过表达BMAL1对脓毒症小鼠心肌损伤的保护作用及机制研究

Protective effect and mechanism of overexpression of BMAL1 on muscle injury in sepsis mice

目的:探究生物钟基因大脑和肌肉芳香烃受体核转运蛋白样蛋白1(BMAL1)过表达对脓毒症小鼠心肌损伤中线粒体自噬途径的影响。方法:将40只小鼠随机分为模型对照组、模型组、阴性对照(NC)组、BMAL1组,每组10只。超声心动图检测小鼠心功能指标左室短轴缩短率(LVFS)、左室射血分数(LVEF)、左室舒张末期内径(LVIDd)、左室收缩末期内径(LVIDs),ELISA检测血清肌酸激酶同工酶(CK-MB)、肌钙蛋白I(cTnI)水平,HE染色观察心肌组织病理变化,TUNEL染色检测心肌细胞凋亡,透射电镜观察心肌线粒体超微结构变化,Western blotting测定心肌组织中微管相关蛋白1轻链3(LC3)、BH3域自噬蛋白(Beclin1)、PTEN诱导激酶(Pink1)、E3泛素-连接酶活性帕金森病蛋白2(Parkin)、电压依赖性阴离子通道蛋白1(VDAC1)表达水平。结果:与模型组和NC组比较,BMAL1组小鼠LVFS、LVEF升高(P<0.05),LVIDd、LVIDs减小(P<0.05),血清中CK-MB、cTnI降低(P<0.05),心肌组织病变得到改善,TUNEL阳性细胞率减少(P<0.05),线粒体损伤减轻,心肌组织中LC3Ⅱ/LC3Ⅰ值升高(P<0.05),Beclin1、Pink1、Parkin、VDAC1蛋白表达上调(P<0.05)。结论:BMAL1过表达能够明显改善脓毒症小鼠心功能,减轻心肌损伤,该作用可能与其促进线粒体自噬有关。

Objective:To explore the effect of circadian clock gene aromatic hydrocarbon receptor nuclear transport-like protein 1(BMAL1)overexpression on mitochondrial autophagy pathway in myocardial injury of sepsis mice.Methods:Forty mice were randomly divided into model control group,model group,negative control(NC)group and BMAL1 group,with 10 mice in each group.Left ventricular short-axis shortening rate(LVFS),left ventricular ejection fraction(LVEF),left ventricular end-diastolic diameter(LVIDd),left ventricular end-systolic diameter(LVIDs)were detected by echocardiography in mice,serum creatine kinase isoenzyme(CK-MB)and troponin I(cTnI)were detected by ELISA,pathological changes of myocardial tissue were observed by HE staining,myocardial apoptosis was detected by TUNEL staining,the ultrastructural changes of myocardial mitochondria were observed by transmission electron microscopy,the expression levels of microtubule-associated protein 1 light chain 3(LC3),BH3-domain autophagy protein(Beclin1),PTEN-induced kinase(Pink1),E3 ubiquitin-ligase active Parkinson s disease protein 2(Parkin)and voltage-dependent anion channel protein 1(VDAC1)in myocardial tissue were determined by Western blotting.Results:Compared with model group and NC group,LVFS and LVEF in BMAL1 group were increased(P<0.05),LVIDd and LVIDs were decreased(P<0.05),serum CK-MB and cTnI were decreased(P<0.05),myocardial tissue lesions were improved,the rate of TUNEL positive cells was decreased(P<0.05),the damage to mitochondria was reduced,LC3Ⅱ/LC3Ⅰratio in myocardial tissue was further increased(P<0.05),and Beclin1,Pink1,Parkin and VDAC1 protein expression levels were further up-regulated(P<0.05).Conclusion:Overexpression of BMAL1 can significantly improve cardiac function and alleviate myocardial injury in sepsis mice,which may be related to the promotion of mitochondrial autophagy.