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自噬和PI3K-Akt-mTOR信号转导通路在缺氧预处理对高糖心肌细胞缺氧/复氧损伤保护中的作用

The roles of autophagy and PI3K-Akt-mTOR signaling transduction pathway in protection of high-glucose cultured cardiomyocytes against hypoxia/reoxygenation injury by hypoxia preconditioning

摘要:

目的 探讨自噬和磷脂酰肌醇3激酶(phosphatidylinositol 3 hydroxy kinase,PI3K)-蛋白激酶B(protein kinase B,Akt)-雷帕霉素靶蛋白(mammalian target of rapamycin,mTOR)信号转导通路在缺氧预处理(hypoxia preconditioning,HPC)对高糖心肌细胞缺氧/复氧(anoxia/reoxygenation,AR)损伤中的作用及机制.方法 将采用高糖培养基培养72 h的心肌细胞用随机数字表法分为5组:空白对照组(S组)、AR损伤对照组(AR组)、HPC组、渥曼青霉素+HPC组(Wo+HPC组)、雷帕霉素+HPC组(Ra+HPC组).采用乳酸脱氢酶(lactate dehydrogenase,LDH)检测试剂盒检测心肌细胞LDH漏出率,Annexin V/PI双染流式细胞术检测心肌细胞凋亡情况,蛋白印迹法检测心肌细胞微管相关蛋白1轻链3(microtubulesas sociated protein light,LC3)-Ⅱ、Beclin-1、mTOR、PI3K表达和磷酸化(phospho,p)蛋白激酶B/蛋白激酶B(p-Akt/Akt)比值.结果 与AR组比较,Wo+HPC组LDH漏出率、早期和晚期凋亡率降低(P<0.05),LC3-Ⅱ和Beclin-1表达降低(P<0.05),mTOR、PI3K表达和p-Akt/Akt比值升高(P<0.05).HPC组各指标与AR组比较,差异均无统计学意义(P>0.05).与HPC组比较,Wo+HPC组LDH漏出率、早期和晚期凋亡率降低(P<0.05),Beclin-1表达降低(P<0.05),mTOR、PI3K表达和p-Akt/Akt比值升高(P<0.05).结论 激活PI3K-Akt-mTOR信号转导通路抑制自噬可明显改善HPC对高糖心肌细胞AR损伤的保护作用.

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abstracts:

Objective To assess the roles of autophagy and phosphatidylinositol 3 hydroxy kinase (PI3K)-protein kinase B (Akt)-mammalian target of rapamycin (mTOR) signaling transduction pathway in protection of high-glucose cultured cardiomyocyte against anoxia/reoxygenation (AR) injury by hypoxia preconditioning (HPC). Methods Ccardiomyocytes cultured in high-glucose medium for 72 h were randomly divided into 5 groups: high glucose control group (S group), AR injury control group (AR group), HPC group, wortmannin+HPC group (Wo+HPC group), rapamycin+HPC group. Lactate dehydrogenase (LDH) leakage and cardiomyocyte apoptosis were analyzed. Microtubulesas sociated protein light(LC3)-Ⅱ, Beclin-1, PI3K, mTOR and phospho(p)-Akt/Akt ratio in cardiomyocytes were assessed by Western-blot assay. Results Compared with AR group, Wo+HPC group showed decrease in the leakage rate of LDH, the percentages of early and late apoptosis, and the levels of LC3-Ⅱand Beclin-1 (P<0.05), but increases in levels of mTOR and PI3K, and p-Akt/Akt ratio in cardiomyocytes (P<0.05), however, HPC group exhibited no significant difference in these parameters (P>0.05). Compared with the HPC group, Wo+HPC group displayed significant decreases in the leakage rate of LDH, the percentages of early and late apoptosis, and the levels of Beclin-1(P<0.05), but elevation of mTOR and PI3K, and p-Akt/Aktratio (P<0.05). Conclusions Activation of PI3K-Akt-mTOR signaling transduction pathway and inhibition of autophagy can significantly improve the protection of high-glucose cultured cardiomyocytes against AR injury by HPC.

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作者: 孙超 [1] 杨桂珍 [1] 薛富善 [1] 李慧娴 [1] 刘亚洋 [1] 廖旭 [1]
栏目名称: 论著
DOI: 10.3760/cma.j.issn.1673-4378.2018.05.001
发布时间: 2018-07-04
基金项目:
国家自然科学基金面上项目 中国医学科学院整形外科医院2015年科研基金重点项目(CZ2015002) National Natural Science Foundation of China (General Program) Key Project of Scientific Research Program of Plastic Surgery Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College of 2015
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