Role of Inositol requiring enzyme 1 in regulating angiogenesis for diabetic wound repair.

需要酶 1 的肌醇在调节糖尿病伤口修复血管生成中的作用。

• 批准号: 9222758
• 负责人: Jiemei Wang
• 金额: $ 34.65万
• 依托单位: WAYNE STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-03-01 至 2021-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9222758
• 关键词: ANGPT1 gene; Affect; American; Amputation; Angiogenic Factor; Attenuated; Biochemical; Biological Assay; Bone Marrow; Cell Therapy; Cell physiology; Data; Diabetes Mellitus; Diabetic mouse; Diabetic wound; Down-Regulation; Endoplasmic Reticulum; Endothelium; Enzymes; Family; Family member; Functional disorder; Future; Gene Targeting; Gene Transfer; Genes; Genetic; Health; Homing; Human; Impaired wound healing; Impairment; In Vitro; Inositol; Intervention; Investigation; Knockout Mice; Knowledge; Lead; Link; LoxP-flanked allele; Mediating; Metabolic; MicroRNAs; Microarray Analysis; Molecular; Mus; Non-Insulin-Dependent Diabetes Mellitus; Outcome Study; Pathogenesis; Pathway interactions; Patient Recruitments; Patients; Pharmacology; Pilot Projects; Proteins; Publishing; RNA; RNA Splicing; Refractory; Regulation; Research; Role; Site; Skin Ulcer; Stem cells; Stress; Testing; Virus; Work; Wound Healing; adenoviral-mediated; angiogenesis; base; biological adaptation to stress; cell type; coping; db/db mouse; diabetic; diabetic patient; diabetic wound healing; endoplasmic reticulum stress; functional loss; gain of function; improved; in vivo; interest; loss of function; neovascularization; novel; novel therapeutic intervention; overexpression; peripheral blood; pre-miRNA; public health relevance; response; restoration; sensor; success; therapeutic development; tissue repair; vector; whole genome; wound; wound closure

 DESCRIPTION (provided by applicant): Refractory wounds in diabetic patients often result in amputation. Endothelial progenitors cells (EPC) actively participate in wound repair through angiogenesis after homing to the wounding site. However, EPC functions are impaired in diabetes with mechanisms poorly understood. Our pilot studies demonstrate a reduced activity of an important ER sensor ER sensor, Inositol requiring enzyme 1 (IRE1), and a decreased expression of a potent angiogenic factor angiopoietin-1 (ANGPT1) in EPCs from human and mice with type 2 diabetes, contribute to EPC dysfunction. Most importantly, our preliminary studies show that IRE1α suppresses a subset of microRNA (miR) clusters known to be detrimental to angiogenesis. We speculate that this suppression is due to IRE1α's direct splicing activity to precursor miRs (pre-miRs), resulting in depletion of functional miRs. Of particular interest, our preliminary data demonstrate that IRE1α deficiency significantly increased miR-200 family in EPCs from diabetic mice, leading to down-regulation of one of their target genes - ANGPT1. Yet how IRE1 regulates EPC function and wound repair in diabetes is unknown. Therefore, the objective in this proposal is to determine the mechanisms of IRE1 regulation of angiogenesis and wound repair. The hypothesis, built upon our extensive preliminary studies and published work, is that deficiency in IRE1α, leads to insufficient degradation of pre-miRNA in diabetes, resulting in impaired endothelial progenitor cell (EPC) angiogenesis and delayed wound healing. Our hypothesis will be tested in three specific aims: 1) Determine the molecular mechanisms underlying the essential role of IRE1α in maintaining EPC function in diabetes in vitro; 2) Determine how IRE1α-targeted pre-miR modulates EPC function in vitro; 3) Determine how IRE1α improves wound healing in diabetes in vivo. Our approaches encompass in vitro and in vivo studies utilizing adenovirus-mediated gene manipulations, whole genome RNA profiling, IRE1α floxed mice and newly generated endothelium-specific IRE1α knockout mice. Furthermore, human EPCs will be obtained from type 2 diabetic patients and healthy subjects in order to determine the levels of IRE1α pathway and miRs. The proposed study is significant, because it will uncover a previously unrecognized role of the ER stress response in impaired angiogenesis and wound healing in diabetes at the translational level. The investigation of IRE1α-mediated regulation of stress miRs will open a new paradigm for the study of the molecular mechanisms responsible for pathogenesis of refractory wounds, which will enable future development of therapeutics for this devastating situation affecting millions of Americans.

 描述(申请人提供)：糖尿病患者难治性伤口通常会导致截肢。内皮祖细胞(EPC)在归巢到创面后，通过血管生成积极参与创面修复。然而，糖尿病患者的EPC功能受损，其机制尚不清楚。我们的初步研究表明，重要的ER传感器ER传感器肌醇需要酶1(IRE1)的活性降低，以及有效的血管生成因子血管生成素-1(ANGPT1)在人和2型糖尿病小鼠的EPC中的表达减少，导致EPC功能障碍。最重要的是，我们的初步研究表明，IRE1α抑制了已知对血管生成有害的microRNA(MiR)簇的子集。我们推测，这种抑制是由于IRE1α的S直接剪接活性与前体miRs(前miRs)，导致功能miRs耗尽所致。特别有趣的是，我们的初步数据显示，IRE1α缺乏显著增加了糖尿病小鼠内皮祖细胞中的miR-200家族，导致其靶基因之一-NGPT1下调。然而，IRE1如何调节糖尿病患者的EPC功能和伤口修复尚不清楚。因此，本研究的目的是确定IRE1对血管生成和伤口修复的调控机制。这一假说建立在我们广泛的前期研究和已发表的工作基础上，即IRE1miRNA缺乏导致糖尿病患者前α降解不足，导致内皮祖细胞血管生成受损，延迟伤口愈合。我们的假设将在三个特定目标进行验证：1)在体外确定IRE1α在维持糖尿病内皮细胞功能中的关键作用的分子机制；2)确定IRE1α靶向的Pre-miR在体外如何调节EPC功能；3)在体内确定IRE1α如何促进糖尿病伤口的愈合。我们的方法包括在体和体外研究，使用腺病毒介导的基因操作，全基因组核糖核酸图谱，IRE1α小鼠和新产生的内皮特异性IRE1α基因敲除小鼠。此外，还将从2型糖尿病患者和健康人身上获取人内皮祖细胞，以确定IRE1α通路和MIR的水平。这项拟议的研究意义重大，因为它将在翻译水平上揭示内质网应激反应在糖尿病血管生成受损和伤口愈合中先前未被认识到的作用。对IRE1α介导的应激MIR调控的研究将为研究难治性创面发病的分子机制开辟新的范式，这将使未来针对这一影响数以百万计美国人的破坏性局势的治疗方法的发展成为可能。