The broad objective of this research is to test the hypothesis that, in response to elevated intraocular pressure (IOP), there are specific changes in extracellular matrix (ECM) structure or composition due to biosynthesis or degradation, which underlie the remodeling of the optic nerve head characteristic of glaucomatous optic neuropathy. By comparison with non-glaucomatous optic neuropathies, we plan to differentiate changes of ECM in glaucoma due to elevated intraocular pressure. Once specific, pressure-related glaucomatous changes are identified, we will investigate potential regulatory mechanisms using in vitro models. Optic nerve heads from enucleated human eyes with well-documented glaucomatous or non-glaucomatous optic neuropathies and from normal age- matched controls will be used to pursue the following Specific Aims: 1) To differentiate specific changes in ECM structure, composition and/or distribution due to elevated intraocular pressure, that may contribute to the pathogenesis of glaucomatous optic neuropathies, POAG and secondary glaucomas, from changes in ECM that are common to non- glaucomatous optic neuropathies. 2) To determine whether pressure-related changes in the ECM components of the optic nerve head in glaucomatous optic neuropathies are due to altered gene expression of ECM macromolecules. 3) To determine whether pressure-related changes in ECM in the lamina cribrosa of glaucomatous optic neuropathies are due to degradative enzyme activity such as metalloproteinases and elastases. 4) To characterize the changes in proteoglycans in the ECM of the human optic nerve head that occur in glaucomatous optic neuropathies due to elevated IOP, using cytochemical and immunocytochemical staining, and biochemical quantitation of glycosaminoglycans. 5) To determine which cellular mediators or environmental factors regulate the synthesis, gene expression and degradation of ECM macromolecules that appear altered by elevated IOP, using cultured lamina cribrosa cells and astrocytes. We will use state of the art morphological, cell and molecular biological and biochemical techniques specifically appropriate to analyze small tissue samples such as individual human optic nerve heads. These techniques include: immunocytochemistry at the light and ultrastructural level, electron microscopy, morphometry, in situ hybridization, reverse transcriptase polymerase chain reaction, differential gene expression, zymography, immunoprecipitation, quantitation of glycosaminoglycans, tissue culture, etc. These studies are directly health-related because the work addresses the underlying causes of clinically relevant optic nerve degeneration due to elevated IOP. Furthermore, knowledge of the remodeling of the extracellular matrix due to elevated IOP will provide the cellular link between mechanical and vascular events and the clinical findings of glaucomatous optic neuropathy. The results from these studies may form the basis for explaining individual, age-related, and/or racial differences in the clinical response of the human optic head to elevated IOP. This could lead to improved therapeutic approaches to stabilize the connective tissue of the optic nerve head in certain individuals susceptible to elevated IOP.
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