Cardiovascular disease (CVD) is the leading cause of mortality in the United States and the rest of the developed world. Individuals with CVD often suffer massive heart injuries, leading to the loss of billions of cardiac muscle cells and associated vasculature. Critical work published in the last two decades demonstrated that these lost cells can be partially regenerated by the epicardium, the outermost mesothelial layer of the heart, in a process that highly recapitulates its role in heart development. Upon cardiac injury, mature epicardial cells activate and undergo an epithelial-to-mesenchymal transition (EMT) to form epicardium-derived progenitor cells (EpiPCs), multipotent progenitors that can differentiate into several important cardiac lineages, including cardiomyocytes and vascular cells. In mammals, this process alone is insufficient for significant regeneration, but it may be possible to prime it by administering specific reprogramming factors, leading to enhanced EpiPC function. Here, I differentiated a mature-like model of human induced pluripotent stem cell (hiPSC)-derived epicardial cells (hEpiCs) and used it to conduct a screening of 15 candidate neuroendocrine hormones that may be involved in the epicardial activation process. I identified oxytocin (OXT) as the compound that most strongly induced epicardial cell proliferation, EMT, and dedifferentiation to a progenitor-like state. In addition, I demonstrated that OXT is produced after cardiac cryoinjury in zebrafish, a naturally regenerating animal model. This increased OXT elicited significant epicardial activation, cardiomyocyte proliferation, and neovascularization, thereby promoting heart regeneration. I also found that oxytocin signaling was critical for proper epicardium development in zebrafish embryos. Finally, I differentiated a three-dimensional hiPSC-derived human heart organoid (hHO) model and used it to develop a cryoinjury protocol that simulated cardiac injury. Again, I found that OXT induced cellular proliferation and epicardial activation, features that were consistent with a pro-regenerative phenotype. Remarkably, all the above processes were significantly impaired when OXT signaling was inhibited chemically through receptor antagonism or genetically through RNA interference. RNA sequencing data suggested that the transforming growth factor beta (TGF-Îø) pathway was the primary mediator of OXT-induced epicardial activation. My research combines a naturally regenerating animal model with multiple in vitro models of the human heart to reveal for the first time an evolutionary conserved brain-controlled mechanism that induces cellular reprogramming and regeneration of the injured heart. These findings could yield significant translational advances for the treatment of cardiac injuries and CVD.
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