基于PINK1/Parkin通路探讨丹红注射液对冠心病大鼠心肌线粒体损伤的保护作用及机制  

Protective Effect of Danhong Injection on Myocardial Mitochondrial Damage in Rats with Coronary Heart Disease Based on PINK1/Parkin Pathway

作　　者：徐丽萍 XU Liping(Science City Hospital of Sichuan Province,Mianyang 621900,Sichuan,China)

机构地区：[1]四川省科学城医院,四川绵阳621900

出　　处：《中西医结合心脑血管病杂志》2024年第21期3878-3891,共14页Chinese Journal of Integrative Medicine on Cardio-Cerebrovascular Disease

摘　　要：目的:探讨丹红注射液(DHI)对冠心病(CHD)大鼠心肌线粒体的保护作用,以及其对同源性磷酸酶张力蛋白诱导激酶1(PINK1)/E3泛素连接酶(Parkin)信号通路的调控机制。方法:采用高脂饲料、自由饮水持续喂养大鼠8周构建大鼠CHD模型,以丹红注射液低(DHI-L)、丹红注射液高(DHI-H)剂量进行干预,以过表达PINK1的pcDNA3.1-PINK1(pc)进行功能挽救实验;电镜检测各组大鼠心肌组织中完整线粒体的比例;脱氧核糖核苷酸末端转移酶介导的缺口末端标记试剂盒(TUNEL)检测大鼠心肌组织的细胞凋亡;超氧化物阴离子二氢乙啶(DHE)检测大鼠心肌组织中活性氧(ROS)的含量;实时荧光定量聚合酶链式反应(qRT-PCR)仪检测各组大鼠心肌组织中PINK1、Parkin、线粒体融合蛋白2(MNF2)、线粒体分裂基因动态相关蛋白1(DRP1)、B细胞淋巴瘤-2相互作用蛋白1(Beclin1)、自噬相关基因5(ATG5),微管相关蛋白1轻链3(LC-3)的mRNA表达;蛋白免疫印迹法(Western Blot)实验检测大鼠心肌组织中PINK1、Parkin、B淋巴细胞瘤-2(Bcl-2)和Bcl-2相关X蛋白(Bax)表达。结果:与CHD模型大鼠比较,DHI-L、DHI-H能明显升高CHD大鼠心肌组织中完整线粒体的比例,抑制心肌细胞凋亡,下调心肌组织ROS含量,降低心肌组织PINK1、Parkin、DRP1、Beclin1、ATG5和LC-3的mRNA表达,升高心肌组织中MNF2的mRNA表达;下调PINK1、Parkin和Bax表达,上调心肌组织中Bcl-2表达;pcDNA3.1-PINK1可部分逆转DHI对心肌线粒体的保护作用,差异均有统计学意义(P<0.05)。结论:丹红注射液能明显抑制CHD大鼠心肌组织的氧化应激,降低心肌线粒体的自噬水平,下调心肌组织中的细胞的凋亡率,改善实验动物的心功能,这可能与抑制PINK1/Parkin信号的传导有关。Objective:To investigate the protective effect of Danhong injection(DHI)on myocardial mitochondria in rats with coronary heart disease(CHD)and its regulation mechanism in homologous phosphatase tensin induced kinase 1(PINK1)/E3 ubiquitin ligase(Parkin)signaling pathway.Methods:The rat CHD model was constructed by continuous feeding of high-fat diet and free drinking water for 8 weeks.The intervention was performed with the low dose of Danhong injection(DHI-L)and high dose of Danhong injection(DHI-H).The function rescue experiment was performed with pcDNA3.1-PINK1(pc)which overexpressed PINK1.The proportion of intact mitochondria in myocardium of each group were detected by electron microscopy.T erminal-deoxynucleoitidyl transferase-mediated nick end labeling(TUNEL)was used to detect apoptosis in rat myocardial tissue.The content of reactive oxygen species(ROS)in rat myocardium was detected by superoxide anion dihydroethidine(DHE).The quantitative real-time polymerase chain reaction(qRT-PCR)was used to detect the mRNA expression of PINK1,Parkin,mitochondrial fusion protein 2(MNF2),mitochondrial division gene dynamic associated protein 1(DRP1),B-cell lymphoma-2 interaction protein 1(Beclin1),autophagy related gene 5(ATG5),and microtubule associated protein 1 light chain 3(LC-3)in myocardial tissue of rats in each group.The expressions of PINK1,Parkin,B-lymphoblastoma-2(Bcl-2),and Bcl-2 associated X protein(Bax)in rat myocardial tissue were detected by Western Blot.Results:DHI-L and DHI-H could significantly increase the proportion of intact mitochondria in myocardial tissue of CHD rats,inhibit the apoptosis of cardiomyocyte,down-regulate the content of ROS in myocardial tissue,and decrease the mRNA expression of PINK1,Parkin,DRP1,Beclin1,ATG5,and LC-3 in myocardial tissue,increase the mRNA expression of MNF2 in myocardial tissue,down-regulate the expression of PINK1,Parkin and Bax in myocardial tissue,and up-regulate the expression of Bcl-2 in myocardial tissue,pc DNA3.1-PINK1 could partially reverse the protective

关 键 词：冠心病 线粒体损伤 丹红注射液 同源性磷酸酶张力蛋白诱导激酶1/E3泛素连接酶 实验研究 

分 类 号：R285.5[医药卫生—中药学]