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中国应用生理学杂志 ›› 2019, Vol. 35 ›› Issue (3): 273-278.doi: 10.12047/j.cjap.5834.2019.057

• 研究论文 • 上一篇    下一篇

髓样分化蛋白2基因沉默对高糖诱导的大鼠心肌细胞增殖抑制、凋亡及炎症反应的影响*

林中民1, 陈国荣1, 张泉波1, 王芳1, 项兰婷1, 曹琼洁2△   

  1. 1. 温州医科大学附属第一医院病理科,
    2. 温州医科大学附属眼视光医院 省部共建眼视光学和 视觉科学国家重点实验室, 浙江 温州 325000
  • 出版日期:2019-05-28 发布日期:2019-06-28
  • 通讯作者: Tel: 0577-88067934; E-mail: vicky.cqj@163.com
  • 基金资助:
    温州市科技计划项目(Y20170053);温州市重大科技创新攻关项目(2018ZY019)

Effects of MD2 gene silencing on high glucose-induced proliferation inhibition, apoptosis and inflammation in rat cardiomyocytes

LIN Zhong-min1, CHEN Guo-rong1, ZHANG Quan-bo1, WANG Fang1, XIANG Lan-ting1, CAO Qiong-jie2△   

  1. 1. Department of Pathology, the First Affiliated Hospital, Wenzhou Medical University,
    2. State Key Laboratory of Ophthalmology, Optometry and Vision Science, Eye Hospital of Wenzhou Medical University, Wenzhou 325000, China
  • Online:2019-05-28 Published:2019-06-28

摘要: 目的:研究髓样分化蛋白2(MD2)基因沉默对高糖(HG)诱导的大鼠心肌细胞增殖抑制、凋亡及炎症反应的影响及其机制。方法:体外大鼠心肌细胞系H9C2细胞随机分为4组(n=3):LG组、HG组、HG + NC组、HG + si-MD2组,分别转染MD2基因小干扰RNA(si-MD2)或阴性对照24 h后进行低糖或高糖处理48 h。RT-qPCR检测MD2及细胞内炎症细胞因子TNF-α、IL-1β、IL-6的表达水平,MTS法、流式细胞术检测细胞增殖能力、细胞周期和细胞凋亡率,Western blot法检测细胞内相关蛋白的表达水平及磷酸化水平。结果:转染si-MD2后,H9C2细胞中MD2的表达水平明显下降(P<0.01)。与低糖(LG)组比较,高糖处理后的H9C2细胞中TNF-α、IL-1β、IL-6的mRNA水平显著升高,细胞增殖能力下降并发生G1期阻滞,细胞凋亡率和Cleaved Caspase-3蛋白水平升高(P< 0.01)。而MD2基因沉默可拮抗高糖对H9C2细胞增殖、细胞周期、凋亡及细胞中TNF-α、IL-1β、IL-6 mRNA水平的影响(P<0.05)。Western blot测定结果表明高糖处理后的H9C2细胞中细胞外信号调节激酶(ERK1/2)、P38丝裂原活化蛋白激酶(P38 MAPK)和C-Jun氨基末端激酶(JNK)蛋白的磷酸化水平明显升高,而MD2基因沉默可抑制高糖诱导下的ERK1/2、P38 MAPK和JNK蛋白激活(P<0.01)。结论:MD2基因沉默可能通过抑制ERK、P38 MAPK和JNK信号通路的激活来减少高糖诱导的大鼠心肌细胞炎症细胞因子表达,减少心肌细胞凋亡,促进细胞增殖。

关键词: 髓样分化蛋白2, 大鼠心肌细胞, 炎症因子, 细胞凋亡, 增殖

Abstract: Objective: To investigate the effects of myeloid differentiation-2 (MD2) gene silencing on high glucose-induced proliferation inhibition, apoptosis and inflammation in rat cardiomyocytes. Methods: The immortalized rat cardiomyocyte cell line H9C2 were transfected with MD2 small interfering RNA (si-MD2) and negative control for 24 h, then stimulated with high glucose (HG) for 48 h. RT-qPCR was performed to detect the mRNA levels of MD2 and inflammatory factors TNF-α, IL-1β and IL-6. MTS and flow cytometry were used to evaluate cell proliferation, cell cycle and apoptosis rate. Western blot was used to detect protein expression levels and phosphorylation levels. Results: The mRNA and protein levels of MD2 in H9C2 cells were dramatically decreased after transfected with si-MD2 (P<0.01). After stimulation of high glucose, the mRNA levels of inflammatory factors, the cells in G0/G1 phase , the cell apoptosis rate and the protein level of cleaved Caspase-3 were significantly increased, while the cell proliferation ability was decreased (P<0.01). MD2 gene silencing antagonized the effects of high glucose on cell proliferation, cell cycle, cell apoptosis and the mRNA levels of TNF-α, IL-1β , IL-6(P<0.05). Western blot analysis showed that the phosphorylation levels of extracellular signal-regulated kinase(ERK1/2), P38 mitogen-activated protein kinase(P38 MAPK) and C-Jun N-terminal kinase(JNK) protein were increased significantly in H9C2 cells treated with high glucose, which could be reversed by silencing of MD2 (P<0.01). Conclusion: This study demonstrates that MD2 gene silencing reverses high glucose-induced myocardial inflammation, apoptosis and proliferation inhibition via the mechanisms involving suppression of ERK, P38 MAPK, JNK signaling pathway.

Key words: myeloid differentiation 2, rat cardiomyocyte cell, inflammatory factors apoptosis, proliferation

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