We are studying the ways in which interacting networks of transcription factors and signal transduction molecules guide the development of precursor/stem cells into mature neurons of the mammalian nervous system. We also study the role of the same networks in neurodegenerative diseases and factors that can act therapeutically as neuroprotective agents.

Using monoclonal antibodies, cDNA probes and controlled tissue culture systems, we have mapped out events that lead to the formation of the separate lineages of retina and retinal pigment epithelium from a homogeneous neuroepithelial sheet during optic cup formation. Subsequent development of the retinal epithelium and the formation of retinal cell types occurs through an interplay of extrinsic and intrinsic factors. We have used genes encoding proteins of the visual transduction system to study the ways in which these factors act to regulate photoreceptor development. The onset of transcription of these genes delineates distinct phases of development that are under different regulatory mechanisms. We have identified important promoter elements in some of these genes using both biochemical methods and analysis in transgenic mice, and have purified the transcription factors with which they interact. Current studies are aimed at understanding how some of these transcription factors themselves are regulated. Through these experiments we are addressing the fundamental question of how the fate of a progenitor cell is directed to the production of specific neural cell types. We have shown that, under specific conditions, activation of STAT3 can block the formation of rod photoreceptors and promote the formation of Mller glial cells - two cell types derived from the late retinal progenitors (Zhang et al. IOVS 45:2407-12, 2004). We have also shown that STAT3 is expressed in neuroepithelial cells and is not found in the photoreceptor layer follow rod generation, suggesting that STAT3 withdrawal is key for rod differentiation (Zhang et al. Exp. Eye Res. 76:421-31, 2003; Zhang et al. Exp. Eye Res. 81:103-15, 2005). Studying of the regulatory mechanisms controlling STAT3 expression in retina will extend our understanding of mammalian retina development.

In collaboration with Dr. Samuel Shao-Min Zhang, we are using modern molecular methods to study retinal development at a systems level. From 2001 we have initiated and generated a set of expressed sequence tags (ESTs) from a series of mouse retina libraries. About 30,000 clones have been sequenced and 12,000 clones have been collected as a non-redundant EST set. This is the first and the largest collection of mouse retina transcripts. We have generated a comprehensive mouse retina transcriptome based on 81,000 murine retina transcripts from whole mouse ESTs. About 33,000 sequence-unique retina transcript clusters (RTCs) have been identified and the highest-grade retina-enriched pool covered almost all the known genes in phototransduction processes that are involved in human retinal diseases, suggesting the potential of gene discoveries for human retina disorders (Zhang et al., BMC Genomics 6:40, 2005). We have also generated mouse retina specific cDNA microarrays that represent about 10,000 UniGene clusters for studying the biological process of retina development. These arrays have been used to generate a complete developmental expression profile of the 10,000 genes. Looking at shared gene expression profiles and promoter elements of these genes is allowing us to define patterns of regulation that help explain the development of the retina.

Many forms of neurodegeneration involve oxidative stress. Reactive oxygen species are a natural by product of oxidative phosphorylation and this endogenous production by mitochondria lessens the ability of cells to withstand exogenous reactive oxygen species induced by stress or injury. One group of proteins that can regulate reactive oxygen species generation are the mitochondrial uncoupling proteins. In collaboration with Dr. Tamas Horvath we are studying ways in which mitochondrial uncoupling proteins alter developmental cell death in retinal ganglion cells and provide protection from a variety of neurotoxic insults. Using cells from mice engineered to overexpress or lack uncoupling proteins we have tested how these proteins prevent apoptotic cell death following treatment with various excitotoxins.. We have also tested the range of protection given by enhanced uncoupling protein expression in primate models of Parkinson's disease. (Horvath et al., Endocrinology 144:2757, 2003)

In collaboration with Dr. Joyce Tombran-Tink, we study the mechanisms by which neurotrophic factors protect cells from excitotoxic, oxidative and other insults. Specifically we are examining the transduction pathways activated by PEDF and comparing them with other protective factors such as CNTF, BDNF and GDNF. Rotation students would define active fragments of PEDF that mediate its potent neuroprotective and anti-angiogenic functions and will develop the best of these into new therapeutic agents. (see Tombran-Tink & Barnstable, Nature Reviews Neuroscience 4:628, 2003). PEDF is unique among these factors in that it is also one of the most potent antiangiogenic factors so far identified and we have recently defined the molecular basis of these actions (Barnstable & Tombran-Tink, Prog Ret Eye Res. 23: 561, 2004). We are developing novel therapeutics based on PEDF and are testing their efficacy in animal models of retinal and neurological disease.

The hallmark of glaucoma is death of retinal ganglion cells. We have used a preparation of isolated and purified ganglion cells to study death by excitotoxins such as glutamate and the effects of other molecules, such as arachidonic acid, on glutamate induced cell death. This cell preparation is also being used to test the efficacy of a variety of neuroprotective agents.

The range of causes of the cone photoreceptor loss in Macular Degeneration is not yet known. We recently participated in the study that identified complement Factor H as a major factor in causing this retinal degeneration. Factor H is an important negative regulator of the complement cascade and reduced function might represent excessive activation of the inflammatory system.