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S. S. VanderWyst, W. D. Stamer; Structural Basement Membrane Components and Corresponding Integrins in Human Schlemm's Canal Endothelia. Invest. Ophthalmol. Vis. Sci. 2009;50(13):2033.
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© ARVO (1962-2015); The Authors (2016-present)
The conventional outflow pathway, including Schlemm’s canal (SC), provides the primary source of resistance to aqueous humor drainage. The structural extracellular matrix proteins (ECM) and their cognate integrins that support SC’s inner wall endothelia remain unclear. The purpose of the present study was to characterize integrin-ECM binding partners of the inner wall basement membrane.
We probed frozen sections of conventional outflow tissues from human donor eyes with antibodies that specifically recognize several isoforms for collagen (Type I, III, IV) and laminin (LM-511, LM-332) often found in vascular endothelia. The most common laminin-integrin subunits, 6/3 and β1 were assessed, as was laminin-specific 6β4. We compared staining in ocular samples with specific control tissues, using confocal microscopy. We also analyzed cultured human SC mature cell monolayers for expression of collagen, laminin and integrins via immunocytochemistry and Western blot.
Our in situ immunofluorescence studies show the presence of Types I and IV collagen, but not collagen III, and LM-511 but not LM-332 in the basement membrane of SC inner wall. While 3 subunits were not detectable, 6β4 was found in SC inner wall basement membrane. Immunohistochemical and Western blot experiments with cultured human SC endothelial cells confirmed the presence of Type I collagen, collagen IV, LM-511 and both 6 and β1 integrin subunits. While collagen III was not expressed by the cultured cells, we were surprised to find that LM-332 was present both in SC and control HUVEC cell lines up to day 21 post-confluence.
Basement membrane structural components of human SC in situ and in vitro include Types I and IV collagen and LM-511, matching human vasculature expression patterns. In addition, SC cells in vitro expressed corresponding integrins, 6β1 and 6β4, analogous to vascular endothelial cells. Together our results show that SC cells in culture provide a viable model to study SC ECM-integrin dynamics.
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