Proliferation and Ploidy During Cardiac Differentiation of Human Induced Pluripotent Stem Cells

Abstract

Cardiomyocytes are a highly specialized cell type whose primary function is uninterrupted, rhythmic contraction. Despite the vital nature of the heart, cardiomyocytes do not readily proliferate, and consequently the heart is unable to sufficiently regenerate in response to acute insult such as myocardial infarction, or in response to chronic stressors such as hypertension. Thus there is great interest in the technologies that could stimulate cardiac regeneration and similarly great effort being expended to develop myocardial replacement therapies. Central to both of these endeavors is the need for a detailed understanding of cardiomyocyte cell cycle regulation; those factors stimulating proliferation and those inhibiting proliferative potential Chapter I reviews the current understanding of our knowledge of cardiomyocyte cell cycle regulation, proliferation, ploidy and regenerative potential in animal models and in humans. Chapter II describes the materials and methods used throughout this thesis. Chapter III examines the emergence of cardiomyocytes during the in vitro cardiac differentiation of human induced pluripotent cells, their replicative capacity, cell cycle distribution and gene expression profiles relating to these traits and attempts to align this data with data from in vivo models of murine cardiogenesis. In Chapter IV we demonstrate a flow cytometry based method for the identification of mono- and bi-nucleated cardiomyocytes. In Chapter V we further investigate ploidy and cell cycle arrest in response to endogenous and exogenous stress. Chapter VI summarizes the results presented in this thesis and proposes further studies that would expand our knowledge of the roles and regulation of cardiomyocyte ploidy