Identifying novel pathways targeting endothelial-to-mesenchymal transition during heart failure

项目摘要 在美国，每7例死亡中就有1例是由心肌梗死（MI）引起的。因为心脏几乎没有再生- 由于损伤后的神经再生能力，迫切需要新的MI治疗方法。改善肌- MI后的血管修复需要增强边缘灌注区域的血管供应， 通过形成新的心肌细胞和支持脉管系统进行心肌再生。内皮- 间充质转化（EndoMT）在胚胎发育和成人心血管疾病中起着关键作用。 缓解（心血管疾病）。虽然一些研究已经表明EndoMT在心力衰竭（HF）中的重要性，但我们的总体研究结果表明， 对这一过程的了解仍处于起步阶段，部分原因是缺乏EndoMT 基因受到调控，缺乏使用特定工具确定EndoMT作用的确定性研究， 在成人HF。这项提案的目标是利用我们小组的新发现来定义一个 目的：探讨心肌梗死中EndoMT的新的microRNA（miR）调控机制，并最终应用于临床。是-- 心脏中的β-抑制蛋白（β-ARRESTIN）-偏性β 2-肾上腺素能受体（β 2AR）信号传导增强心脏保护作用 并且可能是β AR拮抗剂（β-阻断剂）卡维地洛（Carv）的有益作用的基础。我们最近 发现Carv通过这种机制起作用，促进miR-532- 5 p（miR-532）的成熟， 它起到了重要的心脏保护作用。MiR-532诱导心脏内皮细胞（CEC）功能和新生血管。 并抑制蛋白酶丝氨酸23（prss 23; EndoMT的关键起始子，以及直接和 CEC中miR-532的功能靶标）。有趣的是，miR-532的表达显著富集于 CEC在缺血心肌分离的CEC中选择性下调，这与CEC的表达呈负相关。 艾德与prss 23的表达。因此，更好地理解miR-532介导的心脏保护作用， 确定miR-532增加对EndoMT结果和心脏功能的重要性可能导致 开发新的有效疗法（例如，β 2 AR-选择性β-AR偏置配体） 心血管疾病。我们的假设是，β 2 AR/β-AR介导的miR-532成熟激活和EC中的功能 通过抑制prss 23和EndoMT在梗塞心脏中导致有益的适应性重构。为了测试我们的- 假设，我们计划追求三个目标：1）确定miR-532在EC中的表达对后 缺血性心脏重塑，2）阐明prss 23的作用，prss 23是miR-532的一种新的直接和功能性CEC靶点 （3）研究β 2受体激动剂对CEC和心功能的影响。 由miR-532/prss 23轴标记。该提议是创新的，因为β 2 AR/β-AR/miR-1的作用 532/prss 23轴在EndoMT相关心脏病理中的作用从未被研究过，并提出了EC特异性 miR-532条件性敲除（KO）和prss 23 KO小鼠模型是全新的。该项目也是一个重要的- 这是因为β 2 AR/β-AR信号传导和miR-532/prss 23轴之间的串扰可以被利用， 一个新的目标，研究潜在的机制和治疗器官纤维化介导的EndoMT。

PROJECT SUMMARY Myocardial infarction (MI) alone causes 1 of every 7 deaths in the United States. As the heart has little regen- erative capacity after injury, there is an urgent need for novel therapeutic approaches to MI. Improving myo- cardial repair after MI will require enhancing vascular supply to regions with marginal perfusion and stimulating myocardial regeneration through formation of new cardiomyocytes and supporting vasculature. Endothelial-to- mesenchymal transition (EndoMT) plays a key role in embryonic development and in adult cardiovascular dis- eases (CVDs). While several studies have shown the importance of EndoMT in heart failure (HF), our overall knowledge of this process remains in its infancy in part due to the lack of mechanistic insight by which EndoMT genes are regulated and the absence of definitive studies using specific tools to establish the role of EndoMT in adult HF. The goal of this proposal is to leverage novel discoveries made by our group to define the role of a new microRNA (miR) regulatory mechanism of EndoMT in MI, with an ultimate view to clinical translation. Be- ta-arrestin (beta-arr)-biased beta2-adrenergic receptor (beta2AR) signaling in the heart elicits cardioprotection and may underlie the beneficial effects of the betaAR antagonist (beta-blocker) carvedilol (Carv). We recently discovered that Carv, acting through this mechanism, promotes the maturation of miR-532-5p (miR-532), which plays a vital cardioprotective role. MiR-532 induces cardiac endothelial cell (CEC) function and neovas- cularization after MI, and represses protease serine 23 (prss23; a key initiator of EndoMT, and a direct and functional target of miR-532 in CECs). Interestingly, the expression of miR-532 is significantly enriched in CECs and selectively downregulated in CECs isolated from ischemic myocardium, which is inversely associat- ed with the expression of prss23. Thus, a greater understanding of miR-532-mediated cardiac protection, and establishing the importance of increased miR-532 to the outcome of EndoMT and cardiac function might lead to the development of novel effective therapies (eg. beta2AR-selective beta-arr biased ligands) for numerous CVDs. Our hypothesis is that beta2AR/beta-arr-mediated activation of miR-532 maturation and function in ECs results in beneficial adaptive remodeling in infarcted hearts by repressing prss23 and EndoMT. To test our hy- pothesis, we plan to pursue three aims: 1) determine the contribution of miR-532 expression in ECs to post- ischemic heart remodeling, 2) elucidate the role of prss23, a novel direct and functional CEC target of miR-532 in MI, and (3) investigate the impact of beta-arr-biased agonism of beta2AR on CEC and heart function modu- lated by the miR-532/prss23 axis. This proposal is innovative because the role of the beta2AR/beta-arr/miR- 532/prss23 axis in EndoMT-related cardiac pathologies has never been studied, and proposed EC-specific miR-532 conditional knockout (KO) and prss23 KO mouse models are entirely novel. This project is also signif- icant because the crosstalk between beta2AR/beta-arr signaling and miR-532/prss23 axis can be exploited as a novel target for studying underlying mechanisms and treatment of organ fibrosis mediated by EndoMT.