An in vivo multiwell-based fluorescent screen for monitoring vertebrate thyroid hormone disruption.
There is a pressing need for high throughput methods to assess potential effects of endocrine disrupting chemicals (EDCs). released into the environment. Currently our ability to identify effects in vitro exceeds that for in vivo monitoring. However, only in vivo analysis provides the full spectrum of physiological impacts exerted by a given chemical. With the aim of finding a physiological system compatible with automatic plate reading we tested the capacity of early embryonic stage Xenopus laevis tadpoles to monitor thyroid hormone (TH) disruption. Fluorescent transgenic X. laevis embryos bearing a TH/bZIP-eGFP construct, placed in 96 well plates, were used for a physiological-based screen for potential TH signaling disruptors. Using stage NF-45 embryos (time of thyroid gland formation) allowed rapid detection of chemical interference with both peripheral TR signaling and production of endogenous TH. Nanomolar concentrations of TH receptor agonists could be detected within 72 h. Moreover, when testing against a 5nM T3 challenge, the effects of inhibitors of TH production were revealed, including inhibitors of TH synthesis, (methimazole: 1 mM or sodium perchlorate: 3.56 microM), as well as antagonists acting at the receptor level (NH3: 2 microM) and a deiodinase inhibitor (iopanoic acid: 10 microM). Finally, we show that the thyroid disrupting activities of BPA (10 microM) and TBBPA (1 microM) can also be detected in this rapid screening protocol. Finally, this noninvasive technology using an automatic reading system shows low variability (around 5%) and permits detection of subtle changes in signaling by EDCs that either inhibit or activate TH signaling in vivo.
Fini
Jean-Baptiste
JB
UMR CNRS 5166, Evolution des Régulations Endocriniennes, Department of Regulations, Development and Molecular Diversity, Muséum National d'Histoire Naturelle, 7 Rue Cuvier, 75231 Paris 05, France.
Le Mevel
Sebastien
S
Turque
Nathalie
N
Palmier
Karima
K
Zalko
Daniel
D
Cravedi
Jean-Pierre
JP
Demeneix
Barbara A
BA
D000818:Animals; D030801:Animals, Genetically Modified; D013956:Antithyroid Agents / Q000378:metabolism* / Q000494:pharmacology; D001559:Benzhydryl Compounds; D004625:Embryo, Nonmammalian / Q000187:drug effects; D004784:Environmental Monitoring; D005260:Female; D007814:Larva / Q000187:drug effects; D008163:Luminescent Measurements / Q000379:methods*; D010636:Phenols / Q000494:pharmacology; D011075:Polybrominated Biphenyls / Q000494:pharmacology; D013963:Thyroid Hormones / Q000819:agonists / Q000378:metabolism*; D014158:Transcription, Genetic / Q000187:drug effects; D014284:Triiodothyronine / Q000494:pharmacology; D014714:Vertebrates / Q000196:embryology / Q000378:metabolism; D014982:Xenopus laevis; D000098826:Bisphenol A Compounds
D013956:Antithyroid Agents; D001559:Benzhydryl Compounds; D010636:Phenols; D011075:Polybrominated Biphenyls; D013963:Thyroid Hormones; D014284:Triiodothyronine; C020806:tetrabromobisphenol A; C006780:bisphenol A; D000098826:Bisphenol A Compounds