经皮穴位电刺激对小鼠内毒素急性肺损伤时线粒体质量控制的影响  

Effects of transcutaneous electrical acupoint stimulation on mitochondrial quality control during endotoxin-induced acute lung injury in mice

作　　者：刘华洋 史佳[1] 刘沙沙[1] 吴晓炀 黄炎 武雅 张蓝天 余剑波[1] Liu Huayang;Shi Jia;Liu Shasha;Wu Xiaoyang;Huang Yan;Wu Ya;Zhang Lantian;Yu Jianbo(Department of Anesthesiology and Critical Care Medicine,Nankai Clinical College of Tianjin Medical University(Tianjin Nankai Hospital),Tianjin 300100,China)

机构地区：[1]天津医科大学南开临床学院(天津市南开医院)麻醉科与重症医学科,天津300100

出　　处：《中华麻醉学杂志》2022年第7期866-871,共6页Chinese Journal of Anesthesiology

基　　金：国家自然科学基金(82004076,82074153)。

摘　　要：目的评价经皮穴位电刺激对小鼠内毒素急性肺损伤时线粒体质量控制的影响。方法清洁级健康雄性C57BL/6J小鼠24只,4~6周龄,体重15~20 g,采用随机数字表法分为4组(n=6):对照组(C组)、内毒素急性肺损伤组(L-ALI组)、内毒素急性肺损伤+穴位电针组(L-ALI+EA组)和内毒素急性肺损伤+非穴位电针组(L-ALI+SEA组)。采用尾静脉注射LPS 15 mg/kg的方法建立小鼠内毒素急性肺损伤模型。L-ALI+EA组于建模前5 d每天电针刺激双侧足三里穴、肺俞穴30 min,刺激电流为1~2 mA、频率为2/15 Hz的疏密波,以小鼠下肢出现轻微肌颤为宜,1次/d,每次持续30 min,并于造模前30 min针刺留针至实验结束。L-ALI+SEA组选取双侧足三里和肺俞穴旁开5 mm的非经非穴部位采取浅刺法刺激,其他处理方法皆同电针组。C组未给任何处理。给予LPS后12 h时处死小鼠取肺组织,光镜下观察病理学结果,透射电镜下观察线粒体结构和形态,测定肺组织活性氧(ROS)水平及线粒体DNA(mtDNA)含量;测定肺组织还原型谷胱甘肽(GSH)、氧化型谷胱甘肽(GSSG)含量并计算其比值。Western blot法测定线粒体融合蛋白1(Mfn1)、Mfn2、视神经萎缩蛋白1(OPA1)、动力相关蛋白1(Drp1)、分裂相关蛋白1(Fis1)、过氧化物酶体增殖物激活受体γ共激活因子-1α(PGC-1α)、核呼吸因子-1(NRF1)、NRF2、线粒体自噬标志蛋白PTEN诱导激酶1(PINK1)和E3泛素-蛋白连接酶Parkin的表达。结果与C组比较,L-ALI组、L-ALI+EA组和L-ALI+SEA组肺组织ROS水平、GSSG和mtDNA含量升高,GSH含量和GSH/GSSG比值降低,Mfn1、Mfn2、OPA1、NRF1、NRF2和PGC-1α表达下调,Drp1、Fis1、PINK1和Parkin表达上调(P<0.05);与L-ALI组比较,L-ALI+EA组肺组织ROS水平、GSSG和mtDNA含量降低,GSH含量和GSH/GSSG比值升高,Mfn1、Mfn2、OPA1、NRF1、NRF2和PGC-1α表达上调,Drp1、Fis1、PINK1和Parkin表达下调(P<0.05);与L-ALI+EA组比较,L-ALI+SEA组肺组织ROS水平、GSSG和mtDNA含量升高,GSHObjective To evaluate the effect of transcutaneous electrical acupoint stimulation(TEAS)on mitochondrial quality control during endotoxin-induced acute lung injury(ALI)in mice.Methods Twenty-four clean-grade healthy male C57BL/6J mice,aged 4-6 weeks,weighing 15-20 g,were divided into 4 groups(n=6 each)according to the random number table method:control group(group C),endotoxin-induced ALI group(group L-ALI),endotoxin-induced ALI plus acupoint electroacupuncture group(group L-ALI+EA),and endotoxin-induced ALI plus non-acupoint electroacupuncture group(group L-ALI+SEA).Lipopolysaccharide(LPS)15 mg/kg was injected via the caudal vein to develop the model of endotoxin-induced ALI in anesthetized mice.In group L-ALI+EA,at 5 days before LPS injection,bilateral Zusanli and Feishu acupoints were stimulated with an electric stimulator for 30 min each time at a voltage of 1-2 mA and a frequency of 2/15 Hz until the end of the experiment.In group L-ALI+SEA,stimulation was performed at the points 0.5 cm lateral to the acupoints of bilateral Zusanli and Feishu non-meridian and non-acupoint sites using the shallow puncture method,and the other treatment methods were the same as those previously described in group EA.Group C received no treatment.The mice were sacrificed by euthanasia at 12 h after LPS administration,and lung tissues were obtained for microscopic examination of the pathological changes(with a light microscope)and structure and morphology of mitochondria(with a transmission electron microscope)and for determination of the levels of reactive oxygen species(ROS)and mitochondrial DNA(mtDNA)and contents of glutathione(GSH)and glutathione oxidized(GSSG).The GSH/GSSG ratio was calculated.The expression of mitochondrial fusion proteins mitofusin 1(Mfn1),Mfn2,optic atrophy1(OPA1),dynamin-related protein 1(Drp1),fission protein 1(Fis1),peroxisome-proliferator-activated receptorγcoactivator-1α(PGC-1α),nuclear respiratory factor-1(NRF1),NRF2,PTEN-induced putative protein kinase 1(PINK1)and the E3 ubiquitin ligase(Parkin

关 键 词：针刺疗法 内毒素血症 急性肺损伤 线粒体 

分 类 号：R245[医药卫生—针灸推拿学]