基础医学与临床 ›› 2022, Vol. 42 ›› Issue (11): 1690-1696.doi: 10.16352/j.issn.1001-6325.2022.11.1690

• 研究论文 • 上一篇    下一篇

敲低miR-34c-5p抑制低氧诱导的大鼠骨髓间充质干细胞的凋亡

董扬, 张芬, 李幸幸, 吴杰, 徐忠诚*   

  1. 金华市人民医院 心内二科, 浙江 金华 321099
  • 收稿日期:2021-05-24 修回日期:2022-02-18 出版日期:2022-11-05 发布日期:2022-11-01
  • 通讯作者: * sino100@outlook.com
  • 基金资助:
    金华市科技计划项目(2019-3-021)

miR-34c-5p knockdown inhibits hypoxia induced cell apoptosis of bone marrow derived mesenchymal stem cells in rats

DONG Yang, ZHANG Fen, LI Xing-xing, WU Jie, XU Zhong-cheng*   

  1. The Second Department of Cardiology, Jinhua People's Hospital, Jinhua 321099,China
  • Received:2021-05-24 Revised:2022-02-18 Online:2022-11-05 Published:2022-11-01
  • Contact: * sino100@outlook.com

摘要: 目的 探讨敲低miR-34c-5p对提高骨髓间充质干细胞(BM-MSCs)存活率的作用。方法 将大鼠BM-MSCs分为常氧组(normoxia组)、低氧组(hypoxia组)、常氧抑制剂阴性对照组(normoxia inhibitor NC组)、低氧抑制剂阴性对照组(hypoxia inhibitor NC组)、常氧微RNA(miR)抑制剂组(normoxia miR inhibitor组)和低氧miR抑制剂组(hypoxia inhibitor miR组);LipofectamineTM2000将 miR-34c-5p inhibitor转染至BM-MSCs细胞;实时荧光定量PCR(RT-qPCR)和ELISA检测miR-34c-5p、IGF、HGF、bFGF和VEGF表达;流式细胞测量术和TUNEL检测细胞凋亡,Western blot检测cleaved caspase-3, -9,Bcl-2和Bax蛋白水平;生化试剂盒检测ATP/ADP。流式细胞测量术检测细胞摄取葡萄糖能力。结果 与hypoxia inhibitor NC组相比,敲低miR-34c-5p可明显抑制MSCs凋亡(P<0.01),显著降低cleaved caspase-3,-9和Bax蛋白表达,提高Bcl-2蛋白表达(P<0.01); IGF、HGF、bFGF和VEGF表达显著升高(P<0.05,P<0.01);ATP/ADP比率和细胞摄取葡萄糖的能力显著提高(P<0.05,P<0.01)。结论 敲低miR-34c-5p可抑制低氧诱导的大鼠BM-MSCs的凋亡。

关键词: 敲低miR-34c-5p, 骨髓间充质干细胞, 低氧, 细胞凋亡

Abstract: Objective To explore the effect of miR-34c-5p knockdown on improving the survival rate of bone marrow derived mesenchymal stem cells (BM-MSCs). Methods The BM-MSCs were divided into normoxia group, hypoxia group, normoxia inhibitor NC group, hypoxia inhibitor NC group, normoxia microRNA(miR) inhibitor group and hypoxia miR inhibitor group. The expression level of miR-34c-5p, IGF, HGF, bFGF and VEGF was detected by real-time quantitative PCR (RT-qPCR) and ELISA. Cell apoptosis was detected by flow cytometry and TUNEL. The protein expression levels of cleaved caspase-3, -9, Bcl-2 and Bax were detected by Western blot. ATP/ADP was detected by biochemical kits, and glucose uptake was detected by flow cytometry. Results Compared with hypoxic inhibitor NC group, miR-34c-5p knockdown significantly inhibited BM-MSCs apoptosis(P<0.01), reduced protein expression of cleaved caspase-3, -9 and Bax and increased protein expression by Bcl-2 (P<0.01). The expression of IGF, HGF, bFGF and VEGF (P<0.05, P<0.01) were significantly increased; ATP/ADP rate and cell capacity of glucose transporting also increased (P<0.05, P<0.01). Conclusions miR-34c-5p knockdown may antagonize hypoxia induced cell apoptosis of BM-MSCs in rat.

Key words: miR-34c-5p knockdown, bone marrow derived mesenchymal stem cells, hypoxia, cell apoptosis

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