<strong>on</strong>g>f inflow water acts as a


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

c<strong>on</strong>diti<strong>on</strong>ed stimulus to provoke increased cortisol, and elevated dopamine and serot<strong>on</strong>in in amygdala,<br />

hypothalamus and raphé. Increased anxiety induced by CRF stimulates increased attacks, but also increased<br />

retreat and escape behavior. Coping strategies appear to be influenced by CRF 1 receptor activity. Fear<br />

c<strong>on</strong>diti<strong>on</strong>ing in n<strong>on</strong>-escaping fish is manifest by increased neural and horm<strong>on</strong>al stress resp<strong>on</strong>siveness,<br />

whereas those that learn to escape utilize spatial learning to cope with social stress.<br />

7A_06_S<br />

(poster secti<strong>on</strong> A2, poster board #76, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ARE EXTREMES IN ANXIETY LINKED TO COGNITIVE ABILITIES<br />

1M. Bunck, 2 S. B. Sartori, 1 R. Zurmühlen, 1 C. T.Wotjak, 2 N. Singewald, 1 R. Landgraf<br />

1 Max Planck Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychiatry, Munich, Germany, 2 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmacology and Toxicology, University <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Innsbruck, Austria; e-mail: Bunck@mpipsykl.mpg.de<br />

Anxiety and learning/memory are hypothesized to be linked phenomena, as patients suffering from anxiety<br />

disorders and depressi<strong>on</strong> display changes in cognitive abilities. To mimic the clinical situati<strong>on</strong>, wild type CD1<br />

mice (NAB) were selected and inbred <str<strong>on</strong>g>for</str<strong>on</strong>g> extremes in anxiety-related behaviour in the elevated plus maze test<br />

to generate (HAB) versus low anxiety-related behaviour (LAB) mice. To study the interplay between<br />

emoti<strong>on</strong>ality and cogniti<strong>on</strong>, HAB, NAB and LAB mice were subjected to various social and n<strong>on</strong>-social<br />

learning tasks. The general ability to recognise a familiar stimulus animal was assessed by the social<br />

recogniti<strong>on</strong> test. As a more sophisticated measure <str<strong>on</strong>g>of</str<strong>on</strong>g> social memory, the social discriminati<strong>on</strong> task (SD) was<br />

per<str<strong>on</strong>g>for</str<strong>on</strong>g>med. Furthermore, we used a c<strong>on</strong>textual and cued fear-c<strong>on</strong>diti<strong>on</strong>ing (FC) paradigm to investigate<br />

extincti<strong>on</strong> abilities. The results clearly indicate that all three lines show social recogniti<strong>on</strong> abilities with a<br />

similar decline in olfactory investigati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the same stimulus. Remarkably, in the SD <strong>on</strong>ly HAB mice<br />

succeeded in identifying a familiar animal even after 120 min interexposure interval, indicating that the SD<br />

can be used to reveal differences in cognitive abilities between the mouse lines. Interestingly, in both FC<br />

paradigms HAB mice displayed delayed extincti<strong>on</strong> compared to both, NAB and LAB animals. The data<br />

suggest that hyper-anxiety typical <str<strong>on</strong>g>of</str<strong>on</strong>g> HABs is linked to enhanced social memory abilities and resistance to<br />

extincti<strong>on</strong>. Further analyses <str<strong>on</strong>g>of</str<strong>on</strong>g> the interacti<strong>on</strong> between emoti<strong>on</strong>ality and cogniti<strong>on</strong> will focus <strong>on</strong> anxiety- and<br />

cogniti<strong>on</strong>-related neuropeptide systems, the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> antidepressants and extincti<strong>on</strong>-facilitating drugs.<br />

393


23-26 August 2007,<br />

Budapest, Hungary<br />

394<br />

7A_02_P<br />

(poster secti<strong>on</strong> A2, poster board #77, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

COGNITIVE DEFICITS CAUSED BY CHRONIC STRESS CAN BE PREVENTED BY<br />

THE GINKGO BILOBA AND HYPERICUM PERFORATUM ADMINISTRATION<br />

Jan J. Braszko, Anna Walesiuk, Emil Tr<str<strong>on</strong>g>of</str<strong>on</strong>g>imiuk<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Clinical Pharmacology, Medical University <str<strong>on</strong>g>of</str<strong>on</strong>g> Bialystok, Waszyngt<strong>on</strong>a 15A, 15-274<br />

Białystok, Poland<br />

Exposure to chr<strong>on</strong>ic restraint stress in rats as well as the psychosocial stress in humans has been shown to<br />

alter cognitive functi<strong>on</strong>s such as learning and memory and has been linked to the pathophysiology <str<strong>on</strong>g>of</str<strong>on</strong>g> mood<br />

and anxiety disorders. Anxiolytic/sedative/antidepressant agents used in the management <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress-related<br />

disorders have several side effects and are not cost-effective.<br />

There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, in this study we investigated efficacy <str<strong>on</strong>g>of</str<strong>on</strong>g> the orally given two natural medicines: extract <str<strong>on</strong>g>of</str<strong>on</strong>g> Ginkgo<br />

biloba (EGB 761, 100 mg/kg) and dried, standardized c<strong>on</strong>fecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> crude Hypericum per<str<strong>on</strong>g>for</str<strong>on</strong>g>atum (350 mg/kg), in<br />

the preventi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the post-stress memory dysfuncti<strong>on</strong>s.<br />

Male Wistar rats (150-160 g) were stressed <str<strong>on</strong>g>for</str<strong>on</strong>g> 21 days by placing them <str<strong>on</strong>g>for</str<strong>on</strong>g> 2 h daily in a tight restraint tubes.<br />

There were separate sets <str<strong>on</strong>g>of</str<strong>on</strong>g> experiments testing effects <str<strong>on</strong>g>of</str<strong>on</strong>g> EGB 761 and H. per<str<strong>on</strong>g>for</str<strong>on</strong>g>atum. To define specific role<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the rat stress horm<strong>on</strong>e (cortisol in humans) we ran also groups injected daily with corticoster<strong>on</strong>e (5 mg/kg<br />

subcutaneously).<br />

Each experimental set c<strong>on</strong>sisted <str<strong>on</strong>g>of</str<strong>on</strong>g> 6 groups <str<strong>on</strong>g>of</str<strong>on</strong>g> animals: CONTROL, STRESS, corticoster<strong>on</strong>e (CORT),<br />

EXTRACT (EGB 761 or H. per<str<strong>on</strong>g>for</str<strong>on</strong>g>atum), EXTRACT + STRESS, EXTRACT + CORT.<br />

Both, STRESS and CORT groups displayed c<strong>on</strong>siderable and statistically significant deficits in memory:<br />

spatial [measured in the Morris Water Maze (MWM) (p


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

synaptotagmin, a Ca 2+ -dependent synaptic vesicle protein, brain-derived neurotrophic factor (BDNF) and its<br />

receptor tropomyosin-related kinase B (TrkB) play important roles in hippocampus-dependent learning and<br />

memory behavior. However, whether chr<strong>on</strong>ic exercise can improve learning and memory by upregulating<br />

these molecules remains unraveled. To answer this questi<strong>on</strong>, male BALB/c mice were used as the animal<br />

model. After 4 weeks <str<strong>on</strong>g>of</str<strong>on</strong>g> treadmill exercise training, the ability <str<strong>on</strong>g>of</str<strong>on</strong>g> learning and memory was evaluated by <strong>on</strong>etrial<br />

passive avoidance test (PA), an aversive learning task. Hippocampal synaptotagmin and TrkB protein<br />

expressi<strong>on</strong>s were determined by Western blotting, and BDNF was measured by ELISA. Our results showed<br />

that after chr<strong>on</strong>ic treadmill exercise, 1) the retenti<strong>on</strong> latency <str<strong>on</strong>g>of</str<strong>on</strong>g> PA was increased; 2) protein levels <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

hippocampal TrkB and synaptotagmin were elevated; 3) TrkB or synaptotagmin protein expressi<strong>on</strong> was<br />

positively correlated with PA per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance. These data suggest that the upregulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> synaptotagmin and<br />

TrkB may c<strong>on</strong>tribute to the chr<strong>on</strong>ic exercise-facilitated hippocampus-dependent cognitive functi<strong>on</strong>.<br />

7A_05_P<br />

(poster secti<strong>on</strong> A2, poster board #79, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

TOWARDS THE GENETIC DISSECTION OF ANXIETY<br />

L. Czibere, M. Bunck, E. Frank, M. S. Keßler, M. Kohli, T. Bettecken, R. Landgraf<br />

Max-Planck-Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychiatry, 80804 Kraepelinstr. 2-10, Munich, Germany,<br />

e-mail: czibere@mpipsykl.mpg.de<br />

To investigate the genetic background <str<strong>on</strong>g>of</str<strong>on</strong>g> anxiety-related and depressi<strong>on</strong>-like behaviour in mice, two lines,<br />

selectively inbred <str<strong>on</strong>g>for</str<strong>on</strong>g> high (“HAB”) and low anxiety-related behaviour (“LAB”), starting out from outbred<br />

CD1 mice. As mutati<strong>on</strong>s in the genome are assumed to make up <str<strong>on</strong>g>for</str<strong>on</strong>g> the majority <str<strong>on</strong>g>of</str<strong>on</strong>g> genetic differences, we<br />

plan to pinpoint them with a linkage analysis approach. For this, the ‘Mouse Medium Density Linkage Panel’<br />

by Illumina, was used to genotype 1449 single nucleotide polymorphisms (SNPs) in HAB and LAB mice, as<br />

well as in F1 progeny (HABxLAB).<br />

The animals tested were derived from generati<strong>on</strong> 20 or higher, providing a solid basis <str<strong>on</strong>g>for</str<strong>on</strong>g> genetically<br />

homogenous lines.<br />

In a first analysis, out <str<strong>on</strong>g>of</str<strong>on</strong>g> the 1449 loci tested, <str<strong>on</strong>g>for</str<strong>on</strong>g> 225 autosomal loci HAB and LAB mice both displayed<br />

homozygosity, but <str<strong>on</strong>g>for</str<strong>on</strong>g> different alleles. Cross-mated animals (F1, HABxLAB) were all heterozygous <str<strong>on</strong>g>for</str<strong>on</strong>g> these<br />

loci.<br />

Compared to an inter-strain analysis approach using two standard inbred mouse strains (e.g. BALB/c and<br />

C57BL6), where animals do not <strong>on</strong>ly differ in their anxiety-related behaviour, our intra-strain analysis<br />

c<strong>on</strong>siderably reduces the number <str<strong>on</strong>g>of</str<strong>on</strong>g> in<str<strong>on</strong>g>for</str<strong>on</strong>g>mative SNPs. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, in our mouse lines, the number <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

in<str<strong>on</strong>g>for</str<strong>on</strong>g>mative false positive SNPs, relevant to the behavioural phenotype, will be much lower.<br />

Interestingly, about 1/3 <str<strong>on</strong>g>of</str<strong>on</strong>g> the opposite homozygous SNPs are c<strong>on</strong>centrated <strong>on</strong> <strong>on</strong>ly 5 chromosomes,<br />

whereas two chromosomes c<strong>on</strong>tained just 3 and 6 <str<strong>on</strong>g>of</str<strong>on</strong>g> these SNPs.<br />

Altogether, the Illumina Mouse Medium Density Linkage Panel seems to provide a sufficient number <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

in<str<strong>on</strong>g>for</str<strong>on</strong>g>mative SNPs as to allow the identificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a linkage regi<strong>on</strong>, causative <str<strong>on</strong>g>for</str<strong>on</strong>g> the development <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

anxiety trait. We are planning to genotype a larger number <str<strong>on</strong>g>of</str<strong>on</strong>g> freely segregating F2 generati<strong>on</strong> animals<br />

(F1xF1). From this we expect being able to identify true candidate regi<strong>on</strong>s <str<strong>on</strong>g>for</str<strong>on</strong>g> anxiety-related behaviour.<br />

395


23-26 August 2007,<br />

Budapest, Hungary<br />

7A_06_P<br />

(poster secti<strong>on</strong> A2, poster board #80, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

A REWARD-SEEKING MODEL FOR DISTINGUISHING ADAPTATION AND STRESS<br />

Seamus Decker<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> AnthropologyUniversity <str<strong>on</strong>g>of</str<strong>on</strong>g> Massachusetts215 Machmer HallAmherst, MA 01003 USA<br />

e-mail: sdecker@anthro.umass.edu<br />

Lack <str<strong>on</strong>g>of</str<strong>on</strong>g> a unifying c<strong>on</strong>cept <str<strong>on</strong>g>for</str<strong>on</strong>g> distinguishing adaptati<strong>on</strong> from stress with research <strong>on</strong> psych<strong>on</strong>euroendocrine<br />

(PNE) processes such as hypothalamic-pituitary-adrenal axis (HPA) resp<strong>on</strong>se to psychosocial c<strong>on</strong>diti<strong>on</strong>s has<br />

led to at least four distinct types <str<strong>on</strong>g>of</str<strong>on</strong>g> errors: (i) failure to distinguish adaptive PNE activati<strong>on</strong>, i.e., “arousal,”<br />

from maladaptive PNE activati<strong>on</strong>, i.e., “stress;” (ii) failure to distinguish resp<strong>on</strong>se from c<strong>on</strong>diti<strong>on</strong>s that<br />

provoked the resp<strong>on</strong>se, i.e., either stressors or a vulnerability <str<strong>on</strong>g>of</str<strong>on</strong>g> some sort; (iii) assumpti<strong>on</strong> that subjective<br />

distress and psychobiological activati<strong>on</strong> are equivalent and interchangeable; and (iv) failure to distinguish<br />

differences in the causes and c<strong>on</strong>sequences <str<strong>on</strong>g>of</str<strong>on</strong>g> resp<strong>on</strong>se which is episodic, ephemeral and moderated<br />

compared to resp<strong>on</strong>se which is repetitious, prol<strong>on</strong>ged, or exaggerated. Distinguishing adaptive from<br />

maladaptive resp<strong>on</strong>se depends <strong>on</strong> a clear understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> the adaptive purposes <str<strong>on</strong>g>of</str<strong>on</strong>g> PNE processes such as<br />

HPA resp<strong>on</strong>se to psychosocial c<strong>on</strong>diti<strong>on</strong>s, and attenti<strong>on</strong> to the c<strong>on</strong>text and c<strong>on</strong>sequences <str<strong>on</strong>g>of</str<strong>on</strong>g> any specific<br />

pattern <str<strong>on</strong>g>of</str<strong>on</strong>g> resp<strong>on</strong>se. This paper presents a novel and unifying model in which episodic cortisol resp<strong>on</strong>se to<br />

psychosocial factors is c<strong>on</strong>ceived as an adaptati<strong>on</strong> to perceived loss <str<strong>on</strong>g>of</str<strong>on</strong>g> reward. Reward is c<strong>on</strong>ceived broadly<br />

as being the myriad <str<strong>on</strong>g>of</str<strong>on</strong>g> social or behavioral factors that may lead to specific psychobiological states <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

“reward” (e.g., increased activity in specific dopaminergic regi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> the pre-fr<strong>on</strong>tal cortex). By testing this<br />

c<strong>on</strong>ceptual model, researchers stand to make substantial c<strong>on</strong>tributi<strong>on</strong>s to understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> how the HPA<br />

works in everyday life, and effects <strong>on</strong> human health and well-being.<br />

7A_07_P<br />

(poster secti<strong>on</strong> A2, poster board #81, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

RESPONSIVENESS OF THE OK-LIST: A NEW RATING SCALE FOR CLINICAL<br />

DIAGNOSIS AND TREATMENT EVALUATION OF PSYCHOLOGICAL SYMPTOMS<br />

OF CHRONIC STRESS<br />

E. Dorant, A. J. M. Schmidt, G. M. van der Molen<br />

Maastricht University, Health Organizati<strong>on</strong>, Policy and <str<strong>on</strong>g>Ec<strong>on</strong>omic</str<strong>on</strong>g>s, PO Box 616, 6200 MD Maastricht,<br />

The Netherlands; e-mail: E.Dorant@beoz.unimaas.nl<br />

Objective. A new rating scale <str<strong>on</strong>g>for</str<strong>on</strong>g> clinical diagnosis and treatment evaluati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> complaints <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic stress<br />

was developed in The Netherlands. This OK-list c<strong>on</strong>sists <str<strong>on</strong>g>of</str<strong>on</strong>g> 22 universally applicable complaint-like items,<br />

scorable <strong>on</strong> a 5-point Likert scale. Psychometric evaluati<strong>on</strong> showed high validity, reliability and<br />

unidimensi<strong>on</strong>ality. In this study we evaluate the resp<strong>on</strong>siveness <str<strong>on</strong>g>of</str<strong>on</strong>g> the OK-list in patients suffering from<br />

chr<strong>on</strong>ic stress syndrome, equivalent to the clinical end state <str<strong>on</strong>g>of</str<strong>on</strong>g> severe burnout.<br />

Methods. A single group repeated measurements design was used. Intake and end-<str<strong>on</strong>g>of</str<strong>on</strong>g>-treatment in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong><br />

<strong>on</strong> level <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic stress complaints was collected with the OK-list. Resp<strong>on</strong>siveness was assessed using<br />

paired T-test, SRM and a graphical illustrati<strong>on</strong> presenting means <str<strong>on</strong>g>for</str<strong>on</strong>g> each chr<strong>on</strong>ic stress complaint. For<br />

parallel comparis<strong>on</strong> SCL-90R data were used. Results were calculated <str<strong>on</strong>g>for</str<strong>on</strong>g> two subgroups defined by<br />

c<strong>on</strong>current depressi<strong>on</strong> using data collected with the Beck Depressi<strong>on</strong> Inventory.<br />

396


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Results. Comparis<strong>on</strong> with SCL-90R revealed parallel lowering <str<strong>on</strong>g>of</str<strong>on</strong>g> OK-scores between intake and end-<str<strong>on</strong>g>of</str<strong>on</strong>g>treatment<br />

in both subgroups. All paired T-tests were statistically significant (p-values


23-26 August 2007,<br />

Budapest, Hungary<br />

398<br />

7A_09_P<br />

(poster secti<strong>on</strong> A2, poster board #83, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE EFFECTS OF SEX, AGE AND POPULATION ON BASELINE<br />

GLUCOCORTICOID LEVELS IN SPINY MICE (ACOMYS CAHIRINUS)<br />

M. Nováková 1 , H. Kutalová 1 , R. Palme 2 , D. Frynta 1 , L. Janský 3<br />

1Dept.<str<strong>on</strong>g>of</str<strong>on</strong>g> Zoology, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Science, Charles Univ., Prague, Czech Republic<br />

2Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Natural Sciences, Univ. <str<strong>on</strong>g>of</str<strong>on</strong>g> Veterinary Medicine, Vienna, Austria<br />

3Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Animal Physiology, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Biological Sciences, Univ. <str<strong>on</strong>g>of</str<strong>on</strong>g> South Bohemia, České Budějovice,<br />

Czech Republic, e-mail: frynta@centrum.cz<br />

Baseline glucocortiocoid (GC) levels may be related to the social status in small animal species. Here we<br />

examined GC levels in a moderately social rodent the Egyptian spiny mouse (Acomys cahirinus). Studied<br />

animals originated from 2 populati<strong>on</strong>s, differing c<strong>on</strong>siderably in their ecology: the populati<strong>on</strong> from Abu<br />

Simbel (S Egypt) is a desert-dweller, while that from Cairo is commensal. To assess baseline GC levels we<br />

collected faecal samples from 68 individuals bel<strong>on</strong>ging to 10 social groups. Each social group c<strong>on</strong>sisted <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

animals <str<strong>on</strong>g>of</str<strong>on</strong>g> both sexes and two age grades (directly corresp<strong>on</strong>ding to dominance ranks). Levels <str<strong>on</strong>g>of</str<strong>on</strong>g> GC<br />

metabolites were assessed by a 5α-pregnane-3β,11β,21-triol-20-<strong>on</strong>e enzyme immunoassays (EIA). The results<br />

showed no effect <str<strong>on</strong>g>of</str<strong>on</strong>g> rank and <strong>on</strong>ly a small effect <str<strong>on</strong>g>of</str<strong>on</strong>g> sex (higher levels in females) <strong>on</strong> GC metabolites levels.<br />

Thus, the hypothesis that dominant males are more stressed than other functi<strong>on</strong>al groups may be rejected.<br />

Surprisingly, there was a c<strong>on</strong>siderable difference am<strong>on</strong>g groups. This may be interpreted as a substantial<br />

effect <str<strong>on</strong>g>of</str<strong>on</strong>g> social relati<strong>on</strong>ships within each particular group. The commensal populati<strong>on</strong> exhibited much higher<br />

levels <str<strong>on</strong>g>of</str<strong>on</strong>g> GC metabolites than the desert <strong>on</strong>e. This is c<strong>on</strong>sistent with behavioural differences <str<strong>on</strong>g>of</str<strong>on</strong>g> studied<br />

populati<strong>on</strong>s - commensals are more active, but simultaneously also more anxious. Evoluti<strong>on</strong>ary adaptati<strong>on</strong> to<br />

living in buildings is suggested as the ultimate cause <str<strong>on</strong>g>of</str<strong>on</strong>g> these physiological and behavioural differences.<br />

7A_10_P<br />

(poster secti<strong>on</strong> A2, poster board #84, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECTS OF STRESS APPLIED AT THE LATE STAGE OF PREGNANCY ON THE<br />

FEEDING BEHAVIOR IN THE RAT<br />

Mari Kimoto 1* , Rie Amano 1 , Reiko Ishikawa 1 ,Yuki Ooe 1 , Mie Shimura 1 ,Tami Niinuma 1 ,<br />

Hiromi Hirakawa 1 , Yasuhiro Kumei 2 , Kazuo Toda 1,3<br />

Physiological Lab., Japan Women's University, Mejirodai, Bunkyo-ku, Tokyo 112-8681 1 , Biochemistry, Tokyo<br />

Medical and Dental University, Yushima, Bunkyo-ku, Tokyo113-8549, Japan 2 , Integrative Sensory Physiology,<br />

Nagasaki University, Nagasaki 852-8588, Japan 3<br />

e-mail: kimoto@fc.jwu.ac.jp<br />

The purpose <str<strong>on</strong>g>of</str<strong>on</strong>g> the present study is to examine how stress that is imposed at the fetal stage could influence<br />

the same Wistar rat at the adult stage. The pregnant rats were exposed to either gravitati<strong>on</strong>al stress (1.5 and<br />

2.0G) or wire-net restraint during the 15 th through 21 st day <str<strong>on</strong>g>of</str<strong>on</strong>g> pregnancy <str<strong>on</strong>g>for</str<strong>on</strong>g> 10 min daily. The male <str<strong>on</strong>g>of</str<strong>on</strong>g>fspring<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the wire-net restraint mother rat showed significant decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> slope in growth curve at the 15 th through<br />

66 th day. The 1.5G gravitati<strong>on</strong>al stress <strong>on</strong> mother rats did not influence the body weight <str<strong>on</strong>g>of</str<strong>on</strong>g> the male <str<strong>on</strong>g>of</str<strong>on</strong>g>fspring,<br />

while 2.0G gravitati<strong>on</strong>al stress <strong>on</strong> mother rats increased the slope <str<strong>on</strong>g>of</str<strong>on</strong>g> growth curve <str<strong>on</strong>g>of</str<strong>on</strong>g> 6 to 10 day-old male<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g>fspring rats. By c<strong>on</strong>trast, 1.5G gravitati<strong>on</strong>al stress and wire-net restraint did not influence the feeding


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

behavior in the adult stage <str<strong>on</strong>g>of</str<strong>on</strong>g> the female <str<strong>on</strong>g>of</str<strong>on</strong>g>fspring rats. However, 2.0G stress increased the slope <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

growth curve. The somatostatin level in the blood was decreased in gravitati<strong>on</strong>al stress group in the male, but<br />

not in the female <str<strong>on</strong>g>of</str<strong>on</strong>g>fspring rats. Data showed that the stress imposed at the fetal stage modified the growth<br />

pattern even at the adult stage and that the gender difference was observed in the anti-stress resp<strong>on</strong>se. In<br />

additi<strong>on</strong>, type <str<strong>on</strong>g>of</str<strong>on</strong>g> stress may be an important factor <str<strong>on</strong>g>for</str<strong>on</strong>g> producing emoti<strong>on</strong>al changes, such as, feeding<br />

behavior.<br />

7A_11_P<br />

(poster secti<strong>on</strong> A2, poster board #85, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE ROLE OF CORTICOSTERONE IN EXERCISE-INDUCED<br />

HIPPOCAMPAL NEUROGENESIS<br />

Yu-Min Kuo, Ya-Ting Chang, Yi-Chieh Chen, Chih-Wei Wu<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Cell Biology and Anatomy, Medical College, Nati<strong>on</strong>al Cheng Kung University, 1 Ta-Hsueh Road,<br />

Tainan, Taiwan, e-mail: kuoym@mail.ncku.edu.tw<br />

Neurogenesis has been suggested to correlate with certain aspects <str<strong>on</strong>g>of</str<strong>on</strong>g> brain cognitive functi<strong>on</strong>. Several<br />

extrinsic stimuli, including physical exercise is known to have advantageous effects <strong>on</strong> neurogenesis and brain<br />

functi<strong>on</strong>. However, why exercise promotes neurogenesis remains unknown. Previously, stress-induced high<br />

level <str<strong>on</strong>g>of</str<strong>on</strong>g> corticoster<strong>on</strong>e was shown to inhibit neurogenesis, while adrenalectomy enhance neurogenesis. As<br />

exercise modulates the level <str<strong>on</strong>g>of</str<strong>on</strong>g> serum corticoster<strong>on</strong>e, we hypothesize that corticoster<strong>on</strong>e signaling pathway is<br />

involved in the exercise-induced neurogenesis. In this study, hippocampal neurogenesis is estimated by<br />

double-labeling <str<strong>on</strong>g>of</str<strong>on</strong>g> mitotic marker (BrdU) and neur<strong>on</strong>al progenitor marker (DCX) in the subgranule z<strong>on</strong>e.<br />

Our results showed that treadmill running exercise (TRE) significantly enhanced neurogenesis. The levels <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

serum corticoster<strong>on</strong>e were elevated immediately and decreased 4 h after TRE. Five-week TRE downregulated<br />

the protein levels <str<strong>on</strong>g>of</str<strong>on</strong>g> mineralocoticoid receptor (MR), while glucocorticoid receptor (GR) remained<br />

unaltered. To mimic the exercise-elicited corticoster<strong>on</strong>e resp<strong>on</strong>se, mice were injected with low dose <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

corticoster<strong>on</strong>e (4mg/kg/day) <str<strong>on</strong>g>for</str<strong>on</strong>g> three weeks. Our results showed that mice received corticoster<strong>on</strong>e injecti<strong>on</strong><br />

had slightly more BrdU/DCX positive cells than those <str<strong>on</strong>g>of</str<strong>on</strong>g> vehicle c<strong>on</strong>trols with no changes in the MR and<br />

GR levels. Alternatively, mice received MR antag<strong>on</strong>ist, spir<strong>on</strong>olact<strong>on</strong>e, treatment 90 min be<str<strong>on</strong>g>for</str<strong>on</strong>g>e TRE<br />

exhibited more neurogenesis than that <str<strong>on</strong>g>of</str<strong>on</strong>g> TRE. Spir<strong>on</strong>olact<strong>on</strong>e al<strong>on</strong>e also expressed significant effect <strong>on</strong><br />

neurogenesis. Taken together, our results suggest that reduced corticoster<strong>on</strong>e signaling pathway is involved in<br />

the TRE-induced hippocampal neurogenesis.<br />

399


23-26 August 2007,<br />

Budapest, Hungary<br />

7A_12_P<br />

(poster secti<strong>on</strong> A2, poster board #86, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ADOLESCENTS COPING WITH PERSONAL SECURITY STRESS:<br />

STRATEGIES AND ADJUSTMENT<br />

Hanna Laufer<br />

The Academic College <str<strong>on</strong>g>of</str<strong>on</strong>g> Emek Yezreel, Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Human Services, P.O. box 277, Timrat, 23840, Israel<br />

e-mail: hlaufer@research.haifa.ac.il<br />

Living in areas <str<strong>on</strong>g>of</str<strong>on</strong>g> intensive combat and c<strong>on</strong>flict z<strong>on</strong>es, may be accompanied by psychological stress am<strong>on</strong>g<br />

adolescents. The present study makes an attempt to explore the influence <str<strong>on</strong>g>of</str<strong>on</strong>g> the combat z<strong>on</strong>e geographical<br />

proximity <strong>on</strong> the stress assessment, and to detect and define the specific resulting resp<strong>on</strong>ses <str<strong>on</strong>g>of</str<strong>on</strong>g> adolescents.<br />

A comparis<strong>on</strong> was d<strong>on</strong>e between adolescents living in communities al<strong>on</strong>g the seam-z<strong>on</strong>e, and adolescents<br />

living in communities remote from Israel’s borders, in the purpose to investigate whether a difference exists<br />

between the adaptati<strong>on</strong> levels <str<strong>on</strong>g>of</str<strong>on</strong>g> adolescents exposed to military stress - to those who are not.<br />

The test sample included 279 males and females, ages 14 to 18.<br />

It was found that adolescents living in the seam-z<strong>on</strong>e vicinity reported a primary stress evaluati<strong>on</strong> level higher<br />

than that <str<strong>on</strong>g>of</str<strong>on</strong>g> adolescents from the comparis<strong>on</strong> group. In spite <str<strong>on</strong>g>of</str<strong>on</strong>g> this finding - no significant differences <strong>on</strong><br />

the sec<strong>on</strong>dary stress evaluati<strong>on</strong> (c<strong>on</strong>trol estimati<strong>on</strong> level) was observed between the two groups. It was found<br />

that adolescents, who revealed the lowest adaptati<strong>on</strong> level, were those who reported the highest level <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

unc<strong>on</strong>trollability.<br />

Another finding indicates that stress situati<strong>on</strong>s, evaluated as unc<strong>on</strong>trollable, lead to an intense use <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

sec<strong>on</strong>dary c<strong>on</strong>trol engagement strategies (e.g. distracti<strong>on</strong>, positive thinking, cognitive restructuring and<br />

acceptance) and the disengagement coping strategies (e.g. avoidance, denial, wishful thinking). It was also<br />

found that <str<strong>on</strong>g>for</str<strong>on</strong>g> both groups - coping strategies <str<strong>on</strong>g>of</str<strong>on</strong>g> the primary c<strong>on</strong>trol types (problem solving; emoti<strong>on</strong>al<br />

regulati<strong>on</strong> and emoti<strong>on</strong>al expressi<strong>on</strong>) predict a higher level <str<strong>on</strong>g>of</str<strong>on</strong>g> adaptati<strong>on</strong>.<br />

Based <strong>on</strong> these findings it was c<strong>on</strong>cluded that the stress type (c<strong>on</strong>trolled or unc<strong>on</strong>trolled) has an important<br />

effect <strong>on</strong> the predicti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> selected coping strategies and adaptive results. Gender differences were observed:<br />

the males, living in the vicinity <str<strong>on</strong>g>of</str<strong>on</strong>g> the seam-z<strong>on</strong>e, revealed the highest, while the females - the lowest level <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

adaptati<strong>on</strong>.<br />

It will be possible to utilize the present study findings in their adaptati<strong>on</strong> to the treatment and the educati<strong>on</strong><br />

policy <str<strong>on</strong>g>for</str<strong>on</strong>g> the more vulnerable part <str<strong>on</strong>g>of</str<strong>on</strong>g> the populati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> adolescents: the females.<br />

400


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7A_13_P<br />

(poster secti<strong>on</strong> A2, poster board #87, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECTS OF A PSYCHOSOCIAL STRESSOR ON SELF-INJURIOUS BEHAVIOR AND<br />

HYPOTHALAMIC-PITUITARY-ADRENOCORTICAL FUNCTION<br />

IN RHESUS MONKEYS<br />

J. S. Meyer, M. D. Davenport, C. K. Lutz, S. Tiefenbacher, M. A. Novak<br />

Univ. <str<strong>on</strong>g>of</str<strong>on</strong>g> Massachusetts, Amherst, MA 01003 USA; New England Primate <str<strong>on</strong>g>Research</str<strong>on</strong>g> Center<br />

Southborough, MA 01772 USA<br />

e-mail: jmeyer@psych.umass.edu<br />

Self-injurious behavior (SIB) is a disorder that occurs in both clinical and n<strong>on</strong>-clinical populati<strong>on</strong>s. Although<br />

a role <str<strong>on</strong>g>for</str<strong>on</strong>g> stress in the etiology and/or maintenance <str<strong>on</strong>g>of</str<strong>on</strong>g> SIB has been proposed, there is little empirical<br />

evidence <str<strong>on</strong>g>for</str<strong>on</strong>g> this hypothesis. Our laboratory has been studying the pathophysiology <str<strong>on</strong>g>of</str<strong>on</strong>g> sp<strong>on</strong>taneously<br />

occurring SIB (manifested as self-biting and occasi<strong>on</strong>al self-wounding) in singly housed male rhesus m<strong>on</strong>keys<br />

at the New England Primate <str<strong>on</strong>g>Research</str<strong>on</strong>g> Center. In the present study, we examined the short- and l<strong>on</strong>g-term<br />

effects <str<strong>on</strong>g>of</str<strong>on</strong>g> an administratively mandated relocati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> our m<strong>on</strong>keys (both SIB and c<strong>on</strong>trols) <strong>on</strong> the animals’<br />

behavior and hypothalamic-pituitary-adrenocortical (HPA) functi<strong>on</strong>. The relocati<strong>on</strong> involved movement <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the subjects to different cages in a new col<strong>on</strong>y room with unfamiliar animals. Daytime and nighttime<br />

behaviors were recorded, as well as time-dependent changes in salivary, serum, and hair cortisol<br />

c<strong>on</strong>centrati<strong>on</strong>s, and serum corticosteroid binding globulin (CBG). Relocati<strong>on</strong> stress induced l<strong>on</strong>g-lasting<br />

behavioral abnormalities in the SIB group, including an increase in self-biting and a disrupti<strong>on</strong> in sleep. Both<br />

groups exhibited rapid increases in salivary and serum cortisol following the move. Hair cortisol, which we<br />

have validated as an index <str<strong>on</strong>g>of</str<strong>on</strong>g> l<strong>on</strong>g-term HPA system activity (Davenport et al., 2006), was elevated over<br />

baseline at 4 m<strong>on</strong>ths post-move but not at 1 year. In c<strong>on</strong>trast, serum cortisol remained increased over<br />

baseline at 1 year, a point at which we also observed a compensatory rise in serum CBG. These results may<br />

have relevance <str<strong>on</strong>g>for</str<strong>on</strong>g> humans exposed to major life stressors.<br />

7A_14_P<br />

(poster secti<strong>on</strong> A2, poster board #88, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ASSOCIATION OF STAGE OF STRESS MANAGEMENT BEHAVIOR WITH<br />

PERCEIVED STRESS AND COPING IN JAPANESE COLLEGE STUDENTS<br />

Hiroyoshi Murayama 1 , Yoshiyuki Tanaka 2 , Sakurako Ito 1 , Junpei Yajima 3 , Akira Tsuda 1 ,<br />

Shigeko Tsuda 4<br />

1 Kurume University, Kurume, Japan<br />

2 Tokyo University <str<strong>on</strong>g>of</str<strong>on</strong>g> Social Welfare, Isezaki, Japan<br />

3 Beppu University, Beppu, Japan<br />

4 Fukuoka Jogakuin University, Fukuoka, Japan<br />

e-mail: hiro-murayama@umin.net<br />

Background Transtheoretical model (TTM) is a theory <str<strong>on</strong>g>of</str<strong>on</strong>g> behavior change that can be applied to single,<br />

multiple, and complex behavior targets (Prochaska & DiClemente, 1986). The TTM is best known <str<strong>on</strong>g>for</str<strong>on</strong>g> its<br />

applicati<strong>on</strong>s to smoking, high-fat diet, and drinking. TTM assumes five stages <str<strong>on</strong>g>of</str<strong>on</strong>g> change, which are named as<br />

Pre-c<strong>on</strong>templati<strong>on</strong> (PC), C<strong>on</strong>templati<strong>on</strong> (C), Preparati<strong>on</strong> (P), Acti<strong>on</strong> (A), and Maintenance (M) stage. There<br />

401


23-26 August 2007,<br />

Budapest, Hungary<br />

are thus far few attempts that apply TTM to change in effective stress management behavior in Japan. To<br />

understand change in stress management behavior by TTM, this study is to examine associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> stage <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

stress management behavior with perceived stress and coping in Japanese college students.<br />

Method Participants were 699 (243 male and 456 female) college students in Fukuoka. The mean ages were<br />

19.7 and 19.3 years, respectively. The measures were 1) Stage <str<strong>on</strong>g>of</str<strong>on</strong>g> change algorithm, 2) a Japanese versi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Rode Island Stress and Coping Inventory, and 3) a Japanese versi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress Management Behaviors<br />

Inventory c<strong>on</strong>sisting <str<strong>on</strong>g>of</str<strong>on</strong>g> four subscales such as setting a limit, planning, reframing, and unhealthy behaviors.<br />

Result and Discussi<strong>on</strong> Porti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> students were 35 % in PC, 15 % in C, 16 % in P, 11 % in A, and 23 % in<br />

M stage, respectively. Degree <str<strong>on</strong>g>of</str<strong>on</strong>g> perceived stress declined as stage proceeds, whereas perceived frequency <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

effective coping increased across proceeding <str<strong>on</strong>g>of</str<strong>on</strong>g> stages. Subjects in A and M stage showed lower frequency <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

unhealthy behaviors, and higher planning and reframing behaviors. These results replicate previous findings<br />

using American sample that effective stress management behaviors are associated with lower levels <str<strong>on</strong>g>of</str<strong>on</strong>g> stress<br />

and higher levels <str<strong>on</strong>g>of</str<strong>on</strong>g> appropriate coping, and provided support <str<strong>on</strong>g>for</str<strong>on</strong>g> usefulness <str<strong>on</strong>g>of</str<strong>on</strong>g> applying TTM to stress<br />

management behavior in Japan.<br />

7A_15_P<br />

(poster secti<strong>on</strong> A2, poster board #89, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

RELIABILITY AND VALIDITY OF LAYERED VOICE ANALYSIS TECHNOLOGY IN<br />

THE DETECTION OF MENTAL STRESS<br />

K. Nemoto 1 , H. Tachikawa 1 , T. Takao 1 , H. Sato 1 , Y. Ashizawa 1 , G. Endo 1 , K. Tanaka 1 , R. Ishii 1 ,<br />

N. Ishii 1 , K. Hashimoto 1 , T. Iguchi 1 , S. Hada 2 , M. Hori 3 , T. Asada 3<br />

1<br />

Psycholos<str<strong>on</strong>g>of</str<strong>on</strong>g>t, 1-1-1 Tennodai, Tsukuba, Japan; 2 Alegria Co., Ltd., Tokyo, Japan; 3 Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychiatry,<br />

Univ. <str<strong>on</strong>g>of</str<strong>on</strong>g> Tsukuba, Tsukuba, Japan<br />

e-mail: kiyotaka@nemotos.net<br />

It is known that speech signal c<strong>on</strong>tains features which provide in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> about a human speaker. Although<br />

several technologies to detect stress using human voice are available, reports <strong>on</strong> the reliability and validity <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

these technologies are c<strong>on</strong>troversial. In this study, we investigated the reliability and validity <str<strong>on</strong>g>of</str<strong>on</strong>g> the Layered<br />

Voice Analysis (LVA) technology. Methods: One-hundred and six healthy subjects participated this study.<br />

First, stress was assessed by using Speilberger State-Trait Anxiety Inventory (STAI). Blood pressure (BP) was<br />

also measured. Then, subjects were randomly assigned to the anagram task group and c<strong>on</strong>trol group. Be<str<strong>on</strong>g>for</str<strong>on</strong>g>e<br />

task begins, all <str<strong>on</strong>g>of</str<strong>on</strong>g> the subjects were asked to answer 10 questi<strong>on</strong>s vocally, and they were all recorded. After<br />

answering questi<strong>on</strong>s, task group underwent anagram task whereas c<strong>on</strong>trol group just read aloud series <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

words. After the task, STAI-S and BP were measured again. Answers to each questi<strong>on</strong> were analyzed using<br />

LVA and 22 parameters were computed. The internal c<strong>on</strong>sistency was assessed <str<strong>on</strong>g>for</str<strong>on</strong>g> each parameter using<br />

answers be<str<strong>on</strong>g>for</str<strong>on</strong>g>e task. Two-sample t-test was per<str<strong>on</strong>g>for</str<strong>on</strong>g>med to see if parameters change significantly due to<br />

anagram task. Results: Of 22 parameters, Cr<strong>on</strong>bach’s alpha <str<strong>on</strong>g>of</str<strong>on</strong>g> 18 parameters was more than 0.6. Two-sample<br />

t-test showed that 10 <str<strong>on</strong>g>of</str<strong>on</strong>g> 18 parameters al<strong>on</strong>g with STAI-S and systolic BP changed significantly during the<br />

anagram task. C<strong>on</strong>clusi<strong>on</strong>: Most <str<strong>on</strong>g>of</str<strong>on</strong>g> the parameters LVA computed are reliable and the value <str<strong>on</strong>g>of</str<strong>on</strong>g> these<br />

parameters changed significantly under stressful c<strong>on</strong>diti<strong>on</strong>s. LVA might be useful in the detecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> mental<br />

stress.<br />

402


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7A_16_P<br />

(poster secti<strong>on</strong> A2, poster board #90, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

WIDOWHOOD INCREASES RISK FOR SUBSEQUENT DEMENTIA, ESPECIALLY<br />

FOR WOMEN: THE CACHE COUNTY STUDY<br />

M. C. Nort<strong>on</strong> 1 , J. T. Tschanz 1 , T. Østbye 2 , C. Corcoran 1 , P. P. Zandi 3 , J. C. S. Breitner 4 ,<br />

K. A. Welsh-Bohmer 2<br />

1 Utah State University<br />

2 Duke University<br />

3 The Johns Hopkins University<br />

4 University <str<strong>on</strong>g>of</str<strong>on</strong>g> Washingt<strong>on</strong><br />

A host <str<strong>on</strong>g>of</str<strong>on</strong>g> life stressors increase risk <str<strong>on</strong>g>for</str<strong>on</strong>g> depressi<strong>on</strong> in late life, most notably widowhood. Although<br />

depressi<strong>on</strong>, in turn, increases risk <str<strong>on</strong>g>for</str<strong>on</strong>g> dementia, few studies have examined the direct associati<strong>on</strong> between<br />

late-life stressors and subsequent dementia <strong>on</strong>set. Even more rare are large, populati<strong>on</strong>-based l<strong>on</strong>gitudinal<br />

studies addressing this research questi<strong>on</strong>, with their ability to minimize the selecti<strong>on</strong> bias inherent in clinic<br />

based samples. In Cache County, Utah (USA) elderly (aged 65+) residents were assessed <str<strong>on</strong>g>for</str<strong>on</strong>g> dementia (90%<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> entire county participated) with a multistage case ascertainment protocol in 1995-6 (baseline) and those<br />

without dementia at baseline were again assessed in 1998-9 and 2002-3. The l<strong>on</strong>gitudinal sample with at least<br />

two evaluati<strong>on</strong>s included 3,117 pers<strong>on</strong>s and <str<strong>on</strong>g>of</str<strong>on</strong>g> these, 2,231 were married, 789 were widowed, and 97 were<br />

separated or divorced at baseline. Cox proporti<strong>on</strong>al hazards regressi<strong>on</strong> was employed to model effect <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

marital status, <strong>on</strong> dementia-free survival (years) be<str<strong>on</strong>g>for</str<strong>on</strong>g>e and after adjustment <str<strong>on</strong>g>for</str<strong>on</strong>g> gender, age, and presence <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

APOE e4 allele. Be<str<strong>on</strong>g>for</str<strong>on</strong>g>e adjustment <str<strong>on</strong>g>for</str<strong>on</strong>g> covariates, widowhood (at baseline) was associated with significantly<br />

higher incident dementia compared to being married (HR=2.37, 95%CI: 1.96-2.87). A trend <str<strong>on</strong>g>for</str<strong>on</strong>g> the marital<br />

status*gender interacti<strong>on</strong> was observed (p=.140) after covariate adjustments. Compared to being married,<br />

widowhood was associated with increased hazard <str<strong>on</strong>g>for</str<strong>on</strong>g> dementia in women <strong>on</strong>ly (HR=1.77, 95%CI: 1.00-3.12).<br />

Future studies will examine factors that may mitigate increased risk, and the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> more recent<br />

widowhood and multiple marital transiti<strong>on</strong>s.<br />

7A_17_P<br />

(poster secti<strong>on</strong> A2, poster board #91, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STRESS AND MEMORY INTERACTION : DECLARATIVE MEMORY IMPAIRMENT<br />

FOLLOWING A 5-DAY MILITARY COMMANDO OPERATION<br />

C. Piérard 1 , M. Pérès 1 , J. C. Jouanin 1 , P. Liscia 1 , C. Y. Guézennec, D. Béracochéa 2<br />

1IMASSA - BP 73 - F-91223 Brétigny-sur-Orge Cedex<br />

2LNC - Université Bordeaux 1 - F-33405 Talence Cedex<br />

A commando operati<strong>on</strong> is composed <str<strong>on</strong>g>of</str<strong>on</strong>g> multiple routines and skills that require alertness, attenti<strong>on</strong> to<br />

external envir<strong>on</strong>ment, memory per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance and reactivity to events. As the declarative memory system is<br />

very sensitive to envir<strong>on</strong>mental interacti<strong>on</strong>s and stress-generating situati<strong>on</strong>s, we decided to specifically study<br />

the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> a 5-day commando operati<strong>on</strong> <strong>on</strong> this system.<br />

Cognitive and memory per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance <str<strong>on</strong>g>of</str<strong>on</strong>g> 21 male cadets was examined 3 weeks be<str<strong>on</strong>g>for</str<strong>on</strong>g>e and at the end <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

operati<strong>on</strong>, using an original computerized cognitive test battery allowing field investigati<strong>on</strong>. In a first step, the<br />

battery evaluated psychomotor and cognitive per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance with : a subjective vigilance and mood testing, a<br />

visuomotor coordinati<strong>on</strong> test, a selective attenti<strong>on</strong> test, and finally a planning test. In a sec<strong>on</strong>d step, more<br />

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23-26 August 2007,<br />

Budapest, Hungary<br />

specific memory tests were administered to investigate short-term comp<strong>on</strong>ents <strong>on</strong> the <strong>on</strong>e hand, and l<strong>on</strong>gterm<br />

comp<strong>on</strong>ents <strong>on</strong> the other hand. For each item, the battery measured the reacti<strong>on</strong> time and calculated the<br />

percentages <str<strong>on</strong>g>of</str<strong>on</strong>g> good and wr<strong>on</strong>g resp<strong>on</strong>ses, as well as the percentage <str<strong>on</strong>g>of</str<strong>on</strong>g> omissi<strong>on</strong>s.<br />

Our results evidenced that a 5-day commando operati<strong>on</strong> significantly increased reacti<strong>on</strong> times. In accordance<br />

with the relative but significant hypovigilance <str<strong>on</strong>g>of</str<strong>on</strong>g> the subjects, selective attenti<strong>on</strong> and short-term memory were<br />

affected (memory span, visual memory and audiovisual associati<strong>on</strong>). More surprisingly, l<strong>on</strong>g-term memory<br />

(semantic memory) was also impaired. On the c<strong>on</strong>trary, spatial working memory and planning tasks were<br />

spared.<br />

Finally, this field study dem<strong>on</strong>strates that the stress induced by a prol<strong>on</strong>ged commando operati<strong>on</strong> is able to<br />

selectively impair several comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g> declarative memory.<br />

7A_18_P<br />

(poster secti<strong>on</strong> A2, poster board #92, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

LEARN TO SURVIVE CHRONIC STRESS! LONG-TERM BENEFITS OF A<br />

STRUCTURED BEHAVIOURAL INTERVENTION PROGRAM<br />

Adrienne Stauder*, Piroska Balog*, Tamás Martos*, Mónika Kovács*, Virginia Williams**,<br />

Red<str<strong>on</strong>g>for</str<strong>on</strong>g>d Williams***<br />

* Semmelweis University, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Behavioural Sciences, 1089 Budapest, Nagyvárad tér 4<br />

e-mail: staadr@net.sote.hu<br />

**Williams LifeSkills Inc, Durham, NC, USA<br />

***Psychiatry and Behavioural Sciences, Duke University Medical Center, Durham, NC, USA<br />

We studied the effectiveness <str<strong>on</strong>g>of</str<strong>on</strong>g> a 16-hour structured stress management training, the Hungarian versi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the Williams LifeSkills program. Since November 2004 more than 400 pers<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> diverse background were<br />

trained. We m<strong>on</strong>itored short-term and l<strong>on</strong>g-term outcomes in a sample <str<strong>on</strong>g>of</str<strong>on</strong>g> 24 people with high distress (mean<br />

age 34,4 ys, sd=10,35). Data were collected be<str<strong>on</strong>g>for</str<strong>on</strong>g>e (t1), right after (t2) and 6 m<strong>on</strong>th after (t3) the interventi<strong>on</strong>.<br />

We used standardized questi<strong>on</strong>naires: Cohen Perceived Stress scale (PSS), Spielberger Trait Anxiety Inventory<br />

(STAIT), shortened Beck Depressi<strong>on</strong> Inventory (BDI), Patients Health Questi<strong>on</strong>naires (PHQ15), 5-item<br />

WHO Well-being Index (WWB), Rahe Life Meaning (MEAN) scale, and 1-item <strong>on</strong> “satisfacti<strong>on</strong> with life”<br />

(SAT). Statistics: paired sample T test, levels <str<strong>on</strong>g>of</str<strong>on</strong>g> significance are * p


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7A_20_P<br />

(poster secti<strong>on</strong> A2, poster board #93, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CIRCULAR CAUSATION BETWEEN MAJOR LIFE EVENTS, DEPRESSION,<br />

AND WELLBEING<br />

Gabor Szabo, Zsuzsa Szanto, Maria Kopp<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Behavioural Sciences, Semmelweis University, H-1089 Budapest Nagyvarad ter 4, Hungary<br />

e-mail: szabgab@net.sote.hu<br />

Object: Study <str<strong>on</strong>g>of</str<strong>on</strong>g> the circular causati<strong>on</strong> between Rahe-Holmes Major Life Events (MLE), depressi<strong>on</strong> and<br />

wellbeing using data from a Hungarian follow-up survey Hungarostudy Epidemiological Panel (HEP2006).<br />

Sample and methods: From participants <str<strong>on</strong>g>of</str<strong>on</strong>g> a nati<strong>on</strong>wide questi<strong>on</strong>naire survey in 2002, 4528 adult pers<strong>on</strong>s<br />

participated in the sec<strong>on</strong>d wave <str<strong>on</strong>g>of</str<strong>on</strong>g> the survey in 2005. Data were collected about the MLE experienced<br />

during the periods preceding the first and the sec<strong>on</strong>d waves, and the shortened WHO Wellbeing Score (WS)<br />

and Beck Depressi<strong>on</strong> Inventory (BDI). The follow-up enabled us to examine the circular cause-and-effect<br />

relati<strong>on</strong>ships between MLE and psychological states. Results: The relati<strong>on</strong>ships between MLE stress scores<br />

and the number <str<strong>on</strong>g>of</str<strong>on</strong>g> stressful MLE were uncertain because they did hardly or not correlated with the BDI and<br />

the WS. However, including <strong>on</strong>ly the number <str<strong>on</strong>g>of</str<strong>on</strong>g> the negative MLE we found str<strong>on</strong>g relati<strong>on</strong>ships. (1) In<br />

cross-secti<strong>on</strong>al terms, in both waves we found str<strong>on</strong>g relati<strong>on</strong>ships between the number <str<strong>on</strong>g>of</str<strong>on</strong>g> negative MLE,<br />

BDI (0,23***; 0,17***) and WS (−0,15***,−0,14***). (2) The number <str<strong>on</strong>g>of</str<strong>on</strong>g> negative MLE, BDI and WS showed<br />

stability between waves (0,15***; 0,27***; 0,26***). (3) The cross-correlati<strong>on</strong>s showed that previous negative<br />

MLE were str<strong>on</strong>gly related to later BDI and WS (0,17***, −0,12***) while earlier BDI and WS showed a<br />

weaker but significant relati<strong>on</strong>ship with later negative MLE (0,09***, −0,07***). Discussi<strong>on</strong>: Negative MLE<br />

result in permanent stress while the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> positive MLE are uncertain. Although the psychological status<br />

has some effect <strong>on</strong> MLE much more robust effects can be found in the opposite directi<strong>on</strong>: negative MLE<br />

result in the increase <str<strong>on</strong>g>of</str<strong>on</strong>g> BDI and decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> WS.<br />

7A_21_P<br />

(poster secti<strong>on</strong> A2, poster board #94, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SUBJECTIVE WELL-BEING AFFECTION TOWARD DEVELOPMENT OF STRESS-<br />

MANAGEMENT BEHAVIORS<br />

Tanaka Yoshiyuki 1 , Tsuda Akira 2 , Egami Yuko 3 , Jingu Sumie 3<br />

1Tokyo University <str<strong>on</strong>g>of</str<strong>on</strong>g> Social Welfare, Isesaki, Japan<br />

2Kurume University, Kurume, Japan<br />

3Fukuoka Health Promoti<strong>on</strong> Foundati<strong>on</strong>, Fukuoka, Japan<br />

e-mail: ytanaka0519@yahoo.co.jp<br />

The present study describes how development <str<strong>on</strong>g>of</str<strong>on</strong>g> stress-management behaviors (SMBs) was different depend<br />

<strong>on</strong> the level <str<strong>on</strong>g>of</str<strong>on</strong>g> individual subjective well-being. The subjects were 282 participants (56 male and 226 female)<br />

in the 3 m<strong>on</strong>ths health promoti<strong>on</strong> program. Be<str<strong>on</strong>g>for</str<strong>on</strong>g>e and after the program, subjects were asked whether they<br />

had each <str<strong>on</strong>g>of</str<strong>on</strong>g> 8 SMBs. Subjective well-being was assessed at the pre-program by the revised versi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Psychological Lively Scale (PLS-R) which c<strong>on</strong>sists <str<strong>on</strong>g>of</str<strong>on</strong>g> 4 subscales: life satisfacti<strong>on</strong>, negative mood, vitality to<br />

challenge, and emoti<strong>on</strong>al stability. In the result, the development depended <strong>on</strong> the level <str<strong>on</strong>g>of</str<strong>on</strong>g> PLS-R scores.<br />

Most <str<strong>on</strong>g>of</str<strong>on</strong>g> the subjects with high pre-life satisfacti<strong>on</strong> could have passed <str<strong>on</strong>g>of</str<strong>on</strong>g>f their weariness easily, got something<br />

to change their bad mood or daily stress, and had some pleasure after the program; even though, low pre-life<br />

405


23-26 August 2007,<br />

Budapest, Hungary<br />

satisfacti<strong>on</strong> subjects could not. Same relati<strong>on</strong> was found between the level <str<strong>on</strong>g>of</str<strong>on</strong>g> vitality to challenge and their<br />

weariness, emoti<strong>on</strong>al stability and their weariness, emoti<strong>on</strong>al stability and their good sleep. The low prenegative<br />

mood, c<strong>on</strong>versely, related to the change <str<strong>on</strong>g>of</str<strong>on</strong>g> their good sleep, some pleasure, and nice way to pass <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

weariness. The augmentati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> SMB number was also related to the PLS-R score. This result was also<br />

related to the pre-number <str<strong>on</strong>g>of</str<strong>on</strong>g> SMBs. All subjects having few pre-SMBs multiplied the number, though<br />

subjects with high pre-PLS-R changed much more (4.3 to 6.2). While the number <str<strong>on</strong>g>of</str<strong>on</strong>g> SMBs with low pre-PLS-<br />

R in many pre-SMBs was not significantly changed, high pre-PLS-R subjects developed their SMBs even their<br />

pre-SMBs were many. These results revealed that high subjective well-being accelerate the development <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

SMBs.<br />

7A_22_P<br />

(poster secti<strong>on</strong> A2, poster board #95, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

IS AN INCREASE IN STRESS TOLERANCE CAPACITY CONDUCIVE TO<br />

LESSENING PSYCHOSOCIAL STRESS IN AN APPARENTLY HEALTHY<br />

POPULATION<br />

Kazuhiko Yamamoto 1 , Yoko Sakamoto 1 , Masahiro Irie 1 , Susumu Ohmori 2 , Mototaka Yoshinari 2<br />

1Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Health Science, Kyushu University, Fukuoka, 815-8540 Japan<br />

2Kyushu Central Hospital, Fukuoka, 815-8588 Japan<br />

The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to examine the associati<strong>on</strong>s between stress tolerance capacity measured using IMST<br />

(Inventory to measure stress tolerance capacity with 20 items) and a stress score measured using IMPS<br />

(Inventory to measure psychosocial stress with 40 items) am<strong>on</strong>g apparently healthy adults. 1) A total <str<strong>on</strong>g>of</str<strong>on</strong>g> 684<br />

male (47.1 ± 7.9 years) and 517 female public school workers (47.5 ± 7.0 years) resp<strong>on</strong>ded to questi<strong>on</strong>naires<br />

which assessed the degree <str<strong>on</strong>g>of</str<strong>on</strong>g> stress resp<strong>on</strong>se (i.e., the stress score) and the degree <str<strong>on</strong>g>of</str<strong>on</strong>g> stress tolerance capacity<br />

(i.e., the stress intolerance score). 2) The higher the stress intolerance score was, the higher the stress score was<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> both men and women. Stress scores <str<strong>on</strong>g>of</str<strong>on</strong>g> men who answered yes to 17 items <strong>on</strong> the IMST and <str<strong>on</strong>g>of</str<strong>on</strong>g> women<br />

who answered yes to 14 items were lower than those <str<strong>on</strong>g>of</str<strong>on</strong>g> men and women who answered no to the same items<br />

<strong>on</strong> this test. The higher the stress intolerance scores <str<strong>on</strong>g>of</str<strong>on</strong>g> men and women were, the higher were the<br />

proporti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> men who answered yes to 38 items and women who answered yes to 39 items <strong>on</strong> the IMPS. A<br />

stepwise regressi<strong>on</strong> model yielded 8 items <strong>on</strong> the IMST which influenced the stress score <str<strong>on</strong>g>for</str<strong>on</strong>g> men and 4 items<br />

which influenced it <str<strong>on</strong>g>for</str<strong>on</strong>g> women. An inverse associati<strong>on</strong> between the stress score and physical exercise was<br />

found to be significant in men, while this associati<strong>on</strong> was not found in women. The results suggest that<br />

ef<str<strong>on</strong>g>for</str<strong>on</strong>g>ts to increase the stress tolerance capacity, such as having appropriate social support, a healthy lifestyle,<br />

and a positive attitude toward life, may be c<strong>on</strong>ducive to lessening psychosocial stress am<strong>on</strong>g an otherwise<br />

healthy populati<strong>on</strong>. They also imply that a stress management program needs to treat men and women<br />

differentially. References: 1) Yamamoto K (2005) Development <str<strong>on</strong>g>of</str<strong>on</strong>g> the inventories to measure psychosocial<br />

stress and stress tolerance. Jpn J Physiol Anthropol 10: 67-77 [in Japanese], 2) Yamamoto, K. el al. (2007) The<br />

Relati<strong>on</strong>ship between IMPS-measured stress score and biomedical parameters regarding health status am<strong>on</strong>g<br />

public school workers. J Physiol Anthropol (in press).<br />

406


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7A_23_P<br />

(poster secti<strong>on</strong> A2, poster board #96, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DETERMINANTS OF ALCOHOL USE AMONG UNIVERSITY STUDENTS: THE<br />

ROLE OF STRESS, COPING AND EXPECTANCIES<br />

Cecilia Chau<br />

P<strong>on</strong>tificia Universidad Catolica del Perú, Lima, Lima 32, Av. Universitaria s/n cuadra18, Lima, Paru<br />

e-mail: cchau@pucp.edu.pe<br />

The purpose <str<strong>on</strong>g>of</str<strong>on</strong>g> this investigati<strong>on</strong> were two: (1) to obtain a descripti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> alcohol c<strong>on</strong>sumpti<strong>on</strong> patterns<br />

am<strong>on</strong>g Peruvian university students in general, and <str<strong>on</strong>g>of</str<strong>on</strong>g> subgroups defined by gender and socioec<strong>on</strong>omic status<br />

(SES); and (2) to investigate the determinants <str<strong>on</strong>g>of</str<strong>on</strong>g> n<strong>on</strong>-problematic and problematic alcohol use am<strong>on</strong>g<br />

Peruvian university students, taking into account both characteristics <str<strong>on</strong>g>of</str<strong>on</strong>g> the social c<strong>on</strong>text and the<br />

psychosocial variables which mediate their influence.<br />

We c<strong>on</strong>sider as predictor variables: Socioec<strong>on</strong>omic status, gender, parents’ use <str<strong>on</strong>g>of</str<strong>on</strong>g> alcohol and best male and<br />

female friends. The mediating variables were: perceived stress, coping styles and alcohol expectancies. These<br />

two sets <str<strong>on</strong>g>of</str<strong>on</strong>g> variables were related to use <str<strong>on</strong>g>of</str<strong>on</strong>g> alcohol during the last six m<strong>on</strong>ths. Questi<strong>on</strong>naires were<br />

administered to the participants from three universities (N = 1081): <strong>on</strong>e public university and two private<br />

universities.<br />

The main findings indicate that most university students in Peru c<strong>on</strong>sume alcoholic beverages. The<br />

prevalence <str<strong>on</strong>g>of</str<strong>on</strong>g> alcohol use appears to be lower <str<strong>on</strong>g>for</str<strong>on</strong>g> students who attend a public university, whereas almost all<br />

students from the private universities drink alcohol. Students present a low frequency <str<strong>on</strong>g>of</str<strong>on</strong>g> drinking during<br />

weekdays, but c<strong>on</strong>sumpti<strong>on</strong> increases during the weekends, beer being the preferred beverage. C<strong>on</strong>sumpti<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> alcohol is also related to tobacco and marihuana. Significant differences in alcohol c<strong>on</strong>sumpti<strong>on</strong> patterns<br />

were found <str<strong>on</strong>g>for</str<strong>on</strong>g> gender and SES. Parental drinking was found to have a significant influence <strong>on</strong> their<br />

children’s alcohol use in general. More best male friends than best female friends drink alcohol.<br />

To discriminate between drinkers and n<strong>on</strong>-drinkers, the most significant variables were: alcohol c<strong>on</strong>sumpti<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> best female or male friends, positive expectancies in pers<strong>on</strong>al/social aspects <str<strong>on</strong>g>of</str<strong>on</strong>g> drinking, SES, negative<br />

expectancies in pers<strong>on</strong>al/social aspects <str<strong>on</strong>g>of</str<strong>on</strong>g> drinking, and gender.<br />

The results <str<strong>on</strong>g>of</str<strong>on</strong>g> a discriminant analysis between risky and n<strong>on</strong>-risky drinkers indicate that positive alcohol<br />

expectancies in pers<strong>on</strong>al/ social aspects play a major role, al<strong>on</strong>g with gender, female and best female friends’<br />

alcohol use, and avoidant coping styles.<br />

7A_25_P<br />

(poster secti<strong>on</strong> A2, poster board #97, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SEX DIFFERENCES AND THE ROLES OF TESTOSTERONE IN FEAR<br />

POTENTIATED STARTLE<br />

Lung Yu<br />

Inst <str<strong>on</strong>g>of</str<strong>on</strong>g> Behavioral Medicine, Nat’l Cheng Kung Univ Coll <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, 1 University Rd.,<br />

Tainan, Taiwan 70101, ROC<br />

e-mail: lungyu@mail.ncku.edu.tw<br />

This study aims to examine the sex differences in the acquisiti<strong>on</strong> and extincti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the fear potentiated startle<br />

and the roles <str<strong>on</strong>g>of</str<strong>on</strong>g> testoster<strong>on</strong>e in central amygdala in mediating such sex differences. Male and female S-D rats<br />

aged 7-8 weeks <str<strong>on</strong>g>of</str<strong>on</strong>g> age were first accustomed to the c<strong>on</strong>straint chamber in the dark and their startle resp<strong>on</strong>ses<br />

407


23-26 August 2007,<br />

Budapest, Hungary<br />

were recorded then <str<strong>on</strong>g>for</str<strong>on</strong>g> the sporadic t<strong>on</strong>e presentati<strong>on</strong> (95dB). Our fear c<strong>on</strong>diti<strong>on</strong>ing protocol c<strong>on</strong>sisted <str<strong>on</strong>g>of</str<strong>on</strong>g> an<br />

episode <str<strong>on</strong>g>of</str<strong>on</strong>g> 10-trial light-foot shock pairings. Three c<strong>on</strong>secutive days (30 trials/day) <str<strong>on</strong>g>of</str<strong>on</strong>g> light al<strong>on</strong>e presentati<strong>on</strong><br />

served as <strong>on</strong>e bout <str<strong>on</strong>g>of</str<strong>on</strong>g> the extincti<strong>on</strong> protocol. A total <str<strong>on</strong>g>of</str<strong>on</strong>g> two bouts <str<strong>on</strong>g>of</str<strong>on</strong>g> extincti<strong>on</strong> protocol were used. Bilateral<br />

g<strong>on</strong>adectomy, systemic estradiol and testoster<strong>on</strong>e supplement, and intra-accumbal injecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> g<strong>on</strong>adal<br />

horm<strong>on</strong>es were used in appropriate groups. We found that female rats exhibited greater fear potentiated<br />

startle resp<strong>on</strong>ses in the acquisiti<strong>on</strong> test and lesser degree <str<strong>on</strong>g>of</str<strong>on</strong>g> extincti<strong>on</strong> after the sec<strong>on</strong>d bout <str<strong>on</strong>g>of</str<strong>on</strong>g> the extincti<strong>on</strong><br />

protocol as compared to male rats. We further dem<strong>on</strong>strated that ovariectomized rats displayed similar fear<br />

potentiated startle acquisiti<strong>on</strong> and extincti<strong>on</strong> as the intact female rats, while orchidectomized rats displayed<br />

greater degree <str<strong>on</strong>g>of</str<strong>on</strong>g> potentiated startle resp<strong>on</strong>ses and more resistant to the extincti<strong>on</strong> protocol as compared to<br />

the intact male rats. Finally, intra-accumbal testoster<strong>on</strong>e infusi<strong>on</strong> was found to effectively reverse such<br />

sensitized potentiated startle resp<strong>on</strong>ses and resistence to extincti<strong>on</strong>. We c<strong>on</strong>clude that testoster<strong>on</strong>e may play a<br />

pivotal role in determining the sexual differences <str<strong>on</strong>g>of</str<strong>on</strong>g> fear memory acquisiti<strong>on</strong> and <str<strong>on</strong>g>for</str<strong>on</strong>g>getting process.<br />

408


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7B. PSYCHOSOCIAL STRESS AND DISEASE<br />

(HOLGER URSIN)<br />

7B_01_S<br />

STRESS AND COMMON HEALTH COMPLAINTS<br />

Holger Ursin<br />

Unifob health, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Bergen, Norway<br />

According to the Cognitive Activati<strong>on</strong> Theory <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress (CATS – Ursin and Eriksen 2004), a <str<strong>on</strong>g>for</str<strong>on</strong>g>mal system<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> systematic definiti<strong>on</strong>s, the term “stress” is used <str<strong>on</strong>g>for</str<strong>on</strong>g> stress stimuli, the stress experience, the n<strong>on</strong>-specific,<br />

general stress resp<strong>on</strong>se, and the experience <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress resp<strong>on</strong>se. The stress resp<strong>on</strong>se is normal, healthy, and<br />

necessary alarm. If sustained there may be a risk <str<strong>on</strong>g>of</str<strong>on</strong>g> illness and disease. The level and durati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the alarm<br />

depends <strong>on</strong> the expectancy <str<strong>on</strong>g>of</str<strong>on</strong>g> the outcome <str<strong>on</strong>g>of</str<strong>on</strong>g> stimuli and the specific resp<strong>on</strong>ses available <str<strong>on</strong>g>for</str<strong>on</strong>g> coping.<br />

The most comm<strong>on</strong> health complaints are subjective health complaints like muscle pain, tiredness and mood<br />

changes. These are normal aches <str<strong>on</strong>g>of</str<strong>on</strong>g> short durati<strong>on</strong> and low intensity <str<strong>on</strong>g>for</str<strong>on</strong>g> most people. For some the pains and<br />

complaints are substantial and l<strong>on</strong>g-lasting with serious implicati<strong>on</strong>s <str<strong>on</strong>g>for</str<strong>on</strong>g> functi<strong>on</strong>ing. There are no sharp or<br />

obvious limits in the distributi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> health complaints, separating ‘normal’ and endurable pain and<br />

complaints, and intolerable complaints that need pr<str<strong>on</strong>g>of</str<strong>on</strong>g>essi<strong>on</strong>al help. These c<strong>on</strong>diti<strong>on</strong>s are most <str<strong>on</strong>g>of</str<strong>on</strong>g>ten<br />

unspecific, and are the most comm<strong>on</strong> reas<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> encounters with health pr<str<strong>on</strong>g>of</str<strong>on</strong>g>essi<strong>on</strong>als, and the most frequent<br />

reas<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> sick leave and disability. There is a striking comorbidity <str<strong>on</strong>g>for</str<strong>on</strong>g> all these c<strong>on</strong>diti<strong>on</strong>s. This may be<br />

explained by psychobiological sensitizati<strong>on</strong> within neural loops, which has been suggested as a mechanism <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

these c<strong>on</strong>diti<strong>on</strong>s (Ursin 1997).<br />

7B_02_S<br />

LOW-ENERGY STRESS-RELATED DISORDERS. DIAGNOSTIC UTILITY OF BIO-<br />

PSYCHOSOCIAL MARKERS<br />

Bengt B. Arnetz 1,2 , Ulla Maria Anderberg 2 , Ingrid Anderzen 2 , Christina Hal<str<strong>on</strong>g>for</str<strong>on</strong>g>d 2<br />

1Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Occupati<strong>on</strong>al and Envir<strong>on</strong>mental Health, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Family Medicine and Public Health Sciences,<br />

Wayne State University, Detroit, Michigan 48009, USA<br />

e-mail: barnetz@med.wayne.edu<br />

2 Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Social Medicine, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Public Health Sciences, Uppsala University, Uppsala Sciences Park,<br />

751 05 Uppsala, Sweden<br />

It is comm<strong>on</strong>ly difficult to differentiate low energy states due to stress-related disorders from depressi<strong>on</strong>. 1<br />

The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> the present study was to evaluate possible differences in symptoms, psychiatric co-diagnosis, and<br />

biological stress markers in patients suffering from stress-related disorders versus depressi<strong>on</strong>. 150 patients<br />

that had been referred by primary care physicians to an academic stress medicine center were evaluated by a<br />

physician using structured psychiatric tools, the center’s validated stress-assessment visual analogue scales,<br />

biological stress markers, blood pressure, and Body Mass Index. Patients with depressi<strong>on</strong>, >20 points <strong>on</strong><br />

MADRS, scored significantly lower <strong>on</strong> the global energy scale, self rated health, and quality <str<strong>on</strong>g>of</str<strong>on</strong>g> sleep as<br />

compared to patients suffering from stress-related disorders. Depressed patients scored higher <strong>on</strong> the global<br />

stress scale. Depressed patients rated themselves to be less rested following a night’s sleep. Depressed<br />

patients scored lower with regard to satisfacti<strong>on</strong> with their family and job situati<strong>on</strong>s. It is suggested that<br />

studies involving either stress-related disorders or depressi<strong>on</strong> more carefully screen the participants with<br />

409


23-26 August 2007,<br />

Budapest, Hungary<br />

regard to biopsychosocial characteristics. A substantial number <str<strong>on</strong>g>of</str<strong>on</strong>g> patients being referred <str<strong>on</strong>g>for</str<strong>on</strong>g> stress-related<br />

disorders also suffer from psychiatric co-morbidity.<br />

410<br />

7B_03_S<br />

STRESS AND THE HEART: THE PHYSIOLOGICAL BASIS FOR THE<br />

DEVELOPMENT OF CARDIAC RISK IN DEPRESSION AND ANXIETY DISODERS<br />

Gavin Lambert<br />

Baker Heart <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute, Melbourne, Australia, e-mail: gavin.lambert@baker.edu.au<br />

Until recently it was thought that no more than 50% <str<strong>on</strong>g>of</str<strong>on</strong>g> clinical cor<strong>on</strong>ary heart disease was explicable in terms<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> classical cardiac risk factors such as dyslipidemia, cigarette smoking, high blood pressure and diabetes.<br />

Recent large scale epidemiological studies have increased our understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> the mechanisms generating<br />

cardiac risk and have provided evidence indicating that psychosocial factors, particularly depressive illness<br />

(MDD), anxiety states, and acute and chr<strong>on</strong>ic mental stress are involved here, “triggering” clinical<br />

cardiovascular events, and possibly also c<strong>on</strong>tributing to hypertensi<strong>on</strong> and atherosclerosis development.<br />

Although the underlying mechanisms in play are most likely multi factorial in origin the sympathetic nervous<br />

system is undoubtedly paramount in many cases. Using noradrenaline isotope diluti<strong>on</strong> methodology and<br />

direct nerve recording coupled with invasive blood sampling techniques we have examined cardiac<br />

sympathetic activity in patients with MDD and also panic disorder (PD). By sampling blood also from the<br />

internal jugular vein we have been able to gain insight into brain m<strong>on</strong>oamine turnover in these c<strong>on</strong>diti<strong>on</strong>s and<br />

have directly evaluated the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> SSRI therapy <strong>on</strong> brain m<strong>on</strong>oamines in these c<strong>on</strong>diti<strong>on</strong>s. The pattern <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

sympathetic activati<strong>on</strong> is very different in patients with MDD (bimodal) or PD (normal). Brain m<strong>on</strong>omaine<br />

turnover is different in each group, with brain noradrenaline turnover being reduced in MDD yet normal in<br />

PD. Interestingly, in both MDD and PD brain serot<strong>on</strong>in turnover, surprisingly, is markedly elevated be<str<strong>on</strong>g>for</str<strong>on</strong>g>e<br />

treatment and is significantly diminished following SSRI therapy. Clearly, the role <str<strong>on</strong>g>of</str<strong>on</strong>g> brain m<strong>on</strong>oamines and<br />

their relati<strong>on</strong> to generating increased cardiac risk merits further attenti<strong>on</strong>.<br />

7B_04_S<br />

INTERMITTENT NEUROGENIC STRESS DELAYS ONSET OF DIABETES IN RATS:<br />

SELYE’S EUSTRESS<br />

Holly E. Bates, A. Sirek, M. A. Kiraly, J. T. Y. Yue, D. Goche M<strong>on</strong>tes, S. G. Matthews, M. Vranic<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Tor<strong>on</strong>to, Tor<strong>on</strong>to, Canada, e-mail: holdoug@yahoo.ca<br />

Acute stress deteriorates glycemic c<strong>on</strong>trol in diabetes. We hypothesized that chr<strong>on</strong>ic intermittent restraint<br />

stress (1hr/d, 5d/wk <str<strong>on</strong>g>for</str<strong>on</strong>g> 13wks) <str<strong>on</strong>g>of</str<strong>on</strong>g> male ZDF rats, a model <str<strong>on</strong>g>of</str<strong>on</strong>g> Type 2 Diabetes, would accelerate<br />

development <str<strong>on</strong>g>of</str<strong>on</strong>g> diabetes. Intermittent stress lowered food intake by 15%. Thus, pair fed rats were included<br />

to distinguish between the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> reduced food intake and <str<strong>on</strong>g>of</str<strong>on</strong>g> stress per se. Surprisingly, intermittent stress<br />

delayed development <str<strong>on</strong>g>of</str<strong>on</strong>g> fed and fasting hyperglycemia, effects mediated partly by reduced food intake, but<br />

also by intermittent stress per se. The stress-induced reducti<strong>on</strong> in food intake improved insulin secreti<strong>on</strong> and<br />

β-cell mass, suggesting these as mechanisms <str<strong>on</strong>g>for</str<strong>on</strong>g> reduced food intake to improve glycemia. However,<br />

intermittent stress per se also led to HPA axis adaptati<strong>on</strong>s. Although basal ACTH levels did not differ,<br />

intermittent stress prevented the 30% increase in basal corticoster<strong>on</strong>e (CORT) with food restricti<strong>on</strong>, which<br />

could predict 20% <str<strong>on</strong>g>of</str<strong>on</strong>g> the variati<strong>on</strong> in fed glycemia. In additi<strong>on</strong>, intermittent stress led to habituati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

restraint-induced CORT resp<strong>on</strong>ses to lower levels than those induced by food removal. These CORT


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

adaptati<strong>on</strong>s with intermittent stress were c<strong>on</strong>sistent with adaptati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> mRNA levels <str<strong>on</strong>g>for</str<strong>on</strong>g> hippocampal MR,<br />

paraventricular nucleus AVP, and anterior pituitary POMC. Thus, intermittent restraint stress per se delays<br />

hyperglycemia, presumably via adaptati<strong>on</strong>s in the HPA axis that prevent the hypercorticoster<strong>on</strong>emia caused<br />

by food restricti<strong>on</strong>. Since hypercortisolemia deteriorates glycemic c<strong>on</strong>trol, adaptati<strong>on</strong> to repetitive,<br />

predictable stress may be beneficial <str<strong>on</strong>g>for</str<strong>on</strong>g> glucose regulati<strong>on</strong>. Thus, as Hans Selye noted when coining eustress,<br />

not all stress is deleterious.<br />

7B_05_S<br />

(poster secti<strong>on</strong> A2, poster board #98, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STRESS, QUALITY OF LIFE AND MULTIPLE CHRONIC DISEASES: PATTERNS<br />

EMERGING FROM A LARGE NATIONAL SAMPLE, AUSTRALIA<br />

Agnes Walker<br />

<str<strong>on</strong>g>Australian</str<strong>on</strong>g> <str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g> <str<strong>on</strong>g>Ec<strong>on</strong>omic</str<strong>on</strong>g> <str<strong>on</strong>g>Research</str<strong>on</strong>g> <strong>on</strong> Health, <str<strong>on</strong>g>Australian</str<strong>on</strong>g> Nati<strong>on</strong>al University Canberra 0200 Australia<br />

e-mail: Agnes.Walker@anu.edu.au<br />

Objectives: to study the associati<strong>on</strong>s between multiple chr<strong>on</strong>ic diseases and quality <str<strong>on</strong>g>of</str<strong>on</strong>g> life and psychological<br />

distress scores.<br />

Methods: analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> unit record cross secti<strong>on</strong>al data from <str<strong>on</strong>g>Australian</str<strong>on</strong>g> nati<strong>on</strong>al surveys <str<strong>on</strong>g>for</str<strong>on</strong>g> the populati<strong>on</strong> aged<br />

20 years or more. Identificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> an appropriate indicator <str<strong>on</strong>g>of</str<strong>on</strong>g> multiple chr<strong>on</strong>ic diseases (ie comorbidities).<br />

Use <str<strong>on</strong>g>of</str<strong>on</strong>g> logistic regressi<strong>on</strong> techniques to study associati<strong>on</strong>s between (a) comorbidities and demographic,<br />

socioec<strong>on</strong>omic and risk factor variables and (b) quality <str<strong>on</strong>g>of</str<strong>on</strong>g> life (general and psychological distress) and<br />

demographic, socioec<strong>on</strong>omic and health status indicators.<br />

Results: older people, obese pers<strong>on</strong>s, women, pers<strong>on</strong>s with low socioec<strong>on</strong>omic status and those living al<strong>on</strong>e<br />

had significantly greater probability <str<strong>on</strong>g>of</str<strong>on</strong>g> having three or more chr<strong>on</strong>ic illnesses than did other 20+ year olds<br />

(p


23-26 August 2007,<br />

Budapest, Hungary<br />

the equati<strong>on</strong>, while perceived stress at baseline (measure by the 10-item versi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> PSS, aiming at stress the<br />

last m<strong>on</strong>th) was entered in the third step. The result showed that perceived stress level at baseline significantly<br />

predicted (R 2 =.049, p


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7B_01_P<br />

(poster secti<strong>on</strong> A2, poster board #100, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DEPRESSION IN RELATION WITH DIFFERENT SOURCES OF STRESS<br />

Piroska Balog, Szilvia Adam, Tamas Martos, Anna Susanszky, Adrienn Stauder, Krisztina Neculai,<br />

Maria S. Kopp<br />

Semmelweis University, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Behavioral Sciences, 1089, Budapest, Nagyvarad ter 4.,<br />

e-mail: balopir@net.sote.hu<br />

The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to investigate the role <str<strong>on</strong>g>of</str<strong>on</strong>g> different sources <str<strong>on</strong>g>of</str<strong>on</strong>g> stress (work-, marital stress, and the<br />

role <str<strong>on</strong>g>of</str<strong>on</strong>g> life goals <str<strong>on</strong>g>for</str<strong>on</strong>g> both sexes, and gender-role stress <str<strong>on</strong>g>for</str<strong>on</strong>g> men, and work-family c<strong>on</strong>flict <str<strong>on</strong>g>for</str<strong>on</strong>g> women) in<br />

c<strong>on</strong>necti<strong>on</strong> with depressive symptoms. The study is based <strong>on</strong> Hungarostudy Epidemiological Panel 2006 (N<br />

4528). We analyzed a subsample <str<strong>on</strong>g>of</str<strong>on</strong>g> N= 1679, aged 18-65, actively working and living with partner (men<br />

47.4% women 52.6%. Am<strong>on</strong>g them 17% men and 20% women reported elevated depressi<strong>on</strong> score<br />

(Shortened Beck Depressi<strong>on</strong> Scale). To measure sources <str<strong>on</strong>g>of</str<strong>on</strong>g> stress we used shortened versi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g>: Marital<br />

Stress Scale, Ef<str<strong>on</strong>g>for</str<strong>on</strong>g>t – Reward Imbalance Questi<strong>on</strong>naire, Aspirati<strong>on</strong> Index, Masculine-Gender Role Stress<br />

Scale, and <strong>on</strong>e questi<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> work-family c<strong>on</strong>flict. Hierarchical logistic regressi<strong>on</strong> analyses were per<str<strong>on</strong>g>for</str<strong>on</strong>g>med to<br />

study the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> different sources <str<strong>on</strong>g>of</str<strong>on</strong>g> stress <strong>on</strong> depressi<strong>on</strong>, and we calculated odds ratios (OR) with 95%<br />

c<strong>on</strong>fidence intervals. Age and educati<strong>on</strong> were also included in the model. For women age (OR 1, 04 (1,02-<br />

1,06), educati<strong>on</strong> (OR 0,86 (0,75-0,98), marital stress (OR 1,97 (1,24-3,12), work stress (OR 2,40 (1,66-3,45),<br />

and work-family c<strong>on</strong>flict (OR 1,27 (1,05-1,55) were all related to elevated depressi<strong>on</strong> scores at step 2. At step<br />

3 when extrinsic life goals were included in the model, age, marital stress, work stress and stress from workfamily<br />

c<strong>on</strong>flict were still significantly related to depressi<strong>on</strong>, but not the educati<strong>on</strong>. We found extrinsic life<br />

goals as possible mediator between educati<strong>on</strong> and depressi<strong>on</strong> (OR 2,14 (1,44-3,18). In men extrinsic life goals<br />

have no effect <strong>on</strong> depressi<strong>on</strong>. Neither age nor educati<strong>on</strong> was related to depressi<strong>on</strong>. We found in men <strong>on</strong>ly<br />

marital stress (OR 1,91 (1,05-3,48) and work stress (OR 2,45 (1,60-3,73) significantly related to depressi<strong>on</strong>.<br />

7B_03_P<br />

(poster secti<strong>on</strong> A2, poster board #101, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DAILY WORRY PREDICTS HOSPITALIZATION AFTER CORONARY ARTERY AND<br />

VALVE SURGERY<br />

Piroska Balog, Zsuzsa Cserep, Aniko Malik, Eszter Los<strong>on</strong>cz, Andrea Szekely<br />

Semmelweis University, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Behavioral Sciences and Gottsegen Gyorgy Hungarian Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Cardiology,<br />

e-mail: balopir@net.sote.hu<br />

The relati<strong>on</strong>ships between psychosocial factors and future cardiac events have been investigated mainly in<br />

populati<strong>on</strong>-based studies. Few data are available about the l<strong>on</strong>g-term effects <str<strong>on</strong>g>of</str<strong>on</strong>g> n<strong>on</strong>-clinical variables as worry<br />

<strong>on</strong> the outcome after cardiac surgery. 180 patients who underwent cardiac surgery using cardiopulm<strong>on</strong>ary<br />

bypass were prospectively studied and followed up <str<strong>on</strong>g>for</str<strong>on</strong>g> 4 years. Age, educati<strong>on</strong>, living status, worry (4 items<br />

from Spielberger State-Trait Anxiety Inventory), depressi<strong>on</strong> (Beck Depressi<strong>on</strong> Inventory), perioperative<br />

characteristics and clinical risk factors (EUROscore, postoperative c<strong>on</strong>gestive heart failure, durati<strong>on</strong> at ICU<br />

and hospital stay) were also assessed. Psychological self-report questi<strong>on</strong>naires were completed preoperatively<br />

and 6, 12, 24, 36, 48 m<strong>on</strong>th after discharge. Clinical end-points were cardiac events requiring hospitalizati<strong>on</strong><br />

during follow-up. Hierarchical logistic regressi<strong>on</strong> analyses were per<str<strong>on</strong>g>for</str<strong>on</strong>g>med to study the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> daily worry<br />

<strong>on</strong> hospitalizati<strong>on</strong>, and we calculated odds ratios (OR) with 95% c<strong>on</strong>fidence intervals. In the last step were all<br />

413


23-26 August 2007,<br />

Budapest, Hungary<br />

analyzed variables were included in the model, postoperative c<strong>on</strong>gestive heart failure (OR 4,77 (1,27-17,93)),<br />

EUROscore (OR 1,84 (0,71-1,004)) and the mean <str<strong>on</strong>g>of</str<strong>on</strong>g> daily worries measured at different time points (OR 1,40<br />

(1,03-3,18)) were significantly related to hospitalizati<strong>on</strong> during follow-up. Our results support the noti<strong>on</strong> that<br />

daily stressors as worry would predict hospitalizati<strong>on</strong> after cor<strong>on</strong>ary artery and valve surgery.<br />

7B_04_P<br />

(poster secti<strong>on</strong> A2, poster board #102, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

GENETIC VARIATION IN THE HUMAN NORADRENALINE<br />

TRANSPORTER GENE<br />

Richard Bayles, Elisabeth Lambert, Josefin Forstrom, Katherine Shields*, Amanda Eddy*,<br />

Jeremy Jowett*, John Blangero**, Tye Dawood, Gavin Lambert<br />

Baker Heart <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute & *Internati<strong>on</strong>al Diabetes Institute, Melbourne, Australia. **Southwest<br />

Foundati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> Biomedical <str<strong>on</strong>g>Research</str<strong>on</strong>g>, San Ant<strong>on</strong>io, TX, USA<br />

e-mail: richard.bayles@baker.edu.au<br />

The functi<strong>on</strong>al integrity <str<strong>on</strong>g>of</str<strong>on</strong>g> the noradrenaline transporter (NET) is essential <str<strong>on</strong>g>for</str<strong>on</strong>g> the inactivati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> neur<strong>on</strong>ally<br />

released noradrenaline. A number <str<strong>on</strong>g>of</str<strong>on</strong>g> clinical c<strong>on</strong>diti<strong>on</strong>s have been identified in which there is phenotypic<br />

evidence <str<strong>on</strong>g>of</str<strong>on</strong>g> a cardiac defect in NET functi<strong>on</strong>, as determined by the extracti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> tritiated noradrenaline<br />

across the heart. An abnormality in neur<strong>on</strong>al noradrenaline reuptake could sensitise the heart to sympathetic<br />

activati<strong>on</strong>. Patients with panic disorder, major depressive disorder, postural orthostatic tachycardia syndrome<br />

and essential hypertensi<strong>on</strong> were identified as having phenotypic evidence <str<strong>on</strong>g>of</str<strong>on</strong>g> a defect in functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> NET. The<br />

mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> reduced NET activity in these patients remains unknown but it might have a genetic origin.<br />

The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this project is to investigate the possible genetic mechanisms resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> impaired NET<br />

functi<strong>on</strong>. In the present study, 97 publicly available single nucleotide polymorphisms (SNPs) were selected<br />

from NCBI’s dbSNP database within the NET gene. SNPs were selected based <strong>on</strong> their probability <str<strong>on</strong>g>of</str<strong>on</strong>g> having<br />

a functi<strong>on</strong>al role. These were genotyped in 210 unrelated individuals and variati<strong>on</strong> assessed <str<strong>on</strong>g>for</str<strong>on</strong>g> their influence<br />

<strong>on</strong> several phenotypic measures related to noradrenergic metabolism. Of the 97 SNPs selected, 37 were<br />

found to be polymorphic in our sample group. Based <strong>on</strong> preliminary robust Bayesian quantitative trait<br />

nucleotide analysis, several SNPs show c<strong>on</strong>sistent evidence <str<strong>on</strong>g>for</str<strong>on</strong>g> associati<strong>on</strong> with clinical diagnosis and<br />

catecholamine levels.<br />

7B_05_P<br />

(poster secti<strong>on</strong> A2, poster board #103, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PSYCHOSOCIAL STRESS IN AN OUTPATIENT REHABILITATION CLINIC<br />

E. Benkö, C. Kovacs, B. Novy, D. Kargl, G. Beier, S. Kowatsch, R. W. Kurz<br />

Zentrum für Ambulante Rehabilitati<strong>on</strong> der Pensi<strong>on</strong>sversicherungsanstalt<br />

Ärztlicher Leiter: Prim. Univ.-Doz. Dr. Robert Kurz, Wehlistr. 127, 1027 Vienna<br />

e-mail: edith.benkoe@pva.sozvers.at<br />

Psychosocial stress plays an important role in the development <str<strong>on</strong>g>of</str<strong>on</strong>g> physical ailments. Significant gender effects<br />

have been shown to exist be<str<strong>on</strong>g>for</str<strong>on</strong>g>e the manifestati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these ailments. The following study seeks to explore the<br />

type and extent <str<strong>on</strong>g>of</str<strong>on</strong>g> acute and chr<strong>on</strong>ic stress am<strong>on</strong>g male and female patients with chr<strong>on</strong>ic illnesses in<br />

414


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

outpatient rehabilitati<strong>on</strong> clinics. It also provides normative data <str<strong>on</strong>g>for</str<strong>on</strong>g> the measurement <str<strong>on</strong>g>of</str<strong>on</strong>g> acute stress using the<br />

analog scale “stress barometer ” and the Trierer Inventory <str<strong>on</strong>g>for</str<strong>on</strong>g> the Assessment <str<strong>on</strong>g>of</str<strong>on</strong>g> Chr<strong>on</strong>ic Stress.<br />

Method: Cross-secti<strong>on</strong>al study using a self-administered questi<strong>on</strong>naire<br />

Instruments: Trier Inventory <str<strong>on</strong>g>for</str<strong>on</strong>g> the Assessment <str<strong>on</strong>g>of</str<strong>on</strong>g> Chr<strong>on</strong>ic Stress (TICS), an analog scale “stress barometer”<br />

as a measure <str<strong>on</strong>g>of</str<strong>on</strong>g> acute stress<br />

Statistics: Multivariate analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> variance<br />

Participants: 353 patients <str<strong>on</strong>g>of</str<strong>on</strong>g> an outpatient rehabilitati<strong>on</strong> clinic (ZAR: Zentrum für Ambulante Rehabilitati<strong>on</strong>)<br />

in Vienna, Austria with cardiovascular diseases and orthopedic illnesses. 58% <str<strong>on</strong>g>of</str<strong>on</strong>g> participants were male, 31%<br />

were currently employed. The mean age was 59.4 years.<br />

Results: 654 questi<strong>on</strong>naires were distributed, 509 returned, 353 complete data sets (resp<strong>on</strong>se rate 77 %). After<br />

correcting <str<strong>on</strong>g>for</str<strong>on</strong>g> the influence <str<strong>on</strong>g>of</str<strong>on</strong>g> age, multivariate analyses showed significant differences between men and<br />

women (p < .001), as well as between employed and retired patients (p < .001). In terms <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic stress,<br />

female patients reported being more overworked (p < .01), experiencing more social stress (p < .05), feeling a<br />

str<strong>on</strong>ger sense <str<strong>on</strong>g>of</str<strong>on</strong>g> insufficient support (p < .05) and having more chr<strong>on</strong>ic anxiety (p < .001). They also<br />

showed higher overall chr<strong>on</strong>ic stress screening scores than male patients (p < .01). Employed patients<br />

reported higher levels <str<strong>on</strong>g>of</str<strong>on</strong>g> acute stress than retired patients (p < .001). In terms <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic stress, they also had<br />

higher overall screening scores (p < .001). They reported being more overworked (p < .001) and experiencing<br />

more social stress (p < .001), more intense pressure to succeed (p < .001) as well as a str<strong>on</strong>ger sense <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

insufficient support (p < .05). . While working men reported the highest levels <str<strong>on</strong>g>of</str<strong>on</strong>g> stress from pressure <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

success, working women were most burdened by social stress. For both sexes, chr<strong>on</strong>ic anxiety was the sec<strong>on</strong>d<br />

most important source <str<strong>on</strong>g>of</str<strong>on</strong>g> stress.<br />

Discussi<strong>on</strong>: Psychosocial stress can be found not <strong>on</strong>ly be<str<strong>on</strong>g>for</str<strong>on</strong>g>e the manifestati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> illness, but also during<br />

rehabilitati<strong>on</strong>. There are significant differences depending <strong>on</strong> gender and employment. Psychological<br />

interventi<strong>on</strong>s aimed at helping patients cope with stress should take these differences into account.<br />

7B_06_P<br />

(poster secti<strong>on</strong> A2, poster board #104, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

FUNCTIONAL DECLINE: PHYSIOLOGICAL AND PSYCHOSOCIAL STRESS IN<br />

OLDER HOSPITALIZED PATIENTS<br />

Cheryl Chia-Hui Chen, Guan-Hua Huang, Charlotte Wang<br />

Nati<strong>on</strong>al Taiwan University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Nursing; Nati<strong>on</strong>al Chiao Tung University Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Statistics;<br />

Nati<strong>on</strong>al Taiwan University, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Epidemiology, Taipei, Taiwan<br />

e-mail: cheryl.chen@ha.mc.ntu.edu.tw<br />

Hospitalizati<strong>on</strong> remains a stress event <str<strong>on</strong>g>for</str<strong>on</strong>g> older people. Functi<strong>on</strong>al decline is a leading complicati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

hospitalizati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> older patients. Such decline is very stressful both in ec<strong>on</strong>omic and human terms. The<br />

purpose <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to describe and identify patterns <str<strong>on</strong>g>of</str<strong>on</strong>g> functi<strong>on</strong>al decline during and 6-m<strong>on</strong>th post<br />

hospitalizati<strong>on</strong> and to ascertain discrete indicators that signal different patterns <str<strong>on</strong>g>of</str<strong>on</strong>g> functi<strong>on</strong>al trajectories,<br />

using latent class analysis.<br />

A perspective cohort study was c<strong>on</strong>ducted <strong>on</strong> 296 older hospitalized patients aged 65 years and older who<br />

c<strong>on</strong>secutively admitted to five surgical and medical units at a tertiary medical center in northern Taiwan.<br />

Participants were assessed during their hospitalizati<strong>on</strong> (48 hours within admissi<strong>on</strong> and be<str<strong>on</strong>g>for</str<strong>on</strong>g>e discharge) and 3<br />

and 6 m<strong>on</strong>ths post discharge. Demographics, medicati<strong>on</strong> taken, co-morbidities, cognitive status, oral health,<br />

nutriti<strong>on</strong>al status, presence <str<strong>on</strong>g>of</str<strong>on</strong>g> social support and depressive symptoms, admissi<strong>on</strong> diagnosis, and length <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

stay were assessed in order to test their relati<strong>on</strong>ship with patterns <str<strong>on</strong>g>of</str<strong>on</strong>g> functi<strong>on</strong>al trajectories.<br />

415


23-26 August 2007,<br />

Budapest, Hungary<br />

Functi<strong>on</strong>al decline was comm<strong>on</strong> with the worse point occurred at discharge. Six m<strong>on</strong>ths post discharge, more<br />

than half <str<strong>on</strong>g>of</str<strong>on</strong>g> subjects (n=149, 50.3%) never returned to their admissi<strong>on</strong> functi<strong>on</strong>al status. Three patterns <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

functi<strong>on</strong>al trajectories were identified and age, gender, number <str<strong>on</strong>g>of</str<strong>on</strong>g> co-morbidities, cognitive status, nutriti<strong>on</strong>al<br />

status, and oral health status at admissi<strong>on</strong> were statistically associated with patterns <str<strong>on</strong>g>of</str<strong>on</strong>g> decline.<br />

Visualizing three different patterns <str<strong>on</strong>g>of</str<strong>on</strong>g> functi<strong>on</strong>al trajectories and studying indicators that signal such<br />

differences will help care providers understand how functi<strong>on</strong> changed and possible ways to mitigate<br />

physiological and psychosocial stress in older hospitalized patients.<br />

7B_07_P<br />

(poster secti<strong>on</strong> A2, poster board #105, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ACTIVATION OF THE HPA AXIS FOLLOWING SSRI ADMINISTRATION IN<br />

PATIENTS WITH MDD<br />

T. Dawood, D. Bart<strong>on</strong>, E. Lambert, A. Tilbrook * , J. Loose * , M. Esler, D. Haikerwal, D. Kaye,<br />

G. Lambert<br />

Baker Heart <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute and *M<strong>on</strong>ash University, Melbourne, Australia<br />

e-mail: tye.dawood@baker.edu.au<br />

Major depressive disorder (MDD) has been linked with hypothalamic-pituitary-adrenal (HPA) axis activati<strong>on</strong>.<br />

However, the interacti<strong>on</strong>s between the sympathoadrenal system and HPA axis in MDD are equivocal.<br />

Further, the acti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> selective serot<strong>on</strong>in reuptake inhibitors (SSRIs) <strong>on</strong> these systems are yet to be<br />

identified. Nineteen patients with MDD, but otherwise healthy, and 18 healthy c<strong>on</strong>trols were recruited from<br />

the community <str<strong>on</strong>g>for</str<strong>on</strong>g> plasma cortisol, ACTH and adrenaline measurements. Following 12 weeks <str<strong>on</strong>g>of</str<strong>on</strong>g> SSRI<br />

treatment, levels <str<strong>on</strong>g>of</str<strong>on</strong>g> these horm<strong>on</strong>es were again measured in MDD patients. Compared with c<strong>on</strong>trol subjects,<br />

patients with MDD had c<strong>on</strong>siderably higher state (32±2 vs 58±2, p=0.001) and trait (33±2 vs 63±1,<br />

p=0.001) anxiety scores. Indicative <str<strong>on</strong>g>of</str<strong>on</strong>g> HPA axis activati<strong>on</strong>, plasma cortisol c<strong>on</strong>centrati<strong>on</strong>s were significantly<br />

increased in untreated MDD [53(37-68) in c<strong>on</strong>trols and 69(64-92) in MDD, p=0.01, median (25-75<br />

percentile)]. Following treatment, state and trait anxiety levels were reduced in MDD patients (p=0.001).<br />

C<strong>on</strong>sistent with the decrease in anxiety scores, arterial adrenaline c<strong>on</strong>centrati<strong>on</strong>s decreased in MDD<br />

(49±6pg/mL to 36±5pg/mL, p=0.05). Surprisingly, and c<strong>on</strong>trary to adrenaline c<strong>on</strong>centrati<strong>on</strong>s, cortisol<br />

c<strong>on</strong>centrati<strong>on</strong>s increased following therapy (to 103±60ng/mL, p=0.075). Plasma ACTH levels also increased<br />

slightly after treatment. No correlati<strong>on</strong>s between HPA axis activity, symapthoadrenal activity and degree <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

depressi<strong>on</strong> or anxiety were observed, except <str<strong>on</strong>g>for</str<strong>on</strong>g> a significant relati<strong>on</strong>ship between plasma cortisol and ACTH<br />

following SSRI administrati<strong>on</strong> (r=0.7, p


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7B_08_P<br />

(poster secti<strong>on</strong> A2, poster board #106, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ASSESSMENT OF PERCEIVED STRESS: VALIDITY AND PROGNOSTIC<br />

PERFORMANCE OF THE ORIGINAL AND A SHORTER VERSION OF THE<br />

PERCEIVED STRESS QUESTIONNAIRE (PSQ)<br />

Herbert Fliege, Ricarda Joachim, Eva Peters, Petra Arck, Susan Levenstein*, Burghard F. Klapp<br />

Dpt. <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychosomatic Medicine, Charité Universitätsmedizin Berlin, Luisenstr. 13 A, D-10117 Berlin, Germany<br />

e-mail: herbert.fliege@charite.de<br />

Aventino Medical Group, Via della F<strong>on</strong>te di Fauno, 22, 00153 Rome, Italy<br />

The Perceived Stress Questi<strong>on</strong>naire (PSQ) by Levenstein et al. (1993) assesses subjectively experienced stress<br />

independently <str<strong>on</strong>g>of</str<strong>on</strong>g> a specific occasi<strong>on</strong>. It has been translated from English into many other languages, with<br />

validated adaptati<strong>on</strong>s in Italian, Spanish, Portuguese, and Swedish, and translati<strong>on</strong>s in Thai and Korean. In<br />

our own studies we translated, adapted and validated it in German.<br />

The paper presents data from several studies that give evidence <str<strong>on</strong>g>of</str<strong>on</strong>g> the psychometric qualities in different<br />

groups. A special emphasis is <strong>on</strong> how well the slightly shortened 20-item versi<strong>on</strong> (PSQ-20) per<str<strong>on</strong>g>for</str<strong>on</strong>g>ms with<br />

respect to stress and somatic outcomes as compared to the overall 30-item versi<strong>on</strong> (PSQ-30).<br />

Samples comprise patients with asthma, atopic dermatitis, tinnitus, inflammatory bowel diseases,<br />

somat<str<strong>on</strong>g>of</str<strong>on</strong>g>orm, affective or eating disorders, women during pregnancy, after sp<strong>on</strong>taneous aborti<strong>on</strong> or after<br />

regular delivery, healthy adults, and students, with over 3000 participants. Comparis<strong>on</strong>s between the 20-item<br />

and the original 30-item versi<strong>on</strong>s include a re-analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> the original ulcerative colitis sample’s prospective<br />

data.<br />

A structure <str<strong>on</strong>g>of</str<strong>on</strong>g> 4 factors (“worries”, “tensi<strong>on</strong>” and “joy” (reversed) as indicating stress reacti<strong>on</strong>, and<br />

“demands” indicating perceived stressors) proved fairly stable across different groups. Psychometric<br />

properties were good. Associati<strong>on</strong>s between stress, disease symptoms, physiological functi<strong>on</strong>ing (e.g. lung<br />

functi<strong>on</strong> in asthmatics) and immunological parameters are reported. A comparis<strong>on</strong> between the 30-item and<br />

the 20-item versi<strong>on</strong>s shows that the prognostic per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance c<strong>on</strong>cerning an exacerbati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ulcerative colitis<br />

over a period <str<strong>on</strong>g>of</str<strong>on</strong>g> up to 68 m<strong>on</strong>ths is maintained despite item reducti<strong>on</strong>.<br />

We propose the PSQ as a valid and ec<strong>on</strong>omic tool <str<strong>on</strong>g>for</str<strong>on</strong>g> research <strong>on</strong> perceived stress. The overall score <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

revised PSQ-20 permits comparis<strong>on</strong> with results from studies with the PSQ-30.<br />

7B_09_P<br />

(poster secti<strong>on</strong> A2, poster board #107, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DIFFERENTIAL EFFECTS OF FLUOXETINE ON ENERGY BALANCE AND<br />

CORTICOSTERONE RESPONSES TO REPEATED RESTRAINT STRESS IN 40D-<br />

AND 60D-OLD MALE RATS<br />

Sheila Pinto 2 , Francisca Gómez 1,2<br />

1Pharmacology Department, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmacy and 2 Brain Mapping Unit, Pluridiciplinary Institute,<br />

Complutense University <str<strong>on</strong>g>of</str<strong>on</strong>g> Madrid, Paseo Juan XXIII, 1, 28040 Madrid, Spain<br />

e-mail: gomez<str<strong>on</strong>g>of</str<strong>on</strong>g>@farm.ucm.es<br />

Fluoxetine (FLX) is widely used to treat depressi<strong>on</strong> in adolescent and adult depressive patients. FLX affects<br />

activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the hypothalamic-pituitary-adrenal (HPA) axis in rats but little is known about whether agedependent<br />

differences may exist regarding the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> FLX <strong>on</strong> HPA axis activity, as well as the possible<br />

417


23-26 August 2007,<br />

Budapest, Hungary<br />

interacti<strong>on</strong>s with the stress resp<strong>on</strong>se. We investigated the pubertal (40d) and adult (60d) rodent energy<br />

balance and corticoster<strong>on</strong>e (B) resp<strong>on</strong>ses to repeated restraint stress [(3h/day) x 3days], after daily injecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

either vehicle (VEH) or FLX (10mg/kg; 1ml/kg, ip). We show that independently <str<strong>on</strong>g>of</str<strong>on</strong>g> age, the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> FLX<br />

<strong>on</strong> food intake and body weight <str<strong>on</strong>g>of</str<strong>on</strong>g> unrestrained rats was comparable to the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> restraint in VEH-treated<br />

rats. Combinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> FLX and restraint resulted in additive-like effects. However, 40d rats maintained<br />

positive p<strong>on</strong>deral growth whereas 60d rats lost weight. Acutely, FLX blocked restraint-induced acute weight<br />

loss during the 3h <str<strong>on</strong>g>of</str<strong>on</strong>g> daily restraint <strong>on</strong>ly in 60d-old rats suggesting that FLX reduced energy expenditure. It is<br />

likely that this effect may alter compensatory mechanisms involved in energy balance regulati<strong>on</strong>. FLX<br />

increased plasma corticoster<strong>on</strong>e (B) levels, measured 1h after injecti<strong>on</strong>, similarly in 40d- and 60d-old rats.<br />

However, FLX prol<strong>on</strong>ged the acute and repeated restraint-induced increases in plasma B levels <strong>on</strong>ly in 60dold<br />

rats. Nevertheless, the B resp<strong>on</strong>se adapted with repetiti<strong>on</strong>. We c<strong>on</strong>clude that acute and repeated FLX<br />

treatment in unrestrained rats has stress-like effects <strong>on</strong> energy balance and HPA axis activity. Moreover, acute<br />

and repeated FLX treatment alters energy balance and HPA axis resp<strong>on</strong>ses to acute and repeated restraint in<br />

an age-dependent manner. Thus, 40d-old male rats are able to maintain positive energy balance and normal B<br />

resp<strong>on</strong>se to restraint whereas 60d-old male rats lose weight and show prol<strong>on</strong>ged B resp<strong>on</strong>ses to repeated<br />

restraint.<br />

7B_11_P<br />

(poster secti<strong>on</strong> A2, poster board #108, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

NEONATAL ENVIRONMENTS DIFFERENTIALLY AFFECT ANXIETY<br />

BEHAVIOUR AND MAMMARY GLAND DEVELOPMENT IN BALB/C MICE: A NEW<br />

LOOK AT THE STRESS AND BREAST CANCER RELATIONSHIP<br />

Leslie R. Kerr 1 , Ayehsa Salleh 1 , Tricia Ralph 1 , Chi Yip Ho 2<br />

1 Trent University, Peterborough, Ontario, Canada, K9J 7B8<br />

2 Microarray Laboratory, Samuel Lunenfeld <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute, Mount Sinai Hospital, Tor<strong>on</strong>to, Ontario, Canada<br />

Early life stressors may increase the risk <str<strong>on</strong>g>of</str<strong>on</strong>g> breast cancer. Although, several studies have produced<br />

inc<strong>on</strong>sistent support <str<strong>on</strong>g>for</str<strong>on</strong>g> the role <str<strong>on</strong>g>of</str<strong>on</strong>g> stressors in breast cancer risk. Because the mammary gland is <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> a<br />

few organ systems that completes its development postnatally, it may be uniquely susceptible to genetic and<br />

epigenetic modificati<strong>on</strong>s caused by ne<strong>on</strong>atal experiences. In rodents, as in humans, mother and infant<br />

interacti<strong>on</strong>s influence the development <str<strong>on</strong>g>of</str<strong>on</strong>g> the HPA axis, impacting <strong>on</strong> horm<strong>on</strong>e levels, immune resp<strong>on</strong>ses,<br />

reactivity to stressors, and possibly mammary gland development. Here, we examine how early life<br />

experiences differentially influence both anxiety-related behaviour and mammary gland development. Similar<br />

to other studies, we dem<strong>on</strong>strated that immobility in the <str<strong>on</strong>g>for</str<strong>on</strong>g>ced swim test was higher in maternally separated<br />

(MS; 4hrs/d, PND 2-22) than in handled (H; 15min/d; PND2-22) and unmanipulated, typically reared (TR)<br />

females, indicating increased anxiety and HPA axis reactivity in MS mice. Gross morphological analyses <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

mammary glands dem<strong>on</strong>strated that differentiati<strong>on</strong> rates (LAU /TEB) in mammary glands <str<strong>on</strong>g>of</str<strong>on</strong>g> adult H mice<br />

were higher than in both TR and MS mice (p’s


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7B_13_P<br />

(poster secti<strong>on</strong> A2, poster board #109, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SHAKER STRESS – A PSYCHOGENIC MODEL OF STRESS WITH USE IN<br />

TOXICOLOGY<br />

James B. Lucot 1 , Mojmir Mach 2 , Michal Dubovický 2<br />

1Dept. Pharmacology, Wright State University, Dayt<strong>on</strong>, OH, USA, e-mail: james.lucot@wright.edu<br />

2 Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Experimental Pharmacology, Slovak Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences, Bratislava 84104, Slovakia<br />

Animal models <str<strong>on</strong>g>of</str<strong>on</strong>g> stress can provide in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> about the course and etiology <str<strong>on</strong>g>of</str<strong>on</strong>g> stress related<br />

disorders, such as depressi<strong>on</strong> and anxiety. The incidence <str<strong>on</strong>g>of</str<strong>on</strong>g> these disorders c<strong>on</strong>stantly increases, with serious<br />

medical and socio-ec<strong>on</strong>omic c<strong>on</strong>sequences. Shaker stress with an unpredictable <strong>on</strong>set can be delivered over<br />

l<strong>on</strong>g periods <str<strong>on</strong>g>of</str<strong>on</strong>g> time and is a model <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic psychogenic disturbances. It is thus useful in studies <str<strong>on</strong>g>for</str<strong>on</strong>g> which<br />

classic stress models are not able to be used. We refined this model and increased its durati<strong>on</strong> to a week <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

our studies <str<strong>on</strong>g>of</str<strong>on</strong>g> the Gulf War Syndrome, where n<strong>on</strong>-predictability <str<strong>on</strong>g>of</str<strong>on</strong>g> stress was necessary to mimic battlefield<br />

uncertainties. Our results showed that exposure to chemical warfare agents (i.e. sarin) is more dangerous in<br />

combinati<strong>on</strong> with chr<strong>on</strong>ic intermittent shaker stress and that some toxicities have a delayed <strong>on</strong>set, as was<br />

observed in those with the Syndrome. This model also was used <str<strong>on</strong>g>for</str<strong>on</strong>g> behavioral phenotyping <str<strong>on</strong>g>of</str<strong>on</strong>g> stress<br />

resp<strong>on</strong>ses in oxytocin knockout (OTKO) mice. Acute shaker stress in OTKO male mice induced a larger<br />

stress resp<strong>on</strong>se compared to wild type mice. Neurochemical analysis revealed neural substrates <str<strong>on</strong>g>for</str<strong>on</strong>g> both<br />

psychopathology and interacti<strong>on</strong>s with toxins. These data dem<strong>on</strong>strate that this stress model is reproducible<br />

and well c<strong>on</strong>trolled. It is a relevant and flexible stress model which can be used in various studies in<br />

toxicology and behavioral phenotyping.<br />

7B_14_P<br />

(poster secti<strong>on</strong> A2, poster board #110, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

A QUANTITATIVE STUDY ON STRESS OF DISABLED AND RETIRED PEOPLE IN<br />

EYES OF POPULATION OF THE CZECH REPUBLIC<br />

Michal Matyska<br />

Masarykian University, Joštova 10, 602 00 Brno, CZE<br />

e-mail: balu@mail.muni.cz<br />

Sociological research is focused <strong>on</strong> relati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> public opini<strong>on</strong> and real stress status <str<strong>on</strong>g>of</str<strong>on</strong>g> disabled and retired<br />

people. <str<strong>on</strong>g>Research</str<strong>on</strong>g> was d<strong>on</strong>e by answering questi<strong>on</strong>s by targeted pers<strong>on</strong>s. First part was completed <strong>on</strong> healthful<br />

populati<strong>on</strong> (age +18, ec<strong>on</strong>omically productive, physically unhandicapped, total number <str<strong>on</strong>g>of</str<strong>on</strong>g> in<str<strong>on</strong>g>for</str<strong>on</strong>g>mants: 150<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> disabled and 132 <str<strong>on</strong>g>for</str<strong>on</strong>g> retired). Those people were put the questi<strong>on</strong>: „Do you think, that disabled/retired<br />

people are suffering from stress“ Five-points scale was every time used. Answer results: disabled people:<br />

61.3% yes (92), 16.7% rather so (25), 2% do not know (3), 16% rather not (24) and 4% no (4). Retired<br />

people: 64.4% yes (85), 14.4% rather so (19), 1.5% do not know (2), 13.6% rather not (18) and 6% no (8). As<br />

a whole, disabled people are supposed to be suffering from stress by 78% <str<strong>on</strong>g>of</str<strong>on</strong>g> populati<strong>on</strong> and retired people by<br />

78.8% <str<strong>on</strong>g>of</str<strong>on</strong>g> populati<strong>on</strong>. Sec<strong>on</strong>d part was d<strong>on</strong>e with disabled (total 48) and retired (total 75) people. Firstly, they<br />

were inquired, if they suffer from stress. Method is the same as above. As a whole, 31.2% (11+4) disabled<br />

people subjectively think, that they suffer from stress at least sometimes and 68.6% <str<strong>on</strong>g>of</str<strong>on</strong>g> disabled people state<br />

the minimal incidence <str<strong>on</strong>g>of</str<strong>on</strong>g> stress (25+8). 49.3% <str<strong>on</strong>g>of</str<strong>on</strong>g> retired people subjectively think, that they suffer from stress<br />

(6+31) and 50.6% <str<strong>on</strong>g>of</str<strong>on</strong>g> retired people do not c<strong>on</strong>firm this statement (26+12). Finally, psychological stress<br />

419


23-26 August 2007,<br />

Budapest, Hungary<br />

analysis was d<strong>on</strong>e (method in Cungi, Ch.. 1998. Savoir gérer s<strong>on</strong> stress. Paris: RETZ). Objectively 27.1% (13)<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> disabled and 56% (42) <str<strong>on</strong>g>of</str<strong>on</strong>g> retired people suffer from stress. Result is, that the most <str<strong>on</strong>g>of</str<strong>on</strong>g> in<str<strong>on</strong>g>for</str<strong>on</strong>g>mants in<br />

unhandicapped populati<strong>on</strong> do not correctly apprehend the real stress situati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> disabled people, but the<br />

opini<strong>on</strong> was right about retired.<br />

7B_15_P<br />

(poster secti<strong>on</strong> A2, poster board #111, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PSYCHOSOCIAL STRESS AND CHRONIC HEPATITIS C VIRUS (HCV)<br />

Elvira Micali, Giovanni Squadrito, Giuliana Amaddeo, Serena Callipari, Carmelo Abbate,<br />

Mario Barbaro, Giovanni Raim<strong>on</strong>do<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Messina, Medicine and Surgery, Messina, Italy<br />

Chr<strong>on</strong>ic hepatitis C produces extra-hepatic disorders and troubles. Symptomatic patients, that haven’t got<br />

absolute and/or relative c<strong>on</strong>tro-indicati<strong>on</strong>s, could be underwent to pharmacological treatment with<br />

interfer<strong>on</strong>e. The study means to give prominence to the <str<strong>on</strong>g>for</str<strong>on</strong>g>ms with which the HCV chr<strong>on</strong>ic subjects ( with<br />

genotype 2 and 3) react to the appearances <str<strong>on</strong>g>of</str<strong>on</strong>g> their physics functi<strong>on</strong>ality and the neuro-psychic<br />

symptomatology that comes out after the therapy with the interfer<strong>on</strong>e. It was administred by a clinical<br />

psychologist to 20 patients naïve (M=12 F=8) mean age 46,15 years, mean <str<strong>on</strong>g>of</str<strong>on</strong>g> schooling 12 years, all sick with<br />

chr<strong>on</strong>ic HCV, the Illness Behaviour Questi<strong>on</strong>naire (IBQ) mental reactive standardized and validated ;62<br />

items, that <str<strong>on</strong>g>for</str<strong>on</strong>g>m 7 scales, carry <strong>on</strong> the reacti<strong>on</strong>s and the feeling <str<strong>on</strong>g>of</str<strong>on</strong>g> the patients respect the disease and the<br />

percepti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> psychosocial situati<strong>on</strong>. The evalutati<strong>on</strong> was carried out at the beginning <str<strong>on</strong>g>of</str<strong>on</strong>g> the treatment with<br />

interfer<strong>on</strong>e and after 6 m<strong>on</strong>ths <str<strong>on</strong>g>of</str<strong>on</strong>g> therapy. IBQ, at the beginning <str<strong>on</strong>g>of</str<strong>on</strong>g> the pharmacological treatment, with<br />

positive value <str<strong>on</strong>g>of</str<strong>on</strong>g> HCV-RNA, registered that 9 subjects <str<strong>on</strong>g>of</str<strong>on</strong>g> 20 had significant score <str<strong>on</strong>g>for</str<strong>on</strong>g> the scales “c<strong>on</strong>victi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

disease” and “affective inhibiti<strong>on</strong>”.<br />

After six m<strong>on</strong>ths <str<strong>on</strong>g>of</str<strong>on</strong>g> assumpti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> interfer<strong>on</strong>e the haematologic exams gave prominence to a persisten<br />

reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HCV-RNA, while the IBQ scale: “hypoch<strong>on</strong>dria”, “psycho-somatic percepti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> disease”, “dysphoria”,<br />

“negati<strong>on</strong>”, ” irritability”, gave prominence to a significant clinically score <str<strong>on</strong>g>for</str<strong>on</strong>g> all <str<strong>on</strong>g>of</str<strong>on</strong>g> the sample. Our results,<br />

obtained with IBQ, they make assume that aspecific physiological answer <str<strong>on</strong>g>of</str<strong>on</strong>g> the human physical and the<br />

complicated reacti<strong>on</strong>s that follow to a pathological c<strong>on</strong>diti<strong>on</strong> and the specific pharmacological treatment<br />

produce psycho-emoti<strong>on</strong>al modificati<strong>on</strong>s and psyco-somatic stress.<br />

7B_16_P<br />

(poster secti<strong>on</strong> A2, poster board #112, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE STRESS-ASTHMA RELATIONSHIP IN CHILDREN: SOME PROGRESS<br />

TOWARDS SOLVING THE PUZZLE<br />

Seija Sandberg<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Mental Health Sciences, University College L<strong>on</strong>d<strong>on</strong><br />

While there is still an open verdict <str<strong>on</strong>g>for</str<strong>on</strong>g> the role <str<strong>on</strong>g>of</str<strong>on</strong>g> stress in the pathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> childhood asthma, a growing<br />

body <str<strong>on</strong>g>of</str<strong>on</strong>g> research suggests that psychosocial stress is a c<strong>on</strong>tributory factor development <str<strong>on</strong>g>of</str<strong>on</strong>g> asthma especially<br />

during early childhood. However, the area <str<strong>on</strong>g>of</str<strong>on</strong>g> study where the evidence is fairly robust c<strong>on</strong>cerns stressful<br />

experiences exacerbating the symptoms <str<strong>on</strong>g>of</str<strong>on</strong>g> asthma in children.<br />

420


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Despite this robust pattern <str<strong>on</strong>g>of</str<strong>on</strong>g> findings, little is known about the underlying mechanisms resp<strong>on</strong>sible.<br />

Experimental studies have produced inc<strong>on</strong>sistent findings. One reas<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> this may lie in the difficulty <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

experimentally creating true stress c<strong>on</strong>diti<strong>on</strong>s – instead, the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> emoti<strong>on</strong>al arousal have mostly been<br />

measured. Only animal studies have so far managed to examine mediators <str<strong>on</strong>g>of</str<strong>on</strong>g> stress-induced pathways in<br />

some detail.<br />

The presentati<strong>on</strong> will first briefly review the results <str<strong>on</strong>g>of</str<strong>on</strong>g> a prospective study by the author and colleagues<br />

(Sandberg et al, 2000, 2004) in c<strong>on</strong>juncti<strong>on</strong> with those <str<strong>on</strong>g>of</str<strong>on</strong>g> a subsequent case - c<strong>on</strong>trol study by Miller & Chen<br />

(2006). The <str<strong>on</strong>g>for</str<strong>on</strong>g>mer showed that a severely negative life event significantly increases the risk <str<strong>on</strong>g>of</str<strong>on</strong>g> an acute<br />

exacerbati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> asthma immediately afterwards, and again after a period <str<strong>on</strong>g>of</str<strong>on</strong>g> a few weeks. Chr<strong>on</strong>ic background<br />

stress both magnified the risk related to stressful life events and affected the timing <str<strong>on</strong>g>of</str<strong>on</strong>g> maximum risk. The<br />

latter added a possibly important validati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these results. It showed that children with asthma who<br />

experienced a severe life event in the c<strong>on</strong>text <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic stress had diminished expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> genes encoding<br />

glucocorticoid and β2-adrenergic receptors relative to children without comparable stress exposure.<br />

An attempt will then be made to integrate the above findings in the c<strong>on</strong>text <str<strong>on</strong>g>of</str<strong>on</strong>g> stress affecting the immune<br />

processes implicated in asthma, and within the framework <str<strong>on</strong>g>of</str<strong>on</strong>g> emoti<strong>on</strong>al/ physiological dysregulati<strong>on</strong>, possibly<br />

reflecting a comm<strong>on</strong> underlying genetic vulnerability to atopy and asthma.<br />

7B_17_P<br />

(poster secti<strong>on</strong> A2, poster board #113, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

MPM, A SYNTHETIC CCK ANTAGONIST, ANTAGONIZED THE EFFECTS OF<br />

STRESS ON HIPPOCAMPAL DENDRITIC REMODELING IN RATS<br />

Sattayasai Jintana 1 , Bo<strong>on</strong>prakob Yodchai 2 , Lattmann Eric 3<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmacology, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, 2 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Physical therapy, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Associated<br />

Medicine,Kh<strong>on</strong> Kaen University, Kh<strong>on</strong> Kaen, Thailand, 40002<br />

3The School <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmacy, Ast<strong>on</strong> University, Ast<strong>on</strong> Triangle, Birmingham B4 7ET, UK<br />

Stress may be defined as any type <str<strong>on</strong>g>of</str<strong>on</strong>g> threat, either real or perceived, that requires compensatory resp<strong>on</strong>ses <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

the maintenance <str<strong>on</strong>g>of</str<strong>on</strong>g> homeostasis and the resp<strong>on</strong>ses may be adaptive or maladaptive. The maladaptive<br />

resp<strong>on</strong>ses to a stressor could become a stress and a pathophysiological cascade may ensue, <str<strong>on</strong>g>for</str<strong>on</strong>g> example,<br />

depressi<strong>on</strong>, increased anxiety or post-traumatic stress disorder. The hippocampal <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> is <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> a plastic<br />

and vulnerable brain structure and highly susceptible to stress and glucocorticoids. Stress and chr<strong>on</strong>ic<br />

glucocorticoid administrati<strong>on</strong> have been shown to induce hippocampal damage, such as neur<strong>on</strong> death, gliosis<br />

and dendritic atrophy found in many animal models. In this study, MPM [N-(5-methyl-3-oxo-1,2-diphenyl-<br />

1H-pyrazol-4-yl)-N’-3-methoxyphenylurea], a synthetic CCK (cholecystokinin) antag<strong>on</strong>ist, was tested <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

antag<strong>on</strong>istic effect against stress. Male Spraque-Dawley rats were subjected to chr<strong>on</strong>ic restraint stress <str<strong>on</strong>g>for</str<strong>on</strong>g> 6<br />

h/day, <str<strong>on</strong>g>for</str<strong>on</strong>g> 28 days. Prior to the restraint sessi<strong>on</strong>s, rats were orally fed with either 5% DMSO or MPM<br />

(0.5 mg/kg). On day 1, 7, 14, 21 and 28, the animals were tested in elevated plus maze and <str<strong>on</strong>g>for</str<strong>on</strong>g>ced swim<br />

model. At the end <str<strong>on</strong>g>of</str<strong>on</strong>g> the treatment, rat’s brains were removed and processed <str<strong>on</strong>g>for</str<strong>on</strong>g> Golgi-Cox method and the<br />

dendritic trees <str<strong>on</strong>g>of</str<strong>on</strong>g> CA3 hippocampal pyramidal neur<strong>on</strong>s were observed under light microscope. The results<br />

showed that MPM could antag<strong>on</strong>ize all the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic restraint stress, including mood disorders and<br />

hippocampal dendritic atrophy. According to the receptor binding property, MPM might antag<strong>on</strong>ize stress<br />

effects through both types <str<strong>on</strong>g>of</str<strong>on</strong>g> receptors at hippocampus, pituitary and adrenal glands and break the LHPA<br />

axis in resp<strong>on</strong>se to stress.<br />

421


23-26 August 2007,<br />

Budapest, Hungary<br />

422<br />

7B_18_P<br />

(poster secti<strong>on</strong> A2, poster board #114, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

LIVING WITH HIV- IS IT STRESSFUL AS IT SOUNDS<br />

Batia Sheffy<br />

Caesarea, Israel<br />

HIV can produce psycosocial stress. HIV/AIDS has characters that are associated with stress, like secrets and<br />

stigma al<strong>on</strong>g with other variables <str<strong>on</strong>g>of</str<strong>on</strong>g> coping with chr<strong>on</strong>ic-terminal disease.<br />

Living with disease, can produce psychological stress, and since the relati<strong>on</strong>s between mind and body are<br />

known, awareness to coping mechanisms, and using certain coping styles can reduce stress and improve life<br />

satisfacti<strong>on</strong> and psychological well being.<br />

The study presented collected data from 100 people living with HIV/AIDS in Israel. The study examined<br />

correlati<strong>on</strong>s between ways <str<strong>on</strong>g>of</str<strong>on</strong>g> coping (emoti<strong>on</strong>al focused or problem solving) and quality <str<strong>on</strong>g>of</str<strong>on</strong>g> life and well<br />

being.<br />

The main assumpti<strong>on</strong> was that using problem solving coping style will lead to higher life satisfacti<strong>on</strong>, which<br />

are both rati<strong>on</strong>al variables and <strong>on</strong> the other hand, emoti<strong>on</strong>al focused coping style will lead to well being<br />

which is an emoti<strong>on</strong>al variable. Also were examined stress related variables and their associati<strong>on</strong> with the<br />

depended variables.<br />

The assumpti<strong>on</strong> was partly c<strong>on</strong>firmed and the correlati<strong>on</strong>s between the affective variables were not<br />

c<strong>on</strong>firmed. Other variables, related to psychological stress were also correlated with the depended variables.<br />

7B_19_P<br />

(poster secti<strong>on</strong> A2, poster board #115, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PSYCHOLOGICAL RISK FACTORS IN RHEUMATOID ARTHRITIS,<br />

PRELIMINARY STUDY<br />

Agnieszka Wlazło 1 , Jacek Kleszczyński 2 , Grażyna Dolińska 1 , Jerzy Leszek 2<br />

1Uniwersytet Wrocławski<br />

2Akademia Medyczna we Wrocławiu, e-mail: aga.ag@vp.pl<br />

Introducti<strong>on</strong>:<br />

Psychological factors – by intact <strong>on</strong> the <strong>on</strong>e hand <strong>on</strong> assessment <str<strong>on</strong>g>of</str<strong>on</strong>g> difficult situati<strong>on</strong>s and <strong>on</strong> the other <strong>on</strong><br />

stress coping - can promote pathophysiological reacti<strong>on</strong>s, which are associated with rheumatoid arthritis. Up<br />

to now, features <str<strong>on</strong>g>of</str<strong>on</strong>g> pers<strong>on</strong>ality and temperament, typical <str<strong>on</strong>g>for</str<strong>on</strong>g> rheumatoid arthritis patients, were not assess.<br />

Aim <str<strong>on</strong>g>of</str<strong>on</strong>g> the study:<br />

Appreciati<strong>on</strong> psychological risk factors characteristics <str<strong>on</strong>g>for</str<strong>on</strong>g> rheumatoid arthritis patients.<br />

Methods:<br />

Thirty patients (16 F, 14 M) in age 16-19 (mean age 46) with diagnosis <str<strong>on</strong>g>of</str<strong>on</strong>g> rheumatoid arthritis (according to<br />

ARA) were included to the study. C<strong>on</strong>trol group were health people with family load with autoimmunological<br />

disorders. The all patients in preliminary period <str<strong>on</strong>g>of</str<strong>on</strong>g> the disease treated with pharmacotherapy and<br />

rehabilitati<strong>on</strong> and people bel<strong>on</strong>g to c<strong>on</strong>trol group used following questi<strong>on</strong>naires:<br />

Life Orientati<strong>on</strong> Test (M. F. Scheier, C. S. Carver, M. W. Bridges)<br />

The Sense <str<strong>on</strong>g>of</str<strong>on</strong>g> Coherence Questi<strong>on</strong>naire (A. Ant<strong>on</strong>ovsky)<br />

Formal Characteristic Behavior: Temperament Questi<strong>on</strong>naire (B. Zawadzki, J. Strelau)<br />

Eysenck Pers<strong>on</strong>ality Questi<strong>on</strong>naire – Revised (S. B. G. Eysenck, H. J. Eysenck, P. Barett)


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Syndrome <str<strong>on</strong>g>of</str<strong>on</strong>g> Aggressi<strong>on</strong>: Psychological Questi<strong>on</strong>naire (Z. B. Gaś)<br />

Results:<br />

Firstly, we have found that level <str<strong>on</strong>g>of</str<strong>on</strong>g> optimism, sensivity, activity and endurance were statistical significantly<br />

lower in group <str<strong>on</strong>g>of</str<strong>on</strong>g> rheumatoid arthritis patients; sec<strong>on</strong>dly level <str<strong>on</strong>g>of</str<strong>on</strong>g> emoti<strong>on</strong>al reactivity and neurotism were<br />

statistical significantly higher.<br />

C<strong>on</strong>clusi<strong>on</strong>:<br />

These results seem to support hypothesis that psychological factors may play significant role in inducti<strong>on</strong> and<br />

progressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> rheumatoid arthritis. However, taking into c<strong>on</strong>siderati<strong>on</strong> multifactor etiopathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> this<br />

diseases, our results should be c<strong>on</strong>firm in future investigati<strong>on</strong>s.<br />

7B_20_P<br />

(poster secti<strong>on</strong> A2, poster board #116, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

POSTTRANSLATIONAL MODIFICATIONS AND SUBCELLULAR<br />

REDISTRIBUTION OF GLUCOCORTICOID RECEPTOR<br />

IN RESPONSE TO ACUTE STRESS IN WISTAR RAT BRAIN<br />

Miroslav Adzic 1 , Jelena Djordjevic 1 , Ana Nici<str<strong>on</strong>g>for</str<strong>on</strong>g>ovic 1 , Marija Radojcic 1 , Marija Krstic-Dem<strong>on</strong>acos 2<br />

1Dept.Mol.Biol.Endocrinol., VINCA Inst.Nucl.Sci., Belgrade, Serbia<br />

2Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Life Sciences, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Manchester, Manchester, UK<br />

e-mail: marija@vin.bg.ac.yu<br />

Prefr<strong>on</strong>tal cortex (PFC) and hippocampus (HIP) are main limbic structures involved in inhibitory feedback to<br />

HPA axis during stress. Inhibitory feed-back is provided by glucocorticoid receptor (GR), which acts as<br />

suppressor <str<strong>on</strong>g>of</str<strong>on</strong>g> excessive stress resp<strong>on</strong>se. The level <str<strong>on</strong>g>of</str<strong>on</strong>g> blood plasma corticoster<strong>on</strong>e is elevated in resp<strong>on</strong>se to<br />

stress, leading to GR activati<strong>on</strong>. Furthermore, the GR functi<strong>on</strong>s are c<strong>on</strong>trolled by inhibitory phosporylati<strong>on</strong><br />

at ser246 (S246) by JNK and stimulatory phosphorylati<strong>on</strong>s at ser224 and ser232 (S224, S232) by CDK.<br />

Molecular mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> GR posttranslati<strong>on</strong>al modificati<strong>on</strong>s, as well as, subcellular locati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> its<br />

phosphorylated is<str<strong>on</strong>g>of</str<strong>on</strong>g>orms in resp<strong>on</strong>se to acute stress in vivo have not been precisely defined yet. We studied<br />

these mechanisms in PFC and HIP <str<strong>on</strong>g>of</str<strong>on</strong>g> male Wistar rats exposed to acute stress by immobilizati<strong>on</strong>. The<br />

decreased level <str<strong>on</strong>g>of</str<strong>on</strong>g> total GR in the cytoplasm and increased level in the nucleus was accompanied by marked<br />

decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> its phosphorylated is<str<strong>on</strong>g>of</str<strong>on</strong>g>orm S246pGR in both compartments. The levels <str<strong>on</strong>g>of</str<strong>on</strong>g> active, phosphorylated<br />

JNK is<str<strong>on</strong>g>of</str<strong>on</strong>g>orms were elevated in the cytoplasm and significantly decreased in the nucleus. The GR activati<strong>on</strong><br />

by acute stress was indicated by the elevated phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> GR at S232 in the nucleus <str<strong>on</strong>g>of</str<strong>on</strong>g> HIP. Overall,<br />

GR cytoplasmic-nucleus shuttling in PFC and HIP is accompanied by changed ratios <str<strong>on</strong>g>of</str<strong>on</strong>g> inhibitory and<br />

stimulatory phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> GR, pointing out to their significance <str<strong>on</strong>g>for</str<strong>on</strong>g> GR subcellular locati<strong>on</strong> and<br />

resp<strong>on</strong>se to acute stress.<br />

423


23-26 August 2007,<br />

Budapest, Hungary<br />

7B_21_P<br />

(poster secti<strong>on</strong> A2, poster board #117, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

GENETIC AND NEUROENDOCRINE MECHANISMS OF STRESS SENSITIVE<br />

HYPERTENSION IN ISIAH RAT STRAIN<br />

O. E. Redina, S. E.Smolenskaya, A. L. Markel<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Cytology and Genetics, Siberian Branch <str<strong>on</strong>g>of</str<strong>on</strong>g> Russian Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences,<br />

10 Lavrentieva ave., Novosibirsk, Russia 630090<br />

The genetic and neuroendocrine mechanisms underlying hypertensi<strong>on</strong> development in the ISIAH rats with<br />

inherited stress-induced arterial hypertensi<strong>on</strong> were studied. This strain was selected <str<strong>on</strong>g>for</str<strong>on</strong>g> increased resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

systolic arterial blood pressure (ABP) to a mild emoti<strong>on</strong>al stress caused by 0.5 h restricti<strong>on</strong> in a cylindrical<br />

wire-mesh cage. As a result <str<strong>on</strong>g>of</str<strong>on</strong>g> the selecti<strong>on</strong>, the ISIAH rats acquired the number <str<strong>on</strong>g>of</str<strong>on</strong>g> characteristic features<br />

c<strong>on</strong>cerned the hypertensive status. The ISIAH rats were characterized by: the increased mRNA producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

CRH and POMC genes in hypothalamus and pituitary, corresp<strong>on</strong>dingly; increased resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> the plasma<br />

corticoster<strong>on</strong>e to some stressful stimuli; increase in norepinephrine and dopamine level in hypothalamus and<br />

locus coeruleus. Epinephrine c<strong>on</strong>tent in the adrenals as well as norepinephrine plasma c<strong>on</strong>centrati<strong>on</strong> was<br />

more than two fold higher in ISIAH than in normotensive WAG rats. Two F 2 male populati<strong>on</strong>s (WAG x<br />

ISIAH) were used in QTL analysis to identify the genetic loci <str<strong>on</strong>g>for</str<strong>on</strong>g> ABP (basal and under stress) and <str<strong>on</strong>g>for</str<strong>on</strong>g> some<br />

related traits. The highest LOD scores were found in 3-4-m<strong>on</strong>th old populati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g>: body weight <strong>on</strong> Chr.1<br />

(D1Rat76 marker, LOD score 3.23), and relative kidney weight <strong>on</strong> Chr.1 (D1Rat117 marker, LOD score<br />

3.41) and Chr.12 (D12Rat93, LOD score 3.66); <str<strong>on</strong>g>for</str<strong>on</strong>g> the rat locomoti<strong>on</strong> in the periphery <str<strong>on</strong>g>of</str<strong>on</strong>g> the open field <strong>on</strong><br />

Chr.2 (D2Rat157- D2Rat88 markers, LOD score 4.83) and Chr.16 (D16rat32 marker, LOD score 3.71). In 6-<br />

m<strong>on</strong>th old rats the follow QTL were found: <str<strong>on</strong>g>for</str<strong>on</strong>g> basal and stress induced ABP <strong>on</strong> Chr.1 in the regi<strong>on</strong><br />

D1Rat168-D1Rat76 (LOD score 3.42), <str<strong>on</strong>g>for</str<strong>on</strong>g> adrenals weight <strong>on</strong> Chr.6 and <str<strong>on</strong>g>for</str<strong>on</strong>g> dopamine in medulla <strong>on</strong> Chr. 8.<br />

7B_22_P<br />

(poster secti<strong>on</strong> A2, poster board #118, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE EFFECTS OF PARENT TRAINING PROGRAM COMBINED COGNITIVE<br />

BEHAVIORAL SOCIAL SKILL TRAINING ON ADHD SYMPTOMS AND SOCIAL<br />

BEHAVIORS OF CHILDREN WITH ADHD<br />

Hyun Myoungho<br />

Chung-ang University, Psychology, 156-756, Heukseuk D<strong>on</strong>g, D<strong>on</strong>gjak Gu, Seoul, South Korea<br />

e-mail: hyunmh@cau.ac.kr<br />

We c<strong>on</strong>ducted the comparative study <str<strong>on</strong>g>of</str<strong>on</strong>g> the treatment effects between parent training program (PTP)<br />

combined cognitive behavioral social skill training(SST) and PTP n<strong>on</strong>-combined SST <strong>on</strong> self-c<strong>on</strong>cept and<br />

behaviors <str<strong>on</strong>g>of</str<strong>on</strong>g> children with ADHD.<br />

21 children with ADHD (8.84±1.06 years old, 20 boys and 1 girls) were recruited <str<strong>on</strong>g>for</str<strong>on</strong>g> the study. Subjects were<br />

randomly assigned to two groups: combined treatment group (n=8; PTP+SST+pharmacotherapy with<br />

stimulants) or n<strong>on</strong>-combined group(n=13 ; SST+ pharmacotherapy with stimulants).<br />

C<strong>on</strong>ner's Parent Rating Scale (CPRS), C<strong>on</strong>ner's Teacher Rating Scale(CTRS), Social Skill Rating System, and<br />

Piers-Harris‘ self-c<strong>on</strong>cept scale <str<strong>on</strong>g>for</str<strong>on</strong>g> children were used <str<strong>on</strong>g>for</str<strong>on</strong>g> objectively measuring the treatment effects<br />

between groups.<br />

424


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

The data were analyzed by 2 (n<strong>on</strong>combined treatment group/combined treatment group: between subjject<br />

variable)×2(pre/post treatment programs : within subject variable) repeated measures ANOVA.<br />

The results showed that the combined treatment programs significantly reduced CPRS scores and<br />

significantly improved self-insistence, sense <str<strong>on</strong>g>of</str<strong>on</strong>g> resp<strong>on</strong>sibility and cooperati<strong>on</strong> subscale scores in social skill<br />

rating system.<br />

This study supported that parenting skill enhancing combined treatment programs improve not <strong>on</strong>ly ADHD<br />

symptoms but also social behaviors and self-c<strong>on</strong>cept in children with ADHD.<br />

7B_23_P<br />

(poster secti<strong>on</strong> A2, poster board #119, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PSYCHOSOCIAL AND PHYSICAL DIMENSIONS OF STRESS RELATED TO A<br />

CHRONIC DISEASE IN WORKING AND NOT-WORKING PATIENTS<br />

Maria K<strong>on</strong>arska, Joanna Bugajska, Dorota Zołnierczyk-Zreda, Anna Jędryka- Góral<br />

Central Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Labour Protecti<strong>on</strong>, 00-701 Warsaw, Poland<br />

Background: This survey study was c<strong>on</strong>ducted by the Polish group as a task “Socio - ec<strong>on</strong>omic studies”<br />

within the EC 6 FP project „Advanced Interactive Materials by Design - AIMs” coordinated by the<br />

Dortmund University (C<strong>on</strong>tract NoNMP3-CT-2004-500160). The project overall aims to optimise<br />

producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> m<strong>on</strong>ocl<strong>on</strong>al antibodies used in the treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> a range <str<strong>on</strong>g>of</str<strong>on</strong>g> diseases causing significant social<br />

and ec<strong>on</strong>omic burden to the populati<strong>on</strong>.<br />

The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to evaluate the impact <str<strong>on</strong>g>of</str<strong>on</strong>g> different aspects <str<strong>on</strong>g>of</str<strong>on</strong>g> a chr<strong>on</strong>ic disease like rheumatoid<br />

arthritis (RA) into patient’s assessment <str<strong>on</strong>g>of</str<strong>on</strong>g> quality <str<strong>on</strong>g>of</str<strong>on</strong>g> life and ability to work.<br />

Methods: The study covered 300 c<strong>on</strong>secutive outpatients, all fulfilling the ARC (1987) criteria <str<strong>on</strong>g>for</str<strong>on</strong>g> the RA<br />

diagnosis, aged 18 - 65 ys, treated traditi<strong>on</strong>ally i.e. without biological drugs, and 200 healthy c<strong>on</strong>trols<br />

matching the group <str<strong>on</strong>g>of</str<strong>on</strong>g> patients in terms <str<strong>on</strong>g>of</str<strong>on</strong>g> age, gender and employment status, requited from people who<br />

never required a regular l<strong>on</strong>g-term treatment.<br />

The survey questi<strong>on</strong>naire included questi<strong>on</strong>s addressing key aspects <str<strong>on</strong>g>of</str<strong>on</strong>g> stress related to a chr<strong>on</strong>ic disease in<br />

working and not-working patients: quality <str<strong>on</strong>g>of</str<strong>on</strong>g> life (based <strong>on</strong> SF-36 questi<strong>on</strong>naire), work ability (based <strong>on</strong> the<br />

Work Ability Index questi<strong>on</strong>naire) and costs borne by the patient (based <strong>on</strong> a set <str<strong>on</strong>g>of</str<strong>on</strong>g> questi<strong>on</strong>s developed by<br />

the CIOP-PIB team <str<strong>on</strong>g>for</str<strong>on</strong>g> the purpose <str<strong>on</strong>g>of</str<strong>on</strong>g> this study).<br />

Results: The discrete factor analysis showed that disease durati<strong>on</strong>, radiological stage and clinical picture <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

RA correlated at significant level with patients assessment <str<strong>on</strong>g>of</str<strong>on</strong>g> ability to work and quality <str<strong>on</strong>g>of</str<strong>on</strong>g> life, the latter<br />

especially in the physical (bodily pain, role limitati<strong>on</strong> due to physical health problems) and mental (vitality,<br />

social functi<strong>on</strong>ing, role limitati<strong>on</strong> due to emoti<strong>on</strong>al problems,) aspects <str<strong>on</strong>g>of</str<strong>on</strong>g> health. Differences between<br />

working and not working pers<strong>on</strong>s will be analysed in details.<br />

425


23-26 August 2007,<br />

Budapest, Hungary<br />

7B_24_P<br />

(poster secti<strong>on</strong> A2, poster board #120, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFICACY OF A NURSE-SUPPORTED COMMUNICATION SKILLS<br />

INTERVENTION ON PATIENT PSYCHOSOCIAL DISTRESS AFTER CANCER<br />

DIAGNOSIS<br />

Fukui Sakiko<br />

Chiba University, School <str<strong>on</strong>g>of</str<strong>on</strong>g> Nursing, 260-8672, Chou-ku Inohana 1-8-1, Chiba, Japan<br />

e-mail: sfukui@faculty.chiba-u.jp<br />

Background: There had been no evidence <str<strong>on</strong>g>for</str<strong>on</strong>g> the efficacy <str<strong>on</strong>g>of</str<strong>on</strong>g> communicati<strong>on</strong> skills interventi<strong>on</strong> <strong>on</strong> improving<br />

psychosocial distress am<strong>on</strong>g patients after cancer diagnosis. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to determine the effect<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> a nurse-supported communicati<strong>on</strong> skills interventi<strong>on</strong> program in reducing psychological distress and<br />

improving QOL in this populati<strong>on</strong> in a randomized c<strong>on</strong>trolled trial.<br />

Methods: The subjects who were in<str<strong>on</strong>g>for</str<strong>on</strong>g>med cancer diagnosis at a cancer screening center were randomly<br />

assigned either an interventi<strong>on</strong> group or a c<strong>on</strong>trol group. We c<strong>on</strong>ducted a nurse-supported communicati<strong>on</strong><br />

skills interventi<strong>on</strong> program <str<strong>on</strong>g>for</str<strong>on</strong>g> patients <str<strong>on</strong>g>of</str<strong>on</strong>g> an interventi<strong>on</strong> group <str<strong>on</strong>g>for</str<strong>on</strong>g> 1 m<strong>on</strong>th. Nurses, who were trained 6-<br />

step communicati<strong>on</strong> skills “SPIKES” to support cancer patients <str<strong>on</strong>g>for</str<strong>on</strong>g> 2 days, c<strong>on</strong>ducted an interventi<strong>on</strong><br />

program <str<strong>on</strong>g>for</str<strong>on</strong>g> the interventi<strong>on</strong>-group subjects <str<strong>on</strong>g>for</str<strong>on</strong>g> three times according to a manual. Subjects <str<strong>on</strong>g>of</str<strong>on</strong>g> both groups<br />

were assessed <str<strong>on</strong>g>for</str<strong>on</strong>g> psychological distress and QOL by administering the Hospital Anxiety and Depressi<strong>on</strong><br />

Scale (HADS) and 4-itemed VAS scale at the baseline, at 1 m<strong>on</strong>th, and at 3 m<strong>on</strong>ths.<br />

Results: Sixty four patients participated and were randomized in the study. The interventi<strong>on</strong> group (n=31)<br />

had significantly lower scores than the c<strong>on</strong>trols (n=33) <str<strong>on</strong>g>for</str<strong>on</strong>g> total psychological distress and significantly higher<br />

scores <str<strong>on</strong>g>for</str<strong>on</strong>g> total QOL at the end <str<strong>on</strong>g>of</str<strong>on</strong>g> the 1-m<strong>on</strong>th interventi<strong>on</strong>. These improvements were sustained over 3<br />

m<strong>on</strong>ths <str<strong>on</strong>g>of</str<strong>on</strong>g> follow-up.<br />

C<strong>on</strong>clusi<strong>on</strong>s: The results <str<strong>on</strong>g>of</str<strong>on</strong>g> this study suggest that a nurse-supported communicati<strong>on</strong> skills interventi<strong>on</strong><br />

produces significant l<strong>on</strong>g-term improvement in the psychosocial distress <str<strong>on</strong>g>of</str<strong>on</strong>g> patients after cancer diagnoses in<br />

Japan.<br />

426


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7C. PSYCHO-NEURO-IMMUNOLOGY<br />

(JUDIT SZELÉNYI)<br />

7C_01_S<br />

ALTERATIONS OF THE CATECHOLAMINE-CYTOKINE BALANCE<br />

IN DEPRESSION<br />

J. Szelényi, E. S. Vizi<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Experimental Medicine, Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Neuroimmunology, 43. Szig<strong>on</strong>y street H-1083<br />

Budapest, Hungary<br />

Cytokines are involved both in various immune reacti<strong>on</strong>s and in c<strong>on</strong>trolling certain events in the central<br />

nervous system. In our earlier studies it was shown that m<strong>on</strong>oamine neurotransmitters, released in stress<br />

situati<strong>on</strong>s, represent a t<strong>on</strong>ic sympathetic c<strong>on</strong>trol <strong>on</strong> cytokine producti<strong>on</strong> and <strong>on</strong> the balance <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

proinflammatory/antiinflammatory cytokines. Basic and clinical studies have provided evidence that the<br />

biophase level <str<strong>on</strong>g>of</str<strong>on</strong>g> m<strong>on</strong>oamines, determined by the balance <str<strong>on</strong>g>of</str<strong>on</strong>g> their release and uptake, is involved in the<br />

pathophysiology and treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> depressi<strong>on</strong>, while inflammatory mediators might also have role in its<br />

etiology. In this work we studied the role <str<strong>on</strong>g>of</str<strong>on</strong>g> changes in norepinephrine (NE) level <strong>on</strong> the LPS evoked TNF-α<br />

and IL-10 resp<strong>on</strong>se both in the plasma and in the hippocampus <str<strong>on</strong>g>of</str<strong>on</strong>g> mice. We dem<strong>on</strong>strated that the LPS<br />

induced TNF-α resp<strong>on</strong>se is in direct correlati<strong>on</strong> with the biophase level <str<strong>on</strong>g>of</str<strong>on</strong>g> NE as it is significantly higher<br />

when the release <str<strong>on</strong>g>of</str<strong>on</strong>g> NE <str<strong>on</strong>g>of</str<strong>on</strong>g> vesicular origin was completely inhibited in an animal model <str<strong>on</strong>g>of</str<strong>on</strong>g> depressi<strong>on</strong><br />

(reserpine treatment) and it is significantly lower in case <str<strong>on</strong>g>of</str<strong>on</strong>g> increasing biophase level <str<strong>on</strong>g>of</str<strong>on</strong>g> NE by genetical<br />

(NETKO) or chemical (desipramine) disrupti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> NE reuptake. IL-10 was changed inversely to the TNF-α<br />

level <strong>on</strong>ly in the desipramine treated animals. Our results showed that depressi<strong>on</strong> is related both to changes in<br />

peripheral and in hippocampal inflammatory cytokine producti<strong>on</strong> and to m<strong>on</strong>oamine neurotransmitter levels.<br />

Since several antiinflammatory drugs have also antidepressant effect we hypothesized that inhibitors <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

m<strong>on</strong>oamine uptake system might have multiple targets and are also able to modulate the LPS-induced<br />

inflammatory resp<strong>on</strong>ses, which might c<strong>on</strong>tribute to their antidepressant effect.<br />

7C_02_S<br />

CYTOKINE ACTIONS IN THE BRAIN MEDIATE THE INCREASED PREVALENCE<br />

OF DEPRESSION IN DISEASES WITH AN INFLAMMATORY COMPONENT<br />

Robert Dantzer<br />

Integrative Immunology & Behavior Program, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Illinois at Urbana-Champaign, USA<br />

Physically ill patients experience a high prevalence <str<strong>on</strong>g>of</str<strong>on</strong>g> affect disorders that are exacerbated by relatively minor<br />

infecti<strong>on</strong>s. Activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the peripheral innate immune system induces signs <str<strong>on</strong>g>of</str<strong>on</strong>g> sickness that culminate in<br />

depressive-like behavior in adult mice. This resp<strong>on</strong>se is exaggerated in aged mice whose innate immune<br />

system is chr<strong>on</strong>ically activated. Peripheral inflammati<strong>on</strong> is relayed to the brain by both the sensory nerves that<br />

innervate the site <str<strong>on</strong>g>of</str<strong>on</strong>g> inflammati<strong>on</strong> and the overflow <str<strong>on</strong>g>of</str<strong>on</strong>g> inflammatory mediators that enters the general<br />

circulati<strong>on</strong>. In resp<strong>on</strong>se to these stimuli, brain macrophage-like cells produces cytokines that directly or<br />

indirectly affect neur<strong>on</strong>al functi<strong>on</strong>. Activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> indoleamine 2,3 dioxygenase (IDO), a key enzyme in the<br />

metabolism <str<strong>on</strong>g>of</str<strong>on</strong>g> tryptophan, is an important mechanism <str<strong>on</strong>g>for</str<strong>on</strong>g> the depressive-like effects <str<strong>on</strong>g>of</str<strong>on</strong>g> immune activati<strong>on</strong>.<br />

Enhanced activity <str<strong>on</strong>g>of</str<strong>on</strong>g> this enzyme in resp<strong>on</strong>se to tumor necrosis factor-alpha and/or interfer<strong>on</strong>-gamma<br />

induces the producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> neurotoxic kynurenine metabolites and decreases the bioavailability <str<strong>on</strong>g>of</str<strong>on</strong>g> tryptophan,<br />

427


23-26 August 2007,<br />

Budapest, Hungary<br />

which ultimately impacts <strong>on</strong> serot<strong>on</strong>inergic neurotransmissi<strong>on</strong>. The development <str<strong>on</strong>g>of</str<strong>on</strong>g> depressive-like behavior<br />

in mouse in resp<strong>on</strong>se to acute or chr<strong>on</strong>ic activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the peripheral innate immune system is temporarily<br />

correlated with IDO activati<strong>on</strong> in the brain and at the periphery. Furthermore pretreatment with 1-methyltryptophan,<br />

a competitive antag<strong>on</strong>ist <str<strong>on</strong>g>of</str<strong>on</strong>g> IDO, abrogates depressive-like behavior but not sickness behavior in<br />

mice during the course <str<strong>on</strong>g>of</str<strong>on</strong>g> inflammati<strong>on</strong>. These preclinical findings emphasize the role <str<strong>on</strong>g>of</str<strong>on</strong>g> cytokines and their<br />

metabolic effects in the pathophysiology <str<strong>on</strong>g>of</str<strong>on</strong>g> inflammati<strong>on</strong>-associated depressi<strong>on</strong>.<br />

428<br />

7C_03_S<br />

LONG PHOTOPERIOD-INDUCED GLUCOCORTICOID RESISTANCE AND<br />

DECREASED NEGATIVE SELECTION IN DOUBLE-POSITIVE THYMOCYTES OF<br />

FEMALE BUT NOT MALE MICE. MECHANISMS AND POSSIBLE IMPLICATION IN<br />

THE GEOGRAPHICAL DISTRIBUTION OF AUTOIMMUNE DISEASES<br />

Georges Maestr<strong>on</strong>i<br />

Istituto Cant<strong>on</strong>ale di Patologia, Experimantal Pathology, 6601 Via in Selva 24, Locarno, Switzerland<br />

e-mail: georges.maestr<strong>on</strong>i@ti.ch<br />

The mammalian immune resp<strong>on</strong>se is circadian and is regulated through daily alterati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> darkness and<br />

recently defined specific pathway <str<strong>on</strong>g>of</str<strong>on</strong>g> n<strong>on</strong>-visual light. The endogenous pace maker that drives this circadian<br />

cycle is located in the suprachiasmatic nucleus (SCN) <str<strong>on</strong>g>of</str<strong>on</strong>g> the hypothalamus. Light reaches the SCN through<br />

n<strong>on</strong>-rod, n<strong>on</strong>-c<strong>on</strong>e intrinsically photosensitive retinal gangli<strong>on</strong> cells. The nervous pathway c<strong>on</strong>necting the<br />

n<strong>on</strong>-visual light detecti<strong>on</strong> in the SCN <str<strong>on</strong>g>of</str<strong>on</strong>g> the hypothalamus leads to a release <str<strong>on</strong>g>of</str<strong>on</strong>g> neurohorm<strong>on</strong>es from the<br />

pituitary, pineal, adrenal glands, and the g<strong>on</strong>ads that finally reach the lymphoid organs. We found that<br />

exposure <str<strong>on</strong>g>of</str<strong>on</strong>g> adult female but not male mice to l<strong>on</strong>g photoperiod (22 hours light: 2 hours dark, LP) <str<strong>on</strong>g>for</str<strong>on</strong>g> 8 days<br />

decreased the glucocorticoid sensitivity <str<strong>on</strong>g>of</str<strong>on</strong>g> the thymus gland. Dexamethas<strong>on</strong>e –induced apoptosis was<br />

decreased in double positive thymocytes and this effect was apparent <strong>on</strong>ly in vivo but not in vitro. This effect<br />

was not observed in prebubertal female mice and in adult g<strong>on</strong>adectomized mice as well as in beta-2<br />

adrenergic receptor gene deficient mice. We exposed to the LP DO11.10 mice transgenic <str<strong>on</strong>g>for</str<strong>on</strong>g> a T cell<br />

receptor recognizing an ovalbumin peptide. The peptide was then injected intraperit<strong>on</strong>eally to induce<br />

intrathymic negative selecti<strong>on</strong>. The LP-exposed mice showed a significantly reduced negative selecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

double positive thymocytes. Intrathymic negative selecti<strong>on</strong> is mediated by endogenous glucocorticoids and<br />

generates tolerance to self –antigens. As the seas<strong>on</strong>al variati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the photoperiod increases with the latitude,<br />

we might have disclosed a basic mechanism explaining the gender and geographical distributi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

autoimmune disorders which are more frequent in women and show a north-south gradient <str<strong>on</strong>g>of</str<strong>on</strong>g> incidence.<br />

7C_04_S<br />

MATERNAL STRESS AND THE PRENATAL PROGRAMMING OF<br />

INFANT IMMUNITY<br />

Christopher L. Coe, Gabriele R. Lubach<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Wisc<strong>on</strong>sin, 22 N. Charter Street, Madis<strong>on</strong>, WI, USA<br />

e-mail: ccoe@wisc.edu<br />

Background. While stress can influence immune resp<strong>on</strong>ses at any point in the life span, our research has<br />

shown that the effects are larger and more lasting in the young and aged host. In particular, during fetal and


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

infant development, the maturati<strong>on</strong>al trajectory <str<strong>on</strong>g>of</str<strong>on</strong>g> immunity can be permanently altered. This presentati<strong>on</strong><br />

will focus <strong>on</strong> changes seen after manipulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the in utero envir<strong>on</strong>ment and the role <str<strong>on</strong>g>of</str<strong>on</strong>g> placental and<br />

neuroendocrine processes in mediating the immune alterati<strong>on</strong>s. Methods. Pregnancy c<strong>on</strong>diti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> rhesus<br />

m<strong>on</strong>keys were manipulated in several ways, including by psychological disturbance <str<strong>on</strong>g>of</str<strong>on</strong>g> the gravid female,<br />

administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> dexamethas<strong>on</strong>e acutely, dietary treatments, or viral infecti<strong>on</strong>. The impact <strong>on</strong> the infants’<br />

immune, endocrine, brain, and behavioral development was assessed.<br />

Results. Maternal stress, or antenatal administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> corticosteroids, significantly affected immune<br />

resp<strong>on</strong>ses at birth and c<strong>on</strong>tinued to have an effect <strong>on</strong> lymphocyte proliferati<strong>on</strong> and cytokine resp<strong>on</strong>ses up to<br />

2 years <str<strong>on</strong>g>of</str<strong>on</strong>g> age. These immune changes were associated with changes in the gut microbiota (including reduced<br />

Lactobacilli and Bifidobacter), which increased the risk <str<strong>on</strong>g>for</str<strong>on</strong>g> infecti<strong>on</strong> with enteric pathogens. A novel pathway<br />

mediating the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> maternal stress <strong>on</strong> the fetus was also identified: a reducti<strong>on</strong> in the placental transfer <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

ir<strong>on</strong>. Lower ir<strong>on</strong> stores at birth increased the risk <str<strong>on</strong>g>for</str<strong>on</strong>g> an ir<strong>on</strong> deficiency anemia (IDA) in the growing infants.<br />

The anemia emerging at 4-8 m<strong>on</strong>ths <str<strong>on</strong>g>of</str<strong>on</strong>g> age provided a sec<strong>on</strong>d postnatal hit to the immune system. In<br />

additi<strong>on</strong>, infant m<strong>on</strong>keys from stressed pregnancies evinced behavioral and neural changes, including<br />

immature neuromotor reflexes at birth, greater emoti<strong>on</strong>ality during the first year <str<strong>on</strong>g>of</str<strong>on</strong>g> life, and a smaller<br />

hippocampus as juveniles. Smaller hippocampal size was associated with less neurogenesis, and a more<br />

reactive hypothalamic-pituitary-adrenal axis. C<strong>on</strong>clusi<strong>on</strong>s. Stressful and challenging events during fetal life<br />

can significantly impact the development <str<strong>on</strong>g>of</str<strong>on</strong>g> immunity at a vulnerable point in <strong>on</strong>togeny and change the<br />

regulatory set points <str<strong>on</strong>g>for</str<strong>on</strong>g> several physiological systems postnatally.<br />

7C_06_S<br />

(poster secti<strong>on</strong> A2, poster board #121, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STRESS TRIGGERED NEURONAL PLASTICITY IN SPLEEN IN INTERACTION<br />

WITH IMMUNOCYTES<br />

C. Liezmann 1 , M. Daniltchenko 1 , S. Pavlovic Masnikosa 1 , B. F. Klapp 1 , E. M. J. Peters 1<br />

1 University-Medicine Charité, Charité Center 12 <str<strong>on</strong>g>for</str<strong>on</strong>g> Internal Medicine and Dermatology, Psych<strong>on</strong>euroimmunology,<br />

Campus Virchow, Berlin, Germany; e-mail: eva.peters@charite.de<br />

Stress is c<strong>on</strong>sidered as a factor which induces or aggravates inflammatory skin diseases such as atopic<br />

dermatitis. An influence <str<strong>on</strong>g>of</str<strong>on</strong>g> stress <strong>on</strong> the interacti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> peripheral nerve fibers with cells <str<strong>on</strong>g>of</str<strong>on</strong>g> the cutaneous<br />

immune system (mast cells, dendritic cells) with following modulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> an inflammati<strong>on</strong> reacti<strong>on</strong> could be<br />

proven recently. In this c<strong>on</strong>text, Substance P (SP) - a sensory neuropeptide - was revealed as an important<br />

stress mediator with its own stress axis in the skin. Here we postulate stress-dependent communicati<strong>on</strong><br />

between nerve fibers and immune-competent cells with effect <strong>on</strong> the course <str<strong>on</strong>g>of</str<strong>on</strong>g> inflammatory skin diseases in<br />

the spleen. To address this questi<strong>on</strong>, we employed a combined mouse model <str<strong>on</strong>g>of</str<strong>on</strong>g> experimental allergic<br />

dermatitis (AD) and stress. AD was induced in C57BL/6 mice by double sensitizati<strong>on</strong> (i.p) and an<br />

intradermal challenge using chicken egg ovalbumin. Animals were additi<strong>on</strong>ally exposed to s<strong>on</strong>ic stress <str<strong>on</strong>g>for</str<strong>on</strong>g> 24h<br />

prior to challenge. In this model stress leads to a relative hyperinnervati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the immune-competent areas <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the spleen. At the same time, an increased number <str<strong>on</strong>g>of</str<strong>on</strong>g> antigen-presenting cells (APC) can be observed in these<br />

arreas and c<strong>on</strong>tacts between nerve fibers and APC were found. Further analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> quality and functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

neuro-immune interacti<strong>on</strong> will reveal the role <str<strong>on</strong>g>of</str<strong>on</strong>g> the observed stress-induced alterati<strong>on</strong>s in the spleen in<br />

atopic disease .<br />

429


23-26 August 2007,<br />

Budapest, Hungary<br />

7C_01_P<br />

(poster secti<strong>on</strong> A2, poster board #122, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STRESS POTENTIATES MIGRATION AND MATURATION OF SKIN DENDRITIC<br />

CELLS IN ALLERGIC DERMATITIS<br />

S. Pavlovic Masnikosa 1 , M. Daniltchenko 1 , S. Blois 1 , B. F. Klapp 1 , E. M. J. Peters 1<br />

1 University-Medicine Charité, Charité Center 12 <str<strong>on</strong>g>for</str<strong>on</strong>g> Internal Medicine and Dermatology, Psych<strong>on</strong>euroimmunology,<br />

Campus Virchow, Berlin, Germany, e-mail: eva.peters@charite.de<br />

Altered TH1/TH2 immune resp<strong>on</strong>ses are held resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> atopic disease. Stress appears to be a potent<br />

immunmodulator as well as aggravator <str<strong>on</strong>g>of</str<strong>on</strong>g> atopic disease. To examine stress effects <strong>on</strong> immunity in atopy, we<br />

employed a combined mouse model <str<strong>on</strong>g>of</str<strong>on</strong>g> experimental allergic dermatitis (AD) and sound stress. In a previous<br />

study we had shown that stress applied prior to challenge induced neur<strong>on</strong>al plasticity, increased neurogenic<br />

inflammati<strong>on</strong>. We also observed an altered cytokine balance indicating altered antigen processing. Here we<br />

show, that stress prior to sensitizati<strong>on</strong> alters dendritic cell (DC) functi<strong>on</strong>. In stressed animals the number <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

MHCII + cells in epidermis and dermis is reduced 48h after challenge. DCs, mainly Langerhans cells, migrate<br />

more frequently from epidermal sheets cultured from treated-skin biopsies obtained immediately after<br />

challenge when AD animals were exposed to stress. The like was observed after treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> AD epidermal<br />

sheets with the stress mediator substance P (SP), but not with calcit<strong>on</strong>in gene related protein. Moreover, the<br />

effect <str<strong>on</strong>g>of</str<strong>on</strong>g> stress was abolished when animals were treated with NK1 antag<strong>on</strong>ist prior and after stress<br />

applicati<strong>on</strong>. Using flow cytometry, we found that these cells migrate to the draining lymph nodes, where DCs<br />

(CD11c+) from stressed AD animals show significant up-regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> co-stimulatory molecules CD80 and<br />

CD86. This correlated with an enhanced expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> LFA-1 and VLA-4, adhesi<strong>on</strong> molecules implicated in<br />

facilitati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> TH1 and TH2 resp<strong>on</strong>ses, respectively. Taken together, we show that stress activates DC<br />

migrati<strong>on</strong> and maturati<strong>on</strong> in atopic dermatitis-like allergic dermatitis in a SP dependent fashi<strong>on</strong>. Stress may<br />

there<str<strong>on</strong>g>for</str<strong>on</strong>g>e be involved in the sensitisati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> atopic individuals to an allergen and thereby determine the course<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the disease.<br />

7C_04_P<br />

(poster secti<strong>on</strong> A2, poster board #123, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SALIVARY CORTISOL AND LONG TERMED STRESS: TWO YEARS FOLLOW UP ON<br />

LONG TERMED STRESSED EMPLOYEES<br />

Bo Netterstrøm, Aase Marie Hansen<br />

Clinic <str<strong>on</strong>g>of</str<strong>on</strong>g> Occupati<strong>on</strong>al Medicine, Hillerød Hospital, Denmark<br />

Danish Nati<strong>on</strong>al Center <str<strong>on</strong>g>for</str<strong>on</strong>g> Work Envir<strong>on</strong>ment Reasearch<br />

Aim: To explore in the development <str<strong>on</strong>g>of</str<strong>on</strong>g> awakening cortisol reactivity during recovery from l<strong>on</strong>g termed stress.<br />

Methods: Seventy employed pers<strong>on</strong>s aged 25 to 61 years were referred to a stress management programme at<br />

the Clinic <str<strong>on</strong>g>of</str<strong>on</strong>g> Occupati<strong>on</strong>al Medicine in 2003 and 2004.<br />

Questi<strong>on</strong>naires regarding stress symptoms, SF36 and depressi<strong>on</strong> (MDI-10, WHO scale) were filled out at<br />

baseline, after 4 m<strong>on</strong>ths and <strong>on</strong>e and two years. Salivary cortisol was collected at awakening and ½ hour later.<br />

TSH, HbA1C, fibrinogen and serum lipids were measured at baseline and 4 m<strong>on</strong>ths later.<br />

Results:Behavioural and somatic symptoms as well as percepti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> general health (SF36) correlated to<br />

awakening cortisol reactivity (ACR). ACR decreased during the follow up period and correlated to decrease in<br />

symptom score. ACR did not correlate to TSH, HbA1C, fibrinogen or serum lipids.<br />

430


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

C<strong>on</strong>clusi<strong>on</strong>: An almost complete recovery was obtain in <strong>on</strong>e year<br />

The HPA axis functi<strong>on</strong> seems to be depressed during l<strong>on</strong>g term stress<br />

A change in metabolism in an anabolic directi<strong>on</strong> was followed by recovery<br />

7C_05_P<br />

(poster secti<strong>on</strong> A2, poster board #124, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

RELATIONSHIP BETWEEN PSYCHOPATHOLOGICAL SYMPTOMS AND<br />

PSYCHONEUROENEDOCRINIMMUNOLOGICAL INDICATORS IN PATIENTS<br />

WITH PANIC DISORDER<br />

Hisayoshi Okamura 1 , Akira Tsuda 2 , Jumpei Yajima 4 , Hiroo Fukuyama 3<br />

1Cognitive and Molecular <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Brain Diseases, Kurume, Japan, e-mail: oka1013@yahoo.co.jp<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychology<br />

3Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Social Welfare, Kurume University<br />

4Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Human Studies, Beppu University<br />

Many studies have reported that the accelerati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the noradrenergic neur<strong>on</strong>s in the brain have been<br />

recognized <strong>on</strong> the patients with panic disorder. There is substantial evidence that the anxiety state in human is<br />

associated with a high level <str<strong>on</strong>g>of</str<strong>on</strong>g> 3-methoxy-4-hydroxyphenylglychol(MHPG) in the plasma. These finding<br />

suggested that saliva level <str<strong>on</strong>g>of</str<strong>on</strong>g> free-MHPG seemed to be useful indicator <str<strong>on</strong>g>for</str<strong>on</strong>g> assessing the pschopathological<br />

symptoms <str<strong>on</strong>g>of</str<strong>on</strong>g> panic disorder.<br />

The purpose <str<strong>on</strong>g>of</str<strong>on</strong>g> this study is to investigate the relati<strong>on</strong>ships between the psychopathological symptoms with<br />

SSRI treatment and the saliva <str<strong>on</strong>g>of</str<strong>on</strong>g> Psych<strong>on</strong>euroendocrinimmunological (PNEI) indicators. Participants were 10<br />

first-visit outpatients with panic disorder (8 males, 2 females, 18-46 <str<strong>on</strong>g>of</str<strong>on</strong>g> age) and 20 age-matched healthy<br />

volunteers. In order to assess the level <str<strong>on</strong>g>of</str<strong>on</strong>g> free-MHPG, cortisol and s-IgA, the saliva was collected. In additi<strong>on</strong><br />

to this, the participants’ anxiety and mood levels were evaluated with HAS, POMS, and GHQ-28 by the<br />

psychiatrics in charge at their first visit, first week, third week and fifth week after beginning <str<strong>on</strong>g>of</str<strong>on</strong>g> SSRI drug<br />

treatment.<br />

The saliva levels <str<strong>on</strong>g>of</str<strong>on</strong>g> free-MHPG and cortisol at subjects’ first visit to the hospital were significantly higher and<br />

saliva level <str<strong>on</strong>g>of</str<strong>on</strong>g> s-IgA was lower than those <str<strong>on</strong>g>of</str<strong>on</strong>g> c<strong>on</strong>trol subjects. Following the 5 weeks <str<strong>on</strong>g>of</str<strong>on</strong>g> SSRI treatment, the<br />

saliva levels <str<strong>on</strong>g>of</str<strong>on</strong>g> free-MHPG and cortisol were decreased, while the saliva level <str<strong>on</strong>g>of</str<strong>on</strong>g> s-IgA was increased and the<br />

scores <str<strong>on</strong>g>of</str<strong>on</strong>g> HAS, POMS and GHQ-28 subscale <str<strong>on</strong>g>of</str<strong>on</strong>g> anxiety were improved. In additi<strong>on</strong> to this, the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> free-<br />

MHPG and cortisol were associated with the reduced anxiety and depressi<strong>on</strong>.<br />

These results indicated that free-MHPG, cortisol and s-IgA level can be useful indicators <str<strong>on</strong>g>for</str<strong>on</strong>g> assessing the<br />

pschopathological symptoms <str<strong>on</strong>g>of</str<strong>on</strong>g> panic disorder, and also the resp<strong>on</strong>se to drug treatment in these patients.<br />

431


23-26 August 2007,<br />

Budapest, Hungary<br />

432<br />

7C_06_P<br />

(poster secti<strong>on</strong> A2, poster board #125, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ALLERGIC DERMATITIS REDUCES STRESS-COPING SKILLS: INDICATIONS<br />

FROM A MOUSE MODEL<br />

M. Daniltchenko 1 , S. Pavlovic Masnikosa 1 , A. Örsal 1 , B. F. Klapp 1 , E. M. J. Peters 1<br />

1 University-Medicine Charité, Charité Center 12 <str<strong>on</strong>g>for</str<strong>on</strong>g> Internal Medicine and Dermatology, Psych<strong>on</strong>euroimmunology,<br />

Campus Virchow, Berlin, Germany; e-mail: eva.peters@charite.de<br />

Stress is said to cause, trigger and aggravate allergic diseases such as atopic dermatitis. Atopic patients show<br />

an altered HPA-axis reactivity and increased depressi<strong>on</strong> and anxiety which have been related to disease<br />

severity. However, if stress affects dermatitis or dermatitis causes stress remains unclear. To address this<br />

questi<strong>on</strong>, we employed a combined mouse model <str<strong>on</strong>g>of</str<strong>on</strong>g> experimental allergic dermatitis (AD) and stress. AD was<br />

induced in C57BL/6 mice by double sensitizati<strong>on</strong> (i.p) and an intradermal challenge using chicken egg<br />

ovalbumin. Animals were additi<strong>on</strong>ally exposed to s<strong>on</strong>ic stress <str<strong>on</strong>g>for</str<strong>on</strong>g> 24h prior to challenge. We m<strong>on</strong>itored the<br />

alterati<strong>on</strong>s in anxiety- and depressi<strong>on</strong>- like behaviour, locomotor activity, explorati<strong>on</strong>, and “approach/avoid<br />

c<strong>on</strong>flict” behaviour using elevated plus maze (EPM) and tail suspensi<strong>on</strong> test (TST). AD inducti<strong>on</strong> caused<br />

slight increase in anxiety-like behaviour with tendency to avoid c<strong>on</strong>flict situati<strong>on</strong>. Interestingly, stress itself<br />

promoted locomoti<strong>on</strong> and explorati<strong>on</strong>. However, this effect <str<strong>on</strong>g>of</str<strong>on</strong>g> stress was reduced when AD mice had been<br />

exposed to stress. No significant differences in depressi<strong>on</strong>-like behaviour were observed am<strong>on</strong>g the examined<br />

groups as measured in TST. At the same time we observed a significant decrease in c-Fos+ activated neur<strong>on</strong>s<br />

in the hypothalamus in AD mice exposed to stress. Taken together, the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> a cutaneous inflammati<strong>on</strong><br />

in the AD mice affected both behaviour and HPA reactivity suggest that altered reacti<strong>on</strong> to stress stimuli and<br />

coping abilities is a result <str<strong>on</strong>g>of</str<strong>on</strong>g> AD. Enhancement <str<strong>on</strong>g>of</str<strong>on</strong>g> stress-coping skills there<str<strong>on</strong>g>for</str<strong>on</strong>g>e appears a useful measure to<br />

balance AD-induced behavioural changes.<br />

7C_07_P<br />

(poster secti<strong>on</strong> A2, poster board #126, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CNS-IMMUNE INTERACTIONS DURING STRESS : POSSIBLE ROLE FOR<br />

FREE RADICALS<br />

Arunabha Ray, Kavita Gulati<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmacology, Vallabhbhai Patel Chest Institute, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Delhi, Delhi-110 007, India<br />

e-mail: arunabha14@yahoo.co.in<br />

Free radicals are highly reactive moieties and both ROS and RNS have been implicated in a variety <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

pathophysiological states. CNS-Immune interacti<strong>on</strong>s are important regulators <str<strong>on</strong>g>of</str<strong>on</strong>g> stress resp<strong>on</strong>ses and the<br />

present study was designed to investigate the possible role <str<strong>on</strong>g>of</str<strong>on</strong>g> free radicals and their interacti<strong>on</strong>s during stressinduced<br />

behavioral and immunological resp<strong>on</strong>ses in rats. Exposure to restraint stress (RS) persistently<br />

suppressed behavioral activity in the EPM test, as evidenced by the reduced open arm entries and time spent<br />

in the open arms as compared to the c<strong>on</strong>trol (no RS). Pretreatment <str<strong>on</strong>g>of</str<strong>on</strong>g> rats with the antioxidants, ascorbic<br />

acid, melat<strong>on</strong>in and NO mimetics, L-arginine and isosorbide dinitrate, all differentially reversed RS-induced<br />

behavioral suppressi<strong>on</strong> in the EPM test. The NO synthase inhibitors, L-NAME and 7-nitroindazole, <strong>on</strong> the<br />

other hand, either had no influence or induced aggravati<strong>on</strong>s in the neurobehavioral suppressi<strong>on</strong> in the EPM<br />

test. Assay <str<strong>on</strong>g>of</str<strong>on</strong>g> brain homogenates showed that RS-induced behavioral changes were closely paralleled by<br />

enhanced brain MDA levels, and lowered brain NOx and glutathi<strong>on</strong>e levels. These changes were significantly<br />

modified by the antioxidant and NO mimetic pretreatments. In rats immunized with sheep RBC, RS


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

suppressed both humoral and cell-mediated immune resp<strong>on</strong>ses, and differentially modulated cytokine (TNFα<br />

and IL-4) levels, which were attenuated by the anti-oxidants and NO precursors/releasers and aggravated<br />

by NO synthase inhibitors, These immunological changes were accompanied by corresp<strong>on</strong>ding changes in<br />

plasma MDA, glutathi<strong>on</strong>e, and NOx. These behavioral, immunological and biochemical data in resp<strong>on</strong>se to<br />

RS and its alterati<strong>on</strong>s by antioxidants and NO-ergic agents are str<strong>on</strong>gly suggestive <str<strong>on</strong>g>of</str<strong>on</strong>g> the involvement <str<strong>on</strong>g>of</str<strong>on</strong>g> ROS<br />

and RNS, in the CNS-Immune interacti<strong>on</strong>s during stress.<br />

7C_08_P<br />

(poster secti<strong>on</strong> A2, poster board #127, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

WALKING THE LABYRINTH: EFFECTS ON MIND, BODY, AND SPIRIT<br />

M. Kay Sandor<br />

The University <str<strong>on</strong>g>of</str<strong>on</strong>g> Texas Medical Branch, School <str<strong>on</strong>g>of</str<strong>on</strong>g> Nursing301 University Blvd. Galvest<strong>on</strong>, Texas 77555-1029, e-<br />

mail: ksandor@utmb.edu<br />

Background: This project examines the physiologic and psychospiritual resp<strong>on</strong>ses to a walking meditati<strong>on</strong>—<br />

the labyrinth. Meditati<strong>on</strong> is a complementary health practice that integrates the mind, the body, and the spirit.<br />

Labyrinths have been reported to reduce stress and induce relaxati<strong>on</strong>, but no studies testing the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> a<br />

labyrinth walk appear in the literature. The purpose <str<strong>on</strong>g>of</str<strong>on</strong>g> this pilot study is to examine the physiologic and<br />

psychospiritual resp<strong>on</strong>ses to a program <str<strong>on</strong>g>of</str<strong>on</strong>g> walking the labyrinth compared to a program <str<strong>on</strong>g>of</str<strong>on</strong>g> walking <strong>on</strong> a track<br />

at a slow pace. The specific aims <str<strong>on</strong>g>of</str<strong>on</strong>g> this study are as follows:<br />

Evaluate the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> a walking meditati<strong>on</strong> (labyrinth) program <strong>on</strong> physiological markers <str<strong>on</strong>g>of</str<strong>on</strong>g> stress (blood<br />

pressure, pulse, respirati<strong>on</strong>s, Cortisol, IL-6 and CRP) in women 55-70 years <str<strong>on</strong>g>of</str<strong>on</strong>g> age.<br />

Determine the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> a walking meditati<strong>on</strong> (labyrinth) program <strong>on</strong> psychospiritual outcomes (affect,<br />

anxiety, aggressi<strong>on</strong>, depressi<strong>on</strong>, and spiritual well-being) in women 55-70 years <str<strong>on</strong>g>of</str<strong>on</strong>g> age.<br />

Method: An experimental, pretest-posttest, repeated-measures design was chosen <str<strong>on</strong>g>for</str<strong>on</strong>g> this pilot study. The<br />

phenomena <str<strong>on</strong>g>of</str<strong>on</strong>g> interest are the physiologic and psychospiritual resp<strong>on</strong>ses <str<strong>on</strong>g>of</str<strong>on</strong>g> women (55-70 years old) to a<br />

facilitated group labyrinth walking program [Interventi<strong>on</strong>] and a facilitated group track walking program<br />

[Comparis<strong>on</strong>]. Data was collected prior to walking (Time 1), at the end <str<strong>on</strong>g>of</str<strong>on</strong>g> a m<strong>on</strong>th-l<strong>on</strong>g facilitated group<br />

walking program (Time 2) and at the end <str<strong>on</strong>g>of</str<strong>on</strong>g> the a m<strong>on</strong>th <str<strong>on</strong>g>of</str<strong>on</strong>g> self-managed walks (Time 3).<br />

Results: Data analysis is in progress and findings will be reported.<br />

Discussi<strong>on</strong> and C<strong>on</strong>clusi<strong>on</strong>s: Holistic and innovative community-based interventi<strong>on</strong>s addressing disease<br />

preventi<strong>on</strong> and health promoti<strong>on</strong> in women 55-70 years <str<strong>on</strong>g>of</str<strong>on</strong>g> age, are an important complementary adjunct to<br />

traditi<strong>on</strong>al care.<br />

7C_09_P<br />

(poster secti<strong>on</strong> A2, poster board #128, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CHARACTERISTICS OF PSYCHOBIOLOGICAL STRESS RESPONSIVENESS ON<br />

MENTAL STRESS TESTING IN DEPRESSIVE SUBJECTS<br />

Jumpei Yajima, Hisayoshi Okamura, Satoshi Horiuchi, Akira Tsuda<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Human Studies, Beppu University, e-mail: yajima@mc.beppu-u.ac.jp<br />

Our previous study indicated that the acute stress caused increases in the saliva level <str<strong>on</strong>g>of</str<strong>on</strong>g> free-3-methoxy-4-<br />

hydroxyphenylglycol (MHPG) and secretory-immunoglobulin-A (s-IgA), cortisol, and the score <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

questi<strong>on</strong>naire (DSSQ: Dundee stress state questi<strong>on</strong>naire). We have clarified that the health state and the daily<br />

433


23-26 August 2007,<br />

Budapest, Hungary<br />

stress influence the Psych<strong>on</strong>euroendocirnoimmunological stress resp<strong>on</strong>se according to the model <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Experimental-field studies. This study investigated relati<strong>on</strong>ship between the psychobiological stress resp<strong>on</strong>se<br />

induced by mental stress testing and the depressi<strong>on</strong> which was evaluated by Beck Depressi<strong>on</strong> Inventory<br />

(BDI).Subjects were healthy 226 volunteers (20.3±3.2 years old). At first, the subjects completed BDI. They<br />

took 10 minutes rest in an armchair prior to the stress sessi<strong>on</strong>, and were exposed to 15 minutes mental stress<br />

testing. Be<str<strong>on</strong>g>for</str<strong>on</strong>g>e and after the stress sessi<strong>on</strong>, saliva was collected and the subject was assessed a written inquiry<br />

subjective scales. Mental stress testing increased the saliva free-MHPG level, s-IgA level, cortisol level and<br />

then gradually reduced to the normal range after the stress sessi<strong>on</strong>. Subjects also showed stress resp<strong>on</strong>ses<br />

such as the tense arousal, the self c<strong>on</strong>sciousness, and task-irrelevant thought. The saliva s-IgA level <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

depressi<strong>on</strong> group (BDI>7) was lower than that <str<strong>on</strong>g>of</str<strong>on</strong>g> the normal group (BDI


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7D. CULTURAL DIFFERENCES, CULTURAL CHANGES AND STRESS<br />

(MÁRIA KOPP)<br />

7D_01_S<br />

SOCIOECONOMIC AND PSYCHOSOCIAL DETERMINANTS OF CHRONIC STRESS<br />

IN A CHANGING SOCIETY<br />

Maria S. Kopp<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Behavioral Sciences, Semmelweis University, H-1089 Budapest, Nagyvárad t. 4, Hungary<br />

e-mail: kopmar@net.sote.hu<br />

In the last decades in the trans<str<strong>on</strong>g>for</str<strong>on</strong>g>ming societies <str<strong>on</strong>g>of</str<strong>on</strong>g> Central and Eastern Europe (CEE) premature mortality<br />

increased dramatically, especially am<strong>on</strong>g men. In Hungarostudy 2002 survey 12.640 pers<strong>on</strong>s were interviewed<br />

in their homes. They represent the Hungarian populati<strong>on</strong> according to age, sex and 150 sub-regi<strong>on</strong>s above<br />

age 18. Socioec<strong>on</strong>omic, psychosocial, work and family related factors, behavioral and self-reported health<br />

measures were recorded. From the latest Hungarostudy 2006 follow-up study 1130 men and 1529 women<br />

were included into the present study, who in 2002 were between the age <str<strong>on</strong>g>of</str<strong>on</strong>g> 40-69. By 2006 99 men (8.8%)<br />

and 53 women (3.6%) died in the 40-69 age group. After adjustment <str<strong>on</strong>g>for</str<strong>on</strong>g> traditi<strong>on</strong>al risk factors, work related<br />

measures, such as job insecurity and low social support from co-workers predicted significantly the all-cause<br />

mortality am<strong>on</strong>g men. Low educati<strong>on</strong> and low pers<strong>on</strong>al income were c<strong>on</strong>nected with premature mortality<br />

<strong>on</strong>ly am<strong>on</strong>g men. Living with the spouse was significant protective factor against early death <strong>on</strong>ly am<strong>on</strong>g<br />

men, whereas dissatisfacti<strong>on</strong> with pers<strong>on</strong>al relati<strong>on</strong>s was a significant risk factor am<strong>on</strong>g women. Beck<br />

Depressi<strong>on</strong> score and self-rated health were much more important predictors <str<strong>on</strong>g>of</str<strong>on</strong>g> premature death am<strong>on</strong>g men<br />

than am<strong>on</strong>g women. In the trans<str<strong>on</strong>g>for</str<strong>on</strong>g>ming society <str<strong>on</strong>g>of</str<strong>on</strong>g> Hungary middle aged men are much more vulnerable to<br />

the work and socioec<strong>on</strong>omic deprivati<strong>on</strong> related chr<strong>on</strong>ic stress factors than women in the same age groups.<br />

Chr<strong>on</strong>ic stress is proposed as an integrating model that can be applied to understand the gender differences<br />

in premature mortality during social trans<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>.<br />

7D_02_S<br />

JOB STRAIN COMPONENTS AND ADRENERGIC ALFA-2 AND Β1-RECEPTOR-<br />

POLYMORPHISMS - A PUTATIVE STRESS-GENE INTERACTION AFFECTING<br />

OFFICE BLOOD PRESSURE LEVELS IN MEN<br />

Bertil Öhlin, Peter M. Nilss<strong>on</strong>, Olle Melander<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Clinical Sciences, Lund University, University Hospital, Malmö, S-205 02 Sweden<br />

Background: Job strain and the polymorphisms <str<strong>on</strong>g>of</str<strong>on</strong>g> both the alpha-2 and beta-1-adrenergic receptor genes<br />

have both been linked to blood pressure elevati<strong>on</strong>, respectively. We aimed at studying a potential interacti<strong>on</strong><br />

between job strain and its comp<strong>on</strong>ents (job demand and decisi<strong>on</strong> latitude), and the beta-1 receptor<br />

(Arg389Gly) polymorphism in relati<strong>on</strong> to <str<strong>on</strong>g>of</str<strong>on</strong>g>fice blood pressure.<br />

Methods: From the Malmö Diet and Cancer populati<strong>on</strong> cohort a total <str<strong>on</strong>g>of</str<strong>on</strong>g> 6095 subjects were randomly<br />

selected to be followed regarding cardiovascular risk factors. From this group, employed individuals with<br />

baseline questi<strong>on</strong>naire data regarding work characteristics were included (1338 men, 1707 women).<br />

Determinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the adrenergic β1-receptor Arg389Gly polymorphism was possible in 1271 men and 1601<br />

women, and these individuals <str<strong>on</strong>g>for</str<strong>on</strong>g>med the study group. Associati<strong>on</strong>s with the alpha-2 receptor have already<br />

been reported (Öhlin B, et al. J Hypertens 2007, accepted <str<strong>on</strong>g>for</str<strong>on</strong>g> publicati<strong>on</strong>)<br />

435


23-26 August 2007,<br />

Budapest, Hungary<br />

Results: Men with job strain and the Arg389Arg genotype had a n<strong>on</strong>-significant higher SBP (p=0.18) and<br />

DBP (p=0.34) than those with a Gly allele with or without job strain. The interacti<strong>on</strong> term genotype x job<br />

strain was <str<strong>on</strong>g>of</str<strong>on</strong>g> borderline significance <str<strong>on</strong>g>for</str<strong>on</strong>g> SBP (p=0.07) after adjustments <str<strong>on</strong>g>for</str<strong>on</strong>g> age, country <str<strong>on</strong>g>of</str<strong>on</strong>g> birth, and job<br />

status. The demand score was significantly interacting with genotype in men (p=0.01 <str<strong>on</strong>g>for</str<strong>on</strong>g> SBP, and p=0.009<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> DBP), after adjustments <str<strong>on</strong>g>for</str<strong>on</strong>g> age, country <str<strong>on</strong>g>of</str<strong>on</strong>g> birth, job status, antihypertensive treatment, and BMI. Men<br />

with a Gly allele had lower blood pressures with increasing demand score, whereas men homozygous <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

Arg allele had lower blood pressures with increasing latitude score. In women, those with job strain had<br />

borderline significantly higher blood pressures than those without job strain, regardless <str<strong>on</strong>g>of</str<strong>on</strong>g> genetic variants.<br />

C<strong>on</strong>clusi<strong>on</strong>s: Job strain, and in particular job demands, seems to interact with the Arg389Gly polymorphism<br />

in men, resulting in higher blood pressures in men with genotype Arg389Arg. The genotypes interact<br />

differently with the comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g> job strain (demand and decisi<strong>on</strong> latitude) in terms <str<strong>on</strong>g>of</str<strong>on</strong>g> blood pressure<br />

levels. These preliminary findings need to be addressed in future studies.<br />

436<br />

7D_03_S<br />

WORK STRESS AND HEALTH IN THE CULTURAL CONTEXT<br />

OF GLOBALISATION<br />

Johannes Siegrist<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Duesseldorf, Germany<br />

<str<strong>on</strong>g>Ec<strong>on</strong>omic</str<strong>on</strong>g> globalisati<strong>on</strong> has far-reaching impact <strong>on</strong> the socio-cultural life and the health <str<strong>on</strong>g>of</str<strong>on</strong>g> populati<strong>on</strong>s in<br />

modern and rapidly developing societies. Technological progress goes al<strong>on</strong>g with the spread <str<strong>on</strong>g>of</str<strong>on</strong>g> a Western<br />

lifestyle that increases the risk <str<strong>on</strong>g>of</str<strong>on</strong>g> n<strong>on</strong>-communicable diseases. Importantly, a globally expanding market <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

capital, goods and labour results in an increase <str<strong>on</strong>g>of</str<strong>on</strong>g> stressful working c<strong>on</strong>diti<strong>on</strong>s am<strong>on</strong>g large parts <str<strong>on</strong>g>of</str<strong>on</strong>g> employed<br />

people, in additi<strong>on</strong> to the afflicti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> unemployment.<br />

Based <strong>on</strong> the discoveries <str<strong>on</strong>g>of</str<strong>on</strong>g> two leading theoretical models, a wide range <str<strong>on</strong>g>of</str<strong>on</strong>g> health-adverse effects <str<strong>on</strong>g>of</str<strong>on</strong>g> stressful<br />

work has now been identified. In these models, stressful work is defined as either a combinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> high<br />

demand and low c<strong>on</strong>trol in <strong>on</strong>e's job, and as an imbalance between high ef<str<strong>on</strong>g>for</str<strong>on</strong>g>t spent and low reward received<br />

in turn, where rewards include m<strong>on</strong>ey, esteem, and promoti<strong>on</strong> prospects including job security. This latter<br />

model is <str<strong>on</strong>g>of</str<strong>on</strong>g> particular interest in the c<strong>on</strong>text <str<strong>on</strong>g>of</str<strong>on</strong>g> globalisati<strong>on</strong> as it points to violati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> a fundamental<br />

principle <str<strong>on</strong>g>of</str<strong>on</strong>g> ec<strong>on</strong>omic and social exchange, reciprocity.<br />

Major research findings from epidemiological and experimental studies <str<strong>on</strong>g>of</str<strong>on</strong>g> work stress and health are<br />

dem<strong>on</strong>strated, with special emphasis <strong>on</strong> rapidly trans<str<strong>on</strong>g>for</str<strong>on</strong>g>ming societies. In the final part, policy implicati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

current scientific evidence are discussed, with a focus <strong>on</strong> worksite health promoti<strong>on</strong>, and promising<br />

preliminary findings from interventi<strong>on</strong> studies are reported. Yet, given the scale <str<strong>on</strong>g>of</str<strong>on</strong>g> challenge, coordinated<br />

internati<strong>on</strong>al strategies towards healthy work will be needed.<br />

7D_04_S<br />

PSYCHOSOCIAL STRESS MANAGEMENT PROGRAMS IN<br />

TRANSFORMING SOCIETIES<br />

Red<str<strong>on</strong>g>for</str<strong>on</strong>g>d B. Williams<br />

Duke University Medical Center, Durham, NC, USA<br />

Populati<strong>on</strong>s living in societies undergoing major transiti<strong>on</strong> can be subjected to mounting levels <str<strong>on</strong>g>of</str<strong>on</strong>g> stress that<br />

have the potential to produce serious health problems in some or all segments <str<strong>on</strong>g>of</str<strong>on</strong>g> the populati<strong>on</strong>. Since the


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

breakup <str<strong>on</strong>g>of</str<strong>on</strong>g> the Soviet Uni<strong>on</strong> in 1989, <str<strong>on</strong>g>for</str<strong>on</strong>g> example, there has been a marked increase in annual mortality in<br />

countries <str<strong>on</strong>g>of</str<strong>on</strong>g> the <str<strong>on</strong>g>for</str<strong>on</strong>g>mer Soviet Block, most marked in Russia where the annual mortality increased from<br />

600/100,000 to 900/100,000 – producing an excess <str<strong>on</strong>g>of</str<strong>on</strong>g> 400,000 deaths per year. Another example is India,<br />

where a c<strong>on</strong>tinuing dramatic expansi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the ec<strong>on</strong>omy has created another set <str<strong>on</strong>g>of</str<strong>on</strong>g> social, political and<br />

ec<strong>on</strong>omic upheavals. A recent WHO report estimates that by 2010, 65% <str<strong>on</strong>g>of</str<strong>on</strong>g> the world’s cardiac cases will be<br />

pers<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> Indian descent, both in India and elsewhere. It is likely that a significant proporti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these<br />

increasing mortality and cardiac problems are a result <str<strong>on</strong>g>of</str<strong>on</strong>g> the stresses encountered in these trans<str<strong>on</strong>g>for</str<strong>on</strong>g>ming<br />

societies. While all might agree that the ideal soluti<strong>on</strong> would be to ameliorate the social, political, and<br />

ec<strong>on</strong>omic c<strong>on</strong>diti<strong>on</strong>s that are creating the health-damaging stress, that may be a soluti<strong>on</strong> that is easier to<br />

c<strong>on</strong>template than to implement. In this presentati<strong>on</strong> I will review experiences with <strong>on</strong>e structured stress<br />

management program, the Williams LifeSkills Workshop, that has been implemented, with appropriate<br />

adaptati<strong>on</strong>s to the different cultures, in the Far East and Hungary. In both regi<strong>on</strong>s, results indicate that<br />

training in coping skills can be implemented with results that clearly document reduced stress levels. To have<br />

a favorable impact <strong>on</strong> the health problems in these regi<strong>on</strong>s at the public health level, however, it will be<br />

necessary to develop and implement delivery systems that can disseminate this training <strong>on</strong> a mass scale. I will<br />

c<strong>on</strong>clude by c<strong>on</strong>sidering how mass media and the internet might be used to do this.<br />

7D_05_S<br />

(poster secti<strong>on</strong> A2, poster board #130, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PSYCHOSOCIAL STRESS AT WORK AND OUTSIDE WORK AND PROBLEM<br />

DRINKING: THE HAPIEE STUDY<br />

H. Pikhart 1 , M. Bobak 1 , A. Pajak 2 , S. Malyutina 3 , R. Kubinova 4 , A. Peasey 1 , M. Marmot 1<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Epidemiology and Public Health, UCL, UK<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Epidemiology and Populati<strong>on</strong> Studies, Jagiell<strong>on</strong>ian University, Krakow, Poland<br />

3Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Internal Medicine, RAMS, Novosibirsk, Russia<br />

4NIPH, Prague, Czech Republic<br />

Background: It has been shown in the past that psychosocial factors (at work and outside work) are<br />

associated with various health outcomes. It is thought that they influence health partly through health<br />

behaviours. Aims: To examine the associati<strong>on</strong> between the ef<str<strong>on</strong>g>for</str<strong>on</strong>g>t-reward imbalance (ERI), overcommitment<br />

and c<strong>on</strong>trol over general life, and several alcohol related measures in three populati<strong>on</strong>s in Central and Eastern<br />

Europe in the HAPIEE Study. Methods: The sample <str<strong>on</strong>g>of</str<strong>on</strong>g> populati<strong>on</strong> aged 45–69 years old in Novosibirsk<br />

(Russia), Krakow (Poland), and 6 Czech towns completed a questi<strong>on</strong>naire in 2002-2005 that included ERI,<br />

overcommitment (OC), c<strong>on</strong>trol over life, and a number <str<strong>on</strong>g>of</str<strong>on</strong>g> sociodemographic variables. Annual alcohol<br />

intake, annual number <str<strong>on</strong>g>of</str<strong>on</strong>g> drinking sessi<strong>on</strong>s, the mean dose <str<strong>on</strong>g>of</str<strong>on</strong>g> alcohol per drinking sessi<strong>on</strong>, and binge drinking<br />

were calculated from graduated frequencies in the questi<strong>on</strong>naire. Data were also available <strong>on</strong> problem<br />

drinking (>2 positive answers <strong>on</strong> CAGE questi<strong>on</strong>naire). Only men who were full-time working at the time <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

interview were used <str<strong>on</strong>g>for</str<strong>on</strong>g> the analysis (N=6700). Results: After c<strong>on</strong>trolling <str<strong>on</strong>g>for</str<strong>on</strong>g> age and country, the measures <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

problem drinking were associated with ERI, OC and c<strong>on</strong>trol over life. Adjustment <str<strong>on</strong>g>for</str<strong>on</strong>g> socioec<strong>on</strong>omic<br />

positi<strong>on</strong> did not substantially change the results. When all 3 psychosocial c<strong>on</strong>structs were used at the same<br />

model, the estimated effects were reduced but still associated with the alcohol related outcomes. C<strong>on</strong>clusi<strong>on</strong>s:<br />

Stress at work and outside work expressed by ERI, OC and c<strong>on</strong>trol over life is associated with problem<br />

drinking in these populati<strong>on</strong>s.<br />

437


23-26 August 2007,<br />

Budapest, Hungary<br />

7D_01_P<br />

(poster secti<strong>on</strong> A2, poster board #131, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

INTERNET MAIL COUNSELING FOR SOCIAL WITHDRAWAL<br />

IN JAPANESE YOUTH<br />

Kuramoto Hidehiko<br />

Kita-no-Maru Clinic, Tokyo<br />

Social withdrawal (SW) <str<strong>on</strong>g>of</str<strong>on</strong>g> the youth, defined by 1)remaining at home <str<strong>on</strong>g>for</str<strong>on</strong>g> a durati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> six m<strong>on</strong>ths or l<strong>on</strong>ger<br />

with no social participati<strong>on</strong> and by 2)not being diagnosed as schizophrenia or other mental disorders, has<br />

become a serious problem in Japan. One <str<strong>on</strong>g>of</str<strong>on</strong>g> the major therapeutic difficulties resides in the SW case’s<br />

reluctance to meet the others. The internet mail counseling helps overcome those difficulties. In order to<br />

support SW cases, “the net counseling room campaign” was carried out intermittently by NHK (Japan Broadcasting<br />

Corporati<strong>on</strong>) from October 2002 to March 2005. The characteristics <str<strong>on</strong>g>of</str<strong>on</strong>g> the cases c<strong>on</strong>sulted from April 2004 to<br />

March 2005 were analyzed. Out <str<strong>on</strong>g>of</str<strong>on</strong>g> the total 767 individual cases, there were 334 males (43.5%) and 433<br />

females (56.5%). The average ages were 26.9 years old <str<strong>on</strong>g>for</str<strong>on</strong>g> males and 24.8 years old <str<strong>on</strong>g>for</str<strong>on</strong>g> females (p


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Our study has revealed important fact that can be c<strong>on</strong>sidered to be particular to Ukrainian unemployment -<br />

about 50% <str<strong>on</strong>g>of</str<strong>on</strong>g> the group showed low level <str<strong>on</strong>g>of</str<strong>on</strong>g> stress. This was an unexpected result because the western<br />

culture c<strong>on</strong>siders the unemployment as <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the worst mis<str<strong>on</strong>g>for</str<strong>on</strong>g>tunes, after death and disease. We can also<br />

state the cultural differences c<strong>on</strong>cerning the life events.<br />

The values <str<strong>on</strong>g>of</str<strong>on</strong>g> social intelligence and <str<strong>on</strong>g>of</str<strong>on</strong>g> workability (the overall estimate and the qualitative characteristics:<br />

reliability and accuracy) are significantly lower in the group <str<strong>on</strong>g>of</str<strong>on</strong>g> unemployed compared to the works. The<br />

unemployed pers<strong>on</strong>s’ attitude to work is very specific. The length <str<strong>on</strong>g>of</str<strong>on</strong>g> the unemployment and the level <str<strong>on</strong>g>of</str<strong>on</strong>g> stress<br />

affect both the workability <str<strong>on</strong>g>of</str<strong>on</strong>g> individuals and their attitude to work.<br />

7D_03_P<br />

(poster secti<strong>on</strong> A2, poster board #133, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STUDY THE INFLUENCE OF INDIAN AND WESTERN MUSIC ON EMOTIONAL<br />

MATURITY TO REDUCE THE STRESS OF INDIVIDUALS<br />

Urvashi Shrivastava<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Guidance and Counseling, Urvashi Kala Srishti, Pune, India<br />

The psychology <str<strong>on</strong>g>of</str<strong>on</strong>g> music has inspirati<strong>on</strong>al, therapeutic and spiritual values which are useful <str<strong>on</strong>g>for</str<strong>on</strong>g> stress<br />

management. Music not <strong>on</strong>ly pleases the mind but also gives pleasure and ecstasy to the soul. The present<br />

study intends to explore the influence <str<strong>on</strong>g>of</str<strong>on</strong>g> Indian and western music <strong>on</strong> Emoti<strong>on</strong>al Maturity to reduce<br />

‘STRESS” <str<strong>on</strong>g>of</str<strong>on</strong>g> individuals. The sample was comprised <str<strong>on</strong>g>of</str<strong>on</strong>g> 60 adolescents (13-20 years), which had 30 Indianmusic<br />

and 30 Western-music listeners. Following hypotheses was made:<br />

There would be difference in the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> Emoti<strong>on</strong>al maturity <str<strong>on</strong>g>of</str<strong>on</strong>g> Indian and Western music listeners.<br />

There would be difference in the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> Emoti<strong>on</strong>al maturity <str<strong>on</strong>g>of</str<strong>on</strong>g> male and female Indian and Western<br />

music listeners.<br />

“Musical Interest Test” was used to differentiate the music listeners. To measure the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> Emoti<strong>on</strong>al<br />

Maturity “EMS” (Emoti<strong>on</strong>al Maturity Scale – Mahesh Bhargava) was used. The important attributes <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

“Emoti<strong>on</strong>al Maturity (EM)” are;<br />

Firm sense <str<strong>on</strong>g>of</str<strong>on</strong>g> reality<br />

Flexibility<br />

Adoptability.<br />

As per the EMS scale higher the score lesser is the maturity level. The results indicated that by taking the<br />

c<strong>on</strong>solidated data <str<strong>on</strong>g>of</str<strong>on</strong>g> mean values <str<strong>on</strong>g>of</str<strong>on</strong>g> “EMS”, the highest emoti<strong>on</strong>al maturity is indicated by male listeners <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Indian music with the value <str<strong>on</strong>g>of</str<strong>on</strong>g> 71.2 and the lowest Emoti<strong>on</strong>al maturity is indicated by male listeners <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

western music with the value <str<strong>on</strong>g>of</str<strong>on</strong>g> 86.4. Just by c<strong>on</strong>sidering the “Mean Values” <str<strong>on</strong>g>of</str<strong>on</strong>g> listeners, Indian music<br />

listeners are highly emoti<strong>on</strong>ally matured than western music listeners. “t – test” was applied <strong>on</strong> the data <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> significance. Results also indicated that the maximum significant t-values in the grand total <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

“Emoti<strong>on</strong>al Instability, Emoti<strong>on</strong>al Regressi<strong>on</strong>, Maladjustment, Pers<strong>on</strong>ality Disintegrati<strong>on</strong> and lack <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Independence” is by the Male listeners <str<strong>on</strong>g>of</str<strong>on</strong>g> Indian and Western music with value <str<strong>on</strong>g>of</str<strong>on</strong>g> 7.73. The lowest<br />

significant value is observed between the “Female” listeners <str<strong>on</strong>g>of</str<strong>on</strong>g> Indian and Western music with t-value <str<strong>on</strong>g>of</str<strong>on</strong>g> <strong>on</strong>ly<br />

2.4.<br />

439


23-26 August 2007,<br />

Budapest, Hungary<br />

7D_04_P<br />

(poster secti<strong>on</strong> A2, poster board #134, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

MODELLING HOLISTIC HEALTH AND PREVENTION ETHICS IN<br />

MENTAL CARE<br />

Hannele Ylilehto 1 , Arja Rautio 2<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Public Health Science and General Practice, Box 5000, 90014 University <str<strong>on</strong>g>of</str<strong>on</strong>g> Oulu, Finland<br />

e-mail: hannele.ylilehto@oulu.fi, Tel.: +358 (0)8 537 5655<br />

2<str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g> Arctic Medicine, Thule Institute, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Oulu<br />

Understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> the aetiology <str<strong>on</strong>g>of</str<strong>on</strong>g> postpartum mood disorders requires the integrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> both psychosocial<br />

and biological risk factors. The purpose <str<strong>on</strong>g>of</str<strong>on</strong>g> the previous study was to examine the time after a child is born<br />

from the salutogenic perspective. The study evaluated the mood <str<strong>on</strong>g>of</str<strong>on</strong>g> the mothers using the Edinburgh<br />

Postnatal Depressi<strong>on</strong> Scale (EPDS) during pregnancy and after childbirth. The relati<strong>on</strong>ships <str<strong>on</strong>g>of</str<strong>on</strong>g> the couples<br />

were studied using parts <str<strong>on</strong>g>of</str<strong>on</strong>g> Spaniers’s Dyadic Adjustment Scale and part <str<strong>on</strong>g>of</str<strong>on</strong>g> Bienvenu’s Marital<br />

Communicati<strong>on</strong> Inventory. If the relati<strong>on</strong>ship was c<strong>on</strong>sidered bad, the risk <str<strong>on</strong>g>of</str<strong>on</strong>g> developing depressi<strong>on</strong> during<br />

pregnancy was 4.7 times higher (RR = 4.7, 95%, CI 2.8–8), and after childbirth 5.5 times higher (RR = 5.5,<br />

95%, CI 3.1–9.6). The qualitative secti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the study identifies the resources <str<strong>on</strong>g>for</str<strong>on</strong>g> recovery used by the<br />

subjects. A focused interview was carried out with 29 mothers 3–10 m<strong>on</strong>ths after childbirth. Many <str<strong>on</strong>g>of</str<strong>on</strong>g> those<br />

who had exceeded the cut-<str<strong>on</strong>g>of</str<strong>on</strong>g>f point felt they had suffered from passing melancholy or they had problems in<br />

their marital relati<strong>on</strong>ship. The objective is to study stress and recovery in a patient date c<strong>on</strong>sisting <str<strong>on</strong>g>of</str<strong>on</strong>g> subjects<br />

who feel they live in a difficult relati<strong>on</strong>ship. Themes <str<strong>on</strong>g>of</str<strong>on</strong>g> research: Recovery from depressi<strong>on</strong>, c<strong>on</strong>necti<strong>on</strong><br />

between breast infecti<strong>on</strong> and postpartum depressi<strong>on</strong>, c<strong>on</strong>necti<strong>on</strong> between postpartum depressi<strong>on</strong> and<br />

violence, relati<strong>on</strong>s between recovery and molecular phenomena. Preventi<strong>on</strong> ethics secti<strong>on</strong> will be applied in<br />

the background during the entire research project. This applicati<strong>on</strong> supports a wider project in modelling<br />

holistic health.<br />

440<br />

7D_05_P<br />

(poster secti<strong>on</strong> A2, poster board #135, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HIGH PREVALENCE OF WORK-FAMILY CONFLICT IN THE HUNGARIAN<br />

POPULATION: POTENTIAL STRESSORS AND CONSEQUENCES<br />

Szilvia Adam, Zsuzsa Gyorffy, Krisztina Neculai, Piroska Balog, Tamas Martos, Maria Kopp<br />

Semmelweis University, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Behavioural Sciences, Nagyvarad ter 4., HU-1089 Budapest, Hungary<br />

e-mail: adaszil@net.sote.hu<br />

Aims: This study explored the prevalence, stressors and potential c<strong>on</strong>sequences <str<strong>on</strong>g>of</str<strong>on</strong>g> work-family c<strong>on</strong>flict<br />

(WFC) in the Hungarian populati<strong>on</strong>.<br />

Methods: Cross-secti<strong>on</strong>al study in a subset (1248 men, 1471 women) <str<strong>on</strong>g>of</str<strong>on</strong>g> the Hungarostudy Epidemiological<br />

Panel (HEP 2006) using validated questi<strong>on</strong>naires to explore the prevalence <str<strong>on</strong>g>of</str<strong>on</strong>g> WFC, and its associati<strong>on</strong>s with<br />

work stress (Ef<str<strong>on</strong>g>for</str<strong>on</strong>g>t – Reward Imbalance Questi<strong>on</strong>naire) and social stress (Bergen Social Relati<strong>on</strong>ship Scale,<br />

Marital Stress Scale), intrinsic and extrinsic life goals or aspirati<strong>on</strong>s (Aspirati<strong>on</strong>s Index Scale), depressi<strong>on</strong><br />

(shortened Beck depressi<strong>on</strong> scale), and well-being (WHO well-being questi<strong>on</strong>naire).<br />

Results: The prevalence <str<strong>on</strong>g>of</str<strong>on</strong>g> WFC was 82% am<strong>on</strong>g subjects. Severe WFC (defined as WFC reported <str<strong>on</strong>g>of</str<strong>on</strong>g>ten or<br />

very <str<strong>on</strong>g>of</str<strong>on</strong>g>ten) was experienced by 19% <str<strong>on</strong>g>of</str<strong>on</strong>g> the subjects. Stepwise logistic regressi<strong>on</strong> analyses revealed that work<br />

stress, high work-related ef<str<strong>on</strong>g>for</str<strong>on</strong>g>t (β=0.298, p=0.000), lack <str<strong>on</strong>g>of</str<strong>on</strong>g> reward at work (β=-0.140, p=0.000), over-


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

commitment (β=0.114, p=0.000), social stress (β=0.105, p=0.000), marital stress (β=0.093, p=0.000), age<br />

(β=-0.062, p=0.003) and surplus <str<strong>on</strong>g>of</str<strong>on</strong>g> extrinsic life goals (β=0.059, p=0.004) predicted WFC best. WFC,<br />

gender, and age were identified as significant predictors <str<strong>on</strong>g>of</str<strong>on</strong>g> well-being and depressi<strong>on</strong>.<br />

C<strong>on</strong>clusi<strong>on</strong>: These findings suggest a potential path from WFC to adverse psychological outcomes in the<br />

Hungarian populati<strong>on</strong> and provide further data <str<strong>on</strong>g>for</str<strong>on</strong>g> cross-cultural occupati<strong>on</strong>al stress research <strong>on</strong> the workhome<br />

interface.<br />

7D_06_P<br />

(poster secti<strong>on</strong> A2, poster board #136, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EARLY AGEING AS A CONSEQUENCE OF ADVERSE PSYCHOSOCIAL<br />

STRESS EXPOSURE<br />

Peter M. Nilss<strong>on</strong><br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Clinical Sciences, Lund University, University Hospital, S-205 02 Malmö, Sweden<br />

e-mail: Peter.Nilss<strong>on</strong>@med.lu.se<br />

Normal ageing can be further divided into “successful” and “n<strong>on</strong>-successful” ageing, the latter linked to<br />

different manifestati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> early morbidity and mortality shaped by intricate gene-envir<strong>on</strong>mental interacti<strong>on</strong>s.<br />

There are well-known examples <str<strong>on</strong>g>of</str<strong>on</strong>g> pure genetic models <str<strong>on</strong>g>of</str<strong>on</strong>g> early ageing, such as progeria in childhood. On the<br />

other end <str<strong>on</strong>g>of</str<strong>on</strong>g> the ageing spectrum there are many examples <strong>on</strong> how a detrimental mix <str<strong>on</strong>g>of</str<strong>on</strong>g> poor living<br />

c<strong>on</strong>diti<strong>on</strong>s, increased psychosocial stress and unhealthy lifestyle habits add up to early biological ageing and a<br />

reduced lifespan. One important c<strong>on</strong>sequence <str<strong>on</strong>g>of</str<strong>on</strong>g> early ageing is cardiovascular disease (CVD) caused by risk<br />

factor changes <str<strong>on</strong>g>of</str<strong>on</strong>g> arterial stiffening and endothelial dysfuncti<strong>on</strong> as part <str<strong>on</strong>g>of</str<strong>on</strong>g> the Early Vascular Ageing (EVA)<br />

syndrome. Normal vascular ageing is associated with a gradual change <str<strong>on</strong>g>of</str<strong>on</strong>g> the vascular structure and functi<strong>on</strong>,<br />

resulting in decreased arterial compliance and increased arterial stiffening. The process is measurable directly<br />

by use <str<strong>on</strong>g>of</str<strong>on</strong>g> imaging devices such as ultrasound measurements <str<strong>on</strong>g>of</str<strong>on</strong>g> the arterial wall <str<strong>on</strong>g>for</str<strong>on</strong>g> evaluati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> IMT and<br />

plaque <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>. Another approach is to use physiological methods such as pulse wave velocity (PWV) or<br />

the arterial augmentati<strong>on</strong> index, as markers <str<strong>on</strong>g>of</str<strong>on</strong>g> arterial stiffening. Other methods include the measurements <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

different biomarkers reflecting endothelial functi<strong>on</strong>, as an important part <str<strong>on</strong>g>of</str<strong>on</strong>g> the vascular ageing process, or<br />

the measurement <str<strong>on</strong>g>of</str<strong>on</strong>g> telomere length. Telomeres <str<strong>on</strong>g>for</str<strong>on</strong>g>m the end cap <str<strong>on</strong>g>of</str<strong>on</strong>g> the DNA helix and are shortened by<br />

each cell divisi<strong>on</strong> until the cell can divide no more. It has been repeatedly shown that telomere length is<br />

shorter in subjects with arterial ageing, atherosclerosis, CHD, insulin resistance or reporting poor lifestyle<br />

habits as reflected in obesity and excessive smoking. Even psychosocial stress exposure has been shown to be<br />

associated with shorter telomere length, as well as with increased risk <str<strong>on</strong>g>of</str<strong>on</strong>g> CVD. In summary, differential<br />

ageing is influenced by genes and envir<strong>on</strong>mental factors, not at least increased psychosocial stress exposure.<br />

This will influence the risk <str<strong>on</strong>g>of</str<strong>on</strong>g> CVD, the major health hazard, in the EVA syndrome based <strong>on</strong> early vascular<br />

ageing and telomere attriti<strong>on</strong>.<br />

441


23-26 August 2007,<br />

Budapest, Hungary<br />

7D_07_P<br />

(poster secti<strong>on</strong> A2, poster board #137, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HEALTH BEHAVIORS AND HEALTH RISK AWARENESS IN JAPANESE AND<br />

ENGLISH COLLEGE STUDENTS<br />

Akira Tsuda<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychology, Kurume University, 1635 Miimachi, 839-8502 Kurume, Japan<br />

e-mail: tsuda_akira@kurume-u.ac.jp<br />

This study is to asses health behaviors and health risk awareness (i.e., knowledge) in college students <str<strong>on</strong>g>of</str<strong>on</strong>g> Japan<br />

(N=631) and England (N=707), using a standardized protocol suitable <str<strong>on</strong>g>for</str<strong>on</strong>g> translati<strong>on</strong> and administrati<strong>on</strong>. A<br />

Japanese versi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the inventory was basically adapted from The European Health and Behavior Survey<br />

(Wardle & Steptoe, 1991) which had evaluated diet-eating habits, positive health practices, preventive health<br />

care, driving behavior and substance use, and awareness about the influence <str<strong>on</strong>g>of</str<strong>on</strong>g> several behavioral risk factors<br />

including eating animal fat, smoking, stress, excessive alcohol c<strong>on</strong>sumpti<strong>on</strong>, heredity and exercise and so <strong>on</strong>,<br />

<strong>on</strong> some major life style-related disease such as heart disease, high blood pressure, lung cancer and breast<br />

cancer. Differences between Japanese and English students, as well as gender differences, were observed in<br />

a number <str<strong>on</strong>g>of</str<strong>on</strong>g> health behaviors. Generally, females were healthier than males in both countries, with the<br />

excepti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> sleeping time. The importance gaps in health risk awareness were identified in students <str<strong>on</strong>g>of</str<strong>on</strong>g> both<br />

countries, as well as gender differences. It seems to be complex associati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> health behavior prevalence<br />

and health risk awareness. These findings c<strong>on</strong>firmed previous reports <strong>on</strong> the relati<strong>on</strong>ships am<strong>on</strong>g sociocultural<br />

factors, health behaviors and health knowledge.<br />

442


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7E. DENTAL STRESS<br />

(TIBOR KÁROLY FÁBIÁN)<br />

7E_01_S<br />

ACHIEVING STRESSLESS PREDICTABLE PERIO / IMPLANT EXCELLENCE<br />

André P. Saadoun<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Southern Cali<str<strong>on</strong>g>for</str<strong>on</strong>g>nia, 1441 Eastlake Ave, CA 91108, Los Angeles, USA<br />

e-mail: andre.p.saadoun@wanadoo.fr<br />

The creati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> an esthetic implant restorati<strong>on</strong> with gingival architecture in harm<strong>on</strong>y with the adjacent<br />

dentiti<strong>on</strong> is a <str<strong>on</strong>g>for</str<strong>on</strong>g>midable challenge. The process <str<strong>on</strong>g>of</str<strong>on</strong>g> s<str<strong>on</strong>g>of</str<strong>on</strong>g>t and hard tissue healing must be understood and<br />

incorporated into a carefully coordinated sequence <str<strong>on</strong>g>of</str<strong>on</strong>g> therapy.<br />

The essential prerequisites in order to establish an optimal aesthetic implant restorati<strong>on</strong> should always remain<br />

a precise, comprehensive biological and prosthetic diagnosis as well as the choice <str<strong>on</strong>g>of</str<strong>on</strong>g> the most c<strong>on</strong>servative,<br />

appropriate, and least traumatic treatment <str<strong>on</strong>g>for</str<strong>on</strong>g> the patient to prevent any injury to the period<strong>on</strong>tal and dental<br />

structure and achieve a successful outcome.<br />

The final objective is to achieve an optimal crown restorati<strong>on</strong> surrounded by its natural gingival envir<strong>on</strong>ment,<br />

using delicate ossoeus and/or muco-gingival plastic surgery.<br />

A major evoluti<strong>on</strong> in implantology has taken place with tapered and rough surface implants, inserted in a <strong>on</strong>estep,<br />

n<strong>on</strong>-submerged surgical protocol.<br />

It is important to identify complicati<strong>on</strong>s and clinical mistakes and their implicati<strong>on</strong>s <strong>on</strong> the final esthetic<br />

outcome.<br />

7E_02_S<br />

THE ROLE OF STRESS IN PERIODONTAL DISEASE<br />

B. Willershausen, A. Kasaj, A. Pistorius<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Operative Dentistry, Johannes Gutenberg University, Mainz, Germany<br />

In the last two decades several investigati<strong>on</strong>s have been carried out <strong>on</strong> associati<strong>on</strong>s between stress factors and<br />

period<strong>on</strong>titis and a number pathogenesis models have been discussed and examined. As a physiologic<br />

measure <str<strong>on</strong>g>of</str<strong>on</strong>g> stress urin levels <str<strong>on</strong>g>of</str<strong>on</strong>g> corticosteroids have been used, and increased free cortisol levels in the urine<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> period<strong>on</strong>titis patients as compared with c<strong>on</strong>trols have been encountered. It was shown that emoti<strong>on</strong>al<br />

stress may interfere with normal immune functi<strong>on</strong> and may result in increased levels <str<strong>on</strong>g>of</str<strong>on</strong>g> circulating horm<strong>on</strong>es<br />

that can effect the period<strong>on</strong>tium. Stressful life events appear to lead to a greater prevalence <str<strong>on</strong>g>of</str<strong>on</strong>g> period<strong>on</strong>tal<br />

disease and individuals with financial worries, distress, depressi<strong>on</strong> or inadequate coping mechanisms have a<br />

more sever loss <str<strong>on</strong>g>of</str<strong>on</strong>g> clinical attachment. Furthermore, it could be dem<strong>on</strong>strated that anxiety and stress may<br />

have a negative influence <strong>on</strong> the resp<strong>on</strong>se to period<strong>on</strong>tal therapy and that patients resp<strong>on</strong>ding less well to<br />

period<strong>on</strong>tal treatment have more psychosocial strain and a more passivedependent pers<strong>on</strong>ality. Although<br />

epidemiologic data <strong>on</strong> the associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> stress and period<strong>on</strong>tal disease is still limited, it appears that stress<br />

may be a putative risk factor <str<strong>on</strong>g>for</str<strong>on</strong>g> period<strong>on</strong>titis.<br />

The purpose <str<strong>on</strong>g>of</str<strong>on</strong>g> this presentati<strong>on</strong> is to provide an overview, based <strong>on</strong> own data obtained from c<strong>on</strong>trolled<br />

clinical studies, <strong>on</strong> the associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> stress, distress and coping behaviours with period<strong>on</strong>tal disease.<br />

443


23-26 August 2007,<br />

Budapest, Hungary<br />

7E_03_S<br />

HYPNOSIS IN DENTISTRY<br />

W. R. Krause<br />

Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychiatry and Psychotherapy, Harz-Klinikum Wernigerode-Blankenburg, Germany<br />

e-mail: psychiatrie@harz-klinikum.de<br />

In daily practice dentists are faced with a wide array <str<strong>on</strong>g>of</str<strong>on</strong>g> psychological problems including dental fear, gagging,<br />

psychogenic denture intolerance, bruxism, atypical facial pain etc. Although several hypnotherapeutic<br />

methods can be used rather effectively to solve most <str<strong>on</strong>g>of</str<strong>on</strong>g> these problems [1,2], <strong>on</strong>ly few dentists use such<br />

techniques in his/her practice. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e the aim <str<strong>on</strong>g>of</str<strong>on</strong>g> the lecture is to introduce the possibilities <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

hypnotherapy in dentistry and to inspire the audience's ef<str<strong>on</strong>g>for</str<strong>on</strong>g>t to use this method in the dental practice. For<br />

this purpose author will summarize the theoretical background and basic clinical knowledge <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

hypnotherapy, and will describe how hypnosis can be used <str<strong>on</strong>g>for</str<strong>on</strong>g> a wide spectrum <str<strong>on</strong>g>of</str<strong>on</strong>g> dental problems.<br />

References: (1): Krause, W-R.: Hypnose und Autogenes Training. In Schultz, JH.: Hypnosetechnik.<br />

Praktische Anleitung zum Hypnotisieren für Ärzte. 9. Auflage, bearbeitet und ergänzt v<strong>on</strong> G. Iversen und W-<br />

R. Krause. Gustav Fischer Verlag, Stuttgart, 1994. (2): Staats, J., Krause, W-R.: Hypnotherapie in der<br />

zahnärztlichen Praxis. Hüthig Verlag, Heidelberg, 1995.<br />

7E_04_S<br />

PSYCHOSOMATIC DENTISTRY IN HUNGARY. TRENDS AND PROGRESS<br />

IN THE LAST 10 YEARS<br />

T. K. Fábián<br />

Clinic <str<strong>on</strong>g>of</str<strong>on</strong>g> Prosthetic Dentistry, Semmelweis University, Budapest, Hungary<br />

Following a rather l<strong>on</strong>g brake, an increase <str<strong>on</strong>g>of</str<strong>on</strong>g> interest in psychosomatic dentistry occurred in the middle <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

nineties in Hungary. From this time up to the present 67 written publicati<strong>on</strong>s were published from the<br />

Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Dentistry <str<strong>on</strong>g>of</str<strong>on</strong>g> the Semmelweis University Budapest related to this topic. During this time all<br />

together 32 researchers participated in this multidisciplinary project including 13 dentists, 6 psychologists, 5<br />

biochemists, 1 psychiatrist, 1 teacher, 5 dental student and 1 psychology student. <str<strong>on</strong>g>Research</str<strong>on</strong>g> topics included<br />

epidemiology <str<strong>on</strong>g>of</str<strong>on</strong>g> dental fear, clinical treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> dental fear and several oro-facial psychogenic symptoms,<br />

and stress related biochemical changes <str<strong>on</strong>g>of</str<strong>on</strong>g> saliva. The lecture will summarize the most important data and<br />

results <str<strong>on</strong>g>of</str<strong>on</strong>g> this 10 years project and give a broad outline <str<strong>on</strong>g>of</str<strong>on</strong>g> research possibilities in the next future.<br />

444


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7E_05_S<br />

(poster secti<strong>on</strong> A2, poster board #138, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DIFFERENTIAL ELICITATION OF THE SALIVA LEVEL OF 3-METHOXY-4-<br />

HYDROXYPHENYLGLYCOL (MHPG), A METABOLITE OF NORADRENALINE,<br />

INDUCED BY MENTAL STRESS TESTING<br />

Satoshi Horiuchi, Akira Tsuda, Hisayoshi Okamura, Jumpei Yajima<br />

Graduate School <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychology, Kurume University, 1635, Mii-machi, Kurume-shi, Fukuoka-ken, 839-8502, Japan<br />

e-mail: satosato.007@nifty.com<br />

3-methoxy-4-hydroxyphenylglycol (MHPG) is a principal metabolite <str<strong>on</strong>g>of</str<strong>on</strong>g> noradrenaline derived from the brain.<br />

There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, to assess the saliva levels <str<strong>on</strong>g>of</str<strong>on</strong>g> MHPG is c<strong>on</strong>sidered as a n<strong>on</strong>-invasive tool <str<strong>on</strong>g>for</str<strong>on</strong>g> measuring central<br />

noradrenergic activity. From previous findings, although it is suspected that changes in the saliva level <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

MHPG varies am<strong>on</strong>g modes <str<strong>on</strong>g>of</str<strong>on</strong>g> mental stress testing, very few studies have tested this possibility directly.<br />

There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, the purpose <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to test different elicitati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the saliva level <str<strong>on</strong>g>of</str<strong>on</strong>g> MHPG induced<br />

Stroop Color Word C<strong>on</strong>flict Test (SCWCT) and Uchida-Kraepelin Test (UKT) as modes <str<strong>on</strong>g>of</str<strong>on</strong>g> mental stress<br />

testing in ten male healthy volunteers. The participants per<str<strong>on</strong>g>for</str<strong>on</strong>g>med these two tasks at about the same time <strong>on</strong><br />

two different days according to their schedule available. The order <str<strong>on</strong>g>of</str<strong>on</strong>g> the tasks was counterbalanced. MHPG<br />

resp<strong>on</strong>ses varied am<strong>on</strong>g tasks; <strong>on</strong>ly loading SCWCT resulted in the significant increase in MHPG level in the<br />

saliva even while both SCWCT and UKT induced similar degree <str<strong>on</strong>g>of</str<strong>on</strong>g> subjective stress state. These results<br />

provide evidence that mental work load induces increase in levels <str<strong>on</strong>g>of</str<strong>on</strong>g> MHPG specifically, suggesting that there<br />

are specific situati<strong>on</strong>al dimensi<strong>on</strong>(s) which activates the central noradrenergic nervous system. Combined<br />

with previous studies focusing <strong>on</strong> individual difference, future attempts to clarify the dimensi<strong>on</strong> (s) would<br />

improve our understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> brain noradrenergic activity induced by acute stress.<br />

7E_06_S<br />

(poster secti<strong>on</strong> A2, poster board #139, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

AUTOGENIC TRAINING IN THE TREATMENT OF ATYPICAL FACIAL PAIN<br />

1M. Krause, 2 O. Müller, 3 M. T. Nguyen, 3 T. K. Fábián, 3 P. Fejérdy, 1 W. R. Krause<br />

1Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychiatry and Psychotherapy, Harz-Klinikum Wernigerode-Blankenburg, Germany<br />

2Clinic <str<strong>on</strong>g>of</str<strong>on</strong>g> Pediatric Dentistry and Orthod<strong>on</strong>tics, Semmelweis University Budapest, Hungary<br />

3Clinic <str<strong>on</strong>g>of</str<strong>on</strong>g> Prosthetic Dentistry, Semmelweis University, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Dentistry, Budapest, Hungary<br />

In this study 10 female patients (age between 29-58 yr.) with atypical oro-facial pain (ICD-10 category: F40-<br />

48) were treated with autogenic training in a small group, <strong>on</strong>ce a week <str<strong>on</strong>g>for</str<strong>on</strong>g> 10 weeks. The method was<br />

administered as an initial therapy in the early phase <str<strong>on</strong>g>of</str<strong>on</strong>g> treatment. The first two basic exercises (feeling <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

heaviness and warmth) were practiced <strong>on</strong>ly, in sitting positi<strong>on</strong> [1]. Suggesti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> the therapist [1,2]were given<br />

via microph<strong>on</strong>e with the use <str<strong>on</strong>g>of</str<strong>on</strong>g> loud-speakers. A relaxing background music were also given. Symptom<br />

palliati<strong>on</strong> was measured with numerical analogue scale. In 3 cases (30%) symptoms disappeared totally. At<br />

least partial symptom palliati<strong>on</strong> occurred in another 4 cases (40%). In 3 cases (30%) the treatment was<br />

unsuccessful. Belief <str<strong>on</strong>g>of</str<strong>on</strong>g> the patients in psychotherapy and patient-therapist relati<strong>on</strong>ship str<strong>on</strong>g enough <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

entering to the exploratory part <str<strong>on</strong>g>of</str<strong>on</strong>g> psychotherapy were developed in all cases (100%). References: (1): Krause,<br />

W-R.: Hypnose und Autogenes Training. In: Schultz, JH.: Hypnosetechnik. Praktische Anleitung zum<br />

Hypnotisieren für Ärzte. 9. Auflage, bearbeitet und ergänzt v<strong>on</strong> G. Iversen und W-R. Krause. Gustav Fischer<br />

Verlag, Stuttgart, 1994 Pp. 71-80. (2): Staats, J., Krause, W-R.: Hypnotherapie in der zahnärztlichen Praxis.<br />

Hüthig Verlag, Heidelberg, 1995. Pp 120-123.<br />

445


23-26 August 2007,<br />

Budapest, Hungary<br />

7E_01_P<br />

(poster secti<strong>on</strong> A2, poster board #140, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

INFLUENCE OF EXPERIMENTAL DEVIATED MANDIBULAR POSITIONING ON<br />

PREFRONTAL CORTEX<br />

ANALYSIS BY FUNCTIONAL NEAR-INFRARED SPECTROSCOPY<br />

Mami Shibusawa 1 , Keiichi Ishigami 1 , Kazuhiko Takayama 1 , Tomotaka Takeda 1 , Kazunori Nakajima 1 ,<br />

Kaoru Sakatani 2 .<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Sports Dentistry, Tokyo Dental College 1 , Chiba, Japan<br />

e-mail: sibusawa@tdc.ac.jp<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Neurological Surgery Nih<strong>on</strong> University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine 2 , Tokyo, Japan<br />

Exposure to stress increases dopaminergic activity in several regi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> the brain. This phenomen<strong>on</strong> is most<br />

pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ound in the mesocortical dopamine system, where stress has been shown to increase selectively the<br />

release and turnover rate <str<strong>on</strong>g>of</str<strong>on</strong>g> dopamine. Recent in vitro studies employing micro dialysis have indicated that<br />

amino acids can increase the release <str<strong>on</strong>g>of</str<strong>on</strong>g> dopamine from prefr<strong>on</strong>tal cortex (PFC) in rat or small animals.<br />

Disturbance <str<strong>on</strong>g>of</str<strong>on</strong>g> masticati<strong>on</strong> following occlusal dysfuncti<strong>on</strong> has been reported to induce mental stress.<br />

However the relati<strong>on</strong>ship between a dysfuncti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the stomatognathic system and stress remains to be<br />

clarified in human. Functi<strong>on</strong>al near infrared spectroscopy (fNIRS) is a powerful tool <str<strong>on</strong>g>for</str<strong>on</strong>g> n<strong>on</strong>invasive imaging.<br />

We used fNIRS to examine the PFC in healthy volunteers (N =5) under experimental deviated mandibular<br />

positi<strong>on</strong>ing (EDMP) achieved with a resin bite block. The block was positi<strong>on</strong>ed where the n<strong>on</strong>-masticatingside<br />

canine cusp came into c<strong>on</strong>tact. The trial, 300s in total, started with a 60s pre-rest block, and was followed<br />

by a 180s EDMP task block and a 60s post-rest block. We found that EDMP resulted in an increase in Oxy<br />

Hb in the bilateral PFC, indicating neural activati<strong>on</strong> in these areas. However, the area activated and degree <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

increase in Oxy Hb varied am<strong>on</strong>g subjects. Many studies have found that stress resulted in activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

PFC. Taken together with our results, this suggests that the EDMP cause stress, leading to activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

PFC.<br />

446


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7F. POST TRAUMATIC STRESS DISORDER<br />

(MARCUS ISING, CHRISTOPH CZERMAK)<br />

7F_01_S<br />

GENE - ENVIRONMENT INTERACTIONS IN PTSD AND OTHER STRESS-<br />

RELATED DISORDERS<br />

Marcus Ising<br />

Max Planck Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychiatry, Kraepelinstr. 2-10, 80804 Munich, Germany<br />

e-mail: ising@mpipsykl.mpg.de<br />

Stress related disorders can be defined as illnesses whose causati<strong>on</strong>, <strong>on</strong>set, or development is substantially<br />

influenced by stress and its neurobiological correlates. Am<strong>on</strong>g mental disorders, depressi<strong>on</strong>, anxiety, and<br />

post-traumatic stress disorder (PTSD) are typical examples <str<strong>on</strong>g>for</str<strong>on</strong>g> stress-related disorders. They are characterized<br />

by a moderate heritability suggesting the involvement <str<strong>on</strong>g>of</str<strong>on</strong>g> genetic vulnerability factors. Additi<strong>on</strong>ally,<br />

envir<strong>on</strong>mental influences including exposure to stressful life events and trauma c<strong>on</strong>tribute to the disease risk<br />

and can act as triggers <str<strong>on</strong>g>for</str<strong>on</strong>g> the <strong>on</strong>set <str<strong>on</strong>g>of</str<strong>on</strong>g> the disorder. Taking both into account, the investigati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> gene –<br />

envir<strong>on</strong>ment interacti<strong>on</strong>s is an important approach to elucidate the aetiology <str<strong>on</strong>g>of</str<strong>on</strong>g> stress-related disorders. We<br />

will discuss different approaches to examine gene – envir<strong>on</strong>ment interacti<strong>on</strong>s in depressi<strong>on</strong>, anxiety, and<br />

PTSD, summarize the findings from the literature and present own results. The investigati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> gene –<br />

envir<strong>on</strong>ment interacti<strong>on</strong>s could prove as a promising approach to extend our knowledge about the aetiology<br />

and to identify new treatment targets in stress-related disorders.<br />

7F_02_S<br />

PREDICTORS OF RESPONSE TO A NOVEL NK-1 RECEPTOR<br />

ANTAGONIST IN PTSD<br />

Sanjay J. Mathew, Kathryn Keegan, Kate Collins, Jin Fan, Dennis Charney<br />

Mount Sinai School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, One Gustave L. Levy Place Box 1217, NY, NY 10029<br />

e-mail: sanjay.mathew@mssm.edu<br />

Few robust predictors <str<strong>on</strong>g>of</str<strong>on</strong>g> resp<strong>on</strong>se to pharmacological treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> PTSD have been identified. In this study,<br />

we sought to identify demographic, clinical, neurochemical (CSF), structural magnetic res<strong>on</strong>ance imaging<br />

(MRI) moderators <str<strong>on</strong>g>of</str<strong>on</strong>g> resp<strong>on</strong>se to a novel selective neurokinin-1 receptor antag<strong>on</strong>ist (GR205171) or pill<br />

placebo in patients with PTSD who participated in a 8 week randomized placebo-c<strong>on</strong>trolled double-blind<br />

clinical trial. Inasmuch as NKI receptors are highly expressed in medial temporal lobe (MTL) structures<br />

such amygdala and hippocampus, and elevated levels <str<strong>on</strong>g>of</str<strong>on</strong>g> CSF substance P have been reported in PTSD, we<br />

hypothesized that clinical resp<strong>on</strong>se would be associated with morphometric changes in MTL regi<strong>on</strong>s and<br />

reducti<strong>on</strong>s in CSF levels <str<strong>on</strong>g>of</str<strong>on</strong>g> substance P.<br />

Patients with PTSD related to a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> traumas (childhood abuse, combat, witnessing violence) were<br />

treated <str<strong>on</strong>g>for</str<strong>on</strong>g> 8 weeks with GR205171 (5 mg fixed dose) or matching pill placebo. All patients received 2 weeks<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> single-blind placebo prior to randomizati<strong>on</strong>, during which MRI and clinical ratings, using the Clinician<br />

Administered PTSD Scale (CAPS) and M<strong>on</strong>tgomery Asberg Depressi<strong>on</strong> Rating Scale (MADRS) was<br />

per<str<strong>on</strong>g>for</str<strong>on</strong>g>med. MRI was repeated prior to the end <str<strong>on</strong>g>of</str<strong>on</strong>g> the study, while patients were taking active drug or placebo.<br />

Lumbar punctures were also per<str<strong>on</strong>g>for</str<strong>on</strong>g>med at the beginning and endpoint <str<strong>on</strong>g>of</str<strong>on</strong>g> the study.<br />

447


23-26 August 2007,<br />

Budapest, Hungary<br />

448<br />

7F_03_S<br />

THE SEARCH TO CURE PTSD: EVIDENCE FOR NOVEL MOLECULAR TARGETS<br />

Christoph Czermak, Alexander Neumeister<br />

Yale University, Dept <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychiatry, West Haven, CT, USA<br />

e-mail: alexander.neumeister@yale.edu<br />

Indirect support exists <str<strong>on</strong>g>for</str<strong>on</strong>g> an involvement <str<strong>on</strong>g>of</str<strong>on</strong>g> the noradrenergic system, the serot<strong>on</strong>ergic system and nicotinic<br />

acetylcholine receptors in the pathophysiology <str<strong>on</strong>g>of</str<strong>on</strong>g> posttraumatic stress disorder (PTSD). To untangle the<br />

relative c<strong>on</strong>tributi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these transmitter systems to the pathophysiology <str<strong>on</strong>g>of</str<strong>on</strong>g> PTSD and to identify potential<br />

novel targets to treat PTSD, we c<strong>on</strong>ducted a series <str<strong>on</strong>g>of</str<strong>on</strong>g> functi<strong>on</strong>al imaging studies using positr<strong>on</strong> emissi<strong>on</strong><br />

tomography (PET) and single phot<strong>on</strong> emissi<strong>on</strong> computed tomography (SPECT). Using novel radioligands we<br />

studied <str<strong>on</strong>g>for</str<strong>on</strong>g> the first time the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> serot<strong>on</strong>in (5-HT) receptors types 1A and 1B, the norepinephrine<br />

transporter and nicotinic acetylcholine receptors in the brain <str<strong>on</strong>g>of</str<strong>on</strong>g> PTSD patients, trauma-exposed and n<strong>on</strong>traumatized<br />

healthy c<strong>on</strong>trol subjects. We also studied changes in electrodermal skin activity and<br />

neurochemistry following infusi<strong>on</strong> with the α 2 -antag<strong>on</strong>ist yohimbine in PTSD subjects relative to healthy<br />

c<strong>on</strong>trols. We found evidence <str<strong>on</strong>g>for</str<strong>on</strong>g> a role <str<strong>on</strong>g>of</str<strong>on</strong>g> the 5-HT1B receptor but not the 5-HT1A receptor in PTSD<br />

suggesting the 5-HT1B receptor as a potential target <str<strong>on</strong>g>for</str<strong>on</strong>g> drug development. We further substantiated the<br />

important role <str<strong>on</strong>g>of</str<strong>on</strong>g> noradrenergic mechanisms in PTSD. Altogether, our studies provide evidence <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

involvement <str<strong>on</strong>g>of</str<strong>on</strong>g> m<strong>on</strong>oaminergic mechanisms in the pathophysiology <str<strong>on</strong>g>of</str<strong>on</strong>g> PTSD and identify novel targets <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

drug development.<br />

7F_04_S<br />

THE PSYCHOBIOLOGY OF COPING WITH PTSD<br />

Miranda Olff<br />

Center <str<strong>on</strong>g>for</str<strong>on</strong>g> Psychological Trauma, Academic Medical Center University <str<strong>on</strong>g>of</str<strong>on</strong>g> Amsterdam, Meibergdreef 5, 1105 AZ<br />

Amsterdam, The Netherlands, e-mail: m.olff@amc.uva.nl<br />

Although many people are exposed to trauma, <strong>on</strong>ly part develop post traumatic stress disorder (PTSD) or<br />

other posttrauma psychopathology. It is possible that humans differ in the degree to which trauma induces<br />

neurobiological perturbati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> their threat resp<strong>on</strong>se systems, which may result in a differential degree <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

psychopathology. Posttraumatic stress disorder (PTSD) has been associated with dysregulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

hypothalamus-pituitary-adrenal (HPA) axis as well as <str<strong>on</strong>g>of</str<strong>on</strong>g> the hypothalamus-pituitary-thyroid (HPT) axis.<br />

Findings have not been c<strong>on</strong>sistent and may depend <strong>on</strong> methodological issues like c<strong>on</strong>trolling <str<strong>on</strong>g>for</str<strong>on</strong>g> relevant<br />

variables. Results <str<strong>on</strong>g>of</str<strong>on</strong>g> a meta-analysis will be shown. In this paper we also present data <strong>on</strong> six HPA and HPTaxis<br />

related horm<strong>on</strong>es in civilian chr<strong>on</strong>ic PTSD patients. Comparing chr<strong>on</strong>ic PTSD patients to healthy<br />

volunteers we found that patients had significantly lower plasma cortisol, prolactin and TSH levels. When<br />

studying the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> psychotherapy in chr<strong>on</strong>ic PTSD patients we found that after Brief Eclectic<br />

Psychotherapy (BEP) significant changes occurred in levels <str<strong>on</strong>g>of</str<strong>on</strong>g> cortisol and DHEA. Resp<strong>on</strong>ders showed an<br />

increase <str<strong>on</strong>g>of</str<strong>on</strong>g> cortisol and DHEA levels, while in n<strong>on</strong>-resp<strong>on</strong>ders both horm<strong>on</strong>e levels decreased. Differences<br />

were <strong>on</strong>ly found after c<strong>on</strong>trolling <str<strong>on</strong>g>for</str<strong>on</strong>g> depressive symptoms. In c<strong>on</strong>clusi<strong>on</strong>, learning to cope with trauma<br />

through psychotherapy <str<strong>on</strong>g>for</str<strong>on</strong>g> PTSD may alter dysregulati<strong>on</strong>s in the HPA-axis, but comorbid depressive<br />

symptoms should be taken into account. C<strong>on</strong>tinued study <str<strong>on</strong>g>of</str<strong>on</strong>g> the psychobiology <str<strong>on</strong>g>of</str<strong>on</strong>g> trauma and PTSD will<br />

enhance our understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> adaptati<strong>on</strong> to psychosocial stressors and support ef<str<strong>on</strong>g>for</str<strong>on</strong>g>ts to treat associated<br />

psychological and biological sequelae.


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7F_05_S<br />

(poster secti<strong>on</strong> A2, poster board #141, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE INFLUENCE OF PRECLINICAL ANALGESIA AND SEDATION ON THE<br />

ONSET, SEVERITY AND COURSE OF POSTTRAUMATIC STRESS SYMPTOMS<br />

IN ACCIDENT VICTIMS<br />

Michael Schönenberg, Ursula Reichwald, Kathrin Wilk, Heike Jacob, Gregor Domes,<br />

Andreas Badke, Martin Hautzinger<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Clinical and Developmental Psychology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Tübingen, Germany<br />

e-mail: michael.schoenenberg@uni-tuebingen.de<br />

We have previously dem<strong>on</strong>strated a str<strong>on</strong>g associati<strong>on</strong> between the applicati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a single or fracti<strong>on</strong>ated<br />

dose <str<strong>on</strong>g>of</str<strong>on</strong>g> the N-methyl-D-aspartate antag<strong>on</strong>ist ketamine and symptoms <str<strong>on</strong>g>of</str<strong>on</strong>g> acute (Schönenberg et al., 2007) and<br />

posttraumatic stress disorder (Schönenberg et al., 2005) in moderately injured accident victims. Ketamine has<br />

analgesic and sedative properties and <str<strong>on</strong>g>for</str<strong>on</strong>g> that reas<strong>on</strong> it is widely administered in emergency care. Based <strong>on</strong><br />

our previous data there is evidence to suggest that ketamine might pr<str<strong>on</strong>g>of</str<strong>on</strong>g>oundly augment early stress<br />

symptoms, probably via the enhancement <str<strong>on</strong>g>of</str<strong>on</strong>g> glutamatergic neurotransmissi<strong>on</strong>, resulting in worsened l<strong>on</strong>gterm<br />

posttraumatic sequelae. The objective <str<strong>on</strong>g>of</str<strong>on</strong>g> the study to be portrayed at the c<strong>on</strong>ference is to extend prior<br />

findings by prospectively examining ketamines’ effects <strong>on</strong> psychological, endocrinological and neurocognitive<br />

variables. Moderately injured accident victims were c<strong>on</strong>secutively recruited in two urban trauma<br />

centres. Based <strong>on</strong> their preclinical analgosedati<strong>on</strong>, patients were divided into groups (ketamine vs. opioids vs.<br />

no medicati<strong>on</strong>). Initial assessments <str<strong>on</strong>g>of</str<strong>on</strong>g> posttraumatic psychopathology were carried out within 48 hrs postevent<br />

during hospital-stay and were repeated two m<strong>on</strong>ths, six m<strong>on</strong>ths and <strong>on</strong>e year post-event. Further,<br />

neuropsychological assessments and a diurnal pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ile <str<strong>on</strong>g>of</str<strong>on</strong>g> the biological stress markers cortisol and<br />

dehydroepiandroster<strong>on</strong>e (DHEA) were determined at each time point. First results <str<strong>on</strong>g>of</str<strong>on</strong>g> the <strong>on</strong>going study will<br />

be presented at the c<strong>on</strong>ference.<br />

7F_06_S<br />

(poster secti<strong>on</strong> A2, poster board #142, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

RISK AND PROTECTIVE FACTORS IN RELATION TO PTSD SYMPTOMS<br />

Beth Spenciner Rosenthal, W. Cody Wils<strong>on</strong><br />

York College-City Univ. <str<strong>on</strong>g>of</str<strong>on</strong>g> NY, 94-20 Guy R. Brewer Blvd., Jamaica, NY 11451<br />

e-mail: rosenthal@york.cuny.edu<br />

The issues <str<strong>on</strong>g>of</str<strong>on</strong>g> risk and protective factors in relati<strong>on</strong>ship to manifesting PTSD symptoms have begun to be<br />

raised in the literature. A cross-secti<strong>on</strong>al correlati<strong>on</strong>al study <str<strong>on</strong>g>of</str<strong>on</strong>g> 848 adolescents recently graduated from high<br />

school in New York City tested three hypotheses: 1) witnessing the physical assault <str<strong>on</strong>g>of</str<strong>on</strong>g> another (independently<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> being a victim) is traumatic and is associated with PTSD symptoms; 2) having a sympathetic adult c<strong>on</strong>fidant<br />

available is a protective factor with regard to PTSD symptoms <str<strong>on</strong>g>for</str<strong>on</strong>g> older adolescents; and 3) having a sense <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

pers<strong>on</strong>al self efficacy is a protective factor with regard to PTSD symptoms. Theoretical analysis indicates that a<br />

“protective factor” may operate at three levels: 1) by reducing exposure to the “trauma event”; 2) by reducing<br />

levels <str<strong>on</strong>g>of</str<strong>on</strong>g> PTSD symptoms; and 3) by buffering or moderating the impact <str<strong>on</strong>g>of</str<strong>on</strong>g> the traumatic event <strong>on</strong> PTSD<br />

symptoms. The results <str<strong>on</strong>g>of</str<strong>on</strong>g> the empirical study indicate: first, that witnessing the physical assault <str<strong>on</strong>g>of</str<strong>on</strong>g> others<br />

produces PTSD symptoms (medium effect size); and sec<strong>on</strong>d, that both “availability <str<strong>on</strong>g>of</str<strong>on</strong>g> an adult c<strong>on</strong>fidant”<br />

and “feeling <str<strong>on</strong>g>of</str<strong>on</strong>g> self efficacy” reduce the likelihood <str<strong>on</strong>g>of</str<strong>on</strong>g> an adolescent being exposed to physical assault (small<br />

449


23-26 August 2007,<br />

Budapest, Hungary<br />

effect size), reduce the level <str<strong>on</strong>g>of</str<strong>on</strong>g> PTSD symptoms (medium effect size), but do not significantly moderate the<br />

impact <str<strong>on</strong>g>of</str<strong>on</strong>g> exposure to physical assault <strong>on</strong> PTSD. Thus, both “having available an adult c<strong>on</strong>fidant” and<br />

“feeling <str<strong>on</strong>g>of</str<strong>on</strong>g> self efficacy” act as protective factors <str<strong>on</strong>g>for</str<strong>on</strong>g> older adolescents with respect to manifesting PTSD<br />

symptoms. The principal protective mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> both “availability <str<strong>on</strong>g>of</str<strong>on</strong>g> an adult c<strong>on</strong>fidant” and “feeling <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

self efficacy” appears to be through a general reducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the level <str<strong>on</strong>g>of</str<strong>on</strong>g> PTSD symptoms.<br />

450


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7F_01_P<br />

(poster secti<strong>on</strong> A2, poster board #143, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ABSENCE OF ASSOCIATION BETWEEN GENETIC POLYMORPHISMS IN<br />

CRFR1 AND PTSD<br />

Dinko Pavlinić 1 , Goran Ćurić 1 , Marijana Braš 2 , Marta Đurković 3 , Rudolf Gregurek 2 , Pavo Filaković 3 ,<br />

Gordan Lauc 1<br />

1University <str<strong>on</strong>g>of</str<strong>on</strong>g> Osijek School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Chemistry and Biochemistry, Osijek, Croatia<br />

e-mail: glauc@pharma.hr<br />

2University <str<strong>on</strong>g>of</str<strong>on</strong>g> Osijek School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychiatry, Osijek, Croatia<br />

3University <str<strong>on</strong>g>of</str<strong>on</strong>g> Zagreb School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Clinic <str<strong>on</strong>g>for</str<strong>on</strong>g> Psychological Medicine, Zagreb, Croatia<br />

One <str<strong>on</strong>g>of</str<strong>on</strong>g> the main mechanisms in the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the hypothalamo–pituitary–adrenocortical (HPA) axis is the<br />

release <str<strong>on</strong>g>of</str<strong>on</strong>g> corticotropin releasing horm<strong>on</strong>e horm<strong>on</strong>e (CRH) from the hypothalamus and it’s binding to the<br />

corticotropin releasing horm<strong>on</strong>e receptor (CRHR1) in the pituitary gland. Recently it was reported that two<br />

haplotype tagging SNPs (htSNP) in CRHR1 are associated with patterns <str<strong>on</strong>g>of</str<strong>on</strong>g> human alcohol drinking and<br />

could potentially c<strong>on</strong>tribute to the development <str<strong>on</strong>g>of</str<strong>on</strong>g> alcohol dependence. Aiming to see whether same genetic<br />

variati<strong>on</strong>s are associated with potential predispositi<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> the development <str<strong>on</strong>g>of</str<strong>on</strong>g> post-traumatic stress disorder<br />

(PTSD) we have investigated associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these two haplotypes <str<strong>on</strong>g>of</str<strong>on</strong>g> CRHR1 and PTSD. DNA was isolated<br />

from blood <str<strong>on</strong>g>of</str<strong>on</strong>g> 200 patients with diagnosed PTSD and 200 matching c<strong>on</strong>trol individuals. TaqMan Predesigned<br />

SNP genotyping assays were used to genotype two htSNPs (rs242939 corresp<strong>on</strong>ding to T to C<br />

exchange at positi<strong>on</strong> 44371356 and rs1876830 corresp<strong>on</strong>ding to C to T exchange at positi<strong>on</strong> 44386772 <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Chromosome 17). C<strong>on</strong>trary to the situati<strong>on</strong> observed in alcoholism, we were not able to find any associati<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> CRFR1 genotype with PTSD since frequencies <str<strong>on</strong>g>of</str<strong>on</strong>g> all alleles were nearly the same in both studied<br />

populati<strong>on</strong>s. Interestingly, the observed frequency <str<strong>on</strong>g>of</str<strong>on</strong>g> the major allele at rs242939 corresp<strong>on</strong>ded to the<br />

frequency reported to correlate with low alcohol c<strong>on</strong>sumpti<strong>on</strong>, while the observed frequency <str<strong>on</strong>g>of</str<strong>on</strong>g> the major<br />

allele at rs1876830 was between the reported frequencies <str<strong>on</strong>g>for</str<strong>on</strong>g> low and high alcohol c<strong>on</strong>sumers.<br />

7F_02_P<br />

(poster secti<strong>on</strong> A2, poster board #144, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

RISK FACTORS FOR POSTTRAUMATIC STRESS DISORDER<br />

Virginia Olga B. Emery, Paul E. Emery<br />

Dartmouth Medical School, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychiatry HB 7750 USA<br />

<str<strong>on</strong>g>Research</str<strong>on</strong>g> <strong>on</strong> risk factors in the development <str<strong>on</strong>g>of</str<strong>on</strong>g> Posttraumatic Stress Disorder (PTSD) can be c<strong>on</strong>ceptualized<br />

using three major paradigms: (1) risk due to preexisting psychopathology; (2) risk due to preexisting traits or<br />

characteristics, which fall al<strong>on</strong>g a normal-subclinical c<strong>on</strong>tinuum; (3) risk due to preexisting childhood stress<br />

factors in family <str<strong>on</strong>g>of</str<strong>on</strong>g> origin. The research to be reported can be categorized under this third paradigm. The<br />

study to be reported was designed to compare antecedent stressors in homes <str<strong>on</strong>g>of</str<strong>on</strong>g> origin in a group <str<strong>on</strong>g>of</str<strong>on</strong>g> 20<br />

Vietnam veterans diagnosed as having PTSD and 20 Vietnam veterans not meeting criteria <str<strong>on</strong>g>for</str<strong>on</strong>g> PTSD.<br />

Samples were balanced <strong>on</strong> demographic variables, draft status, branch <str<strong>on</strong>g>of</str<strong>on</strong>g> service, rank, and type <str<strong>on</strong>g>of</str<strong>on</strong>g> discharge.<br />

Also, exposure to combat was c<strong>on</strong>trolled across samples. Findings suggest that veterans who developed<br />

PTSD had greater childhood stress related to parental alcoholism and parental unemployment than did n<strong>on</strong>-<br />

PTSD counterparts. Additi<strong>on</strong>al findings are discussed as are methodological refinements needed to better<br />

h<strong>on</strong>e in <strong>on</strong> relati<strong>on</strong>ship between family system stressors during primary socializati<strong>on</strong> and risk <str<strong>on</strong>g>for</str<strong>on</strong>g> PTSD.<br />

451


23-26 August 2007,<br />

Budapest, Hungary<br />

7F_03_P<br />

(poster secti<strong>on</strong> A2, poster board #145, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

RTM - NON VERBAL APPROACH IN HELPING CHILDREN VICTIMIZED BY<br />

THE TSUNAMI<br />

Moshe Farchi<br />

Tel Hai Academic College, Israel<br />

Rauma ( circle) -Trauma Model (R.T.M) was the main interventi<strong>on</strong> model that was developed <strong>on</strong>e m<strong>on</strong>th<br />

after the Tsunami in Srilnka by Israeli delegati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> psychologist and Social workers.<br />

Strategy planning:<br />

The interventi<strong>on</strong> strategy was based <strong>on</strong> 3 basic interventi<strong>on</strong> models:<br />

- Prol<strong>on</strong>ged Expusure (Foa 1992).<br />

- C<strong>on</strong>servati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> resources (Hobfol, 1988).<br />

- Basic-ph (Lahad, 1986).<br />

On sight assessment and insights:<br />

1. The disaster accrued <strong>on</strong>e m<strong>on</strong>th be<str<strong>on</strong>g>for</str<strong>on</strong>g>e the arrival <str<strong>on</strong>g>of</str<strong>on</strong>g> the delegati<strong>on</strong>s. symptoms <str<strong>on</strong>g>of</str<strong>on</strong>g> PTSD are about<br />

rise up: Interventi<strong>on</strong> must be trauma oriented with PTSD privati<strong>on</strong> techniques.<br />

2. No debriefing was d<strong>on</strong>e nor any kind <str<strong>on</strong>g>of</str<strong>on</strong>g> trauma processing.<br />

3. Trauma processing techniques must be – n<strong>on</strong> verbal since therapist are not familiar with the local<br />

languish .<br />

4. Local culture and coping recourses are deferent then therapist culture and beliefs: The interventi<strong>on</strong><br />

will use local n<strong>on</strong> verbal symbols.<br />

5. There is no secure that the interventi<strong>on</strong> will be c<strong>on</strong>tinued after the end <str<strong>on</strong>g>of</str<strong>on</strong>g> the delegati<strong>on</strong> missi<strong>on</strong>: The<br />

interventi<strong>on</strong> will be focused <strong>on</strong> One single meeting – with the opti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> c<strong>on</strong>tinuity.<br />

Field interventi<strong>on</strong>:<br />

Taking all the above c<strong>on</strong>siderati<strong>on</strong>s – including the theoretical point <str<strong>on</strong>g>of</str<strong>on</strong>g> view, 7 stages <str<strong>on</strong>g>of</str<strong>on</strong>g> interventi<strong>on</strong> model<br />

was drown called Rauma - Trauma Model (R.T.M).<br />

The main point <str<strong>on</strong>g>of</str<strong>on</strong>g> the rauma-trauma model is c<strong>on</strong>necting the children to the very initial coping resources –<br />

based <strong>on</strong> the above described models:<br />

Stage 1: Getting together:<br />

Stage 2: OKing – feeling the circle as a secure place<br />

Stage 3 Expressing and legitimating feelings<br />

Stage 4. Regaining coping resources<br />

Stage 5 Tsunami rec<strong>on</strong>structi<strong>on</strong><br />

Stage 6: Reliving the event :Back to the sea<br />

Stage 7 Ending<br />

The stages order dem<strong>on</strong>strate actual n<strong>on</strong> verbal debriefing with adjustments to the local culture.<br />

Outcomes and c<strong>on</strong>clusi<strong>on</strong>s:<br />

Main Outcomes: * After <strong>on</strong>ly <strong>on</strong>e interventi<strong>on</strong> the children where reported by their parents to sleep<br />

through the night without nightmares. Some declining in violence was reported as<br />

well.<br />

* R.T.M was found to be most useful in the first two time <str<strong>on</strong>g>of</str<strong>on</strong>g> per<str<strong>on</strong>g>for</str<strong>on</strong>g>ming – in the<br />

same group. More then 3 time caused the children to be passive and uncooperative.<br />

452


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7F_04_P<br />

(poster secti<strong>on</strong> A2, poster board #146, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

POSTTRAUMATIC STRESS DISORDER AND TERRORISM: WHAT DO WE KNOW<br />

J<strong>on</strong>athan D. E. Laugharne<br />

School <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychiatry and Clinical Neurosciences, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Western Australia, 16 The Terrace, Fremantle,<br />

WA 6160, Australia<br />

There is an increasing body <str<strong>on</strong>g>of</str<strong>on</strong>g> data relating to the psychological effects <str<strong>on</strong>g>of</str<strong>on</strong>g> terrorism, in particular relating to<br />

Posttraumatic Stress Disorder (PTSD). Evidence includes studies in the wake <str<strong>on</strong>g>of</str<strong>on</strong>g> the 9/11 attacks in the US in<br />

2001, the Oklahoma bombing, as well as terrorist activity in Israel, and Kenya. Some cauti<strong>on</strong> must be<br />

exercised in drawing general c<strong>on</strong>clusi<strong>on</strong>s from the data as numerous variables are operating including<br />

different socio-political c<strong>on</strong>texts within which the attacks occur. Elevated rates <str<strong>on</strong>g>of</str<strong>on</strong>g> PTSD in the general<br />

populati<strong>on</strong> follow terrorist attacks but so<strong>on</strong> normalize whereas directly exposed populati<strong>on</strong>s have higher rates<br />

(20-38%) and more persistent symptoms. An increased risk <str<strong>on</strong>g>of</str<strong>on</strong>g> PTSD is associated with direct exposure,<br />

geographical proximity, female sex, low income, poor educati<strong>on</strong>, poor social supports, prior psychotropic<br />

drug use and high-level media reporting <str<strong>on</strong>g>of</str<strong>on</strong>g> events (<str<strong>on</strong>g>for</str<strong>on</strong>g> vulnerable individuals). Studies to date indicate a high<br />

degree <str<strong>on</strong>g>of</str<strong>on</strong>g> psychological resilience in all populati<strong>on</strong>s affected by terrorism.<br />

7F_05_P<br />

(poster secti<strong>on</strong> A2, poster board #147, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

RELATIONSHIP BETWEEN TRAUMATIC BIOGRAPHIC EVENTS AND CURRENT<br />

PERSONALITY ORGANIZATION IN A SAMPLE OF REFUGEES<br />

AND ASYLUM SEEKERS<br />

L<strong>on</strong>y Schiltz<br />

CRP-Santé, 18, rue Dicks, L-1417 Luxembourg, e-mail: l<strong>on</strong>y.schiltz@educati<strong>on</strong>.lu<br />

To estimate the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> traumatic events, from the beginning <str<strong>on</strong>g>of</str<strong>on</strong>g> life up to recent stressors linked to natural<br />

catastrophes, war, political persecuti<strong>on</strong> and migrati<strong>on</strong>, we have undertaken an integrated clinical and<br />

experimental study with a sample <str<strong>on</strong>g>of</str<strong>on</strong>g> 73 refugees and asylum seekers. Every pers<strong>on</strong> had been seen individually<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> at least 8 hours. We used a mixed methodology, combining a semi-structured biographical interview, a<br />

projective test, i.e. the Sentences Completi<strong>on</strong> Test (Rotter & Willerman, 1949), <str<strong>on</strong>g>for</str<strong>on</strong>g> which we have developed<br />

a new manner <str<strong>on</strong>g>of</str<strong>on</strong>g> interpretati<strong>on</strong>, and psychometric scales, i.e. the HADS (Zigm<strong>on</strong>d & Snaith, 1983) and the<br />

Index <str<strong>on</strong>g>of</str<strong>on</strong>g> Wellbeing (Campbell & al, 1976). Furthermore, we analysed the artistic producti<strong>on</strong> (pictures, stories<br />

written under musical inducti<strong>on</strong>), realized by the people during the arts therapeutic sessi<strong>on</strong>s we <str<strong>on</strong>g>of</str<strong>on</strong>g>fered to<br />

them after the test sessi<strong>on</strong>, with the objective <str<strong>on</strong>g>of</str<strong>on</strong>g> helping them overcoming the break-up <str<strong>on</strong>g>of</str<strong>on</strong>g> their life project.<br />

As our data bel<strong>on</strong>ged partly to the nominal and ordinal level <str<strong>on</strong>g>of</str<strong>on</strong>g> measurement, we largely used n<strong>on</strong> parametric<br />

multidimensi<strong>on</strong>al statistics, like N<strong>on</strong> Linear Principal Comp<strong>on</strong>ents Analysis and multidimensi<strong>on</strong>al scaling. We<br />

could ex tract two pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iles <str<strong>on</strong>g>of</str<strong>on</strong>g> pers<strong>on</strong>ality functi<strong>on</strong>ing, linked either to repeated breaks, negligence and<br />

maltreatment from the beginning <str<strong>on</strong>g>of</str<strong>on</strong>g> life, or either to a recent external catastrophe, interrupting a c<strong>on</strong>tinuous<br />

life course. Through the evaluati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the arts therapeutic sessi<strong>on</strong>s, we could note the first signs <str<strong>on</strong>g>of</str<strong>on</strong>g> a<br />

resumpti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the blocked process <str<strong>on</strong>g>of</str<strong>on</strong>g> subjectivati<strong>on</strong>. Our theoretical discussi<strong>on</strong>, focused <strong>on</strong> the mode <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> arts psychotherapy, is based <strong>on</strong> an integrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> recent research results in psychopathology and<br />

neurobiology. The results <str<strong>on</strong>g>of</str<strong>on</strong>g> the studyallowed us proposing differential psychotherapeutic measures <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

two pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iles <str<strong>on</strong>g>of</str<strong>on</strong>g> traumatized people.<br />

453


23-26 August 2007,<br />

Budapest, Hungary<br />

454<br />

7F_06_P<br />

(poster secti<strong>on</strong> A2, poster board #148, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CORTISOL METABOLIC CLEARANCE IN POSTTRAUMATIC STRESS DISORDER<br />

G. H. Trevor Wheler, David Brand<strong>on</strong>, Aar<strong>on</strong> Clem<strong>on</strong>s, Crystal Riley, John Kendall,<br />

D. Lynn Loriaux, J. David Kinzie<br />

Oreg<strong>on</strong> Health and Sciences University, Portland, Oreg<strong>on</strong> 97239 USA<br />

Studies <str<strong>on</strong>g>of</str<strong>on</strong>g> glucocorticoid status in Posttraumatic Stress Disorder (PTSD) have hypothesized that these<br />

patients are maladapted to stress due to distorted feed back regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the HPA axis and/or altered<br />

sensitivity to glucocorticoid. However, cortisol levels in various biological fluids from PTSD subjects have<br />

been reported to be low, high or unchanged compared to c<strong>on</strong>trols. To address this issue, we have studied a<br />

group <str<strong>on</strong>g>of</str<strong>on</strong>g> patients with active PTSD symptoms, not currently <strong>on</strong> medicati<strong>on</strong>, in a c<strong>on</strong>trolled clinical study to<br />

determine the metabolic clearance rate <str<strong>on</strong>g>of</str<strong>on</strong>g> cortisol. PTSD symptomatology was c<strong>on</strong>firmed by DSM IV<br />

criteria, and by the Clinician Administered PTSD Scale (CAPS). The producti<strong>on</strong> rate <str<strong>on</strong>g>of</str<strong>on</strong>g> cortisol (PR) was<br />

measured after c<strong>on</strong>stant infusi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> tri-deuterated (d3) cortisol in normal saline administered at 20 µg per<br />

hour. Isotope diluti<strong>on</strong> ratios <str<strong>on</strong>g>of</str<strong>on</strong>g> cortisol (d0) and d3-cortisol were determined by stable isotope diluti<strong>on</strong> gas<br />

chromatography mass spectrometry using pooled serum samples as previously described (JCEM 91(9): 3486,<br />

2006). Blood samples (3ml) were taken every 30 minutes <str<strong>on</strong>g>for</str<strong>on</strong>g> 24 hours. Serum cortisol was measured by<br />

radioimmunoassay. Daily metabolic clearance rate (MCR24h) was calculated from the daily PR using the<br />

following <str<strong>on</strong>g>for</str<strong>on</strong>g>mula: MCR24h=PR/[cortisol]24h. The MCR24h <str<strong>on</strong>g>for</str<strong>on</strong>g> PTSD and c<strong>on</strong>trols was 122 ± 46.7<br />

L/d·m2 (n=10) and 94.6 ± 33.4 L/d·m2 (n=10) respectively in agreement with reported values. There was<br />

no statistically significant difference between the MCR24h in PTSD compared with c<strong>on</strong>trols. We c<strong>on</strong>clude<br />

that in the chr<strong>on</strong>ic and unprovoked state there is no dem<strong>on</strong>strable difference in producti<strong>on</strong> rate or metabolic<br />

clearance rate <str<strong>on</strong>g>of</str<strong>on</strong>g> cortisol, measured <strong>on</strong> a daily basis, in this group <str<strong>on</strong>g>of</str<strong>on</strong>g> PTSD subjects. This method provides a<br />

way to measure cortisol dynamics more specifically, and should be valuable in detailed assessments <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

psychological c<strong>on</strong>diti<strong>on</strong>s such as PTSD.<br />

7F_07_P<br />

(poster secti<strong>on</strong> A2, poster board #149, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

WORKLOAD, STRESS AND HEALTH AMONG MEDICAL STUDENTS<br />

Eda Merisalu, Vuokko Vahtera, Martin Täll<br />

Deparment <str<strong>on</strong>g>of</str<strong>on</strong>g> Public Health, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Tartu<br />

Str Ravila 19, Tartu 50411, Est<strong>on</strong>ia<br />

The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> the study was to analyse relati<strong>on</strong>ships between work load, stress and health complaints am<strong>on</strong>g<br />

medical students in Est<strong>on</strong>ia<br />

Method: The student study was carried out in late autumn 2005. The total study group c<strong>on</strong>sisted <str<strong>on</strong>g>of</str<strong>on</strong>g> 650<br />

participants from the sec<strong>on</strong>d to sixth course, whereas 470 medicine, 110 dentistry and 70 pharmacy students<br />

were included. Because <str<strong>on</strong>g>of</str<strong>on</strong>g> short learning experience the first course was not included into the study. The<br />

adapted an<strong>on</strong>ymous questi<strong>on</strong>naire Influence <str<strong>on</strong>g>of</str<strong>on</strong>g> studying <strong>on</strong> medical student’s health was used. The instrument was<br />

based <strong>on</strong> the questi<strong>on</strong>naire, used in <str<strong>on</strong>g>of</str<strong>on</strong>g>ficial IFEMSA project since 1998. There were 66 questi<strong>on</strong>s subdivided<br />

into 8 subgroups: demographic data, general health evaluati<strong>on</strong>, health, life style, work load, working<br />

c<strong>on</strong>diti<strong>on</strong>s, communicati<strong>on</strong> and future visi<strong>on</strong>. Yes/no answers, 10-point scale <str<strong>on</strong>g>for</str<strong>on</strong>g> stress and mainly 5-point<br />

scales were used. Also, opened questi<strong>on</strong>s were included.


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Results: The resp<strong>on</strong>se rate was 65,7%. Study group c<strong>on</strong>sisted <str<strong>on</strong>g>of</str<strong>on</strong>g> 427 resp<strong>on</strong>dents - 69% medicine, 19%<br />

dentistry and 12% farmacy students. Most part <str<strong>on</strong>g>of</str<strong>on</strong>g> them (82%) were females. Medicine students <str<strong>on</strong>g>of</str<strong>on</strong>g> younger<br />

courses had significantly higher work load than their older colleagues. A half <str<strong>on</strong>g>of</str<strong>on</strong>g> them spent <strong>on</strong> studies 8-10<br />

and more hours. The students with l<strong>on</strong>g working days described significantly higher stress than the others<br />

(p9) was measured am<strong>on</strong>g 7%, high (6-8) - 45% and moderate (3-5) - 35,5% <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

resp<strong>on</strong>dents (average 5,4±2,3 points). The most part <str<strong>on</strong>g>of</str<strong>on</strong>g> students assessed their health good (61%) and average<br />

(22%). Tiredness (85%), sore eyes (57%) and headache (54%) were the main health complaints am<strong>on</strong>g<br />

medical students. Than more health complaints, the higher stress and lack <str<strong>on</strong>g>of</str<strong>on</strong>g> physical activities and hobbies<br />

(p


23-26 August 2007,<br />

Budapest, Hungary<br />

7G. STRESS AND REPRODUCTION: FROM SEXUAL DYSFUNCTION TO<br />

SPONTANEOUS ABORTION<br />

(PABLO NEPOMNASCHY)<br />

7G_01_S<br />

STRESS-TRIGGERED REPRODUCTIVE SUPRESSION: AN<br />

EVOLVED ADAPTATION<br />

Pablo Nepomnaschy<br />

Nati<strong>on</strong>al Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Envir<strong>on</strong>mental Health Sciences-NIH, 27709-2233 PO BOX 12233, MD A3-05,<br />

<str<strong>on</strong>g>Research</str<strong>on</strong>g> Triangle Park, NC, USA, e-mail: nepomnaschyp@niehs.nih.gov<br />

It is estimated that <strong>on</strong>ly about 23% <str<strong>on</strong>g>of</str<strong>on</strong>g> women trying to c<strong>on</strong>ceive will begin a successful pregnancy during the<br />

first menstrual cycle <str<strong>on</strong>g>of</str<strong>on</strong>g> their attempt. This low rate <str<strong>on</strong>g>of</str<strong>on</strong>g> reproductive success has led some health scientists to<br />

describe human reproducti<strong>on</strong> as inefficient and, there<str<strong>on</strong>g>for</str<strong>on</strong>g>e, an evoluti<strong>on</strong>ary paradox. In c<strong>on</strong>trast to this<br />

pathological visi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> women's reproducti<strong>on</strong>, evoluti<strong>on</strong>ary theorists propose that reproductive suppressi<strong>on</strong><br />

may have originated as an adaptati<strong>on</strong> that, in dire circumstances, helps prevent pregnancy. As practiti<strong>on</strong>ers’<br />

positi<strong>on</strong>s <strong>on</strong> this issue can affect both diagnoses and treatments, the debate <str<strong>on</strong>g>of</str<strong>on</strong>g> these opposing paradigms is <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

critical importance.<br />

This talk c<strong>on</strong>tributes to the a<str<strong>on</strong>g>for</str<strong>on</strong>g>ementi<strong>on</strong>ed debate through an examinati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the physiologic effects <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

stress <strong>on</strong> women's reproductive functi<strong>on</strong>. Stress is comm<strong>on</strong>ly reported to lead to reproductive suppressi<strong>on</strong>.<br />

Most <str<strong>on</strong>g>of</str<strong>on</strong>g> the available evidence to support this claim, however, has been derived from animal and clinical or<br />

retrospective studies. Here I present data from a populati<strong>on</strong>-based, l<strong>on</strong>gitudinal study I c<strong>on</strong>ducted am<strong>on</strong>g<br />

Kaqchikel Maya in the southwestern highlands <str<strong>on</strong>g>of</str<strong>on</strong>g> Guatemala. The relati<strong>on</strong>ship between stress axis<br />

(hypothalmic-pituitary-adrenal) activati<strong>on</strong> and ovarian functi<strong>on</strong>, implantati<strong>on</strong> and early pregnancy fate is<br />

discussed. Evoluti<strong>on</strong>ary and pathologic aspects <str<strong>on</strong>g>of</str<strong>on</strong>g> the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> stress <strong>on</strong> reproductive functi<strong>on</strong> are evaluated.<br />

7G_02_S<br />

THE RELATIONSHIP BETWEEN PSYCHOLOGICAL STRESS AND<br />

FEMALE FERTILITY<br />

Sheiner Eyal<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Obstetrics and Gynecology, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Health Sciences & Soroka University Medical Center,<br />

Ben-Guri<strong>on</strong> University <str<strong>on</strong>g>of</str<strong>on</strong>g> the Negev, Beer-Sheva, Israel<br />

Fertility treatments carry emoti<strong>on</strong>al burdens <strong>on</strong> women and their partners. Although the psychosocial aspects<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> infertility have not been adequately addressed in clinical practice, there is c<strong>on</strong>sensus in the literature that<br />

attenti<strong>on</strong> to the psychological aspects <str<strong>on</strong>g>of</str<strong>on</strong>g> infertility is str<strong>on</strong>gly advisable. Psychological factors such as<br />

depressi<strong>on</strong>, state-anxiety, and stress-induced changes in heart rate might be predictive <str<strong>on</strong>g>of</str<strong>on</strong>g> a decreased<br />

probability <str<strong>on</strong>g>of</str<strong>on</strong>g> achieving a viable pregnancy in various types <str<strong>on</strong>g>of</str<strong>on</strong>g> infertility and fertility treatments. Previous<br />

interventi<strong>on</strong> trials did not include screening <str<strong>on</strong>g>for</str<strong>on</strong>g> psychological nor physiological stress markers known to<br />

predict pregnancy in women experiencing difficulties achieving pregnancy. It is not clear whether the<br />

psychological stress is a part <str<strong>on</strong>g>of</str<strong>on</strong>g> the infertility etiology as a causative factor (psychogenic hypothesis), or rather<br />

represents a development <str<strong>on</strong>g>of</str<strong>on</strong>g> the infertility problem (psychological c<strong>on</strong>sequences hypothesis). Nevertheless, it<br />

is more likely that there is an interactive causal associati<strong>on</strong> between infertility and psychosocial distress.<br />

456


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7G_03_S<br />

THE HYPOTHALAMIC-PITUITARY-ADRENAL AND THE HYPOTHALAMIC-<br />

PITUITARY-GONADAL AXES INTERPLAY IN REPRODUCTION<br />

George Mastorakos<br />

Endocrine Unit, Sec<strong>on</strong>d Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Obstretics and Gynecology, Aretaiei<strong>on</strong> Hospital,<br />

Athens University Medical School, Athens<br />

e-mail: mastorak@mail.kapatel.gr<br />

Vertebrates resp<strong>on</strong>d to stress with activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the HPA axis, the adrenergic and the aut<strong>on</strong>omic nervous<br />

systems. The principal central nervous system regulators <str<strong>on</strong>g>of</str<strong>on</strong>g> the HPA axis are corticotropin releasing horm<strong>on</strong>e<br />

(CRH) and antidiuretic horm<strong>on</strong>e (AVP). Apart central nervous system, CRH has been found in the adrenal<br />

medulla, ovaries, myometrium, endometrium, placenta, testis and elsewhere. The activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the HPA axis<br />

during stress affects all body systems. The reproductive axis is inhibited by HPA axis, <str<strong>on</strong>g>for</str<strong>on</strong>g> the sake <str<strong>on</strong>g>of</str<strong>on</strong>g> saving<br />

energy. The changes <str<strong>on</strong>g>of</str<strong>on</strong>g> the HPG axis during stress are species-specific, and depend <strong>on</strong> type and durati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the stimulus. Several c<strong>on</strong>diti<strong>on</strong>s may be associated with altered regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the HPA axis. Polycystic ovary<br />

syndrome, anorexia nervosa and pregnancy in the third trimester are all characterized by HPA axis activati<strong>on</strong>.<br />

In c<strong>on</strong>trast, during postpartum period, HPA axis suppressi<strong>on</strong> is implicated in the “postpartum blues”. The<br />

acti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> CRH are also essential in fetal development and ne<strong>on</strong>atal survival.<br />

7G_04_S<br />

STRESS DURING PREGNANCY: CONSEQUENCES FOR MOTHER AND CHILD<br />

P. C. Arck 1 , M. Ruecke 1 , K. Nakamura 2 , H. Fliege 1 , M. Rose 3 , S. Blois 1 , M. Pincus 1 , B. F. Klapp 1<br />

1 Charité, University Medicine Berlin, Germany<br />

2 University <str<strong>on</strong>g>of</str<strong>on</strong>g> Vancouver, Canada<br />

3 Qualitymetric, USA<br />

Maternal stress percepti<strong>on</strong> has l<strong>on</strong>g been suspected as a possible cause <str<strong>on</strong>g>of</str<strong>on</strong>g> infertility, implantati<strong>on</strong> failure, late<br />

pregnancy complicati<strong>on</strong>s and impaired fetal development, noti<strong>on</strong>s that exist since ancient times and across all<br />

cultures. In view <str<strong>on</strong>g>of</str<strong>on</strong>g> the enormous complexity <str<strong>on</strong>g>of</str<strong>on</strong>g> the regulatory nervous, endocrine and immune mechanisms<br />

involved in pregnancy maintenance, it is evident that pregnancy failure is not a single entity c<strong>on</strong>diti<strong>on</strong>, but<br />

most likely the result <str<strong>on</strong>g>of</str<strong>on</strong>g> complex dysregulati<strong>on</strong>. This dysregulati<strong>on</strong> can be initiated or aggravated by stress.<br />

While there are still more open questi<strong>on</strong>s than answers, the neuro-endocrine-immune circuitry <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress<br />

resp<strong>on</strong>se during pregnancy is becoming increasingly defined, e.g. due to the development <str<strong>on</strong>g>of</str<strong>on</strong>g> particularly<br />

instructive rodent models and prospectively designed human cohort trials. Subsequently, clinicians are<br />

becoming far more attentive to the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> psychological stress <strong>on</strong> pregnancy complicati<strong>on</strong>s. As a reseult <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

emerging basic science research endeavours elucidating hierarchical, temporal and spatial interacti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> key<br />

parameters during central and peripheral resp<strong>on</strong>ses to psychological stress, a list <str<strong>on</strong>g>of</str<strong>on</strong>g> candidate molecular<br />

targets <str<strong>on</strong>g>for</str<strong>on</strong>g> clinically useful therapeutic interventi<strong>on</strong> has become available by and should be tested in<br />

interdisciplinary research approaches.<br />

457


23-26 August 2007,<br />

Budapest, Hungary<br />

7G_05_S<br />

(poster secti<strong>on</strong> A2, poster board #150, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

REPRODUCTION OF SOUTHERN MURIQUIS IN CAPTIVITY (BRACHYTELES<br />

ARACHNOIDES): ENDOCRINOLOGICAL ASSESSMENT OF REPRODUCTIVE<br />

FUNCTION BY MEASUREMENT OF FECAL STEROID METABOLITES<br />

A. B. F. Lima 1 , P. Viau 1 , A. Pissinati 2 , M. O. M. Chelini 1 , C. A. Oliveira 1*<br />

1Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Horm<strong>on</strong>e Measurements <str<strong>on</strong>g>of</str<strong>on</strong>g> the Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Animal Reproducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the Faculty<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> Veterinary Medicine <str<strong>on</strong>g>of</str<strong>on</strong>g> the University <str<strong>on</strong>g>of</str<strong>on</strong>g> Sao Paulo, Brazil<br />

2Primate Center <str<strong>on</strong>g>of</str<strong>on</strong>g> Rio de Janeiro, Brazil<br />

e-mail: cadolive@usp.br<br />

The southern muriqui (Brachyteles. arachnoides), the largest Neotropical primate species, is <strong>on</strong> the verge <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

extincti<strong>on</strong> with critically endangered status (IUCN 2000). We validated n<strong>on</strong>invasive endocrine m<strong>on</strong>itoring<br />

techniques <str<strong>on</strong>g>for</str<strong>on</strong>g> this species by quantifying fecal estrogens and progestins in 4 adult females as well as fecal<br />

metabolites <str<strong>on</strong>g>of</str<strong>on</strong>g> testoster<strong>on</strong>e and glucocorticoids in 4 adult males and 1 sub-adult male. The study was<br />

c<strong>on</strong>ducted over a period <str<strong>on</strong>g>of</str<strong>on</strong>g> 11 m<strong>on</strong>ths under two different envir<strong>on</strong>mental c<strong>on</strong>diti<strong>on</strong>s: a) Two adult males,<br />

two adult females and a young male living <strong>on</strong> an island <str<strong>on</strong>g>of</str<strong>on</strong>g> 600 m 2, with natural vegetati<strong>on</strong>, at Curitiba Zoo<br />

(PPC) and b) two adult couples housed in a large cage <str<strong>on</strong>g>of</str<strong>on</strong>g> 15.40X5.85X4.70m at Rio de Janeiro Primate Center<br />

(CPRJ). Fecal samples were collected at least every other day over four c<strong>on</strong>tinuous periods <str<strong>on</strong>g>of</str<strong>on</strong>g> 20 to 24 days<br />

with 45-day intervals between to subsequent sampling periods. Reproductive behavior was observed <strong>on</strong> the<br />

same schedule. Fecal steroids were extracted by diluti<strong>on</strong> in ethanol. We used radioimmunoassay (RIA) in<br />

solid phase to quantify fecal estradiol, progester<strong>on</strong>e, testoster<strong>on</strong>e, and glucocorticoid metabolites. Estrogen<br />

and progestin c<strong>on</strong>centrati<strong>on</strong>s in female feces varied individually over a wide range (0.03 to 145.56µg/g <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

dried feces <str<strong>on</strong>g>for</str<strong>on</strong>g> progestins and 0.002 to 71.57µg/g <str<strong>on</strong>g>of</str<strong>on</strong>g> dried feces <str<strong>on</strong>g>for</str<strong>on</strong>g> estrogens). Three females did not show<br />

any ovarian activity over specific periods <str<strong>on</strong>g>of</str<strong>on</strong>g> the study, while ovarian cyclicity was observed over all sample<br />

periods in <strong>on</strong>e female from CPRJ. Despite <str<strong>on</strong>g>of</str<strong>on</strong>g> their low levels <str<strong>on</strong>g>of</str<strong>on</strong>g> fecal reproductive steroids, all females<br />

displayed proceptive behavior and copulati<strong>on</strong>s. Surprisingly, despite <str<strong>on</strong>g>of</str<strong>on</strong>g> their semi-free ranging c<strong>on</strong>diti<strong>on</strong>, the<br />

PPC males showed significantly higher c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> fecal glucocorticoids and lower c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

fecal testoster<strong>on</strong>e than the caged CPRJ males (p


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7G_06_S<br />

(poster secti<strong>on</strong> A2, poster board #151, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE IMPACT OF SECONDARY SEX CHARACTERISTICS (SSC) AND SOCIAL<br />

BEHAVIOR TO CORTISOL AND SEXUAL HORMONES IN FEMALE PRIMATES<br />

B. Wallner 1 , J. Dittami 2 , I. H. Machatschke 2<br />

1Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Anthropology, 2 Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Behavioural Biology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Vienna, Austria<br />

Primates living in multi male–female associati<strong>on</strong>s can develop SSC during sexual active periods. Size <str<strong>on</strong>g>of</str<strong>on</strong>g> SSC is<br />

dependent <strong>on</strong> fluctuating sex-steroids. Males intensify socio-sexual c<strong>on</strong>tact with females showing exaggerated<br />

SSC.<br />

We investigated the socio-endocrine impact <str<strong>on</strong>g>of</str<strong>on</strong>g> SSC in three different species: Facial and anogenital redness in<br />

Japanese macaques (JM) during the breeding and n<strong>on</strong>-breeding seas<strong>on</strong>, perineal swellings in Barbary<br />

macaques (BM) treated with the c<strong>on</strong>traceptive lev<strong>on</strong>orgestrel during n<strong>on</strong>-sexual periods, and perineal<br />

swellings in chimpanzees (Ch) during intact cycle periods. Data from JM and BM were collected under seminatural,<br />

from Ch under caged c<strong>on</strong>diti<strong>on</strong>s. Fecal samples were used <str<strong>on</strong>g>for</str<strong>on</strong>g> analyses <str<strong>on</strong>g>of</str<strong>on</strong>g> sex steroid and cortisol<br />

(CORT) metabolites.<br />

In BM, multiple regressi<strong>on</strong>s showed a negative relati<strong>on</strong>ship between SSC size and CORT and a positive <strong>on</strong>e<br />

between swelling size and intersexual socio-positive c<strong>on</strong>tact. Females with enlarged SSC had lower<br />

progester<strong>on</strong>e levels and increased estradiol-progester<strong>on</strong>e ratios.<br />

Ch females housed with males showed decreased CORT during late and decreasing tumescence <str<strong>on</strong>g>of</str<strong>on</strong>g> the SSC.<br />

In single housed individuals, increased CORT was related to decreased plasma FSH. Paired females had<br />

shorter cycle length.<br />

In JM, a light and a dark group could be discerned with regard to redness. In the dark group, intensity <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

redness increased during the breeding seas<strong>on</strong> and correlated with the amount <str<strong>on</strong>g>of</str<strong>on</strong>g> socio-sexual behavior. These<br />

females had significantly elevated CORT and sex steroid titers.<br />

In c<strong>on</strong>clusi<strong>on</strong>, reduced HPA activity in females is associated with exaggerated SSC and intensified male<br />

c<strong>on</strong>tact when male mating competiti<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> females is diminished (e.g., BM, Ch).<br />

459


23-26 August 2007,<br />

Budapest, Hungary<br />

460<br />

7I. WORK STRESS<br />

7I_01_P<br />

(poster secti<strong>on</strong> A2, poster board #152, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE IMPACT OF A SURVEY BASED WORKPLACE INTERVENTION PROGRAM ON<br />

EMPLOYEE HEALTH, BIOLOGICAL STRESS MARKERS, AND PRODUCTIVITY<br />

Ingrid L. Anderzén, Bengt B. Arnetz 1<br />

Uppsala University, Uppsala Sweden and 1 Uppsala University and Wayne State University, Detroit<br />

Increasing employee health and participati<strong>on</strong> have been identified as crucial to achieve productive<br />

organizati<strong>on</strong>s. However, there are few studies assessing methods to improve psychosocial working<br />

c<strong>on</strong>diti<strong>on</strong>s, employee health and well-being as well as productivity. This study assessed the impact <strong>on</strong> health,<br />

biological stress markers, absenteeism and productivity from a <strong>on</strong>e year structured interventi<strong>on</strong> program<br />

based <strong>on</strong> group-specific psychosocial results from 22 work units in five different <str<strong>on</strong>g>of</str<strong>on</strong>g>fices. White collar<br />

employees representing 22 different work units were assessed be<str<strong>on</strong>g>for</str<strong>on</strong>g>e, and <strong>on</strong>e year after, a <strong>on</strong>e-year<br />

interventi<strong>on</strong> program based <strong>on</strong> group-specific psychosocial work-related results. Self-rated questi<strong>on</strong>naire data<br />

(QWC), biological blood measurements and hard-core registry data <strong>on</strong> sickness absenteeism and productivity<br />

were analyzed at baseline and at the <strong>on</strong>e-year follow-up. Employee ratings <str<strong>on</strong>g>of</str<strong>on</strong>g> the psychosocial work<br />

envir<strong>on</strong>ment showed a significant statistical improvement in per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance feedback, participatory<br />

management, employeeship, skills development, efficiency, leadership, employee well-being, and work-related<br />

exhausti<strong>on</strong>, (p < 0.05). Restorative horm<strong>on</strong>es, such as serum testoster<strong>on</strong>e levels, increased during the<br />

interventi<strong>on</strong> (p < 0.01) and changed levels <str<strong>on</strong>g>of</str<strong>on</strong>g> serum testoster<strong>on</strong>e were significantly correlated to overall<br />

changes in organizati<strong>on</strong>al well-being during the year (r = .51, p


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

to 110. A score


23-26 August 2007,<br />

Budapest, Hungary<br />

7I_04_P<br />

(poster secti<strong>on</strong> A2, poster board #155, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

WORK-RELATED STRESS AMONG FEMALE STAFF IN CROATIAN HOSPITALS<br />

Rajna Golubic 1 , Bojana Knezevic 2 , Milan Milosevic 1 , Ana Rad<strong>on</strong>ic 2 , Marin Belak 3 ,<br />

Predrag Knezevic 4 , Jadranka Mustajbegovic 1<br />

1Department <str<strong>on</strong>g>for</str<strong>on</strong>g> Envir<strong>on</strong>mental and Occupati<strong>on</strong>al Health, "Andrija Stampar" School <str<strong>on</strong>g>of</str<strong>on</strong>g> Public Health,<br />

Rockefellerova 4, Zagreb, CROATIA<br />

2Clinical Hospital <str<strong>on</strong>g>Centre</str<strong>on</strong>g> Zagreb<br />

3Clinical Hospital Sisters <str<strong>on</strong>g>of</str<strong>on</strong>g> Mercy, Zagreb<br />

4Clinical Hospital Dubrava, Zagreb<br />

e-mail: rajna.golubic@gmail.com<br />

Aim: To determine the difference in stress percepti<strong>on</strong> am<strong>on</strong>g female staff at various hospital departments.<br />

Methods: 984 female employees in 4 Croatian hospitals an<strong>on</strong>ymously and voluntarily completed the<br />

Occupati<strong>on</strong>al Stress Assessment Questi<strong>on</strong>naire. According to the factor analysis the stressors were classified<br />

into 6 groups: Work management, Public criticism, Shift work, Pr<str<strong>on</strong>g>of</str<strong>on</strong>g>essi<strong>on</strong>al demands, Interpers<strong>on</strong>al<br />

communicati<strong>on</strong> and Pr<str<strong>on</strong>g>of</str<strong>on</strong>g>essi<strong>on</strong>al health hazards. The differences were analyzed by Mann Whitney U test and<br />

P


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

c<strong>on</strong>cerned <str<strong>on</strong>g>of</str<strong>on</strong>g> the changes and a c<strong>on</strong>trol group. The resp<strong>on</strong>se rate was 81% and 74% respectively at baseline<br />

and follow up measurements. Group differences were analyzed both <str<strong>on</strong>g>for</str<strong>on</strong>g> an open (n= 226) and closed cohort<br />

(n= 156) using <strong>on</strong>e-way ANOVA as well as a two-way ANOVA <str<strong>on</strong>g>for</str<strong>on</strong>g> repeated measurement in a closed<br />

cohort. To explain predictors <str<strong>on</strong>g>for</str<strong>on</strong>g> changed health a stepwise linear regressi<strong>on</strong> analysis was used. The results in<br />

the open cohort showed that the study group experienced significantly worse self-rated health and worse<br />

work satisfacti<strong>on</strong> after the reorganizati<strong>on</strong> compared with the c<strong>on</strong>trol group. The study group had also<br />

increased their level <str<strong>on</strong>g>of</str<strong>on</strong>g> work related exhausti<strong>on</strong>. The results from the closed cohort showed that the recovery<br />

horm<strong>on</strong>e DHEA-S had significantly decreased and sickness absence increased am<strong>on</strong>g employees in the study<br />

group compared to those in the c<strong>on</strong>trol group. Factors that best predicted changed health after the<br />

reorganizati<strong>on</strong> were work related exhausti<strong>on</strong>, age and coping ability. Sickness absence had increased significant <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

the study group at the 1-year follow up (7% and 2% respectively).<br />

7I_06_P<br />

(poster secti<strong>on</strong> A2, poster board #157, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STRESSOR TRAITS&STRESS-REDUCTION IN WORKING AND<br />

NON-WORKING MOTHERS<br />

Mohhamad Hatami<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Tehran, 14174-6619, Enghlab Eslami, Tehran, Iran<br />

e-mail: hatami@nyoir.org<br />

The present study aims to evaluate the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> stress-reducti<strong>on</strong> meichenbaum technique <strong>on</strong> working nurses,<br />

teachers and n<strong>on</strong>-working housewifes mothers according to mother–child stressor traits-Adaptability,<br />

Acceptability, Demandingness, Mood, Hyperactivity / Distractibility, Rein<str<strong>on</strong>g>for</str<strong>on</strong>g>ces, Depressi<strong>on</strong>, Attachment,<br />

Restricti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> role, Sense <str<strong>on</strong>g>of</str<strong>on</strong>g> competence, Social isolati<strong>on</strong> scale, Relati<strong>on</strong> ship with spouse, Parental health.<br />

First,228 working and n<strong>on</strong>-working mothers matched and they were evaluated using "parenting stress index"<br />

and "occcupati<strong>on</strong>al stress index". Sec<strong>on</strong>d ,24 mothers with high level stress shared in two matched groups <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

experimental and c<strong>on</strong>trol.experimental group were under treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> stress-reducti<strong>on</strong> technique <str<strong>on</strong>g>for</str<strong>on</strong>g> 13<br />

weeks but c<strong>on</strong>trol group didn,t received any treatment.the level <str<strong>on</strong>g>of</str<strong>on</strong>g> stress in both group was assessed be<str<strong>on</strong>g>for</str<strong>on</strong>g>e<br />

and after the treatment In order to analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> data,besides <str<strong>on</strong>g>of</str<strong>on</strong>g> the basic statistical methods. The result <str<strong>on</strong>g>of</str<strong>on</strong>g> this<br />

study showed that.<br />

(1)there was significant diference between stress am<strong>on</strong>g working mothers and n<strong>on</strong>-working mothers as well.<br />

(2)the nature <str<strong>on</strong>g>of</str<strong>on</strong>g> stressor in expanding <str<strong>on</strong>g>of</str<strong>on</strong>g> the level <str<strong>on</strong>g>of</str<strong>on</strong>g> stress in mothers had different<br />

effect,while,demandingness and relati<strong>on</strong>ship with spouse in nurses, demandingness and restricti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> role in<br />

teachers and demandingness and depressi<strong>on</strong> in n<strong>on</strong>-working mothers had more c<strong>on</strong>tributi<strong>on</strong> in these<br />

subjects.<br />

(3)the result <str<strong>on</strong>g>of</str<strong>on</strong>g> stress-reducti<strong>on</strong> technique was effective.<br />

In reducting <str<strong>on</strong>g>of</str<strong>on</strong>g> life, occupati<strong>on</strong>al and a few stressor traits.<br />

463


23-26 August 2007,<br />

Budapest, Hungary<br />

7I_07_P<br />

(poster secti<strong>on</strong> A2, poster board #158, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

WORK STRESS OF SOCIAL WORK MANAGERS IN TURKEY<br />

Vedat Isikhan<br />

Hacettepe University Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Social Work, 06 Kecioren, Anakara, Turkey<br />

e-mail: visikhan@hacettepe.edu.tr<br />

Management process <str<strong>on</strong>g>of</str<strong>on</strong>g> the instituti<strong>on</strong>s which <str<strong>on</strong>g>of</str<strong>on</strong>g>fer service to solve the problems <str<strong>on</strong>g>of</str<strong>on</strong>g> individuals and families<br />

may have some particular difficulties. Foremost am<strong>on</strong>g these are problems faced by the managers when they<br />

try to bring together the different type <str<strong>on</strong>g>of</str<strong>on</strong>g> pr<str<strong>on</strong>g>of</str<strong>on</strong>g>essi<strong>on</strong>als <strong>on</strong> behalf <str<strong>on</strong>g>of</str<strong>on</strong>g> individuals who have c<strong>on</strong>stant psychosocial<br />

and ec<strong>on</strong>omic problems . Managers are c<strong>on</strong>stantly experiencing stress when trying to deal with these<br />

problems.<br />

This study aims to determine the factors that influence work stress <str<strong>on</strong>g>of</str<strong>on</strong>g> social work managers in Turkey. The<br />

data have been collected by questi<strong>on</strong>naries sent to 554 social work managers by mail. For statistical analysis,<br />

questi<strong>on</strong>nary <str<strong>on</strong>g>for</str<strong>on</strong>g>ms <str<strong>on</strong>g>of</str<strong>on</strong>g> 320 managers were c<strong>on</strong>sidered to be eligible.<br />

The important results <str<strong>on</strong>g>of</str<strong>on</strong>g> the study are as follows: The great majority <str<strong>on</strong>g>of</str<strong>on</strong>g> managers have high level <str<strong>on</strong>g>of</str<strong>on</strong>g> work<br />

stress and Type A pers<strong>on</strong>ality. In work stress, marital status, job satisfacti<strong>on</strong>, overall durati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> work,<br />

administrative positi<strong>on</strong>, lack <str<strong>on</strong>g>of</str<strong>on</strong>g> aut<strong>on</strong>omy, lack <str<strong>on</strong>g>of</str<strong>on</strong>g> pers<strong>on</strong>al support from the colleagues and obligati<strong>on</strong> to<br />

attend many meetings and interviews are found to be influential factors (p


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

average age <str<strong>on</strong>g>of</str<strong>on</strong>g> male physicians is 43 years (26-63). Male physicians are more frequently clustered in surgery<br />

and radiology. Both groups <str<strong>on</strong>g>of</str<strong>on</strong>g> physicians reported: Work organizati<strong>on</strong>, Public criticism and law suits and<br />

Pr<str<strong>on</strong>g>of</str<strong>on</strong>g>essi<strong>on</strong>al demands as stressful or very stressful. Shift work is significantly more frequently (P=0.003)<br />

reported by male doctors vs. female doctors. C<strong>on</strong>clusi<strong>on</strong>: Both groups, male and female physicians, in<br />

hospitals are experiencing high level <str<strong>on</strong>g>of</str<strong>on</strong>g> stress. Male physicians c<strong>on</strong>sidered shift work more stressful than<br />

females, because they are clustered in more demanding specialties.<br />

7I_09_P<br />

(poster secti<strong>on</strong> A2, poster board #160, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HEART RATE VARIABILITY IN PHYSICIANS WORKING ON NIGHT CALL<br />

B. Malmberg 1 , R. Perss<strong>on</strong> 2 , P. Ørbaek 2<br />

1 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Occupati<strong>on</strong>al and Envir<strong>on</strong>mental Medicine, Lund University Hospital, Sweden<br />

2 Nati<strong>on</strong>al <str<strong>on</strong>g>Research</str<strong>on</strong>g> <str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g> the Working Envir<strong>on</strong>ment, Copenhagen, Denmark<br />

e-mail: Birgitta.Malmberg@med.lu.se<br />

Shiftwork and stress have negative effects <strong>on</strong> per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance, well-being and sleep, and are associated with an<br />

increased risk <str<strong>on</strong>g>for</str<strong>on</strong>g> ischaemic heart disease and metabolic disturbances. Physicians <str<strong>on</strong>g>of</str<strong>on</strong>g>ten work in shifts, but<br />

little is known about possible physiological effects <str<strong>on</strong>g>of</str<strong>on</strong>g> night call duty. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to investigate<br />

the impact <str<strong>on</strong>g>of</str<strong>on</strong>g> physicians’ night call duty <strong>on</strong> aut<strong>on</strong>omic balance, measured by heart rate variability (HRV), and<br />

to determine whether there were differences between physician specialities. For this reas<strong>on</strong> 19<br />

anaesthesiologists were compared with 17 paediatricians and ENT surge<strong>on</strong>s. M<strong>on</strong>itoring by digital holter<br />

electrocardiogram was made during night call, the recovery period after night call, and <strong>on</strong> an ordinary<br />

workday. HRV analyses were made in 24 hour periods <str<strong>on</strong>g>for</str<strong>on</strong>g> SDNN (time domain measurement <str<strong>on</strong>g>of</str<strong>on</strong>g> total<br />

variability), and in 10 min strips during 1 h at 21:00-22:00 <str<strong>on</strong>g>for</str<strong>on</strong>g> HFn (frequency domain variable, reflecting<br />

vagal influence <strong>on</strong> HRV). Statistically significant effects <str<strong>on</strong>g>of</str<strong>on</strong>g> day in HFnu were seen <strong>on</strong>ly am<strong>on</strong>g the<br />

anaesthesiologists, who had 21% lower levels during night call compared with day work, and 38% lower<br />

compared with post night call (P


23-26 August 2007,<br />

Budapest, Hungary<br />

466<br />

7I_10_P<br />

(poster secti<strong>on</strong> A2, poster board #161, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DIFFERENCES IN STRESS PERCEPTION BETWEEN PHYSICIANS IN SURGICAL<br />

AND NON-SURGICAL SPECIALTIES<br />

Milan Milosevic 1 , Bojana Knezevic 2 , Rajna Golubic 1 , Jadranka Mustajbegovic 1 , Lana Matec 3 ,<br />

Maja Debeljak 3<br />

1Department <str<strong>on</strong>g>for</str<strong>on</strong>g> Envir<strong>on</strong>mental and Occupati<strong>on</strong>al Health, "Andrija Stampar" School <str<strong>on</strong>g>of</str<strong>on</strong>g> Public Health,<br />

Rockefellerova 4, Zagreb, CROATIA;<br />

e-mail: milan.milosevic@mef.hr<br />

2Clinical Hospital <str<strong>on</strong>g>Centre</str<strong>on</strong>g> Zagreb, 3 Medical School University <str<strong>on</strong>g>of</str<strong>on</strong>g> Zagreb<br />

Aim: To investigate differences in stress percepti<strong>on</strong> between physicians in surgical and n<strong>on</strong>-surgical<br />

specialties. Subjects and Methods: 212 physicians in surgical and 158 in n<strong>on</strong>-surgical specialties an<strong>on</strong>ymously<br />

and voluntarily completed the Occupati<strong>on</strong>al Stress Assessment Questi<strong>on</strong>naire in four Croatian clinics.<br />

Questi<strong>on</strong>s regarding stress were graded by the Likert Scale from 1 to 5 and due to factor analysis classified in<br />

six items: Work management, Public criticism, Shift work, Pr<str<strong>on</strong>g>of</str<strong>on</strong>g>essi<strong>on</strong>al demands, Interpers<strong>on</strong>al relati<strong>on</strong>ship<br />

at workplace and Pr<str<strong>on</strong>g>of</str<strong>on</strong>g>essi<strong>on</strong>al health hazards. Items were used as dependent variables in multivariate analysis<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> variance. Results: Significant difference was found between surgical and n<strong>on</strong>-surgical specialties <strong>on</strong> the<br />

combined dependent variables: F (6, 363)=11.6, P


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

changed after night shift work (264.3 +/- 29.6 to 259.0 +/- 15.2). Plasma c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> adrenaline were<br />

increased after both works <str<strong>on</strong>g>of</str<strong>on</strong>g> daytime shift (31.5 +/- 3.0 to 48.3 +/- 7.1 pg/ml) and night shift (34.7 +/- 6.4<br />

to 40.8 +/- 5.0). Total amounts <str<strong>on</strong>g>of</str<strong>on</strong>g> urinary secreti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> dopamine, noradrenaline and adrenaline were<br />

significantly lower in nurses <str<strong>on</strong>g>of</str<strong>on</strong>g> night shift work than those in nurses <str<strong>on</strong>g>of</str<strong>on</strong>g> daytime shift work, respectively.<br />

These results suggest the possibility that working at night would affect central nervous system and adrenal<br />

functi<strong>on</strong>, and intensifies metabolic clearance rate <str<strong>on</strong>g>of</str<strong>on</strong>g> catecholamine.<br />

7I_12_P<br />

(poster secti<strong>on</strong> A2, poster board #163, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DAILY LIFE STRESS AND MENTAL HEALTH IN HOSPITAL EMPLOYEES<br />

Yoshio Ohtani 1 , Eiko Shimada 2 , Makihiko Suzuki 3<br />

1 Kitasato University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Allied Health Sciences; Sagamihara-Shi, Kanagawa, Japan;<br />

e-mail: y-ohtani@kitasato-u.ac.jp<br />

2 Knanagawa Prefectural Center <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychiatry, 3 Kitasato University College <str<strong>on</strong>g>of</str<strong>on</strong>g> Liberal Arts and Sciences<br />

Introducti<strong>on</strong>: Workers in a medical facilities may be greatly affected in their mental and/or somatic<br />

c<strong>on</strong>diti<strong>on</strong>s, although they labor to help patients with various diseases. In this study we investigated how daily<br />

life stresses in the hospital employees influence <strong>on</strong> their mental health, sleep and several phases <str<strong>on</strong>g>of</str<strong>on</strong>g> anger.<br />

Methods: 1Subjects: One hundred and <str<strong>on</strong>g>for</str<strong>on</strong>g>ty-eight emloyees in number <str<strong>on</strong>g>of</str<strong>on</strong>g> the Hospital in a big city <str<strong>on</strong>g>of</str<strong>on</strong>g> Japan<br />

(Male 39, Female 80, Unclear 29; 46.2 years old <str<strong>on</strong>g>of</str<strong>on</strong>g> mean age) c<strong>on</strong>sist <str<strong>on</strong>g>of</str<strong>on</strong>g> 18 in Medical, 94 in Nursing, 17 in<br />

Secretary Divisi<strong>on</strong> and Nuclear 19.<br />

2) Materials <str<strong>on</strong>g>for</str<strong>on</strong>g> the assessment <str<strong>on</strong>g>of</str<strong>on</strong>g> mental and/or psychological c<strong>on</strong>diti<strong>on</strong>s<br />

(1) Daily Life Stress Scale (Munakata et al., 1985), (2) Genral Health Questi<strong>on</strong>aire (GHQ-60), (3) Athens<br />

Insomnia Scale, (4) State-Trait Anger Expressi<strong>on</strong> Inventory<br />

3) Statistical analysys: Multiple comparis<strong>on</strong> by B<strong>on</strong>fer<strong>on</strong>i method using the score <str<strong>on</strong>g>of</str<strong>on</strong>g> Daily Life Stress Scale in<br />

4 step groups [Weak(W);0-4 points, Moderate(M); 5-9, Rather Str<strong>on</strong>g(RS);10-18. Str<strong>on</strong>g(S);19] as an<br />

independent variable and the total points <str<strong>on</strong>g>of</str<strong>on</strong>g> GHQ, Insomnia scale and 5 phases <str<strong>on</strong>g>of</str<strong>on</strong>g> STAXI[State(STX-<br />

S),Trait(TX-T,Anger Expressi<strong>on</strong>-Cnotrol (AE-C)AE-Out (AE-O)AE-In (AE-I)] as dependent<br />

variables.<br />

Results and Discussi<strong>on</strong>s: In the total score <str<strong>on</strong>g>of</str<strong>on</strong>g> GHQ, the points <str<strong>on</strong>g>of</str<strong>on</strong>g> S group was significantly higher than those<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> W, M and RS, as well as RS group’s was higher than W group’sp


23-26 August 2007,<br />

Budapest, Hungary<br />

7I_13_P<br />

(poster secti<strong>on</strong> A2, poster board #164, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

JOB STRAIN AND CARDIOVASCULAR RISK FACTORS IN PHYSICIANS<br />

Laura Poanta 1 , Mihai Ober 2 , Anca Farcas 2<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Internal Medicine II, 2 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Internal Medicine I, Clinicilor 2-4 street,<br />

Cluj-Napoca, Romania<br />

e-mail: laurapoanta@yahoo.com<br />

Background: with the development <str<strong>on</strong>g>of</str<strong>on</strong>g> modern civilizati<strong>on</strong>, occupati<strong>on</strong>al stress becomes increasingly<br />

important. Physicians are an exposed category and they are pr<strong>on</strong>e to develop stress-related disease and risk<br />

behaviors.<br />

Methods: we c<strong>on</strong>duct a study am<strong>on</strong>g 118 physicians, 78 women and 40 men. The Romanian versi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Job<br />

C<strong>on</strong>tent Questi<strong>on</strong>naire (JCQ), was mailed to all the subjects. We investigate the main scales <str<strong>on</strong>g>of</str<strong>on</strong>g> JCQ: decisi<strong>on</strong><br />

latitude (DL), psychological job demands (PJD) and social support (SoS), al<strong>on</strong>g with the prevalence <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

smoking, sedentary behavior, cholesterol, triglycerides, fibrinogen, CRP and plasma cortisol.<br />

Results: The final resp<strong>on</strong>se rate was 112 (94.9%), 72 (64.2 %) women and 40 (35.7 %) men aged between 30<br />

and 60. Results are shown in the table.<br />

Smoking (32% <str<strong>on</strong>g>of</str<strong>on</strong>g> women and 56% <str<strong>on</strong>g>of</str<strong>on</strong>g> men) and sedentary behavior (over 60 % both in women and men)<br />

positively correlated with PJD and negatively with SoS. Total cholesterol positively correlated with PJD in<br />

both women (r=0.33) and men (r=0.51), and negatively with coworker support and depressi<strong>on</strong> (r=-0.21, r=-<br />

0.28 in women, and r=-0.57, r=-0.19 in men, respectively). We didn’t find significant correlati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> those<br />

items with proinflammatory state, but we did find interesting correlati<strong>on</strong>s with plasma cortisol (e.g. negative<br />

correlati<strong>on</strong> with coworker support both in men and women, P


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

study was designed to evaluate an integrative approach (IA) that aimed to address the different individual<br />

needs and allowed the individual to use his/her own creative powers <str<strong>on</strong>g>of</str<strong>on</strong>g> visualizati<strong>on</strong> to overcome public<br />

speaking anxiety at work. 150 public speaking anxious employees were randomly assigned to two treatments<br />

and a c<strong>on</strong>trol group. Measurements included self report and physiological measures. Physiological measures<br />

(skin temperature, galvanic skin resp<strong>on</strong>se and heart rate) were taken be<str<strong>on</strong>g>for</str<strong>on</strong>g>e and after treatment. Results<br />

indicated that the IA approach produced significantly lower self report measures <str<strong>on</strong>g>of</str<strong>on</strong>g> stress than either the<br />

c<strong>on</strong>trol group or the PMR group. All treatment groups showed a significant lowering <str<strong>on</strong>g>of</str<strong>on</strong>g> psysiological<br />

measures from pre-test to post-test.<br />

7I_15_P<br />

(poster secti<strong>on</strong> A2, poster board #165, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

OCCUPATIONAL STRESSORS AND ITS ORGANIZATIONAL AND INDIVIDUAL<br />

CORRELATES: A NATIONWIDE STUDY OF NORWEGIAN<br />

AMBULANCE PERSONNEL<br />

Tom Sterud a , Erlend Hem a , Øivind Ekeberg a , Bjørn Lau b<br />

aDepartment <str<strong>on</strong>g>of</str<strong>on</strong>g> Behavioural Sciences in Medicine, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Basic Medical Sciences, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine,<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Oslo, Norway<br />

bNati<strong>on</strong>al Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Occupati<strong>on</strong>al Health, Oslo, Norway<br />

e-mail: tom.sterud@medisin.uio.no<br />

Objectives: We compared the severity and frequency level <str<strong>on</strong>g>of</str<strong>on</strong>g> organizati<strong>on</strong>al and ambulance-specific stressors,<br />

and studied their relati<strong>on</strong>ship to structural c<strong>on</strong>diti<strong>on</strong>s and individual differences in a nati<strong>on</strong>wide sample <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

operati<strong>on</strong>al ambulance pers<strong>on</strong>nel (n = 1180). Methods: The questi<strong>on</strong>naire included the Job Stress Survey, the<br />

Norwegian Ambulance Stress Survey, the Basic Character Inventory, General Self-Efficacy Scale, and<br />

questi<strong>on</strong>s addressing structural work c<strong>on</strong>diti<strong>on</strong>s. Results: Serious operati<strong>on</strong>al tasks and physical demands were<br />

identified as the two most severe stressors. Lack <str<strong>on</strong>g>of</str<strong>on</strong>g> support from co-workers was the most severe and<br />

frequent organizati<strong>on</strong>al stressor. Higher frequency <str<strong>on</strong>g>of</str<strong>on</strong>g> stressors was most str<strong>on</strong>gly associated with size <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

service districts (β ranging between .18 and .30, p < .01) and working overtime (β ranging from .13 to .27,<br />

p < .05). Neuroticism (β ranging from .09 to .17, p < .01) and low general self-efficacy (β ranging from –.12<br />

to –.16, p < .001) were equally str<strong>on</strong>gly related to severity <str<strong>on</strong>g>of</str<strong>on</strong>g> stressors, as were structural c<strong>on</strong>diti<strong>on</strong>s.<br />

C<strong>on</strong>clusi<strong>on</strong>s: Ambulance-specific stressors were reported as both more severe and more frequently occurring<br />

stressors than were organizati<strong>on</strong>al stressors. Structural working c<strong>on</strong>diti<strong>on</strong>s were more str<strong>on</strong>gly related to<br />

frequency <str<strong>on</strong>g>of</str<strong>on</strong>g> job stressors, but higher levels <str<strong>on</strong>g>of</str<strong>on</strong>g> neuroticism, and lower levels <str<strong>on</strong>g>of</str<strong>on</strong>g> self-efficacy were equally<br />

str<strong>on</strong>gly related to stressor severity, as was structural working c<strong>on</strong>diti<strong>on</strong>s.<br />

469


23-26 August 2007,<br />

Budapest, Hungary<br />

7I_16_P<br />

(poster secti<strong>on</strong> A2, poster board #166, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

WORKPLACE STRESS AMONG HEALTH CARE WORKERS<br />

Nóra Szabó 1,2 , Katalin Hegedűs 2 , Gábor Szabó 2 , Krisztina D. Neculai 2 , Gyöngyvér Salavecz 2<br />

1Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Basic and C<strong>on</strong>tinuing Educati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Health Workers, Budapest, Hungary<br />

2Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Behavioral Sciences, Semmelweis Universit,y, Budapest, Hungary<br />

1089, Budapest, Nagyvárad tér 4, Hungary<br />

e-mail: enoraszabo@freemail.hu<br />

Antecedents: Work <str<strong>on</strong>g>of</str<strong>on</strong>g> nurses is extremely stressful and is combined with low pr<str<strong>on</strong>g>of</str<strong>on</strong>g>essi<strong>on</strong>al and social<br />

acknowledgement, but is there any difference in terms <str<strong>on</strong>g>of</str<strong>on</strong>g> workplace stress between two groups <str<strong>on</strong>g>of</str<strong>on</strong>g> nurses in<br />

similar positi<strong>on</strong> Aim and hypothesis: The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> the survey is to investigate comparatively hospice nurses<br />

and nurses caring <str<strong>on</strong>g>for</str<strong>on</strong>g> elderly patients. Hypothesis: due to interdisciplinary approach hospice nurses are in a<br />

more favourable situati<strong>on</strong> than nurses caring <str<strong>on</strong>g>for</str<strong>on</strong>g> elderly patients in terms <str<strong>on</strong>g>of</str<strong>on</strong>g> degree <str<strong>on</strong>g>of</str<strong>on</strong>g> workplace stress and<br />

social support. Methods: A cross-secti<strong>on</strong>al study was per<str<strong>on</strong>g>for</str<strong>on</strong>g>med am<strong>on</strong>g hospice nurses (N=25) and nurses<br />

caring <str<strong>on</strong>g>for</str<strong>on</strong>g> elderly patients (N=50) using a test battery comprising the Brief Stress and Coping Inventory<br />

(Rahe, Tolles, 2002), the Vital Exhausti<strong>on</strong> Questi<strong>on</strong>naire (Appels, Mulder, 1988), the Support Dimensi<strong>on</strong><br />

Scale (Caldwell, 1987), and the Ef<str<strong>on</strong>g>for</str<strong>on</strong>g>t-Reward Imbalance Questi<strong>on</strong>naire (Siegrist, 1996). Expansi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

sample is being in progress with data <str<strong>on</strong>g>of</str<strong>on</strong>g> Hungarostudy Epidemiological Panel. Results: In terms <str<strong>on</strong>g>of</str<strong>on</strong>g> social<br />

support hospice nurses are in a more favourable positi<strong>on</strong> (P=0.048). Workplace stress is significantly higher<br />

in nurses caring <str<strong>on</strong>g>for</str<strong>on</strong>g> elderly patients than in hospice nurses (P=0,035 <str<strong>on</strong>g>for</str<strong>on</strong>g> overcommitment scale, P=0.034 <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

extrinsic ef<str<strong>on</strong>g>for</str<strong>on</strong>g>t scale). C<strong>on</strong>clusi<strong>on</strong>s: Interdisciplinary approach <str<strong>on</strong>g>of</str<strong>on</strong>g> hospice may promote nurses’ acceptance<br />

and appreciati<strong>on</strong>, whereas higher social support may reduce nurses’ vital exhausti<strong>on</strong> and the degree <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

workplace stress. This model might be applicable <str<strong>on</strong>g>for</str<strong>on</strong>g> other groups <str<strong>on</strong>g>of</str<strong>on</strong>g> health care staff as well.<br />

470<br />

7I_17_P<br />

(poster secti<strong>on</strong> A2, poster board #167, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STAFF STRESS IN BATHING RESIDENTS IN JAPANESE AGED CARE FACILITIES<br />

Edit Nagy-Tanaka, Mio Ito, Ryutaro Takahashi<br />

Tokyo Metropolitan Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Ger<strong>on</strong>tology, 35-2 Sakae-cho, Tokyo 173-0015 Japan<br />

e-mail: edit@tmig.or.jp<br />

Bathing residents in l<strong>on</strong>g-term aged care facilities, especially residents with dementia, is a stressful task.<br />

Bathing residents in a bathtub, usually twice a week, which is a comm<strong>on</strong> practice in Japan, is particularly<br />

demanding. The purpose <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to examine the reas<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> staff stress in the bathing situati<strong>on</strong> in<br />

l<strong>on</strong>g-term aged care facilities in Japan. Three large-scale (LS) settings and 3 small-scale (SS) settings were<br />

studied. The methods used were observati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the bathing routines <str<strong>on</strong>g>of</str<strong>on</strong>g> 21 staff (19 LS staff and 2 SS staff)<br />

and a questi<strong>on</strong>naire survey to 80 staff (61 LS staff and 19 SS staff). Data analysis was d<strong>on</strong>e by descriptive<br />

analysis and c<strong>on</strong>tent analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> free answers. The observati<strong>on</strong> results revealed a stressful atmosphere in the<br />

LS settings, with noisy envir<strong>on</strong>ment, hurried pace <str<strong>on</strong>g>of</str<strong>on</strong>g> work and c<strong>on</strong>fusi<strong>on</strong> about task divisi<strong>on</strong>. The<br />

questi<strong>on</strong>naire ratings showed that the LS staff were significantly more dissatisfied with most aspects <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

bathing, especially with the number <str<strong>on</strong>g>of</str<strong>on</strong>g> residents to be bathed a day and bathing time <str<strong>on</strong>g>for</str<strong>on</strong>g> each resident.<br />

According to preliminary results <str<strong>on</strong>g>of</str<strong>on</strong>g> the free answers, the LS staff felt task-oriented stress, such as large<br />

number <str<strong>on</strong>g>of</str<strong>on</strong>g> residents, time c<strong>on</strong>straints and the resp<strong>on</strong>sibility to prevent accidents, while the SS staff


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

menti<strong>on</strong>ed pers<strong>on</strong>-oriented stress, like persuading demented residents to bathe and handling reluctant and<br />

violent residents. Nursing implicati<strong>on</strong>s and stress coping methods will be discussed.<br />

7I_18_P<br />

(poster secti<strong>on</strong> A2, poster board #168, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STRESSFUL LIFE EVENTS AND OCCUPATIONAL ACCIDENTS<br />

Ricardo Cordeiro 1 , Adriano Dias 2<br />

1Campinas State University, School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Sciences, e-mail: ricacordeiro@gmail.com<br />

2Paulista State University, Botucatu School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, e-mail: adias@fmb.unesp.br<br />

Objective: The purpose <str<strong>on</strong>g>of</str<strong>on</strong>g> this study war to examine the associati<strong>on</strong> between stressful life events and<br />

occupati<strong>on</strong>al accidents.<br />

Methods: This was a populati<strong>on</strong>-based case-c<strong>on</strong>trol study, carried out in the city <str<strong>on</strong>g>of</str<strong>on</strong>g> Botucatu, in southeast<br />

Brazil. The cases c<strong>on</strong>sisted <str<strong>on</strong>g>of</str<strong>on</strong>g> 108 workers who had recently experienced occupati<strong>on</strong>al accidents. Each case<br />

was matched with three c<strong>on</strong>trols. The cases and c<strong>on</strong>trols answered a questi<strong>on</strong>naire about recent exposure to<br />

stressful life events.<br />

Results: Reporting <str<strong>on</strong>g>of</str<strong>on</strong>g> “envir<strong>on</strong>mental problems”, “being a victim <str<strong>on</strong>g>of</str<strong>on</strong>g> assault”, “not having enough food at<br />

home” and “n<strong>on</strong>occupati<strong>on</strong>al fatigue” were found to be risk factors <str<strong>on</strong>g>for</str<strong>on</strong>g> work related accidents with estimated<br />

incidence rate ratios <str<strong>on</strong>g>of</str<strong>on</strong>g> 1.4 [95% c<strong>on</strong>fidence interval (95% CI) 1.1-1.7], 1.3 (955 CI 1.1-1.7), 1.3 (95% ci 1.1-<br />

1.6), and 1.4 (95% ci 1.2-1.7), respectively.<br />

C<strong>on</strong>clusi<strong>on</strong>: The findings <str<strong>on</strong>g>of</str<strong>on</strong>g> the study suggested that n<strong>on</strong>work variables c<strong>on</strong>tribute to occupati<strong>on</strong>al accidents,<br />

thus broadening the understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> these phenomena, which can support new approaches to the<br />

preventi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> occupati<strong>on</strong>al accidents.<br />

7I_19_P<br />

(poster secti<strong>on</strong> A2, poster board #169, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFORT-REWARD IMBALANCE AND MENSTRUAL PAIN: RESULTS FROM THE<br />

HUNGAROSTUDY EPIDEMIOLOGICAL PANEL 2006<br />

Krisztina Neculai, Gyöngyvér Salavecz, Szilvia Ádám, Zsuzsa Győrffy, Nóra Szabó, Mária Kopp<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Behavioural Sciences, Semmelweis University, 1089, Budapest, Nagyvárad tér 4, Hungary<br />

e-mail: neckri@net.sote.hu<br />

Background: Painful menstruati<strong>on</strong> is c<strong>on</strong>sidered to be the most comm<strong>on</strong> gynaecological disorder. While<br />

compelling research has been c<strong>on</strong>ducted with respect to its biological determinants, little is known about the<br />

psychosocial factors, including work-related stress, which might influence menstrual pain. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, the<br />

objective <str<strong>on</strong>g>of</str<strong>on</strong>g> the present study was to analyse the relati<strong>on</strong>ship between work stress, defined by means <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

Ef<str<strong>on</strong>g>for</str<strong>on</strong>g>t-Reward Imbalance Model, and menstrual pain. Methods: Self-reported data <str<strong>on</strong>g>of</str<strong>on</strong>g> 1174 working women,<br />

participants in the Hungarostudy Epidemiological Panel 2006 was analysed. Post-menopausal (n=165) and<br />

pregnant women (n=8) and those with missing data (n=215) were excluded from the analyses, resulting in<br />

786 subjects. Binary logistic regressi<strong>on</strong> was used to determine the associati<strong>on</strong> between ef<str<strong>on</strong>g>for</str<strong>on</strong>g>t-reward<br />

imbalance, overcommitment and painful menstruati<strong>on</strong>. Results: Altogether 19% <str<strong>on</strong>g>of</str<strong>on</strong>g> women reported to<br />

experience menstrual pain that limits their daily activity. After c<strong>on</strong>trolling <str<strong>on</strong>g>for</str<strong>on</strong>g> the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> age, educati<strong>on</strong>al<br />

attainment, parity status, smoking and body-mass index ef<str<strong>on</strong>g>for</str<strong>on</strong>g>t-reward imbalance was found to be associated<br />

with a higher risk <str<strong>on</strong>g>for</str<strong>on</strong>g> menstrual pain (OR (95% CI) = 1.39 (1.04-1.87)). Overcommitment, the other<br />

471


23-26 August 2007,<br />

Budapest, Hungary<br />

comp<strong>on</strong>ent <str<strong>on</strong>g>of</str<strong>on</strong>g> the Ef<str<strong>on</strong>g>for</str<strong>on</strong>g>t-Reward Imbalance Model was not related to dysmenorrhea (OR (95% CI) = 1.01<br />

(0.93-1.10)). C<strong>on</strong>clusi<strong>on</strong>s: An imbalance between ef<str<strong>on</strong>g>for</str<strong>on</strong>g>ts and rewards at work is associated with<br />

dysmenorrhea. The relati<strong>on</strong>ship between work-related psychosocial factors and painful menstruati<strong>on</strong> needs to<br />

be further investigated to determine causality and factors that may explain this relati<strong>on</strong>ship.<br />

7I_20_P<br />

(poster secti<strong>on</strong> A2, poster board #170, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

OCCUPATIONAL STRESS AMONG DIFFERENT STUDENTS FROM<br />

OUR UNIVERSITY<br />

Liana Mos, Delia Podea, Ram<strong>on</strong>a Chenderes<br />

Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine Western University "Vasile Goldis" from Arad, City Clinical Hospital,<br />

Departament <str<strong>on</strong>g>of</str<strong>on</strong>g> Internal Medicine, 2900 Piata Mihai viteazul 7-8, Arad, Romania<br />

e-mail: limmos2001@yahoo.com<br />

Aims This study is the first step <str<strong>on</strong>g>of</str<strong>on</strong>g> evaluating the stress am<strong>on</strong>g the students from our university. This study<br />

examined occupati<strong>on</strong>al stress am<strong>on</strong>g students from our University.<br />

Materials and method The sample c<strong>on</strong>sisted <str<strong>on</strong>g>of</str<strong>on</strong>g> 186 students from various specialties from Arad. The<br />

variables evaluated included stress situati<strong>on</strong>s at school and some social indicators. A questi<strong>on</strong>naire was given<br />

to the target populati<strong>on</strong> O.S.I. Occupati<strong>on</strong>al Stress Indicator. The final questi<strong>on</strong>naire comprised the following<br />

aspects: anxiety, socio demographic aspects, psycho physiological symptoms, and satisfacti<strong>on</strong>.<br />

Results. The difference between the scours is signifiant in the group <str<strong>on</strong>g>of</str<strong>on</strong>g> medicine students compare to others<br />

specialties, with a hy incidence <str<strong>on</strong>g>of</str<strong>on</strong>g> psycho physiological symptoms (physical suprasolicitati<strong>on</strong>) . Our results<br />

showed specific associati<strong>on</strong>s between situati<strong>on</strong>al stressors, socio demographic aspects state and satisfacti<strong>on</strong>.<br />

In particular, frustrati<strong>on</strong> in carrying out their work is linked to high levels <str<strong>on</strong>g>of</str<strong>on</strong>g> self-reported symptom.<br />

C<strong>on</strong>clusi<strong>on</strong>s<br />

Self-efficacy moderates the stress-strain relati<strong>on</strong>ship in general, in the sense that low levels <str<strong>on</strong>g>of</str<strong>on</strong>g> self-efficacy are<br />

related to high levels <str<strong>on</strong>g>of</str<strong>on</strong>g> occupati<strong>on</strong>al stress am<strong>on</strong>g students from medical faculty.<br />

472


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7J. STRESS AND COLLECTIVE BEHAVIOR:<br />

OUTBREAKS OF PANIC, WAR, MARKET CRASH<br />

7J_01_P<br />

(poster secti<strong>on</strong> A2, poster board #171, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE ROLE OF PERSONAL RESILIENCE AND COGNITIVE APPRAISAL<br />

IN UNDERGRADUATES’ COPING WITH DAILY HASSLES<br />

B. M. S. Bruneau<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Greenwich, L<strong>on</strong>d<strong>on</strong>, United Kingdom<br />

e-mail: B.S.Bruneau@gre.ac.uk<br />

The pers<strong>on</strong>al resources a pers<strong>on</strong> brings to a situati<strong>on</strong> determine how an event is appraised (Lazarus &<br />

Folkman, 1984; Frydenberg, 2002; Dev<strong>on</strong>port & Lane, 2006). The role <str<strong>on</strong>g>of</str<strong>on</strong>g> the process in how undergraduates<br />

manage stress has yet to be established. The present study hypothesised that when undergraduates cope with<br />

daily hassles their cognitive appraisal is mediated by their resilience (optimism, self-esteem, and perceived<br />

c<strong>on</strong>trol) levels. Undergraduates are a relevant study group as there are c<strong>on</strong>cerns that their health is more at<br />

risk than that <str<strong>on</strong>g>of</str<strong>on</strong>g> other people, after c<strong>on</strong>trolling <str<strong>on</strong>g>for</str<strong>on</strong>g> age and sex (Stewart-Brown et al., 2000). The present<br />

l<strong>on</strong>gitudinal study involved 511 undergraduates across two UK universities who completed a questi<strong>on</strong>naire at<br />

four periods during the first fourteen m<strong>on</strong>ths <str<strong>on</strong>g>of</str<strong>on</strong>g> their degree programme. The questi<strong>on</strong>naire comprised the<br />

Student-Life Stress Inventory (SLSI), Coping Operati<strong>on</strong>s Preference Enquiry (COPE), Resilience Indicator<br />

(RI) and the General Health Questi<strong>on</strong>naire (GHQ-12). Undergraduates’ mean stress score was repeatedly at<br />

around 2.45, <strong>on</strong> a scale <str<strong>on</strong>g>of</str<strong>on</strong>g> 1 to 5, where 5 represents extreme stress. High self-esteem and the seeking <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

instrumental support were the <strong>on</strong>ly variables which correlated significantly at each time period (r(373)=.11,<br />

p


23-26 August 2007,<br />

Budapest, Hungary<br />

7K. STRESS AND GENDER<br />

7K_01_P<br />

(poster secti<strong>on</strong> A2, poster board #172, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DIFFERENCES BETWEEN MEN AND WOMEN IN MENTAL AND PHYSICAL<br />

HEALTH IN THE RURAL POPULATION<br />

Marta Evelia Aparicio García, Mª Pilar Sánchez-López, Violeta Cardenal Herráiz,<br />

Virginia Dresch<br />

Universidad Complutense de Madrid<br />

In this research project differences have been analysed in physical and psychological health between men and<br />

women in the Spanish rural populati<strong>on</strong>, taking into account the work situati<strong>on</strong> (workers, unemployed or<br />

housewives), psychological variables (anxiety, self-esteem and satisfacti<strong>on</strong>) and physical variables (substance<br />

abuse, number <str<strong>on</strong>g>of</str<strong>on</strong>g> ailments and self-perceived state <str<strong>on</strong>g>of</str<strong>on</strong>g> health). 1000 subjects have taken part, representing the<br />

Spanish rural populati<strong>on</strong> and bel<strong>on</strong>ging to the five labour categories studied. The findings show that women<br />

have worse health than men, since they score higher <strong>on</strong> variables such as Anxiety or Ailments). Depending <strong>on</strong><br />

the labour situati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the participants, unemployed men prove to be the healthiest and unemployed women<br />

and housewives the least healthy.<br />

7K_03_P<br />

(poster secti<strong>on</strong> A2, poster board #173, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EXTRINSIC LIFE GOAL ORIENTATION IS ASSOCIATED WITH MARITAL STRESS<br />

FOR WOMEN BUT NOT FOR MEN - EVIDENCE FROM THE HUNGAROSTUDY<br />

EPIDEMIOLOGICAL PANEL 2006<br />

Tamas Martos, Piroska Balog, Maria S. Kopp<br />

Semmelweis University, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Behavioural Sciences, 1089, Budapest, Nagyvarad ter 4.<br />

e-mail: martos.tamas@btk.ppke.hu<br />

Marital stress has been proved to be an important factor in predicting various physical and mental health<br />

outcomes, including cardiovascular morbidity, depressi<strong>on</strong> etc. Motivati<strong>on</strong>al factors, like life goals and<br />

aspirati<strong>on</strong>s were also associated with health outcomes. Pursuing intrinsic goals (e.g. pers<strong>on</strong>al growth and good<br />

relati<strong>on</strong>ships) was shown to be associated with improved psychological functi<strong>on</strong>ing and better health<br />

outcomes while extrinsic aspirati<strong>on</strong>s (financial success and good appearance) were not. C<strong>on</strong>necti<strong>on</strong>s between<br />

life goals and marital stress have not been studied be<str<strong>on</strong>g>for</str<strong>on</strong>g>e. The study is based <strong>on</strong> Hungarostudy<br />

Epidemiological Panel 2006 (N=4528). We analysed a subsample <str<strong>on</strong>g>of</str<strong>on</strong>g> married/cohabiting people (male<br />

N=1550, female N=1875) assessing marital stress (MS, via Shortened Marital Stress Scale) and life goal<br />

orientati<strong>on</strong> (relative importance <str<strong>on</strong>g>of</str<strong>on</strong>g> extrinsic vs. intrinsic life goals, via Aspirati<strong>on</strong> Index). 167 men (10,8%)<br />

and 267 women (14,2%) showed elevated level <str<strong>on</strong>g>of</str<strong>on</strong>g> MS. Hierarchical logistic regressi<strong>on</strong> analyses were<br />

per<str<strong>on</strong>g>for</str<strong>on</strong>g>med, and odds ratios <str<strong>on</strong>g>for</str<strong>on</strong>g> marital stress (with 95% CI) were calculated both <str<strong>on</strong>g>for</str<strong>on</strong>g> men and women. For<br />

men relative importance <str<strong>on</strong>g>of</str<strong>on</strong>g> extrinsic life goals was not significantly related to MS (OR=1,31, 95% CI 0,94-<br />

1,81). For women relative importance <str<strong>on</strong>g>of</str<strong>on</strong>g> extrinsic life goals was significantly related to MS (OR=1,38, 95%<br />

CI 1,06-1,80) even after c<strong>on</strong>trolling <str<strong>on</strong>g>for</str<strong>on</strong>g> sociodemographic variables (age and educati<strong>on</strong>) and average<br />

importance <str<strong>on</strong>g>of</str<strong>on</strong>g> life goals. Results indicate gender differences regarding the relati<strong>on</strong>ship between marital stress<br />

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3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

and life goals: perceived marital stress may be c<strong>on</strong>nected to extrinsic life goal orientati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> women but not<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> men.<br />

7K_04_P<br />

(poster secti<strong>on</strong> A2, poster board #174, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

RISK FACTORS OF MASCULINE GENDER ROLE STRESS AMONG MIDDLE-AGED<br />

MEN IN HUNGARY<br />

Anna Susanszky, Eva Susanszky, Maria S. Kopp<br />

Semmelweis University, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Behavioral Sciences, 1089, Budapest, Nagyvarad ter 4.<br />

e-mail: susana@net.sote.hu<br />

Aim: To investigate relati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> masculine gender role stress to work-, marital,- social stress, rival behavior,<br />

self rated health and socio-ec<strong>on</strong>omical status. Methods: Survey am<strong>on</strong>g middle-aged (age 36-64) men (N=614)<br />

selected from Hungarostudy Epidemiological Panel 2006 (N= 4528) using the shortened Masculine Gender<br />

Role Stress Scale (Eisler&Skidmore, 1987), the shortened marital Stress Scale, shortened Ef<str<strong>on</strong>g>for</str<strong>on</strong>g>t – Reward<br />

Imbalance Questi<strong>on</strong>naire, and Bergen Social Relati<strong>on</strong>ship Scale. Stepwise linear regressi<strong>on</strong> analyses (stepwise<br />

method) were per<str<strong>on</strong>g>for</str<strong>on</strong>g>med to study the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> different sources <str<strong>on</strong>g>of</str<strong>on</strong>g> stress and the rival behavior <strong>on</strong> the<br />

masculine gender role stress. Age, health and socio-ec<strong>on</strong>omical status were also included in the model.<br />

Results: Using stepwise linear regressi<strong>on</strong> analyses, work stress, (standardized beta=0.10), educati<strong>on</strong>al level<br />

(standardized beta=-0.12), and rival behavior (standardized beta=0.08) predicted masculine gender role stress<br />

best (F=6.68; p=0.00). C<strong>on</strong>clusi<strong>on</strong>: These results highlight that rival behavior and work stress may serve as<br />

risk factors <str<strong>on</strong>g>of</str<strong>on</strong>g> gender role stress. High educati<strong>on</strong>al level emerged as a protective factor <str<strong>on</strong>g>of</str<strong>on</strong>g> gender role stress.<br />

7K_05_P<br />

(poster secti<strong>on</strong> A2, poster board #175, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

GENDER DIFFERENCES IN ATTITUDES TOWARD DEATH AND ANXIETY<br />

Agnes Zana, Katalin Hegedus, Gabor Szabo<br />

Semmelweis University, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Behavioural Sciences, 4. Nagyvarad ter, 1024. Budapest, Hungary<br />

e-mail: zanagi@net.sote.hu<br />

Objective: The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> our research is to investigate whether there are gender differences in anxiety and death<br />

related attitudes. According to our hypothesis the attitudes and anxiety toward death and the pattern <str<strong>on</strong>g>of</str<strong>on</strong>g> fear<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> death differs <str<strong>on</strong>g>for</str<strong>on</strong>g> the two genders; we assume that women have higher fear <str<strong>on</strong>g>of</str<strong>on</strong>g> death and anxiety compared<br />

to men.<br />

Method: For the questi<strong>on</strong>naire survey (N=246, female:167, male 76 /3 missing/, average age: 41,5) we used<br />

the Neimeyer and Moore’s Multidimensi<strong>on</strong>al Fear <str<strong>on</strong>g>of</str<strong>on</strong>g> Death Scale (MFODS, Neimeyer and Moore 1994, Zana 2006)<br />

and the STAI-T (State-Trait Anxiety Inventory, Spielberger 1970, Sipos 1978).<br />

Results: The three death fear factors rated highest are Fear <str<strong>on</strong>g>for</str<strong>on</strong>g> significant others, Fear <str<strong>on</strong>g>of</str<strong>on</strong>g> the dying process and Fear <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the dead in both groups. However, we found significant gender differences in fear intensity <strong>on</strong> these factors<br />

(p


476<br />

23-26 August 2007,<br />

Budapest, Hungary


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

MODULE 8<br />

OTHERS<br />

477


23-26 August 2007,<br />

Budapest, Hungary<br />

Poster: August 24th Saturday, poster secti<strong>on</strong> A1, building A groundfloor<br />

478


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

8_01_P<br />

(poster secti<strong>on</strong> A1, poster board #15, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

GENETICS OF SALINITY TOLERANCE IN WHEAT<br />

H. Dashti¹, M. Naghavi², A. Tajabadipur 3<br />

1,3Valiasr University, Rafsanjan, Iran<br />

2Tehran University, Karaj, Iran<br />

Six generati<strong>on</strong>s (P1, P2, F1, F2, BC1 and BC2) <str<strong>on</strong>g>of</str<strong>on</strong>g> cross Rovshan × Falat was grown in greenhouse c<strong>on</strong>diti<strong>on</strong><br />

in order to evaluate gene effects <str<strong>on</strong>g>of</str<strong>on</strong>g> resistance to salinity in wheat. Five characters including: sodium and<br />

potassium c<strong>on</strong>centrati<strong>on</strong>s, K/Na ratios, plant height and heading date were recorded and subjected to<br />

estimate means and variances pooled over replicati<strong>on</strong>s. Gene effects were calculated by using the six<br />

parameter model, viz., m (average effect), d (additive), h (dominance), i (additive_additive) j (additive_<br />

dominance) and l (dominance_dominance) according to Hayman (1958) after testing adequacy <str<strong>on</strong>g>of</str<strong>on</strong>g> additivedominance<br />

(three parameter) model by using joint scaling test <str<strong>on</strong>g>of</str<strong>on</strong>g> Cavalli (1952). The six-parameter model was<br />

adequate in most <str<strong>on</strong>g>of</str<strong>on</strong>g> the cases to explain genetic variati<strong>on</strong> am<strong>on</strong>g the generati<strong>on</strong> means. Generati<strong>on</strong> mean<br />

analysis revealed that additive comp<strong>on</strong>ents played a major role, but that dominance comp<strong>on</strong>ents also<br />

c<strong>on</strong>tributed significantly in c<strong>on</strong>trolling the variability <str<strong>on</strong>g>of</str<strong>on</strong>g> the recorded characters. The dominant and dominant<br />

× dominant comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g> the model <str<strong>on</strong>g>for</str<strong>on</strong>g> most characters showed values in the opposite directi<strong>on</strong> as it<br />

indicated the duplicate epistasis, which denoted little role <str<strong>on</strong>g>of</str<strong>on</strong>g> this epistasis in selecti<strong>on</strong> as it decreased heterosis<br />

in selecti<strong>on</strong>. This study revealed that these characters showed all three types <str<strong>on</strong>g>of</str<strong>on</strong>g> gene acti<strong>on</strong> i.e. additive,<br />

dominance and epistasis and should be c<strong>on</strong>sidered to cumulate the resistance genes to salinity in <strong>on</strong>e<br />

genotype. In order to utilize all three types <str<strong>on</strong>g>of</str<strong>on</strong>g> gene effects simultaneously, reciprocal recurrent selecti<strong>on</strong><br />

breeding procedure would be the best opti<strong>on</strong>. Nevertheless, the in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> obtained in this study provides a<br />

better understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> the genetic resistance to salinity and, it is a prerequisite to apply pyramiding <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

resistance genes. The development <str<strong>on</strong>g>of</str<strong>on</strong>g> appropriate markers linked to resistance genes would greatly enhance<br />

the feasibility <str<strong>on</strong>g>of</str<strong>on</strong>g> such a strategy.<br />

8_03_P<br />

(poster secti<strong>on</strong> A1, poster board #16, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

A NOVEL HAND-HELD MONITOR OF SYMPATHETIC<br />

NERVOUS SYSTEM USING BIOMARKER<br />

Masaki Yamaguchi, Kazutaka Teramura, Yusuke Tahara, Josaku Sakakima<br />

Graduate School <str<strong>on</strong>g>of</str<strong>on</strong>g> Science and Engineering, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Toyama, Japan<br />

e-mail: yamag@eng.u-toyama.ac.jp<br />

Objective: Salivary amylase activity (sAMY) can be a useful index <str<strong>on</strong>g>of</str<strong>on</strong>g> plasma norepinephrine c<strong>on</strong>centrati<strong>on</strong><br />

under a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> stressful c<strong>on</strong>diti<strong>on</strong>s, since it appears that increased sympathetic nervous activity is a major<br />

stimulator <str<strong>on</strong>g>of</str<strong>on</strong>g> amylase secreti<strong>on</strong>. In order to realize a hand-held m<strong>on</strong>itor <str<strong>on</strong>g>of</str<strong>on</strong>g> the sympathetic nervous system,<br />

we fabricated a completely automated analytical system <str<strong>on</strong>g>for</str<strong>on</strong>g> sAMY using a dry-chemistry system. Materials and<br />

Methods: The m<strong>on</strong>itor c<strong>on</strong>sisted <str<strong>on</strong>g>of</str<strong>on</strong>g> a disposable test-strip and an optical analyser (126 × 130 × 48 mm3; 350<br />

g), which was incorporated within an automatic saliva transfer device. The test-strip c<strong>on</strong>sisted <str<strong>on</strong>g>of</str<strong>on</strong>g> a collecting<br />

paper and a reagent paper c<strong>on</strong>taining 2-chloro-4-nitrophenyl-4-O-β-D-galactopyranosylmaltoside (Gal-G2-<br />

CNP), a substrate <str<strong>on</strong>g>for</str<strong>on</strong>g> amylase. The collecting paper is directly inserted into an oral cavity, and approximately<br />

30 µl <str<strong>on</strong>g>of</str<strong>on</strong>g> whole saliva is collected from under the t<strong>on</strong>gue. When Gal-G2-CNP is hydrolyzed by amylase, the<br />

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Budapest, Hungary<br />

hydrolyzed product (CNP) develops a yellow color and the reflectance is measured by the optical analyser.<br />

Results and Discussi<strong>on</strong>: When this m<strong>on</strong>itor was used, it took 30 s <str<strong>on</strong>g>for</str<strong>on</strong>g> saliva sampling, 30 s <str<strong>on</strong>g>for</str<strong>on</strong>g> saliva transfer<br />

and measurement, and a total <str<strong>on</strong>g>of</str<strong>on</strong>g> <strong>on</strong>e minute was enough to measure the sAMY. The calibrati<strong>on</strong> curve <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

m<strong>on</strong>itor was measured <str<strong>on</strong>g>for</str<strong>on</strong>g> sAMY. Within a range <str<strong>on</strong>g>of</str<strong>on</strong>g> sAMY between 10 and 140 kU/l, the calibrati<strong>on</strong> curve<br />

showed a coefficient with R 2 = 0.97. With regard to reproducibility <str<strong>on</strong>g>of</str<strong>on</strong>g> the measured results <str<strong>on</strong>g>for</str<strong>on</strong>g> the saliva<br />

transfer volume <str<strong>on</strong>g>of</str<strong>on</strong>g> the same samples, the coefficient variati<strong>on</strong> (CV) was 5.5%. C<strong>on</strong>clusi<strong>on</strong>: It was<br />

dem<strong>on</strong>strated that the manufactured m<strong>on</strong>itor enabled a user to automatically measure the sAMY with a high<br />

accuracy.<br />

8_04_P<br />

(poster secti<strong>on</strong> A1, poster board #17, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PREJUNCTIONAL AT1 RECEPTOR MEDIATES SYMPATHETIC RESPONSE TO<br />

COLD PRESSOR STRESS IN HUMANS<br />

L. E. Zavala*, M. Cierco*, A. Gutiérrez**, M. R. Garrido***, A. E. Israel***<br />

*Unidad de Investigaci<strong>on</strong>es Farmacológicas y Toxicológicas, UNEFM<br />

** Postgrado de Anestesiología, Hospital Universitario “Dr. Alfredo Van Grieken”, Coro, Estado Falcón<br />

*** Laboratorio de Neuropéptidos, Facultad de Farmacia UCV, Caracas, Venezuela<br />

e-mail: lidazavala@hotmail.com<br />

Angiotensin II (ANG) has a role in stress-induced cardiovascular resp<strong>on</strong>se by supporting sympathetic<br />

outflow through the stimulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> prejuncti<strong>on</strong>al AT 1 receptors. Thus, blockade <str<strong>on</strong>g>of</str<strong>on</strong>g> AT 1 receptor should<br />

decrease sympathetic resp<strong>on</strong>se to stress. To assessed this hypothesis, we compared the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> losartan with<br />

the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> eprosartan (EPRO), a selective prejuncti<strong>on</strong>al AT 1 receptor antag<strong>on</strong>ist, <strong>on</strong> the cardiovascular<br />

resp<strong>on</strong>se to cold pressor test (CPT) applied to healthy human volunteers. CPT is a model <str<strong>on</strong>g>of</str<strong>on</strong>g> acute stress<br />

known to increase arterial blood pressure (BP) and heart rate (HR). CPT was per<str<strong>on</strong>g>for</str<strong>on</strong>g>med by placing the<br />

subject’s left hand up to the wrist in iced water (4°C) <str<strong>on</strong>g>for</str<strong>on</strong>g> 90 sec. 82 healthy normotensive volunteers were<br />

recruited <str<strong>on</strong>g>for</str<strong>on</strong>g> the study. In<str<strong>on</strong>g>for</str<strong>on</strong>g>med c<strong>on</strong>sent was obtained from all subjects. They were divided in three<br />

treatment groups: placebo (n=37), losartan (50 mg) (n=14) and eprosartan (600 mg) (n=18). Cardiovascular<br />

parameters (BP and HR) were measured in a double-blind placebo-c<strong>on</strong>trolled fashi<strong>on</strong> in the groups be<str<strong>on</strong>g>for</str<strong>on</strong>g>e<br />

(basal), 175 minutes after oral single treatment (basal-2) and post CPT. CPT was accompanied by a significant<br />

increase in HR, systolic, diastolic and mean BP in placebo-treated subjects. Pretreatment with any <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

AT 1 R antag<strong>on</strong>ists completely suppressed the pressor resp<strong>on</strong>se to CPT without changes in HR resp<strong>on</strong>se. Our<br />

results dem<strong>on</strong>strate a role <str<strong>on</strong>g>of</str<strong>on</strong>g> prejuncti<strong>on</strong>al AT 1 receptor in the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> sympathetic resp<strong>on</strong>se to acute<br />

stress.<br />

480


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

8_05_P<br />

(poster secti<strong>on</strong> A1, poster board #18, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

WHAT CHARACTERIZES MEN AND WOMEN WITH HIGH LEVELS OF STRESS<br />

Line Nielsen, Tine Curtis, Tage S. Kristensen, Naja Rod Nielsen<br />

Nati<strong>on</strong>al Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Public Health, Øster Farimagsgade 5A, DK-1399 Copenhagen K, Denmark<br />

e-mail: lin@si-folkesundhed.dk<br />

Background: Despite the fact that stress is a growing public health problem little is known about what<br />

characterizes individuals with stress. In order to prevent stress it is essential to be aware <str<strong>on</strong>g>of</str<strong>on</strong>g> factors that are<br />

predictive <str<strong>on</strong>g>of</str<strong>on</strong>g> stress. The objective <str<strong>on</strong>g>of</str<strong>on</strong>g> the study is to assess individual and neighborhood level factors that are<br />

associated with stress in a nati<strong>on</strong>al representative sample <str<strong>on</strong>g>of</str<strong>on</strong>g> the Danish populati<strong>on</strong>.<br />

Methods: The Nati<strong>on</strong>al Health Interview Survey 2005 c<strong>on</strong>sisted <str<strong>on</strong>g>of</str<strong>on</strong>g> a representative sample <str<strong>on</strong>g>of</str<strong>on</strong>g> the Danish<br />

populati<strong>on</strong>. The 9.708 men and women were asked about perceived stress, health related behavior, working<br />

c<strong>on</strong>diti<strong>on</strong>s, and sociodemographic factors. Data <strong>on</strong> neighborhood factors like crime rate or material<br />

deprivati<strong>on</strong> were derived from nati<strong>on</strong>al registries. Stress was assessed by the Perceived Stress Scale. Data were<br />

analyzed by means <str<strong>on</strong>g>of</str<strong>on</strong>g> logistic regressi<strong>on</strong> models.<br />

Results: Women had a higher mean score <str<strong>on</strong>g>of</str<strong>on</strong>g> stress than men. Low educati<strong>on</strong>, heavy smoking, physical<br />

inactivity, lack <str<strong>on</strong>g>of</str<strong>on</strong>g> social network, and poor working c<strong>on</strong>diti<strong>on</strong>s were associated with higher perceived stress<br />

am<strong>on</strong>g both men and women. Living in a deprived neighborhood was associated with higher stress am<strong>on</strong>g<br />

women, while living in a neighborhood with a high rate <str<strong>on</strong>g>of</str<strong>on</strong>g> crime was associated with higher stress am<strong>on</strong>g<br />

men.<br />

C<strong>on</strong>clusi<strong>on</strong>s: This study c<strong>on</strong>tributes to the understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> what characterizes individuals with high stress and<br />

may thereby help in guiding future preventi<strong>on</strong>. To strengthen the knowledge <str<strong>on</strong>g>of</str<strong>on</strong>g> the relati<strong>on</strong> between<br />

individual factors, neighborhood factors and stress prospective studies and interventi<strong>on</strong> studies are needed.<br />

8_10_P<br />

(poster secti<strong>on</strong> A1, poster board #19, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

FUNCTIONAL RESPONSES OF LAND SNAIL DEFENCE CELLS RELATED TO<br />

ANTI-MICROBIAL ACTIVITY AND TOXICANT EXPOSURE<br />

Lisa Johns<strong>on</strong>, Jacqueline Russo, Luc Madec<br />

Instituti<strong>on</strong>: UMR 6553 Ecobio, Univ Rennes1, CNRS, Campus de Beaulieu, avenue du Général Leclerc,<br />

35042 Rennes, France<br />

e-mail: lisajohns<strong>on</strong>@libertysurf.fr<br />

Am<strong>on</strong>g many approaches to study stress in invertebrate, especially molluscs, we have chosen to focus <strong>on</strong> cells<br />

involved in immunity, in the interplay between host-pathogen interacti<strong>on</strong>s and disturbance in their habitat,<br />

such as c<strong>on</strong>taminati<strong>on</strong> by xenobiotics. Phagocytic cells called hemocytes, circulating in body fluid <str<strong>on</strong>g>of</str<strong>on</strong>g> molluscs,<br />

c<strong>on</strong>stitute the first line <str<strong>on</strong>g>of</str<strong>on</strong>g> defence against <str<strong>on</strong>g>for</str<strong>on</strong>g>eign materials, but they also serve a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> other important<br />

physiological functi<strong>on</strong>s, all relevant <str<strong>on</strong>g>of</str<strong>on</strong>g> homeostasis maintenance. C<strong>on</strong>sequently, toxicant threatening could<br />

potentially affect individual survival. In the present experimental study, we compared the immune resp<strong>on</strong>se<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> natural populati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> the invasive land snails Cantareus aspersus living in anthropized ecosystems <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

North-western Europe, and Cepaea nemoralis, native from this regi<strong>on</strong> and living in ruderal or <str<strong>on</strong>g>for</str<strong>on</strong>g>est habitats<br />

but not in agrosystems. Such invasive success <str<strong>on</strong>g>of</str<strong>on</strong>g> C. aspersus led to the hypothesis that the differential<br />

reactivity to toxicants used in agrosystems may be partially due to species differences in immune features. In<br />

481


23-26 August 2007,<br />

Budapest, Hungary<br />

fact, the immune system is known to be a target <str<strong>on</strong>g>for</str<strong>on</strong>g> toxicant and phagocytes to be mediators <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress<br />

resp<strong>on</strong>se. From an appreciati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> immune reactivity and related allocati<strong>on</strong> trade-<str<strong>on</strong>g>of</str<strong>on</strong>g>fs in the two species, we<br />

might understand why C. aspersus populati<strong>on</strong>s are able to live <strong>on</strong> polluti<strong>on</strong> c<strong>on</strong>diti<strong>on</strong>s which impose severe<br />

stress where the other species can not. Using experimental living Escherichia coli infecti<strong>on</strong>, we compared invivo<br />

phagocytic capacity <str<strong>on</strong>g>of</str<strong>on</strong>g> snails (subjected to the herbicide fomesafen) by means <str<strong>on</strong>g>of</str<strong>on</strong>g> the following<br />

measurements: (i) circulating hemocyte c<strong>on</strong>centrati<strong>on</strong>, (ii) hemocyte viability, (iii) subsequent cytotoxic<br />

activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the cells by releasing reactive oxygen species (ROS) and (iv) bacterial clearance. The immune<br />

disorders were investigated at envir<strong>on</strong>mental c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 90 µ/l, after short (24hrs) and l<strong>on</strong>g term (five<br />

days) exposure.<br />

8_11_P<br />

(poster secti<strong>on</strong> A1, poster board #20, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PROTECTIVE ROLE OF N-PROCALCITONIN IN ENDOTOXIC SHOCK<br />

Eva Tavares, Rosario Mald<strong>on</strong>ado, Inmaculada Capilla, Cesar Sevillano, Francisco J. Miñano<br />

Pharmacology <str<strong>on</strong>g>Research</str<strong>on</strong>g> Unit, Valme University Hospital, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Seville, 41014 Seville, Spain<br />

e-mail: jminano@us.es<br />

The overzealous producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proinflammatory cytokines in sepsis can result in shock, multiorgan<br />

dysfuncti<strong>on</strong>, and even death. Procalcit<strong>on</strong>in (PCT), a 14-kDa propeptide <str<strong>on</strong>g>of</str<strong>on</strong>g> calcit<strong>on</strong>in, is a circulating<br />

biomarker <str<strong>on</strong>g>of</str<strong>on</strong>g> bacterial infecti<strong>on</strong> but PCT itself has no known activity. Circulating levels <str<strong>on</strong>g>of</str<strong>on</strong>g> PCT and its free<br />

aminopetide N-procalcit<strong>on</strong>in (N-PCT), have been found dramatically increased in septic patients, and these<br />

rises are correlated with severity and mortality. Importantly, in sepsis, the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> N-PCT may be even<br />

higher than the PCT values. In vitro studies suggest that N-PCT may functi<strong>on</strong> as a factor suppressing the<br />

propagati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> inflammati<strong>on</strong> through the inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> several processes involved during a resp<strong>on</strong>se to a<br />

bacterial stimulus.These findings together with N-PCT's sequence c<strong>on</strong>servati<strong>on</strong> during evoluti<strong>on</strong>, suggest that<br />

N-PCT has a critical, and as yet undefined, pathophysiological functi<strong>on</strong>. In this study, we assessed the role <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

N-PCT as a mediator <str<strong>on</strong>g>of</str<strong>on</strong>g> sepsis in endotoxin-challenged rats. Intraperit<strong>on</strong>eal administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> an lethal dose<br />

(LD 100 ) <str<strong>on</strong>g>of</str<strong>on</strong>g> E. coli endotoxin to normal rats induced a substantial increase in PCT, IL-1β and TNF-α in<br />

plasma. The administrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> recombinant human N-PCT intraperit<strong>on</strong>eally significantly protected rats from<br />

endotoxin-induced mortality from 100% to 15%, and resulted in a decrease in PCT, IL-1β and TNF-α levels.<br />

By c<strong>on</strong>trast, N-PCT did not modify IL-10 levels. These results c<strong>on</strong>firm a critical part <str<strong>on</strong>g>for</str<strong>on</strong>g> PCT in the<br />

pathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> endotoxic shock and indicate that N-PCT is a protective peptide expressed in murine<br />

endotoxemia, and does so by down-regulating the systemic producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proinflammatory cytokines in<br />

endotoxin-challenged animals.<br />

482


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

AUTHORS’ INDEX<br />

483


484<br />

23-26 August 2007,<br />

Budapest, Hungary


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

A<br />

Aaboe, M. 324<br />

Abbate, C. 420<br />

Abbruzzese, A. 285<br />

Abdiová, H. 302<br />

Abdrakhmanov, O. 230<br />

Abraham, G. N. 337<br />

Abreu-Ramos, A. 284<br />

Adam, Sz. 413, 440, 471<br />

Adam, Z. 114<br />

Adeli, K. 101<br />

Ad<strong>on</strong>yeva, N. V. 273<br />

Adzic, M. 423<br />

Afanasyeva, E. 315<br />

Aggeli, C. 261<br />

Agnew, L. L. 32<br />

Aguayo, C. 295<br />

Aguilar, R. 295<br />

Ahn, J.-Y. 266<br />

Ahn, S.-G. 45, 23<br />

Ahrens, F. 290<br />

Akana, F. S. 8<br />

Åkerfelt, M. 44, 268<br />

Akhalaya, M. Y. 166<br />

Akin, S. 38<br />

Akira, T. 405<br />

Aksy<strong>on</strong>ova, O. V. 164<br />

Alavez, S. 152, 239<br />

Albani, S. 330<br />

Aldikactioglu, M. 130, 134<br />

Alekperov, E. A. 83<br />

Alekseeva, L. G. 33<br />

Aleo, M. F. 254<br />

Alexzander Asea 49<br />

Alfieri, R. R. 78<br />

Almeida, M. 226, 284<br />

Altaie, M. B. O. 140<br />

Altug, T. 130, 132, 134<br />

Alyane, M. 142<br />

Amaddeo, G. 420<br />

Amano, R. 398<br />

Amodio, G. 104<br />

Am<strong>on</strong>s, R. 167<br />

Anderberg, U. M. 409<br />

Anders<strong>on</strong>, R. L. 79<br />

Anderzen, I. 409, 462, 460<br />

Andréass<strong>on</strong>, C. 28<br />

Andrés, M. E. 44<br />

Ang, D. 28, 356<br />

Angel, J. 261<br />

Ans<strong>on</strong>, O. 334<br />

Anth<strong>on</strong>y, N. B. 289<br />

Antune, F. 152<br />

Anzal<strong>on</strong>e, R. 158<br />

Aparicio García, M. E. 474<br />

Apel, K. 188<br />

Aquilina, J. A. 24, 26<br />

Arad, Z. 302<br />

Aratanha, C. B. 113<br />

Araújo, S. S. 220<br />

Archbold, G. P. 237<br />

Arck, C. P. 417, 457<br />

Ardizz<strong>on</strong>e, M. N. 85<br />

Areias, F. M. 128<br />

Arispe, N. 51<br />

Armay, Ö. 134<br />

Armentano, N. 221<br />

Arnas<strong>on</strong>, T. J. 200<br />

Arnetz, B. B. 409, 460, 462<br />

Aro, E.-M. 10<br />

Ar<strong>on</strong>z<strong>on</strong>, C. 255<br />

Arrigo, A.-P. 17<br />

Arslan, M. A. 365<br />

Arun, C. P. 175<br />

Arunabha, R. 432, 434<br />

Arzola, J. 132<br />

Arzt, E. 257<br />

Asada, T. 402<br />

Asai, M. 68<br />

Asard, H., 190<br />

Ashizawa, Y. 402<br />

Atukeren, P. 130, 134<br />

Au, D. W. T. 111<br />

Aung, Z. 460<br />

Auvinen, P. 161<br />

Avezov, E. 102<br />

Azevedo, A. R. 190<br />

Azevedo, M. F. 284<br />

Azevedo, R. A. 191, 228<br />

B<br />

Baas, F. 364<br />

Baban, N. 106<br />

Back, R. - J. 47<br />

Badesha, P. 180<br />

Badke, A. 449<br />

Bagaeva, T. 375<br />

Bai, Z.-l. 143<br />

485


23-26 August 2007,<br />

Budapest, Hungary<br />

Baizhumanov, A. A. 166<br />

Bakogiannis, C. 280<br />

Balbis, E. 238<br />

Balch, E. W. 354<br />

Bálint, Z. 147<br />

Balkanci, D. Z. 265<br />

Balla, I. 212<br />

Balog, P. 404, 413, 440, 474<br />

Baltopoulos, G. 71<br />

Bánhegyi, G. 98<br />

Barancik, M. 341<br />

Barbaro, M. 420<br />

Barreto, A. 261<br />

Barriere, H. 118<br />

Barsy, B. 391<br />

Bartels, D. 187<br />

Bartfai, T. 257<br />

Bartha, B. 226<br />

Bartha, B. 294<br />

Bart<strong>on</strong>, D. 416<br />

Bar-Zvi, D. 212<br />

Basa, B. 227<br />

Basha, E. 11<br />

Bashenko, Y. 319<br />

Basini, G. 127, 130, 282<br />

Bassi, A. 340<br />

Bates, E. H. 410<br />

Battaglioli, E. 44<br />

Baumeister, P. 98<br />

Bauza, Y. 138<br />

Bayles, R. 414<br />

Bayrak, I. 130, 134<br />

Bayrak, S. 265<br />

Bazhan, N. M. 259<br />

Beattie, A. E. 361<br />

Bedja, D. 328<br />

Beier, G. 414<br />

Beishuizen, A. 347<br />

Belak, M 462<br />

Belardo, G. 39<br />

Bellafiore, M. 85<br />

Belosevic, L. 464<br />

Bellyei, Sz. 19<br />

Bender, T. 115, 118<br />

Benedek, G. 370<br />

Benedetti, M. 240<br />

Benesch, J. L. P. 24, 26<br />

Benguedouar, L. 142<br />

Benham, A. 101<br />

Benjamin, I. 41, 308<br />

Benkö, E. 414<br />

Benndorf, R. 92, 310<br />

Ben-Zvi, A. 236<br />

Béracochéa, D. 403<br />

Bérczi, A. 190<br />

Berczi, I. 255<br />

Beregova, T. 373<br />

Bergh<str<strong>on</strong>g>of</str<strong>on</strong>g>er, P. 53<br />

Bergström, S.-A. 281<br />

Bernabucci, U. 342<br />

Bernardes da Silva, A. 220<br />

Bernatova, I. 341<br />

Berra, E. 106<br />

Berry-Kravis, E. M. 241<br />

Bertaud, M. 361<br />

Bertram, H. C. 83<br />

Bettecken, T. 395<br />

Bett<strong>on</strong>i, F. 254<br />

Bhat, M. 20<br />

Bhomkar, P. 222<br />

Bianchi, K. 325<br />

Biasioli, B. 279<br />

Bickeböller, H. 316<br />

Bickle, M. 87<br />

Binder, P. 295<br />

Bini<strong>on</strong>, G. D. 332<br />

Birch, S. C. 129<br />

Birtić, S. 191<br />

Biselli, R. 275<br />

Biss<strong>on</strong>, J.-F. 239<br />

Bistola, V. 261<br />

Biswas, A. 20<br />

Black, S. 322<br />

Blandino, G. 75<br />

Blangero, J. 414<br />

Blasczyk, R. 269<br />

Blois, S. 430, 457<br />

Bobak, M. 437<br />

Bobyk, V. 387<br />

Bogatyrenko, T. N. 241<br />

Bogomolova, E. V. 273<br />

Bohnert, J. H. 202<br />

Bojikova-Fournier, S. 167<br />

Bolz<strong>on</strong>i, E. 357<br />

B<strong>on</strong>atti, S. 104<br />

B<strong>on</strong>elli, M. 78<br />

Borghetti, F. A. 78<br />

Borisov, V. B. 137<br />

486


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Bor<strong>on</strong>kai, A. 19<br />

Bost<strong>on</strong>, S. R. 104<br />

Botanov, Y. 176<br />

Bottari, P. S. 380<br />

Boucherie, H. 162<br />

Boutsikou, M. 261<br />

Boyault, C. 97<br />

Braga, A. P. 104<br />

Brahimi-Horn, M. C. 106<br />

Brandi, A. 165<br />

Brand<strong>on</strong>, D. 454<br />

Braš, M. 451<br />

Brassard, P. 258<br />

Braszko, J. J. 243, 394<br />

Bratt<strong>on</strong>, B. S. 76<br />

Brazier, S. P. 106<br />

Breitner, J. C. S. 403<br />

Bremseth, H. 66, 323<br />

Brenner, I. 277<br />

Bret<strong>on</strong>-H<strong>on</strong>eyman, K. 277<br />

Brkljačić, J. 253, 384<br />

Brocchieri, L. 30, 311<br />

Brockmeier, U. 117<br />

Bross, G. P. 357<br />

Bross, P. 83, 356, 359<br />

Brower, J. B. 241<br />

Brown, I. R. 246<br />

Bruneau, B. M. S. 473<br />

Brunner, E. 316, 336<br />

Brunori, M. 137<br />

Bryer, P. 290<br />

Bučková, M. 140<br />

Bugajska, J. 425<br />

Bughi, A. S. 282, 456<br />

Bukau, B. 28<br />

Bulbovas, P. 191<br />

Bunck, M. 393, 395<br />

Burd, R. 322<br />

Burrows, F. 326<br />

Bussani, A. 279<br />

Bussolanti, S. 127<br />

Bussolati, S. 130<br />

Butterbach, K. 347<br />

Buyse, J. 33<br />

C<br />

Cabello, G. 33<br />

Cadei, M. 254<br />

Cagnacci, A. 141<br />

Cai, G. 112<br />

Cai, Q. 143, 145<br />

Calderwood, K. S. 49, 59<br />

Cali, T. 114<br />

Callipari, S. 420<br />

Calvo, J. H. 361<br />

Campanella, C. 85<br />

Campos, R. A. 154<br />

Campos, V. 156<br />

Candela, M. E. 62<br />

Cannoletta, M. 141<br />

Cao, Y. B. 112<br />

Capilla, I. 482<br />

Cappello, F. 85, 158<br />

Cardenal Herráiz, V. 474<br />

Cardoso, A. I. 194<br />

Carnecka, M. 160<br />

Carpenter, E. R. 392<br />

Carpentier, A. 258<br />

Carra, S. 22, 92, 310, 354<br />

Carragher, B. 354<br />

Carsillo, T. 299<br />

Carver, J. 26<br />

Carver, J. A. 24<br />

Cassese, D. 285<br />

Castañaga, L. A. 255<br />

Castro-Caldas, M. 79<br />

Catalano, D. 328<br />

Cavaill<strong>on</strong>, J.-M. 346<br />

Cavalier, E. 366<br />

Cavazz<strong>on</strong>i, A. 78<br />

Caved<strong>on</strong>, C. 380<br />

Cecere, F. 124<br />

Ceciliani, F. 342<br />

Cellini, F. 221<br />

Chaerle, L. 211<br />

Chafekar, S. M. 364<br />

Chambers, A. D. 151, 262<br />

Chami, M. 325<br />

Chang, Y.-T. 399<br />

Chapman-Smith, A. 108<br />

Charney, D. 447<br />

Charrette, J. S. 147<br />

Chatterji, D. 125<br />

Chau, C 407<br />

Chelini, M. M. O. 287, 458<br />

Chen, R. L. 349<br />

Chen, C. C.-H. 415<br />

Chen, E. X. H. 111<br />

487


23-26 August 2007,<br />

Budapest, Hungary<br />

Chen, H.-i. 342, 394<br />

Chen, L. 373, 374<br />

Chen, R. 143, 145<br />

Chen, S. 246<br />

Chen, S. J. 351<br />

Chen, S. J. 349<br />

Chen, T. 167<br />

Chen, X. Q. 109, 110, 112<br />

Chen, X.-C. 112<br />

Chen, Y.-C. 399<br />

Chen, Y.-X. 177<br />

Chen, Z. F. 351<br />

Chen, Z.-H. 136<br />

Chenderes, R. 472<br />

Cheng, D.-x. 143, 145<br />

Cheng, J. S. 249, 250<br />

Cheng, L. 79<br />

Chentsova, N. A. 273<br />

Cheregi, O. 188<br />

Chernik, S. I. 22<br />

Chern<str<strong>on</strong>g>of</str<strong>on</strong>g>f, O. Y. 353<br />

Chiang, M. W. L. 111<br />

Chiang, P. L. 349<br />

Chiang, P. L. 351<br />

Chiba, S. 359<br />

Chiche, J. 106<br />

Chien, S.-P. 351<br />

Chiocca, S. 97<br />

Chiu, C. T. 351<br />

Chivasa, S. 197<br />

Choi, W.-K. 157<br />

Chow, A. 246<br />

Christensen, H. J. 356, 357, 359<br />

Christiansen, M. 387<br />

Chritians, E. 41<br />

Chrousos, P. G. 171<br />

Chylinski, W. K. 225<br />

Ciafre, S. 39<br />

Ciangherotti, C. 131<br />

Cierco, M. 480<br />

Ciucci, A. 39<br />

Clark, I. J. 86<br />

Clem<strong>on</strong>s, A. 454<br />

Cobo, M. P. 381<br />

Coe, L. C. 428<br />

Cohen, H. 260<br />

Cohen, L. R. 151<br />

Cohen, R. I. 262, 331<br />

Colas, P. 87<br />

Colditz, G. I. 32<br />

Collin, V. 187<br />

Collins, K. 447<br />

Collins, L. 146<br />

C<strong>on</strong>way de Macario, E. 30<br />

Corcoran, C. 403<br />

Cordeiro, R. 471<br />

Corrao, S. 158<br />

Cortes-Denia, P. 381<br />

Coryd<strong>on</strong>, J. T. 357, 359, 364<br />

Costa, A. 226<br />

Costa, D. L. M. 104<br />

Cottalasso, D. 238<br />

Cotta-Pereira, G. 113<br />

Cotugno, R. 124<br />

Couls<strong>on</strong>, T. 376<br />

Couraud, P. O. 147<br />

Coutinho, O. P. 128, 284<br />

Cox, D. 176<br />

Craig, A. E. 29<br />

Crippa, V. 357<br />

Cropley, M. 411<br />

Crumeyrolle-Arias, M. 379<br />

Cruz e Silva, E. 206<br />

Cryns, V. 312<br />

Cuilan, M. 150<br />

Ćurić, G. 451<br />

Curran, M. 237<br />

Curtis, T. 481<br />

Cuypers, A. 198<br />

Cyrne, L. 152<br />

Czarnecka, M. A. 85<br />

Czech, V. 195<br />

Czermak, C. 448<br />

Czibere, L. 395<br />

Czövek, P. 195, 205, 212<br />

Czyz, D. 354<br />

Cseh, E. 195<br />

Csenki, Zs. 292<br />

Cser, K. 188<br />

Cserep, Zs. 413<br />

Cserháti, M. 194, 199<br />

Csermely, P. 54, 105, 308, 365<br />

Cseuz, L. 214<br />

Csibi, A. 131<br />

Csiszár, J. 151, 214<br />

Csizmadiova, Z. 341<br />

488


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

D<br />

D´Eramo J. L. 255<br />

d’Alessio, P. 239<br />

D’Haese, D. 198<br />

Dallman, F. M. 8<br />

Damis, E. 380<br />

Dancsó, B. 365<br />

Daniltchenko, M. 429, 430, 432<br />

Dantzer, R. 427<br />

Darnaudéry, M. 383<br />

Darras, V. M. 304<br />

Dashti, H. 479<br />

Date, A. 244<br />

Daugaard, M. 312, 324<br />

Daugé, V. 379<br />

Dave, K. 108<br />

Davenport, M. D. 401<br />

Davis, H. D. 151<br />

Dawood, T. 414, 416<br />

Day, B. 197<br />

Dayan, F. 106<br />

de Carvalho, D. R. 154<br />

De Groef, B. 304<br />

de Haro, V. 216<br />

De Jesús, S. 132<br />

De J<strong>on</strong>ghe, P. 307, 310<br />

De la Chica-Rodriguez, S. 381<br />

de Macario, E. C. 85, 158, 307, 311<br />

De Maio, A. 51<br />

de Oliveira, A. C. 287<br />

De Sim<strong>on</strong>e, C. 285<br />

de Souza, S. R. 191<br />

De Stefani, D. 325<br />

De Vendittis, E. 124<br />

Debeljak, M. 466<br />

Decker, S. 396<br />

Decuypere, E. 33<br />

DeGuzman, C. 92, 310<br />

Delbello, G. 279<br />

Delespaul, P. 366<br />

Delitheos, B. 161<br />

Demajo, M. 138<br />

Demirel, H. 38<br />

Demoinet, E. 63<br />

Dempsey, C. N. 37, 78, 316<br />

Deng, X. M 373, 374<br />

Denis, L. 40<br />

Derek Middlet<strong>on</strong> 237<br />

Descamps, O. 240<br />

Desco, M. M. 340<br />

Deussing, J. M. 274<br />

Devaney, E. 58<br />

Devereux, J. J. 411<br />

Dewey, E. R. 104<br />

Di Agostino, S. 75<br />

Di Felice, V. 85<br />

Di Martino, L. P. C. M. 144<br />

Diam<strong>on</strong>d, M. D. 391<br />

Dias, A. 471<br />

Díaz, H. 84<br />

Diaz, J.C. 51<br />

Díaz, M. 216<br />

Dicitore, A. 285<br />

Dickins<strong>on</strong>, A. M. 332<br />

Didelot, C. 75<br />

Dierick, I. 307, 310<br />

Dietrich, H. 348<br />

Díez, J. 127<br />

Dimitrijević, M. 264<br />

Dimpfel, W. 176<br />

Dittami, J. 459<br />

Djordjevic, J. 423<br />

Dobrozemsky, G. 348<br />

Dolińska, G. 422<br />

Doller, A. 153<br />

Domany, E. 75<br />

Domenicotti, C. 238<br />

Domes, G. 449<br />

Domingos, M. 191<br />

D<strong>on</strong>alds<strong>on</strong>, P. R. 127<br />

D<strong>on</strong>g, D. 98<br />

D<strong>on</strong>gliang, Q. 150<br />

D<strong>on</strong>oghue, A. M. 289<br />

D<strong>on</strong>zelli, S. 75<br />

Dorant, E. 396<br />

Doshi, B. 384<br />

Doumba, P. 261<br />

Dracup, K. 461<br />

Dresch, V. 474<br />

Dressel, R. 269, 316<br />

Dridi, S. 33<br />

Drobinska, O. 373<br />

Du, J. Z. 109, 110, 112<br />

Dubovický, M. 419<br />

Dudits, D. 151, 189,194, 223, 224<br />

Dulin, E. 340<br />

Dunđerski, J. 215<br />

Dundjerski, J. 253<br />

489


23-26 August 2007,<br />

Budapest, Hungary<br />

Duque Santos, S. 363<br />

Duque, A. S. 220<br />

Durham, D. H. 358<br />

Đurković, M. 451<br />

Dürr, A. 359<br />

E<br />

Eberhardt, W. 153<br />

Ec<strong>on</strong>omou, V. 261<br />

Eddy, A. 414<br />

Eikelenboom, P. 364<br />

Eiz-Vesper, B. 269<br />

Ekeberg, Ø. 469<br />

Ekmekci, H. 132<br />

El Fatimy, R. 40, 47<br />

Elaković,I. 384<br />

Eleutherio, E. C. A. 113<br />

El-Hadi, M. 149<br />

Ellis, J. R. 7<br />

Elsner, L. 316<br />

Emeny, R. T. 251<br />

Emerich, J. 320<br />

Emery, E. P. 451<br />

Emery, V. O. B. 451<br />

Enache, M. 383<br />

Endo, G. 402<br />

Engstrom, P. 221<br />

Eom, C.-Y. 126<br />

Epanchinceva, E. M. 164<br />

Erdei, L. 151, 214, 226, 294<br />

Erhard, M. H. 290<br />

Erikss<strong>on</strong>, E. 159<br />

Erikss<strong>on</strong>, J. E. 47<br />

Erikss<strong>on</strong>, M. 173<br />

Erikss<strong>on</strong>, W. J. 336<br />

Erlichman, C. 317<br />

Esler, M. 416<br />

Esp<strong>on</strong>da, P. 84<br />

Etemad, S. 263<br />

Evans, H. 35<br />

Evd<strong>on</strong>in, L. A. 55<br />

Eyal, S. 456<br />

Eyck, C. T. 317<br />

Eymery, A. 94<br />

F<br />

Fábián, K. T. 54, 444, 445<br />

Falasca, P. 124<br />

Fan, J. M. 110, 447<br />

Farcas, A. 468<br />

Farchi, M. 452<br />

Farina, F. 158<br />

Farkas, A. E. 147<br />

Fasano, A. 104<br />

Febbraio, A. M. 338<br />

Fehrenbacher, N. 76<br />

Fejérdy, P. 445<br />

Ferenci, T. 120<br />

Fernandes, M. 154<br />

Ferranti, P. 285<br />

Ferreira, A. 226<br />

Fevereiro, P. 220<br />

Fiaux, J. 28<br />

Figueira, E. 206<br />

Figueiredo, D. 33<br />

Filaković, P. 451<br />

Filaretova, L. 375<br />

Fildissis, G. 71<br />

Filipović, D. 138<br />

Finol, H. J. 343<br />

Fischer, G. 108<br />

Fishels<strong>on</strong>, Z. 314<br />

Flegar-Meštrić, Z. 291<br />

Flescher, E. 85<br />

Fleshner, M. 51<br />

Fliege, H. 417, 457<br />

Flom, G. 63<br />

Flügge, F. P. 316<br />

Fodor, F. 195<br />

Fodor, J. 195<br />

Foglio, E. 254<br />

F<strong>on</strong>taine, B. 359<br />

F<strong>on</strong>taine, J.-M. 92, 310<br />

F<strong>on</strong>tes, P. B. E. 104<br />

Forstrom, J. 414<br />

Forte, E. 137<br />

Fortin, C. 258<br />

Foster, L. A. 240<br />

Foster, T. M. 8<br />

Fouquaert, E. 205<br />

Franceschelli, S. 104<br />

Francini, A. 192<br />

Franco, M. 261<br />

Frank, E. 395<br />

Frederick, B. 176<br />

French, K. 54<br />

Frey, T. 51<br />

Frisch, F. 258<br />

490


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Fritah, S. 94, 97<br />

From<strong>on</strong>t-Racine, M. 63<br />

Frynta, D. 398<br />

Fu, Y. 98<br />

Fujimoto, M. 46<br />

Fukuoka, Y. 461<br />

Fukuyama, H. 431<br />

Fulop, T. 258<br />

Furfaro, A. L. 238<br />

Furutani, M. 57<br />

Futamura, Y. 70<br />

Füchtbauer, E.-M. 357<br />

Fürnrohr, B. G. 270<br />

G<br />

Gabai, V. L 313<br />

Gabriels<strong>on</strong>, K. 328<br />

Gaestel, M. 25<br />

Gal, H. 75<br />

Gallé, Á. 151, 214<br />

Galli, C. 114<br />

Gallyas, F. 19<br />

Gama, L 328<br />

Gama, M. J. 79<br />

Gammazza, A. M. 85<br />

Ganea, K. 157<br />

Gangoiti, P. 89<br />

Garab, Gy. 207<br />

Garcia-Arocena, D. 241<br />

García-Barreno, P. 340<br />

García-Fernández, M. J. 127<br />

García-Fruitós, E. 116<br />

Garcia-Medina, R. 106<br />

Gárd<strong>on</strong>yi, M. 227<br />

Garrido, C. 75, 87<br />

Garrido, M. R. 131, 476<br />

Gáspár, L. 228, 229<br />

Gaughan, J. B. 288<br />

Gauss, R. 115<br />

Gawr<strong>on</strong> A. 318<br />

Gehrman, M. 51, 313, 314<br />

Gekle, M. 108<br />

Gémes, K. 213, 221<br />

Genevaux, P. 28<br />

Gentil, B. 358<br />

Georgopoulos, C. 28, 356<br />

Geraci, F. 62<br />

Gereš, D. 291<br />

Gergely, H. 372<br />

Gertler, A. 33<br />

Gervasini, E. 130<br />

Gettemans, J. 307<br />

Gheysen, G. 205<br />

Ghirlando, R. 212<br />

Ghosh, G. J. 86<br />

Gianferretti, P. 39<br />

Giangrossi, M. 165<br />

Gianoli, E. 279<br />

Giffard, R. G. 246<br />

Gill, S. M. 240<br />

Gillan, V. 58<br />

Ginouvès, A. 106<br />

Ginsberg, B. A. 8<br />

Giros, B. 379<br />

Giuberti, G. 285<br />

Giudice, G. 62<br />

Giuffre, A. 137<br />

Giuliodori, M. A. 120, 165<br />

Givol, D. 75<br />

Glozman, R. 118<br />

Godočíková, J. 140<br />

Goksör, M. 159<br />

Goldgur, Y. 212<br />

Golub, N. V. 24<br />

Golubic, R. 462, 464, 466<br />

Gombos, T. 37<br />

Gomes-Junior, A. R. 190<br />

Gómez, A. V. 44<br />

Gómez, F. 417<br />

Gómez-Baena, G. 127<br />

Gómez-Muñoz, A. 89<br />

G<strong>on</strong>çalves, M. L. 194<br />

G<strong>on</strong>charova, N. D. 241<br />

G<strong>on</strong>ciarz, M. 82<br />

G<strong>on</strong>g, J. 49<br />

G<strong>on</strong>g, Q. 202<br />

G<strong>on</strong>os, S. E. 235<br />

G<strong>on</strong>zález, C. 361<br />

G<strong>on</strong>zález, E. M. 138<br />

G<strong>on</strong>zález, I. 295<br />

G<strong>on</strong>zález, M. 89<br />

G<strong>on</strong>zàlez-M<strong>on</strong>talbán, N. 116<br />

Gorman, J. 108<br />

Gornik, O. 174<br />

Graf, C. 274<br />

Graf, L. 151, 262<br />

Graham, M. Y. 196<br />

Graham, T. L. 196<br />

491


23-26 August 2007,<br />

Budapest, Hungary<br />

Granado, H. M. 89<br />

Grasselli, F. 127, 282<br />

Gratão, L. P. 190, 228<br />

Gray, J. P. 59<br />

Gray, R. S. 277<br />

Greenberg, T. J. 196<br />

Greensmith, L. 247<br />

Gregersen, N. 83, 356, 357, 359, 364<br />

Gregurek, R. 451<br />

Greguric, I. 53<br />

Gressens, P. 47<br />

Griesen, D. 190<br />

Grigolato, P. 254<br />

Grigoryan, G. Yu. 303<br />

Grimaldi, P. 124<br />

Groah, L. 461<br />

Groeneveld, A. B. J. 347<br />

Groisman, B. 102<br />

Gruenberg, J. 87<br />

Gruntenko, N. E. 273<br />

Gu, X. F. 283<br />

Gualerzi, C. O. 120, 165<br />

Guarneri, C. 61<br />

Guarneri, F. 61<br />

Guerriero, V. 322<br />

Guézennec, C. Y. 403<br />

Guisasola, M. C. 340<br />

Guisez, Y. 198<br />

Gulati, K. 432, 434<br />

Gumustas, K. M. 130, 132<br />

Gunasekara, S. D. 101<br />

Guo, J. 146<br />

Guo, J. C. 249, 250<br />

Guóth, A. 151, 213, 214, 221<br />

Gurtovyy, V. 387<br />

Gusev, B. N. 22, 26<br />

Gutiérrez, A. 480<br />

Gutiérrez, C. J. 156, 231<br />

Gutkevich, E. V. 164<br />

Guzhova, I. 350<br />

Guzhova, I. 265, 315, 362<br />

Gyorffy, Zs. 440, 471<br />

Györgyey, J. 151, 199, 223, 224<br />

H<br />

Habel, U. 366<br />

Hada, S. 402<br />

Hageman, J. 355<br />

Hagerman, P. J. 241<br />

Haikerwal, D. 416<br />

Halasz, J. 391<br />

Halász, K. 199<br />

Hal<str<strong>on</strong>g>for</str<strong>on</strong>g>d, C. 409<br />

Halienova, A. 160<br />

Haller, D. 99, 103<br />

Haller, J. 391<br />

Hambartsumyan, M. 69<br />

Hamilt<strong>on</strong>, M. J. 197<br />

Han, Y. 266<br />

Hänninen, A.-L. 161<br />

Hansen, A. M. 430<br />

Hansen, J. 356, 357, 359<br />

Hanss<strong>on</strong>, A.-S. 462<br />

Hanusova, V. 160<br />

Harris, J. 320<br />

Hart, W. G. 178<br />

Harwood, L. J. 89<br />

Hashimoto, K. 402<br />

Hatami, M. 463<br />

Haubner, R. 348<br />

Hauet-Broere, F. 335<br />

Hautzinger, M. 449<br />

Havaux, M. 187<br />

Hayashi, T. 80, 81<br />

Hayashida, N. 46<br />

Haybatollahi, M. S. 397<br />

Hayes, H. 361<br />

Hayes, T. B. 263<br />

Hecker, J. 386<br />

Hegedus, K. 470, 475<br />

Hegyi, Á. 292<br />

Hem, E. 469<br />

Hendershot, M. L. 12<br />

Henrikss<strong>on</strong>, E. 44, 268<br />

Herkovits, J. 255<br />

Hermine, O. 75<br />

Hezinova, V. 160<br />

Hideg, É. 189<br />

Hidehiko, K. 438<br />

Hightower, L. E. 253, 384<br />

Him, N. A. I. I. N. 58<br />

Hirabayashi, S. 134<br />

Hirakawa, H. 398<br />

Hirohashi, Y. 64<br />

Hir<strong>on</strong>iwa, N. 70<br />

Hirose, K. 64<br />

Hirose, M. 20<br />

Hirsch, C. 115<br />

492


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Ho, C. Y. 418<br />

H<str<strong>on</strong>g>of</str<strong>on</strong>g>man, P. 366<br />

Hohmann, S. 159<br />

Hohn, T. 222<br />

Hojo, M. 30<br />

Hołownia, A. 243<br />

Holsboer, F. 274<br />

Holstege, F. 162<br />

Holtick, U. 332<br />

Hooper, L. P. 66<br />

Hooper, L. Ph. 66<br />

Hoozemans, J. J. M. 364<br />

Hoppe, D. A. 92, 310<br />

Horemans, N. 198<br />

Hori, M. 402<br />

Horiuchi, S. 433, 445<br />

Horváth, Á. 292<br />

Horvath, G. 370<br />

Horváth, M. 203<br />

Horváth, V. G. 189, 194<br />

Horváth, Zs. 292<br />

Hovhanissyan, A. 69<br />

Hoyle, C. 37, 316<br />

Hsu, D.-Z. 349<br />

Hsu, D.-Z. 351<br />

Hsu, Y.-C. 342<br />

Hu, Y. 167<br />

Huang, B.-M. 349<br />

Huang, G.-H. 415<br />

Hubbard, E. A. 152<br />

Huff, G. R. 289<br />

Huff, W. E. 289<br />

Hunter-Lavin, C. 37, 53, 78, 140, 316, 376<br />

Hunyadi-Gulyás, É. 199<br />

Hurwitz Eller, N. 339<br />

Husse, B. 108<br />

Hutt, D. 354<br />

Huwiler, A. 153<br />

Hüttelmaier, S. 154<br />

Hyder, C. 47<br />

I<br />

Iancu, C. T. 319<br />

Iannac<strong>on</strong>e, R. 221<br />

Iguchi, T. 402<br />

Igyártó, B. Z. 292<br />

Iizuka, R. 20, 57, 60<br />

Ikeda, T. 298<br />

Ileri, E. 265<br />

Imai, T. 146<br />

Imoto, M. 70<br />

Ingel, F. I. 179<br />

Inouye, S. 46<br />

Iosefs<strong>on</strong>, O. 32<br />

Ippolitova, M. 362<br />

Ireland, H. E. 376<br />

Ireland, S. B. 117<br />

Irie, M. 406<br />

Irobi, J. 307, 310<br />

Irsigler, S. T. A. 104<br />

Ishigami, K. 446<br />

Ishige, K. 146<br />

Ishii, N. 20, 57, 81, 402<br />

Ishii, R. 402<br />

Ishikawa, R. 398<br />

Isikhan, V. 464<br />

Israel, A. 131, 132, 476<br />

Ito, K. 133<br />

Ito, M. 470<br />

Ito, N. 359<br />

Ito, S. 401<br />

Ito, Y. 146<br />

Ivanov, L. 230<br />

Ivanova, M. 341<br />

Ivanova, S. A. 164<br />

Iwahashi, C. K. 241<br />

Iwase, S. 298<br />

J<br />

Jäättelä, M. 76, 87, 312, 324<br />

Jacob, H. 449<br />

Jacquier, A. 63<br />

Jahng, J. W. 283<br />

Jakobsen, C. H. 66, 323<br />

Jakubowicz-Gil, J. 318<br />

Janský, L. 398<br />

Janssens, S. 310<br />

Jarosch, E. 115<br />

Jarvis, P.G. 208<br />

Jaschke, W. 348<br />

Jędryka- Góral, A. 425<br />

Jeliazkova, V. 328<br />

Jen, J. C. 342, 394<br />

Je<strong>on</strong>g, J. 181<br />

Jia, N. 143, 145<br />

Jin, I. 159<br />

Jin, Y. 12<br />

Joachim, R. 417<br />

493


23-26 August 2007,<br />

Budapest, Hungary<br />

Joëls, M. 271<br />

Johanness<strong>on</strong>, H. 221<br />

Johansen, J. 94<br />

Johns<strong>on</strong>, L. 481<br />

Johns<strong>on</strong>, L. J. 63<br />

Jolly, C. 94, 97<br />

J<strong>on</strong>es, G. 211, 376<br />

J<strong>on</strong>sdottir, I. H. 281<br />

Jóri, B. 223<br />

Jouanin, J. C. 403<br />

Jowett, J. 414<br />

Jung, J.-R. 126<br />

Jung, J.-Y. 126, 301<br />

Juretić, D. 291<br />

K<br />

Kader, A. 206<br />

Kagawa, N. 67<br />

Kaidoglou, A. 112<br />

Kakigi, R. 38<br />

Kalmar, B. 247<br />

Kalyanaraman, B. 295<br />

Kamiguchi, K. 64, 266, 268, 359<br />

Kampinga, H. H. 355<br />

Kanevski, L. M. 33<br />

Kang, Y. J. 252<br />

Kanitz, E. 289, 377<br />

Kanzaki, T. 57<br />

Kapahi, P. 152<br />

Kapustian, L. 387<br />

Kargl, D. 414<br />

Karikas, A. G. 280<br />

Karlss<strong>on</strong>, R. 60<br />

Karolkiewicz, J. 278<br />

Karpova, E. K. 273<br />

Kasaj, A. 443<br />

Kataky, R. 101<br />

Katamoto, S. 38<br />

Katsifis, A. 53<br />

Kaufer, D. 271<br />

Kaur, P. 180<br />

Kaushal, S. 353<br />

Kawai-Yamada, M. 189<br />

Kawasaki, Y. 367<br />

Kawashima, D. 68<br />

Kay, S. M. 433<br />

Kaye, D. 416<br />

Kaznacheeva, A. 362<br />

Keegan, K. 447<br />

Kellner,W. 354<br />

Kelly, W. J. 354<br />

Kemp, P. J. 106<br />

Kempa, K. 135<br />

Kendall, J. 454<br />

Keresztes, Á. 205<br />

Kerr, R. L. 418<br />

Keßler, M. S. 395<br />

Khan, Z. A. 294<br />

Khochbin, S. 97<br />

Khomenko, T 373, 374<br />

Khozhaenko, Yu. 387<br />

Ki, L. T. 300<br />

Kidoaki, S. 163<br />

Kidokoro, S.-I. 20<br />

Kim, D. Y. 203<br />

Kim, H. J. 18, 155, 181<br />

Kim, H.-Y. 126<br />

Kim, I.-S. 159<br />

Kim, J. 301<br />

Kim, J. J. 392<br />

Kim, J.-H. 157<br />

Kim, S. 155<br />

Kim, S. C. 99<br />

Kim, S. J. 203<br />

Kim, S.-A. 23, 45<br />

Kim, Y. H. 81, 181, 301<br />

Kimoto, M. 298, 398<br />

Kim-Parker, Y. 263<br />

Kimsa, E. 135<br />

Kinev, A. 362<br />

Kinnunen, P. 87<br />

Kinzie, J. D. 454<br />

Király, A. 371<br />

Kiraly, D. 360<br />

Király, I. 212<br />

Kiraly, M. A. 410<br />

Kirkegaard, T. 324<br />

Kirkegaard-Sørensen, T. 76, 87, 312<br />

Kirtikara, K. 129, 136<br />

Kis, P. 189, 194<br />

Kittinska, J. 345<br />

Klagas, I. 112<br />

Klapp, B. F. 417, 429, 430, 432, 457<br />

Klasa-Mazurkiewcz, D. 320<br />

Klein Koerkamp, E. 335<br />

Kleszczyński, J. 422<br />

Knepper, C. 197<br />

Knezevic, B. 462, 464, 466<br />

494


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Knezevic, P. 462<br />

Kobayashi, M. 367<br />

Kochetov, A. V. 214<br />

Kocsy, G. 224<br />

Kodavayur, S. 262<br />

Koerkamp, G. M. 162<br />

Kohen, R. 101<br />

Kohli, M. 395<br />

Koizumi, K. 275<br />

Kolbert, Zs. 294<br />

Kollias, G. 103<br />

Kolobov, A. A. 68<br />

Kolodyazhnaya, Ya. S. 214<br />

Komarova, E. 315<br />

Komarova, M. L. 214<br />

K<strong>on</strong>arska, M. 425<br />

K<strong>on</strong>cz, Cs. 203, 217<br />

K<strong>on</strong>cz, Zs. 203<br />

K<strong>on</strong>g, R. Y. C. 111<br />

K<strong>on</strong>rad, Z. 212<br />

K<strong>on</strong>stadoulakis, M. M. 261<br />

K<strong>on</strong>stantinov, A. A. 137<br />

Kopp, M. 405, 413, 435, 440,<br />

471, 474, 475<br />

Korcsmáros, T. 308<br />

Kor<strong>on</strong>a, R. 180<br />

Kosuge, Y. 146<br />

Kotlarz, A. 334<br />

Kotlyarov, A. 25<br />

Koulov, A. 354<br />

Kouspou, M. 320<br />

Kovács, A. I. 308<br />

Kovacs, C. 414<br />

Kovács, E. 205<br />

Kovacs, K. 255<br />

Kovács, L. 64<br />

Kovács, M. 404<br />

Koval, V. S. 214<br />

Kovalenko, E. I. 33, 83<br />

Kovalitskaya, Y. A. 68<br />

Kowatsch, S. 414<br />

Kranner, I. 191<br />

Krause, M. 445<br />

Krause, W. R. 444, 445<br />

Krawiec, A. 82<br />

Krebs, N. 290<br />

Kregzdyte, R. 230<br />

Krishnan, J. 360<br />

Kristensen, S. T. 481<br />

Kritis, A. 112<br />

Krizbai, I. A. 147<br />

Kroemer, G. 87<br />

Krokan, E. H. 66<br />

Kroupskaya, I. 387<br />

Krstic-Dem<strong>on</strong>acos, M. 423<br />

Kubinova, R. 437<br />

Kucharskyij, V. 373<br />

Kuczynska-Wisnik, D. 123, 124<br />

Kukuljan, M. 44<br />

Kullisaar, T. 284<br />

Kumar, A. 139<br />

Kumar, M. 139<br />

Kumei, Y. 298, 398<br />

Kuo, Y.-M. 399<br />

Kurachi, M. 367<br />

Kurganov, B. I. 23<br />

Kurosaka, M. 38<br />

Kurotaki, T. 268<br />

Kurz, R. W. 414<br />

Kuštrimović, N. 264<br />

Kutalová, H. 398<br />

Kutomi, G. 266<br />

Kuwagata, M. 377<br />

Kuwajima, K. 57<br />

Kuzminsky, G. 342<br />

Kvetnansky, R. 172<br />

Kwak, S. 159<br />

Kwiatkowska, J. 123, 124<br />

Kw<strong>on</strong>, S.-O. 159<br />

Kyriakidis, E. 461<br />

L<br />

La Rocca, G. 158<br />

Labun, J. 302<br />

Lacetera, N. 342<br />

Ladenstein, R. 43<br />

Lagae, L. 379<br />

Lahijani, M. S. 263<br />

Lahouel, M. 142<br />

Laiho, A. 44<br />

Lajevic, M. 262<br />

Lakhotia, C. S. 95<br />

Lambert, E. 414, 416<br />

Lambert, G. 410, 414, 416<br />

Lambert, H. 22, 148<br />

Lamperti, L. 295<br />

Landgraf, R. 393, 395<br />

495


23-26 August 2007,<br />

Budapest, Hungary<br />

Landry, J. 17, 22, 90, 92,<br />

147, 148, 310<br />

Lanneau, D. 75<br />

Lapi, E. 75<br />

Lapinska-Szumczyk, S. 320<br />

Lapointe, P. 354<br />

Larbi, A. 258<br />

Larkindale, J. 11<br />

Larss<strong>on</strong>, L.-G. 315<br />

Laskowska, E. 123, 124<br />

Lásztity, D. 218, 223<br />

Lau, B. 469<br />

Lauc, G. 174, 451<br />

Laufer, H. 400<br />

Laugharne, J. D. E. 453<br />

Laurent, P. 361<br />

Lawrence, D. A. 251<br />

Le Mouël, A. 40, 47<br />

Lea, J. P. 190<br />

Lecchi, C. 342<br />

Lechner, M.C. 79<br />

Lederkremer, Z. G. 100, 102<br />

Lee, C.-H. 301<br />

Lee, H. 159<br />

Lee, Je. 81<br />

Lee, Ju. 384<br />

Lee, J. H. 95<br />

Lee, J.-H. 283<br />

Lee, K.-J. 81, 155, 181<br />

Lee, S. A. 98<br />

Lee, U. 11<br />

Lee, Y. H. 203, 344<br />

Leen, B. 304<br />

Lefler, K. K. 292<br />

Lehr, C. 55<br />

Leidhold, C. 115, 118<br />

Lein<strong>on</strong>en, I. 211<br />

Leitman, J. 102<br />

Lendvai, Á. 224<br />

Lentzen, G. 361<br />

Le<strong>on</strong>i, F. 37<br />

Le<strong>on</strong>i, F. 316<br />

Le<strong>on</strong>i, F. 376<br />

Lesauskaite, V. 230<br />

Leszczyński, B. 199<br />

Leszek, J. 422<br />

Lévai, L. 229<br />

Levenstein, S. 417<br />

Levitsky, D. I. 26<br />

Lewitus, G. M 260<br />

Li, H. 143, 145<br />

Li, J. 98<br />

Li, J.-P. 163<br />

Li, P. 202<br />

Li, Q.-h. 143, 145<br />

Li, Y.-H. 351<br />

Liao, M. H. 349<br />

Liaw, W. J. 351<br />

Lichtenthaler, K. H. 208<br />

Liebl, C. 157<br />

Liezmann, C. 429<br />

Lima, A. 206<br />

Lima, A. B. F. 458<br />

Lima, C. F. 284<br />

Lima-Costa, T. 162<br />

Lin, Y.-W. 322<br />

Lindberg, S. 206<br />

Linder, S. 208<br />

Lindner, R. A. 24<br />

Lindsey, K. 197<br />

Lindström, B. 173<br />

Link, D. C. 152<br />

Link, D. C. 365<br />

Linke, S. 108<br />

Lipinska, B. 61, 320, 334<br />

Liscia, P. 403<br />

Lithgow, J. G. 152, 235, 239, 240<br />

Liu, H. L. 349<br />

Liu, M.-Y. 349<br />

Liu, M.-Y. 351<br />

Liu, Q. 267<br />

Liu, W. 43<br />

Liu, X. 385<br />

Liu, Y. 109, 112<br />

Liu, Y.-F. 394<br />

Logvinova, N. 438<br />

Loose, J. 416<br />

López, J. 138<br />

López-Cavanillas, M. 364<br />

López-Lozano, A. 127<br />

Lorenzini, G. 192<br />

Loriaux, D. L. 454<br />

Lorigados, L. 138<br />

Los<strong>on</strong>cz, E. 413<br />

Loureiro, J. 226<br />

Lozano, J. 316<br />

Lu, M. 346<br />

Lu, X. 75<br />

496


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Lubach, R. G. 428<br />

Lubsen, H. N. 18<br />

Lucot, B. J. 419<br />

Lukacs, G. L. 118<br />

Lukács, N. 199<br />

Łukasik, I. 199<br />

Lukaszewska, A. J. 225<br />

Lundemo, G. A. 66<br />

Luria, A. 318<br />

Lutfalla, G. 63<br />

Lutz, C. K. 401<br />

Lyahyai, J. 361<br />

Lynes, M. A. 251<br />

Ly<strong>on</strong>s, B. L. 251<br />

M<br />

Ma, S. 202<br />

Macario, J. L. A. 30, 85, 158, 307, 311<br />

Maccari, S. 383<br />

Mach, M. 419<br />

Machatschke, I. H. 459<br />

MacRae, H. T. 167<br />

Madách, K. 37<br />

Madec, L. 481<br />

Maeda, M. 60, 363<br />

Maeder, L. D. 311<br />

Maestr<strong>on</strong>i, G. 428<br />

Magno, F. 158<br />

Magyar, A. 292<br />

Mahadik, S. 369<br />

Mahajan, M. I. 76<br />

Mah<strong>on</strong>, G. C. 155<br />

Mai, A. 223<br />

Maitland, K. 58<br />

Majid Fekri 213<br />

Makara, G. 375<br />

Makarov, V. 242<br />

Makarova, E. N 259<br />

Makarova, M. N. 242<br />

Makarow, M. 161<br />

Maki, K. 57<br />

Makino, T. 244<br />

Mald<strong>on</strong>ado, R. 482<br />

Malik, A. 413<br />

Mallik, M. 95<br />

Malmberg, B. 465<br />

Maltseva, V. N. 323<br />

Malvar, A. R. 200<br />

Malzahn, D. 316<br />

Malyutina, S. 437<br />

Mambula, S. S. 49<br />

Mandel, G. 44<br />

Mandrekar, P. 328<br />

Maneerat, Y. 129, 136<br />

Manitašević, S. 215<br />

Manninger, K. 199<br />

Mano, T. 298<br />

Manov, I. 319<br />

Mao, C. 98<br />

Marcelis, M. 366<br />

Marchi, P. 165<br />

Marcos-Carcavilla, A. 361<br />

Marcus Ising, M. 447<br />

Marek, V. M. 209<br />

Margulis, B. 350<br />

Margulis, B. 265, 315, 362<br />

Marinari, U. M. 238<br />

Marinho, H. S. 152<br />

Markel, A. L. 424<br />

Markossian, K. A. 23, 24<br />

Markov, D. I. 26<br />

Marmot, M. 437<br />

Marova, I. 160<br />

Marques da Silva, J. 220<br />

Marshall, C. H. 277<br />

Marshall, J. M. 78<br />

Martasek, P. 295<br />

Martin, J. S. 77<br />

Martin, L. J. 92, 310<br />

Martín-Burriel, I. 361<br />

Martínez-Al<strong>on</strong>so, M. 116<br />

Martínez-Martos, J. M. 381<br />

Martín-G<strong>on</strong>zález, A. 156, 231<br />

Martins, L. L. 194<br />

Martos, T. 404, 413, 440, 474<br />

Marusov, G. 251<br />

Masnikosa, S. P. 429, 430, 432<br />

Mass, J. C. 44<br />

Mastorakos, G. 457<br />

Masullo, M. 124<br />

Matec, L. 464, 466<br />

Mathew, J. S. 447<br />

Mathis<strong>on</strong>, Y. 343<br />

Matić, G. 215, 253, 384<br />

Matringe, M. 187<br />

Matsuda, T. 163<br />

Matsukawa, K. 133<br />

Mattanovich, D. 116<br />

497


23-26 August 2007,<br />

Budapest, Hungary<br />

Matthews, S. G. 410<br />

Mattner, F. 53<br />

Matuszewska, E. 123, 124<br />

Matyska, M. 419<br />

Mayas, M. D. 381<br />

Mayer, M. 28<br />

Mayerl, C. 348<br />

Mayora, S. 343<br />

Mazure, N. 106<br />

Mazzafera, P. 190<br />

McAlind<strong>on</strong>, T. 279<br />

McColl, G. 152<br />

McCollum, A. 317<br />

McGl<strong>on</strong>e, J. J. 290<br />

Mckenna, W. J. 387<br />

Medvedeva, D. N. 55<br />

Medzihradszky, K. 199<br />

Meehan, L. 263<br />

Melander, O. 435<br />

Melchers, I. 55<br />

Menut, C. 239<br />

Merisalu, E. 454<br />

Merkel, L. 149<br />

Meskauskiene, R. 188<br />

Messlik, A. 99<br />

Mészáros, I. 228<br />

Meyer, A. 29<br />

Meyer, J. S. 401<br />

Mezger, V. 40, 47<br />

Micali, E. 420<br />

Michalak, E. 278<br />

Michalianou, G. 411<br />

Michaud, S. 18<br />

Micheli, W. 279<br />

Miesfeld, R. 322<br />

Mikelsaar, M. 284<br />

Mikhailov, A. 47<br />

Mikhaylov, I. V. 166<br />

Mikics, E. 391<br />

Miklósi, Á. 292<br />

Miller<strong>on</strong>, S. R. 76<br />

Milosevic, M. 462, 464, 466<br />

Miñano, J. F. 482<br />

Minassian, S. M. 303<br />

Minotti, S. 358<br />

Miskolczi, P. 224<br />

Mitić, K. 264<br />

Miyado, T. 297<br />

Miyashita, T. 134<br />

Miyauchi, F. 466<br />

Mizera, A. 47<br />

Mizusawa, N. 287<br />

Mizushima, T. 45<br />

Moazami-Goudarzi, K. 361<br />

Mogosan, C. 296<br />

Moilanen, I. 87<br />

Mojžiš, M. 302<br />

Mok, H. O. L. 111<br />

Molina, H. 178<br />

Molinari, M. 114<br />

Molnár, G. 203<br />

Molvarec, A. 37<br />

M<strong>on</strong>nazzi, P. 275<br />

M<strong>on</strong>talbano, A. 85<br />

M<strong>on</strong>teiro, C. C. 228<br />

M<strong>on</strong>tes, D. G. 410<br />

Mo<strong>on</strong>, I. 159<br />

Mo<strong>on</strong>, Y. W. 283<br />

Morales, L. 138<br />

Morandini, F. 254<br />

Morano, K. 29<br />

Morenkov, O. S. 323<br />

Morimoto, I. R. 236, 354<br />

Morishige, K. 67<br />

Morozumi, M. 81<br />

Morris, N. J. 332<br />

Morrow, G. 18<br />

Mos, L. 472<br />

Moskovich, O. 314<br />

Mota, A. M. 194<br />

Motoyama, K. 134<br />

Mourato, M. P. 194<br />

Mourl<strong>on</strong>, V. 379<br />

Moutet, M. 239<br />

Mozaffari, V. 217, 219<br />

Mujica-Parodi, L. R. 176<br />

Multh<str<strong>on</strong>g>of</str<strong>on</strong>g>f, G. 313, 314, 316<br />

Mult<str<strong>on</strong>g>of</str<strong>on</strong>g>f, G. 51<br />

Muneoka, K. 377<br />

Mun<str<strong>on</strong>g>for</str<strong>on</strong>g>d, S. R. 346<br />

Muñoz, R. 216<br />

Muppala, V. 316<br />

Murase, Y. 60<br />

Murashko, D. A. 83<br />

Muratore, N. 328<br />

Murayama, H. 401<br />

Mushynski, E. W. 358<br />

Mustajbegovic, J. 462, 464, 466<br />

498


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Müller, C. T. 106<br />

Müller, M. B. 157<br />

Müller, O. 445<br />

Müller, R. 153, 270<br />

Myoungho, H. 424<br />

Myrianthefs, P. 71<br />

N<br />

Nadeau, J. P. 147<br />

Nadeau, S. I. 148<br />

Nagai, H. 297<br />

Nagamune, H. 122<br />

Nagaraj, H. R. 20<br />

Naghavi, M. 479<br />

Nagley, P. 79<br />

Nagy, Z. 210<br />

Nagyőszi, P. 147<br />

Nagy-Tanaka, E. 470<br />

Naito, H 38, 134<br />

Nakai, A. 40, 46<br />

Nakajima, K. 300, 446<br />

Nakamoto, H. 90<br />

Nakamura, K. 457<br />

Nakanishi, K. 64, 359<br />

Nali, C. 192<br />

Nam, M. H. 203<br />

Nard<strong>on</strong>e, A. 342<br />

Narkiewicz, J. 320<br />

Nasab, A. R. 216<br />

Navamani, D. 460<br />

Navolotskaya, E. V. 68<br />

Naylor, A. S. 281<br />

Neckers, L. 326<br />

Neculai, D. K. 470<br />

Neculai, K. 413, 440, 471<br />

Nekrasov, A. N. 33<br />

Nemoto, K. 402<br />

Nepomnaschy, P. 456<br />

Nestor, K. E. 289<br />

Netterstrøm, B. 430<br />

Neumeister, A. 448<br />

Nguyen, M. T. 54, 445<br />

Nguyen-Michel, S. 282<br />

Ni, M. 98<br />

Nici<str<strong>on</strong>g>for</str<strong>on</strong>g>ovic, A. 423<br />

Niehaus, I. 350<br />

Nielsen, L. 481<br />

Nielsen, N. M. 356, 357, 359<br />

Nielsen, N. R. 481<br />

Niinuma, T. 398<br />

Nijs, I. 210<br />

Niki, E. 136, 297<br />

Nilss<strong>on</strong>, M. P. 434, 441<br />

Nimmo, A. M. 277<br />

Nino, D. 51<br />

Nishijima, K. 367<br />

Nollen, E. 13<br />

Noorwez, M. S. 353<br />

Nort<strong>on</strong>, M. C. 403<br />

Nova, D. 295<br />

Novak, M. A. 401<br />

Nováková, M. 398<br />

Novy, B. 414<br />

Nowak, A. 278<br />

Nudler, E. 95<br />

Nyitrai, P. 205<br />

Nylandsted, J. 76, 87<br />

O<br />

O’Brien, R. E. 177<br />

O’Callaghan, C. 313<br />

O’Neill, H. 11<br />

Ober, M. 468<br />

Odell, M. 36<br />

Odjakova, M. 219<br />

Ogawa, T. 377<br />

Oglesbee, M. 299<br />

Ogura, Y. 38<br />

Oh, J.-K. 157<br />

Ohashi, N. 134<br />

Ohmori, S. 406<br />

Ohshima, S. 268<br />

Ohtani, Y. 467<br />

Ohtsuka, K. 68<br />

Ojaimi, J. 320<br />

Okamura, H. 431, 433, 444<br />

Okiy<strong>on</strong>eda, T. 118<br />

Oksbjerg, N. 83<br />

Okuda-Shimizu, Y. 12<br />

Olff, M. 448<br />

Oliveira, C. A. 458<br />

Oliveira-Marques, V. 152<br />

Olsen, A. 240<br />

Olss<strong>on</strong>, T. 281<br />

Onesto, E. 357<br />

Onikienko, S. 350<br />

Onikienko, S. 265<br />

Ooe, Y. 398<br />

499


23-26 August 2007,<br />

Budapest, Hungary<br />

Ørbaek, P. 465<br />

Orendáč, M. 302<br />

Orendáčová, J. 302, 378<br />

Orman, R. 275<br />

Ørnt<str<strong>on</strong>g>of</str<strong>on</strong>g>t, T. F. 324<br />

Orosz, A. 41<br />

Ortega, R. 231<br />

Osapay, K. 373<br />

Østbye, T. 403<br />

Ostenfeld, M. S. 324<br />

Otta, E. 287<br />

Otten, W. 377<br />

Otters<strong>on</strong>, F. M. 332<br />

Oules, B. 325<br />

Ouyang, Y. 246<br />

Óvári, M. 205<br />

Owen, S. J. 34, 35<br />

Ozturk, Z. 130, 134<br />

Öhlin, B. 435<br />

Örsal, A. 432<br />

P<br />

Painter, J. A. 26<br />

Pajak, A. 437<br />

Páldi, E. 218<br />

Palme, R. 398<br />

Palmieri, B. 141<br />

Paloheimo, M. 178<br />

Pandey, A. 178<br />

Pandey, N. 222<br />

Panossian, A. 69, 242<br />

Panwar, M. 139<br />

Paolocci, N. 328<br />

Papamichael, K. 161<br />

Papandreou, V. 71<br />

Papp, D. 105<br />

Paredes, A. J. 162<br />

Park, C. B. 361<br />

Park, S. 266<br />

Park, S.-K. 126<br />

Parkes, L. T. 155<br />

Parthimos, T. 280<br />

Parvu, A. E. 296<br />

Parvu, M. 296<br />

Pastorello, M. 131<br />

Patacchioli, F. R. 275<br />

Paterlini-Brechot, P. 325<br />

Patriarca, S. 238<br />

Paul, J. R. 110<br />

Pauly, S. 290<br />

Pavlinić, D. 451<br />

Pawlik, W. 82<br />

Peasey, A. 437<br />

Pecoraro, C. N. 8<br />

Peet, D. J. 108<br />

Pehlivanoglu, B. 265<br />

Pellegrini, E. 192<br />

Perdrizet, G. A. 253<br />

Pereira-Wils<strong>on</strong>, C. 284<br />

Peres, L. E. P. 228<br />

Pérès, M. 403<br />

Pérez-Coll, C. S. 255<br />

Perišić, T. 253<br />

Perrot, M. 162<br />

Perss<strong>on</strong>, R. 465<br />

Petelenz, E. 159<br />

Peters, E. 417<br />

Peters, E. M. J. 429, 430, 432<br />

Petre, I. 47<br />

Petr<strong>on</strong>ini, G. P. 78<br />

Petrozza, A. 221<br />

Pfeilschifter, J. 153<br />

Phan, H.-A. 460<br />

Piairo, P. C. 284<br />

Picariello, G. 285<br />

Piccinini, C. 279<br />

Piérard, C. 403<br />

Piersiak T. 318<br />

Pierzchalski, P. 82<br />

Pikhart, H. 437<br />

Pilaczyńska-Szcześniak, Ł. 278<br />

Pilzer, D. 314<br />

Pin, S. 328<br />

Pincus, M. 457<br />

Pinto, A. P. 194<br />

Pinto, S. 417<br />

Pissinati, A. 458<br />

Pistorius, A. 443<br />

Pivovarova, A. V. 26<br />

Plasterk, R. 13<br />

Plitzko, D. 347<br />

Plutner, H. 354<br />

Poanta, L. 468<br />

Pockley,A. G. 314<br />

Podea, D. 472<br />

Pokora, I. 135<br />

Pokora, Z. 135<br />

Polek, B. 140<br />

500


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Poletti, A. 357 Ralph, T. 418<br />

P<strong>on</strong>, L. C. 165 Ramachandran, S. 125<br />

Pooggin, M. 222 Ramírez-Exposito, M. J. 381<br />

Po<strong>on</strong>, S. 50 Ramkumar, K. M. 344<br />

Portolos, J. 71 Rampelt, H. 28<br />

Pós, V. 199 Rangel, A. O. 127<br />

Pospieszna, B. 278 Ransy, V. 380<br />

Potter, S. C. 354 Rasmussen, A. A. 94<br />

Pour, A. T. 216, 285 Ratajczak, E. 123<br />

Pourzitaki, C. 112 Rath, N. C. 289<br />

Pouysségur, J. 106 Rattan, S. 236<br />

Pozsgai, E. 19 Rau, S. 108<br />

Prakash, N. S. 222 Rauschenbach, I. Yu. 273<br />

Preoteasa, D. 47 Rautio, A. 440<br />

Price, J. 320 Ravindranath, B. 176<br />

Pritchard, W. H. 191 Rayner, K. 177<br />

Prohászka, Z. 37 Raynes, A. D. 322<br />

Pr<strong>on</strong>zato, M. A. 238 Rea, I. M. 237<br />

Pröls, F. 149 Rechavi, G. 75<br />

Puppe, B. 289, 377 Reddy, P. V. J. 57<br />

Puthur, J. T. 207 Redina, O. E. 424<br />

Putics, Á. 105 Reichwald, U. 449<br />

Puzserova, A. 341 Reid, M. L. 200<br />

Pütz, B. 274 Rein, T. 157<br />

Pyun, K.-H. 157 Rekas, A. 24<br />

Rem<strong>on</strong>delli, P. 104<br />

Q<br />

Ren, Z. 385<br />

Qian, L. 385 Renna, M. 104<br />

Qiu, W. 101 Rérole, A.-L. 87<br />

Qiu, Z. 167 Rey, P. 187<br />

Quarantelli, A. 130 Ribbene, A. 85<br />

Quero, Z. 343 Ribeil, J.-A. 75<br />

Quiles, M. J. 216 Riccardi, D. 106<br />

Righi, F. 130<br />

R<br />

Rigó, J. 37<br />

Raček, Ľ. 378 Riley, C.<br />

454<br />

Račeková, E. 302, 378 Ritchie, J. 321<br />

Rácz, I. 218, 223 Riverón, G. 138<br />

Radeke, H. H. 153 Rizzuto, R. 325<br />

Radojčić, B. M. 138, 423 Rjabovola, N. 192<br />

Rad<strong>on</strong>ic, A. 462 Robberecht, W. 307, 310, 360<br />

Rad<strong>on</strong>s, J. 313 Roberts, S. 34, 35, 39<br />

Raeymaekers, T. 198 Roberts<strong>on</strong>, P. 338<br />

Rafiee, P. 332 Robins<strong>on</strong>, C. V. 24, 26<br />

Rai, L. C. 182 Rodella, L. F. 254<br />

Raimo, G. 124 Rodrigues, E. 79<br />

Raim<strong>on</strong>do, G. 420 Rodrigues, Z. 382<br />

Rajaguru, P. 344 Rodriguez, A. P. M. 228<br />

Rajwanshi, R. 222 Rodriguez, E. 226<br />

501


23-26 August 2007,<br />

Budapest, Hungary<br />

Rodriguez, L. S. 261<br />

Rodríguez, M. 51<br />

Rogers, A. 152<br />

Rohde, M. 312, 324<br />

Rolim, M. 113<br />

Rom, S. 212<br />

Romanova, A. V. 214<br />

Romero, I. A. 147<br />

R<strong>on</strong>, E. 102<br />

R<strong>on</strong>dinelli, E. 154<br />

Rose, M. 457<br />

Rosenthal, B. S. 449<br />

Ross, A. O. 237<br />

Rossi, A. 39<br />

Rossi, A. M. 468<br />

Rossi, M. L. 228<br />

Roth, A. 87<br />

Rothman, S. 460<br />

Roux, D. 106<br />

Rozhko, O. 387<br />

Ruecke, M. 457<br />

Ruiz, P. A. 99<br />

Rujano, M. 355<br />

Ruocco, M. R. 124<br />

Rusmini, P. 357<br />

Russo, J. 481<br />

Rutlege, A. 101<br />

Ryabenko, D. 387<br />

Ryadovaya, A. L. 164<br />

Rydstedt, W. L. 411<br />

Ryselis, S. 230<br />

Ryu, J. 361<br />

Ryu, S.-Y. 301<br />

Ryu, V. 283<br />

Rzepka, R. 55<br />

Rzeski, W. 318<br />

S<br />

Sa, C. M. 284<br />

Saadoun, P. A. 443<br />

Sadauskiene, I. 230<br />

Sadlish, H. 28<br />

Sadovnikov, V. B. 68<br />

Safr<strong>on</strong>ova, V. G. 323<br />

Sagace, G. 221<br />

Saganová, K. 302<br />

Sahakyan, S. G. 303<br />

Sahi, C. 29<br />

Sahlan, M. 57<br />

Saito, Y. 136<br />

Sakakibara, H. 134<br />

Sakakima, J. 479<br />

Sakamoto, Y. 406<br />

Sakatani, K. 446<br />

Sakiko, F. 426<br />

Sak<strong>on</strong>o, M. 363<br />

Salam<strong>on</strong>e, M. 62<br />

Salavecz, Gy. 470, 471<br />

Salleh, A. 418<br />

Salom<strong>on</strong>e, G. 124<br />

Samarth, M. R. 139<br />

Sampayo, N. J. 240<br />

Sanchez, M. J. 340<br />

Sánchez-López, M. P. 474<br />

Sandberg, S. 420<br />

Sandh<str<strong>on</strong>g>of</str<strong>on</strong>g>f, K. 87<br />

Sándor, K. B. G. 243<br />

Sandor, Zs. 373, 374<br />

Sandoya, G. 200<br />

Sanina, N. 92<br />

Santiago, R. 200<br />

Santini, S. E. 127, 130, 282<br />

Santoro, M. G. 39<br />

Santos, A. S. M. 162<br />

Santos, C. 226<br />

Santos, D. 220<br />

Sapozhnikov, A. M. 33, 83<br />

Sarid, O. 334<br />

Sarin, N. B. 222<br />

Sarti, P. 137<br />

Sartor, R. B. 99<br />

Sartorelli, P. 342<br />

Sartori, S. B. 393<br />

Saruta, J. 300<br />

Sárvári, É. 227, 228, 229<br />

Sass, L. 189<br />

Sato, C. 300<br />

Sato, H. 402<br />

Sato, N. 64, 266, 268, 359<br />

Sato, S. 300<br />

Sau, D. 357<br />

Savicka, M. 192<br />

Savickas, A. 230<br />

Savickiene, N. 230<br />

Savoy, R. 176<br />

Sblendorio, V. 141<br />

Schade, F. U. 347<br />

Schanaider, A. 113<br />

502


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Scheideler, A. 290<br />

Scheper, W. 364<br />

Schick, C. 55<br />

Schieber, M. 354<br />

Schilling, D. 314<br />

Schiltz, L. 453<br />

Schlink, K. 182<br />

Schmidt, A. J. M. 396<br />

Schmidt, M. V. 157<br />

Schmidt, S. P. 364<br />

Schmitt, E. 87<br />

Schneider, M. 290<br />

Schønberg, S. A. 66, 102, 323<br />

Schönenberg, M. 449<br />

Schuemann, K. 103<br />

Schulpis, H. K. 280<br />

Schulz, N. 103<br />

Schülke, J. 157<br />

Schwager, F. 28<br />

Schwartz, M. 260<br />

Schweizer, P. 224<br />

Schwenndimann, L. 47<br />

Scire, A. 61<br />

Sc<strong>on</strong>zo, G. 62<br />

Scriven, P. 321<br />

Searle, A. 295<br />

Sečenji, M 151, 224<br />

Seguin, S. 22<br />

Seit-Nebi, S. A. 22<br />

Sekine-Ichinoseki, N. 38<br />

Senese, L. 358<br />

Sensi, S. 252<br />

Seppä, L. 161<br />

Serrano, C. 361<br />

Serrano, M. 361<br />

Sert, S. 130<br />

Šešlija, D. 193<br />

Sevillano, C. 482<br />

Shabtay, A. 302<br />

Shadrin, N. 212<br />

Shahriaripour, R. 217<br />

Shamovsky,I. 95<br />

Shan, Z. 150<br />

Sharma, S. 57<br />

Sharp, C. A. 34, 35<br />

Shatalin, K. 95<br />

Sheffy, B. 422<br />

Shemetov, A. A. 22<br />

Shen, Y. 12<br />

Shenkman, M. 100<br />

Shenoy, K. A. 86<br />

Sherman, M. Y. 313<br />

Shevchenko, A. Yu. 259<br />

Shibusawa, M. 446<br />

Shields, K. 414<br />

Shim, I. 157<br />

Shim, J.-Y. 266<br />

Shimada, E. 467<br />

Shimizu, M. 20, 60<br />

Shimizu, T. 244<br />

Shimizu, Y. 12<br />

Shimohama, S. 359<br />

Shimoi, K. 134<br />

Shimura, M. 398<br />

Shinozaki, K. 201<br />

Shioda, S. 377<br />

Shirani, H. 216<br />

Shirasu, M. 300<br />

Shkolnik, D. 212<br />

Shkute, N. 192<br />

Shrivastava, U. 439<br />

Shumny, V. K. 214<br />

Shustova, O. A. 83<br />

Sidorik, L. 387<br />

Siegrist, J. 436<br />

Sik, A. 22<br />

Silva, J. A. 131<br />

Silva, J. P. 128<br />

Silva, M. R. 162<br />

Silva, P. C. 113<br />

Silva, R. 154<br />

Sime<strong>on</strong>i, S. 275<br />

Simola, M. 161<br />

Sim<strong>on</strong>, M.-P. 106<br />

Sim<strong>on</strong>, S. 92, 310<br />

Sim<strong>on</strong>ini, F. 357<br />

Singewald, N. 393<br />

Singh, M. 54<br />

Sioga, A. 112<br />

Sirek, A. 410<br />

Sist<strong>on</strong>en, L. 39, 44, 47, 268<br />

Sitia, R. 99<br />

Skorko-Gl<strong>on</strong>ek, J. 61<br />

Slabas, R. A. 197<br />

Slagsvold, J. E. 102<br />

Slotta-Bachmayr, L. 290<br />

Smalinskiene, A. 230<br />

Smerd<strong>on</strong>, S. 36<br />

503


23-26 August 2007,<br />

Budapest, Hungary<br />

Smith, D. J. 79<br />

Smits, T. 379<br />

Smolenskaya, S. E. 424<br />

Sobiecka-Szkatula, A. 61<br />

Sokolovskaya, A. 64<br />

Sokolovskya, A. 359<br />

Soldà, T. 114<br />

Solti, Á. 228, 229<br />

Sommer, T. 115<br />

S<strong>on</strong>g, J.-H. 301<br />

S<strong>on</strong>g, J.-Y. 266<br />

S<strong>on</strong>g, L. 143, 145<br />

S<strong>on</strong>g, T.-b. 145<br />

Soós, V. 223<br />

Soreq, H. 172<br />

Soto, S. M. 121<br />

Söderström, I. 281<br />

Sönmez, H. 134<br />

Sőti, Cs. 54, 105, 365<br />

Spira, B. 120<br />

Spruijt-Metz, D. 282<br />

Squadrito, G. 420<br />

Srivastava, A. K. 182<br />

Stacchiotti, A. 254<br />

Stach, C. 270<br />

Stagg, S. 354<br />

Stanojević, S. 264<br />

Stauder, A. 404, 413<br />

Stefanadis, C. 261<br />

Stein, N. 224<br />

Steinem, C. 51, 314<br />

Stenner, E. 279<br />

Stensgard, B. 317<br />

Stephano, A. Q. 255<br />

Steptoe, A. 9<br />

Sterlemann, V. 157<br />

Sternberg, E. M. 258<br />

Sterud, T. 469<br />

Stevens<strong>on</strong>, A. M. 59<br />

Stewart, M. 275<br />

Stiuso, P. 285<br />

Stockert, J. C. 255<br />

Stocki, P. 332<br />

Stoevring, B. 387<br />

Stoloschyck, N. V. 242<br />

Størvold, G. L. 66, 323<br />

Stöhr, N. 154<br />

Straadt, I. K. 83<br />

Strano, S. 75<br />

Strauss, M. 382<br />

Strelnikova, Yu. I. 33<br />

Strey, H. H. 176<br />

Su, T.-P. 80<br />

Subramanian, S. 196<br />

Suckling, J. 366<br />

Sugawara, A. 35, 268<br />

Sugino, C. 20<br />

Suh, J.-H. 126<br />

Suh, S.-C. 203<br />

Sumegi, B. 19<br />

Sumie, J. 405<br />

Summers, H. C. 392<br />

Sun, X. 92, 310<br />

Sun, Y. 167<br />

Suo, Z.-r. 143<br />

Susanszky, A. 413, 474<br />

Susanszky, E. 475<br />

Sutherland, M. A. 290<br />

Suur<strong>on</strong>en, R. 243<br />

Suzuki, M. 367, 467<br />

Svetina, A. 291<br />

Sytykiewicz, H. 199<br />

Szabadkai, G. 325<br />

Szabados, L. 217<br />

Szabo, G. 328, 405, 470, 475<br />

Szabó, N. 470, 471<br />

Szabo, S. 373, 374<br />

Szájli, E. 199<br />

Szak<strong>on</strong>yi, D. 203<br />

Szalay, S. M. 308<br />

Szanto, Zs. 405<br />

Szegi, P. 228, 229<br />

Szegő, D. 218, 223<br />

Szekely, A. 413<br />

Székely, Gy. 217<br />

Szelényi, J. 427<br />

Széll, M. 147<br />

Szepesi, Á. 213, 221<br />

Szigeti, A. 19<br />

Szigeti, Z. 218, 223, 229<br />

Sztrum, A. A. 255<br />

Szűcs, E. 212<br />

T<br />

Tabata, A. 122<br />

Taché, Y. 371<br />

Tachikawa, H. 402<br />

Tadd<strong>on</strong>io, G. 221<br />

504


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Tahara, Y. 244, 479<br />

Tähepõld, A. 435<br />

Tajabadipour, A. 217, 219<br />

Tajabadipur, A. 479<br />

Takahashi, K. 57<br />

Takahashi, R. 470<br />

Takahashi, T. 300, 367<br />

Takaki, E. 46<br />

Takao, T. 402<br />

Takayama, K. 446<br />

Takayuki, N. 59<br />

Takeda, T. 446<br />

Takeuchi, K. 375<br />

Talebi, M. 219<br />

Täll, M. 454<br />

Tamás, L. 227<br />

Tamas, M. 296<br />

Tamura, Y. 266, 268<br />

Tan, K. T. 344<br />

Tanaka, K. 402<br />

Tanaka, K.-I. 45<br />

Tanaka, Y. 297, 401<br />

Tanasa, B. 354<br />

Tanfani, F. 61<br />

Tang, Y.-A. 163<br />

Tanguay, M. R. 18<br />

Tari, I. 151, 213, 214, 221<br />

Tarnawski, A. S. 372<br />

Tashiro, E. 70<br />

Tass<strong>on</strong>e, F. 241<br />

Tavares, E. 482<br />

Taverna, S. 62<br />

Tavío, M. M. 121<br />

Taylor, M. D. 358<br />

Taylor, T. C. 107<br />

Tayubi, I. A. 294<br />

Temiz, Y. 132<br />

Tengfei, P. 150<br />

Tentolouris, C. 261<br />

Teramura, K. 244, 479<br />

Terao, T. 81<br />

Theriault, J. 49<br />

Theuns, J. 310<br />

Thijssen, K. 13<br />

Thomm, M. 43<br />

Thorlin, T. 281<br />

Thurm<strong>on</strong>d, R. L. 161<br />

Tian, M. 197<br />

Tiefenbacher, S. 401<br />

Tilbrook, A. 416<br />

Tiligada, E. 161<br />

Timmerman, V. 307, 310<br />

Timmermans, J.-P. 307<br />

Titov, S. E. 214<br />

Tiwari, P. K. 57<br />

Toda, K. 298, 398<br />

T<str<strong>on</strong>g>of</str<strong>on</strong>g>t, D. 317<br />

Tolstanova, G. 373, 374<br />

Tomala, K. 180<br />

Tomé, F. D. 197<br />

Tomoyasu, T. 122<br />

T<strong>on</strong>g, Y. 267<br />

Torigoe, T. 64, 266, 268, 359<br />

Toro, Y. 132<br />

Toth, M. 391<br />

Tóth, Z. Sz. 207<br />

Török, K. 189, 223<br />

Tradewell, L. M. 358<br />

Trakranrungsie, N. 129, 136<br />

Traverso, N. 238<br />

Triantaphylidès, C. 187<br />

Trif<strong>on</strong>ova, E. A. 214<br />

Tr<str<strong>on</strong>g>of</str<strong>on</strong>g>imiuk, E. 243, 394<br />

Tr<strong>on</strong>che, F. 272<br />

Trouillet, D. 40<br />

Trümbach, D. 157<br />

Tsakiris, S. 280<br />

Tsakiris, T. 280<br />

Tsao, A. 328<br />

Tschanz, J. T. 403<br />

Tsuda, A. 401, 431, 433, 445, 442<br />

Tsuda, S. 401<br />

Tsukinoki, K. 300<br />

Tuba, Z. 210<br />

Tuboly, G. 370<br />

Tuchscherer, A. 289<br />

Tuchscherer, M. 289, 377<br />

Tucić, B. 193, 215<br />

Tukaj, S. 334<br />

Turk, N. 291<br />

Turk, R. 291<br />

Turóczy, Z. 189, 194<br />

Turturici, G. 62<br />

Tuteja, N. 204<br />

Tzacheva, I. 219<br />

Tyborowska, J. 334<br />

Tytell, M. 248<br />

505


23-26 August 2007,<br />

Budapest, Hungary<br />

U<br />

Uchimiya, H. 189<br />

Ueda, H. 363<br />

Ullers, S. R. 28<br />

Unfricht, D. W. 251<br />

Unno, K. 134<br />

Upadhyaya, C. P. 222<br />

Urban, O. 209<br />

Urbányi, B. 292<br />

Ursin, H. 409<br />

Usenko, V. 387<br />

Uslu, E. 130, 132, 134<br />

Ürményi, T. P. 154<br />

V<br />

Vaccaro, M. 61<br />

Vahtera, V. 454<br />

Valcu, C.-M. 182<br />

Valentin-Hansen, P. 94<br />

Valli, M. 116<br />

van Aken, A. 364<br />

Van Beek, K. 190, 198<br />

Van Broeckhoven, C. 310<br />

Van Calster, B. 379<br />

Van Damme, J. M. E. 205<br />

Van den Bergh, R. H. B. 379<br />

Van Den Bosch, L. 307, 310, 360<br />

van der Molen, G. M. 396<br />

Van Der Straeten, D. 211<br />

van der Zee, R. 335<br />

van Eden, W. 330, 335<br />

van Ham, T. 13<br />

Van Huffel, S. 379<br />

Van Lith, M. 101<br />

van Os, J. 366<br />

Van Waes,V. 383<br />

Vang, O. 387<br />

Vanloo, B. 307<br />

Vannuvel, K. 360<br />

Vantipalli, M. 240<br />

Varbanova, M. 197<br />

Varela, M. 132<br />

Varennes, A. 194<br />

Váró, G. 147<br />

Vasquez-Vivar, J. 295<br />

Vass, I. 188<br />

Vavuranakis, E. 261<br />

Vega, V. 51<br />

Vera, M. 95<br />

Vermes, I. 347<br />

Vermeylen, D. 380<br />

Vertii, A. 25<br />

Viau, F. P. 287<br />

Viau, P. 458<br />

Vicart, P. 92, 310<br />

Viergutz, T. 289<br />

Vierke, G. 43<br />

Vierling, E. 24, 26, 11<br />

Vieusseux, J. 320<br />

Vigh, L. 91<br />

Vihalemm, T. 284<br />

Vihervaara, A. 268<br />

Vila, J. 121<br />

Villaverde, A. 116<br />

Villeneuve, S. T. 167<br />

Vinet, J. 22<br />

Vingård, E. 462<br />

Vinokurov, M. 350<br />

Visnovitz, T. 218, 223<br />

Vittorelli, M. L. 62<br />

Vizi, E. S. 427<br />

Vlaskin, P. A. 33<br />

Voisine, C. 354<br />

Voll, R. E. 270<br />

Voos, W. 115, 118<br />

Vos, M. 355<br />

Vourc’h, C. 94, 97<br />

Vranic, M. 410<br />

Vrublevskaya, V. V. 323<br />

Vuleta, A. 193, 215<br />

Vydyanath, A. 36<br />

W<br />

Wach, S. 270<br />

Wagenaar-Hilbers, J. 335<br />

Wakida, S.-i. 297<br />

Waldeck, K. 320<br />

Waldhauser, C. 290<br />

Walesiuk, A. 394<br />

Walker, A. 411<br />

Wallis, T. 108<br />

Wallner, B. 459<br />

Walter, L. 316<br />

Wan, C.-H. 349<br />

Wang, C.-Y. 322<br />

Wang, Cha. 415<br />

Wang, Che. 279<br />

Wang, D. 269<br />

506


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Wang, M. 98<br />

Wang, S. J. 109<br />

Warne, P. J. 8<br />

Watanabe, K. 367<br />

Wayenberg, J.- L. 380<br />

Weber, P. 274<br />

Wechsler, B. 147<br />

Wejksza, K. 318<br />

Welsh, J. M. 92, 310<br />

Welsh-Bohmer, K. A. 403<br />

Wenke, A.-K. 43<br />

Werfel, T. 269<br />

Werner, T. 103<br />

Westwood, J. T. 155<br />

Wheler, G. H. T. 454<br />

Whitelaw, M. L. 108<br />

Wichians<strong>on</strong>, J. 282<br />

Wick, C. M. 348<br />

Wick, G. 331, 348<br />

Wieten, L. 335<br />

Wikman, G. 69, 242<br />

Wildemann, D. 108<br />

Wilhelm, I. 147<br />

Wilhelm, S. 290<br />

Wilk, K. 449<br />

Willemsen, R. 241<br />

Willershausen, B. 443<br />

Williams, B. D. 117<br />

Williams, B. R. 436<br />

Williams, G. 101<br />

Williams, H. 53<br />

Williams, H. H. J. 37, 50, 53, 78, 129,<br />

140, 316, 376<br />

Williams, R. 404<br />

Williams, S. J. 106<br />

Williams, V. 404<br />

Wils<strong>on</strong>, R. M. 50, 53, 54<br />

Wils<strong>on</strong>, W. C. 449<br />

Winkler, T. 270<br />

Wittmann, M. 269<br />

Wlazło, A. 422<br />

Wolfs<strong>on</strong>, M. 318<br />

Workman, P. 327<br />

Wotjak, C. T. 393<br />

Wu, C. C. 351<br />

Wu, C. C. 349<br />

Wu, C.-W. 399<br />

Wu, G. C. 249, 250<br />

Wurst, W. 274<br />

Wyatt, A. R. 50, 53<br />

Wyld, L. 321<br />

X<br />

Xia, Y. 249, 250<br />

Xiao, J. 146<br />

Xiao, P. H. 112<br />

Xi<strong>on</strong>g, Z. 146<br />

Xu, L. 246<br />

Xu, Y. 328<br />

Y<br />

Yaari, A. 334<br />

Yabu, T. 30 , 149<br />

Yaglom, J. 245, 317<br />

Yajima, J. 401, 431, 433, 445<br />

Yakovleva, T. V. 259<br />

Yamada, T. 81<br />

Yamaguchi, M. 244, 479<br />

Yamaguchi, T. 134<br />

Yamaguchi-Shinozaki, K. 201<br />

Yamamoto, K. 406<br />

Yamashita, M. 30, 149, 287<br />

Yang, J.-L. 163, 165, 322<br />

Yang, R. 249<br />

Yang, Z. 385<br />

Yassa, M. 130, 134<br />

Yates III, R. J. 354<br />

Yatmark, P. 129, 136<br />

Ye, R. 98<br />

Yen, A.-H. 165<br />

Yenari, M. 246<br />

Yerbury, J. J. 50, 54<br />

Yin, X. 251<br />

Yip, B. W. P. 111<br />

Ylilehto, H. 440<br />

Yohda, M. 20, 57, 60, 363<br />

Y<strong>on</strong>gyu, L. 150<br />

Yoo, S. B. 283<br />

Yo<strong>on</strong>, I.-S. 203<br />

Yo<strong>on</strong>, J.-H. 23, 45<br />

Yo<strong>on</strong>, T. 392<br />

Yoshida, Y. 136, 297<br />

Yoshinaga, K. 189<br />

Yoshinari, M. 406<br />

Yoshiyuki, T. 405<br />

Young, I. R. 237<br />

Young, J. F. 83<br />

Yu, L. 407<br />

507


23-26 August 2007,<br />

Budapest, Hungary<br />

Yu, O. 196<br />

Yu, R. M. K. 111<br />

Yuan, B. 328<br />

Yue, J. T. Y. 410<br />

Yui, K. 367<br />

Yuko, E. 405<br />

Yun, Y.-S. 266<br />

Z<br />

Zachara, E. N. 178<br />

Zagorchev, L. 219<br />

Zako, T. 60, 363<br />

Zámocký, M. 140<br />

Zana, A. 475<br />

Zandi, P. P. 403<br />

Zatinajchenko, N. 192<br />

Zavala, L. E. 480<br />

Zdzisinska B. 318<br />

Zeis, B. 110<br />

Zelena, D. 368<br />

Zeman, M. 341<br />

Zemlyanoi, A. 265<br />

Zemlyanoi, A. 350<br />

Zeredo, J. 298<br />

Zermati, Y. 75<br />

Zgaga-Griesz, A. 55<br />

Zhang, N. X. 267<br />

Zhang, P. 104<br />

Zheng, J.-b. 143<br />

Zhoa, X. 177<br />

Zhou, F. 249, 250<br />

Zhou, S.-Y. 367<br />

Zhu, J.-K. 201<br />

Zhu, Z.-l. 143, 145<br />

Zhukov, A. 60<br />

Zilmer, M. 284<br />

Zitkevičius, V. 230<br />

Zołnierczyk-Zreda, D. 425<br />

Zombori, Z. 223<br />

Zumbrägel, S. 110<br />

Zummo, G. 85, 158<br />

Zurmühlen, R. 393<br />

Zwart, R. 364<br />

Zsigm<strong>on</strong>d, L. 217<br />

508


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

BOOK OF LATE ABSTRACTS<br />

2 ND WORLD CONFERENCE OF STRESS<br />

3 RD CELL STRESS SOCIETY INTERNATIONAL CONGRESS ON<br />

STRESS RESPONSES IN BIOLOGY AND MEDICINE AND<br />

2 ND WORLD CONFERENCE OF STRESS<br />

CELEBRATION OF THE CENTENNIAL BIRTH ANNIVERSARY<br />

OF HANS SELYE<br />

23-26 AUGUST 2007, BUDAPEST, HUNGARY<br />

1


2<br />

23-26 August 2007,<br />

Budapest, Hungary


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

1A. SMALL HSPS<br />

(ANDRE-PATRICK ARRIGO, ROBERT M. TANGUAY)<br />

1A_10_P<br />

(poster secti<strong>on</strong> A1, poster board #65, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ROLE OF IHF AND NTRC PROTEINS IN THE TRANSCRIPTION IN VIVO OF<br />

ESCHERICHIA COLI IBPB GENE<br />

Beata Nadratowska-Wesolowska, Anna Janaszak, Grazyna K<strong>on</strong>opa, Alina Taylor<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular Biology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Gdansk, Poland<br />

e-mail: nadra@biotech.ug.gda.pl<br />

Expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock protein genes in E. coli depends <strong>on</strong> two regul<strong>on</strong>s c<strong>on</strong>trolled by σ 32 and σ 24 , RNAP<br />

subunits. A third regul<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a stress resp<strong>on</strong>se, which is under c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> the RNAP-σ 54 holoenzyme has been<br />

identified. It includes pspA-E oper<strong>on</strong>, rpoH gene and the gene encoding a small heat shock protein IbpB.<br />

RNAP-σ 54 holoenzyme can <str<strong>on</strong>g>for</str<strong>on</strong>g>m a closed promoter complex but is incapable <str<strong>on</strong>g>of</str<strong>on</strong>g> <str<strong>on</strong>g>for</str<strong>on</strong>g>ming an open complex in<br />

the absence <str<strong>on</strong>g>of</str<strong>on</strong>g> an activator protein and ATP. IHF mediated DNA bend is frequently essential <str<strong>on</strong>g>for</str<strong>on</strong>g> efficient<br />

inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> transcripti<strong>on</strong>.<br />

We analyzed the upstream sequence <str<strong>on</strong>g>of</str<strong>on</strong>g> the ibpB gene and found IHF and NtrC binding sites. Binding <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

NtrC, IHF and RNAP-σ 54 holoenzyme to the ibpB DNA fragments was detected by a gel mobility shift assay.<br />

DNase I footprinting method was employed <str<strong>on</strong>g>for</str<strong>on</strong>g> the identificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the NtrC and IHF binding sequences in<br />

the promoter regi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the ibpB gene. Preliminary experiments have shown that NtrC and IHF provide DNA<br />

protecti<strong>on</strong> against DNase I digesti<strong>on</strong>.<br />

Primer extensi<strong>on</strong> assays revealed three transcripti<strong>on</strong> start sites at 81, 88, 111 nt upstream <str<strong>on</strong>g>of</str<strong>on</strong>g> translati<strong>on</strong>al start<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the ibpB gene. After heat shock in the ∆ IHF mutant strain, the transcripts starting at 81 and 88 nt were<br />

absent. There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, while the transcripts starting at 88 and 81 positi<strong>on</strong>s are IHF-dependent, the transcript at<br />

the 111 positi<strong>on</strong> is IHF-independent.<br />

NtrC activity is usually inducible by nitrogen starvati<strong>on</strong>. However, when such c<strong>on</strong>diti<strong>on</strong>s were applied, the<br />

primer extensi<strong>on</strong> test did not reveal the promoter transcripti<strong>on</strong>al activity. The ATP dependent activators <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the AAA+ protein type act from l<strong>on</strong>g distances, what might be interpreted that the enhancer bound NtrC<br />

acts <strong>on</strong> another gene downstream <str<strong>on</strong>g>of</str<strong>on</strong>g> ibpB sequence.<br />

1A_11_P<br />

(poster secti<strong>on</strong> A1, poster board #66, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CHARACTERIZATION OF SPECIFIC PEPTIDE APTAMERS TARGETING HSP27<br />

ANTI-APOPTOTIC ACTIVITY<br />

B. Gibert, A. Czekalla, A. P. Arrigo, C. Diaz-Latoud<br />

<str<strong>on</strong>g>Centre</str<strong>on</strong>g> de Génétique Moléculaire et Cellulaire. Université Claude Bernard, Ly<strong>on</strong> 1, Villeurbanne, France<br />

e-mail: chantal.diaz@univ-ly<strong>on</strong>1.fr<br />

A high level <str<strong>on</strong>g>of</str<strong>on</strong>g> expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the small heat shock protein Hsp27 enhances the resistance <str<strong>on</strong>g>of</str<strong>on</strong>g> many cancer cell<br />

to death and chemotherapeutic treatments while reducing this level sensitize them. Anti-Hsp27 therapies<br />

could be a potential beneficial additi<strong>on</strong> to existing anti-cancer therapies. In order to identify random specific<br />

Hsp27 peptides inhibitors we used the peptide aptamer technology. Peptide aptamers are made <str<strong>on</strong>g>of</str<strong>on</strong>g> short<br />

peptide sequences presented into a scaffold protein, here the bacterial thioredoxin A (TrxA), which<br />

3


23-26 August 2007,<br />

Budapest, Hungary<br />

c<strong>on</strong>strains their c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>. Here, we present the characterizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> peptides aptamers raised against<br />

Hsp27.<br />

Peptide aptamers were selected from an initial pool <str<strong>on</strong>g>of</str<strong>on</strong>g> around 1 billi<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> their ability to bind Hsp27. 160<br />

positive candidates were isolated from a screen using a yeast two hybrid technology and interacti<strong>on</strong> with<br />

Hsp27 was c<strong>on</strong>firmed <str<strong>on</strong>g>for</str<strong>on</strong>g> 59 aptamers. Retrans<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> assay using Hsp27 or αB-crystallin as a bait<br />

c<strong>on</strong>firmed the specificity <str<strong>on</strong>g>of</str<strong>on</strong>g> interacti<strong>on</strong> with Hsp27 <str<strong>on</strong>g>for</str<strong>on</strong>g> 15 aptamers.<br />

Positive aptamers were tested in HeLa cell to identify those with the best activity and functi<strong>on</strong>al pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ile.<br />

Several aptamers bearing different peptide sequences were found to inhibit Hsp27 anti-apoptotic activity in<br />

staurosporine treated HeLa cells without reducing its level. The efficiency <str<strong>on</strong>g>of</str<strong>on</strong>g> these aptamers was as good as<br />

that <str<strong>on</strong>g>of</str<strong>on</strong>g> shRNA. Furthermore these aptamers also interfere with Hsp27 biochemical properties. Hence, these<br />

peptide aptamers present a new alternative to inhibit anti-apoptotic activity <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp27 without reducing its<br />

level.<br />

Taken together our results suggest that in the future high resoluti<strong>on</strong> modeling <str<strong>on</strong>g>of</str<strong>on</strong>g> new druggable sites based<br />

<strong>on</strong> peptide aptamer binding <strong>on</strong> Hsp27 data will be defined.<br />

1A_12_P<br />

(poster secti<strong>on</strong> A1, poster board #67, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

MUTATIONS IN SYNECHOCYSTIS HSP17 SPECIFICALLY MODIFY THE<br />

MEMBRANE ACTIVITY OF THIS SMALL HEAT SHOCK PROTEIN<br />

1 Z. Balogi, 2 O. Cheregi, 3 K. Giese, 1 M. Debreczeny, 1 K. Juhasz, 2 I. Vass, 3 E. Vierling, 1 L. Vígh,<br />

1 I. Horváth<br />

1Inst. Biochem.; 2 Inst. Plant Biol., Biol. Res. Center, POB 521 Szeged, H-6701 Hungary;<br />

3Dept. Biochem. & Mol. Biophys., Univ. Ariz<strong>on</strong>a, 1007 E. Lowel St., Tucs<strong>on</strong>, AZ 85721, USA<br />

The cellular pool <str<strong>on</strong>g>of</str<strong>on</strong>g> small heat shock proteins (sHsps) is divided into a cytoplasmic subfracti<strong>on</strong>, resp<strong>on</strong>sible<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> regular chaper<strong>on</strong>e activity and a membraneous subfracti<strong>on</strong>, involved in membrane stabilizati<strong>on</strong>. We have<br />

isolated a series <str<strong>on</strong>g>of</str<strong>on</strong>g> Synechocystis Hsp17 point mutants characterized with regards to in vitro chaper<strong>on</strong>e activity<br />

and propensity to <str<strong>on</strong>g>for</str<strong>on</strong>g>m oligomers. Intriguingly, two <str<strong>on</strong>g>of</str<strong>on</strong>g> these mutati<strong>on</strong>s (L9P and Q16R), which did not affect<br />

either the in vitro chaper<strong>on</strong>e activity or the propensity <str<strong>on</strong>g>of</str<strong>on</strong>g> this sHsp to <str<strong>on</strong>g>for</str<strong>on</strong>g>m oligomers, resulted in dramatic<br />

changes in membrane acivity <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp17. We defined particular features <str<strong>on</strong>g>of</str<strong>on</strong>g> these mutants resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

interacting with membrane lipids. L9P and Q16R proteins proved to behave differently with respect to their<br />

ability to associate with lipids and to protect photosynthetic membrane functi<strong>on</strong> against UV-B stress. L9P<br />

had a lesser, Q16R a higher membrane activity than the WT protein both in vivo and in vitro. Our knowledge<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the functi<strong>on</strong>al importance and <str<strong>on</strong>g>of</str<strong>on</strong>g> the mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp17-membrane (lipid) interacti<strong>on</strong> suggests that<br />

modificati<strong>on</strong>s either in protein structure or in lipid compositi<strong>on</strong> can be engineered to achieve higher stress<br />

tolerance <str<strong>on</strong>g>of</str<strong>on</strong>g> photosynthetic organisms.<br />

4


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

1C. HSF ACTIVATION<br />

(LEA SISTONEN)<br />

1C_10_P<br />

(poster secti<strong>on</strong> A1, poster board #68, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DIFFERENTIAL DISPLAY OF DNA-BINDING PROTEINS (DDDP) REVEALS HEAT<br />

SHOCK FACTOR 1 (HSF1) AS NOVEL CIRCADIAN TRANSCRIPTION FACTOR<br />

Hans Reinke, Fabienne Fleury-Olela, Charna Dibner, Ueli Schibler<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Geneva, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Molecular Biology, Quai Ernest-Ansermet 30, 1211 Geneva, Switzerland<br />

The circadian clock enables the anticipati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> daily recurring envir<strong>on</strong>mental changes by pre-setting an<br />

organism's physiology and behavior. Driven and synchr<strong>on</strong>ized by a central pacemaker in the brain, circadian<br />

output genes fine-tune a wide variety <str<strong>on</strong>g>of</str<strong>on</strong>g> physiological parameters in peripheral organs. However, <strong>on</strong>ly a<br />

subset <str<strong>on</strong>g>of</str<strong>on</strong>g> circadianly transcribed genes seems to be directly regulated by core clock proteins. Assuming that<br />

yet unidentified transcripti<strong>on</strong> factors must exist in the circadian transcripti<strong>on</strong>al network we set out to develop<br />

a novel technique, differential display <str<strong>on</strong>g>of</str<strong>on</strong>g> DNA-binding proteins (DDDP), that we used to screen mouse liver<br />

nuclear extracts. In additi<strong>on</strong> to a number <str<strong>on</strong>g>of</str<strong>on</strong>g> well established clock proteins we found DNA-binding <str<strong>on</strong>g>of</str<strong>on</strong>g> heat<br />

shock factor 1 (HSF1) to be highly circadian. HSF1 drives the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock proteins at the <strong>on</strong>set<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the dark phase when the animals start to be behaviorally active and to ingest food. Our results suggest that<br />

every day the mammalian body goes through a proteotoxic stress event that elicits the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cellprotective<br />

proteins. Furthermore, our findings dem<strong>on</strong>strate the power <str<strong>on</strong>g>of</str<strong>on</strong>g> the new screening method DDDP<br />

which is not restricted to the circadian network but can be applied to discover novel differentially regulated<br />

transcripti<strong>on</strong> factors in various biological systems.<br />

1C_11_P<br />

(poster secti<strong>on</strong> A1, poster board #69, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HEAT SHOCK FACTOR 1 REPRESSES ESTROGEN DEPENDENT TRANSCRIPTION<br />

THROUGH ASSOCIATION WITH MTA1<br />

Abdul Khaleque, Ajit Bharti, Phillip J. Gray, Daniel R. Ciocca, Arturo Stati, Stuart Calderwood<br />

Harvard medical School/BIDMC, Oncology, Bost<strong>on</strong>, United States<br />

Heat shock factor 1 (HSF1), the transcripti<strong>on</strong>al activator <str<strong>on</strong>g>of</str<strong>on</strong>g> the heat shock genes, is increasingly implicated in<br />

cancer. We have shown that HSF1 binds to the co-repressor metastasis associated protein 1 (MTA1) in vitro and in<br />

human breast carcinoma samples. HSF1–MTA1 complex <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> was str<strong>on</strong>gly induced by the<br />

trans<str<strong>on</strong>g>for</str<strong>on</strong>g>ming ligand heregulin and complexes incorporated a number <str<strong>on</strong>g>of</str<strong>on</strong>g> additi<strong>on</strong>al proteins including hist<strong>on</strong>e<br />

deacetylases (HDAC1 and 2) and Mi2α, all comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g> the NuRD co-repressor complex. These<br />

complexes were induced to assemble the chromatin <str<strong>on</strong>g>of</str<strong>on</strong>g> MCF7 breast carcinoma cells and associated with the<br />

promoters <str<strong>on</strong>g>of</str<strong>on</strong>g> estrogen resp<strong>on</strong>sive genes. Such HSF1 complexes participate in repressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> estrogendependent<br />

transcripti<strong>on</strong> in breast carcinoma cells treated with heregulin and this effect was inhibited by<br />

MTA1 knockdown. Repressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> estrogen dependent transcripti<strong>on</strong> may c<strong>on</strong>tribute to the role <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF1 in<br />

cancer.<br />

5


23-26 August 2007,<br />

Budapest, Hungary<br />

1F. PHARMACOLOGICAL MODULATORS OF STRESS PROTEIN EXPRESSION<br />

1F_10_P<br />

(poster secti<strong>on</strong> A1, poster board #70, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

NOVEL CHEMICAL ENHANCERS OF HEAT SHOCK INCREASE THERMAL<br />

RADIOSENSITIZATION<br />

K<strong>on</strong>jeti R. Sekhar 1 , Vijayakumar N. S<strong>on</strong>ar 2 , Venkatraj Muthusamy 2 , Soumya Sasi 1 , Andrei Laszlo 3 ,<br />

Jamil Sawani 4 , Nobuo Horikoshi 3 , Ryuji Higashikubo 3 , Robert G. Bristow 4 , Michael J. Borrelli 5 ,<br />

Peter A. Crooks 2 , James R. Lepock 4 , Joseph L. Roti Roti 3 , Michael L. Freeman 1<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Radiati<strong>on</strong> Oncology/Vanderbilt-Ingram Cancer Center,<br />

Vanderbilt University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, Nashville, Tennessee;<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmaceutical Sciences, College <str<strong>on</strong>g>of</str<strong>on</strong>g> Pharmacy, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Kentucky, Lexingt<strong>on</strong>, Kentucky;<br />

3Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Radiati<strong>on</strong> Oncology, Washingt<strong>on</strong> University School <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, St. Louis, Missouri;<br />

4Princess Margaret Hospital (University Health Network) and Departments <str<strong>on</strong>g>of</str<strong>on</strong>g> Radiati<strong>on</strong> Oncology and Medical<br />

Biophysics, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Tor<strong>on</strong>to, Tor<strong>on</strong>to, Ontario, Canada;<br />

5Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Radiology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Arkansas <str<strong>on</strong>g>for</str<strong>on</strong>g> Medical Sciences, Little Rock, Arkansas<br />

Radiati<strong>on</strong> therapy combined with adjuvant hyperthermia has the potential to provide outstanding localregi<strong>on</strong>al<br />

c<strong>on</strong>trol <str<strong>on</strong>g>for</str<strong>on</strong>g> refractory disease. However, achieving therapeutic thermal dose is problematic. We used<br />

a chemistry driven approach to design and synthesize small molecules that could functi<strong>on</strong> as thermal<br />

radiosensitizers. (Z)-(±)-2-(1-Benzenesulf<strong>on</strong>ylindol-3-ylmethylene)-1-azabicyclo[2.2.2]octan-3-ol was<br />

identified as a compound that could lower the threshold <str<strong>on</strong>g>for</str<strong>on</strong>g> the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSF-1. Enhanced thermal<br />

sensitivity resulted in increased heat radiosensitizati<strong>on</strong>. The structural requirements <str<strong>on</strong>g>for</str<strong>on</strong>g> activity <str<strong>on</strong>g>of</str<strong>on</strong>g> this novel<br />

molecule are: the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> an Nbenzenesulf<strong>on</strong>ylindole or N-benzylindole moiety linked at the indolic 3-<br />

positi<strong>on</strong> to a 2-(1-azabicyclo [2.2.2]octan-3-ol) or 2-(1-azabicyclo[2.2.2]octan-3-<strong>on</strong>e) moiety. These small<br />

molecules functi<strong>on</strong>ed by exploiting the underlying biophysical events resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g> thermal sensitizati<strong>on</strong>.<br />

Thermal radiosensitizati<strong>on</strong> was characterized biochemically and found to include loss <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>drial<br />

membrane potential, followed by mitotic catastrophe. Furthermore, heat associated perturbati<strong>on</strong>s in<br />

radiati<strong>on</strong> induced signaling pathways were enhanced by these compounds. Overall, we have identified a novel<br />

class <str<strong>on</strong>g>of</str<strong>on</strong>g> small molecules that are a promising tool <str<strong>on</strong>g>for</str<strong>on</strong>g> the treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> recurrent tumors with i<strong>on</strong>izing<br />

radiati<strong>on</strong><br />

6


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2A. CELL STRESS, HSPS AND APOPTOSIS<br />

(SEAMUS MARTIN)<br />

2A_18_P<br />

(poster secti<strong>on</strong> A2, poster board #169, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HSP60D, A MEMBER OF THE HSP60 FAMILY, IS A NOVEL MODIFIER OF CELL<br />

DEATH IN DROSOPHILA<br />

Richa Arya, Subhash C. Lakhotia<br />

Cytogenetics Laboratory, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Zoology, Banaras Hindu University, Varanasi 221 005, India<br />

Hsps, besides functi<strong>on</strong>ing as molecular chaper<strong>on</strong>es, are now known to be also involved in several other vital<br />

cellular activities, <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> which is apoptosis. Hsp60 family proteins have been reported to display anti- as well<br />

as pro-apoptotic activities. Drosophila melanogaster genome has four Hsp60 genes, the Hsp60D gene (CG16954)<br />

being located at 34C cytogenetic regi<strong>on</strong>. We generated transgenic flies in which the Hsp60D could either be<br />

c<strong>on</strong>diti<strong>on</strong>ally over-expressed or its transcripts c<strong>on</strong>diti<strong>on</strong>ally ablated through RNAi. We found a str<strong>on</strong>g<br />

interacti<strong>on</strong> between Hsp60D and the apoptotic proteins like Reaper, HID, GRIM (RHG proteins), caspases<br />

and DIAP1. Cell death caused by over-expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> RHG and DRONC proteins is remarkably suppressed<br />

by ablati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp60D by RNAi. Though, Hsp60D-RNAi rescues cell death mediated by full-length as well<br />

as activated DRONC, it is unable to suppress the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> unguarded caspases in Diap1-RNAi flies. Coexpressi<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> Diap1-RNAi with Hsp60D-RNAi also prevents rescue <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp60D-ablati<strong>on</strong> mediated apoptosis<br />

in RHG expressing flies. Over-expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp60D enhances apoptotic cell death caused by the above<br />

apoptotic proteins. Our results thus suggest that Hsp60D acts downstream <str<strong>on</strong>g>of</str<strong>on</strong>g> RPR, HID and GRIM but<br />

upstream <str<strong>on</strong>g>of</str<strong>on</strong>g> caspases, indicating a possible interacti<strong>on</strong> between Hsp60D and DIAP1.<br />

2A_19_P<br />

(poster secti<strong>on</strong> A2, poster board #170, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CHAPERONE ACTIVITY REGULATES THE PROTEIN STEADY-STATE LEVELS,<br />

PHOSPHORYLATION, AND ULTIMATE ACTIVITY OF THE ANTI-APOPTOTIC<br />

KINASE, SGK-1<br />

Larissa Belova, Betty Ky, Deanna Brickley, Sanjay Sharma, Suzanne D. C<strong>on</strong>zen*<br />

The Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine and the Ben May Department <str<strong>on</strong>g>for</str<strong>on</strong>g> Cancer <str<strong>on</strong>g>Research</str<strong>on</strong>g><br />

The University <str<strong>on</strong>g>of</str<strong>on</strong>g> Chicago<br />

Chicago, IL 60637 USA<br />

Serum and glucocorticoid-regulated kinase-1 (SGK-1), a member <str<strong>on</strong>g>of</str<strong>on</strong>g> the protein kinase AGC family, plays an<br />

important role in maintaining cellular osmotic homeostasis and c<strong>on</strong>tributes to breast epithelial cell<br />

proliferati<strong>on</strong> and survival. SGK-1 mRNA expressi<strong>on</strong> is transiently induced following cellular stress, and<br />

SGK-1 protein levels are tightly regulated by rapid proteasomal degradati<strong>on</strong>. We have previously reported<br />

that SGK-1 <str<strong>on</strong>g>for</str<strong>on</strong>g>ms a complex with the stress-associated E3 ligase CHIP [C-terminus <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsc (heat-shock<br />

cognate protein) 70-interacting protein] and Hsp70; CHIP is required <str<strong>on</strong>g>for</str<strong>on</strong>g> both the ubiquitin modificati<strong>on</strong> and<br />

rapid proteasomal degradati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> SGK-1. We also showed that CHIP co-localizes with SGK-1 at or near the<br />

endoplasmic reticulum (ER), suggesting that SGK-1 may play a role in the resp<strong>on</strong>se to ER stress. CHIPmediated<br />

regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> SGK-1 steady-state protein levels altered SGK-1 kinase activity. These data suggest a<br />

model that integrates CHIP and Hsp70 chaper<strong>on</strong>e functi<strong>on</strong> with regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the PI3K/SGK-1 pathway in<br />

7


23-26 August 2007,<br />

Budapest, Hungary<br />

the stress resp<strong>on</strong>se. We now report that PI3-K-dependent phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> SGK-1 requires <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the complex between SGK-1 and heat-shock protein 90 (Hsp90). We have observed that inactivati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Hsp90 by geldanamycin lead to decreased SGK-1 phosphorylati<strong>on</strong>, and that inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> PI3-K activity by<br />

LY294002 appears to eliminate SGK-1 phosphorylati<strong>on</strong> at the same residues as those affected by<br />

geldanamycin treatment. Interestingly, geldanamycin-targeted phosphorylati<strong>on</strong> sites are not limited to the<br />

known c<strong>on</strong>served PDK1/PDK2 sites Thr256 and Ser422 in SGK-1, but include additi<strong>on</strong>al residues<br />

phosphorylated in a PI3-K dependent manner. We find that inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Hsp90 also results in complete<br />

loss <str<strong>on</strong>g>of</str<strong>on</strong>g> SGK-1 kinase activity. Taken together, our data suggest that chaper<strong>on</strong>es functi<strong>on</strong> as essential<br />

comp<strong>on</strong>ents modulating the PI3-K/SGK-1 cell signaling resp<strong>on</strong>se via both phosphorylati<strong>on</strong> and ubiquitin<br />

modificati<strong>on</strong>/protein degradati<strong>on</strong>.<br />

2B. MEMBRANE-REGULATION OF THE STRESS RESPONSE<br />

(LÁSZLÓ VÍGH, JOHN L. HARWOOD)<br />

2B_03_P<br />

(poster secti<strong>on</strong> A2, poster board #171, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

HYPERFLUIDIZATION-COUPLED MEMBRANE MICRODOMAIN<br />

REORGANIZATION IS LINKED TO ACTIVATION OF THE HEAT SHOCK<br />

RESPONSE<br />

Enikő Nagy*, Zsolt Balogi*, Imre Gombos*, Malin Åkerfelt + , Gábor Balogh*, Zsolt Török*, Andriy<br />

Maslyanko*, Peter Slotte ++ , Lea Sist<strong>on</strong>en + , Ibolya Horváth*, László Vígh* $<br />

*Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, Biological <str<strong>on</strong>g>Research</str<strong>on</strong>g> Center, Hungarian Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences, Szeged, Hungary<br />

+Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Finland; Abo Akademi University, Turku Center <str<strong>on</strong>g>for</str<strong>on</strong>g> Biotechnology, Turku,<br />

Finland<br />

++Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry and Pharmacy, Abo Akademi University, Turku, Finland<br />

Targeting <str<strong>on</strong>g>of</str<strong>on</strong>g> the Hsp functi<strong>on</strong> in tumor cells is currently being assessed as potential anticancer therapy. An<br />

improved understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> the molecular signals that trigger or attenuate the stress protein resp<strong>on</strong>se is<br />

essential <str<strong>on</strong>g>for</str<strong>on</strong>g> advances to be made in this field.<br />

The present study provides evidences that the membrane fluidizer benzyl alcohol (BA), a documented<br />

n<strong>on</strong>protein denaturant, acts as a chaper<strong>on</strong>e inducer in B16(F10) melanoma cells. It is dem<strong>on</strong>strated that this<br />

effect relies basically <strong>on</strong> HSF1 activati<strong>on</strong>. Under the c<strong>on</strong>diti<strong>on</strong>s tested, the BA-induced Hsp resp<strong>on</strong>se<br />

involves the upregulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> a subset <str<strong>on</strong>g>of</str<strong>on</strong>g> hsp genes. It was previously known that a membrane fluidity increase<br />

may initiate an Hsp resp<strong>on</strong>se. It is now shown that the same level <str<strong>on</strong>g>of</str<strong>on</strong>g> membrane fluidizati<strong>on</strong> (estimated with a<br />

DPH probe in the core membrane regi<strong>on</strong>) attained with the closely analogous phenethyl alcohol (PhA)<br />

instead <str<strong>on</strong>g>of</str<strong>on</strong>g> BA, does not generate a stress protein signal. BA at a c<strong>on</strong>centrati<strong>on</strong> that activates heat shock genes<br />

exerts a pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ound effect <strong>on</strong> the melting <str<strong>on</strong>g>of</str<strong>on</strong>g> raft-like cholesterol-sphingomyelin domains in vitro, whereas PhA<br />

at a c<strong>on</strong>centrati<strong>on</strong> equipotent with BA in membrane fluidizati<strong>on</strong> has no such effect. Further, via the in vivo<br />

fPEG-Chol labeling <str<strong>on</strong>g>of</str<strong>on</strong>g> melanoma cells, we found that, similarly as <str<strong>on</strong>g>for</str<strong>on</strong>g> some other known membrane<br />

intercalators or heat stress per se, BA, but not PhA, initiates pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ound alterati<strong>on</strong>s in the plasma membrane<br />

microdomain structure.<br />

We suggest that, apart from membrane hyperfluidizati<strong>on</strong> in the deep hydrophobic regi<strong>on</strong>, a distinct<br />

reorganizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cholesterol-rich microdomains may also be required <str<strong>on</strong>g>for</str<strong>on</strong>g> the generati<strong>on</strong> and transmissi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

sufficient stress signals to activate hsp genes.<br />

8


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

2G. STRESS OF BACTERIA<br />

(MARÍA M. TAVÍO)<br />

2G_06_P<br />

(poster secti<strong>on</strong> B1, poster board #243, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE BACTERIAL STRESS MECHANISM OF STRINGENT RESPONSE PLAY THE<br />

MAJOR ROLE FOR THE FORMATION OF INORGANIC POLYPHOSPHATE IN<br />

E. COLI AND PSEUDOMONAS AERUGINOSA<br />

Chang-Do Hyun 1 , Chi-Y<strong>on</strong>g Eom 2 , Se<strong>on</strong>g-Kyu Park 3 , Jae-Ri Jung 4 , Je<strong>on</strong>g-Han Suh 1 ,<br />

H<strong>on</strong>g-Yeoul Kim 1,4*<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, College <str<strong>on</strong>g>of</str<strong>on</strong>g> Oriental Medicine, Kyung Hee University, Seoul, Korea (Rep.), 2 Metabolome<br />

Analysis Team, KBSI Seoul Center, 126-16 Anam-D<strong>on</strong>g, Korea University, Seoul, Korea (Rep.), 3 Department <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Prescripti<strong>on</strong>ology, College <str<strong>on</strong>g>of</str<strong>on</strong>g> Oriental Medicine, Kyung Hee University, Seoul, Korea (Rep.), 4 HelixPharms Co., Ltd.,<br />

Seoul, Korea (Rep.) * e-mail: hyk@khu.ac.kr<br />

The sec<strong>on</strong>d poly (p)ase has been identified as guanosine pentaphosphate phosphohydrolase(GPP). Since this<br />

enzyme is <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> key enzyme in the stringent resp<strong>on</strong>se, it has been suggested that a possible interacti<strong>on</strong> with a<br />

poly P <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> in E. coli. In this study, we measured the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> poly P in wild-type <str<strong>on</strong>g>of</str<strong>on</strong>g> E. coli and<br />

Pseudom<strong>on</strong>as aeruginosa when the cells cope with amino acids starvati<strong>on</strong>. In wild-type <str<strong>on</strong>g>of</str<strong>on</strong>g> E. coli, and Pseudom<strong>on</strong>as<br />

aeruginosa, poly P levels increases stable manner al<strong>on</strong>g with ppGpp c<strong>on</strong>centrati<strong>on</strong>. Additi<strong>on</strong>ally, neither relA(∆)<br />

nor relA spoT(∆) mutants <str<strong>on</strong>g>of</str<strong>on</strong>g> E. coli can not accumulate poly P in exp<strong>on</strong>entially growing E. coli. However,<br />

relA(∆) mutant can be accumulated the poly P in low Pi c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> MOPS media, culture c<strong>on</strong>diti<strong>on</strong> that<br />

producing the ppGpp by spoT. Futhermore, the plasmid harboring the relA gene under an inducible promoter<br />

allowed us to increase the ppGpp c<strong>on</strong>centrati<strong>on</strong> without any starvati<strong>on</strong> and thus, to observe directly the<br />

effect <str<strong>on</strong>g>of</str<strong>on</strong>g> ppGpp <str<strong>on</strong>g>for</str<strong>on</strong>g> the poly P <str<strong>on</strong>g>for</str<strong>on</strong>g>matin in E. coli. The significant amount <str<strong>on</strong>g>of</str<strong>on</strong>g> poly P can be maintained much<br />

l<strong>on</strong>ger period than that <str<strong>on</strong>g>of</str<strong>on</strong>g> wild-type in gppA(∆) mutant, c<strong>on</strong>firming that GPP act as a sec<strong>on</strong>d poly (P)ase in<br />

vivo. Based <strong>on</strong> these results, this study suggests the direct evidence that poly P <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> is dependent <strong>on</strong><br />

ppGpp c<strong>on</strong>centrati<strong>on</strong> in Gram-negative bacteria.<br />

2G_07_P<br />

(poster secti<strong>on</strong> B1, poster board #244, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ANTIBACTERIAL FUNCTION OF INORGANIC POLYPHOSPHATE ON STRESS<br />

INDUCED E.COLI<br />

Eun-Hye Kim 1 , Se<strong>on</strong>g-Kyu Park 2 , Jae-Ri Jung 3 , Je<strong>on</strong>g-Han Suh 1 , H<strong>on</strong>g-Yeoul Kim 1,3*<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, 2 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Prescripti<strong>on</strong>ology, College <str<strong>on</strong>g>of</str<strong>on</strong>g> Oriental Medicine, Kyung Hee University,<br />

Seoul, Korea (Rep.), 3 HelixPharms Co., Ltd., Seoul, Korea<br />

* e-mail: hyk@khu.ac.kr<br />

Inorganic polyphosphate polyP is a linear polymer <str<strong>on</strong>g>of</str<strong>on</strong>g> up to hundreds <str<strong>on</strong>g>of</str<strong>on</strong>g> orthophosphate (Pi) residues linked<br />

by high-energy phosphoanhydride b<strong>on</strong>ds. PolyP is ubiquitous in nature, having been found in all organism<br />

examined, yet little is known about its physiological roles. The present study was per<str<strong>on</strong>g>for</str<strong>on</strong>g>med to observe the<br />

antibacterial effect <str<strong>on</strong>g>of</str<strong>on</strong>g> inorganic polyphosphate <strong>on</strong> wild type E.coli MG1655 and ppk/x(∆) mutant CF5802.<br />

Additi<strong>on</strong>ally, the antibacterial effect(s) observed <strong>on</strong> several different stresses <str<strong>on</strong>g>of</str<strong>on</strong>g> starvati<strong>on</strong>. Antibacterial effect<br />

9


23-26 August 2007,<br />

Budapest, Hungary<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> polyP was shown significant effect in MOPS carb<strong>on</strong> starvati<strong>on</strong> media. Intracellular uncleotide release from<br />

the cell was increased by approximately. 20% in the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> polyP but was not reserved by the additi<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> divalent cati<strong>on</strong>s like Ca ++ and Mg ++ .After polyP treatment, it is possible to observe the transcripti<strong>on</strong>al<br />

activity <str<strong>on</strong>g>of</str<strong>on</strong>g> possible polyP related genes <strong>on</strong> cell growth. Interestingly, spoT, relA, l<strong>on</strong>, ftsZ, were shown<br />

significant different <str<strong>on</strong>g>of</str<strong>on</strong>g> transcripti<strong>on</strong>al activity. The overall results suggest that polyP have a str<strong>on</strong>g bactericidal<br />

activity against wild type E. coli MG1655, and these effect may related to depending genes rather than<br />

chelati<strong>on</strong> effects.<br />

2G_08_P<br />

(poster secti<strong>on</strong> B1, poster board #245, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

THE EXOPOLYPHOSPHATASE OPERON CONTROLLED BY ENVELOPE<br />

PROTEIN SYNTHESIS STRESS GENE MYCOBACTERIA TUBERCULOSIS H37Rv<br />

Gun-Wook Kim 1 , Ravi Kumar Vilwanathan 1 , Se<strong>on</strong>g-Kyu Park 2 , Chi-Y<strong>on</strong>g Eom 3 , Jae-Ri Jung 4 ,<br />

Je<strong>on</strong>g-Han Suh 1 , H<strong>on</strong>g-Yeoul Kim 1,4*<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, 2 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Prescripti<strong>on</strong>ology, College <str<strong>on</strong>g>of</str<strong>on</strong>g> Oriental Medicine, Kyung Hee University,<br />

Seoul, Korea (Rep.), 3 Metabolome Analysis Team, KBSI Seoul Center, 126-16 Anam-D<strong>on</strong>g, Korea University,<br />

Seoul, Korea (Rep.), 4 HelixPharms Co., Ltd., Seoul, Korea (Rep.)<br />

*e-mail: hyk@khu.ac.kr<br />

In Mycobacteria tuberculosis, a gene, PPX, that encodes exopolyphosphatase (exopoly(p)ase; EC 3.6.1.11) <str<strong>on</strong>g>of</str<strong>on</strong>g> 385<br />

amino acids was found upstream <str<strong>on</strong>g>of</str<strong>on</strong>g> the gene <str<strong>on</strong>g>for</str<strong>on</strong>g> envelope protein (Env). Since Env is started in C-terminal<br />

porti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> PPX gene, they do c<strong>on</strong>stitute an oper<strong>on</strong>. The gene product <str<strong>on</strong>g>of</str<strong>on</strong>g> PPX(mtPPX) was overproduced in<br />

E. coli and its activity was evaluated. Orthophosphate(Pi) is released from polyphosphate, the average chian<br />

length determined. The amount <str<strong>on</strong>g>of</str<strong>on</strong>g> Pi released matched the amount <str<strong>on</strong>g>of</str<strong>on</strong>g> polyp(p) lost. Thus PPX encodes an<br />

enzyme that has expoly(p)ase activity. Since this enzyme show the oper<strong>on</strong> with envelope gene, futher<br />

c<strong>on</strong>siderati<strong>on</strong> speculated.<br />

2H. OXIDATIVE STRESS<br />

2H_24_P<br />

(poster secti<strong>on</strong> A2, poster board #177, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ALPHA SUBUNIT OF F1FO-ATP SYNTHASE IS ASSOCIATED WITH TRANSPORT<br />

OF FE++/CA++ IN HEART MITOCHONDRIA<br />

Mi-Sun Kim, Eunsook S<strong>on</strong>g<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biological science, Sookmyung women’s university, Seoul, 140-742, Korea<br />

e-mail: erate@sookmyung.ac.kr<br />

Previous proteomic analysis revealed overexpressed F1 subunits in ir<strong>on</strong> overloaded heart mitoch<strong>on</strong>dria. As<br />

abnormal amount or assembly <str<strong>on</strong>g>of</str<strong>on</strong>g> F 1 F o -ATP synthase may affect mitoch<strong>on</strong>drial cristae and functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> other<br />

respiratory enzymes, we tried to detect and found alpha and beta subunits in the matrix <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>dria.<br />

Mimicking ir<strong>on</strong> overload c<strong>on</strong>diti<strong>on</strong>, we did a transfecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> alpha subunit into the primary cardiac myocytes<br />

(PCM) to prepare alpha-overexpressed cells. Mitoch<strong>on</strong>drial labile ir<strong>on</strong> pool (LIP) was reduced by 60%,<br />

10


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

whereas calcium c<strong>on</strong>tent was increased 2-fold. Mitoch<strong>on</strong>drial ATP c<strong>on</strong>tent, membrane potential and<br />

NAD(P)H c<strong>on</strong>tent were reduced by 43%, 59%, and 75%, respectively. By ir<strong>on</strong> treatment <str<strong>on</strong>g>of</str<strong>on</strong>g> the transfected<br />

cells, the mitoch<strong>on</strong>drial LIP and NAD(P)H c<strong>on</strong>tents were augmented by 6- and 1.8-fold. Mitoch<strong>on</strong>drial<br />

calcium and membrane potential, however, were declined by 45% and by 63%. In the cells, these changes<br />

were reversed compared with in mitoch<strong>on</strong>dria. ATP was dramatically reduced in mitoch<strong>on</strong>dria, but, the<br />

cellular level was similar. These alterati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> ir<strong>on</strong> and calcium were inverted by Ruthenium red (blocker <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

mitoch<strong>on</strong>drial calcium uniport). There<str<strong>on</strong>g>for</str<strong>on</strong>g>e, we suggest that alpha subunit <str<strong>on</strong>g>of</str<strong>on</strong>g> the enzyme is associated with<br />

ir<strong>on</strong>/calcium transport <str<strong>on</strong>g>of</str<strong>on</strong>g> mitoch<strong>on</strong>dria.<br />

2H_25_P<br />

(poster secti<strong>on</strong> A2, poster board #178, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

OXIDATIVE STRESS FAILS TO INDUCE PROTEIN MISFOLDING IN<br />

REPLICATING CELLS<br />

Eszter Mária Végh, Ákos Putics, Péter Csermely, Csaba Sőti<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Chemistry, Semmelweis University, Budapest, PO Box 260, Hungary<br />

e-mail: csaba@puskin.sote.hu<br />

A hallmark <str<strong>on</strong>g>of</str<strong>on</strong>g> aging and a number <str<strong>on</strong>g>of</str<strong>on</strong>g> degenerative diseases is the increase in oxidized proteins. In these states<br />

misfolded and aggregated protein species also accumulate, especially in postmitotic cells. Hydrophobic<br />

surface exposure <str<strong>on</strong>g>of</str<strong>on</strong>g> misfolded proteins is a potent inducer <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress resp<strong>on</strong>se, signalling the<br />

c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>al state <str<strong>on</strong>g>of</str<strong>on</strong>g> the proteome towards the maintenance mechanisms. However, the causal<br />

relati<strong>on</strong>ship between oxidative stress and protein misfolding is an open questi<strong>on</strong>. In the present study we<br />

investigated whether a localized protein damage induced by oxidative agents (H2O2 and ir<strong>on</strong>-ascorbate)<br />

would lead to protein unfolding/aggregati<strong>on</strong> and stress resp<strong>on</strong>se in replicative lymphocytic and epithelial cell<br />

cultures. We found that in c<strong>on</strong>trast to heat shock, oxidative stress was unable to induce an increase in<br />

hydrophobic surface exposure in live cells, cell extracts and purified proteins. Similarly, we could not observe<br />

an increase in protein aggregati<strong>on</strong>, a direct c<strong>on</strong>sequence <str<strong>on</strong>g>of</str<strong>on</strong>g> protein unfolding. Moreover, oxidative stress did<br />

not lead to HSF-1-dependent transcripti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the major heat shock genes, another sensitive marker <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

protein unfolding. These results suggest that short term oxidative stress does not lead to bulk protein<br />

misfolding in replicating cells and raise important questi<strong>on</strong>s about the role <str<strong>on</strong>g>of</str<strong>on</strong>g> free radicals in protein<br />

homeostasis in aging postmitotic cells.<br />

11


23-26 August 2007,<br />

Budapest, Hungary<br />

2J. STRESS SIGNALING<br />

2J_10_P<br />

(poster secti<strong>on</strong> A2, poster board #298, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

NF-κB ACTIVATION BY HEAT SHOCK: INVOLVEMENT IN CELL SURVIVAL<br />

Mathieu Niv<strong>on</strong>, André-Patrick Arrigo, Carole Kretz-Remy<br />

Université Claude Bernard Ly<strong>on</strong> I, CGMC UMR5534, Bat. G. Mendel, 16 Rue Dubois,<br />

F-69622 Villeurbanne Cédex, France<br />

e-mail: Kretz@univ-ly<strong>on</strong>1.fr<br />

NF-κB bel<strong>on</strong>gs to the Rel/NF-κB family <str<strong>on</strong>g>of</str<strong>on</strong>g> transcripti<strong>on</strong> factors. It is c<strong>on</strong>sidered as a central regulator <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

immune and inflammatory resp<strong>on</strong>ses but also <str<strong>on</strong>g>of</str<strong>on</strong>g> cell death. More recently, it was shown that NF-κB is also<br />

activated in resp<strong>on</strong>se to various stresses. In this respect, we dem<strong>on</strong>strated that NF-κB is activated by heat<br />

shock, but this activati<strong>on</strong> takes place <strong>on</strong>ly during the recovery period after the heat stress. By analyzing the<br />

transducti<strong>on</strong> pathway leading to NF-κB activati<strong>on</strong>, we dem<strong>on</strong>strated that no new transcripti<strong>on</strong>al and<br />

translati<strong>on</strong>al events are required <str<strong>on</strong>g>for</str<strong>on</strong>g> NF-κB activati<strong>on</strong> during heat shock recovery. Moreover, NF-κB<br />

activati<strong>on</strong> after a heat shock does not depend <strong>on</strong> a prior phosphorylati<strong>on</strong> and/or degradati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> IκB<br />

members but <strong>on</strong> the dissociati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> NF-κB from its inhibitory partner (JBC (2001),276). There<str<strong>on</strong>g>for</str<strong>on</strong>g>e we report<br />

a new mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> NF-κB activati<strong>on</strong> by heat shock that does not follow the can<strong>on</strong>ical signal transducti<strong>on</strong><br />

pathway usually described. NF-κB activati<strong>on</strong> during heat shock recovery raises the questi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> deciphering to<br />

which process (immunity, inflammati<strong>on</strong>, cell survival) NF-κB could participate after heat shock. To answer<br />

this questi<strong>on</strong> we decided to analyze the heat shock resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> cells devoid <str<strong>on</strong>g>of</str<strong>on</strong>g> NF-κB and c<strong>on</strong>structed HeLa<br />

cells depleted in p65 subunit by RNA interference strategy. We first could observe that the “heat shock<br />

resp<strong>on</strong>se” is not altered in the various shRNA cell lines. Moreover, our first experiments show that HeLa<br />

cells with very low levels <str<strong>on</strong>g>of</str<strong>on</strong>g> p65 are much more sensitive to heat shock treatments than the wild type,<br />

suggesting that NF-κB, in additi<strong>on</strong> to Hsps, could help the cell to recover from thermal damage.<br />

2K. GENE EXPRESSION REGULATION UNDER STRESS<br />

2K_16_P<br />

(poster secti<strong>on</strong> A2, poster board #172, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

BASIS FOR THE DEVELOPMENT OF A RT-PCR ASSAY TO DETECT ENDOCRINE<br />

DISRUPTION IN REPTILES<br />

Roldán A. Valverde<br />

Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Biological Sciences, Southeastern Louisiana University, Hamm<strong>on</strong>d, LA 70402, USA<br />

There is an accumulating body <str<strong>on</strong>g>of</str<strong>on</strong>g> evidence indicating that envir<strong>on</strong>mental pollutants may behave as endocrine<br />

disruptors <str<strong>on</strong>g>of</str<strong>on</strong>g> vertebrates. Many <str<strong>on</strong>g>of</str<strong>on</strong>g> these pollutants, such as polychlorinated biphenyls (PCBs) and<br />

hexachlorobencene (HCB), are known to persist in the envir<strong>on</strong>ment and to alter the normal functi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

hypothalamo-pituitary-thryroid and HP-adrenal (HPTA) axes. Thyroid and stress horm<strong>on</strong>es (TSs) play<br />

important roles in the development <str<strong>on</strong>g>of</str<strong>on</strong>g> the nervous system and in the mediati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the endocrine stressresp<strong>on</strong>se<br />

in vertebrates. Disrupti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these systems may significantly impair the ability <str<strong>on</strong>g>of</str<strong>on</strong>g> the wildlife to<br />

12


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

adapt to envir<strong>on</strong>mental changes. To investigate the impact <str<strong>on</strong>g>of</str<strong>on</strong>g> these pollutants <strong>on</strong> the HPTA axes <str<strong>on</strong>g>of</str<strong>on</strong>g> n<strong>on</strong>homeothermic<br />

vertebrates we have cl<strong>on</strong>ed both the thyrotropin-releasing horm<strong>on</strong>e (TSHb) and<br />

proopiomelanocortin (POMC) full length cDNAs from the red-eared slider (Trachemys scripta) freshwater<br />

turtle pituitary. The cl<strong>on</strong>ed cDNA transcripts exhibit extensive sequence c<strong>on</strong>servati<strong>on</strong> relative to other<br />

vertebrate sequences. The elucidati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these cDNA transcript sequences are allowing us to develop a<br />

sensitive and specific real time PCR assay to evaluate the impact <str<strong>on</strong>g>of</str<strong>on</strong>g> envir<strong>on</strong>mental pollutants (PCBs and<br />

HCBs) <strong>on</strong> the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the POMC and TSH genes <str<strong>on</strong>g>of</str<strong>on</strong>g> the reptilian HPTA axes. In additi<strong>on</strong>, the cl<strong>on</strong>es<br />

will help us gain an improved understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> the physiological role <str<strong>on</strong>g>of</str<strong>on</strong>g> TSs in the turtle.<br />

2K_17_P<br />

(poster secti<strong>on</strong> A2, poster board #173, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EVALUATION OF GENES EXPRESSION IN MAIZE GENOTYPES WITH<br />

DIFFERENT DEGREES OF DROUGHT TOLERANCE,<br />

AT THE FIRST STAGES OF DEVELOPMENT<br />

D. Badicean 1 , A. Jacota 1 , N. Barbacaru 1 , M Scholz 2 , J. Gardiner 2 , V. Chandler 2,3<br />

1Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Genetics and Plant Physiology <str<strong>on</strong>g>of</str<strong>on</strong>g> Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Science <str<strong>on</strong>g>of</str<strong>on</strong>g> Republic <str<strong>on</strong>g>of</str<strong>on</strong>g> Moldova,<br />

2University <str<strong>on</strong>g>of</str<strong>on</strong>g> Ariz<strong>on</strong>a, 303 Forbes Building, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Sciences, Tucs<strong>on</strong>, AZ 85721 USA,<br />

3University <str<strong>on</strong>g>of</str<strong>on</strong>g> Ariz<strong>on</strong>a, 303 Forbes Building, BIO5 Institute, Tucs<strong>on</strong>, AZ 85721 USA<br />

Drought stress is c<strong>on</strong>sidered <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> the greatest limiting factors in agriculture and the ability <str<strong>on</strong>g>of</str<strong>on</strong>g> crop plants to<br />

adapt to such c<strong>on</strong>diti<strong>on</strong>s is crucial <str<strong>on</strong>g>for</str<strong>on</strong>g> worldwide crop producti<strong>on</strong>. Because <str<strong>on</strong>g>of</str<strong>on</strong>g> increasing scarcity <str<strong>on</strong>g>of</str<strong>on</strong>g> water<br />

resources today objective is creati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> drought resistant crops and/or the development <str<strong>on</strong>g>of</str<strong>on</strong>g> plant varieties<br />

with efficient water use. This is possible through deeply study <str<strong>on</strong>g>of</str<strong>on</strong>g> genes involved in drought stress resp<strong>on</strong>ses.<br />

The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> our work was the evaluati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> genes expressi<strong>on</strong> in maize genotypes with different degrees <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

drought tolerance, at the first stages <str<strong>on</strong>g>of</str<strong>on</strong>g> development, using 70-mer olig<strong>on</strong>ucleotide microarrays, representing<br />

more than 50000 identifiable maize genes. Were used maize genotypes, in comm<strong>on</strong> use within the Republic<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> Moldova, from the maize germplasm collecti<strong>on</strong> at the Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Genetics and Plant Physiology <str<strong>on</strong>g>of</str<strong>on</strong>g> ASM.<br />

Using 20% false discovery rate, AveGI, AveRI > 500 as a cut<str<strong>on</strong>g>of</str<strong>on</strong>g>f parameters we focused <strong>on</strong> genes with<br />

differentially expressi<strong>on</strong> > 3.5 fold compared with c<strong>on</strong>trol plants. Were identified 73-212 significantly<br />

differentially expressed genes am<strong>on</strong>g analyzed genotypes. Number <str<strong>on</strong>g>of</str<strong>on</strong>g> differentially down regulated genes was<br />

greater than that <str<strong>on</strong>g>of</str<strong>on</strong>g> up regulated. For n<strong>on</strong>resistant and low resistant genotypes the proporti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> differentially<br />

expressed genes was higher than that <str<strong>on</strong>g>for</str<strong>on</strong>g> drought resistant and medium resistant genotypes. From all<br />

identified genes 35%-45%, depending <str<strong>on</strong>g>of</str<strong>on</strong>g> genotype, are genes with unknown functi<strong>on</strong> or not annotated genes.<br />

Elucidating their roles and functi<strong>on</strong>s may help in better understanding the complex plants resp<strong>on</strong>se to<br />

drought stress.<br />

13


23-26 August 2007,<br />

Budapest, Hungary<br />

14<br />

2K_18_P<br />

(poster secti<strong>on</strong> A2, poster board #174, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DIAGNOSIS OF COLD STRESS RESPONSE NETWORK AND CROSS-TALK<br />

MAPPING IN CRUCIFERAE PLANTS<br />

Youn Yung Byun 1 , Hyejun Joo 1 , Shinae Yu 1 , Minkyung Choi 1 , Jihye Hwang 1 , Youngsoo Park 2 ,<br />

Cheehark Harn 2 , James Collins 3 , Tim Gardner 3 , D<strong>on</strong>ghee Lee 1<br />

1. Dept. Biological Sciences, Ewha Womans University, Seoul, Korea<br />

2. Biotechnology Center, N<strong>on</strong>g-Woo Bio Co., Ltd., Ky<strong>on</strong>ggi, Korea<br />

3. Dept. Biomedical Engineering, Bost<strong>on</strong>, USA<br />

High level <str<strong>on</strong>g>of</str<strong>on</strong>g> sequence similarity and genetic c<strong>on</strong>servati<strong>on</strong> within plants <str<strong>on</strong>g>of</str<strong>on</strong>g> same family allow us to use the<br />

in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>s and cDNA microarray obtained from a model plant such as Arabidopsis <str<strong>on</strong>g>for</str<strong>on</strong>g> better<br />

understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> n<strong>on</strong>-model plants within the same family, <str<strong>on</strong>g>for</str<strong>on</strong>g> example, Chinese cabbage. Several lines <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

ec<strong>on</strong>omic plants <str<strong>on</strong>g>of</str<strong>on</strong>g> Cruciferae with different sensitivity to cold stress were selected and the gene expressi<strong>on</strong><br />

pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iles under cold stress were examined using 1.6K specialized cDNA microarray. For the comparative<br />

analysis between Arabidopsis and ec<strong>on</strong>omic plants, we adopted a recently developed computati<strong>on</strong>al method<br />

called "Gene Set Enrichment Analysis (GSEA)" which determines whether defined set <str<strong>on</strong>g>of</str<strong>on</strong>g> genes show<br />

statically significant and c<strong>on</strong>cordant differences between two biological states. Al<strong>on</strong>g this, five different gene<br />

sets including a network gene set based <strong>on</strong> a regulatory gene network model <str<strong>on</strong>g>for</str<strong>on</strong>g> early cold stress resp<strong>on</strong>se and<br />

a co-expressi<strong>on</strong> gene set based <strong>on</strong> 1,500 expressi<strong>on</strong> data were built in this lab. With these gene sets and<br />

GSEA method, the expressi<strong>on</strong> data was analyzed to pinpoint the group <str<strong>on</strong>g>of</str<strong>on</strong>g> genes potentially resp<strong>on</strong>sible <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

the difference <str<strong>on</strong>g>of</str<strong>on</strong>g> stress sensitivity between two different plants. Since the plant encounters stress<br />

combinati<strong>on</strong>s c<strong>on</strong>currently or separated temporally and must present an integrated resp<strong>on</strong>se to them, we built<br />

'Cross-talk map' using 63 expressi<strong>on</strong> data <str<strong>on</strong>g>of</str<strong>on</strong>g> Arabidopsis under 9 different envir<strong>on</strong>mental stresses. Utilizing<br />

this cross-talk map, the significance <str<strong>on</strong>g>of</str<strong>on</strong>g> the identified group <str<strong>on</strong>g>of</str<strong>on</strong>g> genes was evaluated <str<strong>on</strong>g>for</str<strong>on</strong>g> their practical<br />

applicati<strong>on</strong> to enhance stress tolerance. Currently, we identified several promising genes at a cross-talk point<br />

and are pursuing transgenic engineering to enhance the stress tolerance against more than two stress<br />

c<strong>on</strong>diti<strong>on</strong>s.<br />

2K_19_P<br />

(poster secti<strong>on</strong> A2, poster board #175, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECT OF LOW-OXYGEN STRESS ON GENE EXPRESSION IN ARABIDOPSIS<br />

THALIANA ECOTYPES<br />

JI-Hye Hwang, Youn-Jung Byun, D<strong>on</strong>g-Hee Lee *<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Life Science, Ewha Womans University, Seoul, Korea<br />

Flooding, defined as lack <str<strong>on</strong>g>of</str<strong>on</strong>g> oxygen at the plant, is <strong>on</strong>e <str<strong>on</strong>g>of</str<strong>on</strong>g> various envir<strong>on</strong>mental stresses. Plants have<br />

developed adaptati<strong>on</strong> mechanisms at both the physiological and the structural levels to enhance their ability<br />

to survive low-oxygen stress. The effect <str<strong>on</strong>g>of</str<strong>on</strong>g> the low-oxygen stress <strong>on</strong> Arabidopsis was investigated at growth<br />

and gene expressi<strong>on</strong> level. To gain insight into how plants resp<strong>on</strong>d to low-oxygen, we analyzed the gene<br />

expressi<strong>on</strong> patterns <str<strong>on</strong>g>of</str<strong>on</strong>g> Arabidopsis using Oper<strong>on</strong> microarray(70-mer olig<strong>on</strong>ucleotide). Firstly, gene<br />

expressi<strong>on</strong> pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iling was carried out at five time point under hypoxic stress in root <str<strong>on</strong>g>of</str<strong>on</strong>g> Arabidopsis<br />

Columbia(Col-0). Hypoxic-resp<strong>on</strong>sive genes (HRG) were selected in gene expressi<strong>on</strong> pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iling. The time


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

series analysis has enabled to identify a set <str<strong>on</strong>g>of</str<strong>on</strong>g> transcripti<strong>on</strong> factors and signal transducti<strong>on</strong> comp<strong>on</strong>ents that<br />

could play a role in the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the hypoxic resp<strong>on</strong>se. Sec<strong>on</strong>dly, the resp<strong>on</strong>se to hypoxia stress in<br />

Columbia(Col-0) and Cape Verdi Islands(cvi) <str<strong>on</strong>g>of</str<strong>on</strong>g> Arabidopsis ecotypes was compared. Cvi showed more<br />

tolerable than Col-0 under low-oxygen c<strong>on</strong>diti<strong>on</strong>, while the general features <str<strong>on</strong>g>of</str<strong>on</strong>g> morphology and growth are<br />

within normal range. The difference <str<strong>on</strong>g>of</str<strong>on</strong>g> gene expressi<strong>on</strong> pattern in two ecotypes was examined using the<br />

microaray. The coupling <str<strong>on</strong>g>of</str<strong>on</strong>g> microarray data with functi<strong>on</strong>al analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> candidate genes will lead to a more<br />

comprehensive understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> the mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> flooding-tolerance and reveals genes that may be<br />

important to enhance the flooding tolerance and will be used to develop the flooding-resistant plants.<br />

2M. OTHER CELLULAR STRESS RELATED TOPICS (STRESS-RELATED PROTEIN<br />

DEGRADATION, STRESS AND CELL MOVEMENT, STRESS PROTEINS AND THE<br />

CELL CYCLE, EXTREMOPHILES)<br />

2M_08_P<br />

(poster secti<strong>on</strong> A2, poster board #176, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

TRANSGENICS OR GENE UP-REGULATION; DHAC INDUCED INCREASE IN<br />

GENE EXPRESSION LEVELS OF TRANSGENE GSHI AND ENDOGENOUS<br />

POPLAR GENE GSH1 (POPULUS X CANESCENS) UNDER PARAQUAT STRESS<br />

P. Lehoczky 1 , G. Gyulai 1 , R. P. Mal<strong>on</strong>e 2 , A. Bittsánszky 1,3,4 , G. Gullner 3 , J. Kiss 1 , M. Czakó 4 ,<br />

I. Bock 1 , L. Márt<strong>on</strong> 4 , T. Kőmíves 4 , L. Heszky 1<br />

1St. Stephanus University, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Genetics and Plant Breeding, Gödöllő, 2103, Hungary; 2 Dublin Institute<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> Technology, School <str<strong>on</strong>g>of</str<strong>on</strong>g> Food Science and Envir<strong>on</strong>mental Health, Dublin 1, Ireland; 3 Plant Protecti<strong>on</strong> Institute,<br />

Hungarian Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences, Budapest, 1525, Hungary; 4 University <str<strong>on</strong>g>of</str<strong>on</strong>g> South Carolina, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biological<br />

Sciences, Columbia, SC 29208, USA; e-mail: gyulai.gabor@mkk.szie.hu<br />

The resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g> relative gene expressi<strong>on</strong> levels <str<strong>on</strong>g>of</str<strong>on</strong>g> transgene gshI and endogenous poplar gene gsh1 to DHAC<br />

(5,6-dihydro-5'-azacytidine hydrochloride) treatment (at 10 -4 M <str<strong>on</strong>g>for</str<strong>on</strong>g> 7 days) were analyzed by qRT-PCR (reverse<br />

transcriptase quantitative PCR) using two types <str<strong>on</strong>g>of</str<strong>on</strong>g> gshI transgenic poplar (Populus x canescens) cl<strong>on</strong>es (11ggs and<br />

6Lgl) and the wild type (WT). For internal c<strong>on</strong>trols, the c<strong>on</strong>stitutively expressed housekeeping genes a-tubulin<br />

and actin were applied coupled with 2 −∆∆Ct method <str<strong>on</strong>g>for</str<strong>on</strong>g> data analysis. High expressi<strong>on</strong> levels <str<strong>on</strong>g>of</str<strong>on</strong>g> transgene gshI<br />

were observed in the 6Lgl cl<strong>on</strong>e (13.5-fold increase) compared to the 11ggs (1.0) sample. The expressi<strong>on</strong> level<br />

doubled (1.8-fold increase) in the DHAC-treated 6Lgl samples but not in the 11ggs cl<strong>on</strong>e (0.4-fold). C<strong>on</strong>trary<br />

to this, the relative copy number <str<strong>on</strong>g>of</str<strong>on</strong>g> transgene gshI in the 6Lgl cl<strong>on</strong>e was found to be 60 % less (1.0) than in<br />

the 11ggs sample (1.6). Relative expressi<strong>on</strong> level <str<strong>on</strong>g>of</str<strong>on</strong>g> endogenous poplar gene gsh1 showed significantly higher<br />

resp<strong>on</strong>siveness to DHAC-induced demethylati<strong>on</strong> than the transgene gshI, with the highest expressi<strong>on</strong> level in<br />

the untrans<str<strong>on</strong>g>for</str<strong>on</strong>g>med WT poplar (19.7-fold increase) compared to trans<str<strong>on</strong>g>for</str<strong>on</strong>g>med cl<strong>on</strong>es <str<strong>on</strong>g>of</str<strong>on</strong>g> 6Lgl (8.7-fold increase)<br />

and 11ggs (2.5-fold increase), respectively. Competiti<strong>on</strong> in the reactivati<strong>on</strong> capacity between transgene gshI and<br />

poplar gsh1 <str<strong>on</strong>g>of</str<strong>on</strong>g> 6Lgl cl<strong>on</strong>e was also observed as the relative gene expressi<strong>on</strong> level <str<strong>on</strong>g>of</str<strong>on</strong>g> transgene gshI increased<br />

from a high relative expressi<strong>on</strong> level (13.5) up by about two fold (1.8 times) rate (to 23.7) compared to poplar<br />

gsh1 gene that increased by an 8.7 increment from a lower level (1.6 rel. expressi<strong>on</strong>) to 13.9. When leaf<br />

samples <str<strong>on</strong>g>of</str<strong>on</strong>g> the 11ggs cl<strong>on</strong>e were exposed to paraquat (PQ) stress at a bleaching c<strong>on</strong>centrati<strong>on</strong> (4 x 10 -7 M) the<br />

relative gene expressi<strong>on</strong> level <str<strong>on</strong>g>of</str<strong>on</strong>g> gshI increased from 1.0 (without DHAC treatment) and 0.4 (DHAC treated)<br />

to 10.0 (combined treatment with DHAC and PQ). Unexpectedly, the gene expressi<strong>on</strong> levels <str<strong>on</strong>g>of</str<strong>on</strong>g> endogenous<br />

gsh1 <str<strong>on</strong>g>of</str<strong>on</strong>g> 11ggs also showed the same activity range from 0.8 (untreated) to 2.0 (DHAC treated) to 28.0 (DHAC<br />

and PQ treated). These results indicate a more extensive stress activati<strong>on</strong> capacity <str<strong>on</strong>g>of</str<strong>on</strong>g> endogenous poplar gene<br />

15


23-26 August 2007,<br />

Budapest, Hungary<br />

gsh1 over the transgenic gshI in the 11ggs cl<strong>on</strong>e, which may imply an alternative gene technology <str<strong>on</strong>g>for</str<strong>on</strong>g> DHAC<br />

induced up-regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> endogenous genes rather than transgenic technology.<br />

16<br />

3C. CHAPERONE NETWORKS AND STRESS PROTEOMICS<br />

3C_06_P<br />

(poster secti<strong>on</strong> A1, poster board #30, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PROTEOME ANALYSIS OF HUMAN MACROPHAGES REVEALS UP-REGULATION<br />

OF APOPTOSIS AND CELL STRESS ASSOCIATED PROTEINS<br />

IN RESPONSE TO INFLUENZA A VIRUS INFECTION<br />

Tuula A. Nyman 1 , Tiina Öhman 1 , Johanna Rintahaka 2 , Nisse Kalkkinen 1 , Sampsa Matikainen 2<br />

1Protein Chemistry <str<strong>on</strong>g>Research</str<strong>on</strong>g> Group, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Biotechnology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Helsinki and 2 Unit <str<strong>on</strong>g>of</str<strong>on</strong>g> Excellence in<br />

Immunotoxicology, Finnish Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Occupati<strong>on</strong>al Health, Helsinki, Finland.<br />

Alveolar macrophages <str<strong>on</strong>g>of</str<strong>on</strong>g> the lung are the primary targets <str<strong>on</strong>g>for</str<strong>on</strong>g> respiratory viruses. Virus-encoded singlestranded<br />

and double-stranded RNA are recognized by host pattern recogniti<strong>on</strong> receptors. This ssRNA and<br />

dsRNA recogniti<strong>on</strong> leads to the changes in the host cell protein expressi<strong>on</strong>. Here we have studied the<br />

regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> influenza A induced protein expressi<strong>on</strong> in human macrophages. Macrophages were infected<br />

with influenza A virus after which the cytosolic and mitoch<strong>on</strong>drial cell fracti<strong>on</strong>s were prepared and analyzed<br />

by using two-dimensi<strong>on</strong>al electrophoresis <str<strong>on</strong>g>for</str<strong>on</strong>g> protein separati<strong>on</strong> and mass spectrometry <str<strong>on</strong>g>for</str<strong>on</strong>g> protein<br />

identificati<strong>on</strong>. In mitoch<strong>on</strong>drial proteomes expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> actin was highly up-regulated up<strong>on</strong> Influenza A<br />

virus infecti<strong>on</strong>. Actin is known to associate with mitoch<strong>on</strong>dria at early phases <str<strong>on</strong>g>of</str<strong>on</strong>g> apoptosis and accordingly<br />

caspase-3, which is the effector caspase in apoptosis, was activated in infected macrophages. In cytosolic<br />

proteomes expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> several heat shock proteins (Hsp60, 70, and 90) and small calcium binding proteins<br />

(S-100A and S-100 Ca + binding protein) was clearly up-regulated. In additi<strong>on</strong>, expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Rho GDP<br />

dissociati<strong>on</strong> inhibitor as well as cathepsins B and D were clearly down-regulated up<strong>on</strong> Influenza A infecti<strong>on</strong>.<br />

In c<strong>on</strong>clusi<strong>on</strong>, our results show that the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> several key proteins involved in apoptosis and stress<br />

resp<strong>on</strong>se are regulated during influenza A virus infecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> human primary macrophages.<br />

4A. PLANT OXIDATIVE STRESS<br />

(IMRE VASS)<br />

4A_11_P<br />

(poster secti<strong>on</strong> B2, poster board #319, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

IDENTIFICATION OF GENES AFFECTING FREEZING TOLERANCE IN WHEAT<br />

BY TRANSCRIPT PROFILING<br />

G. Kocsy 1* , T. Kellős 1 , P. Schweizer 2 , G. Galiba 1 , N. Stein 2<br />

1 Agricultural <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> HAS, 2462Mart<strong>on</strong>vásár, Hungary<br />

2 Leibniz Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Genetics and Crop Plant <str<strong>on</strong>g>Research</str<strong>on</strong>g> (IPK), 06466 Gatersleben, Germany<br />

*corresp<strong>on</strong>ding author: kocsyg@mail.mgki.hu<br />

Winter frosts can severely damage cereals, and c<strong>on</strong>sequently result in reduced yield. The freezing tolerance <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

winter wheat significantly increases during autumn when the gradual decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> temperature induces cold<br />

hardening. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> our experiments was the identificati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> candidate genes affecting this trait. The


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

transcript pr<str<strong>on</strong>g>of</str<strong>on</strong>g>ile was compared during cold hardening in a freezing-tolerant and in a freezing-sensitive wheat<br />

(Triticum aestivum L.) chromosome 5A (it has a major effect <strong>on</strong> freezing tolerance) substituti<strong>on</strong> line. A barley<br />

cDNA-macroarray representing 10,297 tentatively unique genes (unigenes) was utilized <str<strong>on</strong>g>for</str<strong>on</strong>g> transcript<br />

pr<str<strong>on</strong>g>of</str<strong>on</strong>g>iling. The high level <str<strong>on</strong>g>of</str<strong>on</strong>g> sequence c<strong>on</strong>servati<strong>on</strong> within genes between barley and wheat granted <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

sufficient level <str<strong>on</strong>g>of</str<strong>on</strong>g> cross hybridisati<strong>on</strong>. During a 21-day period <str<strong>on</strong>g>of</str<strong>on</strong>g> cold acclimati<strong>on</strong> at 2 °C, gene expressi<strong>on</strong><br />

was m<strong>on</strong>itored at 0, 1, 7 and 21 days after starting the treatment. The expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 1822 genes (25 %)<br />

changed during cold hardening, and the transcript levels <str<strong>on</strong>g>of</str<strong>on</strong>g> 173 genes were affected by chromosome 5A. A<br />

general relati<strong>on</strong>ship between the overall number <str<strong>on</strong>g>of</str<strong>on</strong>g> genes with altered expressi<strong>on</strong> and the degree <str<strong>on</strong>g>of</str<strong>on</strong>g> freezing<br />

tolerance was observed. The expressi<strong>on</strong> changes <str<strong>on</strong>g>of</str<strong>on</strong>g> the most important candidate genes have been c<strong>on</strong>firmed<br />

by Northern analysis and real time PCR. These genes coded, am<strong>on</strong>g others, <str<strong>on</strong>g>for</str<strong>on</strong>g> a new Ca 2+ -binding protein<br />

and a protein related to the meristem development which may be involved in the signal transducti<strong>on</strong> and<br />

vegetative to reproductive phase transiti<strong>on</strong>, respectively.<br />

4A_12_P<br />

(poster secti<strong>on</strong> B2, poster board #314, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DIFFERENTIAL TOLERANCE OF MAIZE TO HERBICIDES IS RELATED TO<br />

INDUCTION OF GLUTATHIONE AND GLUTATHIONE-ASSOCIATED ENZYMES<br />

Mamdouh M. Nemat Alla*, Abdel-Hakeem M. Badawi, Nemat M. Hassan, Zeinab M. El-Bastawisy,<br />

Enas G. Badran<br />

Botany Department, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Science at Damietta, Mansoura University, Damietta, Egypt<br />

* e-mail: mamnematalla@mans.edu.eg, Fax: +20.57.2403868, Tel.: +20 57 2400233<br />

The recommended field dose <str<strong>on</strong>g>of</str<strong>on</strong>g> metribuzin, pretilachlor and chlorimur<strong>on</strong>-ethyl differentially reduced shoot<br />

fresh and dry weights <str<strong>on</strong>g>of</str<strong>on</strong>g> ten-days-old maize seedlings during the following 16 days. Metribuzin showed great<br />

and c<strong>on</strong>sistent reducti<strong>on</strong>s, however the reducti<strong>on</strong> induced by pretilachlor and chlorimur<strong>on</strong>-ethyl mostly<br />

nullified by the end <str<strong>on</strong>g>of</str<strong>on</strong>g> the experiment. Moreover, there were differential accumulati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> lipid peroxides,<br />

carb<strong>on</strong>yl groups and H 2 O 2 in maize leaves; metribuzin showed the greatest accumulati<strong>on</strong>. Meanwhile, levels<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> thiol <str<strong>on</strong>g>for</str<strong>on</strong>g>ms and reduced glutathi<strong>on</strong>e (GSH) were much more induced by pretilachlor and chlorimur<strong>on</strong>ethyl<br />

than metribuzin; the c<strong>on</strong>trary was true regarding oxidized glutathi<strong>on</strong>e (GSSG). The ratio <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

GSH/GSSG was highest following pretilachlor treatment and least by metribuzin. On the other hand,<br />

activities <str<strong>on</strong>g>of</str<strong>on</strong>g> glutathi<strong>on</strong>e-S-transferases (GSTs, EC 2.5.1.18), γ-glutamyl-cysteine synthetase (γ-GCS, EC<br />

6.3.2.2), glutathi<strong>on</strong>e synthetase (GS, EC 6.3.2.3), glutathi<strong>on</strong>e peroxidase (GPX, EC 1.15.1.1) and glutathi<strong>on</strong>e<br />

reductase (GR, EC 1.6.4.2) were more enhanced in maize leaves by pretilachlor and chlorimur<strong>on</strong>-ethyl than<br />

metribuzin. These findings suggest occurrence <str<strong>on</strong>g>of</str<strong>on</strong>g> an oxidative stress differentially induced in maize by<br />

herbicides, a state that was most pr<strong>on</strong>ounced with metribuzin. Pretilachlor was c<strong>on</strong>cluded to be the least<br />

phytotoxic to maize while metribuzin was the most, this differential tolerance seemed to<br />

17


23-26 August 2007,<br />

Budapest, Hungary<br />

4A_13_P<br />

(poster secti<strong>on</strong> B2, poster board #315, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

REDUCTION OF STRESS RESULTS IN SUCCESSFUL PLANT REGENERATION<br />

FROM UNIQUE INOCULI IN RICE (ORYZA SATIVA L.)<br />

Al<str<strong>on</strong>g>for</str<strong>on</strong>g>a-Stella G<strong>on</strong>zalez-Cor<strong>on</strong>el 1 , Csanád Gurd<strong>on</strong> 1,2 *, Orhan Kurt 2 , Barnabás Jenes 1<br />

1Plant Trans<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> Group, Agricultural Biotechnology Center, H-2100 Gödöllő, Hungary<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Physiology & Molecular Plant Biology, ELTE, Budapest, H-1117 Hungary<br />

3Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Agr<strong>on</strong>omy, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Agriculture, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Ondokuz Mayıs, 55139, Samsun, Turkey<br />

*e-mail: gcsanad@abc.hu<br />

It is crucial to establish a system <str<strong>on</strong>g>for</str<strong>on</strong>g> plant regenerati<strong>on</strong> from callus induced from any seedling part, which<br />

might be integrated into a transgenic rice producti<strong>on</strong> scheme. We induced 8 normal plantlets from callus <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

38 complete root explants, 30 normal plantlets from callus <str<strong>on</strong>g>of</str<strong>on</strong>g> 24 complete tillers, and 111 normal plantlets <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

24 tillers without root explants. We assume that these plantlets were induced by embryogenesis since no<br />

albino plantlet was produced and all the plantlets were phenotypically similar to seedlings raised normally<br />

from seeds. The regenerati<strong>on</strong> frequency calculated as number <str<strong>on</strong>g>of</str<strong>on</strong>g> regenerated plants divided by the original<br />

number <str<strong>on</strong>g>of</str<strong>on</strong>g> explants and multiplied by 100 is 125%, 462%, and 21% in the case <str<strong>on</strong>g>of</str<strong>on</strong>g> complete tillers, tillers<br />

without root, and complete roots, respectively. Use <str<strong>on</strong>g>of</str<strong>on</strong>g> complete tiller, tiller without root, and complete root<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> callus inducti<strong>on</strong> was decided after observing that leaves and culms either died or failed to produce callus<br />

cut to small pieces. Probably the tissues suffered high stress when cut in segments and died be<str<strong>on</strong>g>for</str<strong>on</strong>g>e callus<br />

inducti<strong>on</strong> could take place. In tissue culture, antioxidant protecti<strong>on</strong> is <str<strong>on</strong>g>of</str<strong>on</strong>g>ten compromised. Due to phenolic<br />

compunds, oxidative stress ensues and free radicals and their reacti<strong>on</strong> products react with macromolecules. It<br />

results in cellular dysfuncti<strong>on</strong> and the cultures become recalcitrant. We have developed a protocol, which can<br />

be integrated into any transgenic rice producti<strong>on</strong> scheme.<br />

18


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

4C. STRESS SIGNALING IN PLANTS<br />

(CSABA KONCZ)<br />

4C_07_P<br />

(poster secti<strong>on</strong> B2, poster board #316, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

MOLECULAR AND PHYSIOLOGICAL IMPACT OF COLD STRESS ON COFFEA SP.<br />

P. Eichler 1,2 , A. Fortunato 1,3 , M. M. Gouveia 4 , F. Partelli 5 , M. M. Chaves 2,3 ,<br />

J. C. Ramalho 1,6 , A. Ribeiro 1,6<br />

1Centro de Ec<str<strong>on</strong>g>of</str<strong>on</strong>g>isiologia, Bioquímica e Biotecnologia Vegetal / Instituto de Investigação Científica Tropical, Quinta<br />

do Marquês, 2784-505 Oeiras, Portugal; 2 Inst. Sup. Agr<strong>on</strong>omia / UTL, Tapada da Ajuda, 1349-017 Lisboa,<br />

Portugal; 3 Inst. Tecnol. Química Biológica / UNL, Quinta do Marquês, 2781-901, Oeiras, Portugal;<br />

4Dept. Biologia and Centro de Estudos da Macar<strong>on</strong>ésia,, Univ. Madeira, Campus da Penteada, 9000-390 Funchal,<br />

Portugal; 5 Universidade Estaual do Norte Fluminense Darcy Ribeiro, CCTA/LFIT, Campos dos Goyatacazes,<br />

RJ, Brazil; 6 Unidade de Biotecnologia Ambiental / FCT / UNL, 2829-516 M<strong>on</strong>te de Caparica, Portugal.<br />

Am<strong>on</strong>g different abiotic stresses, low positive temperature (chilling) limits plant metabolism and is <str<strong>on</strong>g>of</str<strong>on</strong>g>ten<br />

associated to yield reducti<strong>on</strong>s, particularly in tropical plants, such as C<str<strong>on</strong>g>of</str<strong>on</strong>g>fea sp. In order to c<strong>on</strong>tribute to the<br />

understanding <str<strong>on</strong>g>of</str<strong>on</strong>g> the molecular and physiological aspects <str<strong>on</strong>g>of</str<strong>on</strong>g> c<str<strong>on</strong>g>of</str<strong>on</strong>g>fee tolerance to cold we are currently<br />

analysing the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> genes related to the oxidative stress resp<strong>on</strong>se and to photosynthesis by semiquantitative<br />

and real-time RT-PCR, using photosynthesis as an integrative metabolism and a sensor to<br />

acclimati<strong>on</strong> ability. This study involves a multidisciplinary analysis <str<strong>on</strong>g>of</str<strong>on</strong>g> the impacts <str<strong>on</strong>g>of</str<strong>on</strong>g> chilling exposure in some<br />

c<str<strong>on</strong>g>of</str<strong>on</strong>g>fee genotypes with different cold tolerance potential, submitted to an acclimati<strong>on</strong> period, to three chilling<br />

cycles (4ºC) and to a rewarming period. Results <str<strong>on</strong>g>of</str<strong>on</strong>g> gene expressi<strong>on</strong> are in accordance with physiological and<br />

biochemical data previously obtained by our group (Campos et al., 2003; Ramalho et al., 2003) and suggest<br />

that the transcripti<strong>on</strong>al activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the genes under study changes according to the degree <str<strong>on</strong>g>of</str<strong>on</strong>g> cold<br />

tolerance/susceptibility <str<strong>on</strong>g>of</str<strong>on</strong>g> c<str<strong>on</strong>g>of</str<strong>on</strong>g>fee genotypes.<br />

4C_08_P<br />

(poster secti<strong>on</strong> B2, poster board #317, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

INDENTIFICATION OF TRANS-ACTING STRESS FACTORS FROM ARABIDOPSIS<br />

USING NOVEL GENETIC APPROACH<br />

Csaba Papdi, Edit Ábrahám, M. Prathiba Joseph, Csaba K<strong>on</strong>cz, László Szabados<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Biology, Biological <str<strong>on</strong>g>Research</str<strong>on</strong>g> Center, Temesvári krt. 62, 6726-Szeged, Hungary<br />

Max-Planck-Institut für Züchtungs<str<strong>on</strong>g>for</str<strong>on</strong>g>schung, Carl-v<strong>on</strong>-Linné-Weg 10, D–50829 Köln, Germany<br />

e-mail: szabados@brc.hu<br />

We have developed a new genetic strategy to determine yet unknown stress regulatory factors in Arabidopsis.<br />

A full-length cDNA expressi<strong>on</strong> library has been c<strong>on</strong>structed from different tissues <str<strong>on</strong>g>of</str<strong>on</strong>g> Arabidopsis subjected to<br />

salt stress. The cDNA library has been cl<strong>on</strong>ed into the estradiol inducible pER8 plant expressi<strong>on</strong> vector and<br />

trans<str<strong>on</strong>g>for</str<strong>on</strong>g>med into Arabidopsis. T1 generati<strong>on</strong> trans<str<strong>on</strong>g>for</str<strong>on</strong>g>med plants were screened <str<strong>on</strong>g>for</str<strong>on</strong>g> salt tolerance and ABA<br />

insensitivity. A number <str<strong>on</strong>g>of</str<strong>on</strong>g> trans<str<strong>on</strong>g>for</str<strong>on</strong>g>mants with altered stress resp<strong>on</strong>ses have been obtained depending to the<br />

presence <str<strong>on</strong>g>of</str<strong>on</strong>g> estradiol. To indentify trans-acting regulators <str<strong>on</strong>g>of</str<strong>on</strong>g> the stress inducible ADH1 gene we have<br />

c<strong>on</strong>structed luciferase reporter gene fusi<strong>on</strong> with the ADH1 promoter. An ABA resp<strong>on</strong>sive ADH-LUC<br />

19


23-26 August 2007,<br />

Budapest, Hungary<br />

expressing line were used <str<strong>on</strong>g>for</str<strong>on</strong>g> large scale trans<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> with the cDNA library. By screening over 10 000<br />

trans<str<strong>on</strong>g>for</str<strong>on</strong>g>mants we have found 16 lines with increased bioluminescence. One <str<strong>on</strong>g>of</str<strong>on</strong>g> the line am<strong>on</strong>g them has been<br />

selected with highest luciferase activity based <strong>on</strong> estradiol dependent inducti<strong>on</strong>. We determined that the<br />

cDNA insert codes <str<strong>on</strong>g>for</str<strong>on</strong>g> an uncharacterized AP2/EREBP type transcript<strong>on</strong> factor, and revealed that its<br />

induced expressi<strong>on</strong> and ABA treatment has a synergistic effect <strong>on</strong> ADH1 promoter. The estradiol dependent<br />

activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ADH1 promoter suggests that this transcripti<strong>on</strong> factor is a positive regulator <str<strong>on</strong>g>of</str<strong>on</strong>g> ADH1 gene.<br />

4G. HEAVY METAL STRESS<br />

4G_09_P<br />

(poster secti<strong>on</strong> B2, poster board #318, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PHYSIOLOGICAL RESPONSE AND PHYTOACCUMULATION OF CHROMIUM BY<br />

PETROSELINUM CRISPUM<br />

H. Ejtehadi, A. Zaker, P. Abrishamchi, M. Lahouti<br />

Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Biology, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Science, Ferdowsi University <str<strong>on</strong>g>of</str<strong>on</strong>g> Mashhad, Mashhad, Iran<br />

e-mail: hejtehadi@science1.um.ac.ir<br />

Chromium is the seventh most abundant element <strong>on</strong> the earth's crust. Cr +3 is an essential micr<strong>on</strong>utrient <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

the health <str<strong>on</strong>g>of</str<strong>on</strong>g> man and animals while Cr +6 is extremely toxic. To study physiological resp<strong>on</strong>se and<br />

phytoaccumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Cr +3 and Cr +6 by Petroselinum crispum, seedlings were grown in hydrop<strong>on</strong>ic culture<br />

c<strong>on</strong>taining different levels <str<strong>on</strong>g>of</str<strong>on</strong>g> Cr +3 (0.1, 0.25, 0.75, 1, 1.5, 2 and 3 mg/l) and Cr +6 (0.1, 0.25, 0.5 and 0.75 mg/l).<br />

After 5 and 3 weeks treatment with Cr +3 and Cr +6 , the effects <strong>on</strong> growth, chlorophyll a and b c<strong>on</strong>tent and<br />

c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> Ca +2 , K + , Na + , Cr +3 and Cr +6 in the roots and shoots were studied. The results showed that<br />

increasing <str<strong>on</strong>g>of</str<strong>on</strong>g> Cr +3 to 3 and 2-3 mg/l caused statistically significant decreasing <str<strong>on</strong>g>of</str<strong>on</strong>g> dry weight in the root and<br />

shoot, respectively. It was the same <str<strong>on</strong>g>for</str<strong>on</strong>g> Cr +6 but at 0.75 mg/l. Length <str<strong>on</strong>g>of</str<strong>on</strong>g> the roots and shoots decreased<br />

significantly at 0.75 mg/l to 3 mg/l Cr +3 and 0.75 mg/l Cr +6 . Chromium reduced significantly total<br />

chlorophyll, chlorophyll a and b at 3 mg/l Cr +3 and all treatments <str<strong>on</strong>g>of</str<strong>on</strong>g> Cr +6 except 0.1 mg/l. Ca +2 and K +<br />

c<strong>on</strong>centrati<strong>on</strong>s in the roots and shoots were also affected by chromium. Significant decrease <str<strong>on</strong>g>of</str<strong>on</strong>g> Ca +2 , K + and<br />

Na + was observed in 2-3 mg/l Cr + 3 and 0.5-0.75 mg/l Cr +6 , 3 mg/l Cr +3 and 0.25-0.75 mg/l Cr +6 and 3 mg/l<br />

Cr +3 , respectively. Accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> chromium in both roots and shoots enhanced with increasing levels <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Cr +3 and Cr +6 in the culture. Roots accumulated more chromium than the shoots. Visible toxicity symptoms<br />

including shrunk leaves, stunted roots and shoots growth, reduced number and surface <str<strong>on</strong>g>of</str<strong>on</strong>g> leaves and<br />

chlorosis <str<strong>on</strong>g>of</str<strong>on</strong>g> the young leaves were observed at the highest level <str<strong>on</strong>g>of</str<strong>on</strong>g> chromium supplied (3mg/lCr +3 and 0.75<br />

mg/lCr +6 ). Generally, Cr +6 indicated toxicity symptoms at lower c<strong>on</strong>centrati<strong>on</strong>s and shorter time than Cr +3 .<br />

Keywords: Phytoaccumulati<strong>on</strong>, Chromium Stress, Petroselinum crispum<br />

20


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5A. STRESS, AGING AND CELLULAR SENESCENCE<br />

(EFSTATHIOS S. GONOS, GORDON J. LITHGOW)<br />

5A_11_P<br />

(poster secti<strong>on</strong> B1, poster board #256, 25th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EXERCISE TRAINING PROMOTES SIRT1 ACTIVITY IN AGED RATS<br />

C<strong>on</strong>ti Valeria 1 , Corbi Graziamaria 2,3 , Ferrara Nicola 2,3 , Rinaldi Barbara 1 , Stiuso Paola 4 ,<br />

Boccuti Silvia 1 , Rengo Giuseppe 3 , Filippelli Walter 5 , Rossi Francesco 1 , Filippelli Amelia 1<br />

1 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Experimental Medicine and Excellence Center <str<strong>on</strong>g>of</str<strong>on</strong>g> Cardiovascular Disease,<br />

Sec<strong>on</strong>d University <str<strong>on</strong>g>of</str<strong>on</strong>g> Naples, Italy<br />

2 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Health Sciences, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Molise, Italy<br />

3 S. Maugeri Foundati<strong>on</strong>, Scientific Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Telese Terme, Italy<br />

4 Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemestry, Sec<strong>on</strong>d University <str<strong>on</strong>g>of</str<strong>on</strong>g> Naples, Italy<br />

5 Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Exercise Sciences Parthenope University <str<strong>on</strong>g>of</str<strong>on</strong>g> Naples, Italy<br />

The objective <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to determine the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> aging and exercise training <strong>on</strong> SIRT1 activity, and<br />

to identify a pathway linking SIRT1 to antioxidant resp<strong>on</strong>se and cell cycle regulati<strong>on</strong> in rats. SIRT1 is an<br />

NAD + -dependent deacetylase involved in the oxidative stress resp<strong>on</strong>se and aging. The effects <str<strong>on</strong>g>of</str<strong>on</strong>g> aging and <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

moderate and prol<strong>on</strong>ged exercise training in rats are unknown. We measured SIRT1 activity in heart and<br />

adipose tissue <str<strong>on</strong>g>of</str<strong>on</strong>g> young (6 m<strong>on</strong>ths old), sedentary old (24 m<strong>on</strong>ths old) and trained old (24 m<strong>on</strong>ths old) rats<br />

using an assay kit. Peroxidative damage was determined by measuring levels <str<strong>on</strong>g>of</str<strong>on</strong>g> ThioBarbituric Reactive<br />

Substances (TBARS) and the protein-aldehyde adduct 4-hydroxyn<strong>on</strong>enal (4-HNE). MnSOD, catalase and<br />

FOXO3a levels were evaluated by western blot, and GADD45a, cyclin D 2 and FOXO3a mRNA by RT-PCR.<br />

Aging significantly reduced SIRT1 activity in heart, but not in adipose tissue, increased TBARS and 4-HNE,<br />

and decreased Mn-SOD and catalase expressi<strong>on</strong> in both heart and adipose tissue. Aging did not affect<br />

FOXO3a protein expressi<strong>on</strong> in the heart, or FOXO3a mRNA in adipose tissue. Exercise training<br />

significantly increased FOXO3a protein in the heart and FOXO3a mRNA in adipose tissue <str<strong>on</strong>g>of</str<strong>on</strong>g> aged rats. It<br />

also significantly increased Mn-SOD and catalase levels in both heart and adipose tissue. The exerciseinduced<br />

increase in SIRT1 activity in the heart caused a decrease in cyclin D 2 and an increase in GADD45a<br />

mRNA expressi<strong>on</strong>. There was a similar decrease in cyclin D 2 , and no changes in GADD45a mRNA<br />

expressi<strong>on</strong> in adipose tissue. Exercise training, which significantly increases SIRT1 activity, could counteract<br />

age-related systems impairment.<br />

21


23-26 August 2007,<br />

Budapest, Hungary<br />

5B. HSP-S IN CEREBRAL PROTECTION<br />

(IAN R. BROWN)<br />

5B_06_P<br />

(poster secti<strong>on</strong> B1, poster board #257, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ARIMOCLOMOL, AN ORALLY AVAILABLE CHAPERONE AMPLIFIER,<br />

ACCELERATES FUNCTIONAL RECOVERY IN RAT MODELS OF STROKE, EVEN<br />

WHEN INITIAL DRUG TREATMENT IS DELAYED UP TO 48 HOURS<br />

POST-INJURY<br />

S. C. Ng, J. M. Ren, S. P. Finklestein, S. Wieland, J. R. Barber<br />

CytRx Corporati<strong>on</strong><br />

Stroke is the third leading cause <str<strong>on</strong>g>of</str<strong>on</strong>g> death and the number <strong>on</strong>e cause <str<strong>on</strong>g>of</str<strong>on</strong>g> l<strong>on</strong>g-term disability am<strong>on</strong>g Americans,<br />

with up to 30% <str<strong>on</strong>g>of</str<strong>on</strong>g> surviving stroke victims permanently disabled. Since the <strong>on</strong>ly drug currently approved <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

stroke, tPA, is <strong>on</strong>ly effective in the relatively rare ischemic stroke victim that is diagnosed in a therapeutic<br />

window no greater than three hours after the <strong>on</strong>set <str<strong>on</strong>g>of</str<strong>on</strong>g> stroke symptoms, there is a great need to identify<br />

pharmaceutical interventi<strong>on</strong>s that can improve the course <str<strong>on</strong>g>of</str<strong>on</strong>g> stroke recovery even after the rapid primary<br />

neur<strong>on</strong>al damage caused by ischemia. A large body <str<strong>on</strong>g>of</str<strong>on</strong>g> evidence suggests a “penumbra” <str<strong>on</strong>g>of</str<strong>on</strong>g> functi<strong>on</strong>ally<br />

impaired but living tissue surrounding the central core <str<strong>on</strong>g>of</str<strong>on</strong>g> a stroke infarct. These cells apparently c<strong>on</strong>tain toxic<br />

misfolded proteins, as dem<strong>on</strong>strated by the presence <str<strong>on</strong>g>of</str<strong>on</strong>g> ubiquitin staining protein aggregates, and they have<br />

been shown to be protected by molecular chaper<strong>on</strong>es. Since arimoclomol, currently under clinical<br />

investigati<strong>on</strong> as a potential therapy <str<strong>on</strong>g>for</str<strong>on</strong>g> ALS, is an amplifier <str<strong>on</strong>g>of</str<strong>on</strong>g> the normal cell stress chaper<strong>on</strong>e resp<strong>on</strong>se, we<br />

decided to test the ability <str<strong>on</strong>g>of</str<strong>on</strong>g> the drug to improve functi<strong>on</strong>al recovery from stroke.<br />

In the present study, stroke was induced in rats by permanent occlusi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the middle cerebral artery,<br />

blocking blood flow to parts <str<strong>on</strong>g>of</str<strong>on</strong>g> the brain causing cerebral oxygen deprivati<strong>on</strong>. Recovery from stroke was<br />

measured by m<strong>on</strong>itoring sensory-motor skills. An initial study indicated that <strong>on</strong>ce daily oral doses <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

arimoclomol beginning 1h after stroke resulted in a highly significant improvement in the functi<strong>on</strong>al recovery,<br />

matching or exceeding the positive c<strong>on</strong>trol, intracisternal injecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> bFGF. Despite dramatically improved<br />

functi<strong>on</strong>al recovery neither treatment significantly affected the resulting infarct volume measured at the end<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the experiment.<br />

To determine the therapeutic window <str<strong>on</strong>g>of</str<strong>on</strong>g> opportunity <str<strong>on</strong>g>for</str<strong>on</strong>g> improvement <str<strong>on</strong>g>of</str<strong>on</strong>g> stroke recovery by arimoclomol,<br />

initiati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> drug treatment was delayed in the subsequent study. Beginning either 6, 12, 24 or 48 hours after<br />

stroke was induced rats were then administered oral doses <str<strong>on</strong>g>of</str<strong>on</strong>g> either arimoclomol or vehicle <strong>on</strong>ce daily <str<strong>on</strong>g>for</str<strong>on</strong>g> 35<br />

days. While motor skills declined dramatically in all study rats immediately after stroke was induced, those<br />

treated with arimoclomol recovered dramatically faster and more completely than vehicle-treated animals<br />

regardless <str<strong>on</strong>g>of</str<strong>on</strong>g> when treatment was initiated. The functi<strong>on</strong>al recovery results were highly statistically significant<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g> all treated groups but <strong>on</strong>ly relatively small trends toward decreased infarct volume were observed and did<br />

not reach statistical significance. In all <str<strong>on</strong>g>of</str<strong>on</strong>g> the sensory and motor skills tested, even animals whose<br />

arimoclomol treatment began 48 hours after stroke were approximately half-way to complete recovery by day<br />

7 after stroke, whereas motor skills <str<strong>on</strong>g>of</str<strong>on</strong>g> untreated animals were not improved.<br />

Based <strong>on</strong> these impressive results, we are planning a Phase 2 clinical trial with arimoclomol to assess the<br />

potential to accelerate and improve functi<strong>on</strong>al recovery in stroke patients. Additi<strong>on</strong>al results from <strong>on</strong>going<br />

studies will also be presented.<br />

22


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

5D. STRESS AND THE IMMUNE RESPONSE<br />

(TAMAS BARTFAI)<br />

5D_17_P<br />

(poster secti<strong>on</strong> B1, poster board #258, 25th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

A NOVEL SIGNALING INTERACTION UNDERLINES THE EFFECTS OF<br />

CORTICOTROPIN RELEASING HORMONE IN INNATE IMMUNE RESPONSES<br />

Thalia Teli 1 , Katia P. Karalis 1,2<br />

1Foundati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> Biomedical Sciences, Developmental Biology, Soranou Ephessiou 4, Athens, 15527, Greece,<br />

2Divisi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Endocrinology, Children’s Hospital, Harvard Medical School, Bost<strong>on</strong>, 02115 MA, USA<br />

e-mail: kkarali@bioacademy.gr<br />

Corticotropin Releasing Horm<strong>on</strong>e (CRH) is the principal coordinator <str<strong>on</strong>g>of</str<strong>on</strong>g> the hypothalamic-pituitary-adrenal<br />

(HPA) axis, whereby any stressful stimuli, including inflammati<strong>on</strong>, is rapidly communicated to the adrenal<br />

cortex, triggering glucocorticoid release. Glucocorticoid release mediates the anti-inflammatory effects <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

central CRH. In c<strong>on</strong>trast, peripherally expressed CRH is largely proinflammatory. CRH mediates its effects<br />

via binding <strong>on</strong> specific CRH receptors namely CRHR1 and CRHR2 which bel<strong>on</strong>g to the family <str<strong>on</strong>g>of</str<strong>on</strong>g> G protein<br />

coupled receptors (GPCRs).<br />

Toll-like receptor (TLR)-4 is a pattern recogniti<strong>on</strong> receptor member <str<strong>on</strong>g>of</str<strong>on</strong>g> the TLRs family that regulate innate<br />

immunity by specific binding <str<strong>on</strong>g>of</str<strong>on</strong>g> a wide range <str<strong>on</strong>g>of</str<strong>on</strong>g> microbial pathogens. TLR4 binds lipopolysaccharide (LPS),<br />

the main pathogenic comp<strong>on</strong>ent <str<strong>on</strong>g>of</str<strong>on</strong>g> the Gram-negative bacteria cell wall, and triggers a cascade <str<strong>on</strong>g>of</str<strong>on</strong>g> cellular<br />

signals that culminates in the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> MAPK and NFκB and the subsequent inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> proinflammatory<br />

factors. Macrophages are critical cells <str<strong>on</strong>g>for</str<strong>on</strong>g> innate immunity and express TLR4 as well as CRH<br />

receptor(s) (Baker et al., 2003; Tsatsanis et al., 2005). The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to explore the role <str<strong>on</strong>g>of</str<strong>on</strong>g> CRH in<br />

the LPS-mediated inducti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> MAPKs in the murine macrophage cell line, RAW264.7. Activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> TLR4<br />

by LPS (10µg/ml) caused a potent, time- dependent phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> p38, ERK1/2, and JNK. Similarly,<br />

CRH (10 -7 M) stimulati<strong>on</strong> dem<strong>on</strong>strated a robust phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> p38 and ERK1/2, although with a<br />

biphasic pattern <str<strong>on</strong>g>of</str<strong>on</strong>g> inducti<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> the latter. However, p38MAPK dem<strong>on</strong>strated a CRH-mediated suppressi<strong>on</strong><br />

in macrophages co-treated with LPS and CRH, evident at both early and late phases <str<strong>on</strong>g>of</str<strong>on</strong>g> inducti<strong>on</strong>.<br />

Our findings reveal a novel CRH mediated signaling mechanism involved in the innate immune resp<strong>on</strong>se <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

macrophages by selective inhibiti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> p38MAPK. Further studies remain to elucidate other comp<strong>on</strong>ents <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

this signaling mechanism and explore its role in vivo.<br />

23


23-26 August 2007,<br />

Budapest, Hungary<br />

24<br />

5E. ISN SYMPOSIUM ON STRESS ENDOCRINOLOGY<br />

(FRANCOIS TRONCHE)<br />

5E_05_P<br />

(poster secti<strong>on</strong> B1, poster board #245, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CENTRAL, BEHAVIOURAL AND AUTONOMIC ACTIONS OF TROUT<br />

CORTICOTROPIN-RELEASING HORMONE AND UROTENSIN-I IN THE TROUT<br />

Jean-Claude Le Mével a , Nagi Mimassi a , Frédéric Lancien a , J. Michael C<strong>on</strong>l<strong>on</strong> b<br />

a INSERM U650, LaTIM, laboratoire de Neurophysiologie, Université de Bretagne Occidentale, Brest, 29238,<br />

France. b Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemistry, United Arab Emirates University, Al Ain, 17666, UAE<br />

e-mail: jean-claude.lemevel@univ-brest.fr<br />

In fish, the neurohorm<strong>on</strong>al peptides, corticotropin-releasing horm<strong>on</strong>e (CRH) and urotensin I (U-I), an<br />

ortholog <str<strong>on</strong>g>of</str<strong>on</strong>g> mammalian urocortin-1, are known to be released primarily during stress to produce cortisol<br />

secreti<strong>on</strong>. The widespread distributi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these peptides and their receptors within the central nervous system<br />

also suggests that CRH and U-I might be involved in behavioural and aut<strong>on</strong>omic resp<strong>on</strong>ses. However, very<br />

little is known regarding the central effects <str<strong>on</strong>g>of</str<strong>on</strong>g> these peptides in n<strong>on</strong>-mammalian species. C<strong>on</strong>sequently, the<br />

present study was undertaken to compare the central acti<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> a low picomole dose (5 pmol) <str<strong>on</strong>g>of</str<strong>on</strong>g> trout CRH<br />

and U-I <strong>on</strong> motor activity, heart rate (HR) and heart rate variability (HRV) in the unanesthetized trout.<br />

Intracerebroventricular (ICV) injecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> CRH and U-I evoked a significant increase (1400 % and 600 %<br />

respectively) in the durati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> motor activity. In additi<strong>on</strong>, the ICV injecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> CRH produced a significant<br />

decrease in the HR (-12 %) <str<strong>on</strong>g>of</str<strong>on</strong>g> the trout together with a significant increase in the HRV (+ 120 %), a potent<br />

index <str<strong>on</strong>g>of</str<strong>on</strong>g> parasympathetic drive to the heart. In c<strong>on</strong>trast, the ICV injecti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> U-I was without acti<strong>on</strong> <strong>on</strong> HR<br />

and HRV. In c<strong>on</strong>clusi<strong>on</strong>, our results dem<strong>on</strong>strate that, after central injecti<strong>on</strong>s, CRH and U-I exert similar<br />

behavioural acti<strong>on</strong>s but differentially affect the aut<strong>on</strong>omic c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> the heart. This suggests that the two<br />

endogenous peptides may be selectively implicated in important neuroregulatory functi<strong>on</strong>s during stressful<br />

c<strong>on</strong>diti<strong>on</strong>s.<br />

5H. STRESS OF DOMESTIC ANIMALS AND FISH<br />

5H_11_P<br />

(poster secti<strong>on</strong> B1, poster board #294, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CORTICOSTERONE INDUCES CHANGES ON THE ULTRASTRUCTURAL<br />

MORPHOLOGY OF CHICKEN HETEROPHILS AND LYMPHOCYTES<br />

Shaniko Shini, Pete Kaiser, Agim Shini, Wayne L. Bryden<br />

School <str<strong>on</strong>g>of</str<strong>on</strong>g> Animal Studies, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Queensland, Gatt<strong>on</strong> QLD 4343, Australia<br />

Institute <str<strong>on</strong>g>for</str<strong>on</strong>g> Animal Health, Compt<strong>on</strong>, Berkshire RG20 7NN, UK<br />

e-mail: s.shini@uq.edu.au<br />

In chickens, as in mammals a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> stressors, including exposure to stress horm<strong>on</strong>es induces an elevati<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> plasma corticoster<strong>on</strong>e c<strong>on</strong>centrati<strong>on</strong>, and subsequently increases heterophil to lymphocyte (H/L) ratio.<br />

In<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> is lacking c<strong>on</strong>cerning morphological alterati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> chicken heterophils and lymphocytes that<br />

accompany an increase in the H/L ratio. We have c<strong>on</strong>ducted series <str<strong>on</strong>g>of</str<strong>on</strong>g> experiments and observed that in<br />

chickens, an increase in heterophils and decrease in lymphocytes in the peripheral blood is associated with


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

morphological changes <str<strong>on</strong>g>of</str<strong>on</strong>g> circulating cells. We further attempt to elucidate and differentiate cellular and<br />

molecular changes that are linked with an increase in H/L ratio in stressed birds. Here we report results <strong>on</strong><br />

ultrastructural characteristics <str<strong>on</strong>g>of</str<strong>on</strong>g> chicken heterophils and lymphocytes isolated from peripheral blood and<br />

b<strong>on</strong>e marrow. Chickens were exposed to corticoster<strong>on</strong>e and samples were taken <str<strong>on</strong>g>for</str<strong>on</strong>g> haematological,<br />

endocrine, and electr<strong>on</strong> microscopy examinati<strong>on</strong>. Results indicated that in additi<strong>on</strong> to the plasma<br />

corticoster<strong>on</strong>e elevati<strong>on</strong> and an increase in H/L ratio, corticoster<strong>on</strong>e induced changes <strong>on</strong> heterophil size,<br />

shape, and granules, and lymphocyte cytoplasmic characteristics. Moreover, the number <str<strong>on</strong>g>of</str<strong>on</strong>g> immature<br />

heterophils in the b<strong>on</strong>e marrow and peripheral blood was increased (heterophilia), which explains why their<br />

proporti<strong>on</strong> increases during stress. The proporti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> lymphocytes in peripheral blood was decreased<br />

(lymphopenia); this was associated with degenerative changes in morphological features <str<strong>on</strong>g>of</str<strong>on</strong>g> lymphocytes and<br />

dem<strong>on</strong>strated that these cells were undergoing apoptosis. It was suggested that corticoster<strong>on</strong>e stimulates a<br />

release <str<strong>on</strong>g>of</str<strong>on</strong>g> heterophils from b<strong>on</strong>e marrow and the marginated pool into the blood circulati<strong>on</strong> where they are<br />

needed in the beginning stage <str<strong>on</strong>g>of</str<strong>on</strong>g> a resp<strong>on</strong>se to the stressor. The suppressive effect <strong>on</strong> lymphocytes is<br />

associated with apoptosis and/or redistributi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> these cells into other compartments <str<strong>on</strong>g>of</str<strong>on</strong>g> the body. It was<br />

c<strong>on</strong>cluded that, corticoster<strong>on</strong>e activates neuroendocrine and cytokine pathways to which leukocytes<br />

c<strong>on</strong>tribute, and upregulates/downregulates their resp<strong>on</strong>se.<br />

5H_12_P<br />

(poster secti<strong>on</strong> B1, poster board #295, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EFFECT OF SLAUGHTERING METHODS ON CAGED BLUEFIN TUNA<br />

(THUNNUS THYNNUS)<br />

C. Messina*, A. Santulli<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Palermo, Dept. <str<strong>on</strong>g>of</str<strong>on</strong>g> Biochemical Science, Marine Biochemistry Secti<strong>on</strong>,<br />

Lungomare D. Alighieri, 91016, Trapani, Italy,<br />

*e-mail: cmessina@unipa.it<br />

Poor in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <strong>on</strong> different aspects <str<strong>on</strong>g>of</str<strong>on</strong>g> reared bluefin tuna, related to slaughtering and its c<strong>on</strong>sequences <strong>on</strong><br />

stress and to the related effects <strong>on</strong> quality product and its shelf-life are available. As slaughtering techniques<br />

influence the quality <str<strong>on</strong>g>of</str<strong>on</strong>g> the product, this project had the scope to verify the eventual exerted effect by<br />

slaughtering method <strong>on</strong> the quality <str<strong>on</strong>g>of</str<strong>on</strong>g> tuna. In particular, the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> slaughtering by gun <strong>on</strong> tuna fish and<br />

the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> slaughtering with submarine bang-stick were evaluated in tuna reared in Castellammare del<br />

Golfo (Trapani, Italy). Results pointed out a greater stress c<strong>on</strong>diti<strong>on</strong> in animals killed by gun compared to<br />

those killed with bang-stick, as it was shown by muscular and ocular pH values and by all haematic<br />

parameters <str<strong>on</strong>g>of</str<strong>on</strong>g> sec<strong>on</strong>dary stress such as hematocrit, glucose, lactate, MDA underlining higher values in animals<br />

killed by gun. Slaughtering stress, inducing a greater glycogen utilizati<strong>on</strong> in animals killed by gun, reduces<br />

muscular pH and determines greater drip loss and lower quality compared to those animals killed by bangstick.<br />

The immediate c<strong>on</strong>diti<strong>on</strong>ing <str<strong>on</strong>g>of</str<strong>on</strong>g> animals in ice and water after death, reducing depleti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> muscular<br />

glycogen and ATP post mortem and further lactate accumulati<strong>on</strong> in muscle, determines positive effects <strong>on</strong><br />

quality, also in terms <str<strong>on</strong>g>of</str<strong>on</strong>g> technological properties. Tunas processed after 24 hours <str<strong>on</strong>g>of</str<strong>on</strong>g> cold c<strong>on</strong>diti<strong>on</strong>ing show a<br />

lower drip loss compared to those processed after slaughtering and lower TVBN and MDA producti<strong>on</strong>. The<br />

results obtained in the present study show that slaughtering processing methods represent a critical point <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

producti<strong>on</strong> cycle <str<strong>on</strong>g>of</str<strong>on</strong>g> reared bluefin tuna and that the investigated parameters guarantee a significant and<br />

reliable c<strong>on</strong>trol <str<strong>on</strong>g>of</str<strong>on</strong>g> stress level <str<strong>on</strong>g>of</str<strong>on</strong>g> reared animals.<br />

25


23-26 August 2007,<br />

Budapest, Hungary<br />

5I. STRESS AND NO<br />

5I_07_P<br />

(poster secti<strong>on</strong> B1, poster board #296, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ROLE OF NITRIC OXIDE IN CHOLINERGIC CELLS OF LATERODOSAL<br />

TEGMENTAL NUCLEI ON RESTRAINT STRESS IN THE RAT<br />

Ji Hyun Kim 1,2 , Hye Young Joung 2 , Eun Yee Jung 2 , Kyu Sop Lee 2 , Insop Shim 2 , Kwang Ho Pyun 2 *<br />

1Immunobiology and Cell Biology Core Laboratory, Catholic <str<strong>on</strong>g>Research</str<strong>on</strong>g> Institutes <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Science, The Catholic<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Korea, Seoul 137-701, Korea<br />

2Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Integrative Medicine, College <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, The Catholic University <str<strong>on</strong>g>of</str<strong>on</strong>g> Korea, Seoul 137-701, Korea<br />

e-mail: pyunkh@catholic.ac.kr<br />

Nitric oxide (NO) as a retrograde intercellular messenger is known to play a crucial role in physiological<br />

processes including stress resp<strong>on</strong>se. The purpose <str<strong>on</strong>g>of</str<strong>on</strong>g> the present study was to determine whether cholinergic<br />

cell in the laterodosal tegmental neuclei (LDTg) cells after acute stress produce NO-c<strong>on</strong>taining neur<strong>on</strong>s using<br />

NADPH-diaphorase staining. Acute restraint stress (2h) significantly increased NADPH-d positive neur<strong>on</strong>s<br />

in the paraventricular nucleus (PVN) and LDTg compared with the n<strong>on</strong>-stressed group. NADPH-d positive<br />

cells doubled with choline acetyltransferase (ChAT) were found in the LDTg, not in the PVN. These results<br />

suggest that stress activates NO-producing cells in cholinergic cells in LDTg and these NO-producing<br />

cholinergic cells may play an important role in modulating a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> stress resp<strong>on</strong>ses.<br />

5I_08_P<br />

(poster secti<strong>on</strong> B1, poster board #297, 25 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

NITRIC OXIDE-MEDIATED VASCULAR ADAPTATION<br />

TO CHRONIC SOCIAL STRESS<br />

Angelika Puzserova, Iveta Bernatova<br />

Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Normal and Pathological Physiology, Slovak Academy <str<strong>on</strong>g>of</str<strong>on</strong>g> Sciences, Bratislava, Slovak Republic,<br />

e-mail: angelika.puzserova@savba.sk<br />

The endothelium <str<strong>on</strong>g>of</str<strong>on</strong>g> blood vessels appears to play a central role in the regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> vascular smooth muscle<br />

t<strong>on</strong>e and thus in blood pressure regulati<strong>on</strong> via the synthesis and release <str<strong>on</strong>g>of</str<strong>on</strong>g> vasoactive substances such as nitric<br />

oxide (NO). The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was there<str<strong>on</strong>g>for</str<strong>on</strong>g>e to investigate the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic social stress produced by<br />

crowding <strong>on</strong> blood pressure and vascular functi<strong>on</strong> in normotensive Wistar-Kyoto rats. Adult, male rats were<br />

divided into c<strong>on</strong>trol (480 cm 2 /rat) or stressed (200 cm 2 /rat) group <str<strong>on</strong>g>for</str<strong>on</strong>g> 8 weeks. Blood pressure and heart rate<br />

were determined using tail-cuff plethysmography and their were not influenced by chr<strong>on</strong>ic stress (stress vs.<br />

c<strong>on</strong>trol: 106±2 mm Hg vs. 107±3 mm Hg, 351±8 bpm vs. 354±8 bpm). Total NO synthase activity in the<br />

aorta (determined by [ 3 H]-L-arginine c<strong>on</strong>versi<strong>on</strong>) was elevated in crowded rats (4.7±0.5 pmol/min/mg vs.<br />

2.6±0.2 pmol/min/mg, p


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

vasorelaxati<strong>on</strong> was increased in stressed rats vs. c<strong>on</strong>trols (32.8±4% vs. 18.2±3%, p


23-26 August 2007,<br />

Budapest, Hungary<br />

28<br />

6F. SEPSIS, ENDOTOXINS, STRESS AND ORGAN DYSFUNCTION<br />

(ROBERT S. MUNFORD, JEAN-MARC CAVAILLON)<br />

6F_10_P<br />

(poster secti<strong>on</strong> B2, poster board #292, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

DECREASE OF ADAMTS13 ACTIVITY IS A COMMON FEATURE OF THE<br />

INFLAMMATORY RESPONSE<br />

C. L. Bockmeyer 1 , W. Lösche 1 , U. Budde 2 , M. Bauer 1 , K. Reinhart 1 , F. M. Brunkhorst 1 , R. A. Claus 1<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Anaesthesiology and Intensive Care Medicine, FSU-Jena, University Hospital, Jena;<br />

2Lab-Associati<strong>on</strong> Pr<str<strong>on</strong>g>of</str<strong>on</strong>g>. Arndt & Partners, Hamburg; Germany. e-mail: Ralf.claus@med.uni-jena.de<br />

ADAMTS13 is a protease that specifically acts <strong>on</strong> the multimeric pattern <str<strong>on</strong>g>of</str<strong>on</strong>g> v<strong>on</strong> Willebrand factor VWF.<br />

C<strong>on</strong>genital deficiency <str<strong>on</strong>g>of</str<strong>on</strong>g> ADAMTS13 results in the appearance <str<strong>on</strong>g>of</str<strong>on</strong>g> ultralarge VWF multimers with activati<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> blood platelets, thrombus <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> and (multi) organ failure. There is some evidence that systemic<br />

inflammati<strong>on</strong> results in restricti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> microcirculati<strong>on</strong> and that the development <str<strong>on</strong>g>of</str<strong>on</strong>g> multiple organ<br />

dysfuncti<strong>on</strong> syndrome (MODS) is associated with decreased ADAMTS13 activity and increased VWF levels.<br />

In patients with various degrees <str<strong>on</strong>g>of</str<strong>on</strong>g> systemic inflammati<strong>on</strong> we aimed to characterize a crucial role <str<strong>on</strong>g>of</str<strong>on</strong>g> an<br />

imbalance between ADAMTS13 activity and VWF level in the development <str<strong>on</strong>g>of</str<strong>on</strong>g> inflammati<strong>on</strong> induced<br />

MODS. Four groups <str<strong>on</strong>g>of</str<strong>on</strong>g> patients were studied: healthy volunteers with moderate SIRS after physical exercise,<br />

heart surgery patients with low risk <str<strong>on</strong>g>for</str<strong>on</strong>g> systemic inflammati<strong>on</strong>/MODS, heart surgery patients with<br />

SIRS/moderate MODS, and patients with severe sepsis/septic shock. C<strong>on</strong>secutive samples were analysed <str<strong>on</strong>g>for</str<strong>on</strong>g><br />

ADAMTS13 and compared with markers <str<strong>on</strong>g>of</str<strong>on</strong>g> systemic inflammati<strong>on</strong>/MODS and activated coagulati<strong>on</strong>.<br />

Decrease in ADAMTS13 activity was related to extend <str<strong>on</strong>g>of</str<strong>on</strong>g> systemic inflammati<strong>on</strong> and pr<strong>on</strong>ounced in patients<br />

with MODS, specifically in patients who died in septic shock. Exemplarily we dem<strong>on</strong>strate an associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

low ADAMTS13 activity/ MOF with the appearance <str<strong>on</strong>g>of</str<strong>on</strong>g> ULVWF multimers. Some <str<strong>on</strong>g>of</str<strong>on</strong>g> the patients with low<br />

ADAMTS13 activity and high VWF revealed evidence <str<strong>on</strong>g>of</str<strong>on</strong>g> DIC and changes in platelet count. Systemic<br />

inflammati<strong>on</strong> may cause an imbalance between ADAMTS13 activity and VWF level, and that this imbalance<br />

may c<strong>on</strong>tribute to inflammati<strong>on</strong>-mediated MODS.<br />

6F_11_P<br />

(poster secti<strong>on</strong> B2, poster board #293, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

EXTRACELLULAR HYDROLYSIS OF SPHINGOMYELIN IN PATIENTS WITH<br />

SYSTEMIC INFLAMMATION AND ORGAN FAILURE<br />

A. C. Bunck 1 , D. Hupe 1 , M. Dorer 1 , F. M. Brunkhorst 1 , H. P. Deigner 2 , R. A. Claus 1<br />

1Department <str<strong>on</strong>g>for</str<strong>on</strong>g> Anesthesiology and Intensive Care Therapy, University Hospital Jena;<br />

2SIRS-Lab GmbH, Winzerlaer Straße 2a, Jena, Germany; e-mail: ralf.claus@med.uni-jena.de<br />

Background and Objectives: Recent data indicate that bioactive lipids such as ceramide (Cer) have significant<br />

effects <strong>on</strong> cells relevant <str<strong>on</strong>g>for</str<strong>on</strong>g> inflammati<strong>on</strong>, which may there<str<strong>on</strong>g>for</str<strong>on</strong>g>e be regarded as candidate mediators in<br />

systemic inflammatory resp<strong>on</strong>se syndrome (SIRS). In order to elucidate the role <str<strong>on</strong>g>of</str<strong>on</strong>g> ceramide <str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> in the<br />

development and progressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> systemic sequelae <str<strong>on</strong>g>of</str<strong>on</strong>g> SIRS, we addressed the questi<strong>on</strong> whether there is a<br />

difference in the sphingolytic activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the secreted is<str<strong>on</strong>g>of</str<strong>on</strong>g>orm <str<strong>on</strong>g>of</str<strong>on</strong>g> sphingomyelinases (pSMPD1) in plasma <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

pts with various degrees <str<strong>on</strong>g>of</str<strong>on</strong>g> SIRS or sepsis <str<strong>on</strong>g>of</str<strong>on</strong>g> different origin.


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Methods Plasma samples were obtained from pts with SIRS 1) after <str<strong>on</strong>g>of</str<strong>on</strong>g>f pump cor<strong>on</strong>ary bypass surgery, 2) after<br />

extracorporeal circuit, 3) from pts with sepsis. Activity was determined by the hydrolysis <str<strong>on</strong>g>of</str<strong>on</strong>g> fluorescently<br />

labelled sphingomyelin. In endothelial cells exposed to patient plasma, we determined the rate <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

sphingomyelin hydrolysis and the resulting Cer pattern by chromatographical separati<strong>on</strong> as well as the<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> Cer enriched macro domains by fluorescence microscopy.<br />

Results Plasma activity <str<strong>on</strong>g>of</str<strong>on</strong>g> pSMPD1 in critically ill patients was significantly increased. In pts with fatal<br />

outcome sphingolytic activity increased, while a decrease in sepsis survivors was observed. Low and high<br />

pSMPD1 activity levels were paralleled by equally low or high values <str<strong>on</strong>g>of</str<strong>on</strong>g> established clinical severity markers.<br />

Comparing the absolute increase <str<strong>on</strong>g>of</str<strong>on</strong>g> pSMPD1 activity 24 hours after surgery we found the increase <str<strong>on</strong>g>of</str<strong>on</strong>g> enzyme<br />

activity to be significantly lower in both the MIDCAB group as well as in patients without SIRS at the first<br />

day post op. We found an increase in breakdown <str<strong>on</strong>g>of</str<strong>on</strong>g> sphingomyelin in endothelial cells after stimulati<strong>on</strong> with<br />

pts plasma as well as endotoxin or prototypic pro-inflammatory cytokines such as TNF-alpha. Also we found<br />

<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ceramide enriched macro domains by immuno-staining using specific antibodies directed against<br />

Cer, CD14, Fas-receptor and TNF-Receptor1.<br />

C<strong>on</strong>clusi<strong>on</strong>s pSMPD1 activity is enhanced in SIRS and sepsis. Together with data from in vitro experiments and<br />

animal models, bio-functi<strong>on</strong>ally relevant role <str<strong>on</strong>g>of</str<strong>on</strong>g> pSMPD1 resulting in an altered signal transducti<strong>on</strong> in<br />

systemic inflammati<strong>on</strong> is c<strong>on</strong>cluded. The associati<strong>on</strong> with mortality suggests that increased pSMPD1 activity<br />

may be involved in the complex network <str<strong>on</strong>g>of</str<strong>on</strong>g> processes finally resulting in an unfavourable outcome.<br />

6M. OTHER STRESS TOPICS IN MEDICINE (GENDER DIFFERENCES IN<br />

PATHOPHYSIOLOGICAL STRESS, HSPS AND WOUND HEALING, STRESS AND<br />

MYOCARDIAL PROTECTION, PRE- AND POSTOPERATIVE STRESS)<br />

6M_07_P<br />

(poster secti<strong>on</strong> B1, poster board #225, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CARDIAC FUNCTION ALTERATION IN CTNI TRANSGENIC MICE AFTER<br />

VARIOUS STRESS STIMULATIONS<br />

J. Du, N. Gobara, C. Nan, P. Jean-Charles, XP. Huang<br />

1Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Biomedical Science and Center <str<strong>on</strong>g>for</str<strong>on</strong>g> Molecular Biology and Biotechnology, Florida Atlantic University,<br />

Boca Rat<strong>on</strong>, FL 33431, USA<br />

Cardiac trop<strong>on</strong>in I (cTnI) plays a critic role in regulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cardiac my<str<strong>on</strong>g>of</str<strong>on</strong>g>ibril c<strong>on</strong>tracti<strong>on</strong> and relaxati<strong>on</strong>. A<br />

linkage study has dem<strong>on</strong>strated that idiopathic restrictive cardiomyopathy (RCM) is part <str<strong>on</strong>g>of</str<strong>on</strong>g> the clinical<br />

expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> cTnI mutati<strong>on</strong>. We modeled <strong>on</strong>e mutati<strong>on</strong> in human cTnI C-terminus, R192H and created<br />

cTnI 193His transgenic mice (Du et al, 2006). The phenotype <str<strong>on</strong>g>of</str<strong>on</strong>g> cTnI 193His transgenic mice is an impaired cardiac<br />

relaxati<strong>on</strong> and progressively development <str<strong>on</strong>g>of</str<strong>on</strong>g> heart failure. In the present study, we have m<strong>on</strong>itored cardiac<br />

functi<strong>on</strong> in these mice after various stress stimulati<strong>on</strong>s. The stress stimulati<strong>on</strong>s include exercise (swimming<br />

assays), hypoxia stimulati<strong>on</strong> and dobutamine stimulati<strong>on</strong> assays. The results have dem<strong>on</strong>strated that the<br />

swimming durati<strong>on</strong> was significantly shortened in transgenic mice compared to their wild type littermates<br />

(1.4±0.3 hours in transgenic mice vs. 2.1±0.5 hours in wild type mice, P


23-26 August 2007,<br />

Budapest, Hungary<br />

in cTnI 193His mice. Our results dem<strong>on</strong>strate that cTnI 193His mice are more vulnerable to various stress<br />

stimulati<strong>on</strong>. The failure <str<strong>on</strong>g>of</str<strong>on</strong>g> dobutamine stimulati<strong>on</strong> indicates that the increase <str<strong>on</strong>g>of</str<strong>on</strong>g> the endogenous my<str<strong>on</strong>g>of</str<strong>on</strong>g>ibril<br />

tensi<strong>on</strong> is probably the main cause <str<strong>on</strong>g>for</str<strong>on</strong>g> the stiff heart and restrictive cardiomyopathy in cTnI 193His mice.<br />

7A. STRESS, COGNITIVE FUNCTION AND BEHAVIOR<br />

(JÓZSEF HALLER, JEANSOK KIM)<br />

7A_26_P<br />

(poster secti<strong>on</strong> A2, poster board #176, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SOCIAL ISOLATION DURING ADOLESCENCE PRODUCES INCREASED<br />

SENSITIVITY TO STRESS IN ADULT FEMALES<br />

Seema Bhatnagar, Ari Weintraub, Vikram Iyer<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Pennsylvania, Philadelphia, Pennsylvania, USA<br />

Adolescence is a period <str<strong>on</strong>g>of</str<strong>on</strong>g> flux and transiti<strong>on</strong> encompassing reproductive, cognitive, emoti<strong>on</strong>al and social<br />

maturati<strong>on</strong>. How an organism negotiates the challenges and stressors <str<strong>on</strong>g>of</str<strong>on</strong>g> adolescence may determine its future<br />

risk or resilience <str<strong>on</strong>g>for</str<strong>on</strong>g> pathology in later life. Interestingly, adolescence is a time when some types <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

psychopathology first emerge and when sex differences in rats <str<strong>on</strong>g>of</str<strong>on</strong>g> psychopathology and stress resp<strong>on</strong>visity<br />

also emerge. In the present studies, we hypothesized that social isolati<strong>on</strong> during adolescence would increase<br />

sensitivity to stress and anxiety-like behavior in a sex-specific manner in adult rats. Sprague-Dawley rats were<br />

purchased from Charles River (Wilmingt<strong>on</strong>, MA) at day 21 <str<strong>on</strong>g>of</str<strong>on</strong>g> age (n=20 males and n=25 females). At day 30<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> age, rats were separated into two groups, c<strong>on</strong>trol or social isolati<strong>on</strong>. C<strong>on</strong>trol rats were housed in same-sex<br />

groups <str<strong>on</strong>g>of</str<strong>on</strong>g> 3 per cage. Socially isolated rats were housed al<strong>on</strong>e until age day 50. Isolated male rats gained more<br />

weight during the period <str<strong>on</strong>g>of</str<strong>on</strong>g> isolati<strong>on</strong> than c<strong>on</strong>trol rats. On day 50, all rats were exposed to acute restraint and<br />

blood sampled <str<strong>on</strong>g>for</str<strong>on</strong>g> ACTH and corticoster<strong>on</strong>e c<strong>on</strong>centrati<strong>on</strong>s. Isolated female rats exhibited delayed recovery<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> ACTH and corticoster<strong>on</strong>e from restraint whereas isolated males exhibited enhanced recovery compared to<br />

their respective c<strong>on</strong>trols. At this time, isolated rats were put in same-sex groups <str<strong>on</strong>g>of</str<strong>on</strong>g> 2 rats per cage and c<strong>on</strong>trol<br />

rats were also re-housed 2/cage. All animals were re-exposed to acute restraint in adulthood, at approximately<br />

day 70 <str<strong>on</strong>g>of</str<strong>on</strong>g> age. Isolated females exhibited enhanced peak corticoster<strong>on</strong>e levels and delayed recovery from<br />

restraint compared to c<strong>on</strong>trol females. In c<strong>on</strong>trast, isolated males exhibited lower peak resp<strong>on</strong>ses to restraint.<br />

Animals were stressed by restraint <str<strong>on</strong>g>for</str<strong>on</strong>g> 6 c<strong>on</strong>secutive days and then tested in the elevated plus maze. Isolated<br />

male rats exhibited indices <str<strong>on</strong>g>of</str<strong>on</strong>g> decreased anxiety-like behavior compared to c<strong>on</strong>trol males. Finally, isolated<br />

females gained less weight over the repeated restraint period and had larger adrenal glands compared to<br />

c<strong>on</strong>trol females. Overall, clear sex differences in the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> adolescent isolati<strong>on</strong> were observed with<br />

female isolated rats exhibiting hyperrep<strong>on</strong>siveness to stress at the end <str<strong>on</strong>g>of</str<strong>on</strong>g> isolati<strong>on</strong> and in adulthood. Whether<br />

g<strong>on</strong>adal horm<strong>on</strong>es and/or changes in stress-related neural circuitry underlie these sex-specific effects <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

adolescent isolati<strong>on</strong> remains to be seen.<br />

30


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7A_27_P<br />

(poster secti<strong>on</strong> A1, poster board #31, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

MUSIC IMPROVES SLEEP QUALITY IN STUDENTS<br />

László Harmat, Johanna Takács, Róbert Bódizs<br />

Semmelweis University, Institite <str<strong>on</strong>g>of</str<strong>on</strong>g> Behavioural Sciences, H-1089 Budapest, Nagyvárad tér 4., Hungary<br />

e-mail: laszloharmat@yahoo.com<br />

Objectives: Investigati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> relaxing classical music <strong>on</strong> students’ sleep quality. Background:<br />

Studies have focused <strong>on</strong> the effects <str<strong>on</strong>g>of</str<strong>on</strong>g> music <strong>on</strong> sleep quality in a variety <str<strong>on</strong>g>of</str<strong>on</strong>g> settings and populati<strong>on</strong>s. Music is<br />

a potential n<strong>on</strong>pharmacological treatment reducing sleep complaints. Methods: We used a three group<br />

repeated measures design. 94 students (ages 19-28) with sleep complaints - Pittsburg Sleep Quality Index<br />

(PSQI) > 5 - were enrolled during 2006. Participants listened to 45 minutes <str<strong>on</strong>g>of</str<strong>on</strong>g> either relaxing classical music<br />

(Group 1) or audiobook (Group 2) at bedtime <str<strong>on</strong>g>for</str<strong>on</strong>g> 3 weeks. In the c<strong>on</strong>trol group (Group 3) there was no<br />

interventi<strong>on</strong>. Sleep quality was measured by PSQI be<str<strong>on</strong>g>for</str<strong>on</strong>g>e the study as well as weekly during the interventi<strong>on</strong>.<br />

We assessed the depressive symptoms <str<strong>on</strong>g>of</str<strong>on</strong>g> the participants in the experimental groups with BDI be<str<strong>on</strong>g>for</str<strong>on</strong>g>e and<br />

after the study. Results: Repeated measures ANOVA revealed a main effect <str<strong>on</strong>g>of</str<strong>on</strong>g> GROUPS (P=0·0028) and<br />

TIME (P


23-26 August 2007,<br />

Budapest, Hungary<br />

In our examinati<strong>on</strong> we applied the below menti<strong>on</strong> tests: Perceived Stress (Cohen), Depressi<strong>on</strong> (Beck),<br />

Somatisati<strong>on</strong> (PHQ-15) (Kroenke) and Burnout (Maslach) and Burn-out and Engagement (student versi<strong>on</strong>)<br />

(Schaufeli), Emphaty (Jeffers<strong>on</strong>), Coping (Folkman-Lazarus), Parental B<strong>on</strong>ding (Parker), Career-Motivati<strong>on</strong><br />

(Molnár) (Super)<br />

Our results: In our survey we have found, that the Perceived Stress and the Burn-out corralates, with the<br />

increaing numbers <str<strong>on</strong>g>of</str<strong>on</strong>g> the Emoti<strong>on</strong>al Exhausti<strong>on</strong> and the Depers<strong>on</strong>alisati<strong>on</strong> /Cinism and the decreasing<br />

numbers <str<strong>on</strong>g>of</str<strong>on</strong>g> the Pr<str<strong>on</strong>g>of</str<strong>on</strong>g>essi<strong>on</strong>al /Pers<strong>on</strong>al Efficacy. Also correlates with the Engagement, with all factors, as<br />

Energy, Dedicati<strong>on</strong> and Absorpti<strong>on</strong>.<br />

Perceived Stress also correlates with the Somatisati<strong>on</strong> and Depressi<strong>on</strong> Am<strong>on</strong>g the coping mechanism<br />

correlates with mostly the emoti<strong>on</strong>al-centered methods. Parental overprotecti<strong>on</strong> also influences the perceived<br />

stress, especially from the part <str<strong>on</strong>g>of</str<strong>on</strong>g> the father.<br />

The perceived stress relates with the career-motivati<strong>on</strong>, mostly with the helper attitude and with the<br />

preference <str<strong>on</strong>g>of</str<strong>on</strong>g> aut<strong>on</strong>omy.<br />

In our survey the above menti<strong>on</strong>ed examinated groups does not differ from each other int he field <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Perceived Stress, but while the technical university students showed more burn-out, the medical students<br />

seemed to be more emphatic and vulnerable <str<strong>on</strong>g>for</str<strong>on</strong>g> somatisati<strong>on</strong>.<br />

7K. STRESS AND GENDER<br />

7K_06_P<br />

(poster secti<strong>on</strong> A2, poster board #321, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

WELL-BEING IN THE LATE PERIOD OF PREGNANCY AND MATERNITY BLUES<br />

FOLLOWING CHILDBIRTH IN JAPANESE WOMEN<br />

Shigeko Tsuda 1 , Akira Tsuda 2 , Yoshiyuki Tanaka 3<br />

1Fukuoka Jo Gakuin, Fukuoka, 2 Kurume University, Kurume, 3 Tokyo University <str<strong>on</strong>g>of</str<strong>on</strong>g> Social Welfare, Isesaki,<br />

Japan, e-mail: t-add02@kumin.ne.jp<br />

This study is to examine any relati<strong>on</strong>ships between subjective well-being in the late period <str<strong>on</strong>g>of</str<strong>on</strong>g> pregnancy and<br />

maternity blues symptoms following childbirth, as well as effects <str<strong>on</strong>g>of</str<strong>on</strong>g> obstetric factors. Subjects were 79<br />

pregnant women, aged from 19 to 41, who c<strong>on</strong>sulted K University Hospital obstetric outpatients department<br />

after 36 weeks <str<strong>on</strong>g>of</str<strong>on</strong>g> pregnancy. Well-being and maternity blues symptoms were assessed by the WHO<br />

Subjective Well-being Inventory (SUBI) and Stein’s maternity blues scale, respectively. The maternity blues<br />

symptoms were associated with lower levels <str<strong>on</strong>g>of</str<strong>on</strong>g> well-being in the late period <str<strong>on</strong>g>of</str<strong>on</strong>g> pregnancy, as well as with<br />

obstetric factors <str<strong>on</strong>g>of</str<strong>on</strong>g> age, complicati<strong>on</strong>s and newborn infant status. These findings suggest that perceived<br />

psychological health, thus, well-being, appears to be a reliable c<strong>on</strong>tributing factor to the differential<br />

susceptibility to maternity blues symptoms in the early postpartum period.<br />

32


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

7K_07_P<br />

(poster secti<strong>on</strong> A2, poster board #322, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PSYCHOSOCIAL STRESS, OBESITY, PREDIABETES AND INSULIN RESISTANCE<br />

IN CHILDREN AND ADOLESCENTS<br />

J. M. Malacara 1 , M. E. Garay-Sevilla 1 , H. Aguilar-Zavala 1 , E. L. Pérez-Luque,<br />

E. de la Cruz-Mendoza 2 , M. Ibarra- Torres 2 , M. Goldaracena-Azuara 2 , C. Aradillas-García 2<br />

1Universidad de Guanajuato, Instituto de Investigaci<strong>on</strong>es Médicas, 2 Universidad Autónoma de San Luis Potosí,<br />

Facultad de Medicina, Mexico<br />

Previous evidence indicate the influence <str<strong>on</strong>g>of</str<strong>on</strong>g> psychosocial stress with the <strong>on</strong>set <str<strong>on</strong>g>of</str<strong>on</strong>g> obesity and diabetes. It is<br />

important to study that associati<strong>on</strong> in children and adolescents.<br />

Objective: To study the relati<strong>on</strong>ship <str<strong>on</strong>g>of</str<strong>on</strong>g> psychological stress with obesity, insulin resistance and prediabetes in<br />

children and adolescents.<br />

Materia and methods: total <str<strong>on</strong>g>of</str<strong>on</strong>g> 309 subjects 6 to 17 years old were randomly selected from urban schools <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

León and San Luis Potosí (SLP) in Central Mexico. We collected pers<strong>on</strong>al and anthropometric data, blood<br />

pressure and psychological measurements <str<strong>on</strong>g>for</str<strong>on</strong>g>: perceived stress, dominance, submissive behavior and anxiety<br />

and depressi<strong>on</strong> (MTAD). A venous blood sample was drawn after 12 hrs. fasting to measure glucose, totalcholesterol,<br />

LDL-cholesterol, HDL-Cholesterol, triglycerides, insulin, IGF-1 and testoster<strong>on</strong>e. Insulin<br />

resistance (HOMA-IR) was calculated. When available their parents were also studied.<br />

Results: Overwight was found in 22.9%, obesity in 15.0%, insulin resistance in 29.3%, and prediabetes in 7.6%<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> the cases. We found significant associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> perceived stress with age <str<strong>on</strong>g>of</str<strong>on</strong>g> the mother (p


23-26 August 2007,<br />

Budapest, Hungary<br />

8. OTHER<br />

8_06_P<br />

(poster secti<strong>on</strong> A1, poster board #21, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STRESS PROTEIN αB-CRYSTALLIN KEEPING CYTOSKELETON DYNAMICS WITH<br />

TENSION AND PHYSICAL ACTIVITIES AS AN ESSENTIAL FACTOR OF QOL –<br />

A SIGNIFICANCE OF CONSTITUTIVE EXPRESSION IN ACTIVE STRIATED<br />

MUSCLES<br />

Atomi Yoriko, Eri Fujita, Tetsuo Yamaguchi, Takashi Sakurai, Yoshinobu Fujita<br />

N1304 Exp. Res. Building <str<strong>on</strong>g>of</str<strong>on</strong>g> Dep. Med., The Univ. <str<strong>on</strong>g>of</str<strong>on</strong>g> Tokyo, Tokyo, 113-0033, Japan<br />

The cytoskelet<strong>on</strong> dynamically maintains tensi<strong>on</strong> and cell shape resp<strong>on</strong>ding mechanical stress in our body and<br />

aut<strong>on</strong>omously organizes cell activities. Subtle protein c<strong>on</strong><str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong>al change with the assembly/disassembly<br />

dynamics needs molecular chaper<strong>on</strong>e. Recently actin filament disassembly resulting in associati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> another<br />

factor <str<strong>on</strong>g>of</str<strong>on</strong>g> heat sock RNA (HSR) was included into gene expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock proteins (HSPs) by an<br />

activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> heat shock factor (HSF). Small HSPs (sHPSs) have been known to work as chaper<strong>on</strong>e <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

cytoskelet<strong>on</strong> like actin, tubulin/microtubule (MT), and intermediate filament, however the molecular<br />

mechanism has not been elucidated yet. All members <str<strong>on</strong>g>of</str<strong>on</strong>g> sHSPs express more in especially stressful slow and<br />

cardiac striated muscle cells, which c<strong>on</strong>tinues to c<strong>on</strong>tract mainly with oxidative metabolism. This study shows<br />

recent findings in my laboratory and the essential role <str<strong>on</strong>g>of</str<strong>on</strong>g> αB-crystallin keeping mechanical (structural)<br />

homeostasis using GFP-αB-crystallin as well as protein homeostasis in c<strong>on</strong>stitutive active muscles. Decisive<br />

role <str<strong>on</strong>g>of</str<strong>on</strong>g> the C-terminal <strong>on</strong>e-third <str<strong>on</strong>g>of</str<strong>on</strong>g> αB-crystallin (a c<strong>on</strong>served α-crytallin domain) is to inhibit tubulin<br />

aggregati<strong>on</strong> and to maintain “dynamic instability”, as well the N-terminal domain also works keeping dynamic<br />

stabilizati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> MT via MT associating proteins. The decreases <str<strong>on</strong>g>of</str<strong>on</strong>g> both protein and mRNA <str<strong>on</strong>g>of</str<strong>on</strong>g> αB-crystallin<br />

were followed by <strong>on</strong>ly protein tubulin decrease in rat unloaded slow muscle atrophy, showing an important<br />

role <str<strong>on</strong>g>of</str<strong>on</strong>g> chaper<strong>on</strong>e maintaining the protein cytoskelet<strong>on</strong> system. The c<strong>on</strong>centrati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> free tubulin dimer is<br />

autoregulated in the balance <str<strong>on</strong>g>of</str<strong>on</strong>g> free dimer and polymeric <str<strong>on</strong>g>for</str<strong>on</strong>g>ms <str<strong>on</strong>g>of</str<strong>on</strong>g> MT protein, having an intrinsic property <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

“dynamic instability”, and through cotranslati<strong>on</strong>al β-tubulin mRNA degradati<strong>on</strong>. Innactive human muscles<br />

increases phophorylated <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> aB-crystallin as well as the total protein without mRNA levels, indicates that<br />

inactivity cannot induce gene expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> αB-crystallin nor maintain protein homeostasis without good<br />

stress to induce sHSPs and cytoskelet<strong>on</strong> dynamics. This dynamic biological system is based <strong>on</strong> stress<br />

resp<strong>on</strong>se in our active human body.<br />

34


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

8_14_P<br />

(poster secti<strong>on</strong> A1, poster board #22, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

STRESS-INDUCED CHANGE IN GLUTAMATERGIC VESICLE MOBILITY IN<br />

ASTROCYTES<br />

Maja Potokar 1,2 , Matjaž Stenovec 1,2 , Marko Kreft 1,2 , Mateja Erdani Kreft 3 , Robert Zorec 1,2<br />

1Celica Biomedical Center, Proletarska cesta 4, 1000 Ljubljana, Slovenia<br />

2Laboratory <str<strong>on</strong>g>of</str<strong>on</strong>g> Neuroendocrinology-Molecular Cell Physiology, Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Pathophysiology, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine,<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Ljubljana, Zaloška 4, 1000 Ljubljana<br />

3Institute <str<strong>on</strong>g>of</str<strong>on</strong>g> Cell Biology, Faculty <str<strong>on</strong>g>of</str<strong>on</strong>g> Medicine, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Ljubljana, Lipičeva 2, 1000 Ljubljana, Slovenia<br />

In the pathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> neurodegenerative disorders as well as in traumatic and ischemic injuries <str<strong>on</strong>g>of</str<strong>on</strong>g> the central<br />

nervous system (CNS) excitotoxic stress is developed due to highly increased extracellular c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

glutamate. Astrocytes are, in additi<strong>on</strong> to neur<strong>on</strong>s, sensitive to excitotoxic stress, leading to a substantial<br />

increase in the intracellular free calcium c<strong>on</strong>centrati<strong>on</strong> ([Ca 2+ ] i ). A rise in the [Ca 2+ ] i elicits a discharge <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

several neuroactive molecules from membrane-bound vesicles, releasing more glutamate into the extracellular<br />

space. Several aspects <str<strong>on</strong>g>of</str<strong>on</strong>g> the trafficking <str<strong>on</strong>g>of</str<strong>on</strong>g> glutamate c<strong>on</strong>taining vesicles have been studied, but the recycling<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> these vesicles is poorly defined. We exposed astrocytes to antibodies against the vesicular glutamate<br />

transporter 1 (VGLUT1) to fluorescently label vesicles undergoing Ca 2+ -dependent exocytosis and examined<br />

their number, fluorescence intensity and mobility by c<strong>on</strong>focal microscopy. We show that in n<strong>on</strong>-stimulated<br />

cells VGLUT1 vesicles cycle slowly between the plasma membrane and the cytoplasm. A rise in [Ca 2+ ] i<br />

elicited an increase in the number and fluorescence intensity <str<strong>on</strong>g>of</str<strong>on</strong>g> the puncta, likely because the VGLUT1<br />

epitopes were more accessible to the extracellularly applied antibodies. In c<strong>on</strong>trast to n<strong>on</strong>-stimulated cells, in<br />

stimulated cells many vesicles exhibited higher, directi<strong>on</strong>al mobility. In CNS pathologies astrocytes change<br />

the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> many genes, including genes encoding intermediate filament proteins. Since cytoskelet<strong>on</strong>severing<br />

agents abolished vesicle mobility, this indicates a dependence <str<strong>on</strong>g>of</str<strong>on</strong>g> glutamatergic vesicle recycling <strong>on</strong><br />

the cytoskelet<strong>on</strong>. Our findings importantly c<strong>on</strong>tribute to the understanding how this mobility is regulated.<br />

8_15_P<br />

(poster secti<strong>on</strong> A1, poster board #23, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ADRENAL REGULATION FOR CIRCADIAN GLUCOCORTICOID SYNTHESIS BY<br />

CHRONIC AND ACUTE EXERCISE, AND POSSIBLE ROLE OF<br />

EXERCISE-INDUCED HSP70<br />

Atomi Yoriko, Mayumi Ohtawa, Hideaki Arai<br />

N1304 Exp. Res. Building <str<strong>on</strong>g>of</str<strong>on</strong>g> Dep. Med., The Univ. <str<strong>on</strong>g>of</str<strong>on</strong>g> Tokyo, Tokyo, 113-0033, Japan<br />

Mild physical exercise stress is essential <str<strong>on</strong>g>for</str<strong>on</strong>g> keeping homeostasis <str<strong>on</strong>g>of</str<strong>on</strong>g> our whole body system via HPA axis. The<br />

adrenal cortex produces glucocortidoid as end-product regulator <str<strong>on</strong>g>of</str<strong>on</strong>g> HPA axis. We studied the adaptati<strong>on</strong><br />

mechanism <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic exercise via HPA axis and a possible relati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> HSP70 in glucocorticoid synthesis in<br />

adrenal cortex. Two weeks <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic voluntary wheel running <str<strong>on</strong>g>of</str<strong>on</strong>g> mice c<strong>on</strong>firmed the significant increase <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the circadian peak <str<strong>on</strong>g>of</str<strong>on</strong>g> plasma corticoster<strong>on</strong>e without alterati<strong>on</strong> in adrenocorticotropic horm<strong>on</strong>e (ACTH) level,<br />

as previously reported be<str<strong>on</strong>g>for</str<strong>on</strong>g>e. To elucidate the mechanisms <str<strong>on</strong>g>of</str<strong>on</strong>g> exercise modulati<strong>on</strong> <strong>on</strong> corticoster<strong>on</strong>e<br />

synthesis, we first examined the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> transcripts involved in corticoster<strong>on</strong>e synthesis <str<strong>on</strong>g>of</str<strong>on</strong>g> the adrenal gland.<br />

Am<strong>on</strong>g them, <strong>on</strong>ly steroidogenic acute regulatory protein (StAR), the rate-limiting factor that transfers<br />

35


23-26 August 2007,<br />

Budapest, Hungary<br />

substrate cholesterol into inner mitoch<strong>on</strong>drial membrane, showed significantly higher expressi<strong>on</strong> in the<br />

exercise group. Since the splanchnic nerve input to the adrenal gland has been reported as a factor involved in<br />

the direct modulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> corticoster<strong>on</strong>e synthesis, we next examined the expressi<strong>on</strong> levels <str<strong>on</strong>g>of</str<strong>on</strong>g> enzymes <str<strong>on</strong>g>for</str<strong>on</strong>g> the<br />

catecholamine synthesis as indices <str<strong>on</strong>g>of</str<strong>on</strong>g> sympatho-adrenomedullary activity. We found that the <strong>on</strong>ly rate-limiting<br />

enzyme, tyrosine hydroxylase (TH), was significantly higher in the adrenals <str<strong>on</strong>g>of</str<strong>on</strong>g> exercise group. In additi<strong>on</strong> to<br />

the increment <str<strong>on</strong>g>of</str<strong>on</strong>g> StAR and TH mRNAs in resp<strong>on</strong>se to the chr<strong>on</strong>ic exercise, we found <strong>on</strong>ly these factors<br />

showed the circadian variati<strong>on</strong> in those expressi<strong>on</strong> levels that was correlated to the circadian rhythm <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

corticoster<strong>on</strong>e. Chr<strong>on</strong>ic exercise seems to alter the circadian corticoster<strong>on</strong>e synthesis, at least partially via<br />

altering the levels <str<strong>on</strong>g>of</str<strong>on</strong>g> circadian-regulated transcripts, StAR and TH <str<strong>on</strong>g>of</str<strong>on</strong>g> the adrenal gland. Finally,<br />

immunoprecipitati<strong>on</strong> experiments using adrenal homogenates and these antibodies to find causal relati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

HSP70 in adrenal cortex and StAR showed the associati<strong>on</strong> each other. This study shows molecular aspects <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

stress resp<strong>on</strong>ses including molecular chaper<strong>on</strong>e and key enzyme’s synthesis cascade, which c<strong>on</strong>nects intraand<br />

inter-cellular and tissue regulati<strong>on</strong>s during adaptati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> whole body stress such as endurance exercise.<br />

8_16_P<br />

(poster secti<strong>on</strong> A1, poster board #24, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

CONNECTION BETWEEN ENDOPLASMIC RETICULUM GLUCOSE METABOLISM<br />

AND OXIDATIVE STRESS RESISTANCE:<br />

THE ROLE OF PYRIDINE NUCLEOTIDES<br />

Péter Száraz 1 , Gábor Bánhegyi 2 , Angelo Benedetti 1<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Pathophysiology, Experimental Medicine and Public Health, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Siena, 53100 Siena,<br />

Italy 1 , Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Medical Chemistry, Pathobiochemistry and Molecular Biology, Semmelweis University, 1444<br />

Budapest, Hungary 2 . E-mail: benedetti@unisi.it<br />

The endoplasmic reticulum is a highly multifuncti<strong>on</strong>al compartment <str<strong>on</strong>g>of</str<strong>on</strong>g> the eukaryotic cell. Besides its well<br />

known roles – like synthesis and maturati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> extracellularly exposed proteins, calcium homeostasis and<br />

signalling, glucose metabolism etc. – it has a crucial role providing a buffer capacity against oxidative cellular<br />

stress. Our recent studies suggest that this functi<strong>on</strong> is not exclusively related to the well characterized<br />

thiol/disulphide system, but is also tightly in c<strong>on</strong>necti<strong>on</strong> with the NADP + /NADPH pyridine nucleotide<br />

redox couple. According to our examinati<strong>on</strong>s <strong>on</strong> cell cultures either chemical inhibiti<strong>on</strong> or RNA silencing<br />

treatments against the key proteins <str<strong>on</strong>g>of</str<strong>on</strong>g> the glucose metabolism related intraluminal NADPH producti<strong>on</strong> in the<br />

endoplasmic reticulum, namely glucose-6-phosphate transporter and hexose-6-phosphate dehydrogenase, as<br />

well as the direct depleti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the luminal NADPH pool causes an enhanced sensitivity against oxidative<br />

stress.<br />

36


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

8_17_P<br />

(poster secti<strong>on</strong> A1, poster board #25, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

SILYMARIN INCREASES THE RENAL ACTIVITY OF ANTIOXIDANT ENZYMES IN<br />

EXPERIMENTAL DIABETES MELLITUS<br />

C. Soto 1* , E. Uría 2 , J. Pérez 1 , C. Alvarez 1 , V. García 1<br />

1Departamento de Sistemas Biológicos, Universidad Autónoma Metropolitana-Xochimilco. 2 Departamento de<br />

Morfología, Escuela Naci<strong>on</strong>al de Ciencias Biológicas, I.P.N. Calz. Del Hueso 1100, Col. Villa Quietud, Coyoacán,<br />

C.P. 04960, México, D.F. México. *e-mail: casoto@correo.xoc.uam.mx<br />

Oxidative stress plays a relevant role in the pathogenesis <str<strong>on</strong>g>of</str<strong>on</strong>g> diabetes mellitus and its sequels as nephropathy.<br />

Many studies have dem<strong>on</strong>strated that the kidneys are especially pr<strong>on</strong>e to the acti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> free radicals which<br />

would suggest that may c<strong>on</strong>tribute to the renal damage. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to analyze the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> the<br />

flav<strong>on</strong>oid silymarin a free radical scavenger that prevents lipoperoxidati<strong>on</strong> in the renal activity <str<strong>on</strong>g>of</str<strong>on</strong>g> superoxide<br />

dismutase, glutathi<strong>on</strong>e peroxidase and catalase in rats with alloxan-induced diabetes mellitus. Alloxan<br />

intoxicated rats were treated with silymarin in two manners, simultaneously (four or eight doses) or 20 days<br />

after alloxan administrati<strong>on</strong> <str<strong>on</strong>g>for</str<strong>on</strong>g> nine weeks. Alloxan elicited a transient increase in the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the three<br />

enzymes which decreased after five days <str<strong>on</strong>g>of</str<strong>on</strong>g> treatment. On its own, silymarin significantly increased the<br />

activity <str<strong>on</strong>g>of</str<strong>on</strong>g> these enzymes. Simultaneous treatment with alloxan and silymarin also induced an increment in the<br />

activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the enzymes followed by a delayed decrease (four doses). However a l<strong>on</strong>ger treatment with<br />

silymarin (eight doses) induced a more sustained effect. Interestingly silymarin treatment recovered to c<strong>on</strong>trol<br />

values the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> the three antioxidant enzymes that were significantly diminished after twenty days <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

alloxan administrati<strong>on</strong>. It is suggested that the protective effect <str<strong>on</strong>g>of</str<strong>on</strong>g> silymarin <strong>on</strong> the pancreatic damage<br />

induced by alloxan may be due to an increase in the activity <str<strong>on</strong>g>of</str<strong>on</strong>g> antioxidant enzymes which in additi<strong>on</strong> to<br />

glutathi<strong>on</strong>e system c<strong>on</strong>stitute the more important defense mechanisms against free radicals damage.<br />

8_18_P<br />

(poster secti<strong>on</strong> A1, poster board #26, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PSYCHO-SOCIAL FACTORS INFLUENECE STROKE OUTCOME<br />

Géza Szabó, László Molnár, Csaba Óvary, Zoltán Nagy<br />

Nati<strong>on</strong>al Stroke <str<strong>on</strong>g>Centre</str<strong>on</strong>g>, Budapest, Hungary<br />

Background. Factors influencing stroke outcome are subject <str<strong>on</strong>g>of</str<strong>on</strong>g> interest. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> present study was to<br />

compare the impact <str<strong>on</strong>g>of</str<strong>on</strong>g> “traditi<strong>on</strong>al” risks factors and psychosocial factors <strong>on</strong> modified Rankin scale.<br />

Methods. In the stroke unite <str<strong>on</strong>g>of</str<strong>on</strong>g> a county hospital, 646 c<strong>on</strong>secutive stroke patients were include in this study.<br />

Determining disease outcome we used the modified Rankin scale after 28 days <str<strong>on</strong>g>of</str<strong>on</strong>g> stroke <strong>on</strong>set. Stroke risk<br />

factors were recorded and psychosocial data (Beck depressi<strong>on</strong> scale, coping skills, educati<strong>on</strong>al attainment, and<br />

isolati<strong>on</strong>) were collected using the standardized questi<strong>on</strong>naires. For statistics we used a multiway c<strong>on</strong>tingency<br />

tables with log linear modelling.<br />

Results. A log linear model fits well (p=0.786) to our data. Patients suffering <str<strong>on</strong>g>for</str<strong>on</strong>g>m repeated stroke showed<br />

significantly worse outcome (p


23-26 August 2007,<br />

Budapest, Hungary<br />

The 2-factors interacti<strong>on</strong> between educati<strong>on</strong>al attainment and isolati<strong>on</strong>, and between educati<strong>on</strong>al attainment<br />

and depressi<strong>on</strong> could be also proved.<br />

Discussi<strong>on</strong>. The significant effects <str<strong>on</strong>g>of</str<strong>on</strong>g> depressi<strong>on</strong>, insufficient coping skills, low educati<strong>on</strong>al level, and<br />

isolati<strong>on</strong> <strong>on</strong> stroke outcome were documented. Better managing <str<strong>on</strong>g>of</str<strong>on</strong>g> these factors may improve the quality <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

life after stroke.<br />

8_19_P<br />

(poster secti<strong>on</strong> A1, poster board #27, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

PSYCHOTRAUMATIC STRESSORS AS TRIGGER OF CHRONIC INSOMNIA<br />

Elena Rasskazova<br />

Moscow Municipal University <str<strong>on</strong>g>of</str<strong>on</strong>g> Psychology and Pedagogics, Russia<br />

The research supported by Russian Fund <str<strong>on</strong>g>for</str<strong>on</strong>g> Humanitarian Sciences project 07-06-00407а<br />

Background. There are close relati<strong>on</strong>ships between psychosocial stress and insomnia proved in different investigati<strong>on</strong>s. From <strong>on</strong>e hand,<br />

psychosocial stress is a comm<strong>on</strong> factor precipitating and perpetuating insomnia. Lots <str<strong>on</strong>g>of</str<strong>on</strong>g> empirical researches including prospective showed that<br />

stress experience triggers subjective insomnia complaints and objective symptoms (i.e. Vein, Kovrov, 2004, Kaeppler et al., 2003, Lint<strong>on</strong>, 2004).<br />

Insomnia complaints are usual symptom in posttraumatic stress disorder (Principles…, 2005, Horowitz, 1979, Allaert et al, 2004). When asking<br />

about possible reas<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> their insomnia about 80% patients usually call stress and str<strong>on</strong>g emoti<strong>on</strong>s (Tkhostov et al, in print). From the other<br />

hand, insomnia symptoms itself <str<strong>on</strong>g>of</str<strong>on</strong>g>ten lead to stress which allowed researchers to describe an insomnia as a model <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic stress (Vein,<br />

Kovrov, 2004).<br />

However, most <str<strong>on</strong>g>of</str<strong>on</strong>g> the studies didn’t distinguish between serious life events and everyday stressful situati<strong>on</strong>s which do have different impact <strong>on</strong><br />

the health and per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance (Johns, Bright, 2001) and could have different impact <strong>on</strong> insomnia. There is also lack <str<strong>on</strong>g>of</str<strong>on</strong>g> researches showing specificity<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> psychotraumatic influence <strong>on</strong> subjective and objective sleep in insomnia patients.<br />

Aims. The aim <str<strong>on</strong>g>of</str<strong>on</strong>g> this study was to identify the peculiarities <str<strong>on</strong>g>of</str<strong>on</strong>g> subjective and objective symptoms and perpetuating factors <str<strong>on</strong>g>of</str<strong>on</strong>g> chr<strong>on</strong>ic insomnia<br />

triggered by serious life event comparing to chr<strong>on</strong>ic insomnia triggered by other factors.<br />

Method. 73 insomniac patients (23 male and 50 female, mean age 46±13 years old) receiving ambulatory treatment in the Somnology <str<strong>on</strong>g>Centre</str<strong>on</strong>g> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Moscow Medical Academy took part in our research. The procedure included clinical and psychological diagnostics and polysomnography<br />

recording.<br />

Clinical diagnostics were provided by somnologists according to Internati<strong>on</strong>al Sleep Disorders Classificati<strong>on</strong> in the sec<strong>on</strong>d revise (Principles…,<br />

2005). Only patients with chr<strong>on</strong>ic insomnia without any psychiatric and sleep disorders were included.<br />

Psychological diagnostics included diagnostic <str<strong>on</strong>g>of</str<strong>on</strong>g> the serious life event tended to trigger insomnia and diagnostics <str<strong>on</strong>g>of</str<strong>on</strong>g> subjective sleep quality and<br />

insomnia perpetuating factors. First, we asked patients when their symptoms began and what could be the possible reas<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> them. Then, we<br />

asked them whether they had experienced something traumatic just be<str<strong>on</strong>g>for</str<strong>on</strong>g>e <str<strong>on</strong>g>of</str<strong>on</strong>g> a year be<str<strong>on</strong>g>for</str<strong>on</strong>g>e first insomnia symptoms and whether it could be<br />

related to insomnia. Then patients filled Russian versi<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> Insomnia Severity Index (ISI, Bastien et al, 2001, Savard et al, 2005), Hospital Scale <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

Anxiety and Depressi<strong>on</strong> (Hamilt<strong>on</strong>), Dysfuncti<strong>on</strong>al Beliefs about Sleep Inventory (DBAS, Espie et al, 2000) and Glasgow C<strong>on</strong>tent <str<strong>on</strong>g>of</str<strong>on</strong>g> Thoughts<br />

Inventory (GCTI, Harvey & Espie, 2004). All the tests were validated in Russian samples <str<strong>on</strong>g>of</str<strong>on</strong>g> good sleepers and insomniacs in our researches and<br />

had moderate to high reliability and validity scores.<br />

Polysomnography were c<strong>on</strong>ducted by electroencephalography (EEG), electromyography (EMG) and electrooculography (EOG) registrati<strong>on</strong><br />

during the night. The next parameters were evaluated: total sleep time and sleep time without time <str<strong>on</strong>g>of</str<strong>on</strong>g> awake, durati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> all the sleep phases and<br />

stages (and their percent porti<strong>on</strong> in sleep), latent periods <str<strong>on</strong>g>of</str<strong>on</strong>g> stages, awake time and it percent porti<strong>on</strong> in sleep, number <str<strong>on</strong>g>of</str<strong>on</strong>g> awakenings.<br />

Results. Most <str<strong>on</strong>g>of</str<strong>on</strong>g> the serious life events reported by patients were deaths <str<strong>on</strong>g>of</str<strong>on</strong>g> relatives (parents, children or partners) and problems with children<br />

(drug dependence, in<str<strong>on</strong>g>for</str<strong>on</strong>g>mati<strong>on</strong> about child’s sexual orientati<strong>on</strong> etc). In 4 cases we included loss <str<strong>on</strong>g>of</str<strong>on</strong>g> job because it was describes by patients as a<br />

loss <str<strong>on</strong>g>of</str<strong>on</strong>g> meaning <str<strong>on</strong>g>of</str<strong>on</strong>g> their life and activity.<br />

There were 27% insomniacs (20 people) who reported a serious life event just be<str<strong>on</strong>g>for</str<strong>on</strong>g>e insomnia began and other 27% (20 people) who reported<br />

more than <strong>on</strong>e serious life be<str<strong>on</strong>g>for</str<strong>on</strong>g>e and after the beginning <str<strong>on</strong>g>of</str<strong>on</strong>g> insomnia that worsen symptoms.<br />

Patients experienced more than <strong>on</strong>e serious life event have more dysfuncti<strong>on</strong>al attitudes to sleep (p


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Also our c<strong>on</strong>clusi<strong>on</strong>s are limited by the retrospective character <str<strong>on</strong>g>of</str<strong>on</strong>g> the study we found that subjective sleep complaints d<strong>on</strong>’t depend <strong>on</strong> whether<br />

insomnia appeared as a result <str<strong>on</strong>g>of</str<strong>on</strong>g> serious life event or not. However, stressful experience triggered insomnia is related to anxiety level and<br />

dysfuncti<strong>on</strong>al beliefs about sleep which means that it is easier <str<strong>on</strong>g>for</str<strong>on</strong>g> insomnia to become chr<strong>on</strong>ic and harder to treat it (Bastein et al, 2001).<br />

Experiencing a number <str<strong>on</strong>g>of</str<strong>on</strong>g> serious life events triggered insomnia is related to worse objective sleep, such patients have less delta sleep and l<strong>on</strong>ger<br />

awake period. Although it doesn’t reflect in sleep complaints <str<strong>on</strong>g>of</str<strong>on</strong>g> such patients, severity <str<strong>on</strong>g>of</str<strong>on</strong>g> their insomnia might be higher and they may require<br />

different approach in treatment.<br />

8_20_P<br />

(poster secti<strong>on</strong> A1, poster board #28, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ROLE OF PHOTOSYNTHETIC PERFORMANCE IN SALT STRESS ACCLIMATION<br />

OF TOMATO AFTER SALICYLIC ACID PRE-TREATMENT<br />

Katalin Gémes, Ágnes Szepesi, Adrienn Guóth, Irma Tari<br />

Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Plant Physiology, University <str<strong>on</strong>g>of</str<strong>on</strong>g> Szeged, 6701 Szeged POB 654, Hungary<br />

pigsy17@freemail.hu, tari@bio.u-szeged.hu<br />

Impositi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> salt stress reduced the net CO 2 assimilati<strong>on</strong> rate, chlorophyll (Chl) and carotenoid c<strong>on</strong>tents,<br />

stomatal c<strong>on</strong>ductance and biomass producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> tomato (Lycopersic<strong>on</strong> esculentum Mill. L. cv. Rio Fuego). Pretreatments<br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> plants with 10 -4 M, but not with 10 -7 M salicylic acid (SA), could partially restore the CO 2<br />

fixati<strong>on</strong> rate, Chl a/b ratio, carotenoid levels under 100 mM NaCl exposure. Accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> soluble sugars,<br />

a biochemical marker <str<strong>on</strong>g>of</str<strong>on</strong>g> salinity tolerance <str<strong>on</strong>g>of</str<strong>on</strong>g> tomato, could be detected in pre-treated plants. After SA<br />

applicati<strong>on</strong>, c<strong>on</strong>centrati<strong>on</strong>s <str<strong>on</strong>g>of</str<strong>on</strong>g> hexoses (glucose and fructose) remained high in the leaves and that <str<strong>on</strong>g>of</str<strong>on</strong>g> sucrose<br />

in the roots during salt stress. Both SA and salt stress increased the H 2 O 2 producti<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> tissues. Activities <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

superoxide dismutase and catalase (CAT), a H 2 O 2 generating and scavenging enzymes, respectively, decreased<br />

significantly under salt stress, but in 10 -4 M SA pre-treated plants, CAT activity was significantly induced both<br />

in the root and shoot tissues. The improved photosynthetic per<str<strong>on</strong>g>for</str<strong>on</strong>g>mance, the accumulati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> soluble sugars<br />

as compatible osmolytes and the effective eliminati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> H 2 O 2 by CAT c<strong>on</strong>tributed to the successful<br />

acclimati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> 10 -4 M SA pre-treated tomato plants to high salinity.<br />

8_21_P<br />

(poster secti<strong>on</strong> A1, poster board #29, 24 th <str<strong>on</strong>g>of</str<strong>on</strong>g> August)<br />

ACTIVATION OF MITOGEN ACTIVATED PROTEIN KINASES FOLLOWING PACAP<br />

TREATMENT IN WARM ISCHEMIA/REPERFUSION INTESTINAL TISSUE<br />

A. Ferencz, B. Racz, A. Tamas * , A. Lubics * , D. Reglodi * , E. Roth<br />

University <str<strong>on</strong>g>of</str<strong>on</strong>g> Pecs, Medical School, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Surgical <str<strong>on</strong>g>Research</str<strong>on</strong>g> and Techniques, * Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Anatomy,<br />

H-7624 Pecs Szigeti street 12, Hungary<br />

During warm ischemia/reperfusi<strong>on</strong> (I/R) the oxidative stress triggers several intracellular pathways via<br />

mitogen activated protein kinases (MAPK). MAPK include the extracellular signal related kinase (ERK),<br />

c-Jun N-terminal kinase (JNK), and p38 MAPK. Pituitary adenylate cyclase-activating polypeptide (PACAP)<br />

is present and plays a central role in the intestinal physiology. We investigated the effect <str<strong>on</strong>g>of</str<strong>on</strong>g> PACAP <strong>on</strong> the<br />

activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> MAPK during small bowel warm I/R <strong>on</strong> Wistar rats. Group (G) I: no-ischemia; GII: 1 hour<br />

ischemia; GIII: 2 hs ischemia; GIV: 3 hs ischemia; GV: no-ischemia+PACAP; GVI: 1 hour<br />

ischemia+PACAP; GVII: 2 hs ischemia+PACAP; GVIII: 3 hs ischemia+PACAP. Reperfusi<strong>on</strong> was 3 hs in all<br />

groups. In GV-GVIII 5 µg bolus and 25 µg c<strong>on</strong>tinuous PACAP were administrated during reperfusi<strong>on</strong><br />

intravenously. Small bowel biopsies were collected after laparotomy, at the end <str<strong>on</strong>g>of</str<strong>on</strong>g> ischemia and reperfusi<strong>on</strong><br />

39


23-26 August 2007,<br />

Budapest, Hungary<br />

periods. In small bowel tissue the activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> ERK1/2, JNK1/2 and p38 MAPK were investigated by<br />

immunocytochemistry and Western blot analysis. Our results showed that in GII-GIV the phosphorilated<br />

ERK1/2 level decreased, meanwhile the phosphorilated JNK1/2 and p38 MAPK activati<strong>on</strong> elevated<br />

compare to c<strong>on</strong>trol level. In GVI-GVIII PACAP enhanced the phospho-ERK1/2 appearance and decreased<br />

JNK and p38 MAPK activity by the end <str<strong>on</strong>g>of</str<strong>on</strong>g> reperfusi<strong>on</strong>. We have provided evidence that warm I/R decreased<br />

phosphorilated ERK1/2 level and increased JNK1/2 and p38 MAPK activity. Which means, that warm I/R<br />

caused a trigger effect to apoptotic cell death. In c<strong>on</strong>trast, PACAP treatment induced the protective signalling<br />

pathways against oxidative injury by MAPK in bowel tissue.<br />

40


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

AUTHORS’ INDEX<br />

A<br />

Ábrahám, E. 19<br />

Abrishamchi, P. 20<br />

Aguilar-Zavala, H. 33<br />

Alvarez, C. 37<br />

Aradillas-García, C. 33<br />

Arai, H. 35<br />

Arrigo, A. P. 3, 12<br />

Arya, R. 7<br />

B<br />

Badawi, M. A.-H. 17<br />

Badicean, D. 13<br />

Badran, G. E. 17<br />

Balogi, Z. 4<br />

Bánhegyi, G. 36<br />

Barbacaru, N. 13<br />

Barber, J. R. 22<br />

Bauer, M. 28<br />

Belova, L. 7<br />

Benedetti, A. 36<br />

Bernatova, I. 26<br />

Bharti, A. 5<br />

Bhatnagar, S. 30<br />

Bittsánszky, A. 15<br />

Boccuti, S. 21<br />

Bock, I. 15<br />

Bockmeyer, C. L. 28<br />

Bódizs, R. 31<br />

Borrelli, J. M. 6<br />

Brickley, D. 7<br />

Bristow, G. R. 6<br />

Brunkhorst, F. M. 28<br />

Bryden, L. W. 24<br />

Budde, U. 28<br />

Bugán, A. 31<br />

Bunck, A. C. 28<br />

Byun, Y.-J. 14<br />

C<br />

Calderwood, S. 5<br />

Chandler, V. 13<br />

Chaves, M. M. 19<br />

Cheregi, O. 4<br />

Choi, M. 14<br />

Ciocca, R. D. 5<br />

Claus, R. A. 28<br />

Collins, J. 14<br />

C<strong>on</strong>l<strong>on</strong>, M. J. 24<br />

C<strong>on</strong>ti, V. 21<br />

C<strong>on</strong>zen, D. S. 7<br />

Corbi, G. 21<br />

Crooks, A. P. 6<br />

Czakó, M. 15<br />

Czekalla, A. 3<br />

Csermely, P. 11<br />

D<br />

de la Cruz-Mendoza, E. 33<br />

Debreczeny, M. 4<br />

Deigner, H. P. 28<br />

Diaz-Latoud, C. 3<br />

Dibner, C. 5<br />

Dorer, M. 28<br />

Du, J. 29<br />

E<br />

Eichler, P. 19<br />

Ejtehadi, H. 20<br />

El-Bastawisy, Z. M. 17<br />

Eom, C.-Y. 9, 10<br />

F<br />

Ferencz, A. 39<br />

Ferrara, N. 21<br />

Filippelli, A. 21<br />

Filippelli, W. 21<br />

Finklestein, S. P. 22<br />

Fleury-Olela, F. 5<br />

Fortunato, A. 19<br />

Freeman, L. M. 6<br />

Fujita, E. 34<br />

Fujita, Y. 34<br />

G<br />

Galiba, G. 16<br />

Garay-Sevilla, M. E. 33<br />

García, V. 37<br />

Gardiner, J. 13<br />

Gardner, T. 14<br />

41


23-26 August 2007,<br />

Budapest, Hungary<br />

Gémes, K. 39<br />

Gibert, B. 3<br />

Giese, K. 4<br />

Gobara, N. 29<br />

Goldaracena-Azuara, M. 33<br />

G<strong>on</strong>zalez-Cor<strong>on</strong>el, A.-S. 18<br />

Goto, K. 27<br />

Gouveia, M. M. 19<br />

Gray, J. P. 5<br />

Gullner, G. 15<br />

Guóth, A. 39<br />

Gurd<strong>on</strong>, Cs. 18<br />

Gyulai, G. 15<br />

H<br />

Harmat, L. 31<br />

Harn, C. 14<br />

Hassan, M. N. 17<br />

Hazag, A. 31<br />

Heszky, L. 15<br />

Higashikubo, R. 6<br />

Horikoshi, N. 6<br />

Horváth, I. 4<br />

Huang, X.-P. 29<br />

Hupe, D. 28<br />

Hwang, J. 14<br />

Hwang, J.-H. * 14<br />

Hyun, C.-D. 9<br />

I<br />

Ibarra- Torres, M. 33<br />

Iyer, V. 30<br />

J<br />

Jacota, A. 13<br />

Janaszak, A. 3<br />

Jean-Charles, P. 29<br />

Jenes, B. 18<br />

Joo, H. 14<br />

Joung, Y. H. 26<br />

Juhasz, K. 4<br />

Jung, J. R. 9, 10<br />

Jung, Y. E. 26<br />

K<br />

Kaiser, P. 24<br />

Kalkkinen, N. 16<br />

Karalis, P. K. 23<br />

Kellős, T. 16<br />

Khaleque, A. 5<br />

Kim, E.-H. 9<br />

Kim, G.-W. 10<br />

Kim, H.-Y. 9, 10<br />

Kim, J.-H. 26<br />

Kim, M.-S. 10<br />

Kiss, J. 15<br />

Kocsy, G. 16<br />

Kojima, A. 27<br />

K<strong>on</strong>cz, Cs. 19<br />

K<strong>on</strong>opa, G. 3<br />

Kőmíves, T. 15<br />

Kreft, M. 35<br />

Kreft, M. E. 35<br />

Kretz-Remy, C. 12<br />

Kurt, O. 18<br />

Ky, B. 7<br />

L<br />

Lahouti, M. 20<br />

Lakhotia, C. S. 7<br />

Lancien, F. 24<br />

Laszlo, A. 6<br />

Le Mével, J-C. 24<br />

Lee, D. 14<br />

Lee, D.-H. 14<br />

Lee, S. K. 26<br />

Lehoczky, P. 15<br />

Lepock, R. J. 6<br />

Lösche, W. 28<br />

Lubics, A. 39<br />

M<br />

Major, J. 31<br />

Malacara, J. M. 33<br />

Mal<strong>on</strong>e, R. P. 15<br />

Márt<strong>on</strong>, L. 15<br />

Matikainen, S. 16<br />

Matsuba, Y. 27<br />

Messina, C. 25<br />

Mimassi, N. 24<br />

Molnár, L. 37<br />

Molnár, R. 31<br />

Morioka, S. 27<br />

Muthusamy, V. 6<br />

N<br />

Nadratowska-Wesolowska, B. 3<br />

42


3rd Cell Stress Society Internati<strong>on</strong>al C<strong>on</strong>gress <strong>on</strong><br />

Stress Resp<strong>on</strong>ses in Biology and Medicine and<br />

2nd World C<strong>on</strong>ference <str<strong>on</strong>g>of</str<strong>on</strong>g> Stress<br />

Nagy, Z. 37<br />

Naito, T. 27<br />

Nan, C. 29<br />

Nemat Alla, M. M. 17<br />

Ng, S. C. 22<br />

Niv<strong>on</strong>, M. 12<br />

Nyman, A. T. 16<br />

O<br />

Ohira, Y. 27<br />

Ohtawa, M. 35<br />

Óvary, Cs. 37<br />

Öhman, T. 16<br />

P<br />

Papdi, Cs. 19<br />

Park, S.-K. 9, 10<br />

Park, Y. 14<br />

Partelli, F. 19<br />

Pérez, J. 37<br />

Pérez-Luque, E. L. 33<br />

Potokar, M. 35<br />

Prathiba, J. M. 19<br />

Putics, Á. 11<br />

Puzserova, A. 26<br />

Pyun, H. K. 26<br />

R<br />

Racz, B. 39<br />

Ramalho, J. C. 19<br />

Rasskazova, E. 38<br />

Reglodi, D. 39<br />

Reinhart, K. 28<br />

Reinke, H. 5<br />

Ren, J. M. 22<br />

Rengo, G. 21<br />

Ribeiro, A. 19<br />

Rinaldi, B. 21<br />

Rintahaka, J. 16<br />

Rossi, F. 21<br />

Roth, E. 39<br />

Roti, R. L. J. 6<br />

S<br />

Sakurai, T. 34<br />

Santulli, A. 25<br />

Sasi, S. 6<br />

Sawani, J. 6<br />

Schibler, U. 5<br />

Scholz, M. 13<br />

Schweizer, P. 16<br />

Sekhar, R. K. 6<br />

Sharma, S. 7<br />

Shim, I. 26<br />

Shini, A. 24<br />

Shini, S. 24<br />

S<strong>on</strong>ar, N. V. 6<br />

S<strong>on</strong>g, E. 10<br />

Soto, C. 37<br />

Sőti, Cs. 11<br />

Stati, A. 5<br />

Stein, N. 16<br />

Stenovec, M. 35<br />

Stiuso, P. 21<br />

Sugiura, T. 27<br />

Suh, J.-H. 9, 10<br />

Szabados, L. 19<br />

Szabó, G. 37<br />

Száraz, P. 36<br />

Szemán, A. 31<br />

Szepesi, Á. 39<br />

T<br />

Takács, J. 31<br />

Tamas, A. 39<br />

Tanaka, Y. 32<br />

Tari, I. 39<br />

Taylor, A. 3<br />

Teli, T. 23<br />

Tsuda, A. 32<br />

Tsuda, S. 32<br />

U<br />

Uría, E. 37<br />

V<br />

Valverde, A. R. 12<br />

Vass, I. 4<br />

Végh, M. E. 11<br />

Vierling, E. 4<br />

Vígh, L. 4<br />

Vilwanathan, K. R. 10<br />

43


23-26 August 2007,<br />

Budapest, Hungary<br />

W<br />

Weintraub, A. 30<br />

Wieland, S. 22<br />

Y<br />

Yamaguchi, T. 34<br />

Yoriko, A. 34, 35<br />

Yoshioka, T. 27<br />

Yu, S. 14<br />

Z<br />

Zaker, A. 20<br />

Zorec, R. 35<br />

44


ERRATUM LIST OF THE ABSTRACT BOOK OF 2ND WORLD CONFERENCE OF STRESS<br />

2D_03_S<br />

ISCHEMIA/STROKE IMPAIRS ENDOPLASMIC RETICULUM FUNCTION<br />

Wulf Paschen, Ph.D.<br />

Multidisciplinary Neuroprotecti<strong>on</strong> Laboratories, Department <str<strong>on</strong>g>of</str<strong>on</strong>g> Anesthesiology, Duke University Medical Center,<br />

Durham, North Carolina, USA, e-mail: wulf.paschen@duke.edu<br />

Cerebral Ischemia/Stroke is a severe <str<strong>on</strong>g>for</str<strong>on</strong>g>m <str<strong>on</strong>g>of</str<strong>on</strong>g> metabolic stress that interferes with most biochemical and<br />

molecular biology pathways. A prominent feature <str<strong>on</strong>g>of</str<strong>on</strong>g> transient cerebral ischemia is an irreversible<br />

suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> protein synthesis in vulnerable cells. Ischemia-induced shutdown <str<strong>on</strong>g>of</str<strong>on</strong>g> translati<strong>on</strong> is caused<br />

by phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the alpha subunit <str<strong>on</strong>g>of</str<strong>on</strong>g> the eukaryotic initiati<strong>on</strong> factor 2 (eIF2a) resulting in<br />

disaggregati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> polyribosomes and a block <str<strong>on</strong>g>of</str<strong>on</strong>g> translati<strong>on</strong> at the initiati<strong>on</strong> step. Am<strong>on</strong>g the four eIF2a<br />

kinases identified so far, the double stranded RNA-activated kinase-like endoplasmic reticulum (ER)<br />

kinase (PERK) was the <strong>on</strong>ly kinase found to be activated after ischemia. This implies that cerebral<br />

ischemia impairs ER functi<strong>on</strong>. When ER functi<strong>on</strong> is impaired, the unfolded protein resp<strong>on</strong>se is activated<br />

resulting in phosphorylati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> eIF2a-induced shutdown <str<strong>on</strong>g>of</str<strong>on</strong>g> translati<strong>on</strong> and activati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> the expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

genes coding <str<strong>on</strong>g>for</str<strong>on</strong>g> ER stress genes. Expressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> genes coding <str<strong>on</strong>g>for</str<strong>on</strong>g> ER stress genes is activated after<br />

ischemia. Since suppressi<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> protein synthesis is an irreversible process in vulnerable brain structures<br />

such as the hippocampal CA1 subfield, the rise in mRNA levels is not followed by an increase in levels <str<strong>on</strong>g>of</str<strong>on</strong>g><br />

the respective proteins in CA1 neur<strong>on</strong>s. ER stress-induced gene expressi<strong>on</strong> requires programmed<br />

recovery from translati<strong>on</strong>al repressi<strong>on</strong>. This is a process induced by GADD34, a protein activating the<br />

type 1 protein phosphatase that de-phosphorylates eIF2a-P. Post-ischemic translati<strong>on</strong> <str<strong>on</strong>g>of</str<strong>on</strong>g> gadd34 mRNA is<br />

activated in the resistant cortex but not in the vulnerable CA1 subfield. Forced post-ischemic activati<strong>on</strong><br />

<str<strong>on</strong>g>of</str<strong>on</strong>g> GADD34 expressi<strong>on</strong> may help vulnerable cells to restore protein synthesis and thus to recover from<br />

transient cerebral ischemia.


1D_04_S => Justin Yerbury instead <str<strong>on</strong>g>of</str<strong>on</strong>g> Mark R. Wils<strong>on</strong><br />

1F_02_P => Building A, Poster Secti<strong>on</strong> A1, Board A#57<br />

1F_03_P => Building A, Poster Secti<strong>on</strong> A1, Board A#58<br />

1F_04_P => Building A, Poster Secti<strong>on</strong> A1, Board A#59<br />

1F_05_P => Building A, Poster Secti<strong>on</strong> A1, Board A#60<br />

1F_06_P => Building A, Poster Secti<strong>on</strong> A1, Board A#61<br />

1F_07_P => Building A, Poster Secti<strong>on</strong> A1, Board A#62<br />

1F_08_P => Building A, Poster Secti<strong>on</strong> A1, Board A#63<br />

1F_09_P => Building A, Poster Secti<strong>on</strong> A1, Board A#64<br />

1F_10_P => Building A, Poster Secti<strong>on</strong> A1, Board A#70<br />

2A_10_P => Building B, Poster Secti<strong>on</strong> B1, Board B#168<br />

2C_04_S => cancelled<br />

2G_06_P => Building B, Poster Secti<strong>on</strong> B1, Board B#242<br />

2G_07_P => Building B, Poster Secti<strong>on</strong> B1, Board B#243<br />

2G_08_P => Building B, Poster Secti<strong>on</strong> B1, Board B#244<br />

5C_03_S => cancelled<br />

5E_05_S => Building B, Poster Secti<strong>on</strong> B1, Board B#245<br />

5E_06_S => Building B, Poster Secti<strong>on</strong> B1, Board B#246<br />

5E_02_P => Building B, Poster Secti<strong>on</strong> B1, Board B#247<br />

5E_03_P => Building B, Poster Secti<strong>on</strong> B1, Board B#248<br />

5E_05_P => Building B, Poster Secti<strong>on</strong> B1, Board B#249<br />

5J_01_P => Building B, Poster Secti<strong>on</strong> B1, Board B#245<br />

5J_02_P => Building B, Poster Secti<strong>on</strong> B1, Board B#246<br />

5J_03_P => Building B, Poster Secti<strong>on</strong> B1, Board B#247<br />

5J_04_P => Building B, Poster Secti<strong>on</strong> B1, Board B#248<br />

5J_05_P => Building B, Poster Secti<strong>on</strong> B1, Board B#249<br />

5J_06_P => Building B, Poster Secti<strong>on</strong> B1, Board B#250<br />

5J_07_P => Building B, Poster Secti<strong>on</strong> B1, Board B#251<br />

5J_08_P => Building B, Poster Secti<strong>on</strong> B1, Board B#252<br />

5J_09_P => Building B, Poster Secti<strong>on</strong> B1, Board B#253<br />

5J_10_P => Building B, Poster Secti<strong>on</strong> B1, Board B#254<br />

5J_11_P => Building B, Poster Secti<strong>on</strong> B1, Board B#255<br />

6D_04_S => Yaakov Naparstek instead <str<strong>on</strong>g>of</str<strong>on</strong>g> Irun R. Cohen<br />

6E_06_S => cancelled<br />

6F_01_P => Building B, Poster Secti<strong>on</strong> B2, Board B#286<br />

6F_02_P => Building B, Poster Secti<strong>on</strong> B2, Board B#287<br />

6F_03_P => Building B, Poster Secti<strong>on</strong> B2, Board B#288<br />

6F_04_P => Building B, Poster Secti<strong>on</strong> B2, Board B#289<br />

6F_08_P => Building B, Poster Secti<strong>on</strong> B2, Board B#290<br />

6F_09_P => Building B, Poster Secti<strong>on</strong> B2, Board B#291<br />

6G_02_S => cancelled<br />

8_11_P => 6F_12_P, Building B, Poster Secti<strong>on</strong> B2, Board B#299<br />

7I_13_P => Building A, Poster Secti<strong>on</strong> A2, Board A#164<br />

7I_14_P => Building A, Poster Secti<strong>on</strong> A2, Board A#319<br />

7I_20_P => Building A, Poster Secti<strong>on</strong> A2, Board A#170<br />

7J_01_P => Building A, Poster Secti<strong>on</strong> A2, Board A#320

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