miR-615, AKT/eNOS signaling, and angiogenesis

miR-615、AKT/eNOS 信号传导和血管生成

• 批准号: 9973357
• 负责人: MARK W FEINBERG
• 金额: $ 68.56万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-15 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9973357
• 关键词: 3' Untranslated Regions; AKT inhibition; Acute; Base Pairing; Binding; Biological Assay; Biology; Blood Vessels; Blood capillaries; Blood flow; Brain; Cardiovascular Diseases; Cardiovascular Physiology; Cardiovascular system; Cell Proliferation; Cell physiology; Cellular biology; Chronic; Clinical Research; Deletion Mutation; Disease; Endothelial Cells; Endothelium; FGF2 gene; Foundations; Gene Expression; Genes; Genetic Transcription; Goals; Growth; Growth Factor; Health Expenditures; Heart; Hindlimb; Human; IGF2 gene; Impairment; Infarction; Injections; Ischemia; Knockout Mice; Leg; Ligation; Limb structure; Link; Mediating; Mediator of activation protein; Messenger RNA; MicroRNAs; Molecular; Morbidity - disease rate; Mus; Myocardial; Myocardial Infarction; Myocardial Ischemia; Nitric Oxide; Organoids; Outcome; Participant; Pathway interactions; Patients; Peripheral arterial disease; Phenocopy; Plasma; Process; Proliferating; Proto-Oncogene Proteins c-akt; Publishing; Recovery; Regulation; Reperfusion Therapy; Reporter Genes; Resistance; Role; Signal Pathway; Signal Transduction; Stimulus; Therapeutic; Thrombospondin 1; Tissue Sample; Tissues; Tube; United States; Untranslated RNA; Untranslated Regions; Vascular Endothelial Cell; Vascular Endothelial Growth Factors; acute coronary syndrome; angiogenesis; artery occlusion; blood vessel development; cell growth; cell motility; crosslinking and immunoprecipitation sequencing; human disease; human subject; human tissue; improved; in vivo; inhibitor/antagonist; insight; intravenous administration; ischemic injury; loss of function; matrigel; migration; molecular imaging; mortality; myocardial infarct sizing; nanoparticle; neovascularization; novel; novel therapeutic intervention; novel therapeutics; overexpression; perfusion imaging; preclinical study; response; response to injury; tissue repair; transcriptome sequencing; transcriptomics; wound closure

Ischemic cardiovascular disease (CVD) due to atherosclerotic occlusion of the arteries to the heart, legs, or brain is associated with considerable morbidity, mortality, and health care expenditure in the United States. The induction and orchestration of new blood vessels is critical for tissue repair in response to injury such as myocardial infarction or peripheral artery disease (PAD). In response to pro-angiogenic stimuli, vascular endothelial cells (ECs) are activated to migrate and proliferate to form primary capillaries. However, despite the importance of ECs in neoangiogenesis, our understanding of the mechanisms regulating this process remains poorly understood. Emerging studies indicate that the inability of angiogenic growth factors to stimulate angiogenesis is likely due to impaired angiogenic signaling and not due to deficiency in these growth factors. MicroRNAs (miRNAs) are small, single-stranded, non-coding RNAs capable of repressing gene expression by base pairing to the 3' untranslated regions (3'-UTRs) of mRNA targets and are involved in a variety of pathophysiological processes in cardiovascular biology, though their function in angiogenesis and angiogenic signaling pathways remains poorly defined. We undertook a microarray profiling approach of plasma from subjects with ischemic CVD and identified that miR-615-5p expression is increased by ischemia and reduced in response to pro-angiogenic stimuli–observations that are recapitulated in both mice and human ischemic paradigms in vivo. Preliminary and published gain and loss-of-function studies reveal that miR-615-5p overexpression markedly impaired EC proliferation, migration, and network tube formation in matrigel, whereas blockade of miR-615-5p had the opposite effects. Mechanistically, using unbiased transcriptomic profiling, we find that miR-615-5p suppressed EC proliferation and binding to 2 unique targets–RASSF2 and IGF2–in their 3'-UTRs and reduced their expression, an effect that selectively regulated the AKT/eNOS signaling pathway in ECs. Finally, systemic intravenous administration of miR-615-5p inhibitors increased blood vessel formation and reduced infarct size and improved blood flow recovery in ischemic legs compared to mice that received scrambled control anti-miR injections. These observations provide the foundation for the central hypothesis that miR-615-5p may serve as a critical regulator of EC proliferation and angiogenic responses. To better understand the precise role of miR-615-5p in AKT/eNOS signaling and angiogenesis, we will in Aim1 delineate the upstream mechanisms governing miR-615-5p expression in ECs. In Aim2, we will determine the molecular basis for miR-615-5p's ability to regulate AKT/eNOS signaling and EC functions critical to angiogenesis. In Aim3, we will explore the effect of altering miR-615-5p expression in the microvasculature on acute and chronic experimental ischemic injury. The results of these studies will provide insights regarding miR-615-5p function in EC biology, pathophysiological angiogenesis, and cardiovascular ischemic states and may provide new targets to rescue impaired angiogenic signaling for a range of ischemic cardiovascular disease states.

缺血性心血管疾病(CVD)，由动脉粥样硬化性闭塞导致的心脏、腿部或 在美国，大脑与相当大的发病率、死亡率和医疗保健支出有关。 新血管的诱导和协调对于组织修复是至关重要的，因为损伤的反应包括 心肌梗死或外周动脉疾病(PAD)。作为对促血管生成刺激的反应，血管 内皮细胞(ECs)被激活以迁移和增殖，形成初级毛细血管。然而，尽管 内皮细胞在新生血管生成中的重要性，我们对这一过程的调控机制的理解仍然 人们对此知之甚少。新出现的研究表明，血管生成生长因子无法刺激 血管生成可能是由于血管生成信号受损，而不是由于这些生长因子的缺乏。 MicroRNAs(MiRNAs)是一种能够抑制基因表达的单链非编码小RNA 通过与mRNA靶标的3‘非翻译区(3’-UTRs)进行碱基配对来表达，并参与 心血管生物学中的各种病理生理过程，尽管它们在血管生成和 血管生成信号通路仍然没有明确的定义。我们进行了微阵列分析方法， 缺血性脑血管病患者血浆中miR-615-5p的表达 对促血管生成刺激的反应减少-在小鼠和人类中都总结了观察到的情况 体内的缺血范例。初步和已发表的功能得失研究表明，miR-615-5p 在Matrigel中，过表达显著损害EC的增殖、迁移和网状管的形成，而 阻断miR-615-5P则有相反的作用。从机制上讲，使用无偏见的转录图谱，我们 发现miR-615-5p抑制了EC的增殖并与2个独特的靶点-RASSF2和IGF2-在其 3‘-UTRs并降低其表达，这种作用选择性地调节AKT/eNOS信号通路 ECS。最后，全身静脉注射miR-615-5P抑制剂可增加血管形成。 与接受注射的小鼠相比，缺血小腿的脑梗塞范围和血流恢复得到了改善 杂乱无章的对照抗miR注射。这些观察结果为中心假说提供了基础 MiR-615-5p可能是EC增殖和血管生成反应的重要调节因子。为了更好地 了解miR-615-5p在AKT/eNOS信号转导和血管生成中的确切作用，我们将在Aim1中描述 调控内皮细胞miR-615-5p表达的上游机制。在AIM2中，我们将确定分子 MiR-615-5p调节AKT/eNOS信号的能力和EC对血管生成至关重要的功能的基础。在……里面 目的探讨改变微血管中miR-615-5p的表达对急性和慢性肾小球疾病的影响。 慢性实验性缺血性损伤。这些研究的结果将提供有关miR-615-5P的见解 在EC生物学、病理生理学血管生成和心血管缺血状态中的作用，并可能提供 新的目标是拯救一系列缺血性心血管疾病状态下受损的血管生成信号。