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山东大学学报 (医学版) ›› 2022, Vol. 60 ›› Issue (5): 16-21.doi: 10.6040/j.issn.1671-7554.0.2021.1504

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孤核受体NR4A1在H2O2诱导小鼠肾脏足细胞损伤中的作用

刘敏1,张玉超2,马小莉2,刘昕宇2,孙露2,左丹2,刘元涛3   

  1. 1. 南京医科大学附属青岛临床医学院 青岛市市立医院, 山东 青岛 266011;2. 青岛市市立医院内分泌科, 山东 青岛 266011;3. 山东大学齐鲁医院内分泌科(青岛), 山东 青岛 266011
  • 发布日期:2022-06-01
  • 通讯作者: 刘元涛. E-mail:sduliuyuantao@126.com张玉超. E-mail:yc.zhang@hotmail.com
  • 基金资助:
    青岛市创新领军人才项目(16-8-24-zhc)

Effects of orphan nuclear receptor NR4A1 on H2O2 induced injury of mouse renal podocytes

LIU Min1, ZHANG Yuchao2, MA Xiaoli2, LIU Xinyu2, SUN Lu2, ZUO Dan2, LIU Yuantao3   

  1. 1. Qingdao Clinical Medical School of Nanjing Medical University, Qingdao Municipal Hospital, Qingdao 266011, Shandong, China;
    2. Department of Endocrinology, Qingdao Municipal Hospital, Qingdao 266011, Shandong, China;
    3. Department of Endocrinology, Qilu Hospital of Shandong University, Qingdao 266011, Shandong, China
  • Published:2022-06-01

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摘要： 目的 观察双氧水(H2O2)诱导的小鼠肾脏足细胞(MPC-5)氧化应激中孤核受体(NR4A1)表达变化,以及其对细胞增殖和凋亡的影响。 方法 MPC-5分为空白对照组(H2O2处理浓度为0 μmol/L或时间为0 h)和处理组(对应H2O2处理各浓度或时间),慢病毒感染后的MPC-5分为过表达组(Ad-NR4A1组)和空载对照组(Ad-Ctrl组)。采用H2O2处理MPC-5建立氧化应激模型;采用Q-PCR法检测H2O2处理MPC-5后,NR4A1 mRNA表达变化;采用Western blotting法检测NR4A1蛋白、凋亡相关蛋白Caspase 3(17 kDa)表达变化;采用慢病毒感染MPC-5,嘌呤霉素筛选并获得稳定过表达NR4A1的MPC-5细胞(Ad-NR4A1组)与空载细胞(Ad-Ctrl组),H2O2处理后,采用Western blotting法检测两组中Caspase 3表达;采用CCK8实验检测两组细胞增殖活性;采用TUNEL染色法检测两组细胞凋亡。 结果 H2O2 处理后,与空白对照组相比,MPC-5中NR4A1 mRNA相对表达量和蛋白表达量均明显增加(P均<0.05),Caspase 3蛋白表达量高于空白对照组(P<0.05)。Ad-NR4A1组中NR4A1蛋白表达量高于Ad-Ctrl组(P<0.001)。H2O2处理后,Ad-NR4A1组中Caspase 3蛋白表达量高于Ad-Ctrl组(P<0.05),细胞凋亡比率高于Ad-Ctrl组(P<0.05),细胞增殖活性低于Ad-Ctrl组(P<0.05)。 结论 H2O2诱导氧化应激抑制MPC-5细胞增殖、促进细胞凋亡,其作用机制可能与诱导NR4A1表达并激活 Caspase 3途径有关。

关键词: 小鼠肾脏足细胞, 孤核受体NR4A1, 氧化应激, 凋亡

Abstract: Objective To explore the expression of orphan nuclear receptor(NR4A1)in mouse renal podocytes(MPC-5)under H2O2 induced oxidative stress and to investigate the effects of NR4A1 on the viability and apoptosis of MPC-5. Methods MPC-5 were divided into blank control group(H2O2 concentration was 0 μmol/L or treatment time was 0 h), and treatment groups(related to different H2O2 concentrations or treatment time). After MPC-5 were treated with H2O2 to establish the oxidative stress model, the mRNA and protein expressions of NR4A1 and Caspase 3(17 kDa)were detected with Q-PCR and Western blotting, respectively. After MPC-5 were infected with lentivirus with cDNA encoding NR4A1, the stable cell lines with NR4A1 over-expression(Ad-NR4A1 group)and the empty vector(Ad-Ctrl group)were selected with puromycin. After H2O2 treatment, the protein expression of Caspase 3(17 kDa), viability and apoptosis were detected with Western blotting, CCK8 assay and TUNEL staining, respectively. Results After H2O2 treatment, both protein and mRNA expressions of NR4A1 were significantly higher in the treatment groups than in the blank control group(P<0.05), and the protein expression of Caspase 3(17 kDa)also significantly increased(P<0.05). The protein expression of NR4A1 was much higher in Ad-NR4A1 group than in Ad-Ctrl group(P<0.001). After H2O2 treatment, the protein expression of caspase 3(17 kDa)was significantly higher in Ad-NR4A1 group than in Ad-Ctrl group, the apoptotic rate was significantly higher(P<0.05), and the cell viability was significantly lower(P<0.05). Conclusion H2O2 induced oxidative stress contributes to the decreased cell viability and increased cell apoptosis in MPC-5. The possible mechanism is related to the activation of Caspase 3 by NR4A1.

Key words: Mouse renal podocytes, Orphan nuclear receptor NR4A1, Oxidative stress, Apoptosis

中图分类号: 

  • R34
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