Structural basement membrane components and corresponding integrins in Schlemm's canal endothelia

PURPOSE The conventional outflow pathway provides the primary source of resistance to aqueous humor drainage, regulating intraocular pressure. Despite large pressure gradients across the inner wall of Schlemm's canal (SC), cells remain attached to their basement membrane. The goal of this study was to examine integrin-extracellular matrix binding partners of the inner wall basement membrane that facilitate attachment. METHODS Human outflow tissues and cultured cells were analyzed by immunofluorescence and western blotting, respectively. Radial sections of human donor eyes or en face preparations of human SC inner wall were probed with antibodies that specifically recognize collagens (Type I, III, and IV), laminins (LM-332 and LM-511) and laminin-specific integrin subunits, α3, α6, β1, and β4, typical of vascular endothelia. RESULTS Immunofluorescence studies showed collagens Type I and IV in the SC basement membrane but not collagen III. As expected with mature vascular endothelia, SC cells in situ expressed LM-511 but not LM-332. Significantly, the integrin α6 subunit was expressed uniquely by SC. En face labeling of the inner wall displayed integrin α6 colocalizing with LM α5 at the cell periphery. Western blots of cultured human SC endothelial cell monolayers confirmed expression of Type I collagen, collagen IV, LM-511, and the α6 integrin subunit. Interestingly, LM-332 was present in cultured SC cells up to 60 days post-confluence. CONCLUSIONS Even though cells of the inner wall endure pressure gradients in the basal to apical direction, opposite of other endothelia, human SC cells express basement membrane proteins and their cognate integrins typical of vascular endothelia.