Estrogenic and anti-estrogenic influences in cultured brown trout hepatocytes: Focus on the expression of some estrogen and peroxisomal related genes and linked phenotypic anchors

•Evidence of crosstalk between estrogens and peroxisomal pathways in brown trout.•VtgA and ERα mRNA levels increased after 1, 10 and 50 μM of ethinylestradiol (EE2).•ERβ-1, catalase and urate oxidase mRNA levels decreased after estrogenic stimuli.•Estrogenic effects in VtgA, ERα and Uox mRNA levels were reverted by ICI 182,780.•Immunofluorescence/electron microscopy shows smaller peroxisomes after 50 μM of EE2.

Estrogens, estrogenic mimics and anti-estrogenic compounds are known to target estrogen receptors (ER) that can modulate other nuclear receptor signaling pathways, such as those controlled by the peroxisome proliferator-activated receptor (PPAR), and alter organelle (inc. peroxisome) morphodynamics. By using primary isolated brown trout (Salmo trutta f. fario) hepatocytes after 72 and 96 h of exposure we evaluated some effects in selected molecular targets and in peroxisomal morphological features caused by: (1) an ER agonist (ethinylestradiol-EE2) at 1, 10 and 50 μM; (2) an ER antagonist (ICI 182,780) at 10 and 50 μM; and (3) mixtures of both (Mix I-10 μM EE2 and 50 μM ICI; Mix II-1 μM EE2 and 10 μM ICI and Mix III-1 μM EE2 and 50 μM ICI). The mRNA levels of the estrogenic targets (ERα, ERβ-1 and vitellogenin A-VtgA) and the peroxisome structure/function related genes (catalase, urate oxidase-Uox, 17β-hydroxysteroid dehydrogenase 4-17β-HSD4, peroxin 11α-Pex11α and PPARα) were analyzed by real-time polymerase chain reaction (RT-PCR).Stereology combined with catalase immunofluorescence revealed a significant reduction in peroxisome volume densities at 50 μM of EE2 exposure. Concomitantly, at the same concentration, electron microscopy showed smaller peroxisome profiles, exacerbated proliferation of rough endoplasmic reticulum, and a generalized cytoplasmic vacuolization of hepatocytes. Catalase and Uox mRNA levels decreased in all estrogenic stimuli conditions. VtgA and ERα mRNA increased after all EE2 treatments, while ERβ-1 had an inverse pattern. The EE2 action was reversed by ICI 182,780 in a concentration-dependent manner, for VtgA, ERα and Uox. Overall, our data show the great value of primary brown trout hepatocytes to study the effects of estrogenic/anti-estrogenic inputs in peroxisome kinetics and in ER and PPARα signaling, backing the still open hypothesis of crosstalk interactions between these pathways and calling for more mechanistic experiments.