Abstract 441: Role of Nitric Oxide and S-nitrosylation in Cardiomyogenesis by iPSCs

Abstract

Introduction: The mechanism by which signaling pathways, such as Wnt and BMP interact and modulate each other’s function is crucial to our understanding of cardiomyogenesis and cardiomyocyte proliferation. Nitric oxide (NO) is a signaling molecule that can trigger cardiac differentiation of stem cells, suggesting a cardiogenic function of NO synthase(s) (NOS). Hypothesis: NO modulates transcription factor function during pluripotency and differentiation toward a cardiac phenotype. Methods: Induced pluripotent stem cells (iPSCs) were derived from fibroblasts from wildtype mice and mice lacking S-nitrosoglutathione reductase (GSNOR -/- ), a denitrosylase that regulates protein S-nitrosylation. iPSCs were differentiated into functional cardiomyocytes from embryoid bodies (EBs) via the hanging-drop method. Results: During differentiation into cardiomyocytes, GSNOR -/- iPSC-derived cardiomyocytes exhibited reduced expression of mesoderm induction-related ( Brachyury ), cardiac mesoderm (Kdr , Isl-1 ) and cardiac progenitor genes ( Nkx2.5 , GATA4 ). Axin-1, an inducer of apoptosis and negative regulator of the Wnt signaling pathway and MAPK pathways, specifically p38, were increased on EB-Day (D)4. In contrast, SMAD1/5/8, members of the BMP canonical signaling pathway, were reduced beginning on EB-D8. Increased p38 is associated with reduced GATA4 expression and differentiation of human ES cells into cardiomyocytes. Decreased SMAD1/5/8 is likely at least in part responsible for the reduced expression of Nkx2.5. Conclusions: Our findings support that the absence of GSNOR modulates Wnt/β-catenin and BMP signaling pathways during cardiogenesis, resulting in reduced expression of mesoderm, cardiac mesoderm and cardiac progenitor genes. These findings are expected to have important implications for regenerative medicine and can provide new targets for iPS cell-based therapy.