二甲双胍联合塞来昔布对肝癌细胞HepG2和Huh7增殖及凋亡的影响

Effects of metformin combined with celecoxib on the proliferation and apoptosis of hepatoma HepG2 and Huh7 cell lines

目的探讨二甲双胍联合塞来昔布对人肝癌细胞HepG2和Huh7增殖、凋亡的影响,并初步研究其作用机制。方法将肝癌细胞HepG2和Huh7分为对照组、二甲双胍组、塞来昔布组和联合用药组培养,CCK-8法检测细胞增殖情况;Hoechst33258荧光染色法观察细胞凋亡;划痕实验检测细胞迁移能力;Transwell侵袭小室实验检测细胞侵袭能力;蛋白印迹法检测AMPK、PI3K、Akt、mTOR蛋白表达情况。结果二甲双胍和塞来昔布作用于HepG2和Huh7细胞后,细胞逐渐圆缩,崩解和凋亡细胞增多,细胞增殖受明显抑制。划痕实验结果显示,二甲双胍和塞来昔布作用后细胞迁移能力明显下降(P<0.05)。Transwell小室结果显示,联合用药对HepG2和Huh7细胞侵袭能力具有显著抑制作用(P<0.05)。蛋白印迹结果显示,联合用药组HepG2细胞内AKT、AMPK、mTOR表达水平呈下降趋势,PI3K表达水平先下降,后上升;Huh7细胞内AKT、AMPK、PI3K、mTOR表达水平均呈下降趋势。结论二甲双胍可协同塞来昔布增强对HepG2和Huh7细胞的增殖、迁移和侵袭的抑制作用,其机制可能与抑制mTOR信号通路的表达相关。

Objective To explore the effects and the mechanism of metformin combined with celecoxib on the proliferation and apoptosis of hepatoma HepG2 and Huh7 cells.Methods Hepatoma cells HepG2 and Huh7 were divided into control group,metformin group,celecoxib group and combination medication group,CCK-8 assay was used to detect cell proliferation;Hoechst33258 staining method was used to investigate the cell apoptosis;wound healing test was used to detect cells migration ability;Transwell invasion chamber test was used to detect cell invasion ability;Western blotting was used to detect the expression of AMPK,PI3K,Akt,mTOR.Results After metformin and celecoxib treatment,HepG2 and Huh7 cells were gradually contracted,disintegrated and more apoptotic cells were noticed,and cell proliferation was significantly inhibited.The wound healing test results showed that the cell migration was significantly decreased(P<0.05)under metformin and celecoxib treatment.The results of the transwell invasion chamber test showed that the metformin and celecoxib treatment inhibited the invasion of HepG2 and Huh7 cells(P<0.05).The expression levels of AKT,AMPK,and mTOR were decreased in HepG2 cells in the combinational treatment group,and the expression level of PI3K was decreased and then increased;the expression levels of AKT,AMPK,PI3K,and mTOR in Huh7 cells were decreased.Conclusions Metformin can cooperate with celecoxib to enhance the inhibitory effect on the proliferation,migration and invasion of HepG2 and Huh7 cells.The mechanism may be related to the inhibition of the expression of mTOR signaling pathway.