解毒消癥饮对肝癌侧群细胞增殖和相关因子表达的影响

Effect of Jiedu Xiaozheng Yin on expression of cancer stem cells-associated genes in human hepatoma hep3B cell line

目的从肝癌干细胞角度探讨解毒消癥饮抑制肝癌的机制。方法解毒消癥饮乙酸乙酯提取物(EE-JXY)体外干预Hep3B细胞后采用MTT检测细胞存活率;平板克隆实验检测细胞的克隆形成能力;流式细胞仪分析细胞中侧群(SP)细胞的含量;RT-PCR检测干细胞相关基因CD133、Oct4、Nanog和Sox2mRNA的表达。不同浓度组细胞存活率值比较采用单因素方差分析。结果与空白组相比,EE-JXY剂量和时间依赖性抑制Hep3B细胞的存活,0.25mg/ml浓度干预24h后,抑制率达34.37﹪(P<0.01)。克隆形成能力也受到明显抑制(吸光度从0.45±0.06下降至0.29±0.03,P<0.05)。EE-JXY组SP细胞的含量显著降低(从2.27﹪下降至0.8﹪),CD133、Oct4、Nanog和Sox2mRNA的表达均受到抑制。结论 EE-JXY能显著抑制Hep3B细胞的增殖,可能的机制与其下调干细胞相关因子CD133、Oct4、Nanog、Sox2mRNA的表达及SP的比例,干扰肝癌干细胞自我更新有关。

Objective To investigate the effect of Jiedu Xiaozheng Yin on proliferation of Hep3B cell line, and explore the underlying mechanism from the aspect of cancer stem cells. Method Ethyl acetate extraction from Jiedu Xiaozheng Yin （EE-JXY） was prepared. Hep3B cells were cultured for 24 h in the present or absence of EE-JXY. The cell viability and colony formation capacity were measured by MTT and flat colony formation assay, respectively. The percentage of side population （SP） cells was analyzed by a flow cytometer. The mRNA levels of CD133, Oct4, Nanog and Sox2 were examined using reverse transcription PCR. Results Compared with control group, the cell viability, colony formation capacity and the percentage of SP cells of Hep3B ceils in EE-JXY group all decreased remarkably. And the mRNA levels of CD133, Oct4, Nanog and Sox2 were downregulated. Conclusion EE-JXY remarkably inhibited the proliferation of Hep3B cells via inhibiting CD133, Oct4, Nanog and Sox2 mRNA expression and decreasing the proportion of SP cells of liver cancer.