17α-Ethinylestradiol decreases expression of multiple hepatic nucleotide excision repair genes in zebrafish (Danio rerio)

Waterborne 17α-ethinylestradiol (EE₂) alters hormone-mediated biological indicators in fish. These alterations include increased plasma vitellogenin, increased intersex individuals, decreased egg and sperm production, reduced gamete quality, and complete feminization of male fish. Together, these observations implicate aquatic estrogens in a broad range of detrimental effects on fish reproduction and fitness. In addition to impairing reproductive processes, EE₂ is also a strong promoter of hepatic tumor formation. Since many ubiquitous, aquatic hepatocarcinogens form DNA adducts that are preferentially repaired by nucleotide excision repair (NER) processes, we hypothesized that EE₂ may exert co-carcinogenic effects by reducing an organisms ability to repair DNA adducts via this mechanism. The present study used fluorescence-based quantitative RT-PCR to examine effects of environmentally relevant concentrations of the semisynthetic estrogen, EE₂, on hepatic nucleotide excision repair (NER) gene expression. Adult male and female zebrafish (Danio rerio) were exposed to 1 ng/L, 10 ng/L or 100 ng/L concentrations of EE₂, or to a solvent control (0.05%, v/v ethanol), for 7 days with static water renewal every 24 h. Effectiveness of EE₂ exposure in the liver was confirmed by examining hepatic expression of two estrogen-responsive biomarkers, vitellogenin-1 and cytochrome P450-1A1 (CYP1A1). Quantitative analysis confirmed that exposure to 100 ng/L EE₂ caused significant decreases in transcript abundance of several hepatic NER genes in male zebrafish, including XPC (>17-fold), XPA (>7-fold), XPD (>8-fold), and XPF (>8-fold). Adult female zebrafish exhibited a four-fold decreased in XPC mRNA abundance at all exposure concentrations. Decreased mRNA abundance of NER genes was also seen to a lesser degree at lower concentrations of EE₂. Adult male zebrafish showed greater reduction of hepatic NER transcript levels than their female counterparts, which is consistent with the sexually dimorphic incidence of hepatocellular carcinoma in many species. Decreased transcript levels of NER genes have been shown to be an important epidemiological marker for increased cancer risk and decreased repair capacity in humans.