白杨素对5- 氟尿嘧啶诱导人肝癌Bel-7402 细胞凋亡的增敏机制研究

Chrysin sensitizes 5-fluorouracil-induced apoptosis in human liver cancer Bel-7402 cells

摘要:

目的 探讨白杨素对5- 氟尿嘧啶（5-FU）诱导肝癌Bel-7402 细胞凋亡的增敏作用。方法 白 杨素单独或联合5-FU 作用Bel-7402 细胞，用MTT 法检测细胞活力；流式细胞术检测细胞凋亡与周期分 布变化；Western blotting 检测Bcl-2、Bax、Bad 及Cleaved Caspase-3 蛋白。结果 50 ～ 250μmol/L 白杨 素作用Bel-7402 细胞48 h 呈浓度依赖性的抑制细胞活力，且100μmol/L 白杨素组细胞存活率为（82.261± 7.793）%。100μmol/L 白杨素单独作用24 h 对Bel-7402 细胞活力和凋亡均无影响（P >0.05）。与空白对照 组比较，100μmol/L 白杨素作用24 h 后，不同浓度5-FU（12.5、25.0 及50.0μg/ml）作用48 h 对Bel-7402 细胞活力的抑制作用增强（P <0.05）；25.0μg/ml 5-FU 诱导Bel-7402 细胞凋亡率增加（P <0.05）；G2/M 期 细胞比例升高（P <0.05）；促凋亡蛋白Bax、Bad 及Cleaved Caspase-3 蛋白表达水平上调（P <0.05）；抗凋亡 蛋白Bcl-2 表达水平下调（P <0.05）。结论 白杨素增强5-FU 诱导人肝癌Bel-7402 细胞凋亡，其机制可能 与线粒体凋亡通路激活有关。

Abstract:

Objective To investigate the effect of chrysin pretreatment on 5-fluorouracil (5-FU) -induced apoptosis human liver cancer cells. Methods Bel-7402 cells were treated with chrysin alone or jointly treated with 5-FU. The cell viability was assessed by MTT assay, the cell apoptosis and cell cycle were determined by flow cytometry (FCM) and the protein expressions of Bcl-2, Bax, Bad, and Cleaved Caspase-3 were analyzed by western blotting. Results Treatment of Bel-7402 cells induced dose-dependent cytotoxicity from 50 to 250μmol/L chrysin for 48 h and (82.261±7.793) % of the cell viability rate in the 100μmol/L chrysin group. The cell viability and apoptosis were no significantly different in Bel-7402 cells treated with 100μmol/L chrysin for 24 h alone comparing with the control group (P > 0.05). Compared with control group, after the pretreatment with 100 μmol/L chrysin for 24h, the cytotoxicity was enhanced by 5-FU (12.5, 25.0, and 50.0μg/ml) to Bel-7402 cells (P < 0.05); the rate of 5-FU-induced apoptosis was increased in 25.0μg/ml group (P < 0.05); accumulation of Bel-7402 cells in the G2/M phase was increased (P < 0.05); the expressions of apoptotic protein Cleaved Caspase-3, Bax, and, Bad upregulated (P < 0.05); the protein level of antiapoptotic protein Bcl-2 downregulated (P < 0.05). Conclusions Chrysin sensitizes 5-FUinduced apoptosis in Bel-7402 cells, which may be closely associated with the activation of mitochondrial apoptosis pathway.