Abstract 12519: Regulation of Cardiac Fibroblast Phenotype by Scleraxis
Cardiac fibroblasts constitute the primary extracellular matrix synthesis machinery in the myocardium. Activation of fibroblasts into a hyper-synthetic and contractile phenotype potentiates fibrosis, impairs cardiac function and contributes to heart failure. Our laboratory previously reported that the transcription factor scleraxis regulates human cardiac collagen Iα2 expression and has shown its up-regulation in the post-infarct scar. Here we demonstrate a novel regulatory role for scleraxis in governing cardiac fibroblast function and phenoconversion. Cell contractility assays using collagen gels demonstrated the abrogation of pro-fibrotic TGF-β-mediated contractility of myofibroblasts in response to scleraxis knockdown. The de novo expression of α-smooth muscle actin (αSMA) and its incorporation into stress fibers is a key feature of myofibroblasts - key causative cells of fibrosis. Scleraxis over-expression in isolated primary cardiac fibroblasts induced αSMA gene expression and stress fiber formation, and rescued the αSMA loss observed in cardiac fibroblasts from scleraxis null mice. Luciferase reporter assays demonstrated a significant transactivation of the αSMA gene promoter by scleraxis. Mutation analysis revealed that scleraxis interacts with two E-boxes within the αSMA promoter, a finding confirmed by chromatin immunoprecipitation of scleraxis in primary cardiac fibroblasts. An increase in scleraxis binding to the αSMA promoter was observed in cardiac myofibroblasts compared to fibroblasts, and also in response to TGF-β, further supporting a direct role of scleraxis in regulation of myofibroblast αSMA expression and its contractile phenotype. Gel shift assays also confirmed the direct interaction of scleraxis with E-boxes within the αSMA gene promoter. Our data indicates that scleraxis plays a required role in cardiac fibroblast phenotype and contractile function. Taken in context with our finding that scleraxis regulates expression of multiple extracellular matrix components, including fibrillar collagens, our data reveals that scleraxis exerts broad and potent pro-fibrotic effects on cardiac fibroblast form and function, and may thus represent a novel target for fibrosis therapy.
Author Disclosures: R. Bagchi: None. P. Roche: None. R. Schweitzer: None. M.P. Czubryt: None.
- © 2014 by American Heart Association, Inc.