Endothelial Regeneration Following Lung Vascular Injury

描述（由申请方提供）：肺血管损伤导致富含蛋白质的水肿形成是危重患者ALI和呼吸衰竭的标志。血管损伤后肺血管内皮再生的机制尚不清楚。然而，随着对内皮祖细胞（EPCs）的不断了解以及我们从诱导的多能干细胞衍生的内皮祖细胞（iPSC）或通过体细胞（成纤维细胞）的直接重编程来鉴定和获得足够数量的EPCs的能力，解决EPCs在促进血管再生中的作用并定义血管再生的机制是可行的。此外，我们已经开发了一种小鼠报告模型，采用他莫昔芬诱导的内皮特异性Scl-Cre（End-Scl-Cre-ER），这使得严格的跟踪内皮细胞损伤后的内皮细胞谱系。使用这种方法，我们也在解决机制的内在内皮再生和恢复肺血管的完整性。该建议的重点是通过内源性细胞修复受损的肺内皮以及移植外源性再生EPCs。在目标1中，我们将研究肺血管损伤后移植iPSC-EPCs（诱导多能干细胞衍生的内皮祖细胞）的疗效和命运。我们将测试iPSC-EPC移植通过急性恢复屏障功能以及通过植入肺微血管系统来预防肺水肿并改善肺损伤后的存活率，从而恢复肺液体平衡的假设。在目标2中，我们将研究小鼠成纤维细胞的肺血管再生潜力，这些成纤维细胞已经经历了谱系转化为内皮细胞（Fib-EPCs）。我们将检验一种假设，即成年成纤维细胞使用一种新的microRNA策略直接转化为功能性增殖性EPCs，恢复肺内皮屏障功能和液体平衡，并预防肺水肿。在目标3中，我们将通过内皮细胞谱系示踪来鉴定内源性修复细胞的群体，并确定它们的活化是否促进肺血管再生和肺液体平衡的恢复。我们将检验这样一个假设，即内源性修复性内皮细胞的活化和增殖在血管损伤后恢复肺内皮屏障和液体平衡。 损伤上述研究将为开发肺血管损伤后内皮再生和恢复的新疗法提供必要的框架。

DESCRIPTION (provided by applicant): Lung vascular injury leading to protein-rich edema formation is a hallmark of ALI and respiratory failure in critically ill patients. Little is known bout the mechanisms of lung vascular endothelial regeneration following vascular injury. However, with an ever-growing understanding of Endothelial Progenitor Cells (EPCs) and our ability to identify and obtain them in sufficient numbers from induced Pluripotent Stem Cell-derived Endothelial Progenitor Cells (iPSC) or through direct reprogramming of somatic cells (fibroblasts), it is feasible to address the role EPCs in promoting vascular regeneration and to define the mechanism of vascular regeneration. In addition, we have developed a mouse reporter model employing the tamoxifen- inducible endothelial specific Scl-Cre (End-Scl-Cre-ER) which enables the rigorous tracing of endothelial lineage following endothelial injury. Using this approach we are also in the position of addressing mechanisms of intrinsic endothelial regeneration and restoration of lung vascular integrity. This proposal focuses on restoration of the injured lung endothelium by endogenous cells as well as transplantation of exogenous regenerative EPCs. In Aim 1 we will investigate the efficacy and fate of transplanted iPSC-EPCs (induced pluripotent stem cells-derived endothelial progenitor cells) following lung vascular injury. We will test the hypothesis that iPSC-EPC transplantation prevents pulmonary edema and improves survival after lung injury by acutely restoring barrier function as well as through engraftment into the lung microvasculature, and thus restores lung fluid balance. In Aim 2 we will study the lung vascular regenerative potential of mouse fibroblasts that have undergone lineage conversion into endothelial cells (Fib-EPCs). We will test the hypothesis that adult fibroblasts converted directly into functional proliferative EPCs using a novel microRNA strategy restore lung endothelial barrier function and fluid balance and prevent pulmonary edema. In Aim 3 we will identify through endothelial lineage tracing populations of endogenous reparative cells and determine whether their activation promotes lung vascular regeneration and restoration of lung fluid balance. We will test the hypothesis that activation and proliferation of endogenous reparative endothelial cells restores lung endothelial barrier and fluid balance following vascular injury. The above studies will provide the essential frame-work needed to develop novel therapies for endothelial regeneration and recovery after lung vascular injury.