染料木素逆转肝癌Bel-7402/5Fu细胞耐药性的机制研究

Mechanism of genistein reversing drug resistance in Bel-7402/5Fu cells by inhibiting autophagy

目的:探讨染料木素逆转肝癌Bel-7402/5Fu细胞耐药性的机制。方法:通过CCK-8法检测染料木素对肝癌Bel-7402/5Fu细胞的杀伤作用。通过蛋白印迹和qPCR法检测染料木素作用后自噬关键基因Beclin1和LC3的表达水平。通过蛋白印迹法检测自噬上游信号通路蛋白Akt和mTOR的活化水平。结果:染料木素能够抑制肝癌Bel-7402/5Fu细胞增殖,并呈剂量依赖性,IC50为30.0±5.9μg/mL。选取低浓度染料木素作用于Bel-7402/5Fu细胞后,再用5Fu处理细胞,发现细胞对5Fu的耐受能力降低。蛋白印迹结果显示,染料木素处理显著降低了细胞自噬关键基因Beclin1和LC3的表达水平,显著上调自噬抑制性通路PI3K/Akt/mTOR信号通路相关蛋白Akt和mTOR的活化水平。结论:染料木素能够降低肝癌Bel-7402/5Fu细胞对5Fu的耐受性,即染料木素处理逆转了肝癌Bel-7402/5Fu细胞的耐药性。该过程可能与染料木素通过上调PI3K/Akt/mTOR信号通路而抑制细胞自噬相关。

Objective:to investigate whether genistein can reverse the drug resistance of Bel-7402/5 Fu cells and whether this process is related to the inhibition of autophagy of Bel-7402/5 Fu cells.Methods:the killing effect of genistein on Bel-7402/5 Fu cells was detected by CCK-8 method.Bel-7402/5 Fu cells were treated with low concentration genistein and then treated with 5 Fu.The survival rate of the cells was determined by CCK-8 method.Detection of key autophagy gene Bec by Western blot and qPCR The expression level of lin1 and LC3.The activation levels of Akt and mTOR in the upstream signaling pathway of autophagy were detected by Western blot.Results:genistein inhibited the proliferation of Bel-7402/5 Fu cells in a dose-dependent manner,and the IC50 was 30±5.9μg/ml.Bel-7402/5 Fu cells were treated with lowconcentration genistein and then treated with 5 Fu.The results showed that the tolerance of cells to 5 Fu decreased.Western blot analysis showed that genistein treatment significantly reduced the surface of Beclin1 and LC3,the key genes of autophagy.Reach a level.Further studies showed that genistein treatment could significantly up-regulate the activation levels of PI3K/Akt/mTOR signaling pathway related proteins Akt and mTOR in autophagy inhibitory pathway.Conclusion:genistein can reduce the tolerance of Bel-7402/5Fu cells to 5-Fu,that is,genistein treatment can reverse the drug resistance of Bel-7402/5Fu cells.Further studies suggest that genistein may inhibit autophagy by up-regulating the PI3K/Akt/mTOR signaling pathway.