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目的探讨饮水消毒副产物二氯乙腈(dichloroacetonitrile,DCAN)对Hep G2细胞凋亡的影响及相关调控机制。方法二甲基亚砜(DMSO)溶解DCAN,以DMSO处理人肝癌细胞(Hep G2细胞)为对照组,分别以50μmol/L、100μmol/L、200μmol/L、400μmol/L DCAN染毒Hep G2细胞为处理组,培养24 h后通过Annexin V/PI流式细胞分析法检测细胞凋亡情况,用荧光测定法观察Caspase 3酶活性,蛋白质印迹法(Western blotting)技术检测Hep G2细胞pro-caspase 3和P53蛋白的表达情况。结果与对照组相比,400μmol/L DCAN处理组可诱导Hep G2细胞凋亡(P<0.05),各处理组细胞内Caspase 3酶活性随着染毒剂量的增加而显著升高(P<0.05)。DCAN作用于Hep G2细胞后pro-Caspase 3和P53蛋白含量呈下降趋势,当染毒浓度达到200μmol/L以上时,pro-Caspase 3和P53蛋白表达明显低于对照组(P<0.05)。结论高浓度的DCAN可通过激活Caspase 3诱导Hep G2细胞凋亡。
Objective To investigate the effect of dichloroacetonitrile (DCAN), a by-product of drinking water disinfection, on the apoptosis of Hep G2 cells and its related regulatory mechanism. Methods DCAN was dissolved in dimethylsulfoxide (DMSO) and Hep G2 cells were treated with DMSO. Hep G2 cells were treated with 50 μmol / L, 100 μmol / L, 200 μmol / L and 400 μmol / L DCAN, For the treatment group, the apoptosis of cells was detected by Annexin V / PI flow cytometry after cultured for 24 h, the activity of Caspase 3 was observed by fluorometry, the expression of pro-caspase 3 in Hep G2 cells was detected by Western blotting And P53 protein expression. Results Compared with the control group, the apoptosis of Hep G2 cells was induced by 400 μmol / L DCAN treatment (P <0.05). The activity of Caspase 3 in each treatment group was significantly increased with the dose of DCA (P <0.05) ). The expression of pro-Caspase 3 and P53 in Hep G2 cells decreased after DCAN treatment. When the concentration reached 200μmol / L, the expression of pro-Caspase 3 and P53 was significantly lower than that of the control group (P <0.05). Conclusion High concentration of DCAN can induce Hep G2 cell apoptosis by activating Caspase 3.