Comparative assessment of thyroid disrupting effects of ethiprole and its metabolites: In silico, in vitro, and in vivo study.

Ethiprole is widely used as a second-generation phenyl pyrazole insecticide. Previous studies indicated that ethiprole exhibited thyroid toxicity while two main metabolites (ethiprole sulfone (M1) and ethiprole sulfide (M2)) of ethiprole showed higher acute toxicity than ethiprole. Therefore, assessing the thyroid toxicity of its metabolites is crucial for safety assessment. In this study, the thyroid toxicity and underlying mechanisms of ethiprole and its metabolites were explored using in silico, in vitro, and in vivo assays, with the aim of conducting a comparative study on thyroid toxicity. Molecular docking analysis showed that ethiprole, M1 and M2 could bind with thyroid receptor isoforms and exhibited higher binding affinity compared to 3,3',5-triiodothyronine (T3). GH3 cell proliferation assays revealed that ethiprole, M1 and M2 all served as thyroid hormone antagonists to hinder the T3-induced cell proliferation. Using the zebrafish model, we further investigated that exposure to ethiprole, M1, and M2 disrupted thyroid hormone levels and the transcriptional expressions of hypothalamus-pituitary-thyroid (HPT) axis-related genes. Ethiprole induced thyroid disrupting effects by binding with the thyroid receptor beta, M1 mainly through binding with the corticotropin releasing factor receptor-1, and M2 exposure firstly inhibited the thyroid peroxidase enzyme activity. M2 showed the highest developmental toxicity and thyroid disrupting effects, which significantly reducing hatching rates, increasing deformity rates, exhibiting the lowest lethal concentration 50 value and showing the most serious transcription inhibitory effects on the HPT axis. This study suggested the risk assessment of metabolites should be considered in assessing potential environmental risk of ethiprole.