Program Director/Principal Investigator (Last, First, Middle): Tyagi, Suresh C., The overall goal of this project is to understand the mechanism of endocardial endothelial-myocyte (E-M) dysfunction in chronic heart failure. Studies from the previous funding period suggested that endocardial endothelial dysfunction is associated with increased oxidized-matrix accumulation (fibrosis), activation of latent resident myocardial matrix metalloproteinases (MMPs) and inactivation of cardiac tissue inhibitor of metalloproteinase (TIMP-4) secondary to oxidative and proteolytic stresses. Administration of TIMP-4 ameliorated both the formation of reactive oxygen species (ROS, oxidative stress) and MMP activation (proteolytic stress). In addition, we discovered the induction of proteinase activated receptor-1 (PAR-1). However, the role of PAR-1 in fibrosis and E-M uncoupling remains poorly defined. H2S gas is the most potent antioxidant in mitigating oxidative stress and recent studies have implicated a cardioprotective role of H2S. The central hypothesis of this competitive renewal proposal is that during chronic heart failure the oxidative and proteolytic stresses induce PAR-1, leading to generate mitochondrial (mt) ROS and reactive nitrogen species (RNS) and mitochondrial nitric oxide synthase (mtNOS), respectively, thus activating the latent resident cardiac MMPs. These events disrupt the MMP/TIMP axis, causing fibrosis between endothelium and myocyte. Treatment with H2S alleviates fibrosis and mitigates E-M uncoupling. Therefore, the specific aims of this proposal are: #1: To determine whether chronic left ventricle (LV) volume overload causes mitochondrial oxidative stress (ROS and RNS) by inducing NADPH oxidase (p47 subunit), mtNOS and PAR-1, and H2S alleviates mitochondrial oxidative stress. #2: To determine whether chronic LV volume overload causes cardiac fibrosis by increasing collagen/elastin ratio, MMP-2, -9, -13, TIMP-1, -3, decreasing TIMP-4, and inducing PAR-1, and H2S mitigates cardiac fibrosis. #3: To determine whether chronic LV volume overload causes E-M dysfunction and LVH by inducing PAR-1 and H2S decreases E-M uncoupling. Chronic heart failure will be created by LV volume overload by aorta-venacava fistula (AVF) in wild type (WT), PAR-1-/+, iNOS-/-, MMP-9-/-, TIMP-3-/-, and TIMP-4++/++ mice, treated with or without NaHS, a H2S donor. PHS 398/2590 (Rev. 11/07) Page Continuation Format Page