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目的:研究己烯雌酚(DES)对卵巢上皮性癌细胞株OVCAR3中HOXA10基因表达及启动子区甲基化状态的影响。方法:体外培养卵巢上皮性癌细胞株OVCAR3,分别加入5×10-6、5×10-7和5×10-8 mol/L的DES培养5d(120h)后收集细胞,采用实时荧光定量聚合酶链反应和蛋白免疫印记技术检测HOXA10基因mRNA和蛋白的表达水平;运用甲基化特异性聚合酶链反应检测HOXA10基因启动子区的甲基化状态。结果:随着DES浓度的升高,OVCAR3细胞中HOXA10基因的表达存在升高的趋势,且各组间的表达差异有统计学意义,HOXA10基因mRNA表达(5×10-8 mol/L DES组与对照组比较,P=0.002;5×10-7 mol/L DES组与5×10-8 mol/L DES组比较,P=0.028;5×10-6 mol/L DES组与5×10-7 mol/L DES组比较,P=0.000 2)、HOXA10基因蛋白表达(5×10-8 mol/L DES组与对照组比较,P=0.032;5×10-7 mol/L DES组与5×10-8 mol/L DES组比较,P=0.049;5×10-6 mol/L DES组与5×10-7 mol/LDES组比较,P=0.009);且DES可诱导HOXA10基因启动子区的甲基化状态发生去甲基化改变。结论:DES可以上调卵巢上皮性癌细胞株OVCAR3细胞中HOXA10基因的表达,这与DES对HOXA10基因启动子区甲基化状态的影响有关。
Objective: To study the effect of diethylstilbestrol (DES) on HOXA10 gene expression and methylation status in ovarian epithelial carcinoma cell line OVCAR3. Methods: Ovarian epithelial carcinoma cell line OVCAR3 was cultured in vitro. Cells were harvested after 5 × 10-6, 5 × 10-7 and 5 × 10-8 mol / L DES for 5 days (120h), respectively. Real-time fluorescence quantitative polymerase chain reaction The mRNA and protein expression of HOXA10 were detected by enzyme-linked immunosorbent assay (ELISA) and Western blotting. The methylation status of HOXA10 promoter was detected by methylation-specific polymerase chain reaction (PCR). Results: With the increase of DES concentration, the expression of HOXA10 gene in OVCAR3 cells tended to increase, and the expression of HOXA10 gene was significantly different (5 × 10-8 mol / L DES group Compared with the control group, P = 0.002; P = 0.028 in 5 × 10-7 mol / L DES group and 5 × 10-8 mol / L DES group; -7 mol / L DES group, P = 0.000 2), HOXA10 gene protein expression (5 × 10-8 mol / L DES group compared with the control group, P = 0.032; 5 × 10-7 mol / L DES group and P = 0.049 in 5 × 10-8 mol / L DES group and P = 0.009 in 5 × 10-6 mol / L DES group compared with 5 × 10-7 mol / LDES group); and DES could induce the activation of HOXA10 gene The methylation status of the subregion is demethylated. Conclusions: DES can up-regulate HOXA10 gene expression in OVCAR3 cells, which is related to the effect of DES on the methylation status of HOXA10 promoter.