二甲双胍对人胆管癌RBE细胞的作用及其机制

目的：探讨二甲双胍对人肝胆管癌细胞的增殖、凋亡和细胞周期的影响及机制。 方法：将人肝胆管癌RBE细胞分别用二甲双胍、Compound C（AMPK抑制剂）、二甲双胍+ Compound C处理，以未处理的RBE细胞作为空白对照，分别用MTT法检测细胞增殖、流式细胞仪检测细胞凋亡和细胞周期、Western blot检测细胞AMPK/mTOR通路蛋白的表达。 结果：与空白对照组比较，二甲双胍作用后的RBE细胞存活率降低；细胞凋亡率升高；G0/G1期比例增加，而S期比例减少；磷酸化AMPK（p-AMPK）蛋白表达上调，而磷酸化mTOR（p-mTOR）蛋白表达明显下调，差异均有统计学意义（均P<0.05）。二甲双胍与Compound C联合作用RBE细胞后，二甲双胍的上述作用均被取消，各项指标与空白对照组差异均无统计学意义（均P>0.05）。单独的Compound C作用RBE细胞后，各项指标未见明显改变，与空白对照组差异均无统计学意义（均P>0.05）。 结论：二甲双胍能抑制人胆管癌RBE细胞的增殖、促进凋亡和细胞周期阻滞，该作用可能其与激活AMPK从而抑制mTOR下游效应分子有关。

Effect of metformin on human cholangiocarcinoma RBE cells and its mechanism

Objective: To investigate the effect of metformin on cell proliferation, apoptosis and cell cycle in human cholangiocarcinoma cells and the mechanism. Methods: Human cholangiocarcinoma RBE cells were exposed to metformin, Compound C (AMPK inhibitor) or metformin plus Compound C respectively, using the untreated RBE cells as blank control. The cell proliferation was determined by MTT assay, apoptosis and cell cycle were measured by flow cytometry, and the expressions of proteins associated with AMPK/mTOR signaling pathway were detected by Western blot analysis, respectively. Results: In RBE cells treated with metformin compared with blank control, the survival rate was decreased, apoptosis rate was increased, the ratio of G0/G1 cells was increased while ratio of S cells was decreased, and the phosphorylated AMPK (p-AMPK) level was increased while the phosphorylated mTOR (p-mTOR) was decreased, and all differences had statistical significance (all P<0.05). The above effects exerted by metformin were all abolished by the combination treatment of Compound C, and the differences in all tested indexes had no statistical significance compared with blank control group (all P>0.05). All the indexes in RBE cells treated with Compound C alone had no statistical significance compared with blank control group (all P<0.05). Conclusion: Metformin can inhibit cell growth and promote apoptosis and cell cycle arrest in human cholangiocarcinoma RBE cells, and the mechanism may be associated with its activating AMPK and thereby suppressing mTOR downstream effector molecules.