Empowering Cardiac Progenitor Cell-Mediated Repair of Injured Myocardium by Overexpressing P2Y2 Nucleotide Receptor

 描述(由申请人提供)：心力衰竭(HF)是美国人的主要死亡原因。目前的药物治疗和植入式心脏辅助装置可能有助于在短期内改善心力衰竭的症状，但从长远来看，它们不足以恢复和维持心脏的完整性和功能。自体干细胞疗法已经被引入作为目前治疗心力衰竭的一种有前途的替代方法。然而，无论是内源性干细胞还是过继转移的干细胞，修复反应都是有限的，这是因为在不利的坏死环境中，植入率极低，细胞死亡的易感性增加。因此，对包括心脏来源的前体细胞在内的不同来源的干细胞进行基因修饰和体外扩增，以提高其生长和存活能力，并最终促进体内的优势再生，一直是人们追求的目标。心肌缺血时释放的细胞外核苷酸通过激活多个跨膜核苷酸受体启动炎症和再生反应，这些跨膜核苷酸受体包括P2Y2受体(P2Y2R)，这是一种G蛋白偶联受体，可被ATP和UTP等效激活。除了在啮齿动物模型和人类心肌细胞中已证实的心脏保护作用外，P2Y2R激动剂ATP和UTP还是有效的刺激人类造血干细胞(HHSC)增殖和迁移的药物。ATP还可诱导人骨髓间充质干细胞(HMSCs)分化。然而，细胞外核苷酸在CPC中的生理作用还没有得到充分的研究。本研究旨在探讨P2核苷酸受体介导的生理反应，重点是CPC中的P2Y2R，并确定其潜在的机制。研究策略包括使用药理学以及功能丧失和功能获得研究来初步确定小鼠和人类CPC中P2Y2R介导的反应的体外特征。通过在梗塞小鼠心肌过继移植之前在人CPC中高表达P2Y2R，体外研究结果将在体内得到扩展，我们假设这会改善CPC介导的心肌梗死后受损心肌的修复能力。我们还假设，P2Y2R诱导的CPC再生反应涉及HIPPO信号通路的激活，该信号通路受各种G蛋白偶联受体的调节。这在心肌缺血时释放的细胞外核苷酸、细胞外基质感应和最近被引入的作为损伤后心脏再生重要调节因子的Hippo信号之间提供了一种新的直接联系。

 DESCRIPTION (provided by applicant): Heart failure (HF) is a major cause of death in the United States. Current pharmacological treatments and implantable cardiac assist devices may contribute to improving symptoms of HF on the short term but they are insufficient to restore and maintain cardiac integrity and function on the long term. Autologous stem cell therapy has been introduced as a promising alternative approach to current therapies for HF. However, the reparative responses of either endogenous or adoptively transferred stem cells are limited due to extremely poor engraftment rates and increased susceptibility to cell death in the unfavorable necrotic environment. Hence, genetic modification and ex vivo expansion of stem cells from diverse origins, including cardiac-derived progenitor cells (CPCs), have been pursued to improve their growth and survival capabilities and eventually promote superior regeneration in vivo. Extracellular nucleotides released during cardiac ischemia initiate inflammatory and regenerative responses required for the healing of injured myocardium through the activation of multiple transmembrane nucleotide receptors, including P2Y2 receptor (P2Y2R), a G protein-coupled receptor that is equipotently activated by ATP and UTP. In addition to its established cardioprotective responses in both rodent animal models and human cardiomyocytes, P2Y2R agonists ATP and UTP are potent stimulants of human hematopoietic stem cell (hHSC) proliferation and migration. ATP also induces the differentiation of human bone-marrow derived mesenchymal stem cells (hMSCs). Nonetheless, the physiological roles of extracellular nucleotides in CPCs have not been adequately addressed. This proposal aims to explore the physiological responses mediated by P2 nucleotide receptors with a focus on the P2Y2R in CPCs and identify the underlying mechanisms. Research strategy involves initial in vitro characterization of P2Y2R-mediated responses in mouse and human CPCs using pharmacological as well as loss- and gain-of- function studies. The in vitro findings will be extended in vivo by overexpressing P2Y2R in human CPCs prior to adoptive transfer in infarcted mouse myocardium, which we hypothesize improves CPC-mediated reparative potential of injured myocardium following myocardial infarction. We also hypothesize that P2Y2R-induced regenerative responses in CPCs involve the activation of Hippo signaling pathway that is known to be regulated by various G protein-coupled receptors. This provides a novel direct link between extracellular nucleotides released during cardiac ischemia, extracellular matrix sensing and Hippo signaling that has been recently introduced as an important regulator for cardiac regeneration following injury.