Department of Pharmacochemistry, Section of Molecular and Computational Toxicology, Leiden/Amsterdam Center for Drug Research (LACDR), Vrije Universiteit Amsterdam, De Boelelaan 1083, 1081 HV Amsterdam, The Netherlands.
Environ Toxicol Pharmacol. 2005 Jan;19(1):41-55. doi: 10.1016/j.etap.2004.03.010.
Metabolism of polycyclic aromatic hydrocarbons (PAHs) has been studied intensively, and potential metabolites with estrogenic activity have been identified previously. However, little attention has been paid to the metabolic pathways in mammalians and to the combined effect of individual metabolites. Several hydroxylated metabolites of benzo[a]pyrene (BaP) and chrysene (CHN) were formed by rat liver microsomal cytochrome P450 (CYP) activity, some of which possess estrogenic activity. All mono- and several dihydroxylated metabolites of BaP and CHN were tested for ER affinity and estrogenic activity in a proliferation assay (E-screen) and in a reporter-gene assay (ER-CALUX). Twelve estrogenic metabolites were identified with EC50 values ranging from 40nM to 0.15mM. The combined effect of a mixture of seven PAH-metabolites was also studied in the ER binding assay. At concentrations that show little activity themselves, their joint action clearly exhibited significant estrogenic activity. BaP itself exhibited estrogenicity in the ER-CALUX assay due to bio-activation into estrogenic metabolites, probably via aryl hydrocarbon receptor (AhR) induced CYP activity. Furthermore, 2-hydroxy-CHN (2-OHCHN) induced supra-maximal (400%) estrogenic effects in the ER-CALUX assay. This effect was entirely ER-mediated, since the response was completely blocked with the ER-antagonist ICI182,780. We showed that 2-OHCHN increased ER-concentration, using ELISA techniques, which may explain the observed supra-maximal effects. Co-treatment with the AhR-antagonist 3',4'-dimethoxyflavone (DMF) enhanced ER-signaling, possibly via blockage of AhR-ER inhibitory cross-talk.
多环芳烃（PAHs）的代谢已得到深入研究，此前已鉴定出具有雌激素活性的潜在代谢产物。然而，人们对哺乳动物中的代谢途径以及单个代谢产物的组合效应关注甚少。几种苯并[a]芘（BaP）和屈（CHN）的羟化代谢物由大鼠肝微粒体细胞色素 P450（CYP）活性形成，其中一些具有雌激素活性。BaP 和 CHN 的所有单羟化和几种二羟化代谢物均在增殖测定（E-screen）和报告基因测定（ER-CALUX）中测试 ER 亲和力和雌激素活性。鉴定出 12 种具有 EC50 值为 40nM 至 0.15mM 的雌激素代谢物。还在 ER 结合测定中研究了七种 PAH 代谢物混合物的组合效应。在自身显示出很少活性的浓度下，它们的共同作用明显表现出明显的雌激素活性。BaP 本身在 ER-CALUX 测定中表现出雌激素活性，因为它被生物转化为雌激素代谢物，可能是通过芳烃受体（AhR）诱导的 CYP 活性。此外，2-羟基-CHN（2-OHCHN）在 ER-CALUX 测定中引起了超最大（400%）的雌激素效应。这种作用完全是 ER 介导的，因为用 ER 拮抗剂 ICI182,780 完全阻断了反应。我们使用 ELISA 技术表明，2-OHCHN 增加了 ER 浓度，这可能解释了观察到的超最大效应。用 AhR 拮抗剂 3'，4'-二甲氧基黄酮（DMF）共同处理增强了 ER 信号转导，可能是通过阻断 AhR-ER 抑制性交叉对话。
