This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. For nearly 4 decades, the human heart has been considered a post-mitotic organ composed of a predetermined number of myocytes, which is established at the end of gestation. According to this Old Paradigm, the generation of myocytes ceases at birth and their number is preserved throughout life until death of the organ and organism. Cardiac growth postnatally and organ hypertrophy in the adult occur only by myocyte enlargement. On this premise, the age of myocytes corresponds to the age of the organ and organism, i.e., cellular, organ and organism age coincide. Recent results from our laboratory and others have documented that tissue specific stem cells reside in the human heart. Human cardiac stem cells (hCSCs) are self-renewing and multipotent in vitro and in vivo; hCSCs differentiate in myocytes, and vascular smooth muscle cells (SMCs) and endothelial cells (ECs) organized in coronary vessels. The recognition that the human heart possesses a stem cell compartment has imposed a reevaluation of cardiac homeostasis, aging and pathology. The New Paradigm refutes the conviction that myocytes are formed only during embryonic development and suggests that the replacement of coronary vascular SMCs and ECs is regulated by differentiation of hCSCs rather than by the ability of these mature cells to divide. A novel conceptual framework of the heart has emerged; the Heart Is a Self-renewing Organ characterized by a compartment of resident stem cells. This discovery has laid the ground work for the use of hCSCs in the treatment of the failing heart. Currently, two phase I clinical trials are in progress (ClinicalTrials.gov Identifier: NCT00474461; Identifier: NCT00893360). Our understanding of the cellular processes implicated in the maturation, homeostasis and repair of the human heart is extremely deficient and the need for basic information is striking. Findings in nematodes, fruit flies, zebra fish and rodents have often been translated to human beings with little caution, emphasizing the necessity to study the fundamental principles that regulate the plasticity of the myocardium during the lifespan of women and men. Moreover, the mechanisms modulating the response of the female and male heart to ischemic and non-ischemic myocardial injury and the principal factors conditioning end-stage heart failure and death in humans are at present unknown. Thus, the major objective of this application is to establish the rate of myocyte and non-myocyte turnover mediated by hCSC activation and differentiation in the developing, adult, aging and failing heart. To achieve this goal, we will employ retrospective 14C birth dating of cardiac cells to establish the average age of myocytes and non-myocytes. This information will be complemented by defining the age distribution of myocytes and non-myocytes utilizing a mathematical model of age-structured cell populations. These data will offer a novel comprehensive perspective of the cellular processes which govern the lifespan of the human heart. This information is critical for the recognition of the mechanisms that control the dynamics of the human heart, its growth reserve, adaptation to stress and failure. Preliminary data were collected from 4 pure preparations of myocyte nuclei obtained by enzymatic digestion of 1 donor, 20 years old, and 3 explanted hearts with ischemic cardiomyopathy; these patients were 59, 63 and 67 years old (Table 1). 14C content was measured in myocyte DNA by accelerator mass spectrometry and found to reflect an average age of cardiomyocytes of 17.8 ¿ 12.3 years. Myocyte turnover varied from 2.5% to 18.3%, averaging 9.2 ¿ 6.7% per year. In the 3 patients with ischemic cardiomyopathy, none of the cardiomyocytes exiting at birth were present at the time of transplantation. Importantly, these data are very preliminary and the variability in the 4 cases clearly indicates the needs to include a large number of samples to obtain reliable information. Technical issues concerning the actual proportion of mononucleated young myocytes and binucleated older cells and the extraction of large amounts of myocyte DNA are currently being resolved.