PI3K/Akt信号通路抑制剂对胃癌细胞SGC7901中Skp2和NAG-1表达的影响及其效应

Influence of Inhibitor of PI3K/Akt Pathway on Expression of Skp2 and NAG-1 in Gastric Carcinoma Cell Line SGC7901

目的研究PI3K/Akt信号转导通路靶向抑制剂LY294002对胃癌细胞SGC7901中S期激酶相关蛋白2(Skp2)和非甾体抗炎药诱导基因-1(NAG-1)表达的影响。方法将LY294002作用于胃癌细胞株SGC7901,四甲基偶氮唑蓝(MTT)法测定细胞增殖抑制率,流式细胞术AnnexinV2FITC/PI染色法检测细胞凋亡率,RT-PCR检测NAG-1、Skp2 mRNA的表达情况,Western blot检测p-Akt(Ser473)和总Akt(t-Akt)的表达情况。结果LY294002可明显抑制SGC7901的生长,呈时间-剂量依赖性,并出现典型的细胞形态学改变;10、30、50μmol/L的LY294002作用12h后细胞凋亡率分别为(12.7±0.42)%、(36.5±0.45)%和(64.1±0.79)%,显著高于空白对照组的(2.3±0.36)%,差异均有统计学意义(P<0.01);LY294002可下调p-Akt的蛋白表达,而t-Akt的表达未发生改变;LY294002可上调NAG-1 mRNA的表达,并下调Skp2 mRNA的表达。结论阻断PI3K/Akt信号转导通路可改变胃癌细胞株SGC7901中Skp2和NAG-1的表达,并能明显抑制细胞增殖,诱导细胞凋亡,其机制可能与其抑制Akt磷酸化及调控NAG-1、Skp2 mRNA表达水平有关。

Objective To explore the influence of PI3K/Akt pathway inhibited by LY294002 on the expression of Skp2 and NAG-1 in gastric carcinoma cell line SGC7901.Methods The effect of LY294002 on the proliferation of gastric cancer cells was observed by MTT assays.The apoptosis rate was determined by flow cytometry analysis,expression of NAG-1（non-steroidal anti-inflammatory drug activated gene-1） and Skp2 （S-phase kinase associated protein2）mRNA by RT-PCR.p-Akt（Ser473） and t-Akt by Western blot.Results LY294002 inhibited the proliferation of SGC7901,showing time-dose dependent,presenting with typical cell morphological changes.The cell apoptosis rate by LY294002 effects of 10,30,50 μmol/L after 12 h were （12.7±0.42）%,（36.5±0.45）%,（64.1±0.79）%,respectively,significantly higher than blank control group （2.3±0.36 %）,the difference was significant （P〈0.05）.LY294002 down-regulated expression of p-Akt protein,but t-Akt expression did not changed.LY294002 up-regulated NAG-1 mRNA expression and down-regulated Skp2 mRNA expression.Conclusion PI3K/Akt pathway inhibited by LY294002 could inhibit proliferation and induce apoptosis of SGC7901 cells,possiblely through regulating the phosphorylation of Akt and the expression of Skp2 and NAG-1 genes.