芪苈强心胶囊对心力衰竭小鼠心肌内皮间充质作用及机制研究

Effect and mechanism of Qipi Qiangxin Capsule on myocardial endothelial mesenchymal in heart failure mice

目的探究芪苈强心胶囊对心肌梗死后心力衰竭小鼠心肌细胞内皮间充质转化的作用及可能机制。方法6~8周龄雄性C57BL/6小鼠21只,分为正常对照组、药物对照组(正常鼠加高剂量芪苈强心胶囊给药)和心力衰竭模型组,其中心力衰竭模型组分为阳性药物对照组[微小RNA21(miR-21)抑制剂干预]、高、中、低剂量芪苈强心胶囊干预组,共计7组,心力衰竭模型组在冠状动脉左前降支实施结扎术,正常对照组和药物对照组手术步骤与模型组一致,但未行左冠状动脉前降支结扎。利用小动物超声影像系统检测小鼠血流动力学参数;HE染色观察小鼠心肌病理学改变;天狼星红染色检测小鼠心肌组织纤维化;麦胚凝集素染色评价心肌细胞扩大程度;采用酶联免疫吸附试验(ELISA)检测小鼠血清心力衰竭标志物N末端脑钠肽前体(NT-proBNP)和肌钙蛋白I(cTnI)表达及小鼠心肌组织中激动蛋白-1(AP-1)和转化生长因子-β1(TGF-β1)的表达水平;实时荧光定量PCR(qRT-PCR)检测小鼠外周血miR-21的表达水平;蛋白质印迹法检测小鼠心肌组织CD31、管内皮钙黏素(VE-cadherin)、免疫组化检测α-平滑肌肌动蛋白(α-SMA)、铁死亡抑制蛋白1(FSP1)、骨形态发生蛋白7(BMP7)、TGF-β1、Smad2/3和p-Smad2/3蛋白表达水平。结果芪苈强心胶囊显著改善心力衰竭小鼠心功能和心肌组织病理损伤,降低心肌组织纤维化表达水平缓解非梗死区心肌细胞代偿性肥大,抑制心肌组织内皮间充质转化,降低miR-21、TGFβ1、α-SMA、FSP1和Smad2/3表达水平,增加BMP7、VE-cadherin、CD31表达水平。结论芪苈强心胶囊可能是通过调控miR-21的表达水平进而影响下游TGF-β1/Smad2/3信号通路的改变从而改善心肌梗死所致心力衰竭小鼠心肌损伤。

Objective To investigate the effect and possible mechanism of Qili Qiangxin Capsule on the endothelial mesenchymal transition of myocardial cells in mice with heart failure after myocardial infarction.Methods A total of 21 male C57BL/6 mice aged 6-8 weeks were divided into a normal control group,a drug control group(normal mice were treated with high-dose Qili Qiangxin Capsule),and a heart failure model group.The heart failure model group was divided into a positive drug control group[microRNA21(miR-21)inhibitor intervention],high,medium,and low dose Qili Qiangxin Capsule intervention groups,a total of seven groups.The heart failure model group underwent ligation of the left anterior descending coronary artery,while the normal control group and the drug control group underwent the same surgical procedure as the model group,but left anterior descending coronary artery ligation was not performed.A small animal ultrasound imaging system was used to detect hemodynamic parameters in mice.HE staining was used to observe pathological changes in mouse myocardium.Sirius red staining was used to detect myocardial tissue fibrosis in mice.Wheat-germ agglutinin staining was used to evaluate the degree of myocardial cell enlargement.Enzyme linked immunosorbent assay(ELISA)was used to detect the expression of N-terminal pro brain natriuretic peptide(NT-proBNP)and cardiac troponin I(cTnI)in mouse serum heart failure markers,as well as the expression levels of activator protein-1(AP-1)and transforming growth factor-β1(TGF-β1)in mouse myocardial tissue.Real time fluorescence quantitative PCR(qRT-PCR)was used to detect the expression level of miR-21 in peripheral blood of mice.Western blot was used to detect CD31 and VE-cadherin in mouse myocardial tissue,and immunohistochemistry was used to detect the expression levels ofα-smooth muscle actin(α-SMA),iron death inhibitory protein 1(FSP1),bone morphogenetic protein 7(BMP7),TGF-β1,Smad2/3,and p-Smad2/3 proteins.Results Qili Qiangxin Capsule significantly improved heart function and myocardial tissue pathological damage in heart failure mice,reduced myocardial tissue fibrosis levels,alleviated compensatory hypertrophy of non infarcted myocardial cells,inhibited endothelial mesenchymal transition in myocardial tissue,decreased the expression of miR-21,TGFβ1,α-SMA,FSP1 and Smad2/3,and increased the expression of BMP7,VE-cadherin,and CD31.Conclusion Qili Qiangxin Capsule may improve myocardial injury in mice with heart failure caused by myocardial infarction by regulating the expression of miR-21 and affecting the downstream TGF-β1/Smad2/3 signaling pathway.