二氯乙腈对HepG2细胞凋亡的影响

翟璐; 罗皓; 叶美洁; 李永崇; 黄嘉华; 丁梦珂; 刘振龙; 杨慧; 唐焕文

doi:10.13421/j.cnki.hjwsxzz.2016.04.001

二氯乙腈对HepG2细胞凋亡的影响

Effect of Dichloroacetonitrile on the Apoptosis of HepG2 Cells

摘要

摘要:
目的探讨饮水消毒副产物二氯乙腈（dichloroacetonitrile,DCAN）对HepG2细胞凋亡的影响及相关的调控机制。
方法二甲基亚砜（DMSO）溶解DCAN，以DMSO处理人肝癌细胞（HepG2细胞）为对照组，分别以50 μmol/L、100 μmol/L、200 μmol/L、400 μmol/L DCAN染毒HepG2细胞为处理组，24 h后通过AnnexinV/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，Western blotting技术检测HepG2细胞pro-Caspase 3和P53蛋白的表达情况。
结果与对照组相比，400 μmol/L DCAN处理组可诱导HepG2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HepG2细胞后pro-Caspase 3和P53蛋白含量呈下降趋势，当染毒浓度达到200 μmol/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HepG2细胞凋亡。

Abstract:
Objectives To investigate the effect of dichloroacetonitrile (DCAN) on apoptosis in HepG2 cells and its related regulatory mechanism.
Methods DCAN was dissolved with dimethyl sulfoxide (DMSO), HepG2 cells were exposed to 50 μmol/L, 100 μmol/L, 200 μmol/L and 400 μmol/L DCAN for 24 h, and HepG2 cells treated with DMSO only were served as the control. Apoptosis was tested by Annexin V/PI double staining and determined by flow cytometry. The activity of Caspase 3 was measured by fluorometric assay. The expression of pro-Caspase and P53 protein was detected by western blotting.
Results Compared with the control group, the proportion of apoptotic cells in the 400 μmol/L DCAN-treated group was increased (P<0.05), the enzyme activity of Caspase 3 was significantly increased in all DCAN-treated groups with the increase of exposure doses, and the expression of pro-Caspase and P53 protein were down-regulated in the 200 μmol/L and 400 μmol/L DCAN-treated groups (P<0.05).
Conclusion High concentration of DCAN might induce the apoptosis of HepG2 cells by raising the activity of Caspase 3.

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