ITGA8 positive cells in the conventional outflow tissue exhibit Schlemm's canal endothelial cell properties

Elevated intraocular pressure is primarily induced by the increased resistance of conventional outflow of aqueous humor. Dysfunction of the juxtacanalicular region of trabecular meshwork (TM) and Schlemm's canal (SC) endothelium, as the main conventional outflow tissue, have been implicated as the major reasons for the increased resistance. Integrins are widespread in these tissues, especially alpha8 integrin (ITGA8). We aim to investigate the properties of cells expressing ITGA8 in the conventional outflow tissue. Fluorescence in situ hybridization (FISH) and immunofluorescence (IF) were performed to detect the mRNA and protein levels of ITGA8 in human conventional outflow tissue. ITGA8-positive cells were isolated from the cultured human TM cells through a magnetic bead-based approach. Flow Cytometry was used to determine the purification efficiency. The expressions of TM and SC biomarkers and dexamethasone-induced myocilin secretion capacity of ITGA8-positive cells was assessed by Real-time PCR, IF and Western blot. A gel contraction assay was performed to evaluate contractility of ITGA8-positive cells after endothelin 1 treatment. ITGA8 was found with robust expression near the inner wall of SC endothelium. After purification, the proportion of ITGA8-positive cells were increased by about 10%. ITGA8-positive cells were identified with the properties as SC endothelial cells, such as more robust expressions of SC biomarkers, less dexamethasone-inducible myocilin expression, and stronger contractility. This study demonstrated that cells expressing ITGA8 in SC region possess more properties as SC endothelial cells. Our data implicate a crucial role of ITGA8 in aqueous humor (AH) outflow resistance regulation.