Department of Cardiology, Xinhua Hospital Affiliated to Shanghai Jiaotong University School of Medicine, Shanghai, China.
PLoS One. 2013 Jul 12;8(7):e68611. doi: 10.1371/journal.pone.0068611. Print 2013.
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.
内皮祖细胞 (EPCs) 的功能障碍导致糖尿病血管疾病。微小 RNA (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，II 型糖尿病 (DM) 患者的 EPCs 中 miR-126、miR-130a、miR-21、miR-27a 和 miR-27b 下调，miR-126 下调可损害 EPC 功能。本研究进一步探讨了失调的 miR-130a 是否也与 EPC 功能障碍有关。从糖尿病患者和健康对照者的外周血单个核细胞中培养 EPCs。进行 EPC 功能（增殖、迁移、分化、凋亡、集落和管形成）检测。生物信息学分析用于鉴定 EPCs 中 miR-130a 的潜在靶标。实时 PCR 检测 miR-103a 和 Runx3 的基因表达，Western blot 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。通过荧光素酶报告基因测定测量 Runx3 启动子活性。将 miR-130a 抑制剂或模拟物和表达 miR-130a、Runx3 或针对 Runx3 的短发夹 RNA 的慢病毒载体转染到 EPCs 中，以操纵 miR-130a 和 Runx3 水平。DM 患者的 EPCs 中 miR-130a 减少。抗 miR-130a 抑制而 miR-130a 过表达促进 EPC 功能。miR-130a 在 EPCs 中负调控 Runx3 (mRNA、蛋白和启动子活性)。Runx3 表达的下调增强了 EPC 功能。miR-130a 还上调了 EPCs 中 ERK/VEGF 和 Akt 的蛋白表达。总之，miR-130a 在维持正常 EPC 功能中起重要作用，DM 患者的 EPCs 中 miR-130a 的减少导致 EPC 功能受损，可能通过其靶标 Runx3 以及通过 ERK/VEGF 和 Akt 途径。
