Endothelial Regeneration Following Lung Vascular Injury

描述(申请人提供)：肺血管损伤导致富含蛋白质的浮肿形成是危重病患者ALI和呼吸衰竭的标志。血管损伤后肺血管内皮细胞再生的机制尚不清楚。然而，随着对内皮祖细胞的了解不断加深，以及我们能够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内皮祖细胞，研究EPC在促进血管再生中的作用和确定血管再生的机制是可行的。此外，我们还利用三苯氧胺诱导的内皮特异性scl-Cre(End-scl-Cre-ER)建立了小鼠报告模型，该模型能够严格追踪内皮损伤后的内皮谱系。使用这种方法，我们也处于解决内在内皮再生和恢复肺血管完整性的机制的地位。这一建议侧重于内源性细胞修复受损的肺内皮细胞以及移植外源性再生内皮祖细胞。在目的1中，我们将研究肺血管损伤后移植的IPSC-EPC(诱导多能干细胞来源的内皮祖细胞)的疗效和命运。我们将验证这样一种假设，即IPSC-EPC移植通过迅速恢复屏障功能以及通过植入肺微血管来预防肺水肿并提高肺损伤后的存活率，从而恢复肺液平衡。在目标2中，我们将研究小鼠成纤维细胞向内皮细胞(Fib-EPC)转化后的肺血管再生能力。我们将使用一种新的microRNA策略来验证这一假设，即成人成纤维细胞直接转化为功能性增殖性内皮祖细胞，恢复肺内皮细胞屏障功能和液体平衡，并预防肺水肿。在目标3中，我们将通过内皮谱系追踪鉴定内源性修复细胞群体，并确定它们的激活是否促进肺血管再生和肺液平衡的恢复。我们将验证内源性修复性内皮细胞的激活和增殖恢复血管后肺内皮屏障和液体平衡的假设 受伤。上述研究将为开发肺血管损伤后内皮细胞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.