栀子苷抗H_2O_2诱导人脐静脉内皮细胞凋亡的实验研究

目的探讨栀子苷对氧化应激诱导人脐静脉内皮细胞(HUVEC)凋亡的抑制作用及可能机制。方法体外培养的HUVEC细胞用不同浓度的栀子苷药物预处理24h后,加入终浓度为500μmol·L-1H2O2氧化损伤12h。荧光染色法观察细胞内DNA损伤情况,流式细胞术检测细胞凋亡率和线粒体膜电位的变化,Western blot检测Bcl-2和Bax蛋白的表达,酶免法测定caspase-3蛋白酶的活性,Real-time PCR检测超氧化物歧化酶(Mn-SOD)和谷胱甘肽过氧化物酶(GPx)的mRNA表达。结果与模型组相比,不同浓度栀子苷(12.5、25、50mg·L-1)可以明显改善H2O2对细胞内的DNA氧化损伤,降低细胞凋亡率,下调Bax蛋白表达,逐步恢复Bcl-2/Bax表达比例和线粒体膜电位的改变,降低caspase-3蛋白酶活性,上调细胞内Mn-SOD和GPx的表达,但对Bcl-2的表达无影响。结论栀子苷可以抑制H2O2诱导的HUVEC细胞凋亡,其机制可能与调节线粒体应激途径和发挥抗氧化损伤功能有关。

Effect and possible mechanism of geniposide on apoptosis of HUVECs induced by H2O2

Aim To investigate the effect and possible mechanism of geniposide on apoptosis of HUVECs induced by H2 O2.Methods HUVECs were pre-incubated with various doses geniposide(12.5,25,50 mg· L -1 ) for 24 h,then exposed to H 2 O2 with concentration of 500μmol·L -1 for 12 h.The morphology changes of chromosome were observed by invert fluorescent microscope.Apoptosis rate and△Ψm were measured byflow cytometry.The protein expressions of Bcl-2 and Bax were determined by Western blot method.The activation of caspase-3 was detected with the caspase-3 activity assay kit.The mRNA expressions of Mn-SOD and GPx were measured by real-time PCR.Results In comparision with the control(cells treated with H 2 O2 but not geniposide) ,the DNA injury of the cells was attenuated,apoptosis rate,△Ψm,protein expression of Bax and activity of caspase-3 decreased,Bcl-2/Bax protein expression rate and expressions of Mn-SOD and GPx increased in a geniposide-dose-dependent manner,but no narked change occurred in Bcl-2 level. Conclusion Geniposide could inhibit the apoptosis of HUVECs induced by H2 O2 which might be correlated with regulation of apoptosis in the path of mitochondrion and regulation of Mn-SOD and GPx expression.