Examination of the Functional Role of Sca1+ Vascular Progenitor Cells following Vascular Injury

项目摘要/摘要 在发达国家，心血管疾病是导致死亡和中风的主要原因。长期结果 严重动脉粥样硬化的后续治疗受到新生内膜增生发展的限制 导致血管再狭窄。因此，需要更好和更持久的疗法来预防 血管介入术后再狭窄。一氧化氮(NO)是一种有效的抑制新生内膜增生的物质。而当 人们对一氧化氮如何调节动脉损伤反应知之甚少，但对一氧化氮如何调节动脉损伤反应知之甚少。 在这一过程中的祖细胞。我们实验室的初步数据显示，祖细胞 驻留在外膜，即Sca1+祖细胞，动脉后内膜和中层增多 受伤。NO在阻止这种增加的同时，也将这些细胞的分化重新定向到内皮细胞。 而Sca1+祖细胞具有分化为多种细胞类型的能力，包括血管 平滑肌细胞和血管内皮细胞，其在新生内膜增生发展中的功能作用是 未知。因此，这项提议的目标是探索这些未知因素。我们假设居民 外膜Sca1+祖细胞分化为血管平滑肌细胞 动脉损伤后新生内膜增生的发展，但NO可重定向分化为 驻留的Sca1+细胞向内皮细胞谱系靠拢，从而限制了 新生内膜增生。为了研究这一假说，我们的具体目的是：1)检验 一氧化氮对体外培养的Sca1+血管前体细胞的影响 不加与不加NO的分化培养液，定量血管和内皮细胞分化。 2)研究Sca1+血管祖细胞在体外的迁移和表型命运。 损伤±无暴露。具体地说，我们将通过永久性的方式追踪血管损伤后Sca1+细胞的命运 利用Cre-lox小鼠模型用绿色荧光蛋白标记Sca1+细胞。然后小鼠将经历 股动脉钢丝损伤±NO暴露后Sca1+祖细胞的行为和命运 血管损伤。3)探讨Sca1+血管前体细胞在血管生成中的作用。 新生内膜增生的发生。具体来说，我们将研究Sca1+细胞丢失的影响 基因工程小鼠，通过以下方式选择性和暂时消除Sca1+祖细胞 一旦Sca1基因被激活，就表达白喉毒素。股动脉钢丝损伤将在 这些小鼠±NO。成功完成本提案中描述的研究将阐明 SCA1+祖细胞在新生内膜增生和再狭窄发生中的作用。这项研究可能会导致 血管手术后调节Sca1+祖细胞的新疗法的发展 防止再狭窄。执行这些研究将帮助我获得发展为一名 独立的医生兼科学家。

PROJECT SUMMARY/ABSTRACT Cardiovascular disease is the leading cause of death and stroke in developed countries. Long-term outcomes following treatments for severe atherosclerosis are limited by the development of neointimal hyperplasia leading to restenosis of the vessel. Thus, there is a need for better and more durable therapies to prevent restenosis after vascular interventions. Nitric oxide (NO) is a potent inhibitor of neointimal hyperplasia. While much is known about how NO regulates the arterial injury response, little is known about how NO may regulate progenitor cells in this process. Preliminary data from our laboratory have shown that progenitor cells that reside in the adventitia, namely Sca1+ progenitor cells, increase in the intima and media following arterial injury. NO prevents this increase, while also redirecting differentiation of these cells toward endothelial cells. While Sca1+ progenitor cells have the capacity to differentiate into multiple cell types, including vascular smooth muscle cells and endothelial cells, their functional role in the development of neointimal hyperplasia is unknown. Thus, the goal of this proposal is to explore these unknowns. We hypothesize that resident adventitial Sca1+ progenitor cells differentiate into vascular smooth muscle cells and contribute to the development of neointimal hyperplasia following arterial injury, but that NO redirects differentiation of the resident Sca1+ cells toward an endothelial cell lineage, thereby limiting the development of neointimal hyperplasia. To investigate this hypothesis, our Specific Aims are: 1) to examine the effect of nitric oxide on Sca1+ vascular progenitor cells in vitro, by exposing cultured Sca1+ progenitor cells to differentiation media with and without NO and quantifying smooth muscle and endothelial cell differentiation. 2) To investigate the migration and phenotypic fate of Sca1+ vascular progenitor cells in response to injury ± NO exposure. Specifically, we will track the fate of Sca1+ cells after vascular injury by permanently labeling Sca1+ cells with green fluorescent protein using a Cre-lox mouse model. Mice will then undergo femoral artery wire injury ± NO exposure in order to study the behavior and fate of Sca1+ progenitor cells after vascular injury. 3) To determine the functional role of Sca1+ vascular progenitor cells in the development of neointimal hyperplasia. Specifically, we will study the effect of loss of Sca1+ cells using a genetically engineered mouse that will selectively and temporally eliminate Sca1+ progenitor cells by expressing diphtheria toxin once the Sca1 gene is activated. Femoral artery wire injury will be performed on these mice ± NO. Successful completion of the studies described in this proposal will elucidate the role of Sca1+ progenitor cells in the development of neointimal hyperplasia and restenosis. This research may lead to the development of novel therapeutics that regulate Sca1+ progenitor cells following vascular procedures to prevent restenosis. Execution of these studies will help me to gain the skills necessary to develop as an independent physician-scientist.