TOFA对人食管鳞癌Eca109和KYSE-450细胞生长的影响

Effects of TOFA on growth of Eca109 and KYSE-450 cells in human esophageal squamous cell carcinoma

目的:探讨5-十四烷氧基-2-呋喃酸(TOFA)对人食管鳞癌(ESCC)细胞Eca109和KYSE-450细胞增殖、周期和凋亡的影响。方法:将Eca-109细胞和KYSE-450细胞分为对照组(DMSO)和实验组(TOFA),细胞(4×10^(3) cells/100μl)接种于96孔板中,每个浓度设置5个复孔,培养24 h后,给予DMSO(对照)和不同浓度(1、3、5、10μg/ml)TOFA处理,继续培养24、48和72 h;MTT检测细胞增殖,流式细胞术检测细胞周期和凋亡,Western blot检测p21、Cleaved caspase-3表达水平及p-AKT、p-mTOR、p-4EBP1修饰水平,专用试剂盒检测细胞内游离脂肪酸。结果:与DMSO组比较,TOFA以浓度和时间依赖性方式抑制Eca109和KYSE-450细胞增殖(P均<0.05),处理48 h的IC_(50)分别为4.65和3.93μg/ml;实验组细胞G2/M期细胞百分比增加,细胞凋亡率增高,p21、Cleaved caspase-3蛋白表达水平上调(P均<0.05),p-AKT、p-mTOR、p-4EBP1修饰水平下调(P均<0.05)。结论:TOFA抑制人食管鳞癌细胞增殖、阻滞细胞周期并促进细胞凋亡,其机制可能与其抑制AKT/mTOR/4EBP1信号通路有关。

Objective:To investigate the effects of 5-tetradecanoxy 2-furanic acid(TOFA)on cell proliferation,cell cycle and apoptosis of esophageal squamous cell carcinoma(ESCC)cells.Methods:Eca-109 cells and KYSE-450 cells were divided into control group(DMSO)and experimental group(TOFA),respectively.The cells(4×10^(3) cells/100μl)were inoculated into 96-well plates with 5 multiple wells at each concentration.After 24 h culture,cells were treated with DMSO or different concentrations(1,3,5,10μg/ml)of TOFA for 24,48 and 72 h.Cell proliferation was detected by MTT,cell cycle and apoptosis were detected by flow cytometry,the expression levels of p21 and Cleaved caspase-3 and modification levels of p-Akt,p-mTOR and p-4EBP1 were detected by Western blot,and intracellular free fatty acids were detected by special kits.Results:MTT results showed that TOFA inhibited the proliferation of Eca109 and KYSE-450 cells in a concentration and time dependent manner(all P<0.05),with IC_(50) of 4.65μg/ml and 3.93μg/ml for 48 h,respectively.Flow cytometry results showed that compared with DMSO group,the percentage of cells in G2/M phase was increased and the apoptosis rate was increased in the experimental group.Western blotting results showed that compared with DMSO group,p21 and Cleaved caspase-3 protein expression levels were up-regulated,and p-AKT,p-mTOR and p-4EBP1 protein expression levels were down-regulated(all P<0.05).Conclusion:TOFA inhibits the proliferation,blocks the cycle progression and promotes apoptosis of ESCC,the mechanism may be related to the AKT/mTOR/4EBP1 signaling pathway.