In vivo Characterization of Endocrine Disrupting Chemical Effects via Thyroid Hormone Action Indicator Mouse

Summary

The Thyroid Hormone Action Indicator mouse model was developed to enable tissue-specific quantification of local thyroid hormone action using its endogenous regulatory machinery. Recently, it has been shown that the model is suitable for characterizing endocrine-disrupting chemicals interacting with thyroid hormone economy, both by ex vivo and in vivo methodologies.

Abstract

Thyroid hormones (TH) play a critical role in cell metabolism and tissue function. TH economy is susceptible to endocrine disrupting chemicals (EDCs) that can disturb hormone production or action. Many environmental pollutants are EDCs, representing an emerging threat to both human health and agricultural production. This has led to an increased demand for proper test systems to examine the effects of potential EDCs. However, current methodologies face challenges. Most test systems use endogenous markers regulated by multiple, often complex regulatory processes, making it difficult to distinguish direct and indirect effects. Moreover, in vitro test systems lack the physiological complexity of EDC metabolism and pharmacokinetics in mammals. Additionally, exposure to environmental EDCs usually involves a mixture of multiple compounds, including in vivo generated metabolites, so the possibility of interactions cannot be ignored. This complexity makes EDC characterization difficult. The Thyroid Hormone Action Indicator (THAI) mouse is a transgenic model that carries a TH-responsive luciferase reporter system, enabling the assessment of tissue-specific TH action. One can evaluate the tissue-specific effects of chemicals on local TH action by quantifying luciferase reporter expression in tissue samples. Furthermore, with in vivo imaging, the THAI mouse model allows for longitudinal studies on the effects of potential EDCs in live animals. This approach provides a powerful tool for testing long-term exposure, complex treatment structures, or withdrawal, as it enables the assessment of changes in local TH action over time in the same animal. This report describes the process of in vivo imaging measurements on THAI mice. The protocol discussed here focuses on developing and imaging hyper- and hypothyroid mice, which can serve as controls. Researchers can adapt or expand the treatments presented to meet their specific needs, offering a foundational approach for further investigation.

Introduction

Thyroid hormone (TH) signaling is a fundamental regulator of cellular metabolism, essential for normal development and optimal tissue function in adulthood¹. Within tissues, TH action is finely controlled by a complex molecular machinery, allowing for tissue-specific maintenance of local TH levels. This autonomy of different tissues from circulating TH levels is of great importance^2,3,4.

Numerous chemicals have the potential to disrupt endocrine functions and are found in the environment as pollutants. It is a growing concer....

Protocol

The present protocol was reviewed and approved by the Animal Welfare Committee at the Institute of Experimental Medicine (PE/EA/1490-7/2017, PE/EA/106-2/2021). The presented data is from FVB/Ant background¹⁴, 3-month-old male THAI mice (n = 3-6/group). FVB/Ant background THAI animals tend to have highly pigmented spots on their skin that may distort measurements. Hence, search for pigmented spots on the skin of the imaged area after fur removal. Animals do not require special housing conditions un.......

Discussion

The threats posed by Endocrine-Disrupting Chemicals (EDCs) to human health are well recognized; however, research on EDCs faces formidable challenges. These challenges are partially a consequence of the complexity of the endocrine system. Many EDCs have been identified to simultaneously disrupt multiple endocrine systems²². Additionally, in the context of Thyroid Hormone (TH) economy, there exists an additional layer of complexity due to tissue-specific differences in regulating TH action. This co.......

Materials

Name	Company	Catalog Number	Comments
3,5,3'-triiodothyronine (T3)	Merck	T2877
Animals, mice			THAI mouse
Eye protection gel	Oculotect		1000 IU/g
Falcon tube	Thermo Fisher Scientific		50 mL volume
Iodine-free chow diet	Research Diets	custom
IVIS Lumina II in vivo imaging system	Perkin Elmer	-
Ketamine	Vetcentre	E1857
Living Image software 4.5	Perkin Elmer	-	provided with the instrument
Measuring cylinder			250 mL
methimazole	Merck	M8506
Microfuge tubes	Eppendorf		For diluting treatment materials
NaClO4	Merck	71852
Na-luciferin, substrate	Goldbio	103404-75-7
NaOH	Merck	101052833
Phoshphate buffer saline	Chem Cruz	sc-362302
Pipette	Gilson		For diluting treatment materials
Pipette tips	Axygen		For diluting treatment materials
Shaving cream/epilator/shaver			Personal preference
Syringe	B Braun		1 mL volume
Syringe needle	B Braun		0.3 x 12 mm
Xylazine	Vetcentre	E1852