Assessment of Thyroid Endocrine Disruption Effects of Parabens Using In Vivo, In Vitro, and In Silico Approaches

Abstract

The extensive applications of parabens in foods, drugs, and cosmetics cause inevitable exposure to humans. Revealing the developmental toxicity of parabens is of utmost importance regarding their safety evaluation. In this study, the effects of four commonly used parabens, including methyl paraben (20 ∼ 200 μM), ethyl paraben (20 ∼ 100 μM), propyl paraben (5 ∼ 20 μM), and butyl paraben (BuP, 2 ∼ 10 μM), were investigated on the early development of zebrafish embryos and larvae. The underlying mechanisms were explored from the aspect of their disturbance in the thyroid endocrine system using in vivo, in vitro, and in silico assays. Paraben exposure caused deleterious effects on the early development of zebrafish, with BuP displaying the highest toxicity among all, resulting in the exposure concentration-related mortality, decreased hatching rate, reduced body length, lowered heart rate, and the incidence of malformation. Further investigation showed that paraben exposure reduced thyroid hormone levels and disturbed the transcriptional expressions of the target genes in the hypothalamic–pituitary–thyroid axis. Molecular docking analysis combined with in vitro GH3 cell proliferation assay testified that all test parabens exhibited thyroid receptor agonistic activities. The findings confirmed the developmental toxicity of the test parabens and their thyroid endocrine disruption effects, providing substantial evidence on the safety control of paraben-based preservatives.

Azole-Induced Color Vision Deficiency Associated with Thyroid Hormone Signaling: An Integrated In Vivo, In Vitro, and In Silico Study

“…The downstream gene transactivation regulated by azole-bound TR was confirmed by a T-screen assay. ,, The T3 positive control enhanced GH3 proliferation indicating that TH-triggered TR-related transactivation is required for this biological response. , In the uniform concentration range, CBZ and TDF hindered T3-triggered GH3 cell proliferation but produced no cytotoxicity in the absence of T3 (Figure ). A dual-luciferase reporter assay using GH3 cells was conducted to rule out false positives due to cell death from compound toxicity and normalize well-to-well variations using an internal control reporter. , Significant decreases in T3-induced luciferase transcription were observed in azole-treated groups as in the T-screen assay (Figure ).…”

“…Many in silico methods, such as molecular docking, have been developed to study the interaction of xenobiotics with target receptors and to be a first step in determining the toxicological mode of action. ,− , Before investigating the interaction between azoles and TR isoforms, we used T3 as a positive control due to its known potent TR agonistic capacity . T3-TRα LBD and T3-TRβ LBD complexes produced low RMSD values (<1.2 Å), demonstrating good predictability of our docking protocol for identification of the binding site and ligand pose compared with the corresponding crystallographic structures. , Based on the lowest affinity score obtained by the blind molecular docking analysis, , CBZ and TDF preferred to occupy the binding pocket where T3, the native ligand for TR isoforms, was located (Figure S6).…”

“…Default parameters were used in the molecular docking calculation except that the number of genetic algorithm runs was configured at 200. The lowest energy represents a protein–ligand complex’s computational binding affinity . The pretreatment of compounds and proteins and the binding energy calculation were conducted in AutoDock Tools .…”

“…The lowest energy represents a protein− ligand complex's computational binding affinity. 17 The pretreatment of compounds and proteins and the binding energy calculation were conducted in AutoDock Tools. 28 The interaction profile between the compound and the amino acid of proteins based on the lowest docking energy was visually depicted by the PyMOL software (http://www.pymol.org).…”

“…The zebrafish retina possesses photoreceptor subtypes, and it is cone-dominant, resulting in richer color vision and higher acuity − …”