二甲双胍对下咽癌Fadu细胞的增殖抑制作用及其对化疗药物敏感性影响的研究

Metformin inhibits the proliferation of hypopharyngeal carcinoma Fadu cells and enhances the chemotherapeutic sensitivity of cells

目的 研究二甲双胍对下咽癌Fadu细胞的作用及其对常用化疗药物敏感性的影响.方法 不同浓度二甲双胍干预Fadu细胞不同时间,四甲基偶氮唑蓝（MTT）比色法测细胞的增殖情况及5 mmol/L的二甲双胍分别联合不同浓度的顺铂、氟尿嘧啶（5-Fu）及紫杉醇作用不同时间细胞增殖情况的变化,流式细胞术检测细胞周期变化,免疫细胞化学检测二甲双胍主要作用靶点一磷酸腺苷活化蛋白激酶（AMP-dependent/activated protein kinase,AMPK）及P21蛋白表达水平的变化.结果 二甲双胍能够抑制Fadu细胞增殖,并且呈浓度-时间依赖性.多因素方差分析显示：药物浓度及作用时间对细胞存活率均有影响（F=97.411,F=323.251,P值均＜0.01）.二甲双胍可诱导Fadu细胞发生G1期阻滞,并且使细胞中AMPK及P21蛋白表达水平均升高.二甲双胍联合顺铂、紫杉醇与单用化疗药物相比,Fadu细胞的存活率下降,差异有统计学意义（P均＜0.01）,二者与二甲双胍联合作用Fadu细胞48 h的联合作用指数（combined index,CI）值分别为0.43、0.37;然而,二甲双胍与5-Fu联合无明显增殖抑制作用,联合作用48 h后CI值为1.15.结论 二甲双胍对Fadu细胞的增殖有抑制作用,这与其诱导的细胞周期G1期阻滞相关.并且,二甲双胍至少部分通过AMPK及P21相关的信号分子发挥作用.另外,在Fadu细胞中,二甲双胍对化疗药物顺铂及紫杉醇具有协同作用,但却拮抗5-Fu的细胞毒性作用.

Objective To investigate the role of metformin on the growth inhibition induced by chemotherapeutic agents in hypopharyngeal carcinoma Fadu cells.Methods Fadu cells were treated with different concentrations of metformin for different time or treated with different concentrations of cisplatin,5-fluorouracil or paclitaxel with or without metformin 5 mmol/L.MTT assay was used to evaluate the influence of metformin on the proliferation of Fadu cells.Cell-cycle was analyzed by flow cytometry.The expressions of AMP-dependent/activated protein kinase （AMPK） and P21 were examined by immunocytochemistry.Results Metformin inhibited the proliferation of Fadu cells in a dose-and timedependent manner.Flow cytometry showed that cell cycle arrest in G1 phase was induced by metformin in Fadu cells.Immunocytochemistry showed the expressions of both AMPK and P21 in cells treated with metformin were higher than those in cells untreated with metformin.The growth inhibition of cells induced by cisplatin or paclitaxel but not 5-fluorouracil was enhanced by metformin.The combined indexes of cisplatin/ paclitaxel/5-fluorouracil and metformin for 48 h were 0.43,0.37,and 1.15,respectively.Conclusions Metformin may inhibit the proliferation of Fadu cells by inducing the cell cycle arrest in G1 phase mediated in part by AMPK and P21.Metformin enhances the sensitivity of Fadu cells to cisplatin and paclitaxel.