Microtubule disruption leads to active cellular contraction in human trabecular meshwork cells

PURPOSE: We have observed pharmacologically produced, cytoskeleton-related changes in cell shape and cell to cell attachment in cultured trabecular and other endothelial cells that have correlated with aqueous outflow (AO) effects in vitro and in vivo. These changes could be produced simply by some form of cellular reorganization or alternatively by active cellular contraction (CC). We wished to specifically study the role of CC in these events and to test the tensegrity hypothesis that microtubule (MT) disruption leads to active CC. METHODS: Human trabecular meshwork (HTM) cells were plated in culture dishes containing a polymerized deformable silicone substrate (SS). After 48 hours the dish was placed on an inverted microscope and treated with ethacrynic acid (ECA) or colchicine (COL). and then recorded on video for 15 minutes. An increase in wrinkle size and/or number of the SS indicated a contraction. Sham controls were performed. RESULTS: We observed ECA-induced CC at dosages between 10 and 100 uM. Above this dosage HTM cells were observed to detach from the SS. We observed COL-induced CC at dosages between 5 and 100 uM. For both agents the time of onset of CC correlated with induced MT changes. CONCLUSION: ECA and COL cause active CC in HTM cells and this effect can be explained by the induced loss of MT rigidity (outward force) that would be predicted by a tensegrity hypothesis. Tensegrity mechanisms through effects on trabecular cell shape could be involved in the normal regulation of AO and this offers the potential for novel outflow therapies. The CC assay could serve as a screening tool for future glaucoma medications.