| Objective:The purpose of this study was to explore the protective effects of Curculigo orchioides and Epimedium("Er-xian")on myocardial ischemia-reperfusion injury(MIRI)in SD rats,and whether the mechanism of protection was associated with mitophagy.Methods:1.Establish I/R model and take material: 60 male SD rats weighing 240-260 g were selected.The animals were randomly divided into 5 groups including the control group(Control group),the ischemia-reperfusion group(I/R group),the low-dose "Er-xian" group(Er-xian 2.5:2.5g/Kg),the medium-dose "Er-xian" group(Er-xian 5:5g/Kg),and the high-dose "Er-xian" group(Er-xian 10:10g/Kg),with 12 rats in each group.The intragastric gavage were continued for 14 days before I/R model established.And the animals ingested food freely during the gavage period.After two hours of the last gavage of the rats,the left anterior descending coronary artery were ligated for 0.5 hours and reperfusion for 2 hours.Abdominal aorta blood and heart tissue of rats were taken for using.2.Experimental Methods: The myocardial infarct size of SD rats was detected by TTC staining.The serum expression level of CK-MB,LDH-a were determined by ELISA.The pathological injury of myocardial tissues was observed by HE staining.The cardiomyocyte apoptosis was detected by TUNEL staining.Western blot was used to assess the expression levels of autophagy proteins LC3-II/LC3-I and Beclin1,and the expression levels of mitophagy-related proteins PINK1,Parkin.Results:1.Effects of "Er-xian" on myocardial infarction size: The proportion of myocardial infarct size were significantly higher in the I/R group.After the intervention of "Er-xian",the proportion of myocardial infarct size were decreased,which appeared a dose-dependent effect.2.Effects of "Er-xian" on the expression of myocardial injury markers CK-MB and LDH-a: CK-MB and LDH-a levels were markedly raised in the I/R group,while the levels of CK-MB and LDH-a were declined after the intervention of Er-xian in a dose-dependent manner.3.Effects of "Er-xian" on pathological injury of myocardial tissue: I/R group presented obvious damage,including disorganized myocardial fibers,edema and vacuolar degeneration of myocardial cells,dilated vessels and infiltration of a great number of inflammatory cells.Compared with I/R group,"Er-xian" significantly reduced myocardial injury.Myocardial fibers disorder and vacuolar degeneration,the infiltration of inflammatory cells,and the damage of the nucleus were ameliorated.4.Effects of "Er-xian" on the apoptosis of cardiomyocytes: The apoptosis rate of cardiomyocytes were significantly added in the I/R group.After "Er-xian" intervention,the apoptosis rate were significantly reduced,and appeared a dose-related effect.5.Effects of "Er-xian" on the expression of Beclin1 and LC3-II/LC3-I autophagyrelated proteins: I/R obviously activated autophagy,and the expression of Beclin1 and LC3-II/LC3-I were enhanced.Compared with the I/R group,the "Er-xian" pretreatment inhibited the over-activation of autophagy and decreased the expression of Beclin1 and LC3-II/LC3-I.6.Effects of "Er-xian" on the expression of PINK1 and Parkin mitophagy-related proteins: The expression of PINK1 and Parkin mitophagy-related proteins were increased in I/R group."Er-xian" reduced the expression of PINK1 and Parkin mitophagy-related proteins and inhibited mitophagy.Conclusion:1."Er-xian" protected SD rats from MIRI by inhibiting autophagy,reducing myocardial pathological injury and decreasing myocardial infarct size,markers of myocardial injury and apoptosis rate.2."Er-xian" reduced myocardial reperfusion injury in rats by inhibiting mitophagy which was related to the PINK1/Parkin signaling pathway. |
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