AR-V7对前列腺癌细胞增长及Akt/mTOR信号通路的影响

The effect of AR-V7 on the growth of prostate cancer cells and Akt/mTOR signaling pathway

目的探讨雄激素受体剪接变异体7(androgen receptor splice variant-7,AR-V7)对前列腺癌(PCA)细胞的生长作用及与PI3K/AKT信号通路的相关性。方法构建pIRES2-EGFP-AR-V7重组质粒并转染人PCA细胞(prostate cancer-3,PC-3)为AR-V7重组载体组,用pIRES2-EGFP空载质粒转染PC-3为空载对照组,正常培养的PC-3为空白对照组,应用实时荧光定量PCR(quantitative real-time PCR,qRT-PCR)及蛋白免疫印迹法(western blot,WB)验证AR-V7基因及蛋白表达水平。用CCK-8细胞计数法及流式细胞仪比较各组细胞的增殖及凋亡差异,用qRT-PCR比较组间Akt、mTOR基因表达水平,用WB比较组间Akt、p-Akt、mTOR及p-mTOR的蛋白表达水平。结果AR-V7重组载体组AR-V7基因及蛋白表达水平明显高于空载对照组和空白对照组(P<0.05)。AR-V7重组载体组细胞增殖活性明显高于空载对照组和空白对照组(P<0.01)。各组细胞凋亡率差异无统计学意义(P>0.05),各组Akt、mTOR基因及蛋白表达水平差异无统计学意义(P>0.05)。ARV7重组载体组p-Akt、p-mTOR蛋白表达水平明显低于空载对照组和空白对照组(P<0.05)。结论AR-V7对前列腺癌细胞增殖有促进作用,对Akt、mTOR基因及蛋白表达无影响,但可以抑制p-Akt及p-mTOR蛋白表达。

Objective To investigate the effect of androgen receptor splice variant-7(AR-V7)on the proliferation of prostate cancer cells and its correlation with PI3K/AKT signaling pathway.Methods The recombinant plasmid of pIRES2-EGFP-AR-V7 was constructed to transfect into human prostate cancer cell-3(PC-3)as the AR-V7 recombination carrier group;PC-3 transfected with pIRES2-EGFP empty plasmid as the empty load control group;normal cultured PC-3 is the blank control group.The expression levels of AR-V7 gene and protein were verified by quantitative real-time PCR(qRT-PCR)and western blot(WB).CCK-8 cell counting method and flow cytometry were used to compare the differences of cell proliferation and apoptosis in each group.The gene expression levels of Akt and mTOR were compared by qRT-PCR among the groups.The protein expression levels of Akt,p-Akt,mTOR and p-mTOR were compared by WBamong the groups.Results The Levels of AR-V7 gene and protein expression in AR-V7 recombinant carrier group were significantly higher than those of the two control groups(P<0.05).The cell proliferation activity of AR-V7 recombinant carrier group was significantly higher than those of the two control groups(P<0.01).There was no significant difference in apoptosis rate among the groups(P>0.05),and there was no significant difference in Akt、mTOR genes and proteins expression level among the groups(P>0.05).The expression level of p-Akt,p-mTOR protein in AR-V7 recombinant carrier group was significantly lower than those of the two control groups(P<0.05).Conclusion AR-V7 can promote the proliferation of prostate cancer cells,while AR-V7 has no effect on the expression of Akt,mTOR genes and proteins,but it can inhibit the expression of p-Akt and p-mTOR proteins.