University of Gothenburg, Department of Zoology, SE-405 30 Göteborg, Sweden.
Aquat Toxicol. 2010 Nov 1;100(3):263-70. doi: 10.1016/j.aquatox.2010.07.024. Epub 2010 Jul 27.
The aryl hydrocarbon receptor (AhR) and the estrogen receptor (ER) are ligand-activated transcription factors, both of which can be activated by environmental pollutants. The AhR regulates cytochrome P450 1A (CYP1A) expression and can be induced by aromatic hydrocarbons. The ER regulates vitellogenin (VTG) expression and can be induced by estrogenic substances. Both receptor responses are established biomarkers used to assess the effects of pollutants in the aquatic environment. The receptors can also be affected in situations of mixed exposure. Cross-talk between these receptor pathways has been suggested, although there are conflicting data in the literature. We investigated cross-talk between ER-VTG and AhR-CYP1A signaling pathways in primary cultures of rainbow trout hepatocytes, using quantitative PCR (qPCR) for mRNA analyses and studies of CYP1A catalytic function and protein expression. The model agonists β-naphthoflavone (BNF) and 17α-ethinylestradiol (EE(2)) were used for AhR and ER activation, respectively. Combined exposure to BNF and EE(2) reduced the EE(2)-mediated induction of VTG mRNA levels by about 40%, but had no effect on the BNF-mediated CYP1A mRNA levels, indicative of a one-way inhibiting AhR-ER cross-talk. However, basal levels of CYP1A mRNA were reduced 40% upon exposure to EE(2) alone, implying different cross-talk mechanism between basal and induced CYP1A mRNA levels. The mammalian ER antagonist fulvestrant (ICI) is commonly described as an absolute ER antagonist. However, ICI failed to reverse the ER activation caused by EE(2) in the present study. The CYP1A-mediated ethoxyresorufin-O-deethylase (EROD) activity was reduced by 80% in cells co-treated with BNF and EE(2), compared to cells exposed to BNF alone. In vitro inhibiting studies suggests that this reduction was a result of inhibition of the CYP1A catalyst by EE(2) since EE(2) acted as a potent inhibitor (IC(50): 4.6μM) of the EROD activity. In addition, ICI also acted as a potent inhibitor of the EROD enzyme (IC(50): 0.6μM). Taken together, our data supports a one-way inhibiting AhR-ER cross-talk in rainbow trout hepatocytes exposed to a mixture of BNF and EE(2).
芳香烃受体 (AhR) 和雌激素受体 (ER) 都是配体激活的转录因子，两者都可以被环境污染物激活。AhR 调节细胞色素 P450 1A (CYP1A) 的表达，并可被芳香烃诱导。ER 调节卵黄蛋白原 (VTG) 的表达，并可被雌激素物质诱导。这两种受体反应都是评估水生环境中污染物影响的既定生物标志物。在混合暴露的情况下，这些受体也可能受到影响。虽然文献中存在相互矛盾的数据，但已经有人提出这些受体途径之间存在交叉对话。我们使用定量 PCR (qPCR) 进行 mRNA 分析以及 CYP1A 催化功能和蛋白表达研究，在虹鳟鱼原代肝细胞中研究了 ER-VTG 和 AhR-CYP1A 信号通路之间的交叉对话。模型激动剂 β-萘黄酮 (BNF) 和 17α-乙炔雌二醇 (EE(2)) 分别用于 AhR 和 ER 的激活。联合暴露于 BNF 和 EE(2) 使 EE(2)介导的 VTG mRNA 水平的诱导降低了约 40%，但对 BNF 介导的 CYP1A mRNA 水平没有影响，表明存在单向抑制 AhR-ER 交叉对话。然而，单独暴露于 EE(2)会使基础 CYP1A mRNA 水平降低 40%，这意味着基础和诱导的 CYP1A mRNA 水平之间存在不同的交叉对话机制。哺乳动物 ER 拮抗剂氟维司群 (ICI) 通常被描述为绝对的 ER 拮抗剂。然而，在本研究中，ICI 未能逆转 EE(2)引起的 ER 激活。与单独暴露于 BNF 的细胞相比，联合处理 BNF 和 EE(2)的细胞中 CYP1A 介导的乙氧基 RESORUFIN-O-去乙基酶 (EROD) 活性降低了 80%。体外抑制研究表明，这种降低是由于 EE(2)抑制 CYP1A 催化剂引起的，因为 EE(2)是 EROD 活性的有效抑制剂 (IC(50)：4.6μM)。此外，ICI 本身也是 EROD 酶的有效抑制剂 (IC(50)：0.6μM)。综上所述，我们的数据支持在暴露于 BNF 和 EE(2)混合物的虹鳟鱼肝细胞中存在单向抑制 AhR-ER 交叉对话。
