Abstract 17995: Inhibition of Myofibroblast Differentiation by FOXO3a - Implications for Acute Myocardial Infarction and Cardiac Remodeling

Introduction: Transdifferentiation of cardiac fibroblasts into myofibroblasts regulated by TGFβ/SMAD3 signaling is a major mechanism of scar formation and adverse remodeling following myocardial infarction. Hypothesis: We hypothesized that the transcription factor FOXO3a, a key regulator of cell differentiation, cycle and size, might inhibit this process. Methods: Acute myocardial infarction was induced in FOXO3-/- and WT mice (FVB) by permanent LAD ligation and myofibroblast transdifferentiation markers were assessed. FOXO3a-/- and WT cardiac fibroblasts were investigated in transdifferentiation assays ex vivo. FOXO3a gene transfer was performed with gain of function adenoviral vectors. IP/IF and Western blotting were used to test for a direct interaction between FOXO3a and SMAD3. Results: FOXO3a-/- mice had significantly higher survival rates compared to WT mice due to reduced rates of ventricle perforation. Myocardial expression of alpha smooth muscle actin (ASMA) and Collagen1A1 (Col1A1) was significantly enhanced in FOXO3a-/- mice 3 and 14 days post infarction. Moreover, Foxo3a-/- mice showed larger fibrotic areas following MI. In line with these results, cardiac fibroblasts isolated from FOXO3a-/- mice showed significantly enhanced expression levels of Col1A1 and ASMA 24 hours following stretch or TGF-β stimulation when compared to WT cells in vitro. Furthermore supernatants of FOXO3a -/- fibroblasts showed significantly higher protein expression of Col1A1 in (p<0.01) while FOXO3a gene-transfer dose-dependently downregulated Col1A1. Immunfluorescence staining for ASMA protein was significantly attenuated following FOXO3a gene transfer in cardiac fibroblasts. Mechanistically, immunoprecipitation showed direct interaction of FOXO3a with SMAD3a that was enhanced following activation of the transcription factor leading to diminished SMAD3 downstream gene expression. Conclusions: Our results identify FOXO3a as a direct inhibitor of TGF- β regulated matrix remodeling via FOXO3a-SMAD3 interaction. FOXO3a regulates fibrosis and scar formation following myocardial infarction and thus targeting the FOXO3a-SMAD3 axis might be of therapeutic interest.