DESCRIPTION (provided by applicant): We will continue our studies to define molecular mechanisms related to development of septic cardiomyopathy in rodents after cecal ligation and puncture (CLP), with an emphasis on the roles of C5a anaphylatoxin and its receptors, C5aR and C5L2. Aim 1 will define the requirements for C3, C5, C5a receptors (C5aR, C5L2) and IL-17 for spontaneous release of cardiosuppressive cytokines (IL-12, TNF1, IL-6) from sham and CLP cardiomyocytes (CMs). These data will be compared to the presence of cytokines in heart homogenates and plasma from CLP mice. Aim 2 will determine the time course after CLP for upregulation of receptors for C5a and the cardiosuppressive cytokines (IL-12, TNF1 and IL-6) on CMs. Aim 3 will assess electrophysiological responses (action potentials, Ca2+ transients, and K+ and Ca2+ currents) as well as functional responses (by measuring intracellular calcium transients and myocyte contraction) in CMs obtained from sham and CLP rodents as a function of time after CLP. In addition, sham and CLP CMs will be studied for changes in electrophysiological and contractile responses after in vitro exposure to C5a in an effort to define molecular mechanisms by which C5a causes contractile dysfunction in CMs. Aim 4 will assess the ability of C5a to induce changes in CMs (as described in Aim 3), using sham and CLP CMs to which varying concentrations of C5a or cardiosuppressive cytokines, or both, have been added. We will determine if there is a synergy in development of electrophysiological dysfunction with combination of C5a and cardiosuppressive cytokines. Aim 5 will employ non-invasive echocardiography approaches to assess systolic and diastolic function and systemic vascular resistance, as well as other parameters, in CLP mice as compared to sham mice. We will attempt to define the roles of C3, C5, C5a and IL-17 receptors in CM dysfunction. These studies will also feature CLP mice that have been treated with neutralizing antibodies to C5a or IL-17. A C5aR knockout mouse model will also be utilized to define the role of the C5a signaling pathways in the pathogenesis of septic cardiomyopathy. Collectively, these studies should define the roles of C5a and its receptors as well as cardiosuppressive cytokines in the development of septic cardiomyopathy and how this complication can be averted or treated.

Relevance Sepsis continues to be a daunting problem in humans with a single FDA approved intervention. Depending on location and clinical details, the mortality rate may be as high as 50-60%, an especially alarming statistic in view of 600,000 or more patients with sepsis in North America each year. It is clear that we have incomplete understanding of sepsis. Until this obstacle is resolved through an understanding of the molecular determinants of sepsis, treatment will be supportive and extremely costly.