Book of abstracts - British Neuroscience Association

6.07
Proteomic identification of biomarkers for neural stem cells
Maltman D J, Przyborski S A
School of Biological and Biomedical Sciences, University of Durham,
South Road, Durham, DH1 3LE, , ReInnervate Ltd, Durham, DH1 3HP
Stem cell research suffers from a lack of protein markers which allow
precise definition of cellular identity. Of the few antigens used to define
neural stem/progenitor cells none are expressed exclusively in these
populations. This highlights a strong requirement for novel specific
biomarkers. Comparative proteomic approaches hold great promise
for the rapid detection of such molecules, and employment of
complementary sample preparation methods will enhance the
likelihood of their detection. High-throughput determination of cellular
status is now achievable through the combined detection of several
differentially expressed proteins. Such profiling employs time-of-flight
(TOF) mass spectrometry and circumvents any requirement for
individual protein identification. Previously, surface-enhanced laser
desorption ionization (SELDI)-TOF biomarker profiling has been
successfully used in our laboratory to distinguish between control and
differentiated cultures of a model stem cell system. Currently we are
applying high accuracy matrix-assisted laser desorption ionization
(MALDI)-TOF to profile a human neural stem cell line and its
differentiated derivatives. Profiles generated will not only enable the
rapid evaluation of sample status, but also provide a lead into specific
biomarkers whose further characterization will increase our
understanding of neurogenesis. These proteins will also form the basis
for the development of immunological tools which may be used for
improved cellular characterization and isolation.
6.08
Neural effects of novel synthetic retinoids.
Christie V.B., Cartmell E, Whiting A, Marder T.B., Przyborski S.A.
Durham University, Biological and Biomedical sciences, South Rd, Durham,
DH1 3LE
Vitamin A and its derivatives, collectively termed retinoids, are essential for
many biologically important processes and are used to modulate cell
proliferation and differentiation in vitro. The importance of optimal retinoid
function in embryonic neural development is well known, and is now being
realised in certain adult neural cell populations. The aim of this research is
to study the role of both natural and synthetic retinoids during neural
differentiation in adult verses embryonic model systems. It has been shown
that all-trans-retinoic acid (ATRA), a naturally occurring retinoid which
activates all retinoic acid receptor (RAR) subtypes, induces neural
differentiation in several cell model systems, including embryonic stem cells
and adult neural progenitors. The in vitro study of ATRA, however, is
complicated by its photo-isomerisation when used under standard
laboratory conditions. To try and overcome this, we have synthesised
ATRA retinoid analogues, EC23 and EC19, which unlike ATRA do not
isomerise in response to light or heat. These compounds could therefore be
potentially advantageous over ATRA for use in in-vitro investigations into
retinoid modes of action. Preliminary data show that EC23 elicits similar
cellular responses to ATRA when tested in vitro, whereas EC19 appears to
induce a higher percentage of glia in adult neural progenitor models. This
work will have the potential to aid research into pharmacological
manipulation of neural differentiation and its associated receptors for
potential therapeutic application.
6.09
Generic scaffolds for 3-dimensional in vitro neural cell co-culture
Bokhari M, Carnachan R, Cameron N, Przyborski S
School of Biological and Biomedical Science; Department of
Chemistry, University of Durham DH1 3LE; 3ReInnervate Limited, Old
Shire Hall, Old Elvet, Durham DH1 3HP
The structure and function of cultured cells are dramatically affected
by the micro-environment in which they are grown. Traditionally, two
dimensional (2-D) polystyrene surfaces are used to support cell
growth in vitro; however such surfaces do not enable the most
favourable cell growth and function. A more thorough understanding of
cell biology and cell-cell interactions requires three dimensional (3-D)
culture systems that more closely represent the natural structure and
function of tissues in vivo. Here we present a cell culture device that
provides a 3-D environment for routine cell culture. We have
developed a polystyrene scaffold which exhibits a well defined and
uniform porous micro-architecture and have adapted these threedimensional
scaffolds for cell culture and/or tissue engineering
applications. These scaffolds are readily adaptable to many different
types of tissue culture plastic-ware including 6- and 24-well plates.
These culture devices are pre-fabricated, sterile, easy to use and are
handled in a similar manner to standard 2-D plastic-ware. Our work
investigates the production of these polymers for routine 3-D cell
growth in-vitro, as well as neuron-glia co-culture. Synergistic effects of
scaffold micro-architecture combined with chemical and/or biological
stimuli on neuronal cell culture have not yet been explored in detail.
The aim of this work is to determine the synergistic effects of physical,
chemical and biological guidance cues on neuronal differentiation,
viability and function in an in vitro microenvironment which more
closely resembles the cellular microenvironment in vivo.
6.10
Neuritogenesis in adult hippocampal neurons in growth- permissive
versus inhibitory environments in vitro
Mellough C B 1, Wood A 2, Przyborski S A 1
1School of Biological and Biomedical Sciences, University of Durham,
South Rd, Durham, UK. 2 Wyeth Neuroscience, 865 Ridge Rd, Monmouth
Junction, New Jersey, USA.
The inability of the adult central nervous system to regenerate following
injury largely depends on the expression of myelin-associated inhibitors.
These ligands are present before the glial scar has formed and bind to the
p75NTR-NgR receptor complex on regenerating neurons, causing growth
cone collapse and axonal retraction. Progress is being made towards
elucidation of the downstream events which result in growth cone collapse
and retraction of the axonal cytoskeleton. The majority of studies that
investigate myelin inhibition employ neuronal populations derived from the
postnatal developmental period, or which lie in close apposition to the
regeneration-permissive peripheral nervous system in vivo. In this study,
the effects of myelin-associated glycoprotein (MAG) on neurite outgrowth
was assessed in a population of differentiating neurons derived from adult
hippocampal neural progenitor cells. We show that MAG does not alter
neural progenitor cell fate but, unlike their developmental counterparts,
neurite outgrowth from differentiating neurons was significantly attenuated
by MAG. We demonstrate that this effect can be partially overcome (by up
to 69%) by activation of the neurotrophin, cAMP and PKA pathways or by
Rho-kinase suppression. We also demonstrate that combining regeneration
promoting methods elicits enhanced neurite outgrowth from differentiating
neurons under myelin inhibitory conditions when compared with solitary
application. This work pertains especially to the facilitation of neural repair
in the compromised adult brain by endogenous mechanisms, such as the
mobilisation and appropriate integration of host stem cells for functional
replacement within depleted neuronal circuitry.
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