三氧化二砷联合丹参酮胶囊抗肝癌细胞bel-7404的研究

Effects of a combination of arsenic trioxide and Tanshinone Capsule on inhibiting hepatoma bel-7404 cells

目的:探讨三氧化二砷(As2O3)联合丹参酮胶囊对体外培养的人肝癌bel-7404细胞增殖和凋亡的影响及其信号转导机制。方法:以不同浓度的As2O3、丹参酮胶囊分别组成单药组和联合组,另设空白对照组。采用CCK-8法检测增殖抑制率,Annexin V-FITC/PI双标法检测凋亡率,Western blot法检测Bcl-2、cleaved caspase-3等凋亡相关蛋白影响。最后检测MAPK的磷酸化水平分析凋亡信号转导机制,MAPK抑制剂实验进一步证实诱导凋亡的分子机制。结果:与单药组比较,两药联合可明显抑制细胞增殖、提高凋亡率(P<0.01);联合组可显著下调抗凋亡蛋白Bcl-2、XIAP、Survivin的表达,同时上调cleaved caspase-3、cleaved caspase-9、cleaved PARP促凋亡蛋白的表达。结论:As2O3联合丹参酮胶囊作用bel-7404细胞具有协同增效作用,其协同机制可能与线粒体途径和激活JNK、p38 MAPK信号通路有关。

Objective： To explore the effects of a combination of arsenic trioxide and Tanshinone Capsule on the proliferation and apoptosis of human bel-7404 cells in vitro and the mechanism of its potential signal transduction. Methods： Human bel-7404 cells were cultured in different drug groups： control group, single-drug groups with arsenic trioxide or Tanshinone Capsule respectively in varied concentrations, combination of the drugs group. The rate of proliferation and the rate of apoptosis were detected with CCK-8 analysis and Annexin V-FITC/PI method, respectively. Western blot method was used to detect the apoptosis-related proteins such as Bcl-2, cleaved caspase-3 and so on. The phosphorylation level of mitogenactivated protein kinase（MAPK） was measured to explore the signal transduction mechanism. Additionally, the inhibitor of MAPK further confirmed the apoptosis mechanism. Results： Compared with the single drug groups, the combination of the drugs group showed that could significantly inhibit the rate of proliferation and increase the rate of apoptosis（P0.01）. What＇s more, the expression of Bcl-2, XIAP, Survivin protein that an inhibitor of apoptosis was obviously down-regulated by the combination of the drugs, meanwhile the expression of cleaved caspase-3, cleaved caspase-9, cleaved PARP protein that a promotor of apoptosis was up-regulated by the combination of the drugs. Conclusion： The combination of arsenic trioxide and Tanshinone Capsule had a synergistic effect on human bel-7404 cells in vitro, which might be related to mitochondrial pathway and active JNK and p38 MAPK signaling pathway.