title: A screening assay for thyroid hormone signaling disruption based on thyroid hormone-response gene expression analysis in the frog Pelophylax nigromaculatus.
abstract: Amphibian metamorphosis provides a wonderful model to study the thyroid hormone (TH) signaling disrupting activity of environmental chemicals, with Xenopus laevis as the most commonly used species. This study aimed to establish a rapid and sensitive screening assay based on TH-response gene expression analysis using Pelophylax nigromaculatus, a native frog species distributed widely in East Asia, especially in China. To achieve this, five candidate TH-response genes that were sensitive to T3 induction were chosen as molecular markers, and T3 induction was determined as 0.2 nmol/L T3 exposure for 48 hr. The developed assay can detect the agonistic activity of T3 with a lowest observed effective concentration of 0.001 nmol/L and EC50 at around 0.118-1.229 nmol/L, exhibiting comparable or higher sensitivity than previously reported assays. We further validated the efficiency of the developed assay by detecting the TH signaling disrupting activity of tetrabromobisphenol A (TBBPA), a known TH signaling disruptor. In accordance with previous reports, we found a weak TH agonistic activity for TBBPA in the absence of T3, whereas a TH antagonistic activity was found for TBBPA at higher concentrations in the presence of T3, showing that the P. nigromaculatus assay is effective for detecting TH signaling disrupting activity. Importantly, we observed non-monotonic dose-dependent disrupting activity of TBBPA in the presence of T3, which is difficult to detect with in vitro reporter gene assays. Overall, the developed P. nigromaculatus assay can be used to screen TH signaling disrupting activity of environmental chemicals with high sensitivity.
authors.affiliation: State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-environmental Sciences, Chinese Academy of Sciences, Beijing 100085, China.; Department of Environmental Bio-Technology, Research Center for Eco-environmental Sciences, Chinese Academy of Sciences, Beijing 100085, China. Electronic address: lingboxian.yin@163.com.
authors.forename: Yinfeng
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-environmental Sciences, Chinese Academy of Sciences, Beijing 100085, China.
authors.forename: Yuanyuan
authors.initials: Y
authors.lastname: Li
authors.affiliation: State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-environmental Sciences, Chinese Academy of Sciences, Beijing 100085, China.. Electronic address: qinzhanfen@rcees.ac.cn.
authors.forename: Zhanfen
authors.initials: Z
authors.lastname: Qin
authors.affiliation: Department of Environmental Bio-Technology, Research Center for Eco-environmental Sciences, Chinese Academy of Sciences, Beijing 100085, China. Electronic address: huiliwang@rcees.ac.cn.
authors.forename: Huili
authors.initials: H
authors.lastname: Wang
authors.affiliation: Department of Environmental Bio-Technology, Research Center for Eco-environmental Sciences, Chinese Academy of Sciences, Beijing 100085, China.
authors.forename: Jianzhong
authors.initials: J
authors.lastname: Li
mesh_terms: D000818:Animals; D052244:Endocrine Disruptors / Q000633:toxicity*; D004784:Environmental Monitoring / Q000379:methods*; D005411:Flame Retardants / Q000633:toxicity; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects*; D007413:Intestinal Mucosa / Q000378:metabolism; D007422:Intestines / Q000187:drug effects; D007814:Larva / Q000187:drug effects / Q000254:growth & development / Q000378:metabolism; D008675:Metamorphosis, Biological / Q000187:drug effects; D011075:Polybrominated Biphenyls / Q000633:toxicity*; D011898:Ranidae / Q000254:growth & development / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D013963:Thyroid Hormones / Q000378:metabolism*; D014874:Water Pollutants, Chemical / Q000633:toxicity*
chemical_list: D052244:Endocrine Disruptors; D005411:Flame Retardants; D011075:Polybrominated Biphenyls; D013963:Thyroid Hormones; D014874:Water Pollutants, Chemical; C020806:tetrabromobisphenol A
keywords: Pelophylax nigromaculatus; Screening assay; TH-response gene expression; Tetrabromobisphenol A; Thyroid hormone signaling disruptor