LY294002对人纤维肉瘤HT1080细胞生长的影响及机制研究

目的 研究PI3K特异性抑制剂LY294002对人纤维肉瘤HT1080细胞生长的影响及机制.方法 将对数生长期的HT1080细胞分组培养,对照组加入不含LY294002的培养液,实验组加入LY294002,浓度分别为5、10、25、50、100 μmol/L,培养12 h和24 h后,用SunBio~(TM) Am-Blue法测定细胞增殖抑制率.100 μmol/L LY294002作用HT1080细胞24 h后,观察细胞形态的改变,用流式细胞仪检测细胞凋亡情况,并收集蛋白标本,Western Blotting检测p-Akt及p-mTOR蛋白的表达.结果 与对照组比较,随着LY294002剂量、作用时间的增加,对HT1080细胞增殖抑制率增大(P＜0.05).LY294002(100 μmol/L)作用24 h后HT1080细胞胞体皱缩,细胞数较对照组减少,细胞早期凋亡率较对照组增加(P＜0.05),p-Akt(0.23±0.01)及p-mTOR(0.32±0.06)蛋白的表达低于对照组p-Akt(0.63±0.02)及p-mTOR(0.71±0.02)蛋白的表达(P＜0.01).结论 LY294002通过抑制PI3K-mTOR信号通路来抑制HT1080细胞的生长,PI3K-mTOR信号通路参与纤维肉瘤细胞的生长调节,该通路可能作为治疗纤维肉瘤的新靶点.

Effect and Mechanism of LY294002 on Growth of Fibrosarcoma Cell Line HT1080

Objective To investigate the effect and mechanism of LY294002 on growth of fibrosarcoma cell line HT1080.Methods The proliferation inhibitory rate of HT1080 cells treated by LY294002 at doses of 5,10,25,50,100 μmol/L for 12 and 24 h,respectively,was evaluated using SunBio~(TM) Am-Blue method.HT1080 cells were divided into two groups,that is,A and B.The A group was control group without treatment.The B group received LY294002(100 μmol/L)for 24 h.The changes of cell morphology and quantity were observed by phase contrast microscope.The apoptosis rate of HT1080 cells was detected by flow cytometry.And the protein expression of p-Akt and p-mTOR in HT1080 cells were detected by Western Blotting.Results The proliferation of HT1080 cells was inhibited in time-and dose-dependent manner by LY294002.After the treatment of 100 μmol/L LY294002 for 24 h,the growth of HT1080 cell line was remarkably inhibited by LY294002.The rate of apoptosis increased.And the protein expression of p-Akt(0.23±0.01)and p-mTOR(0.32±0.06)in LY294002 group was lower than p-Akt(0.63±0.02)and p-mTOR(0.71±0.02)in control group(P＜0.01).Conclusion LY294002 can inhibit the growth of HT1080 cells through PI3K-mTOR pathway.PI3K-mTOR pathway presents an appealing therapeutic target on fibrosarcoma.