| 李琼,王磊,高波.微小RNA-204-5p 靶向蛋白质酪氨酸磷酸酶1B 基因对缺氧复氧诱导的大鼠心肌细胞氧化应激的影响[J].安徽医药,2022,26(5):977-982. |
| 微小RNA-204-5p 靶向蛋白质酪氨酸磷酸酶1B 基因对缺氧复氧诱导的大鼠心肌细胞氧化应激的影响 |
| Effects of miR-204-5p targeting PTP1B gene on oxidative stress of rat cardiomyocytes induced by hypoxia/reoxygenation and its mechanism |
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| DOI：10.3969/j.issn.1009-6469.2022.05.030 |
| 中文关键词: 心肌再灌注损伤 微小RNA-204-5p 蛋白质酪氨酸磷酸酶1B(PTP1B) H9C2细胞 缺氧复氧 氧化应激 |
| 英文关键词: Myocardial reperfusion injury MiR-204-5p The protein tyrosine phosphatase 1B(PTP1B) H9C2 cells Hypoxiareoxygenation Oxidative stress |
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| 中文摘要: |
| 目的探讨miR-204-5p对缺氧复氧诱导大鼠心肌细胞H9C2氧化应激的调控作用机制。方法体外培养大鼠胚胎心肌细胞H9C2，构建心肌细胞缺氧/复氧模型。实验分为空白组、缺氧复氧组、缺氧复氧+miR-con组、缺氧复氧+miR-204-5p组、缺氧复氧+miR-204-5p+pcDNA组、缺氧复氧+miR-204-5p+pcDNA-蛋白质酪氨酸磷酸酶1B(PTP1B)组。实时荧光定量逆转录聚合酶链式反应(qRT-PCR)和蛋白质印迹法(Western blotting)检测miR-204-5p和PTP1B的表达。双荧光素酶报告基因实验和Western blotting验证miR-204-5p和PTP1B的靶向调控关系。四甲基偶氮唑盐微量酶反应比色(MTT)法检测心肌细胞存活率。试剂盒检测乳酸脱氢酶(LDH)、心肌肌钙蛋白(cTnT)、超氧化物歧化酶(SOD)、活性氧簇(ROS)和丙二醛(MDA)水平变化。流式细胞术检测心肌细胞凋亡情况。结果缺氧复氧处理可显著抑制心肌细胞miR-204-5p表达，促进PTP1B表达。PTP1B是miR-204-5p的靶基因，miR-204-5p可负性调控PTP1B的表达。缺氧复氧处理显著抑制细胞存活，降低SOD水平，提高LDH、ROS和MDA水平，促进细胞凋亡。过表达miR-204-5p可降低LDH、ROS和MDA水平，促进细胞存活，降低细胞凋亡；过表达PTP1B可部分削弱miR-204-5p过表达对缺氧复氧诱导心肌细胞凋亡和氧化应激的影响。结论miR-204-5p通过靶向下调PTP1B抑制心肌细胞氧化应激，促进细胞存活，抑制细胞凋亡，对心肌细胞发挥保护作用。 |
| 英文摘要: |
| Objective To investigate the regulatory mechanism of miR-204-5p on hypoxia- reoxygenation-induced rat cardiomyo?cytes oxidative stress.Methods Rat embryonic cardiomyocytes H9c2 were divided into blank group, hypoxia-reoxygenation group, hy?poxia-reoxygenation+ miR-con group, hypoxia-reoxygenation+miR-204-5p group, hypoxia- reoxygenation+miR-204-5p+pcDNA group,and hypoxia reoxygenation+miR-204-5p+ pcDNA-PTP1B group. The expression of miR-204-5p and PTP1B was detected by qRT-PCR and Western blotting. The dual luciferase reporter gene assay and Western blotting were used to verify the targeted regulatory relation?ship between miR-204-5p and PTP1B. Cell survival rate was measured by MTT assay. The levels of lactate dehydrogenase (LDH),cardi?ac troponin (cTnT), superoxide dismutase (SOD), reactive oxygen species (ROS), and malondialdehyde (MDA) were detected by the kits.Flow cytometry was used to detect cell apoptosis.Results Hypoxia-reoxygenation significantly inhibited the expression of miR-204-5p in cardiomyocytes, while promoted the expression of PTP1B. PTP1B was a target gene of miR-204-5p, and miR-204-5p negatively regu?lated the expression of PTP1B. Hypoxia and reoxygenation significantly inhibited cell survival, reduced SOD level, increased LDH,ROS and MDA levels, and promoted cell apoptosis. Over-expression of miR-204-5p could reduce LDH, ROS and MDA levels, promote cell survival and reduce apoptosis. Over-expression of PTP1B could partially rescue the effects of miR-204-5p on hypoxia-reoxygen?ation-induced apoptosis and oxidative stress.Conclusion MiR-204-5p inhibits oxidative stress, promotes cell survival, and inhibits apoptosis by down-regulating PTP1B, thus playing a protective role on cardiomyocytes. |
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