Éditeur(s) : HAL CCSD
Résumé : Benzophenone derivatives (BPs) are used as UV filters in many different products. These lipophilic compounds were recently described as ubiquitous contaminant of different compartiments of aquatic systems, including fish. In this study, we evaluated estrogenic activity of ten BPs in a model fish species, the zebrafish (Danio rerio), by combining newly developed in vitro and in vivo models. In vitro, three specific zebrafish reporter gene models expressing zebrafish estrogen receptors (ZfERs) in zebrafish hepatic cell line (ZFL) were used.Estrogenic activity of BPs was quantify in vivo by using a newly developed transgenic zebrafish line that expresses green fluorescent protein (GFP) under the control of the cyp19a1b promoter. The cyp19a1b gene codes for the brain aromatase B that is under the control of estrogens. Vitellogenin (Vtg), a marker of estrogenic compound, was used to assess the potential estrogenic effect of certain BPs in male fish. Five of 10 BPs induced an estrogenic activity on the three models: BP2, THB, BP1, 4BP, while BP and 234 BP were non active. We noted diffrence in BPs ranking depending on the zfER subtypes suggesting ?/? receptor selectivity by certain BPs. By quantifying GFP expression in live zebrafish, we confirm the estrogenic activity of some BP, i.e., BP1, THB, 4BP and 44'BP, as shown by their ability to induce GFP in a concentration-dependent manner. However, the ranking of BPs as a function of their estrogenic activity assay is different between models. For instance, 4BP was the most estrogenic compound in vivo but not in vitro. Differences were also noticed regarding the estrogenic activity of BP2. In vitro, BP2 has strong activity but had no effect on GFP in cyp19a1b-GFP embryo. However, BP2 induced strongly Vtg in male. The lack of estrogenic activity of BP2 in embryo may be related to difference of biodisponibility and/or metabolisation of this compound between embryo and adult fish. Our approach allows to characterize the estrogenic potency of emerging pollutants, using innovative in vitro and in vivo assays within a unique model fish species. The differences observed between assays deserve further studies to characterize the behavior of compounds in the different biological models in term of biodisponibility and metabolism.
1. European Conference on the Replacement, Reduction and Refinement of Animal Experiments in Ecotoxicology
Dübendorf, Switzerland

ineris-00971019
https://hal-ineris.archives-ouvertes.fr/ineris-00971019
INERIS : EN-2012-243

Éditeur(s) : HAL CCSD Elsevier
Résumé : International audience
Aryl hydrocarbon receptor-dependent genomic effects of environmental polycyclic aromatic hydrocarbons (PAHs) have been shown to be modulated by non-genomic protein kinase C (PKC)-related pathways. The present study was designed to determine whether PKC activation may also impair estrogenic genomic response triggered by PAHs. Treatment by the PKC activator phorbol 12-myristate 13-acetate (PMA) was found to markedly and differentially impair the up-regulation of estrogenic markers triggered by the estrogenic PAH benzanthracene (BZA) in cultured human mammary cells; BZA-mediated mRNA up-regulation of pS2 and amphiregulin was thus increased, whereas that of progesterone receptor and CXCL12 was repressed. BZA/PMA cotreatment however failed to alter BZA-mediated increase of activity of a luciferase gene reporter construct driven by an estrogen response element, thus discarding any global effect of PMA toward BZA-triggered estrogen receptor activation. Various chemicals inhibiting PKCs or extracellular signal-regulated kinase (ERK) as well as the knock-down of PKCδ expression counteracted the PMA-mediated increase of pS2 mRNA up-regulation triggered by BZA, demonstrating that it was dependent on PKCs, including PKCδ isoform, and ERKs. This non-genomic modulation of estrogenic effects of PAHs by PKC activation may have to be considered when considering the deleterious effects of these environmental contaminants towards the endocrine system.
ISSN: 0887-2333

inserm-00872862
http://www.hal.inserm.fr/inserm-00872862
DOI : 10.1016/j.tiv.2012.05.006
PUBMED : 22643241

Éditeur(s) : HAL CCSD Endocrine Society
Résumé : International audience
: Context:Masculinization depends on the fetal testis. Exposure of the human fetus during pregnancy to paracetamol or/and to other mild-analgesics is associated with an increased risk of cryptorchidism.Objective:We aimed at determining whether mild analgesics disrupted the morphology and endocrine function of the human testis.Design:We used an in vitro system based on the culture of human fetal testes exposed or not to paracetamol, its metabolite AM404, aspirin, indomethacin, ketoconazole at 10(-4) M to 10(-7) M.Setting:The study was conducted at the University of Rennes I.Patients/Participants:Human fetal testes were from pregnant women following induced abortion, between 7 and 12 weeks of gestation (GW).Main Outcome Measures:Testosterone (radioimmunoassay: RIA), Anti-Müllerian Hormone (AMH; ELISA), insulin-like factor 3 (INSL3; RIA), prostaglandin D2 and E2 (PGD2 and PGE2, respectively; ELISA), were assayed in the media. Testicular cells were counted using histology and image analysis. The possible nuclear receptor-mediated activities of the analgesics were investigated using reporter cell lines expressing estrogen (ERs), androgen (AR) and peroxysome proliferator-activated γ ( PPARγ) receptors.Results:Indomethacin and aspirin stimulated testosterone production, particularly by the younger testes (8-9 GW vs 10-12 GW). Paracetamol, AM404 and ketoconazole decreased INSL3 levels. Aspirin stimulated, whereas ketoconazole inhibited AMH production. PGE2 levels were inhibited by paracetamol, and aspirin in the 7-12 GW testes, and by indomethacin but only in 7-9.86 GW old testes. The inhibitory trends seen for PGD2 were not statistically significant.Conclusions:Analgesics at concentrations relevant to human exposure cause endocrine disturbances in the fetal testis. We suggest that the fetal human testis displays slight critical age windows for sensitivity to direct exposure to aspirin, indomethacin and paracetamol. The analgesic-induced inhibition of INSL3 may be the mechanism by which analgesics increase the risk of cryptorchidism. Greater caution is required concerning consumption of analgesics during pregnancy.
ISSN: 0021-972X

inserm-00876125
http://www.hal.inserm.fr/inserm-00876125
DOI : 10.1210/jc.2013-2531
PUBMED : 24030937

Éditeur(s) : HAL CCSD Elsevier
Résumé : International audience
: Zebrafish, Danio rerio, is increasingly used as an animal model to study the effects of pharmaceuticals and environmental estrogens. As most of these estrogens have only been tested on human estrogen receptors (ERs), it is necessary to measure their effects on zebrafish ERs. In humans there are two distinct nuclear ERs (hERα and hERβ), whereas the zebrafish genome encodes three ERs, zfERα and two zfERβs (zfERβ1 and zfERβ2). In this study, we established HeLa-based reporter cell lines stably expressing each of the three zfERs. We first reported that estrogens more efficiently activate the zfERs at 28°C as compared to 37°C, thus reflecting the physiological temperature of zebrafish in wildlife. We then showed significant differences in the ability of agonist and antagonist estrogens to modulate activation of the three zfER isotypes in comparison to hERs. Environmental compounds (bisphenol A, alkylphenols, mycoestrogens) which are hER panagonists and hERβ selective agonists displayed greater potency for zfERα as compared to zfERβs. Among hERα selective synthetic agonists, PPT did not activate zfERα while 16α-LE2 was the most zfERα selective compound. Altogether, these results confirm that all hER ligands control in a similar manner the transcriptional activity of zfERs although significant differences in selectivity were observed among subtypes. The zfER subtype selective ligands that we identified thus represent new valuable tools to dissect the physiological roles of the different zfERs. Finally, our work also points out that care has to be taken in transposing the results obtained using the zebrafish as a model for human physiopathology.
ISSN: 0041-008X

inserm-01067804
http://www.hal.inserm.fr/inserm-01067804
http://www.hal.inserm.fr/inserm-01067804/document
http://www.hal.inserm.fr/inserm-01067804/file/pintot_et_al._manuscript.pdf
DOI : 10.1016/j.taap.2014.07.020

Éditeur(s) : HAL CCSD Elsevier
Résumé : International audience
Co-exposure to environmental polycyclic aromatic hydrocarbons (PAHs) and interleukin (IL)-1β induces expression of the tumor-promoting cytokine IL-6 in cancer cells. The present study was designed to determine whether such an IL-6 up-regulation also occurs in response to co-treatment by PAHs and tumor necrosis factor (TNF)-α, an inflammatory cytokine commonly found in tumor microenvironment. Co-exposure to the prototypical PAH benzanthracene (BZA) and TNF-α was found to markedly induce mRNA expression and secretion of IL-6 in human breast cancer cells MCF-7, whereas exposure to either BZA or TNF-α alone was without significant effect. Co-treatment by BZA and TNF-α-containing conditioned media from human inflammatory macrophages similarly up-regulated IL-6 expression in MCF-7 cells. BZA/TNF-α-mediated IL-6 induction in MCF-7 cells was counteracted by silencing aryl hydrocarbon receptor (AhR), known to mediates most of PAH effects. IL-6 up-regulation was moreover associated with NF-κB activation and was abolished by using chemical NF-κB inhibitors or knocking-down expression of the p65/RelA NF-κB subunit. Taken together, these data indicate that co-exposure to BZA/TNF-α induces IL-6 expression by AhR- and NF-κB-dependent pathways in MCF-7 cancer cells. This regulation of IL-6 by environmental PAHs, that is dependent of inflammatory cytokine microenvironment, may contribute to the well-known carcinogenic properties of these organic pollutants.
ISSN: 0009-2797

inserm-00818896
http://www.hal.inserm.fr/inserm-00818896
DOI : 10.1016/j.cbi.2013.01.005
PUBMED : 23396138