ROS在吡那地尔后处理减轻大鼠心肌细胞缺氧复氧损伤中的作用:与Nrf2-ARE信号通路的关系  

Role of ROS in attenuation of hypoxia-reoxygenation injury in rat cardiomyocytes by pinacidil postconditioning:relationship with Nrf2-ARE signaling pathway

作　　者：周雯静 徐鹏 陈伟 喻田 王海英 Zhou Wenjing;Xu Peng;Chen Wei;Yu Tian;Wang Haiying(Department of Anesthesiology,the Affliated Hospital of Zunyi Medical University,Zunyi 563000,China;Key Laboratory of Anesthesia and Organ Protection of Guizhou Province,the Affliated Hospital of Zunyi Medical University,Zunyi 563000,China;Key Laboratory of Anesthesia and Organ Protection(Zunyi Medical University),Ministry of Education,Zunyi 563000,China;Department of Critical Care Medicine,the Second Affiliated Hospital of Zunyi Medical University,Zunyi 563000,China)

机构地区：[1]遵义医科大学附属医院麻醉科,遵义563000 [2]贵州省麻醉与器官保护基础研究重点实验室,遵义563000 [3]遵义医科大学麻醉与器官保护教育部重点实验室,遵义563000 [4]遵义医科大学第二附属医院重症医学科,遵义563000

出　　处：《中华麻醉学杂志》2024年第1期91-96,共6页Chinese Journal of Anesthesiology

基　　金：遵义市科技计划项目(遵市科合HZ字(2023)227号,(2021)84号);国家自然科学基金(30960366)。

摘　　要：目的评价ROS在吡那地尔后处理减轻大鼠心肌细胞缺氧复氧损伤中的作用及其与核因子E2相关因子2(Nrf2)-抗氧化反应元件(ARE)信号通路的关系。方法分离、培养成年大鼠心肌细胞,采用随机数字表法分为4组(n=20):对照组(C组)、缺氧复氧组(H/R组)、吡那地尔后处理组(P组)和活性氧清除剂N-(2-巯基丙酰基)-甘氨酸(MPG)+吡那地尔后处理组(MPG+P组)。C组持续于37℃95%O2+5%CO_(2)培养箱中持续培养105 min;缺氧复氧损伤模型制备:5%CO_(2)+1%O2+94%N2条件下缺氧45 min,复氧60 min;P组缺氧45 min,吡那地尔50μmol/L处理5 min,复氧60 min;MPG+P组缺氧45 min,MPG 2 mmol/L处理10 min,吡那地尔处理5 min,复氧60 min。于复氧末检测心肌细胞Ca^(2+)含量和Nrf2活性;观察心肌细胞超微结构,并行线粒体Flameng评分;采用Western blot法和RT-PCR法分别检测Nrf2、超氧化物歧化酶1(SOD1)、醌氧化还原酶1(NQO1)和血红素加氧酶1(HO-1)及其mRNA的表达。结果与C组比较,H/R组心肌细胞Ca^(2+)含量、Nrf2活性和Flameng评分升高,Nrf2、SOD1、NQO1和HO-1及其mRNA表达下调(P<0.05),心肌细胞超微结构损伤加重;与H/R组比较,P组心肌细胞Ca^(2+)含量和Flameng评分降低,Nrf2活性升高,Nrf2、SOD1、NQO1和HO-1及其mRNA表达上调(P<0.05),心肌细胞超微结构损伤减轻;与P组比较,MPG+P组心肌细胞Ca^(2+)含量和Flameng评分升高,Nrf2活性降低,Nrf2、SOD1、NQO1和HO-1及其mRNA表达下调(P<0.05),心肌细胞超微结构损伤加重。结论ROS参与了吡那地尔后处理减轻大鼠心肌细胞缺氧复氧损伤的过程,与激活Nrf2-ARE信号通路有关。Objective To evaluate the role of reactive oxygen species(ROS)in attenuation of hypoxia-reoxygenation(H/R)injury in rat cardiomyocytes by pinacidil postconditioning and the relationship with nuclear factor erythrid 2-related factor 2(Nrf2)-antioxidant response element(ARE)signaling pathway.Methods Adult rat cardiomyocytes were isolated and cultured and then divided into 4 groups(n=20 each)by a random number table method:control group(group C),H/R group,pinacidil postconditioning group(group P)and reactive oxygen scavenger N-(2-mercaptopropionyl)-glycine(MPG)+pinacidil postconditioning group(group MPG+P).Group C was continuously exposed to 95%O2+5%CO_(2)in an incubator at 37℃for 105 min.The cells were exposed to 5%CO_(2)+1%O2+94%N2 in an incubator at 37℃for 45 min followed by reoxygenation for 60 min to prepare H/R injury model.The cells were exposed to hypoxia for 45 min and then treated with pinacidil 50μmol/L for 5 min followed by reoxygenation for 60 min in group P.The cells were exposed to hypoxia for 45 min,treated with MPG 2 mmol/L for 10 min,and then treated with pinacidil for 5 min followed by reoxygenation for 60 min in group MPG+P.The content of Ca^(2+)and activity of Nrf2 in cardiomyocytes were measured at the end of reoxygenation.The ultrastructure of cardiomyocytes was observed,and mitochondrial ultrastructure was evaluated using mitochondrial Flameng score.The expression of Nrf2,superoxide dismutase(SOD1),quinone oxidoreductase 1(NQO1),and heme oxygenase 1(HO-1)protein and mRNA was detected using Western blot and real-time polymerase chain reaction.Results Compared with group C,the Ca^(2+)content,Nrf2 activity and mitochondrial Flameng score were significantly increased,the expression of Nrf2,SOD1,NQO1 and HO-1 protein and mRNA was down-regulated(P<0.05),and the damage to the ultrastructure of cardiomyocytes was aggravated in group H/R.Compared with H/R group,the Ca^(2+)content and mitochondrial Flameng score were significantly decreased,the Nrf2 activity was increased,the expression of Nrf2,SO

关 键 词：活性氧 吡那地尔 缺血后处理 肌细胞 心脏 低氧 NF-E2相关因子2 抗氧化反应元件 

分 类 号：R614[医药卫生—麻醉学]