Molecular Analysis of the Human Aqueous Outflow Pathway

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The goal of our research program is to identify the proteins involved in aqueous outflow resistance, and characterize whether proteins are altered in the glaucomatouseye. Aqueous outflow resistance can be affected by the proteins in aqueous humor and the proteins and extracellular matrix produced by trabecular cells. Because no animal model of primary open angle glaucoma (POAG) exists, studies of the human aqueous outflow pathway over the past 30 years have used cell and tissue culture techniques to analyze the trabecular meshwork. These techniques use media supplements (serum, etc) that differ from the meshwork's normal aqueous humor environment.Our studies find that current culture conditions alter trabecular meshwork protein expression (in both cell and tissue) when compared with meshworks cultured in aqueous humor. Trabecular meshworks cultured in aqueous humor maintain a protein profile that more closely resembles fresh, non-cultured meshworks. Wehypothesize that proteinsfound in aqueous humor are required to maintain trabecular cells and meshworks near their normal physiologic state. Studies of cells and meshworks not cultured in aqueous humor result in findings that may not be physiologicallyrelevant to understanding the in vivo situation. We further hypothesize that changes in the types and proportions of aqueous proteins (growth factors, cytokines, etc.) can alter trabecular homeostasis resulting in abnormal aqueous outflow resistance and development of glaucoma. The current proposal will investigate our hypothesis in the following manner: ¿ Characterize the effect of aqueous humor on molecular and cellular activities in human trabecular meshwork culture models and determine which aqueous proteins are important for trabecular cell homeostasis. ¿ Create a comprehensive profile of proteins found in aqueous humor obtained from normal and POAG patients. We will also determine gene and protein expression changes in the meshwork induced by normal and POAG aqueous humor. ¿ Develop a medium that can be used to mimic aqueous humor for trabecular meshwork cell culture.