Sensitization of vitellogenin gene expression by low doses of octylphenol is mediated by estrogen receptor autoregulation in the Bombina orientalis (Boulenger) male liver

• OP induced VTG, ERα and ERβ mRNA in the B. orientalis male liver.
• Endogenous estrogen modulates the hepatic ERs gene expression in male liver.
• VTG mRNA cannot be induced by activated ERα or ERβ alone.
• E2- or OP-induced transcription of the ERα gene was more sensitive than that of ERβ.
• Exposure to low concentration OP sensitized VTG expression via autoregulation of ERα.

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 (10 μg/kg; 0.03 μmol/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, 50 μg/kg; 0.12 μmol/kg) or the ERβ-selective agonist 2,3-bis-(4-hydroxyphenyl)-propionitrile (DPN, 50 μg/kg; 0.20 μmol/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 1–100 mg/kg (0.04–484.66 μmol/kg) OP, while 0.1 mg/kg (0.48 μmol/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 1 mg/kg (4.84 μmol/kg). Furthermore, treatment with E2 (10 μg/kg; 0.03 μmol/kg) or OP (1 mg/kg; 4.84 μmol/kg) significantly upregulated ERα transcription, and a 10 mg/kg (48.46 μmol/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 0.1 mg/kg (0.48 μmol/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.