琥珀酸脱氢酶在缺氧后处理减轻大鼠心肌细胞缺氧复氧损伤中的作用及其与mito-KATP的关系  

Role of succinate dehydrogenase in hypoxic postconditioning-induced reduction of hypoxia-reoxygenation injury in myocardial cells of rats and the relationship with mito-KATP

作　　者：周雯静 唐锟 陈伟 杜康 喻田 王海英 Zhou Wenjing;Tang Kun;Chen Wei;Du Kang;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,Zunyi 563003,China;Department of Anesthesiology,the Southwest Hosppital of Army Medical University,Chongqing 400038,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563003 [3]陆军军医大学第一附属医院手术麻醉科,重庆400038

出　　处：《中华麻醉学杂志》2023年第4期422-426,共5页Chinese Journal of Anesthesiology

基　　金：国家自然科学基金项目(81860062);贵州省科技计划项目(黔科合基础【2018】1195)。

摘　　要：目的评价琥珀酸脱氢酶(SDH)在缺氧后处理(HPC)减轻大鼠心肌细胞缺氧复氧损伤中的作用及其与线粒体ATP敏感性钾通道(mito-KATP)的关系。方法分离成年雄性SD大鼠心肌细胞并培养48 h,采用随机数字表法分为7组(n=24):空白对照组(Nor组)、缺氧复氧组(H/R组)、SDHA-siRNA腺病毒+缺氧复氧组(siRNA+H/R组)、HPC组、SDHA-siRNA腺病毒+HPC组(siRNA+HPC组)、5-羟葵酸(5-HD)+HPC组和SDHA-siRNA腺病毒+5-HD+HPC组(siRNA+5-HD+HPC组)。Nor组常氧条件下持续培养195 min。缺氧复氧损伤模型制备:5%CO_(2)+1%O_(2)+94%N2条件下缺氧45 min,复氧150 min。HPC方法:缺氧45 min时行3个循环的复氧5 min-缺氧5 min处理,继续复氧培养120 min。mito-KATP阻断剂5-HD给药方法:缺氧30 min时加入终浓度为100μmol/L的5-HD。各siRNA组心肌细胞成功转染SDHA-siRNA腺病毒,沉默SDHA表达。于复氧末,分别检测细胞活力、钙离子水平、SDH活性、ATP含量、线粒体通透性转换孔(mPTP)开放程度和线粒体膜电位(MMP)。结果与Nor组比较,H/R组细胞活力、ATP含量和MMP降低,mPTP开放程度、钙离子水平和SDH活性升高(P<0.05)。与H/R组比较,siRNA+H/R组和HPC组细胞活力、ATP含量和MMP升高,mPTP的开放程度、钙离子水平和SDH活性降低(P<0.05)。与HPC组比较,5-HD+HPC组细胞活力、ATP含量和MMP降低,mPTP开放程度、钙离子水平和SDH活性升高,siRNA+HPC组细胞活力、ATP含量和MMP升高,mPTP开放程度、钙离子水平和SDH活性降低(P<0.05)。与siRNA+HPC组比较,siRNA+5-HD+HPC组细胞活力、ATP含量和MMP降低,mPTP开放程度和钙离子水平升高(P<0.05),SDH活性差异无统计学意义(P>0.05)。与5-HD+HPC组比较,siRNA+5-HD+HPC组SDH活性降低,其余指标差异无统计学意义(P>0.05)。结论HPC可能通过降低SDH活性,开放mito-KATP,减轻大鼠心肌细胞缺氧复氧损伤。Objective To evaluate the role of succinate dehydrogenase(SDH)in hypoxic postconditioning(HPC)-induced reduction of hypoxia-reoxygenation(H/R)injury in myocardial cells of rats and the relationship with mitochondrial ATP-sensitive potassium channels(mito-KATP).Methods Myocardial cells isolated from adult male Sprague-Dawley rats were cultured for 48 h and then divided into 7 groups(n=24 each)using a random number table method:blank control group(Nor group),H/R group,SDHA-siRNA adenovirus+H/R group(siRNA+H/R group),HPC group,SDHA-siRNA adenovirus+HPC group(siRNA+HPC group),5-HD+HPC group,and SDHA-siRNA adenovirus+5-HD+HPC group(siRNA+5-HD+HPC group).Nor group was continuously cultured for 195 min under normoxic conditions.The H/R injury model was prepared by exposing the cells to hypoxia for 45 min in 5%CO_(2)+1%O_(2)+94%N2,followed by reoxygenation for 150 min.The HPC method involved three cycles of 5 min reoxygenation/5 min hypoxia at the end of 45 min ischemia before 120 min reoxygenation.The mito-KATP blocker 5-HD administration method involved adding 5-HD at a final concentration of 100μmol/L at 30 min of hypoxia.The myocardial cells in each siRNA group were successfully transfected with SDHA-siRNA adenovirus to silence SDHA expression.The cell viability,calcium ion level,SDH activity,ATP content,degree of mitochondrial permeability transition pore(mPTP)opening,and mitochondrial membrane potential(MMP)were measured at the end of reoxygenation.Results Compared with Nor group,the cell viability,ATP content and MMP were significantly decreased,and the degree of mPTP opening,level of calcium ion and activity of SDH were increased in H/R group(P<0.05).Compared with H/R group,the cell viability,ATP content and MMP were significantly increased,and the degree of mPTP opening,calcium ion level and SDH activity were decreased in siRNA+H/R group and HPC group(P<0.05).Compared with HPC group,the cell viability,ATP content and MMP were significantly decreased,and the degree of mPTP opening,calcium ion level and SDH activ

关 键 词：琥珀酸脱氢酶 心肌再灌注损伤 KATP通道 线粒体 缺血后处理 

分 类 号：R614[医药卫生—麻醉学]