基于细胞自噬的荭草苷抗心肌缺血/再灌注损伤的机制研究

Cardioprotection of orientin against myocardial ischemia reperfusion injury through induction of autophagy

目的本研究主要探讨荭草苷抗心肌缺血/再灌注损伤的保护作用与细胞凋亡和细胞自噬的关系。方法采用Langendorff法,建立大鼠离体心脏缺血/再灌注模型,缺血30min,再灌注120 min;随机将Wistar大鼠分为正常对照组、模型组、不同剂量荭草苷组(1.0、2.0、4.0 mg·kg^(-1))、自噬抑制剂组(2.0 mg·kg^(-1)荭草苷+渥曼青霉素)以及阳性对照药白藜芦醇组;采用多道生理记录仪采集血流动力学指标,全自动生化仪检测灌流液中心肌酶活性,电镜观察大鼠心肌细胞超微结构改变及自噬体,TUNEL法检测心肌细胞凋亡,Western blot法检测心肌细胞中自噬相关蛋白LC3和Beclin1水平。结果与模型组比较,荭草苷可明显改善大鼠血流动力学指标,降低乳酸脱氢酶和肌酸激酶水平,明显改善大鼠心肌细胞超微结构变化,同时能降低心肌细胞凋亡水平,增加细胞内自噬体数量以及自噬相关蛋白LC3Ⅱ和Beclin 1表达水平。结论荭草苷对大鼠心脏缺血/再灌注损伤具有明显保护作用,其机制可能与降低心肌细胞凋亡及增强细胞自噬有关。

Aim To investigate the role of autophagy in the cardioprotection by orientin and the relative molecular mechanisms in cell apoptosis and autophay. Methods The isolated ischemia reperfusion( I / R) heart model was built firstly. The Experiments were divided into seven groups: control group,I / R group,different concentrations of orientin-treated group( 1. 0 mg ·kg^(-1),2. 0 mg·kg^(-1),4. 0 mg·kg^(-1)),autophagy inhibitor group and resveratrol group. Hemodynamic index were recorded by Power Lab,the activities of myocardial enzymes were detected through Biochemical analyzer,the ultrastructure changes and autophagosomes in myocardial cells were detected by electron microscopy,the apoptosis was detected by TUNEL,and LC3 and Beclin1 protein levels of left ventricle were measured by Western blot. Results Orientin at middle and high concentrations( 2 and 4 mg·kg^(-1)) induced autophagy shown by increased the number of autophagosomes,LC3-Ⅱ / LC3-Ⅰ ratio and Beclin 1 expression after ischemia-reperfusion. The induction of autophagy by orientin was correlated with enhanced cardiac function and decreased apoptosis. But wortmannin,a kind of autophagy inhibitor,significantly attenuated the orientin-induced autophagy and increased apoptosis. Conclusion The cardioprotection of orientin against myocardial ischemia reperfusion injury may be mediated through the inhibition of apoptosis and induction of autophagy.