雷帕霉素对食管癌EC1细胞mTOR表达及细胞生长与凋亡的影响

Effects of rapamycin on expression of mTOR gene,growth and apoptosis of human esophageal EC1 cells

目的:探讨雷帕霉素对人食管癌EC1细胞哺乳动物雷帕霉素靶蛋白(mTOR)的表达及细胞生长、凋亡的影响。方法:采用低、中、高浓度的雷帕霉素(100、150、200nmol/L)分别作用于EC1细胞24、48、72h,同时设溶剂对照组和空白对照组,用免疫细胞化学及原位杂交法检测EC1细胞中mTOR蛋白与mRNA的表达,MTT法检测EC1细胞的增殖情况,TUNEL法检测各组作用24h后细胞凋亡情况。结果:雷帕霉素作用24、48与72h,5组细胞mTOR蛋白与mRNA的表达差异均有统计学意义(mTOR蛋白:F=29.273、34.328、41.571,P均<0.001;mTORmRNA:F=34.969、53.614、36.943,P均<0.001),细胞生长抑制率差异亦有统计学意义(F=566.732、51.768和186.022,P均<0.001);雷帕霉素低、中、高浓度组EC1细胞mTOR蛋白与mRNA的表达均低于各对照组,且其表达随雷帕霉素浓度的增加及作用时间的延长而减弱(P<0.05)。雷帕霉素低、中、高浓度组细胞生长抑制率均高于各对照组,且随雷帕霉素浓度的增加和作用时间的延长而升高(P<0.05)。作用24h,5组细胞凋亡指数(AI)差异有统计学意义(F=524.563,P<0.001),雷帕霉素低、中、高浓度组AI均高于各对照组,且随雷帕霉素浓度的增加而增高(P<0.05)。结论:雷帕霉素可能通过抑制EC1细胞mTOR蛋白与mRNA的表达从而抑制EC1细胞的增殖,促进细胞凋亡。

Aim：To explore the effect of rapamycin on the expression of mTOR gene and growth and apoptosis of human esophageal EC1 cells.Methods：EC1 cells were treated with different concentration of rapamycin （100,150 and 200 nmol/L）for different time（24,48 and 72 h）,and the control group was established accordingly by using solutes（DMSO） only.The level of mRNA and protein of mTOR in cells was measured by in situ hybridization and immunocytochemistry.MTT was used to detect proliferation of cells,and apoptosis rate was detected using TUNEL.Results：There were significant differences in the expressions of mTOR mRNA,protein and growth inhibition rate of EC1 cells among the 5 groups at 24,48,and 72 h （protein：F=29.273,34.328,41.571,P0.001;mRNA：F=34.969,53.614,36.943,P0.001;growth inhibition rate：F=566.732,51.768 and 186.022,P0.001）.The expressions of mTOR mRNA and protein in the rapamycin treatment groups were lower than those of the control groups,which weakened with the increase of rapamycin concentration and time exposure（P0.05）.Compared with the control groups,growth inhibition rate of EC1 cells in rapamycin treatment groups was higher（P0.05）,and increased with the increase of rapamycin concentration and time exposure（P0.05）.There were significant differences in the apoptosis index among the 5 groups（F=524.563,P0.001）.The apoptosis index of EC1 cells treated with different concentrations of rapamycin for 24 h exposure was higher than those of the control groups（P0.05）.Conclusion：Rapamycin could effectively downregulate the expression of mTOR,which may inhibit the cell proliferation,and promote apoptosis of EC1.