解剖学报

解剖学报 ›› 2018, Vol. 49 ›› Issue (5): 584-590.doi: 10.16098/j.issn.0529-1356.2018.05.004

• 神经生物学 • 上一篇    下一篇

microRNA-25在脑缺血/再灌注损伤诱导的细胞凋亡过程中的作用

张军峰1,2 赵朝华1 郝佳晖2 房智超2 徐仓宝1 徐曦1,2*   

  1. 1.西安医学院陕西省缺血性心血管疾病重点实验室，西安 710021; 2.西安医学院人体解剖学教研室，西安 710021
  • 收稿日期:2018-03-16 修回日期:2018-05-21 出版日期:2018-10-06 发布日期:2018-10-06
  • 通讯作者: 徐曦 E-mail:zhangjf_08@126.com
  • 基金资助:
    陕西省缺血性心血管疾病重点实验室开放基金项目;陕西省大学生创新训练项目;西安医学院国科金培育基金;西安医学院配套基金项目;陕西省优势学科建设经费

Effects of microRNA-25 on cerebral ischemia/reperfusion injury-induced cell apoptosis

ZHANG Jun-feng 1,2 ZHAO Zhao-hua1 HAO Jia-hui2 FANG Zhi-chao2  XU Cang-bao1 XU Xi 1,2*   

  1. 1. Shanxi Key Laboratory of Ischemic Cardiovascular Disease, Xi’an Medical University, Xi’an 710021, China; 2. Department of Human Anatomy, Xi’an Medical University, Xi’an 710021, China
  • Received:2018-03-16 Revised:2018-05-21 Online:2018-10-06 Published:2018-10-06
  • Contact: XU Xi E-mail:zhangjf_08@126.com

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摘要：

目的 在体外培养的SH-SY5Y细胞和IMR-32细胞中，观察microRNA-25（miR-25）对缺血/再灌注（I/R）损伤诱导的细胞凋亡的作用及其可能机制。方法 在体外培养的人SH-SY5Y细胞和IMR-32细胞中，用氧葡萄糖剥夺/再恢复（OGDR）模拟脑缺血/再灌注损伤。合成miR-25过表达片段，并构建慢病毒载体质粒，用脂质体将载体质粒转导入细胞中。MTT法检测细胞活力、TUNEL法检测凋亡、RT-PCR、Real-time PCR和Western blotting分别检测目的基因mRNA和蛋白的表达情况。 结果 与正常培养组相比，OGDR组中细胞内miR-25表达下调，细胞活力明显下降，凋亡明显增多，同时Bax、Caspase-3 mRNA和蛋白表达上调，Bcl-2 mRNA和蛋白表达下调（n=3，P<0.05）；而与OGDR组相比，过表达miR-25组内细胞活力明显升高，凋亡明显减少，同时Bax、Caspase-3 mRNA和蛋白表达下降Bcl-2 mRNA和蛋白表达升高（n=3，P<0.05）。结论 I/R损伤后miR-25表达上调可能通过Bax/Bcl-2-Caspase-3途径进而抑制I/R损伤诱导的凋亡，为I/R损伤的临床治疗提供潜在的治疗靶点。

关键词: 小核糖核酸-25, 缺血/再灌注损伤, Bax, Bcl-2, Caspase-3, 实时定量聚合酶链反应

Abstract:

Objective To evaluate the role and the possible mechanisms of microRNA-25 (miR-25) in regulating oxygen-glucose deprivation (OGD)/reperfusion (OGDR)-induced cell apoptosis in cultured SH-SY5Y and IMR-32 cell. Methods The OGD model was constructed in human SH-SY5Y and IMR-32 cells to mimic I/R injury. Fragments mediated miR-25 overexpression, mi-R25 mimics, were synthesized and constructed into recombined lentivirus plasmid. Using lipofectamine, the plasmids were transfected into the cells. The cell viability and apoptosis were detected with MTT assay and TUNEL. The mRNA and protein expression of the target genes were detected using RT-PCR, Real-time PCR and Western blotting respectively. Results Compared with the normal group, miR-25 was down-regulated in OGDR model group with decreased cell viability and increased apoptosis. Meanwhile, mRNA and protein expression of Bax and Caspase-3 were increased, but with decreased expression of Bcl-2 (n=3, P<0.05). Compared with the OGDR group, cell viability was increased and the apoptosis was decreased in miR-25 transfection group, with decreased expression of Bax and Caspase-3, as well as increased expression of Bcl-2 (n=3, P<0.05). Conclusion Taken together, our result indicate that up-regulation of miR-25 inhibits cerebral I/R injury-induced apoptosis through regulating Bax/Bcl-2 and Caspase-3 signaling pathway, which will provide a promising therapeutic target for cerebral ischemia/reperfusion injury.

Key words: MicroRNA-25, Ischemia/reperfusion injury, Bax, Bcl-2, Caspase-3, Real-time PCR