The heme protein myeloperoxidase (MPO) plays an important role in the host defense response. Myeloperoxidase catalyzes the addition of chloride to hydrogen peroxide, resulting in the formation of the chlorinating species hypochlorous acid (HOCI) Hypochlorous acid reacts avidly with numerous cellular targets including thiols, amines, nucleotides and unsaturated fatty acids. Recent evidence suggests that MPO and HOCI contribute to the pathogenesis of cardiovascular disease. In this application, we present data suggesting that MPO-derived HOCI contributes to the development of endothelial dysfunction by reducing nitric oxide (NO) bioavailability. MPO effectively binds to the arterial wall and is associated with a significant increase in tissue enzymatic activity. Results of in vitro studies showed that MPO-derived HOCI impairs acetylcholine-induced relaxation but does not affect relaxation induced by an NO donor. These data suggest that the endothelium is a target of MPO-dependent injury. HOCI mediates the inhibitory effects of MPO since endothelial dysfunction could be prevented by addition of the HOC scavenger L-methionine. There are several components of the endothelial NO synthetic pathway that may be modified by HOCI including NO synthase (NOS III) activity, L-arginine substrate and NOS Ill-dependent cofactors. In this proposal, we will identify mechanisms underlying the inhibitory effects of MPO-derived HOCI on NO bioavailability by 1) defining the effects of MPO on endothelial NO signaling processes; 2) characterizing the role of L-arginine derived N-chloramines in MPO-dependent endothelial cell injury; and 3) by determining whether inhibitors of MPO prevent endothelial dysfunction in a rodent model of ischemia-reperfusion. The HOCI-dependent modification of L-arginine may represent a common pathogenic mechanism underlying endothelial dysfunction in diverse cardiovascular diseases.