Neuropeptide Y (NPY) induced reentry of differentiated rat neonatal and adult cardiomyocytes into the cell cycle. NPY also induced differentiation of bone marrow derived-mesenchymal stem cells (MSC) into cardiomyocytes following transplantation into infarcted myocardium. In vitro: Rat neonatal and adult cardiomyocytes were treated with vehicle, NPY, fibroblast growth factor (FGF) (100ng/ml), or FGF plus NPY. DNA synthesis, mitosis, and cytokinesis were determined by immunocytochemistry. NPY-induced MSC gene expression, cell migration, tube formation, and endothelial cell differentiation were analyzed. In vivo: Male rat green fluorescent protein (GFP)-MSC (2x10⁶), pretreated with either vehicle or NPY (10^-8 M) for 72 hours, were injected into border zone of the myocardial infarction (MI) in female rats following LAD ligation. On day 30, heart function was assessed, and hearts were harvested for histological and immunochemical analyses. NPY increased BrdU incorporation, and promoted both cytokinesis and mitosis in rat neonatal and adult myocytes. NPY also upregulated several genes required for mitosis in MSC, including aurora B kinase, FGF-2, cycline A2, elF4E, and SDF-1. NPY directly induced neonatal and adult cardiomyocyte cell-cycle reentry and enhanced the number of differentiated cardiomyocytes from MSC in the infarcted myocardium, which corresponded to improve cardiac function, reduce fibrosis, ventricular remodeling, and increase angiomyogenesis. This treatment is a novel approach by promoting MSC-based cardiac repair by NPY.