Cardiovascular Impact of Gata4 Loss in the c-Kit Lineage

项目摘要 缺血性心脏病是世界范围内主要的死亡原因。为了应对这场重大的公共卫生疫情， 出现了两个流行的研究领域：1)旨在提供培养扩展的多能性的细胞疗法 缺血事件后冠状动脉输注细胞；2)内源性心脏祖细胞 以及它们在体内更新和再生心肌方面的潜在作用。起初，这两个主题似乎 千丝万缕的联系，但现在变得明显的是，虽然交付CPC进行治疗可能会提供一些 机制不明的益处，这些细胞在心肌中的滞留是边缘的和重新开始的。 心肌细胞分化更不常见。以酪氨酸激酶为标志的一组推定的CC 受体c-Kit一直是这两个领域研究的共同焦点。我们的实验室最近发表了一份基因谱系 Kit血统在心脏中对心脏贡献的示踪模型，最终表明 Kit来源的细胞很少成为心肌细胞，但有很高的血管形成倾向 内皮细胞。此外，这项研究表明，大约80%的Kit血统追踪的心肌细胞 实际上是从融合事件中衍生出来的，而受到追捧的新生心肌细胞甚至比 最初认为。为了验证这项研究，我们的目标是消除Kit来源的细胞产生任何 通过敲除基本的生心肌转录因子Gata4，理论上允许 仅适用于融合后的有血统记录的心肌细胞。有趣的是，Kit血统的出现 随着Kit谱系中Gata4基因的缺失，融合所致的心肌细胞急剧增加，而且， 我们观察到心脏和整个心脏血管内皮细胞中Kit系细胞的急剧增加。 我们现在假设，联合这些发现的是Gata4在成人血管内皮细胞中的一个新角色， 最初被认为是心肌细胞先天遗传特征的一个有趣的驱动因素是 可能是由于血管通透性增加和新生血管生成所致的偶然读数。虽然许多GATA 很少有研究报道转录因子在血管发育中的作用，包括Gata4 都专注于成人血管的动态平衡。更好地理解Gata4介导的转录调控 血管扩张的研究可以为心脏更新和修复的机制提供有意义的见解。至 针对Gata4在心脏血管内皮细胞中的这一新作用，我们将追求以下目标：(1) 确定内皮细胞特异性缺失Gata4是否导致血管通透性增加和增强 (2)评估成人心脏内皮细胞Gata4缺失后心脏恢复的变化。

Project Summary Ischemic heart disease is the leading cause of death worldwide. To approach this major public health epidemic, two prevailing areas of study have emerged: 1) Cell therapy aimed at delivering culture-expanded pluripotent cells by coronary artery infusion after an ischemic event, and 2) Endogenous cardiac progenitor cells (CPCs) and their potential role for in vivo renewal and regeneration of the myocardium. Initially these two topics seemed inextricably linked, but it is now becoming clear that while delivery of CPCs for therapy may provide some mechanistically-unexplained benefit, the retention of these cells into the myocardium is marginal and de novo cardiomyocyte differentiation is even less common. A population of putative CPCs, marked by the tyrosine kinase receptor, c-Kit, have been a common focus in both areas of study. Our lab recently published a genetic lineage tracing model demonstrating the cardiac contribution of the Kit lineage in the heart which showed conclusively that Kit-derived cells rarely become cardiomyocytes, but have a high propensity for the formation of vascular endothelium. Furthermore, this study demonstrated that approximately 80% of Kit lineage-traced cardiomyocytes were actually derived from fusion events and the sought-after de novo cardiomyocyte was even more rare than originally believed. To validate this study, we aimed to eliminate the ability of Kit-derived cells to make any de novo cardiomyocytes by knocking out essential cardiomyogenic transcription factor, Gata4, theoretically allowing only for lineage-traced cardiomyocytes derived from fusion. Interestingly, the appearance of Kit lineage-traced cardiomyocytes due to fusion increased dramatically with the loss of Gata4 in the Kit lineage, and, furthermore, we observed a dramatic increase of both Kit-lineage cells in the heart and total cardiac vascular endothelium. We now hypothesize that what is uniting these findings is a novel role for Gata4 in the adult vascular endothelium, and what initially appeared to be an interesting driver of innate fusigenic character with cardiomyocytes is most likely an incidental readout due to increased vascular permeability and neoangiogenesis. While many GATA transcription factors have had previously reported roles in vascular development, including Gata4, few studies have focused on adult vascular homeostasis. Better understanding of this Gata4-mediated transcriptional control of vascular expansion could provide meaningful insight into mechanisms of cardiac renewal and repair. To address this novel role of Gata4 in the cardiac vascular endothelium, we will pursue the following aims: (1) Determine whether endothelial-specific loss of Gata4 leads to increased vascular permeability and enhanced angiogenesis; (2) Assess changes in cardiac recovery after deletion of Gata4 in the adult cardiac endothelium.