The Toxicologist - Society of Toxicology

cells with MeHg. N-acetylcysteine effectively reversed the MeHg-induced cell
apoptosis-related signals. Taken together, our data suggest that MeHg-induced oxidative
stress causes pancreatic β-cells apoptosis, and both mitochondrial-dependent
and ER stress- mediated apoptotic signals involve in the MeHg-induced pancreatic
β-cells death.
308 ROLES OF RIP1 AND YIL006W, MITOCHONDRIAL
PROTEINS, IN METHYLMERCURY TOXICITY IN
BUDDING YEAST.
J. Lee, G. Hwang and A. Naganuma. Graduate School of Pharmaceutical Sciences,
Tohoku University, Sendai, Japan.
Methylmercury (MeHg) is an environmental pollutant that causes severe damage in
central nervous system. However, the molecular mechanism of MeHg toxicity is
not fully understood. Several investigators have demonstrated that mitochondria
are potentially involved in MeHg toxicity. In order to characterize functional relationship
between MeHg toxicity and mitochondria, we searched for mitochondrial
proteins involved in MeHg toxicity in budding yeast. Yeast is a genetically wellcharacterized
eukaryotic organism that shares many genes common with mammals,
including humans. We recently found that yeast cells deficient in Yil006w exhibited
resistance to MeHg as compared to wild-type yeast. Yil006w is one of the transporter
families existing in inner-membrane of mitochondria. Overexperssion of
Yil006w conferred hypersensitivity to MeHg. Moreover, not only cellular protein
level, but also mRNA level of Yil006w was increased by MeHg treatment.
Mitochondrial level of Yil006w was also increased by treatment of MeHg. These results
suggested that MeHg might increase the mitochondrial Yil006w level and
strengthen its toxicity. We previously demonstrated that Rip1, a component of mitochondrial
electron transfer chain complex III, was potentially involved in the production
of reactive oxygen species (ROS) in response to MeHg. Therefore, we examined
relationship between Rip1 and Yil006w in MeHg toxicity. We treated yeast
cells, which lacked both Rip1 and Yil006w, with MeHg. The degree of resistance to
MeHg was not increased by the deletion of both Rip1 and Yil006w as compared to
each single-deletion mutants, suggesting that Rip1 and Yil006w strengthen MeHg
toxicity through a mechanism involved in a common pathway. We also found that
the increase of ROS level induced by MeHg was significantly attenuated in
Yil006w-deficient yeast. Thus, Yil006w might have a major role in the mechanism
of Rip1-enhanced ROS generation in response to MeHg.
309 MERCURY MODULATE THE CYTOCHROME P450 1A1
IN C57BL/6 MICE: IN VIVO AND IN VITRO STUDIES.
I. Amara and A. El-kadi. Faculty of Pharmacy and Pharmaceutical Sciences,
University of Alberta, Edmonton, AB, Canada.
Heavy metals, such as mercury (Hg2+), and aryl hydrocarbon receptor (AhR) ligands
such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) are environmental cocontaminants
and their molecular interaction may disrupt the coordinated regulation
of the carcinogen-activating enzyme cytochrome P4501a1 (Cyp1a1).
Therefore, in the current study we examined the effect of Hg2+ on the expression
of Cyp1a1 in C57BL/6 mice livers and isolated mouse hepatocytes. For this purpose,
C57BL/6 mice were injected intraperitoneally with Hg2+ (2.5 mg/kg) in the
absence and presence of TCDD (15 μM/kg). After 6 and 24 h the tissue was harvested
and the expression of Cyp1a1 mRNA, protein and catalytic activity levels
were determined using real time-PCR, Western blot and 7-ethoxyresorufin Odeethylase
(EROD) analyses, respectively. In vitro, isolated mouse hepatocytes were
incubated with TCDD (1 nM) in the presence and absence of various concentrations
of Hg2+ (2.5, 5 and 10 μM). Our results demonstrate that at in vivo level,
Hg2+ significantly decreased the TCDD-mediated induction of Cyp1a1 mRNA at
6 h while potentiating its mRNA, protein, and catalytic activity levels at 24 h. At in
vitro level, Hg2+ significantly inhibited the TCDD-mediated induction of Cyp1a1
at mRNA, protein and activity levels in a concentration-dependent manner.
Investigating the effect of Hg2+ at transcriptional levels revealed that Hg2+ significantly
inhibited the TCDD-mediated induction of AhR-dependent luciferase reporter
gene expression. In conclusion, caution should be taken when extrapolating
in vitro data to in vivo situation. Furthermore, our results indicate more complex
regulation of Cyp1a1 at the in vivo level that warrants further investigation.
Funded by NSERC Discovery Grant RGPIN 250139-07.
66 SOT 2011 ANNUAL MEETING
310 ASSESSING METAL LEVELS IN CHILDREN FROM THE
MECHANISTIC INDICATORS OF CHILDHOOD
ASTHMA (MICA) STUDY.
A. Sanders 1 , J. Gallagher 2 , J. McGee 2 , S. Rhoney 2 , E. Hugens 2 , H. Özkaynak 3
and R. Fry 1 . 1 Environmental Sciences and Engineering, Gillings School of Global
Public Health, University of North Carolina, Chapel Hill, NC, 2 National Health
Environmental Effects Research Laboratory, U.S. EPA, Research Triangle Park, NC
and 3 National Exposure Research Laboratory, U.S. EPA, Research Triangle Park, NC.
Toxic and essential metals levels can be used as health indicators. Here we quantitatively
compare and contrast toxic and essential metals levels in vacuum dust, urine,
and fingernail samples of 109 children in Detroit, Michigan as part of The
Mechanistic Indicators of Childhood Asthma (MICA) study – a study aimed at examining
relationships between exposure and biomarkers of exposure effects and
susceptibility in children ages 9-13. Comparison of the Detroit children’s urinary
levels with the CDC’s Fourth National Report on Human Exposure to
Environmental Chemicals showed that urinary metal levels for As, Cd and Pb, were
similar to nationally reported levels. Creatinine-adjusted urinary metal concentrations
were in the order: Zn >> Se > Cu > As > Ni > Pb > Cd > Cr > V > Mn. We
find no statistical differences in essential metals levels between children with varied
vitamin use status. Our results indicate that levels of toxic metals indeed covary
with each other as well as with essential metals (e.g. arsenic and selenium) in the biological
matrices. Furthermore, among all possible pairwise comparisons of metals
and media types, metals levels in the matrices representing chronic exposure (nails
and dust) were more closely associated relative to the short term exposure indicator
(urine). This study underscores the importance for the consideration of both individual
and multiple metals in health effect studies. This abstract is the text of a proposed
presentation and does not represent EPA policy.
311 BRIDGING THE MISSING LINK IN PRINCIPLE
THEORY – SELENIUM TRANSPORT IN ENTEROCYTES
AND HEPATOCYTES.
S. Misra 1 , R. W.M. Kwong 2 and S. Niyogi 1 . 1 Biology, University of Saskatchewan,
Saskatoon, Saskatchwean, Canada and 2 Toxicology Centre, University of
Saskatchewan, Saskatoon, Saskatchwean, Canada.
Selenium (Se) is a unique trace element with diverse physiological functionality. In
contrast, it exhibits very low margin between essentiality and toxicity. Recent research
reveals potential applications of Se in cancer prevention and treatment.
Nevertheless, a simple, yet basic research question remains to be understood in
depth – how Se is transported in higher eukaryotic cells? To address this, the present
study was designed to evaluate Se transport kinetics at physiological concentrations
using isolated rainbow trout hepatocytes and enterocytes as model systems.
Selenite transport was linear in both cell types at the concentration range tested.
However, pre-incubation of selenite with glutathione (GSH) resulted in increased
Se uptake with saturable uptake kinetics in enterocytes but not in hepatocytes during
early uptake phase. L-cysteine, another thiol-group containing agent, increased
Se uptake in both cell types by ∼10-folds when pre-incubated with selenite.
Inhibitory effects of anionic transporter blocker, DIDS and analogous chemical
compounds (sulfite) suggested specific carrier mediated uptake of both forms.
However, DCCD and orthovanadate couldn’t block Se uptake, indicating an energy-independent
process. We then tested the hypothesis whether mercury can
block both forms of Se uptake or not. Mercury exerted uptake inhibition in both
cell types except reduced form (selenite + GSH) of Se in enterocytes. These observations
indicate the possibility of at least two different pathways of inorganic Se uptake
in enterocytes. However, when both cell types were pre-exposed in combination
with mercury and DIDS, the inhibitory effect was higher than DIDS but was
comparable with mercury alone. Absence of any synergistic effects in combining exposure
raises the possibility of inhibition of anionic transporter by mercury.
Overall, our data suggests a passive carrier mediated uptake of inorganic Se in both
cell types.
312 THE IRON TRANSPORTER FERROPORTIN CAN ALSO
FUNCTION AS A MANGANESE EXPORTER.
M. S. Madejczyk and N. Ballatori. Environmental Medicine, University of Rochester
School of Medicine, Rochester, NY.
Manganese (Mn) is an essential trace element that is utilized in a number of important
cellular processes; however, excess Mn can also be quite toxic. Chronic exposure
to high levels of Mn leads to neurological damage with psychiatric symptoms
as well as disruptions in motor function. The mechanisms by which Mn is transported
across cell membranes to reach its target sites are now beginning to be elucidated,
although much remains to be learned about these processes, and in particu-