【摘要】 目的探讨Fbxw8对人前列腺癌DU145细胞增殖及对增殖相关基因cyclinA、cyclinB、CDK1、p21和p27表达的影响。方法 RT-PCR和Western blot法验证Fbxw8 siRNAs的干扰效率;利用CCK-8检测法检测细胞的增殖能力;流式细胞仪检测细胞周期分布情况;Western blot法检测细胞增殖相关蛋白CDK1、CDK2、cyclinA、cyclinB、P21、P27的表达。结果 Fbxw8 siRNAs显著下调DU145细胞中Fbxw8mRNA及蛋白水平的表达(P<0.01);Fbxw8 siRNAs显著抑制DU145细胞增殖活力,其中72 h抑制最明显(P<0.05);流式细胞术分析显示,Fbxw8 siRNAs可阻滞细胞于G2/M期,G2/M期细胞百分数由15.32%(control siRNA)增加到29.03%(Fbxw8 siRNA 1)和28.39%(Fbxw8 siRNA 2);Western blot结果显示,Fbxw8 siRNAs可促进周期蛋白依赖性激酶抑制剂p21、p27的表达,同时下调G2/M期调控蛋白CDK1、CDK2、cyclin A及cyclin B1的表达。结论靶向抑制DU145细胞中Fbxw8的表达可显著抑制细胞的增殖,使阻滞细胞于G2/M期,并且可能是通过下调CDK1、CDK2、cyclin A及cyclin B1的表达,上调P21、P27的表达实现的。更多还原

【Abstract】 Objective To investigate the effect of F-box and WD repeat domain containing 8(Fbxw8) on cell proliferation and the expression levels of proliferation-related genes such as cyclinA,cyclinB,cdk1,cdk2,p21 and p27 in human prostate cancer DU145 cells.Methods RT-PCR and Western blot were applied to evaluate the interference efficiency of Fbxw8 siRNAs.The cell proliferation was assessed by CCK-8 assay.The cell cycle of DU-145 cells was determined by flow cytometry.The expression levels of CDK1,CDK2,cyclin A,cyclinB,P21 and P27 proteins were detected by Western blot.Results Fbxw8 siRNA significantly decreased the mRNA and protein of Fbxw8 in DU145 cells(P<0.01).The proliferation of DU-145 cells was obviously inhibited after Fbxw8 siRNA intervention,especially in 72 h(P<0.05).Flow cytometry analysis showed that Fbxw8 siRNA caused cell cycle arrest in G2/M phase and cell percent of G2/M phase increased from 15.32%(control siRNA) to 29.03%(Fbxw8 siRNA 1) and 28.39%(Fbxw8 siRNA 2).Westert blot showed that the levels of CDK1,CDK2,CyclinA and CyclinB expression were down-regulated whereas the levels of p21 and p27 were up-regulated.Conclusion Targeted inhibition of Fbxw8 in DU145 cells can significantly inhibit cell proliferation and block cells in G2 / M phase,which may be achieved by decreasing the expressions of CDK1,CDK2,cyclin A and cyclin B1 and increasing the expressions of P21 and P27.更多还原