Transcriptional modulation of brain and hepatic estrogen receptor and P450arom isotypes in juvenile Atlantic salmon (Salmo salar) after waterborne exposure to the xenoestrogen, 4-nonylphenol.

The molecular basis for vitellogenin (Vtg) and Zr-protein gene (and protein) expression shows that the Vtg and Zr-protein gene activations are receptor-mediated responses that are ligand structure-dependent interactions with ER, probably involving all isoforms, in addition to other co-activators. In addition to their effect as direct agonist to the ERs, the endocrine-disrupting effects of environmental chemicals are sometimes interpreted as interference with steroidogenesis and with the steroidal regulation of the normal development and function of the male and female reproductive tracts. In vertebrates, the cytochrome P450 aromatase (P450arom) is a crucial steroidogenic enzyme catalyzing the final step in the conversion of androgens to estrogens. The present study was undertaken to investigate the effects of nonylphenol on brain and liver ER and P450arom isotypes gene expressions as sensitive biomarker of effect. Fish were exposed to static and continuous waterborne nonylphenol at 5, 15 and 50 microg/L. Blood, brain and liver samples were collected after 0 (control), 3 and 7 days post-exposure. Plasma levels of Vtg and Zr-proteins were analyzed using immunoblotting and enzyme linked immunosorbent assay (ELISA) methods. A quantitative RT-PCR method was used to analyse gene expression patterns using cloned plasmid containing amplicons of interest as standards. Our data demonstrate that both ER and P450arom gene isotypes showed differential organ-specific, nonylphenol concentration- and time-dependent expression patterns after exposure to environmental relevant concentrations of nonylphenol. Liver and plasma Vtg and Zr-protein gene and protein expression showed a concentration- and time-dependent induction at day 3 and 7 post-exposure. This is the first study evaluating, in parallel, the ERs, P450aroms and protein xenoestrogen biomarkers of effect in any fish species or lower vertebrate. Therefore, the present study shows that the brain and hepatic ER and P450arom isotypes evaluated using state-of-the-art molecular approaches are sensitive xenoestrogen biomarkers of effect. These responses should be evaluated in parallel with plasma protein biomarkers such as Vtg and Zr-proteins.