Expression of aquaporin-1 in human trabecular meshwork cells: Role in resting cell volume

PURPOSE. Drainage of aqueous humor from the human eye appears dependent on intracellular volume of trabecular meshwork (TM) cells, the predominant cel l type of the human outflow pathway. Thus, the modulation of water and solu te flux across the plasma membrane of TM cells is predicted to be an import ant factor in regulating outflow facility. Aquaporin (AQP)-1 is a hexahelic al integral membrane protein that functions as a regulated channel for wate r and cations in fluid-secreting and -absorbing tissues. AQP1 is present in many tissues of the human eye, including the TM; however, its role in outf low facility is unknown. The purpose of the present study was twofold: to e valuate the prospect of manipulating AQP1 protein levels in TM cells using sense and antisense mRNA and to investigate the functional role of AQP1 in TM cells. METHODS. An adenovirus (AV) expression system was used to alter AQP1 protei n levels. AQP1 protein expression was monitored using immunoblot analysis, and resting cell volume was measured by forward light scatter, electronic c ell sizing, and [C-14]-sucrose/urea equilibration. Permeability of TM monol ayers to [C-14]-sucrose was also assessed as an indirect evaluation of cell volume. RESULTS. AV-mediated gene transfer of AQP1 cDNA to TM cells resulted in a t iter-dependent increase in recombinant AQP1, whereas transfer of antisense cDNA decreased native AQP1 protein by 71.7% +/- 5.5% (P < 0.01) after 5 day s. A novel finding of this study is that mean resting volumes of AQP1(s) AV -infected TM cells in suspension were 8.7% +/- 3.0% greater (P < 0.05) than control cells. Conversely, AQP1 antisense (as) AV-infected cells had resti ng volumes 7.8% +/- 2.9% less than control cells (P < 0.05). Similar effect s of AQP1 expression on resting cell volume were observed in TM monolayers. Consistent with this finding, paracellular permeability of AQP1(s) AV-infe cted TM monolayers to [C-14]-sucrose decreased by 8.0% +/- 1.4% (p < 0.001) . CONCLUSIONS. In addition to influencing the osmotic permeability of TM plas ma membranes, the level of AQP1 protein expression influences resting intra cellular volume and thus paracellular permeability of TM cell monolayers in vitro. These data suggest that AQP1 expression may affect outflow facility in vivo.