Abstract P288: ATP Signaling Stimulates Fibroblast Activation: The Role of Pannexin1 Channels in Fibrosis
BACKGROUND-We have shown that Pannexin1 (Panx1), a novel ATP channel, is found in ventricular cardiac myocytes and upregulated on myocyte sarcolemmal membranes during acute ischemia. We hypothesize that the channel is part of a protein complex that plays a role in downstream cell signaling post myocardial infarction (MI).
METHODS-Using a canine model of MI, where the left anterior descending coronary artery is occluded (CO) we examined epicardial tissue from hearts subjected to 0–3 hrs of CO. Immunoprecipitation (IP) with subsequent silver stain studies revealed novel binding partners for Panx1. Co-immunoprecipitation studies (Co-IP) were performed to confirm Panx1 binding partners. In vitro dye uptake studies and ATP assays were used to determine the function of Panx1 channels under hypoxic conditions.
RESULTS-IP followed by silver staining revealed multiple potential binding partners for Panx1 in the heart. Following 3 hrs CO some interactions increased while others decreased. Co-immunoprecipitation identified one of these partners for Panx1 as the MAGUK scaffolding protein SAP97, an interaction that is significantly increased following 3 hr CO (n=3, p<0.05). Immunostaining showed that CO increased trafficking and stabilization of Panx1 channels at the cell membranes. In vitro dye uptake studies show that cells increase dye uptake following hypoxia, an event that is blocked by the Panx1 channel blocker, Probenecid. Hypoxia also causes increased ATP release likely due to opening of Panx1 channels. ATP release activated fibroblasts and stimulated early fibrosis signaling.
CONCLUSIONS-Our data show the presence of a Panx1 protein complex in cardiac myocytes that is scaffolded at the cell membranes by SAP97. Under cellular stress Panx1 channels release ATP to induce early fibrotic events. We We conclude that Panx1 plays a key functional role in ATP signaling and formation of fibrosis in the post-MI heart.
- © 2011 by American Heart Association, Inc.