纤维连接蛋白—paxillin通路对人胃癌细胞系AGS体外侵袭力的影响

Effect of Fibronectin-paxillin Pathway on Invasiveness of Human Gastric Cancer Cellline AGS

目的研究纤维连接蛋白激活paxillin后对人胃癌细胞侵袭力的影响,探讨抑制paxillin影响胃癌细胞侵袭力的机制。方法以递增浓度的纤维连接蛋白刺激人胃癌细胞株AGS,以免疫沉淀和蛋白质印迹法检测胃癌细胞内paxillin第118位酪氨酸(tyr-118)磷酸化的变化,同时以改良Boy-den小室法检测细胞侵袭力变化。设计合成paxillin siRNA并进行效果比较,观察siRNA抑制纤维连接蛋白促胃癌细胞内paxillin tyr-118磷酸化及细胞侵袭力的改变。结果纤维连接蛋白能促进AGS细胞paxillin tyr-118磷酸化的增强和胃癌细胞侵袭力(P<0.05),并在一定范围内具有剂量依赖性。siRNA干预后,胃癌细胞内paxillin tyr-118磷酸化及细胞侵袭力均有显著降低(P<0.05)。结论纤维连接蛋白可有效增强胃癌细胞的侵袭力,paxillin tyr-118磷酸化在此过程中起关键作用,使用paxillin siRNA可以抑制纤维连接蛋白促人胃癌细胞侵袭的作用。

Objective To investigate the interaction of fibronectin and paxillin in human gastric cancer cell,AGS,and reversal effects of paxillin siRNA on invasiveness of AGS cell induced by fibronectin.Methods A gastric cancer cell line,AGS,was stimulated by fibronectin with gradient concentrations （0,10,100,1 000nmol/L）.The phosphorylation expression of paxillin tyrosine 118（tyr-118）,was detected by immunoprecipitation and Western blot.The invasiveness of AGS cells was measured by the modified Boyden chamber assay.siRNA targeting paxillin was transfected into AGS cells,effect of paxillin silencing on phosphorylation of paxillin（tyr-118）and invasiveness of AGS cells stimulated by fibronectin were detected respectively.Results The AGS cell showed a dose-dependence on fibronectin in phosphorylation of paxillin tyr-118 and its invasiveness.Invasiveness and phosphorylation of paxillin tyr-118 in AGS reached their climax when the concentration of fibronectin reached 100nmol/L,whereas the expression of paxillin remained unchanged after stimulated by fibronectin（P 〈0.05）.siRNA targeting paxillin suppressed phosphorylation of paxillin tyr-118 and the invasiveness of AGS cells significantly,decreased compared with the controls（P〈0.05）.Conclusion Fibronectin promotes paxillin tyr-118 phosphorylation and invasiveness of AGS cells.Paxillin silenced by RNA interference inhibits the cell invasiveness stimulated by fibronectin.Paxillin is a key factor in the fibronectin-stimulated cell invasiveness of AGS cells.