Abstract 926: Regulation of Fibrosis by Phospholipase Cεin Cardiac Fibroblasts

Abstract

Cardiac fibrosis is commonly associated with heart diseases, such as cardiac hypertrophy and myocardial infarction. Upon injury to the heart, cardiac fibroblasts transition to an activated myofibroblast state, driven by TGFβ or angiotensin II (AngII) signaling, characterized by expression of α-smooth muscle actin (αSMA). Fibrosis is driven by the activity of both cardiac myofibroblasts and inflammatory cells. Activation of cardiac fibroblasts leads to the induction of pro-inflammatory cytokines that are important for recruitment of immune cells and regulation of cardiac remodeling. Monocyte Chemoattractant Protein (MCP)-1/CCL2 is a chemokine expressed by cardiac myofibroblasts through activation of Nuclear Factor-kB (NF-kB) that leads to recruitment of mononuclear cells. Phospholipase Cε (PLCε) is highly expressed in cardiac fibroblasts and is activated by multiple upstream stimuli including GPCRs and RTKs, leading to protein kinase C (PKC) and D (PKD) activation and Calcium release. In this study to elucidate the role of PLCε in neonatal rat cardiac fibroblasts adenovirus encoding for a PLCε-shRNA was used to knockdown PLCε in fibroblasts. Treatment with PLCε-shRNA strongly inhibited the transition to myofibroblasts induced by TGFβ as assessed by staining of αSMA. PLCε knockdown did not inhibit the ability of TGFβ to induce SMAD nuclear translocation. PLCε depletion also strongly inhibited, basal, Thrombin, and AngII mediated PLC activity in cardiac fibroblasts, indicating PLCε is a dominant regulator of phosphoinositide hydrolysis in these cells. PLCε shRNA also decreased expression of MCP-1 mRNA and reduced the ability of pro-fibrotic stimuli to induce MCP-1 mRNA expression. Knockdown of PLCε or inhibition of PKD with the small molecule inhibitor NB 142-70 reduced the ability of thrombin to activate NF-kB. These data indicate that PLC signaling, and PLCε in particular, plays an important role in pro-fibrotic processes in cardiac fibroblasts, through both cell autonomous and paracrine mechanisms. Future studies will focus on further elucidating the mechanistic role of PLCε in the transition to myofibroblasts, and will study in in vivo models of cardiac fibrosis.