Our research focuses on identifying mechanisms and molecules that control the development of the normal lens. This is an essential step towards understanding cataract formation. We have already established that both FGF and TGFbeta have major effects on lens cells in epithelial explants; moreover, TGFbeta causes cataract-like changes. This project represents a detailed investigation of the roles of TGFbeta and FGF in governing events in lens development. There are two main sections: (1) The role of FGF receptors (FGFRs) in the control of lens cell behavior. Specific hypotheses are: (a) That spatiotemporal patterns of FGFRs expression in the lens correlate with known patterns of lens cell behavior and contribute to the regulation of FGF action on lens cells throughout life. The expression patterns of the FGFRs will be studied by in situ hybridization using complementary RNA probes; (b) That FGFR expression is influenced by FGF and other growth factors. Changes in mRNAs for FGFRs will be monitored by in situ hybridization in rat lens epithelial explants during culture with growth factors at concentrations known to induce a variety of cellular responses; (c) That different FGF-induced responses are mediated by different FGFRs. Neutralizing antibodies specific for each of the FGFRs found in the lens will be used to block ligand binding. Effects on various FGF-induced responses will be assessed. (2) The role of TGFbeta in lens biology. Specific hypotheses are: (a) That increasing the concentration of active TGFbeta in the vitreous induces cataract-like changes in the lens in vivo. Anesthetized rats will be microinjected with TGFbeta and lenses will be monitored for opacities and morphological and molecular and changes; (b) That expression of TGFbeta occurs in lens cells throughout life. The expression patterns of the TGFbetas will be studied by immunohistochemistry and in situ hybridization; (c) That FGF protects lens cells from TGFbeta-induced cataractous changes. Explants will be cultured with various combinations of TGFbeta and FGF and monitored using molecular markers for cataract and normal lens phenotype. (d) That inhibitors of TGFbeta are present in the lens or ocular media. Fractions will be prepared by chromatography and tested for their ability to inhibit TGFbeta-induced changes in explants; (e) That TGFbeta and FGF have different effects on cell-cell and cell-substratum interactions. Explants will be cultured with TGFbeta or FGF and immunohisto-chemistry and electron microscopy will be used to monitor changes in cytoskeletal and cell surface markers. These studies will provide crucial information about the control of normal lens growth and development and should provide insights into the aetiology of major forms of cataract.
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