佛手苷内酯对H9C2心肌细胞缺氧复氧损伤的保护作用及机制研究

Protective effect and mechanism of bergeninolide on hypoxia-reoxygenation injury of H9C2 cardiomyocytes

目的:研究佛手苷内酯(bergapten,BG)预处理对心肌缺血再灌注损伤的保护作用及机制。方法:将H9C2心肌细胞系传代培养,随机分为正常对照组、缺氧复氧模型组、佛手苷内酯低、中、高浓度预处理组(浓度为5、10、20μmol/L),通过缺氧4 h、复氧24 h模拟构建心肌缺血再灌注模型,采用CCK-8法检测细胞活性,按试剂盒方法检测LDH、CKMB漏出量,ELISA法检测SOD、MDA水平,流式细胞学检测细胞凋亡水平,Western blot法检测IL-6、TNF-α和p-PI3K、p-Akt的蛋白表达。结果:与H/R组比较,5、10、20μmol/L佛手苷内酯均能显著缓解缺氧复氧诱导的细胞存活率下降,减少心肌细胞凋亡,降低LDH、CKMB漏出量,提高SOD活性,下调MDA水平,抑制TNF-α、IL-6的释放,促进p-PI3K、p-AKT的表达,并有浓度依赖性。结论:佛手苷内酯预处理对心肌细胞缺氧复氧损伤有保护作用,并且可能与PI3K/Akt信号通路有关。

Objectives:To investigate the protective effect of bergapten preconditioning on myocardial ischemia-reperfusion injury and its effect on PI3 K/Akt signal pathway.Methods:Cultured H9 C2 cell line of cardiomyocytes was treated with hypoxia and reoxygenation to imitate the model of myocardial ischemia and reperfusion,and was randomly divided into 5 groups:normal control group,anoxia-reoxygenation group,anoxia-reoxygenation+bergapten pretreatment group(the dose was 5,10,20μmol/L),the cytotoxicity was detected by CCK-8 assay;the supernatant was extracted to detect LDH,CK-MB,SOD,MDA by biochemical techniques;the apoptotic rate was calculated by flow cytometry;the expression of IL-6,TNF-α,p-PI3 K,p-Akt of cardiomyocytes was detected by Western blot assay.Results:Compared with HR group,bergapten,at 5,10,20μmol/L,significantly alleviated the decrease of cell survival rate induced by hypoxia and reoxygenation,reduced cardiomyocyte apoptosis,decreased the leakage of LDH,CK-MB,increased the activity of SOD,down-regulated the level of MDA,inhibited the release of TNF-α,IL-6,and promoted the expression of p-PI3,p-AKT,which was concentration-dependent.Conclusion:Preconditioning with bergapten can reduce myocardial ischemia-reperfusion injury through PI3 K-Akt signal transduction.