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R.K. Sanka, D.L. Epstein, P.V. Rao; Actin Depolymerizing Agents Induce Activation of MMP–2 in Trabecular Meshwork Cells . Invest. Ophthalmol. Vis. Sci. 2005;46(13):1352.
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Purpose: We hypothesize that actin cytoskeletal reorganization and changes in cell morphology modulate the secretion and activation of matrix metalloproteinases (MMPs), which in turn, may influence aqueous humor outflow facility. The goal of this study was to examine the relationship between changes in the actin cytoskeletal organization and matrix metalloproteinase–2 (MMP–2) activity in trabecular meshwork (TM) cells. Methods: Activation of MMP–2 was evaluated by gelatin zymography and Western Blot analysis of serum–free culture medium conditioned by porcine TM cells treated with latrunculin A, cytochalasin D, ethacrynic acid or inhibitors of Rho kinase and protein kinase C. Changes in the protein levels of TIMP–2 and MT1–MMP were determined in porcine TM cell cultures by Western Blot analysis. The effects of actin depolymerizing agents on the expression of MT1–MMP was also determined in human TM cells by semiquantitative RT–PCR analysis. Changes in cell morphology and actin cytoskeletal organization were evaluated by staining with rhodamine–phalloidin and phase contrast microscopy, respectively. Results: Treatment of porcine TM cells with ethacrynic acid (100 µM), cytochalasin D (25 µM), and latrunculin A (0.25 µM) led to an activation of MMP–2 that appeared to correlate well with changes in cell morphology and actin cytoskeletal reorganization. Protein levels of TIMP–2 did not appear to be altered under these conditions. Treatment of human TM cells with latrunculin A (0.25 uM) led to an upregulation of MT1–MMP expression, while treatment with Y–27632 (25 uM) resulted in a downregulation of MT1–MMP expression in comparison to controls. Conclusions: These data demonstrate that actin depolymerizing agents such as latrunculin A, ethacrynic acid, and cytochalasin D increase activation of MMP–2 in TM cells implying the existence of a relationship between the integrity of actomyosin organization and ECM turnover in TM cells. Therefore, cytoskeletal–acting drugs may possibly influence aqueous humor outflow facility through multiple potential mechanisms.
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