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A. I. Kupperman, P. K. Russ, S. Zhang, F. R. Haselton, M. S. Chang; Modulation of Trabecular Meshwork Permeability by Bves. Invest. Ophthalmol. Vis. Sci. 2008;49(13):1619.
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Aqueous outflow in the trabecular meshwork (TM) is altered by tight junctions (TJ) and cellular contraction. TJ formation and cellular contraction lead to decreased outflow. We have previously shown that Blood Vessel Epicardial Substance (Bves) increases TJ formation and decreases Rho activation in epithelial cells. Therefore, we hypothesize that Bves in TM cells plays a role in regulating aqueous outflow by modulating both TJ formation and cellular contraction, a downstream target of activated Rho.
TM cell lines from normal (NTM) and glaucomatous (GTM) donors, as well as NTM cells stably transfected to overexpress Bves (NT-w), were evaluated by Western blots to determine levels of TJ protein ZO-1 and phosphorylated myosin light chain (pMLC). pMLC is an indicator of cellular contraction that is a downstream target of Rho. Immunofluorescent (IF) localization of Bves and ZO-1 was also performed. In addition, diffusion of sodium fluorescein in a two-chamber assay was measured to assess paracellular permeability for all cell lines.
Western blots demonstrated that GTM cells had increased Bves and ZO-1 levels compared to NTM cells. Bves and ZO-1 co-localized to the cell border as shown by IF. Functionally, GTM cells showed decreased permeability compared to NTM cells in the two-chamber assay. NT-w cells (stably overexpressing Bves) also exhibited an increase in ZO-1 levels and a decrease in permeability similar to GTM cells. These findings indicate that increased Bves correlates with increased TJs and decreased TM permeability. Because increased TJ formation is also associated with decreased Rho activation, we measured pMLC to determine Bves’ effect on cellular contraction. NT-w cells showed lower levels of pMLC compared to NTM cells, consistent with decreased Rho activation and cellular contraction. GTM cells, however, showed pMLC levels similar to NTM cells, suggesting that the diseased GTM cells have an alteration in the Rho pathway.
These studies indicate that Bves is an important modulator of TM TJ formation and cellular contraction. Increased levels of Bves lead to increased TJs and decreased pMLC. Others have previously demonstrated that increased TJs lead to decreased flow and decreased pMLC leads to increased flow. Our findings suggest that Bves is an important homeostatic regulator of aqueous flow through its dual, opposing effects on outflow, and that alterations in this pathway may lead to dysregulation of intraocular pressure.
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