参附注射液诱导H9C2大鼠心肌细胞热休克蛋白22表达并减轻缺氧/复氧损伤的研究

Shenfu Injection induces heat shock protein 22 expression and alleviates anoxia/reoxygenation injury in H9C2 rat cardiomyocytes

目的观察参附注射液(SFI)对缺氧/复氧(A/R)损伤大鼠心肌细胞热休克蛋白22(HSP22)表达的影响,探讨SFI对心肌缺血再灌注(I/R)损伤的保护作用机制。方法体外培养H9C2大鼠心肌细胞,予以A/R处理模拟心肌I/R损伤,分别以1.5%、2.5%和3.5%的SFI预处理心肌细胞,采用四甲基偶氮唑蓝(MTT)法检测不同复氧时间对心肌细胞存活率的影响,探索最佳复氧时间,流式细胞术检测各组心肌细胞凋亡情况,荧光定量RT-PCR检测各组心肌细胞HSP22 mRNA表达,Western blot检测HSP22蛋白的表达情况。结果复氧2 h、4 h、8 h后H9C2心肌细胞存活率均显著低于对照组(均P<0.05);A/R+2.5%SFI组细胞凋亡率显著低于A/R组、A/R+1.5%SFI组、A/R+3.5%SFI组(P<0.05)。A/R+2.5%SFI组、A/R+3.5%SFI组HSP22蛋白相对表达量均显著高于A/R组(P<0.05);A/R+3.5%SFI组HSP22蛋白相对表达量显著高于A/R组、A/R+1.5%SFI组、A/R+2.5%SFI组(P<0.05)。结论SFI在H9C2大鼠心肌细胞A/R损伤中可诱导HSP22表达上调,并减轻A/R损伤诱导的细胞凋亡。

Objective To observe the effect of Shenfu Injection(SFI)on the expression of heat shock protein 22(HSP22)in myocardial cells injured by anoxia/reoxygenation(A/R),and to preliminarily explore the protective mechanism of SFI on ischemia/reperfusion(I/R)injury.Methods The H9C2 rat cardiomyocytes were cultured in vitro and treated with A/R to simulate myocardial I/R injury.The cardiomyocytes were pretreated with 1.5%,2.5%,and 3.5%SFI.The tetramethylazazole blue method(MTT)was used to detect the effect of different reoxygenation times on H9C2 cell viability to explore the optimal reoxygenation time.The flow cytometry was used to detect cardiomyocyte apoptosis.RT-qPCR was used to detect the level of HSP22 mRNA expression.The Western blot was used to detect the level of HSP22 protein expression.Results The suvival rate of the H9C2 cells after 2,4,8 h of reoxygenation was significantly lower than that of the control group(P<0.05 or P<0.01).The cell apoptotic rate in the A/R+2.5%SFI group was significantly lower than that of the A/R group,A/R+1.5%SFI group and A/R+3.5%SFI group(P<0.01).The relative expression level of HSP22 protein in the A/R+2.5%SFI group and A/R+3.5%SFI group was significantly higher than that in the A/R group(P<0.01).The relative expression of HSP22 protein in the A/R+3.5%SFI group was significantly higher than that in the A/R group,A/R+1.5%SFI group and A/R+2.5%SFI group(P<0.05).Conclusion SFI could induce the up-regulaion of HSP22 expression in H9C2 rat cardiomyocytes A/R injury and reduces apoptosis induced by A/R injury.