葛根素对过氧化氢诱导人心肌细胞损伤的影响

Effect of puerarin on human cardiomyocytes injury induced by H_(2)O_(2)

目的观察葛根素对过氧化氢(H_(2)O_(2))诱导的人心肌细胞的线粒体功能的影响,并探求其机制。方法培养人心肌细胞系AC16,高糖完全培养基培养内加入与H_(2)O_(2)等体积的磷酸缓冲盐溶液作为对照组;通过H_(2)O_(2)(200μmol/L)诱导建立心肌细胞损伤模型(模型组);并采用低、中、高剂量(50μmol/L、100μmol/L、200μmol/L)葛根素处理心肌细胞,给药48 h后,采用MTT法检测细胞活力;以Western blot检测Mzb1、磷酸化-Drp1/Drp1的蛋白表达;以一步法TUNEL细胞凋亡检测试剂盒检测细胞凋亡情况;以ATP检测试剂盒检测细胞内ATP的水平;通过脂质体转染siRNA沉默细胞内Mzb1的表达水平。结果与对照组相比,模型组细胞活力显著下降,细胞凋亡率明显升高,Mzb1表达量显著降低,ATP含量显著降低,磷酸化-Drp1/Drp1表达升高;与模型组相比,葛根素低剂量组、中剂量组和高剂量组AC16细胞中Mzb1及细胞活力有所恢复,细胞凋亡率显著降低,Mzb1表达明显升高,ATP含量显著升高,磷酸化-Drp1/Drp1表达明显下降;沉默Mzb1后,葛根素对细胞活力及磷酸化-Drp1/Drp1表达的影响均被阻断。结论葛根素通过Mzb1/Drp1通路恢复线粒体功能,改善H_(2)O_(2)诱导的心肌细胞损伤。

Objective To detect the effect and mechanism of puerarin on mitochondrial function of human cardiomyocytes induced by H_(2)O_(2).Methods AC16 human cardiomyocytes were cultured,the equal volume of PBS with H_(2)O_(2) was added into the high glucose complete medium as the control group,cells were induced by H_(2)O_(2)(200μmol/L)to establish a model of cardiomyocytes injury(model group),and treated with puerarin at low,medium and high doses(50μmol/L,100μmol/L,200μmol/L).After 48 h administration,the cell viability was detected by MTT assay.The protein expression of Mzb1 and phosphorylation-Drp1/Drp1 were detected by Western Blot.One step TUNEL apoptosis assay kit was used to measure apoptosis level.The content of intracellular ATP was detected by ATP detection kit.siRNA was transfected with lipofectamine to silence the expression of Mzb1.Results Compared with the control group,the cell viability of the model group was significantly decreased,the apoptosis rate was significantly increased,the expression of Mzb1 was significantly decreased,the ATP content was significantly decreased,and the expression of phosphorylation-Drp1/Drp1 was increased.Compared with the model group,Mzb1 and cell viability of AC16 cells in the low dose,middle dose,and high dose puerarin groups were recovered,the apoptosis rate was significantly decreased,the expression of Mzb1 was significantly increased,the ATP content was significantly increased,and the expression of phosphorylation-Drp1/Drp1 was significantly decreased.After silencing Mzb1,puerarin affected the cell vitality and phosphorylation-Drp1/Drp1 expression were blocked.Conclusion Puerarin ameliorates H_(2)O_(2)-induced cardiomyocyte injury and mitochondrial function through Mzb1/Drp1 signal pathway.