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E.R. Doherty, J. Choi, B.Y. Yue, A.M. Miller, X. Shen, P.A. Knepper; Trabecular Meshwork MT1-MMP Is Up-Regulated in Response to Metabolic Stress . Invest. Ophthalmol. Vis. Sci. 2003;44(13):1158.
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Purpose: Maintenance of the extracellular matrix (ECM) requires a balance between proteinases and their endogenous inhibitors. A major ECM regulator, membrane-type 1 matrix metalloproteinase (MT1-MMP), is activated by the ubiquitous proprotein convertase furin. The activated MT1-MMP activates MMP-2 (gelatinase A) by releasing pro-MMP-2 from tissue inhibitor of metalloproteinase-2 (TIMP-2), and undergoes autocatalytic inactivation. MT1-MMP is also known to cleave the ectodomain of CD44 to form soluble CD44 (sCD44), a protein marker of primary open-angle glaucoma (POAG) in aqueous humor. The purpose of this study was to determine if MT1-MMP protein expression or activation was altered in trabecular meshwork (TM) cells challenged with lactate. Methods: Human TM cells were grown in minimum essential medium Eagle (MEM) containing 10% fetal calf serum (FCS) and 5% calf serum until confluent. The cells were washed twice with PBS and incubated in MEM containing 0.1% FCS with 1mM lactate or PBS for 3 and 6 hours. Western blot analysis of cell lysates was performed using antibodies against MT1-MMP. Results: Western blot analysis revealed a greater intensity of MT1-MMP-positive material in lactate-treated cells than in control cells. In addition, the molecular weight of immunopositive material was observed to decrease from 65 to 60 kDa in lactate-treated TM cells as compared to control cells. Conclusions: Lactate treatment of TM cells resulted in increased MT1-MMP protein expression and activation as indicated by the formation of the 60 kDa immunopositive band. The elevated MT1-MMP activity suggests that increased proteolytic cleavage by MT1-MMP may alter ECM turnover and accounts for the increased sCD44 observed in POAG aqueous humor.
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