Proangiogenic microRNA in Microvesicles Released from Adipose-derived Stem Cells

脂肪干细胞释放的微泡中的促血管生成 microRNA

• 批准号: 8642653
• 负责人: Dong Liu
• 金额: $ 14.15万
• 依托单位: MOREHOUSE SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-04-10 至 2015-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8642653
• 关键词: 3' Untranslated Regions; Address; Adipose tissue; Animal Model; Animals; Attention; Binding; Biological Assay; Cardiac; Cell physiology; Cells; Cellular biology; Cerebrum; Clinical; Clinical Trials; Complement; Disease; Distant; Endocrine; Endothelial Cells; Epidemic; Event; Fatty acid glycerol esters; Future; Gene Targeting; Genetic Materials; Goals; Growth Factor; Histocompatibility Antigens; Human; Intervention; Ischemia; Knowledge; Light; Limb structure; Lipids; Location; Luciferases; Mediating; Mediator of activation protein; Medical; Messenger RNA; MicroRNAs; Molecular; Morbidity - disease rate; Myocardial Ischemia; Nucleic Acid Regulatory Sequences; Operative Surgical Procedures; Patients; Play; Process; Property; Protein Microchips; Proteins; Publications; RNA; Recombinant Proteins; Reporter; Reporting; Reverse Transcriptase Polymerase Chain Reaction; Role; Signal Transduction; Small RNA; Source; Stem cells; Stroke; Subfamily lentivirinae; Surface; Therapeutic; Time; Tissues; Transplantation; Tube; Vascular Diseases; Vascular Endothelial Cell; Western Blotting; angiogenesis; base; cell motility; gene therapy; inhibitor/antagonist; innovation; intercellular communication; loss of function; mRNA Expression; migration; mortality; novel; novel therapeutic intervention; paracrine; preconditioning; protein expression; research study; stem cell therapy; transdifferentiation; treatment strategy

DESCRIPTION (provided by applicant): Ischemic cardiac and cerebral vascular diseases continue to represent a significant and growing source of morbidity and mortality despite advances in traditional treatments. Recently, novel emerging therapeutic strategies focus on stem cells. Adipose tissue derived stem cells (ASCs), which are easily acquired and have reduced surface histocompatibility antigens increasing their allo-transplantation potential, were demonstrated to have beneficial effect on ischemic heart and limb in animals likely due to a paracrine function rather than transdifferentiation. Besides secreted soluble growth factors, cell-released microvesicles (MVs) have been recently described as a new mechanism of intercellular communication. MVs play an important role in cell biologic processes not only by specifically targeting recipient cells to deliver proteins, lipids and/or trigger downstream signalng events, but also by transferring genetic material, mRNA and microRNA. Our preliminary studies for the first time demonstrated that 1). MVs are released from human ASCs (hASCs) and promote vascular endothelial cell migration, 2). They contain RNA, mostly small RNA, 3). These small RNAs, rich in angiogenesis-regulating miRNAs, are proangiogenic. Although the importance of cognate miRNA in angiogenesis and endothelial function has been addressed, the mechanism for of angiogenic effect of miRNAs present in stem cell-released MVs is so far unknown. The goal of this proposal is to unravel the mechanistic aspects of the proangiogenic effect of hASCs-MVs in detail. We hypothesize that microRNAs in MVs secreted by hASCs is proangiogenic. In Aim 1 we will examine the angiogenic effects of miRNAs in hASCs-released MVs using two loss-of-function strategies. hASCs will be transduced with Lentivirus-based specific anti-miRNA hairpin expression construct and the angiogenic potential of the ensuing MVs on the endothelial cells will be assessed. Vascular endothelial cells will be co-transfected with specific anti-miRNA inhibitor and small RNAs isolated from hASCs-released MVs and assessed for angiogenesis. In Aim 2, we will determine the target genes of proangiogenic miRNA from hASCs-released MVs in vascular endothelial cells by profiling analysis of mRNA and proteins. The predicted target genes of miRNA from hASCs-released MVs in recipient cells will be confirmed by using a 3'UTR insertion reporter constructs. These studies will expand our currently scant knowledge of mechanistic aspects of proangiogenic miRNA present in stem cell-released MVs as well as shed light on their paracrine/endocrine properties and set the basis for their use as a novel therapeutic approach for cerebro-vascular disease.

描述（由申请人提供）：尽管传统治疗取得了进步，但缺血性心脏和脑血管疾病仍然是发病率和死亡率的重要且不断增长的来源。最近，新出现的治疗策略集中在干细胞上。脂肪组织来源的干细胞（ASC）很容易获得，并且表面组织相容性抗原减少，增加了同种异体移植的潜力，被证明对动物缺血性心脏和肢体具有有益作用，这可能是由于旁分泌功能而不是转分化。除了分泌的可溶性生长因子外，细胞释放的微泡（MV）最近被描述为细胞间通讯的新机制。 MV 在细胞生物学过程中发挥着重要作用，不仅通过特异性靶向受体细胞来递送蛋白质、脂质和/或触发下游信号事件，而且还通过转移遗传物质、mRNA 和 microRNA。我们的初步研究首次证明了这一点：1）。 MV 由人 ASC (hASC) 释放并促进血管内皮细胞迁移，2)。它们含有 RNA，大部分是小 RNA，3)。这些小 RNA 富含血管生成调节 miRNA，具有促血管生成作用。尽管同源 miRNA 在血管生成和内皮功能中的重要性已经得到解决，但干细胞释放的 MV 中存在的 miRNA 的血管生成作用的机制迄今为止尚不清楚。该提案的目标是详细阐明 hASC-MV 促血管生成作用的机制。我们假设 hASC 分泌的 MV 中的 microRNA 具有促血管生成作用。在目标 1 中，我们将使用两种功能丧失策略来检查 miRNA 在 hASC 释放的 MV 中的血管生成作用。 hASC 将用基于慢病毒的特异性抗 miRNA 发夹表达构建体转导，并且将评估随后的 MV 在内皮细胞上的血管生成潜力。血管内皮细胞将与特定的抗 miRNA 抑制剂和从 hASC 释放的 MV 中分离的小 RNA 共转染，并评估血管生成。在目标 2 中，我们将通过 mRNA 和蛋白质的谱分析，确定血管内皮细胞中 hASC 释放的 MV 中促血管生成 miRNA 的靶基因。受体细胞中 hASC 释放的 MV 中预测的 miRNA 靶基因将通过使用 3'UTR 插入报告构建体进行确认。这些研究将扩大我们目前对干细胞释放的 MV 中存在的促血管生成 miRNA 的机制方面的了解，并阐明它们的旁分泌/内分泌特性，并为它们作为脑血管疾病的新型治疗方法奠定基础。