Epicardial progenitors in the developing and postnatal heart

描述（由申请人提供）：心力衰竭是全球发病率和死亡率的主要原因。目前的心力衰竭治疗在增强心肌修复或再生失去的心肌方面无效。改善心肌梗死后的心肌修复需要增强边缘灌注区域的血管供应，并通过形成新的心肌细胞和支持血管系统刺激心肌再生。 心外膜是覆盖心脏的极化上皮，是胎儿心肌生长和冠状动脉血管发生的重要调节因子。心外膜和心肌参与精细的旁分泌信号传导以调节彼此的发育。此外，心外膜细胞经历上皮向间充质转化（EMT），产生心外膜衍生的间充质细胞（EPDC），其迁移到心脏中并分化成成纤维细胞、血管平滑肌细胞、内皮细胞和潜在的心肌细胞。 在成人心脏中，心外膜是心肌损伤反应的重要调节剂，最近的研究表明，心外膜的发育特性可用于治疗性再生。我们的初步数据表明，化学修饰的mRNA（m*RNA）驱动瞬时，高水平的旁分泌因子在心脏中的表达。VEGF-A m*RNA，在实验性心肌梗死（MI）时递送一次，增强毛细血管密度，缩小梗死面积，改善射血分数，并提高长达一年的生存率。以转录因子Wt 1（Wilms Tumor Suppressor 1）表达为标志的心外膜祖细胞是VEGF-A m*RNA活性的主要靶点。VEGF-A扩增并动员MI后WT 1+心外膜祖细胞。值得注意的是，VEGF-A m*RNA改变了这些细胞的命运，增强了它们向内皮细胞和心肌细胞的分化，并减少了它们向肌成纤维细胞的分化。我们的数据表明，一种新的治疗模式，其中短暂激活旁分泌信号通路改变居民祖细胞的命运，以实现持续的治疗效益。这种无细胞治疗范式可以很容易地转化为大型动物和临床研究。 在本研究中，我们进一步研究了成人心外膜细胞行为的调节，并通过以下具体目的发展了我们的初步数据所提出的治疗范式：（1）确定VEGF-A重定向EPDC命运的机制。(2)定义Wt 1在调节正常和受损成人心脏中成人心外膜祖细胞活性中的作用。(3)确定在心肌梗死中具有有益活性的其他因素。通过将新的m*RNA技术与Pu实验室在心外膜祖细胞及其在胎儿和成人心脏中的作用方面的成熟专业知识相结合，该提案将导致对心肌再生的机械见解，推进m*RNA技术在心肌再生中的应用，并为临床转化开辟新的途径。

DESCRIPTION (provided by applicant): Heart failure is the leading cause of morbidity and mortality world-wide. Current heart failure treatments are not effective in enhancing myocardial repair or regenerating lost heart muscle. Improving myocardial repair after myocardial infarction will require enhancing vascular supply to regions with marginal perfusion and stimulating myocardial regeneration through formation of new cardiomyocytes and supporting vasculature. The epicardium, a polarized epithelium covering the heart, is an essential regulator of fetal myocardial growth and coronary vasculogenesis. Epicardium and myocardium engage in elaborate paracrine signaling to regulate each other's development. Furthermore, epicardial cells undergo epithelial to mesenchymal transition (EMT), generating epicardium-derived mesenchymal cells (EPDCs) that migrate into the heart and differentiate into fibroblasts, vascular smooth muscle cells, endothelial cells, and potentially cardiomyocytes. In the adult heart, epicardium is an important modulator of the myocardial injury response, and recent studies indicate that the developmental properties of epicardium may be harnessed for therapeutic regeneration. Our preliminary data show that chemically modified mRNA (m*RNA) drives transient, high level paracrine factor expression in the heart. VEGF-A m*RNA, delivered once at the time of experimental myocardial infarction (MI), enhanced capillary density, reduced infarct size, improved ejection fraction, and enhanced survival for up to one year. Epicardial progenitors, marked by expression of the transcription factor Wt1 (Wilms Tumor Suppressor 1), were a major target of VEGF-A m*RNA activity. VEGF-A expanded and mobilized post-MI WT1+ epicardial progenitors. Remarkably, VEGF-A m*RNA altered the fate of these cells, enhancing their differentiation into endothelial cells and cardiomyocytes and reducing their differentiation into myofibroblasts. Our data suggest a novel therapeutic paradigm, in which brief activation of paracrine signaling pathways alters resident progenitor cell fate to achieve sustained therapeutic benefit. This cell-free therapeutic paradigm can be readily translated to large animal and clinical studies. In this proposal, we further investigate the regulation of adul epicardial cell behavior develop the therapeutic paradigm advanced by our preliminary data, through the following Specific Aims: (1) Determine the mechanism by which VEGF-A redirects EPDC fate. (2) Define the role of Wt1 in regulating adult epicardial progenitor activity in the normal and injured adult heart. (3) Identify additional factors with beneficial activity in myocardal infarction. By combining novel m*RNA technology with the Pu lab's established expertise in epicardial progenitors and their role in fetal and adult heart, this proposal will lead to mechanistic insights into myocardial regeneration, advance application of m*RNA technology to myocardial regeneration, and lead to new avenues for clinical translation.