Rupture of atherosclerotic lesions is a major trigger for myocardial infarction and sudden death. One important mechanism is likely to be activation of matrix metalloproteinases (MMPs), which destroy matrix components in vitro and in atherosclerotic models in vivo. MMPs also play a critical role in regulating cell migration, the recruitment of inflammatory cells, and the immune response. Multiple lines of evidence suggest that oxidants produced by the phagocyte NADPH oxidase and reactive nitrogen species both activate and inactivate MMPs in vitro. These observations indicate that oxidation may regulate the progression of atherosclerotic lesions by a variety of mechanisms. We have shown that oxidants produced by macrophages markedly activate pro-MMP-7 by converting the thiol residue of the cysteine switch to sulfinic acid. This activation mechanism is distinct from the well studied proteolytic cleavage of MMP pro-enzymes. Failure to inhibit MMP activity might also play a role in tissue destruction under inflammatory conditions. We have shown that higher concentrations of oxidants inhibit the activity of human MMP-7 in vitro, suggesting that it might limit proteolytic activity during inflammation. Our findings suggest that local, pericellular production of oxidants by phagocytes is a physiological mechanism for restraining MMP activity during inflammation. We propose to test the hypothesis that oxidants generated by macrophages in the artery wall play a critical role in regulating MMPs, matrix degradation, and lesion progression during the pathogenesis of atherosclerosis.
Our specific aims are: First, to establish the molecular mechanisms by which the NADPH oxidase pathway, the myeloperoxidase pathway and the reactive nitrogen pathway both activate and inactivate MMPs in vitro. Second, to use macrophages isolated from mice deficient in MMPs or oxidant generating pathways to investigate the mechanisms for regulation of proteolysis during atherogenesis. Third, to investigate cellular mechanisms for regulating proteolysis in the artery wall, using mice deficient in pathways that generate oxidants. Fourth, to determine whether human atherosclerotic tissue contain oxidized prodomains of MMPs implicated in proteolytic activation. Collectively, the proposed experiments will address the role of macrophage-derived oxidants in regulating MMP activity during atherogenesis. ? ?
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