摘要
目的:探讨丹参多酚酸盐(salvianolate,SAL)对过氧化氢(hydrogen peroxide,H_(2)O_(2))诱导H9c2心肌细胞凋亡与自噬的影响及其机制。方法:以100μmol/L的H_(2)O_(2)干预对数生长期H9c2细胞4 h,建立心肌细胞氧化应激损伤模型,设H_(2)O_(2)组,SAL50、100、200 mg/L组,另取对数生长期H9c2细胞设为正常组。药物干预24 h后,CCK-8法检测细胞增殖率,流式细胞术检测细胞凋亡,GFP-LC3质粒转染后观察自噬小体,Western blot法检测H9c2细胞蛋白激酶B(protein kinase B,Akt)、磷酸化Akt(phosphorylation Akt,p-Akt)、半胱氨酸蛋白酶3(cysteinyl aspartate specific proteinase 3,Caspase-3)、B细胞淋巴瘤2(B cell lymphoma-2,Bcl-2)、Bcl-2相关X蛋白(Bcl-2 related X protein,Bax)、磷酸化哺乳动物雷帕霉素靶蛋白(phosphorylation mam-malian target of rapamycin,p-mTOR)、Beclin1、微管相关蛋白1轻链3(microtubule-associated protein 1 light chain 3,LC3)、P62蛋白表达。结果:与H_(2)O_(2)组比较,SAL 100、200μg/mL组H9c2细胞增值率显著升高而凋亡率显著降低(P<0.01),细胞内自噬小体数量减少,p-Akt、p-mTOR、bcl-2表达明显上调而Caspase-3、Bax、Beclin1、LC3-I、LC3-Ⅱ、P62表达明显下调(P<0.05或P<0.01),p-Akt/Akt、bcl-2/Bax比值升高而LC3-Ⅱ/LC3-I比值下降(P<0.01)。结论:SAL对H_(2)O_(2)所致H9c2心肌细胞凋亡与自噬具有抑制作用,其机制可能与激活Akt/mTOR/Beclin1通路而抑制促凋亡、促自噬相关蛋白表达和活化有关。
Objective:To discuss the effects of SAL on the apoptosis and autophagy of H9c2 myocardial cells induced by H_(2)O_(2)and its mechanism.Methods:H_(2)O_(2),100μmol/L,was used to intervene H9c2 cells in logari thmic growth phase for four hours,to establish oxidative stress injury models of cardiomyocytes,and the models were divided into H_(2)O_(2)group,SAL group of 50,100 and 200 mg/L,besides,H9c2 cells in logarithmic growth phase were selected as the normal group.After drug intervention for 24 hours,CCK-8 method was used to detect cell proliferation rate,flow cytometry to measure cellular apoptosis,to observe autophagosome after GFP-LC3 plasmid transinfection,Western blot method to measure the expressions of Akt,p-Akt,Caspase-3,Bcl-2,Bax,p-mTOR,Beclin1,LC3 and P62 protein in H9c2 cells.Results:Compared with H_(2)O_(2)group,the proliferation rate of H9c2 cells was increased while the apoptosis rate was significantly reduced in SAL groups of 100 and 200μg/mL(P<0.01),the number of autophagosome was lowered in the cells,the expressions of p-Akt,p-mTOR and Bcl-2 were up-regulated apparently while the expressions of Caspase-3,Bax,Beclin1,LC3-1,LC3-Ⅱand P62 were down regulated remarkably(P<0.05 or P<0.01),the ratios of p-Akt/Akt,Bcl-2/Bax were increased while LC3-Ⅱ/LC3-I ratio was decreased(P<0.01).Conclusion:SAL owns inhibitory effects on the apoptosis and autophagy of H9c2 myocardial cells induced by H_(2)O_(2),and its mechanism might be related to the activation of Akt/mTOR/Beclin1 pathway,and inhibiting the expressions and activation of pro-apoptosis and pro-autophagy related proteins.
作者
李兵
LI Bing(Handan Central Hospital,Handan 056001,China)
出处
《西部中医药》
2022年第10期29-34,共6页
Western Journal of Traditional Chinese Medicine
基金
河北省医学科学研究课题(20191845)。
作者简介
李兵(1978-),男,硕士学位,副主任医师。研究方向:急诊医学。
