Abstract 1039: A Proprotein Convertase Regulates Integrin - MMP Cooperation In VSMCs

ECM remodeling is a key feature in atherosclerosis and restenosis. Integrins link the cytoskeleton with the ECM and regulate adhesion/migration. MMPs are the major degrading enzymes important for cell motility. Binding of the MMP-2 zymogen to alpha v beta 3 is necessary for its full activation, thereby coordinating cell motility with membrane-associated proteolysis. The alpha v integrin chain is synthesized as a proprotein that is endoproteolytically activated by furin. Furin is the prototype proprotein convertase, highly expressed in human atherosclerotic lesions. The aim of this study was to investigate the role of furin in integrin -MMP coordination and cooperation in VSMCs. Methods and Results: Treatment of VSMCs with either the furin inhibitor dec-CMK (50 umol/L) or an MMP-inhibitor (GM6001; 25 umol/L) significantly inhibited VSMC Matrigel invasion (p<0.05 vs. controls). Immunoblotting demonstrated that dec-CMK inhibited alpha v endoproteolytic activation, but this did not affect alpha v membrane expression assessed by FACS analysis. Zymography revealed that in dec-CMK or TSR1265 (10 umol/L; an inhibitor of MMP-2 binding to alpha v beta 3) treated cells, maturation of the intermediate 68 kDa MMP-2 to its fully active 62 kDa form was significantly decreased (p<0.05 vs. controls). Furthermore, dec-CMK significantly inhibited binding of FITC-conjugated pro-MMP-2 to the cell surface of VSMCs. Immunfluorescence and 3D in gel zymography demonstrated that inhibition of endoproteolytic cleavage of the alpha v integrin inhibits actin rearrangement and focal contact formation upon integrin stimulation (matrix adhesion or PMA treatment), as well as membrane-associated proteolysis. Conclusion: Our study demonstrates that endoproteolytic cleavage of alpha v integrin by furin regulates not only integrin activation, but also affects MMP-2 maturation. Endoproteolytic activation of alpha v is required for cytoskeleton rearrangement upon integrin stimulation and focal contact formation, as well as the coordination and cooperation of integrins and MMPs. Therefore furin-convertase may be a novel target in atherosclerosis and restenosis.