Function of Glycocalyx in the Trabecular Outflow Pathway

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Primary open-angle glaucoma (POAG) is a leading cause of blindness worldwide. The most common risk factor for the development and progression of POAG is elevated intraocular pressure, resulting from an increase in aqueous humor outflow resistance. Most outflow resistance is believed to be generated in the juxtacanalicular connective tissue and modulated by the inner wall endothelium of Schlemm's canal (SC), and its pores. Studies also suggest that one-third to one-half of the total outflow resistance is located distal to SC. The mechanisms that regulate the aqueous humor outflow resistance in each of these regions remain unknown in normal and POAG eyes. We propose that a complex of glycoproteins assembled as a glycocalyx may contribute to regulating the aqueous humor outflow resistance. Glycocalyx is composed of glycoproteins, proteoglycans, and glycosaminoglycan chains that are filamentous structures covering endothelial cells of blood vessels. We have recently demonstrated that there is a similar filamentous glycocalyx layer covering endothelial cells throughout the trabecular outflow pathway. More importantly, glycocalyx actually fills 80% of the pores found in the endothelium of SC when stained with Alcian Blue. While the function of the glycocalyx in the endothelial cells of the trabecular outflow pathway is unknown, in the vascular endothelium it plays a role in: 1) mediating the alignment and elongation of endothelial cells in the direction of blood flow, 2) acting as a mechanotransducer of fluid shear stress, leading to activation of endothelial nitric oxide synthase (eNOS), release of nitric oxide (NO), and vessel dilation. In disease states, glycocalyx is lost or damaged, which causes endothelial cells to lose their ability to respond to fluid shear stress. This in turn decreases NO release, preventing vessel dilation, and increasing permeability of vascular and lymphatic vessels. These findings in vascular endothelium prompt us to elucidate the function of glycocalyx in the trabecular outflow pathway. We will test our hypothesis that glycocalyx contributes to the regulation of aqueous humor outflow resistance by attenuating flow through filling the pores in SC endothelium, and dilating SC and distal scleral vessels through mechanotransduction-induced release of NO, similar to vascular endothelium. Our specific aims are: 1) to investigate the differences in distribution and height of glycocalyx and percentage of the pores filled with glycocalyx in high-flow vs. non-flow areas of normal bovine eyes, and the changes in glycocalyx in laser-induced ocular hypertensive monkey eyes by comparison with normal monkey eyes; 2) to investigate the functions of glycocalyx by measuring outflow facility and eNOS activity in bovine eyes after enzymatic removal of glycocalyx or perfusion with eNOS blocker. The results of the proposed study will provide an initial understanding of the role of glycocalyx in regulating the aqueous humor outflow resistance of normal eyes and changes in laser-induced ocular hypertensive monkey model, and a potential for developing new therapeutic strategies to treat POAG.