苓桂术甘汤对心梗后慢性心衰大鼠线粒体分裂-融合及Sirt3/AMPK信号通路的影响  

Effect of Linggui Zhugantang on M itochondrial Fusion-fission and Sirt3/AM PK Signaling Pathway in Chronic Heart Failure Rats After M yocardial Infarction

作　　者：姚娟 丁芮 李向阳 汤同娟 吴婉婉 魏科东 徐少华 王靓[1,4] 周鹏 黄金玲[1,4] YAO Juan;DING Rui;LI Xiangyang;TANG Tongjuan;W U Wanwan;W EI Kedong;XU Shaohua;WANG Liang;ZHOU Peng;HUANG Jinling(School of Integrated Chinese Medicine and Western Medicine,Anhui University of Chinese Medicine,Hefei 230012,China;School of Chinese Medicine,Anhui University of Chinese Medicine,Hefei 230012,China;The First Affiliated Hospital of Anhui University of Chinese Medicine,Hefei 230031,China;Anhui Province Key Laboratory of Chinese Medicinal Formula,Hefei 230012,China)

机构地区：[1]安徽中医药大学中西医结合学院,合肥230012 [2]安徽中医药大学中医学院,合肥230012 [3]安徽中医药大学第一附属医院,合肥230031 [4]中药复方安徽省重点实验室,合肥230012

出　　处：《中国实验方剂学杂志》2024年第3期1-9,共9页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家自然科学基金项目(81973844,82004180,81373533)。

摘　　要：目的:探讨苓桂术甘汤对心肌梗死(MI)后慢性心力衰竭(CHF)大鼠线粒体分裂-融合及沉默信息调节因子3(Sirt3)/单磷酸腺苷依赖的蛋白激酶(AMPK)信号通路的影响。方法:SD大鼠随机分为假手术组(生理盐水,仅穿线不结扎)、模型组(生理盐水,结扎冠状动脉左前降支近心端)、苓桂术甘汤组(4.2 g·kg^(-1))、卡托普利组(0.00257 g·kg^(-1)),每组10只,连续给药28 d。彩色多普勒超声成像仪检测大鼠心功能参数变化;苏木素-伊红(HE)染色观察心脏病理学改变;免疫荧光法检测活性氧(ROS)水平;JC-1法检测线粒体膜电位;比色法检测三磷酸腺苷(ATP)水平、丙二醛(MDA)、超氧化物歧化酶(SOD)、线粒体呼吸链复合物活性(Ⅰ~Ⅳ);原位末端标记法(TUNEL)染色检测心肌组织细胞凋亡率;蛋白免疫印迹法(Western blot)检测Sirt3、磷酸化(p)-AMPK、磷酸化动力相关蛋白1(p-Drp1)、线粒体分裂蛋白1(Fis1)、线粒体分裂因子(MFF)、视神经萎缩蛋白1(OPA1)蛋白表达水平。结果:与假手术组比较,模型组大鼠左室舒张末期内径(LVIDd)、左室收缩末期内径(LVIDs)显著升高(P<0.01),左室短轴缩短率(LVFS)、左室射血分数(LVEF)显著降低(P<0.01);心肌组织出现炎性细胞浸润,病理损伤明显,ROS、MDA水平与心肌细胞凋亡率显著升高(P<0.01),SOD水平、ATP含量、膜电位显著降低(P<0.01);线粒体呼吸链复合物活性(Ⅰ~Ⅳ)显著降低(P<0.01);p-Drp1、Fis1、MFF蛋白水平显著上调(P<0.01),Sirt3、p-AMPK、OPA1蛋白水平显著下调(P<0.01)。与模型组比较,苓桂术甘汤组与卡托普利组LVIDd和LVIDs显著降低(P<0.01),LVEF和LVFS显著升高(P<0.01),心肌组织炎性细胞浸润与病理损伤明显减轻,ROS、MDA水平与心肌细胞凋亡率显著降低(P<0.01),SOD水平、ATP含量、膜电位显著升高(P<0.01);线粒体呼吸链复合物活性(Ⅰ~Ⅳ)显著升高(P<0.01);p-Drp1、Fis1、MFF蛋白表达显著下调(P<0.01),Sirt3、p-AMPK、OPA1蛋白表达显Objective:To investigate the effects of Linggui Zhugantang on mitochondrial fission and fusion and silencing information regulator 3(Sirt3)/adenosine monophosphate dependent protein kinase(AMPK)signaling pathway in chronic heart failure(CHF)rats after myocardial infarction(MI).Method:SD rats randomly divide into sham operation group(normal saline,thread only without ligature),model group(normal saline,ligation of the left anterior descending coronary artery proximal to the heart),Linggui Zhugantang group(4.8 g·kg^(-1))and Captopril group(0.00257 g·kg^(-1)),with 10 rats in each group.Administere drug continuously for 28 days.Echocardiography detected cardiac function parameters.Hematoxylin eosin(HE)staining observed the pathological changes of the heart.Immunofluorescence detected the levels of reactive oxygen species(ROS).JC^(-1) detect mitochondrial membrane potential.Colorimetry measure adenosine triphosphate(ATP),superoxide dismutase(SOD),malondialdehyde(MDA),mitochondrial respiratory chain complex activity(I-IV).TdT-mediated dUTP nick end labeling(TUNEL)staining detected the apoptosis rate of myocardial tissue.Western blot detected protein expression levels of Sirt3,phosphorylated AMPK(p-AMPK),phosphorylated dynamic-related protein 1(p-Drp1),mitochondrial fission protein 1(Fis1),mitochondrial fission factor(MFF),optic atrophy protein 1(OPA1).Result:Compared to the sham group,the left ventricular end diastolic diameter(LVIDd)and left ventricular end systolic diameter(LVIDs)were significantly increased in model group(P<0.01),while the left ventricular short axis shortening rate(LVFS)and left ventricular ejection fraction(LVEF)were significantly decreased(P<0.01).There were inflammatory cell infiltration and obvious pathological injury in myocardial tissue.ROS,MDA levels and myocardial cell apoptosis rate were significantly increased(P<0.01),SOD level,ATP content,and membrane potential were significantly decreased(P<0.01).The activity of mitochondrial respiratory chain complexes(I-IV)was significantly decrease

关 键 词：苓桂术甘汤 慢性心力衰竭 氧化应激 细胞凋亡 线粒体分裂-融合 沉默信息调节因子3(Sirt3)/单磷酸腺苷依赖的蛋白激酶(AMPK)信号通路 

分 类 号：R2-0[医药卫生—中医学] R22R285.5R289R33