miR-10a regulation of regional arterial endothelial phenotypes in atherosclerosis

项目描述（由申请人提供）：本计划书概述了方云博士在Peter Davies博士实验室完成博士后培养，并通过建立心血管病理生理学多学科研究项目，向独立学术职位过渡的综合研究和职业发展计划。PI目前是美国心脏协会会员，在分子生物学，生物工程和血管生物学领域接受过培训。在2年的指导期内，PI将接受来自宾夕法尼亚大学导师和咨询委员会的额外学术指导。职业发展计划旨在为PI提供必要的生物医学研究知识和技能，以便成功过渡为独立院士，并随着R00阶段工作的进展而获得RO1。总体研究目标是确定microRNA-10a （miR-10a）在动脉粥样硬化发生和发展过程中介导内皮表型的作用。microrna介导的转录后调控在动脉生物学和病理学中知之甚少。K99 PI进行的初步研究表明，miR-10a在不同动脉部位的不同地形表达显著促进了与动脉粥样硬化易感性相关的内皮异质性。值得注意的是，在大型动物模型中，暴露于血流紊乱的动脉粥样硬化易感区域的内皮细胞miR-10a在体内被显著抑制。进一步的功能基因组学和生化分析表明，miR-10a通过抑制nf - b介导的炎症促进内皮细胞的动脉粥样硬化保护表型（PNAS报道）。该研究方案验证了血流和/或高胆固醇敏感的miR-10a在动脉粥样硬化的发生和发展过程中动态调节内皮表型的总体假设。目的1将验证动脉粥样硬化保护miR-10a通过直接抑制一组NF-B和未折叠蛋白反应（UPR）阳性反应分子来抑制内皮炎症和内质网应激（ER应激）的假设。目的2将验证动脉粥样硬化相关血流动力学力调节机械敏感转录因子的假设，从而导致体内动脉粥样硬化易感区和动脉粥样硬化保护区内皮细胞miR-10a生物发生的差异控制。Aim 3将构建内皮细胞miR-10a诱导表达的转基因小鼠模型，验证内皮细胞miR-10a表达与动脉粥样硬化之间的因果关系，从而在体内验证具有动脉粥样硬化保护作用的miR-10a抑制内皮细胞炎症和内质网应激，减轻动脉粥样硬化负担的假说。该目标将通过在体外和体内系统中整合系统生物学和分子分析来实现，从而对内皮细胞miR-10a的下游基因网络和上游调节因子与动脉粥样硬化的机制理解。

DESCRIPTION (provided by applicant): The proposal outlines an integrated research and career development plan for Yun Fang, Ph.D to complete postdoctoral training in the laboratory of Dr. Peter Davies and transition to an independent academic position by establishing a multi-disciplinary research program in cardiovascular pathophysiology. The PI is currently an American Heart Association Fellow who is trained in the fields of molecular biology, bioengineering, and vascular biology. During the 2 year mentored period, the PI will receive additional academic guidance from the mentor and the advisory committee at the University of Pennsylvania. The career development plan is designed to equip the PI with necessary knowledge and skills in biomedical research for a successful transition as an independent academician, leading to a RO1 as the R00 phase of the work progresses. The overall research goal is to determine the role of microRNA-10a (miR-10a) in mediating endothelial phenotypes in relation to the initiation and development of atherosclerosis. MicroRNA-mediated post-transcriptional regulation is poorly understood in arterial biology and pathology. Preliminary studies conducted by the K99 PI demonstrate that differential topographic expression of miR-10a in distinct arterial sites significantly contributes to the endothelial heterogeneity associated with susceptibility to atherosclerosis. Notably, endothelial miR-10a is significantly suppressed in vivo in athero-susceptible regions exposed to disturbed blood flow in a large animal model. Further functional genomics and biochemical analyses demonstrated that miR-10a promotes the athero-protective phenotype in endothelial cells by suppressing NF-B-mediated inflammation (PNAS in press). The research proposal tests the overall hypothesis that flow and/or hypercholesterolemia-sensitive miR-10a dynamically modulates endothelial phenotypes in the initiation and progression of atherosclerosis. Aim 1 will test the hypothesis that athero-protective miR-10a suppresses endothelial inflammation and Endoplasmic Reticulum stress (ER stress) by direct inhibition of a cohort of positive NF-B and Unfolded Protein Response (UPR) responsive molecules. Aim 2 will test the hypothesis that athero- relevant hemodynamic force regulates mechano-sensitive transcription factors, leading to differential control of endothelial miR-10a biogenesis at athero-susceptible and athero-protected regions in vivo. And Aim 3 will develop a transgenic mouse model that exhibits inducible expression of endothelial miR-10a to demonstrate the causality of endothelial miR-10a expression and atherosclerosis, thereby testing in vivo the hypothesis that athero-protective miR-10a inhibits endothelial inflammation and ER stress, alleviating atherosclerotic burden. The goal will be achieved by integrating system biology and molecular analysis in both in vitro and in vivo systems, leading to mechanistic understandings of the down-stream gene networks and up-stream regulators of endothelial miR-10a with respect to atherosclerosis. PUBLIC HEALTH RELEVANCE: Atherosclerosis causes most cardiovascular diseases such as heart attack and stroke. The plaques develop in predictable sites of arteries which are susceptible to disease. The sites have unusual blood flow which affects the endothelial cells lining the artery and predisposes them to lesions. My studies analyze the regulatory role of small nucleic acids called microRNAs in cells in relation to atherosusceptibility and blood flow. We hope to define microRNA-related targets for therapeutic intervention in cardiovascular disease.