二甲双胍对人食管癌KYSE450细胞系增殖和凋亡的影响

Effects of metformin on proliferation and apoptosis of human esophageal cancer cells KYSE450

目的:探讨二甲双胍对人食管癌KYSE450细胞系增殖和凋亡的影响及其可能机制。方法:体外常规培养KYSE450细胞,采用MTT法检测不同浓度二甲双胍对KYSE450细胞的增殖抑制率,流式细胞术检测凋亡率,RT-PCR、Western blot检测药物作用后KYSE450细胞中4EBP1和S6K1 mRNA及蛋白表达的变化。结果:5、10、20和40 mmol/L二甲双胍对KYSE450细胞的增殖均有抑制作用,且呈剂量和时间依赖性(F剂量=470.513,F时间=743.696,F交互=22.359,P均<0.001)。5、10和20 mmol/L二甲双胍作用48 h后KYSE450细胞凋亡率显著增加,且呈浓度依赖性(F=282.076,P<0.001)。与对照组相比,5、10和20 mmol/L二甲双胍组KYSE450细胞中4EBP1、S6K1 mRNA和蛋白的表达量均明显下降(mRNA:F4EBP1=86.727,FS6K1=86.486,P均<0.001;蛋白:F4EBP1=43.137,FS6K1=118.924,P均<0.001)。20 mmol/L二甲双胍作用24、48和72 h后KYSE450细胞中4EBP1、S6K1 mRNA和蛋白表达亦降低(mRNA:F4EBP1=146.772,FS6K1=36.500,P均<0.001;蛋白:F4EBP1=73.900,FS6K1=92.950,P均<0.001)。结论:二甲双胍能够抑制人食管癌KYSE450细胞增殖并诱导凋亡,其机制可能与4EBP1和S6K1 mRNA及蛋白表达下调有关。

Aim： To investigate the effects of metformin on the proliferation and apoptosis of esophageal cancer KYSE450 cells and to explore the possible mechanism. Methods： KYSE450 cells were cultured in vitro and treated by metformin at different doses. The inhibition of cellular proliferation was determined by MTT. The apoptosis was assessed by flow cytometry. The mRNA and protein expression levels of 4EBP1 and S6K1 were detected by RT-PCR and Western blot. Results： Different doses of metformin could obviously inhibit the proliferation of KYSE450 in a dose-dependent and time-dependent manner（ Fdose= 470. 513,Ftime= 743. 696,Finteraction= 22. 359, P〈0. 001）,and the apoptosis rate of KYSE450 was increased in a dose-dependent manner（ F = 282. 076, P〈0. 001）. Compared with control group,the expression levels of4EBP1 and S6K1 mRNA and protein were down-regulated with the treatment of 5,10 and 20 mmol / L metformin（ mRNA：F4EBP1= 86. 727,FS6K1= 86. 486, P〈0. 001; protein： F4EBP1= 43. 137,FS6K1= 118. 924, P〈0. 001）. The expression levels of 4EBP1 and S6K1 mRNA and protein were down-regulated with the treatment of 20 mmol / L metformin for 24,48 and 72h（ mRNA： F4EBP1= 146. 772,FS6K1= 36. 500, P〈0. 001; protein： F4EBP1= 73. 900,FS6K1= 92. 950, P〈0. 001）. Conclusion： Metformin can inhibit the proliferation and induce apoptosis of esophageal cancer KYSE450 cells in vitro,probably by down-regulating the expressions of 4EBP1 and S6K1.