This application is in response to the recently announced availability of Recovery Act Funds for Competitive Revision Applications (NOT-OD-09-058). It will create two new jobs in our laboratory and help to retain several jobs at the Johns Hopkins University School of Medicine ES Cell Targeting Core Laboratory, as some of the studies proposed in this revised application will be done at the core facility as a paid service. We previously described a naturally occurring mutation (Nuc1) in the Sprague-Dawley rat with a novel eye phenotype involving cataract, retention of fetal vasculature, and developmental abnormalities in the retina. We have recently reported that the Nuc1 phenotype in which the development of both the lens and retina is abnormal results from mutation of the ?A3/A1-crystallin gene. ?A3/A1-crystallin is expressed in lens fiber cells;our recent studies show that in neural retina it is expressed only in astrocytes. Persistent fetal vasculature (PFV) is a human disease that results from failure of the fetal vasculature to regress. It is a common congenital developmental disorder of the eye found in an otherwise normal child. The underlying cause of PFV disease is not well understood. The Nuc1 spontaneous mutant rat is the only model that accurately resembles all the clinical symptoms of human PFV disease. In the present competitive supplement, we propose to further characterize our recently established ?A3/A1-crystallin transgenic mouse lines and to generate conditional knockout mice where ?A3/A1-crystallin will be selectively deleted in astrocytes, or the lens. These genetically engineered mouse lines will provide additional tools for gain and loss-of-function studies and will generate more definitive data on the effects of eliminating or modulating the expression of ?A3/A1-crystallin, in the etiology of PFV disease. The studies proposed in this revision application are based on progress made to date under the parent grant and are specifically designed to augment the experiments outlined in the Specific Aims of that grant. Our working hypothesis is that """"""""mutation of ?A3/A1- crystallin causes abnormal association of astrocytes with the hyaloid artery, which inhibits regression of the fetal vasculature"""""""". To test this hypothesis, the following specific aims were proposed in the parent grant:
SPECIFIC AIM 1 : To characterize and compare ?A3/A1-crystallin expression in wildtype and in Nuc1 homozygous rats during lens fiber cell and astrocyte development.
SPECIFIC AIM 2 : To investigate if altered motility of Nuc1 homozygous astrocytes during development contributes to the abnormal association between astrocytes and the hyaloid vasculature.
SPECIFIC AIM 3 : To determine if VEGF produced by astrocytes expressing mutant ?A3/A1-crystallin mediates the survival and stabilization of the hyaloid vasculature in the Nuc1 rat. We believe that the proposed studies should provide new insights into the cellular and molecular interactions that regulate hyaloid vascular regression. The possibility that ?A3/A1-crystallin may have a role in hyaloid vascular regression is important;it may help elucidate mechanisms underlying PFV that would have potential clinical implications.
Persistent Fetal Vasculature (PFV) is a common potentially blinding, congenital eye disease. Our data (published and unpublished) indicate that ?A3/A1- crystallin plays a role in PFV. The proposed studies may help elucidate mechanisms that lead to PFV and would have potential therapeutic implications.
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