Effect of H-7 on cultured human trabecular meshwork cells

PURPOSE: To determine the effect of the serine-threonine kinase inhibitor H -7, which blocks actomyosin contractility and increases outflow facility in live monkeys, on morphology, cytoskeleton, and cellular adhesions of human trabecular meshwork (HTM) cells in culture. METHODS: Cultured HTM cells were videographically recorded and evaluated be fore and after exposure to H-7 at different concentrations. The subcellular distribution of the actin-based cytoskeleton and associated anchor protein s including vinculin, paxillin, and beta -catenin, as well as phosphotyrosi ne-containing proteins were evaluated by fluorescence immunocytochemistry a nd digital fluorescence microscopy. RESULTS: H-7 induced pronounced but reversible HTM cell thickening toward t he cell center and deterioration of the actin cytoskeletal network. Cell-ex tracellular matrix (ECM) and cell-cell adhesions were also affected, but th e beta -catenin-rich, vinculin-containing adherens junctions were clearly m ore resistant than focal contacts. Phosphotyrosine labeling in focal contac ts was highly sensitive to H-7. CONCLUSIONS: H-7 induces alterations in cell shape, actin cytoskeleton, and associated focal adhesions in cultured HTM cells, which may be responsible for the effects of H-7 on outflow facility in live monkey eyes.