线粒体通透性转换孔在心肌细胞胆碱能抗炎通路中的作用  

Role of mitochondrial permeability transition pore in cholinergic anti-inflammatory pathway in cardiomyocytes

作　　者：程怡[1] 薛富善[2] 崔昕龙[2] 王世玉[2] 李瑞萍[2] 刘高谱 杨桂珍[2] 孙超[2] 廖旭[2] 

机构地区：[1]首都医科大学附属北京安贞医院麻醉科,100029 [2]中国医学科学院北京协和医学院整形外科医院麻醉科

出　　处：《中华麻醉学杂志》2015年第8期993-997,共5页Chinese Journal of Anesthesiology

基　　金：国家自然科学基金（8117012s）

摘　　要：目的 探讨线粒体通透性转换孔（mPTP）在心肌细胞胆碱能抗炎通路中的作用.方法 乳鼠心肌细胞培养72 h后,采用随机数字表法将其分为5组（n=36）：空白对照组（C组）、缺氧复氧组（A/R组）、特异性α7烟碱型乙酰胆碱受体激动剂PNU282987组（P组）、mPTP开放剂苍术苷组（ATR组）和PNU282987＋ATR组（P＋A组）.采用不含血清的低糖DMEM,置于厌氧培养盒中（O2浓度＜0.1％）缺氧6h后,更换为含10％胎牛血清的高糖培养基,置于37 ℃、常氧、5％ CO2培养箱中复氧6h,以制备缺氧复氧损伤模型.P组和ATR组于复氧即刻,分别在培养基中加PNU282987（终浓度为30 μmol/L）和苍术苷（终浓度为20 μmol/L）.P＋A组于复氧即刻在培养基中加入PNU282987（终浓度30 μmol/L）和苍术苷（终浓度20 μmol/L）.于复氧6h时测定心肌细胞乳酸脱氢酶（LDH）漏出率、凋亡率、NF-κB p65和磷酸化NF-κB p65 Ser536（p-NF-κB p65 Ser536）表达水平、培养液IL-6和TNF-α的浓度和线粒体膜电位.结果 与C组相比,A/R组、P组、ATR组和P＋A组LDH漏出率、早期和晚期细胞凋亡率升高,培养液IL-6和TNF-α浓度升高,心肌细胞NF-κB p65和p-NF-κBp65 Ser536表达上调,线粒体膜电位降低（P＜0.01）;与A/R组相比,P组LDH漏出率和早期细胞凋亡率降低,培养液IL-6和TNF-α浓度降低,心肌细胞p-NF-κB p65 Ser536表达下调,线粒体膜电位升高,ATR组LDH漏出率和早期细胞凋亡率升高,培养液IL-6和TNF-α浓度升高,心肌细胞p-NF-κBp65 Ser536表达上调,线粒体膜电位降低,P＋A组早期细胞凋亡率降低,培养液IL-6和TNF-α浓度降低,心肌细胞p-NF-κBp65 Ser536表达下调（P＜0.05或0.01）,LDH漏出率、晚期细胞凋亡率和线粒体膜电位差异无统计学意义（P＞0.05）;与P组相比,P＋A组LDH漏出率和晚期细胞凋亡率升高,培养液IL-6和TNF-α浓度升高,心肌细胞p-NF-κBp65Ser536表达上调,线粒体膜电位降低（P＜0.05）.�Objective To investigate the role of mitochondrial permeability transition pore （mPTP） in cholinergic anti-inflammatory pathway in cardiomyocytes.Methods After being cultured for 72 h, cardiomyocytes of neonatal Sprague-Dawley rats were randomly divided into 5 groups （n =36 each） using a random number table： control group （group C） , anoxia/reoxygenation （A/R） group, specific α7 nicotinic acetylcholine receptor agonist PNU282987 group （PNU282987 group）, mPTP opener atractyloside group （ATR group） and PNU282987＋ATR group （P＋A group）.The cells were incubated in serum-free and low-glucose DMEM culture medium which was placed in an anaerobic box （O2〈 0.1%） lasting for 6 h, the culture medium was then replaced with high-glucose DMEM culture medium supplemented with 10% fetal bovine serum, and the cells were then re-incubated and exposed to 5% CO2 in normoxia in an incubator at 37 ℃ for 6 h to establish a model of A/R injury.In P and ATR groups, PNU282987 （final concentration 30 μmol/L） and atractyloside （final concentration 20 μmol/L） were added to the culture medium immediately after onset of reoxygenation, respectively, and in P＋A group, the combination of the two drugs previously mentioned was added instead.At 6 h of reoxygenation, lactate dehydrogenase （LDH） release rate, apoptosis, expression of NF-κBp65 and phosphorylated NF-κB p65 Ser536 （p-NF-κB p65 Ser536） in cardiomyocytes, concentrations of interleukin-6 （IL-6） and tumor necrosis factor-α （TNF-α）in the supernatant, and mitochondrial membrane potential （/△ ψm） were detected.Results Compared to group C, the LDH release rate, early and late apoptosis and concentrations of IL-6 and TNF-α in the supernatant were significantly increased, the expression of NF-κB p65 and p-NF-κB p65 Ser536 was up-regulated, and △ψm was decreased in A/R, P, ATR and P＋A groups.Compared to A/R group, the LDH release rate, early apoptosis and concentrations of IL-6 and TNF-α in the supernatant w

关 键 词：线粒体膜转运蛋白质类 乙酰胆碱 迷走神经 心肌再灌注损伤 肌细胞 心脏 

分 类 号：R614[医药卫生—麻醉学]