Rho GTPase-mediated cytoskeletal organization in Schlemm's canal cells play a critical role in the regulation of aqueous humor outflow facility

The increased intraocular pressure (IOP) has been considered to be an increased resistance of the aqueous humor outflow through the inner wall of Schlemm's canal (SC) and/or the juxtacanalicular tissue (JCT). The Rho GTPase-regulated actomyosin organization appears to be an important mechanistic determinant of aqueous humor outflow facility. Therefore, in this study, we have evaluated the effects of modulating Rho GTPase activity on actomyosin cytoskeletal organization, monolayer permeability/barrier function of human SC cells, and aqueous humor outflow facility in enucleated porcine eyes ex vivo. Human SC cells, isolated from cadaver eyes, were treated with either Rho GTPase activators such as thrombin and lysophosphatidic acid (LPA), or a specific inhibitor (C3-exoenzyme) of Rho GTPases. Treatment of SC cells with thrombin and LPA led to increased formation of stress fibers, focal adhesion, and increased myosin light chain phosphorylation, whereas treatment with C3-exoenzyme showed the opposite effects like H-7 and ECA, known for increasing the outflow facility in porcine eyes. The findings presented here suggest that LPA and thrombin, presumably through activation of Rho GTPase-mediated actomyosin cytoskeletal reorganization in SC cells, cause a decrease in monolayer permeability of SC cells as well as a decrease in outflow facility of porcine eyes in ex vivo. Our results suggest that decrease in aqueous humor outflow may be correlated better with the changes in cytoskeletal organizations of SC, which could be the prime locus of the outflow resistance.