雌二醇对子宫内膜癌细胞AKT通路中细胞因子的影响

Effect of Estrogen on the Cytokines of AKT Signaling Pathway in Endometrial Cancer Cells

目的:通过17β雌二醇(E2)诱导人子宫内膜癌HFC-1A细胞产生不同细胞因子,探讨雌激素在子宫内膜癌发展中的作用。方法:以浓度为1×10^(-6)mol/L雌二醇(E2组)作用于HEC-1A细胞8h、12h后,或1×10^(-6)mol/L雌激素受体抑制剂(ER组)、25×10^(-6)mol/L AKT抑制剂(AKT组)分别预处理HEC-1A细胞60min,各加入1×10^(-6)mol/L雌二醇作用8h和12h后;采用荧光定量PCR及ELISA技术,分别检测细胞内VEGF、bFGF、IL-8基因mRNA及细胞培养上清液中蛋白表达情况。Western Blot检测以1×10^(-6)mol/L雌二醇作用HEC-1A细胞15min后,细胞内AKT蛋白表达情况。结果:E_2组的VEGF、bFGF、IL-8 mRNA及蛋白的表达均明显高于对照组(P<0.05);ER组VEGF、bFGF、IL-8 mRNA和蛋白的表达,与E_2组相比较,除了8h的VEGF蛋白和12h的IL-8 mRNA表达无显著性差异及12h的VEGF mRNA表达稍增加外,均明显低于E_2组(P<0.05);AKT组VEGF、bFGF、IL-8 mRNA和蛋白的表达,与E_2组相比较,除了12h的IL-8 mRNA表达无显著性差异外,均明显低于E2组(P<0.05)。雌二醇作用HEC-1A细胞15 min后,与对照组比较,细胞内p-AKT蛋白表达明显增强(P<0.05)。结论:雌二醇诱导子宫内膜癌产生细胞因子VEGF、bFGF、IL-8可能是通过激活AKT通路实现的。

Objective： To investigate the relationship between estrogen and the development of endometrial cancer. Methods： Real-time polymerase chain reaction （ PCR ） was used to detect VEGF, bFGF, and IL-8 mRNA expression in HEC-1A cells and an enzyme-linked immunosorbent assay （ ELISA ） was used to detect the VEGF, bFGF, and IL-8 proteins in the cell culture fluid after stimulation with 1×10-6 mol/L estradiol （ E2 group ） for 8 h or 12 h, and with 60 min pretreatment with 1 ×10-6 mol/L ER inhibitor （ ER group ） or 25 x 10-6 mol/L AKT inhibitor （AKT group ） following stimulation with 1 × 10-6 mol/L estradiol for 8 h or 12 h. Western blot was used to detect AKT protein expression in the HEC-1A cells after stimulation with 1 × 10-6 mol/L estradiol for 15 min. ResultsReal-time PCR and ELISA showed the mRNA and protein expression of VEGF, bFGF, and IL-8 in the E2 group was significantly higher than that in the control group （ P〈 0.05 ）. The mRNA and protein expression ofVEGF, bFGF, and IL-8 in the ER group was significantly lower than that in the E2 group （ P 〈 0.05 ） except for the VEGF protein expression at 8 h and the IL-8 mRNA expression at 12 h. VEGF mRNA expression was slightly high in the E2 group at 12 h. The mRNA and protein expression ofVEGF, bFGF, and IL-8 in the AKT group was significantly lower than that in the E2 group （ P 〈 0.05 ）, except for IL-8 mRNA expression at 12 h. Western blot analysis showed that p-AKT protein expression in the HEC-1A cells after 15 min stimulation with 1 ×l0-6 mol/L estradiol was markedly higher than that in the control group （ P 〈 0.05 ）. Conclusion： Estrogen induced the production of VEGF, bFGF, and IL-8 through activating the AKT signal transmission pathway.