利多卡因对肺癌A549细胞增殖、侵袭及AMPK/mTOR/4EBP1信号通路的影响

Effects of lidocaine on proliferation,invasion and AMPK/mTOR/4EBP1 signaling pathway of lung cancer A549 cells

目的探讨利多卡因对肺癌A549细胞增殖、侵袭及单磷酸腺苷活化蛋白激酶(AMPK)/哺乳动物雷帕霉素靶蛋白(mTOR)/真核细胞翻译起始因子4E结合蛋白1(4EBP1)信号通路的影响。方法设肺癌A549细胞组,5-氟尿嘧啶组(50μmol/L),利多卡因低剂量组(100μmol/L)、高剂量组(200μmol/L),以上各组每组设6个平行孔,培养72 h。试验结束后,采用细胞计数试剂盒8(CCK-8)法测定细胞增殖水平,transwell小室测定细胞侵袭水平,实时荧光定量PCR及蛋白免疫印迹法测定细胞AMPK、mTOR、4EBP1 mRNA和蛋白,以及磷酸化mTOR(p-mTOR)、磷酸化4EBP1(p-4EBP1)蛋白表达水平。结果与肺癌A549细胞组比较,5-氟尿嘧啶组、利多卡因低剂量组、利多卡因高剂量组吸光度值、存活率及mTOR、4EBP1 mRNA和蛋白、p-mTOR蛋白、p-4EBP1蛋白表达水平降低,穿膜数减少(P<0.05),AMPK mRNA和蛋白表达水平升高(P<0.05);与5-氟尿嘧啶组比较,利多卡因低、高剂量组吸光度值、存活率及mTOR、4EBP1 mRNA和蛋白、p-mTOR蛋白、p-4EBP1蛋白表达水平升高,穿膜数增加(P<0.05),AMPK mRNA和蛋白表达水平降低(P<0.05);与利多卡因低剂量组比较,利多卡因高剂量组吸光度值、存活率及mTOR、4EBP1 mRNA和蛋白、p-mTOR蛋白、p-4EBP1蛋白表达水平降低,穿膜数减少(P<0.05),AMPK mRNA和蛋白表达水平升高(P<0.05)。结论利多卡因对肺癌A549细胞增殖、侵袭具有明显抑制作用,其机制可能与利多卡因通过激活AMPK表达进而抑制mTOR/4EBP1信号通路的激活有关。

Objective To investigate the effects of lidocaine on the proliferation and invasion of lung cancer A549 cells and the signaling pathway of adenosine monophosphate activated protein kinase(AMPK)/mammalian target of rapamycin(mTOR)/eukaryotic translation initiation factor 4E binding protein 1(4EBP1).Methods Lung cancer A549 cell group,5-fluorouracil group(50μmol/L),low-and high-dose lidocaine groups(100,200μmol/L respectively)were set up.Six parallel samples were set for each group of the above groups and cultured for 72 h.At the end of the experiment,cell counting kit 8(CCK-8)assay was used to determine cell proliferation,cell invasion was measured by transwell chamber,real time fluorescent quantitative PCR and Western blot were used to determine the mRNA and protein expression levels of AMPK,mTOR and 4EBP1,phosphorylated mTOR(p-mTOR)and phosphorylated 4EBP1(p-4EBP1)protein.Results Compared with the lung cancer A549 cell group,the absorbance value,survival rate,number of transmembrane,mTOR,4EBP1 mRNA and protein,p-mTOR,p-4EBP1 protein expression levels in the 5-fluorouracil group,low-and high-dose lidocaine groups were decreased,AMPK mRNA and protein expression levels were increased(P<0.05).Compared with 5-fluorouracil group,the absorbace value,survival rate,number of transmembrane,mTOR,4EBP1 mRNA and protein,p-mTOR,p-4EBP1 protein expression levels were increased in the low-and high-dose lidocaine groups,AMPK mRNA and protein expression levels were decreased(P<0.05).Compared with the low-dose lidocaine group,the absorbance value,survival rate,number of transmembrane,mTOR,4EBP1 mRNA and protein,p-mTOR,p-4EBP1 protein expression levels of the high-dose lidocaine group were decreased,AMPK mRNA and protein expression levels were increased(P<0.05).Conclusion Lidocaine has a significant inhibitory effect on the proliferation and invasion of lung cancer cell line A549,and its mechanism may be related to the inhibition of mTOR/4EBP1 signaling pathway activation by activating AMPK.