Angiomyogenesis with HIF-1a responsive microRNAs

描述（由申请人提供）：HIF-11应答microRNA的血管肌生成概述：干细胞通过多因素机制促进实验动物模型和心肌梗死患者的功能恢复。我们假设，干细胞重新编程与缺氧诱导因子-11（HIF-11）响应microRNAs（HRM）将促进梗死心脏血管肌生成。从维持谱系定型到血管生成旁分泌活性和分化潜能，MicroRNA（miR）决定了具有离散miR谱的干细胞的固有特性。因此，通过操纵miR对干细胞进行重编程可以改善其旁分泌活性和血管肌生成潜力。我们已经确定了一些HRM的下调或上调的间充质干细胞（MSC）在重编程过程中的短暂缺氧/复氧（A/R）与HIF-11的激活并行的间歇性周期。最显著改变的HRM是miR-210和miR-107，其显著影响MSC的存活和血管生成旁分泌行为（初步结果）。本提案的总体目标是阐明HRM及其靶基因在介导干细胞旁分泌活性、血管肌生成分化和存活的重编程过程中的机制参与。我们的建议旨在验证三个假设。假设1：HRM调节干细胞中的血管生成旁分泌活性。与假设-1相关的具体目标将定义重编程MSC（RePMSC）中离散组HRM的表达谱，特别强调其血管生成旁分泌活性。假设2：HRM是重编程干细胞功能结果的机械决定因素。相关的具体目标将涉及血管生成的体外模型和急性冠状动脉结扎的实验动物模型，以显示重编程在HRM的机械参与下增强干细胞的血管生成潜力。体内研究的终点将包括血管密度分析、成熟指数和改善缺血心脏中的局部血流量以及保存整体心脏功能。还将进行RePMSC植入和HRM直接注射到缺血心脏之间的比较。假设3：HRM机械地参与干细胞存活。我们已经证明，HRM显着提高干细胞存活率。我们的初步研究表明，4种不同的抗凋亡基因在重复循环A/R处理的干细胞中表达上调。我们将阐明每一个已确定的HRM靶基因在干细胞存活中的作用。我们还将确定RePMSC植入后梗死心脏中HRM的变化构成了心脏修复的一个完整和新的机制。总之，我们提出，干细胞重新编程的间歇性周期的A/R与机制参与的HRM，将促进他们的旁分泌，血管平滑肌和细胞保护反应，在梗死的心脏。这些研究将促进基于供体干细胞中HRM操作的新型治疗方法的开发。 公共卫生相关性：该建议旨在研究通过操纵HIF-11响应性microRNA（HRM）重编程的干细胞的血管生成和旁分泌行为，使用我们的新方法，用间歇性多个短循环的缺氧/再给氧处理细胞。我们预期用重编程干细胞治疗后梗死心脏的血管肌生成反应会得到改善。我们的建议将是第一个阐明机制参与的HRM的旁分泌行为，血管平滑肌生成的潜力和细胞保护。

DESCRIPTION (provided by applicant): Angiomyogenesis with HIF-11 responsive microRNAs Summary: Stem cells promote functional recovery in experimental animal models and patients with myocardial infarction through multifactorial mechanisms. We hypothesize that stem cells reprogrammed with hypoxia inducible factor-11 (HIF-11) responsive microRNAs (HRMs) will promote angiomyogenesis in the infarcted heart. Ranging from maintenance of lineage commitment to angiogenic paracrine activity and differentiation potential, MicroRNAs (miRs) determine the inherent properties of stem cells which possess a discrete miRs profile. Hence, reprogramming of stem cells by manipulation of miRs can improve their paracrine activity and angiomyogenic potential. We have identified a number of HRMs which are down- or upregulated in mesenchymal stem cells (MSCs) during reprogramming by intermittent cycles of brief anoxia/re-oxygenation (A/R) in parallel with activation of HIF-11. The most significantly altered HRMs were miR-210 and miR-107 which markedly influenced the survival and angiogenic paracrine behavior of MSCs (Preliminary results). The overall goal of this proposal is to elucidate the mechanistic involvement of HRMs and their target genes during reprogramming which mediate stem cell functions of paracrine activity, angiomyogenic differentiation and survival. Our proposal is aimed to validate three hypotheses. Hypothesis-1: HRMs regulate angiogenic paracrine activity in stem cells. The specific aims pertinent to hypothesis-1 will define the expression profile of the discrete set of HRMs in the reprogrammed MSCs (RePMSCs) with special emphasis on their angiogenic paracrine activity. Hypothesis-2: HRMs are mechanistic determinants of functional consequences of the reprogrammed stem cells. The relevant specific aim will involve in vitro models of angiogenesis and experimental animal model of acute coronary artery ligation to show that the reprogramming accentuate the angiogenic potential of stem cells with the mechanistic participation of HRMs. The end-points of in vivo studies will include blood vessel density analysis, maturation index and improved regional blood flow in the ischemic heart and preservation of global cardiac function. A comparison between RePMSCs engraftment and direct injection of HRMs to the ischemic heart will also be carried out. Hypothesis-3: HRMs mechanistically participate in stem cell survival. We have already shown that HRMs significantly improve stem cell survival. Our preliminary studies showed that 4 different anti-apoptotic genes were up-regulated in the stem cells treated with repeated cycles of A/R. We will elucidate the role of each one of the identified target genes of HRMs in stem cell survival. We will also establish that changes in HRMs in the infarcted heart after RePMSCs engraftment constitute an integral and novel mechanism of cardiac repair. Put together, we propose that stem cells reprogrammed by intermittent cycles of A/R with mechanistic involvement of HRMs, will promote their paracrine, angiomyogenic and cytoprotective responses in the infarcted heart. These studies will facilitate the development of novel therapeutic approaches based on manipulation of HRMs in donor stem cells. PUBLIC HEALTH RELEVANCE: This proposal is designed to investigate angiomyogenic and paracrine behavior of stem cells reprogrammed by manipulation of HIF-11 responsive microRNAs (HRMs) using our novel approach of treating the cells with intermittent multiple short cycles of anoxia/reoxygenation. We anticipate improved angiomyogenic response in the infarcted heart after treatment with reprogrammed stem cells. Our proposal will be the first to elucidate the mechanistic participation of HRMs in paracrine behavior, angiomyogenic potential and cytoprotection.