Downregulation of microRNA-130a contributes to endothelial progenitor cell dysfunction in diabetic patients via its target Runx3.
Downregulation of microRNA-130a contributes to endothelial progenitor cell dysfunction in diabetic patients via its target Runx3.
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DOI：
10.1371/journal.pone.0068611
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发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
Li Y
中科院分区：
文献类型：
--
作者：
Meng S;Cao J;Zhang X;Fan Y;Fang L;Wang C;Lv Z;Fu D;Li Y
内皮祖细胞（EPCs）功能障碍与糖尿病血管疾病有关。MicroRNAs （miRs）已成为包括血管生成在内的多种细胞过程的关键调节因子。我们最近报道了miR-126、miR-130a、miR-21、miR-27a和miR-27b在II型糖尿病（DM）患者的EPCs中下调，miR-126的下调会损害EPC的功能。本研究进一步探讨miR-130a失调是否也与EPC功能障碍有关。从糖尿病患者和健康人外周血单个核细胞中培养EPCs。进行EPC功能测定（增殖、迁移、分化、凋亡、集落和小管形成）。生物信息学分析用于鉴定EPCs中miR-130a的潜在靶点。real-time PCR检测miR-103a和Runx3的基因表达，Western blotting检测Runx3、细胞外信号调节激酶（ERK）、血管内皮生长因子（VEGF）和Akt的蛋白表达。荧光素酶报告基因法测定Runx3启动子活性。将miR-130a抑制剂或模拟物以及表达miR-130a或Runx3的慢病毒载体或靶向Runx3的短发夹RNA转染到EPCs中以操纵miR-130a和Runx3水平。DM患者EPCs中MiR-130a水平降低。Anti-miR-130a抑制EPC功能，而过表达miR-130a促进EPC功能。miR-130a负调控EPCs中的Runx3 （mRNA、蛋白和启动子活性）。敲除Runx3表达增强EPC功能。MiR-130a还上调EPCs中ERK/VEGF和Akt的蛋白表达。综上所述，miR-130a在维持正常的EPC功能中发挥重要作用，DM的EPCs中miR-130a的降低可能通过其靶点Runx3以及ERK/VEGF和Akt通路导致EPC功能受损。
Dysfunction of endothelial progenitor cells (EPCs) contributes to diabetic vascular disease. MicroRNAs (miRs) have emerged as key regulators of diverse cellular processes including angiogenesis. We recently reported that miR-126, miR-130a, miR-21, miR-27a, and miR-27b were downregulated in EPCs from type II diabetes mellitus (DM) patients, and downregulation of miR-126 impairs EPC function. The present study further explored whether dysregulated miR-130a were also related to EPC dysfunction. EPCs were cultured from peripheral blood mononuclear cells of diabetic patients and healthy controls. Assays on EPC function (proliferation, migration, differentiation, apoptosis, and colony and tubule formation) were performed. Bioinformatics analyses were used to identify the potential targets of miR-130a in EPCs. Gene expression of miR-103a and Runx3 was measured by real-time PCR, and protein expression of Runx3, extracellular signal-regulated kinase (ERK), vascular endothelial growth factor (VEGF) and Akt was measured by Western blotting. Runx3 promoter activity was measured by luciferase reporter assay. A miR-130a inhibitor or mimic and lentiviral vectors expressing miR-130a, or Runx3, or a short hairpin RNA targeting Runx3 were transfected into EPCs to manipulate miR-130a and Runx3 levels. MiR-130a was decreased in EPCs from DM patients. Anti-miR-130a inhibited whereas miR-130a overexpression promoted EPC function. miR-130a negatively regulated Runx3 (mRNA, protein and promoter activity) in EPCs. Knockdown of Runx3 expression enhanced EPC function. MiR-130a also upregulated protein expression of ERK/VEGF and Akt in EPCs. In conclusion, miR-130a plays an important role in maintaining normal EPC function, and decreased miR-130a in EPCs from DM contributes to impaired EPC function, likely via its target Runx3 and through ERK/VEGF and Akt pathways.
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