磷脂酰肌醇3-激酶/蛋白激酶B信号通路对CD40介导胃癌细胞侵袭与转移的研究

Blocking PI3K/Akt signaling pathway strengthened the suppression of invasion and migration by CD40 in gastric cancer cells

目的 探讨可溶性CD40配体（sCD40L）联合磷脂酰肌醇3-激酶/蛋白激酶B（P13K/Akt）信号通路特异性阻断剂LY294002对胃癌AGS细胞体外增殖及侵袭、转移的影响;初步探讨其可能的作用机制.方法 将细胞分成对照组（仅加入含10%胎牛血清的RPMI 1640培养液）、LY294002组（20μmol/L）、sCD40L组（2μg/ml）和LY294002＋sCD40L组.四甲基偶氮唑盐法检测不同浓度sCD401,及联合LY294002对细胞增殖的影响;Transwell实验和划痕实验分析细胞侵袭、转移能力的改变;Western印迹检测细胞P13K、p-Akt、血管内皮生长因子（VEGF）蛋白表达的变化.结果 LY294002作用后细胞生存率为65.7%,sCD40L作用后为70.1%,LY294002＋sCD40L组生存率为41.3 0A,差异有统计学意义（P〈0.05）.LY294002＋sCD40L组细胞的划痕愈合速度较sCD40L和LY294002组明显减慢.LY294002＋sCD40L组平均每个视野的细胞数较sCD40L和LY294002组明显减少.与对照组相比,sCD40L使P13K、p-Akt、VEGF蛋白表达升高,与LY294002联合作用后,P13K,p-Akt、VEGF蛋白表达明显降低.结论 阻断P13K/Akt信号途径可显著增强sCD40L对人胃癌AGS细胞增殖及侵袭、转移的抑制作用,为胃癌的生物治疗提供一种新的方法.

Objective To investigate the effects of soluble CD40 ligand combined with LY294002,a specific inhibitor of phosphatidylinositol 3-kinase,on proliferation and invasion of gastric cancer cells （AGS ） and its potential mechanisms. Methods The effect of gradient concentration of sCD40L and its combination with LY294002 on cellular proliferation was determined using MTT assay. The ability of cellular invasion and migration was evaluated by wound healing and Transwell invasion assay. The protein expression of PI3K, p-Akt and vascular endothelia growth factor （VEGF） was tested by Western blotting. Results The inhibitory effect of cellular proliferation in sCD40L, LY294002 and sCD40L ＋ LY294002 groups was 70. 1%, 65. 7% and 41. 3%, respectively, with significant difference （P〈0. 05）. And wound healing was also delayed in cells treated with sCD40L plus LY294002 when compared with those treated with sCD40L or LY294002. These results indicated that the inhibitory effect of cellular proliferation, migration and invasion was enhanced when cells were treated with sCD40L plus LY294002. The protein expression of PI3K,p-Akt and VEGF increased in cells treated with sCD40L, but reduced drastically in cells treated with sCD40L plus LY294002. Conclusion Inhibiting PI3K/Akt signaling pathway may enhance the inhibitory effect of proliferation, migration and invasion by sCD40L on AGS cells, which provide a new way for biochemical therapy of gastric cancer.