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Abstract
The mesothelium, which constitutes the outermost layer of the coelomic organs including the heart, lung, liver and gut, plays a critical role in the development, homeostasis and potentially in repair of the internal organs following injury or disease. During organogenesis, the mesothelium contributes to the vasculature and stroma of the developing organs in addition to acting as an important mitogenic signal. While relatively quiescient in the adult, in response to injury, the mesothelial layer has been shown to reactivate its embryonic developmental program, invade the injured tissue and release cytokines that modulate the injury response. Here, we describe highly efficient methods for the differentiation of human pluripotent stem cells (hPSCs) into mesothelial progenitor cells (MPCs) and define their developmental potential in both in vitro and in vivo models. Differential gene expression analysis of freshly isolated murine embryonic mesothelium was used to validate the characterization of our hPSC-derived MPCs as authentic mesothelium. Clonogenic assays were used to determine the in vitro differentiation potential of hPSC-derived MPCs into fibroblast, smooth muscle and endothelial lineages and the multipotency of hPSC-derived MPCs was evaluated in vivo by assessing integration of hPSC-derived MPCs into embryonic chick hearts and mechanically-damaged neonatal mouse hearts. At the molecular level, hPSC-derived MPCs are indistinguishable from their in vivo counterparts and respond to signaling molecules that are known to impact mesothelial cell fate decisions during development as shown by their in vitro differentiation into fibroblasts, smooth muscle cells and endothelium in response to PDGF-alpha, PDGF-beta and Vegf signaling, respectively. When transplanted onto developing chick hearts, MPCs incorporate into the host mesothelium and invade the underlying myocardium. MPCs transplanted into mechanically-damaged neonatal mouse hearts migrate into damaged tissue along with endogenous epicardium-derived cells and assemble into coronary vessels in the repair zone. In addition to the utility of these cells for modeling mesothelial development and disease, this study opens up new avenues for tissue engineering and regeneration of the coelomic organs.