盐酸小檗胺对心肌梗死小鼠心肌损伤的影响及其分子机制

Effect of berbamine hydrochloride on myocardial injury in myocardial infarction mice and its molecu⁃lar mechanism

目的观察盐酸小檗胺(BA)对心肌梗死(MI)小鼠心肌损伤的影响,并探讨其可能的机制。方法将60只成年雄性C57/BL小鼠随机分Sham组、MI组、BA+MI组、BA+DAPT+MI组,每组15只。MI组、BA+MI组、BA+DAPT+MI组采用结扎法建立MI模型,术后即刻BA+MI组腹腔注射BA 15 mg/(kg·d),BA+DAPT+MI组腹腔注射BA15 mg/(kg·d)及肺组织缺刻基因1(Notch1)抑制剂DAPT 10 mg/(kg·d),1次/d,持续1个月,Sham组和MI组术后不做任何干预。术后1个月,各组通过心脏超声检查计算心脏左心室射血分数(EF),并检测血清乳酸脱氢酶(LDH)、肌酸激酶同工酶(CK-MB)水平。各组处死后取心肌组织,观察心肌病理改变,采用Western blotting法检测心肌组织肌球蛋白样BCL2结合蛋白1(Beclin1)、自噬受体蛋白(p62)、微管相关蛋白轻链3-Ⅱ(LC3-Ⅱ)/LC3-Ⅰ、Notch1、发状分裂相关增强子1(Hes1)蛋白表达。结果 Sham组细胞核无逸出,心肌纤维无断裂,具有清晰的心肌纤维结构且排列有序;MI组和BA+DAPT+MI组均可见细胞核逸出,心肌纤维断裂明显且排列紊乱,心肌纤维化明显;与MI组的心肌纤维排列状态相比,BA+MI组心肌纤维化程度明显减轻。与Sham组比较,MI组EF及心肌组织Notch1、Hes1蛋白表达均降低,血清LDH、CK-MB水平及心肌组织Beclin1、p62表达、LC3-Ⅱ/LC3-Ⅰ均升高(P均<0.05)。与MI组及BA+DAPT+MI组比较,BA+MI组EF及心肌组织Notch1、Hes1蛋白表达均升高,血清LDH、CK-MB水平及心肌组织Beclin1、p62表达、LC3-Ⅱ/LC3-Ⅰ均降低(P均<0.05);MI组与BA+DAPT+MI组上述指标比较均无统计学差异(P均>0.05)。结论 BA可明显改善MI小鼠的左心功能、减轻心肌损伤,其机制可能与通过激活Notch1/Hes1信号通路而减轻心肌细胞自噬有关。

Objective To observe the effect of berbamine hydrochloride(BA)on myocardial injury in myocardial in-farction(MI)mice and to investigate the possible mechanism.Methods Sixty adult male C57/BL mice(8 weeks old)were randomly divided into the control group(Sham group),myocardial infarction group(MI group),BA-treated MI group(BA+MI group),and BA combined with Notch1 inhibitor DAPT-treated MI group(BA+DAPT+MI group),with 15 in each.In the MI group,BA+MI group,BA+DAPT+MI group,we established the MI models by ligation.Immediately af-ter the operation,in the BA+MI group,BA[15 mg/(kg·d)]was injected intraperitoneally,and in the BA+DAPT+MI group,BA[15 mg/(kg·d)]and DAPT[10 mg/(kg·d)]were injected intraperitoneally.The treatment lasted for 1 month in the BA+MI group and BA+DAPT+MI group.However,the mice in the Sham group and MI group were not treated.After treatment for 1 month,serum LDH and CK-MB were detected by special kits.Left ventricular ejection fraction(EF)was detected by echocardiography.Myocardial tissues were taken after death in each group,and pathological changes of myo-cardia were observed.Western blotting was used to detect the expression levels of myosin-like BCL2-binding protein 1(Beclin1),p62,andmicrotubule-associatedproteinlightchain3-Ⅱ(LC3-Ⅱ)/LC3-Ⅰ,Notch1,andHes1protein.Results In the Sham group,there were no nuclei escaping,no broken myocardial fibers,and the myocardial fibers had clear struc-tures and were arranged in order;in the MI group and BA+DAPT+MI group,we could see more nuclei escaping,and the myocardial fibers were broken and arranged disorderly,and the myocardial fibrosis was significant.Compared with the arrangement of myocardial fibers in the MI group,the degree of myocardial fibrosis in the BA+MI group was significantly al-leviated.Compared with the Sham group,EF and Notch1 and Hes1 protein expression levels decreased,and the serum LDH,CK-MB levels,Beclin1,p62 expression and LC3-Ⅱ/LC3-Ⅰin the myocardial tissues increased in the MI group(all P<0.05).Compared with the MI group and BA+DAPT+MI group,EF and Notch1 and Hes1 protein expression levels increased in the myocardial tissues,and the serum LDH,CK-MB levels,Beclin1,p62 expression and LC3-Ⅱ/LC3-Ⅰde-creased in the BA+MI group(all P<0.05).There were no statistical differences in the above indicators between the MI group and the BA+DAPT+MI group(all P>0.05).Conclusion BA can significantly improve the myocardial function and reduce myocardial damage in MI mice by activating Notch1/Hes1 signal and thus alleviating myocardial autophagy.