Journal Article

Selective Activation of Zebrafish Estrogen Receptor Subtypes by Chemicals by Using Stable Reporter Gene Assay Developed in a Zebrafish Liver Cell Line

Anne Cosnefroy, François Brion, Emmanuelle Maillot-Maréchal, Jean-Marc Porcher, Farzad Pakdel, Patrick Balaguer and Selim Aït-Aïssa

in Toxicological Sciences

Volume 125, issue 2, pages 439-449
Published in print February 2012 | ISSN: 1096-6080
Published online November 2011 | e-ISSN: 1096-0929 | DOI: http://dx.doi.org/10.1093/toxsci/kfr297
Selective Activation of Zebrafish Estrogen Receptor Subtypes by Chemicals by Using Stable Reporter Gene Assay Developed in a Zebrafish Liver Cell Line

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The number of environmental chemical contaminants suspected to act as endocrine disruptor compounds by interacting with estrogen receptor (ER) signaling pathway has been continuously increasing. To study such interaction, the use of stable reporter gene assays is relevant, but species-specific in vitro screening assays are still lacking to address hazard assessment of estrogenic chemicals in aquatic vertebrates. Here, we describe the development of stable reporter gene assays based on stable expression of subtypes of zebrafish ER (zfERα, zfERβ1, and zfERβ2) coupled to estrogen response element–driven luciferase in a zebrafish liver (ZFL) cell line. The three established cell models, named ZELH-zfERα, ZELH-zfERβ1, and ZELH-zfERβ2, expressed stable and significant basal luciferase signal, which was induced by 17β-estradiol (E2) in a sensitive and dose-response manner at EC50s of 0.2, 0.03, and 0.05nM, respectively. In addition, E2 significantly altered cell proliferation in ZELH-zfERα and ZELH-zfERβ2 cells, but not in parental ZFL and ZELH-zfERβ1 cells, suggesting a functionality of these two receptors to modulate endogenous gene expression in the transfected clones. The screening of various xenoestrogens from different classes in the three models resulted in different luciferase response patterns. Natural and synthetic estrogens and 1,1,1-trichloro-2-(2 chlorophenyl)-2-(4-chlorophenyl)ethane were active at lower concentrations in ZELH-zfERβ1 and ZELH-zfERβ2 than in ZELH-zfERα cells, whereas genistein and zearalenone metabolites as well as three benzophenone derivatives preferentially activated zfERα. Altogether, the newly established models provide specific and convenient in vitro tool for comparative assessment of zfERs selective activation by chemicals within ZFL cell context.

Keywords: ZFL cells; zebrafish estrogen receptor subtypes; in vitro screening assay

Journal Article.  6406 words.  Illustrated.

Subjects: Medical Toxicology ; Toxicology (Non-medical)

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