RAD001对胃癌MKN-45细胞生长影响的实验研究

目的探讨RAD001对胃癌细胞株MKN-45细胞生长的影响及可能作用机制。方法体外培养MKN-45细胞,给予RAD001干预后通过细胞计数检测MKN-45细胞数目,流式细胞术检测细胞周期分布,Western-blot检测核糖体40S小亚基S6K蛋白激酶（p70S6K）、真核起始因子4E（eIF-4E）结合蛋白1（4E-BP1）、S6蛋白表达及磷酸化情况。结果RAD001作用于MKN-45细胞24 h：RAD001对MKN-45细胞的生长产生抑制作用,抑制作用具有时间依赖性;MKN-45细胞细胞周期发生改变,停滞在G0-G1期细胞比例增加;MKN-45细胞的4E-BP1、p70S6K、S6无明显变化,phosphate-p70S6K、phosphate-S6表达缺失,phosphate 4E-BP1表达下降。结论RAD001是通过抑制mTOR下游通路阻断4E-BP1及p70S6K的磷酸化和蛋白质翻译进而调节细胞周期以发挥抑制MKN-45细胞生长的作用。

Experimental study of the effect of RAD001on gastric carcinoma cell line MKN-45

Objective To investigate the effect of RAD001 on gastric carcinoma cell line MKN-45 and to explore the possible mechanism.Methods Gastric carcinoma MKN-45 cell line were cultured in RPMI1640 with 10% fetal calf serum and treated by RAD001（20 nmol/L）;afterwards,the cell coulter,flow cytometry,western-blot array were used to detect the growth,cell cycle and protein expression of the tumor cells.Results Following RAD001 treatment,the MKN-45 cells were inhibited in a time-dependence manner.There were changes in the cycle of the MKN-45 cells,with an increase in the cell percentage of G0-G1 phase stagnation.Meanwhile,there were no marked changes in the expressions of eukaryotic translation initiation factors eIF-4E（eIF-4E） and phosphate-4E-BP1,ribosomal protein 40S kinase,and there was loss in the expression of phosphate-p70S6K and phosphate-S6,and the expression of phosphate 4E-BP1 was down-regulated.Conclusion RAD001 can inhibit the growth of gastric carcinoma cell line MKN-45 by the adjustment of cell cycle via inhibition of the downstream pathway of mTOR and the consequent blockage of the phosphorylation and protein translation of p70S6K and 4E-BP1