二甲双胍在喉癌Hep-2细胞中对5-氟尿嘧啶、顺铂、紫杉醇的作用及机制探讨

Metformin's effect on 5-fluorouracil,cisplatin,paclitaxel in laryngocarcinoma Hep-2 cells

目的:检测5-氟尿嘧啶联合二甲双胍对喉癌Hep-2细胞增殖抑制作用及化疗敏感影响。探究二甲双胍联合顺铂、紫杉醇、5-氟尿嘧啶作用于Hep-2细胞AMPK通路信号分子的表达变化。方法:体外培养Hep-2细胞,二甲双胍联合5-氟尿嘧啶作用细胞72h,MTT检测增殖情况。二甲双胍联合顺铂、5-氟尿嘧啶、紫杉醇作用细胞72h,Western blot检测P21、Cyclin D1、AMPK-α表达。结果:5-氟尿嘧啶以及二甲双胍可抑制Hep-2细胞增殖。二甲双胍联合低浓度5-氟尿嘧啶,细胞生长抑制率升高;二甲双胍联合高浓度5-氟尿嘧啶生长抑制率无明显差异,两药在Hep-2细胞中相互拮抗。Western blot显示,顺铂、5-氟尿嘧啶增加细胞中上述蛋白表达,紫杉醇减少细胞中Cyclin D1表达。对比单独用药组,联用二甲双胍均引起Hep-2细胞中上述蛋白表达变化。结论:5-氟尿嘧啶抑制Hep-2细胞增殖,二甲双胍拮抗5-氟尿嘧啶对Hep-2细胞抗肿瘤效应,其拮抗作用及对顺铂、紫杉醇的影响部分通过AMPK、P21及Cyclin D1相关通路发生作用且与周期阻滞存在密切关系。

Objective：To investigate metformin＇s effect on chemosensitivity of chemotherapeutic drug 5-fluorouracil in laryngocarcinoma Hep-2cells.Investigate the variation trend of protein expression of AMPK pathway in the combined effect.Method：Laryngocarcinoma Hep-2cells were treated with different concentrations of 5-fluorouracil in vitro together with or without metformin for 72 h.Use MTT assay to investigate the influence on the inhibition rate to Hep-2cells.Hep-2cells were treated with cisplatin,5-fluorouracil or paclitaxel with or without metformin.Use Western blot assay to investigate the expression level of AMPKα,P21 or Cyclin D1 protein.Result：5-fluorouracil and metformin could inhibit the proliferation of Hep-2cells.5-fluorouracil in low concentration combined with metformin could increase the proliferation inhibition rate of Hep-2cells.In the circumstances of using 5-fluorouracil in high concentration with metformin,the cell proliferation inhibition rate of combining group makes no differences with the single-drug group.The combination of metformin and 5-fluorouracil produced an antagonism action in Hep-2cells.Western blot assay showed that metformin,cisplatin,5-fluorouracil could have caused the increase of expression level of AMPK-α,P21 and Cyclin D1 in Hep-2cells while Paclitaxel could have cause the decrease of expression level of Cyclin D1.Using combined drug could cause the change of protein expression.Conclusion：5-fluorouracil has been found to inhibit the proliferation of Hep-2cells.Metformin has an antagonism on the anticancer effect to 5-fluorouracil in Hep-2cells,and this antagonistic effect occurred partially through molecular signal pathways of AMPK-α,P21 and Cyclin D1 and it＇s significantly related to the cell cycle arrest.