低剂量双酚A及其与苯并芘联合染毒对人乳腺上皮细胞MDM2基因的影响

The combined exposure of low dose bisphenol A and benzo[a]pyrene affect the expression of MDM2 gene in human mammary epithelial cells

目的观察BPA与B[a]P联合作用对3种乳腺上皮细胞MDM2及TP53基因表达的影响。方法采用细胞增殖荧光法检测环境相关剂量BPA对3种乳腺上皮细胞S期细胞比例的影响,采用实时荧光定量PCR和蛋白免疫印迹的方法分别检测环境相关剂量水平(10-9、10-7mol/L)BPA及其与10-6mol/L B[a]P联合作用对3种乳腺上皮细胞中MDM2和TP53基因及其编码的蛋白表达水平的影响。结果环境相关剂量的BPA可以引起MCF-7细胞S期细胞比例增高;环境相关剂量的BPA单独作用对3种类型乳腺上皮细胞MDM2和TP53基因表达水平无显著影响;B[a]P单独作用可以引起MCF-10A细胞中MDM2基因表达水平的轻微增加及MCF-7细胞中MDM2基因表达的显著增加;在MCF-7细胞中BPA与B[a]P联合作用可明显增加B[a]P所引起的MDM2基因表达上调,而在HMEC和MCF-10A细胞未发现该现象,蛋白水平与mRNA水平的改变基本一致,同时p53/mdm2比值降低。结论环境相关剂量的BPA与B[a]P联合作用可以在雌激素受体表达阳性的MCF-7细胞中引起MDM2基因及其蛋白产物表达水平的显著上调,提示环境暴露剂量的BPA可能通过雌激素受体依赖的作用途径增加化学致癌物致乳腺癌发生风险。

Objective To observe the effect of combined exposure of BPA and B[a]P on MDM2 and TP53 gene expression in three different kinds of human epithelial cell lines.Methods Cell proliferation flourescence method was used to observe the S-phase cells ratio in three different kinds of human epithelial cells under the effect of low dose BPA.Real-time PCR and western-blotting were applied to detect the relative expression level of MDM2 and TP53 gene in three cell lines under the combined exposure to low dose BPA（ 10-9,10-7mol /L）and B[a]P（ 10-6mol /L）.Results Environment related doses BPA could increase the S-phase cell ratio in MCF-7 cells.There’s no notable effect of low dose BPA on the expression level of both MDM2 and TP53 genes in all of the three kinds of human mammary epithelial cells.The expression level of MDM2 gene was marginally increased in MCF-10A and dramatically increased in MCF-7 cells when exposed to B[a]P.Moreover,compare with the B[a]P only group combination of 10-9mol /L,10-7mol /L BPA and 17-β-estrodiol with B[a]P could increase the expression of MDM2 gene in both mRNA and protein level in MCF-7 cells and decrease the p53 /mdm2 ratio,but not in HMEC and MCF-10A cells.Conclusion Compare with the B[a]P only group the combination of low dose BPA and B[a]P could induce the expression of MDM2 oncogene in estrogen receptor positive MCF-7 cells.The result suggests that environment related doses BPA could increase the risk of carcinogen induced breast cancer through the estrogen receptor dependent pathways.