Department of Life Science and Institute for Natural Sciences, Hanyang University, Seoul 133-791, South Korea.
Department of Life Science and Institute for Natural Sciences, Hanyang University, Seoul 133-791, South Korea.
Aquat Toxicol. 2014 Nov;156:191-200. doi: 10.1016/j.aquatox.2014.08.013. Epub 2014 Sep 6.
This study aimed to elucidate the mechanisms by which alkylphenols disrupt endocrine function in wild amphibians in Korea. To this end, the effects of 4-tert-octylphenol (OP), 17β-estradiol (E2), and estrogen receptor (ER) agonists on the expression profiles of vitellogenin (VTG) and ERs were examined in livers obtained from male Bombina orientalis toads. A single injection of E2 (10μg/kg; 0.03μmol/kg) induced transcription of VTG mRNA at 2 days post injection; however, injection of either the ERα-selective agonist propyl-(1H)-pyrazole-1,3,5-triyl-trisphenol (PPT, 50μg/kg; 0.12μmol/kg) or the ERβ-selective agonist 2,3-bis-(4-hydroxyphenyl)-propionitrile (DPN, 50μg/kg; 0.20μmol/kg) did not affect the expression of VTG. This finding suggests that both ERα and ERβ are required to induce transcription of VTG in the male B. orientalis liver. Interestingly, E2, PPT, and DPN induced transcription of ERα, which was also reflected on the protein level; however, these alkylphenols did not affect ERβ transcription. Similarly, VTG transcription was induced by a single injection of 1-100mg/kg (0.04-484.66μmol/kg) OP, while 0.1mg/kg (0.48μmol/kg) OP had no effect on VTG transcription. This result suggests that the lowest observable effect concentration (LOEC) of OP for induction of VTG transcription in the male liver is 1mg/kg (4.84μmol/kg). Furthermore, treatment with E2 (10μg/kg; 0.03μmol/kg) or OP (1mg/kg; 4.84μmol/kg) significantly upregulated ERα transcription, and a 10mg/kg (48.46μmol/kg) dose of OP significantly upregulated ERβ transcription. The ER antagonist ICI 182,780 decreased the basal levels of ERα and ERβ mRNA, and also prevented E2-mediated and OP-mediated induction of VTG, ERα, and ERβ transcription. A second injection of 0.1mg/kg (0.48μmol/kg) OP after a two-day interval significantly upregulated the transcription of VTG and ERα, but not of ERβ. These results suggest that sensitization of VTG transcription by repeated exposure to OP is mediated by the induction of ERα. Different combinations of alkylphenols that are ubiquitous in the freshwater system in Korea could potentially exert a synergistic effect on endocrine disruption. Thus, chronic exposure to alkylphenols, even at their NOECs, could still disrupt endocrine function in B. orientalis.
本研究旨在阐明烷基酚类物质在韩国野生两栖动物中破坏内分泌功能的机制。为此，我们检测了 4-叔辛基苯酚（OP）、17β-雌二醇（E2）和雌激素受体（ER）激动剂对雄性东方铃蟾肝脏中卵黄蛋白原（VTG）和 ERs 表达谱的影响。单次注射 E2（10μg/kg；0.03μmol/kg）可在注射后 2 天诱导 VTG mRNA 的转录；然而，注射 ERα 选择性激动剂丙基（1H）-吡唑-1,3,5-三基三苯（PPT，50μg/kg；0.12μmol/kg）或 ERβ 选择性激动剂 2,3-双-（4-羟苯基）-丙腈（DPN，50μg/kg；0.20μmol/kg）均未影响 VTG 的表达。这一发现表明，ERα 和 ERβ 都需要诱导雄性 B. orientalis 肝脏中 VTG 的转录。有趣的是，E2、PPT 和 DPN 诱导了 ERα 的转录，这也反映在蛋白质水平上；然而，这些烷基酚类物质并没有影响 ERβ 的转录。同样，单次注射 1-100mg/kg（0.04-484.66μmol/kg）OP 可诱导 VTG 转录，而 0.1mg/kg（0.48μmol/kg）OP 对 VTG 转录没有影响。这表明 OP 诱导雄性肝脏中 VTG 转录的最低可观察效应浓度（LOEC）为 1mg/kg（4.84μmol/kg）。此外，用 E2（10μg/kg；0.03μmol/kg）或 OP（1mg/kg；4.84μmol/kg）处理可显著上调 ERα 的转录，10mg/kg（48.46μmol/kg）OP 可显著上调 ERβ 的转录。ER 拮抗剂 ICI 182,780 降低了 ERα 和 ERβ mRNA 的基础水平，并阻止了 E2 介导和 OP 介导的 VTG、ERα 和 ERβ 转录的诱导。间隔两天后再次注射 0.1mg/kg（0.48μmol/kg）OP 可显著上调 VTG 和 ERα 的转录，但不上调 ERβ 的转录。这些结果表明，OP 重复暴露对 VTG 转录的敏化作用是通过 ERα 的诱导介导的。韩国淡水系统中普遍存在的不同烷基酚类化合物组合可能对内分泌干扰产生协同作用。因此，即使在其无可见效应浓度（NOEC）下，慢性暴露于烷基酚类化合物仍可能破坏 B. orientalis 的内分泌功能。
