A rapid assay of human thyroid peroxidase activity

Dong, Hongyan; Godlewska, Marlena; Wade, Michael G.

doi:https://open-science.canada.ca/handle/123456789/2196

A rapid assay of human thyroid peroxidase activity

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dc.contributor.author: Dong, Hongyan; Godlewska, Marlena; Wade, Michael G.
dc.date.accessioned: 2024-03-25T14:30:49Z
dc.date.available: 2024-03-25T14:30:49Z
dc.date.issued: 2019-10-16
dc.description - en: Health Canada’s mandate requires the review and assessment of potential health risks due to commercial chemical use in Canada to manage and mitigate these risks. For many chemicals in commerce, information about potential toxicity to vulnerable parts of the endocrine system is lacking. Disruption of thyroid hormone synthesis is recognized as a key mechanism by which some endocrine disrupting chemicals can cause impaired brain development and other adverse effects. The current study modified a rapid method to screen chemicals for inhibition of a critically vulnerable enzyme (thyroid peroxidase - TPO) necessary for thyroid hormone production to make this assay more relevant for human hazard characterization and less dependent on lab animal use. Activity of human TPO (hTPO) – harvested from either of two transgenic cell lines expressing high levels of the hTPO - was compared to TPO activity harvested from rat thyroid glands in its ability to identify TPO inhibiting activity among a group of chemicals with known effect. Human TPO was as effective as the rat enzyme in identifying chemicals with moderate to high potency. Chemicals shown to have weak activity on rat TPO were less effectively identified with hTPO from either cell line. These results demonstrate that hTPO from transgenic cells can be used for a rapid assay to reliably identify TPO-inhibiting chemicals. These results will be included in Health Canada’s contribution to international efforts at the Organization of Economic Cooperation and Development to developing test methods to screen and test chemicals for endocrine disrupting activity.
dc.description.abstract - en: Impaired synthesis or action of thyroid hormones (THs) during critically sensitive periods of development can have long term adverse effects on health. Development of rapid assays to identify chemicals that impair THs physiology is an important goal for reducing risks from chemical use. Thyroid peroxidase (TPO) is a key enzyme regulating THs synthesis in thyroid gland and a vulnerable target for chemicals that disrupt THs synthesis. To develop a human-relevant, rapid assay for TPO inhibition, we have engineered two cell lines (CHO and LentiX- 293) to express active human TPO (hTPO) enzyme and applied them in a recently-described assay using a stable fluorescent product (Amplex UltraRed). Assay performance was assessed by comparing activity of 19 reference chemicals with known strong, weak or no TPO inhibitory activity. The assay using hTPO from either cell line consistently identified the relative potency of strong to moderate inhibitors and chemicals known to be inactive. Results were less consistent for chemicals reported to be weak inhibitors of rodent TPO, possibly suggesting some species specificity. Our studies support the use of hTPO from stably transfected cell lines to substitute for animal-derived thyroid microsomes for rapid high throughput screening assays to identify and characterize TPO inhibitors.
dc.description.abstract-fosrctranslation - fr: Une synthèse ou une action altérée des hormones thyroïdiennes (TH) pendant les périodes de développement critiques peuvent avoir des effets néfastes à long terme sur la santé. Le développement de tests rapides pour identifier les produits chimiques qui altèrent la physiologie des TH est un objectif important pour réduire les risques liés à l'utilisation de produits chimiques. La peroxydase thyroïdienne (TPO) est une enzyme clé régulant la synthèse des TH dans la glande thyroïde et une cible vulnérable aux produits chimiques qui perturbent la synthèse des TH. Pour développer un test rapide et pertinent pour l'homme pour l'inhibition de la TPO, nous avons conçu deux lignées cellulaires (CHO et LentiX-293) pour exprimer l'enzyme TPO humaine active (hTPO) et les avons appliquées dans un test récemment décrit utilisant un produit fluorescent stable ( Amplex UltraRouge). Les performances du test ont été évaluées en comparant l’activité de 19 produits chimiques de référence présentant une activité inhibitrice de la TPO forte, faible ou inexistante. Le test utilisant la hTPO de l’une ou l’autre lignée cellulaire a identifié de manière cohérente la puissance relative des inhibiteurs forts à modérés et des produits chimiques connus pour être inactifs. Les résultats étaient moins cohérents pour les produits chimiques signalés comme étant de faibles inhibiteurs de la TPO des rongeurs, suggérant peut-être une certaine spécificité d'espèce. Nos études soutiennent l'utilisation de hTPO provenant de lignées cellulaires transfectées de manière stable pour remplacer les microsomes thyroïdiens d'origine animale pour des tests de criblage rapides à haut débit afin d'identifier et de caractériser les inhibiteurs de la TPO.
dc.description.fosrctranslation - fr: Health Canada’s mandate requires the review and assessment of potential health risks due to commercial chemical use in Canada to manage and mitigate these risks. For many chemicals in commerce, information about potential toxicity to vulnerable parts of the endocrine system is lacking. Disruption of thyroid hormone synthesis is recognized as a key mechanism by which some endocrine disrupting chemicals can cause impaired brain development and other adverse effects. The current study modified a rapid method to screen chemicals for inhibition of a critically vulnerable enzyme (thyroid peroxidase - TPO) necessary for thyroid hormone production to make this assay more relevant for human hazard characterization and less dependent on lab animal use. Activity of human TPO (hTPO) – harvested from either of two transgenic cell lines expressing high levels of the hTPO - was compared to TPO activity harvested from rat thyroid glands in its ability to identify TPO inhibiting activity among a group of chemicals with known effect. Human TPO was as effective as the rat enzyme in identifying chemicals with moderate to high potency. Chemicals shown to have weak activity on rat TPO were less effectively identified with hTPO from either cell line. These results demonstrate that hTPO from transgenic cells can be used for a rapid assay to reliably identify TPO-inhibiting chemicals. These results will be included in Health Canada’s contribution to international efforts at the Organization of Economic Cooperation and Development to developing test methods to screen and test chemicals for endocrine disrupting activity.
dc.identifier.doi: https://doi.org/10.1016/j.tiv.2019.104662
dc.identifier.uri: https://open-science.canada.ca/handle/123456789/2196
dc.language.iso: en
dc.publisher: Elsevier
dc.subject - en: Health; Health and safety
dc.subject - fr: Santé; Santé et sécurité
dc.subject.en - en: Health; Health and safety
dc.subject.fr - fr: Santé; Santé et sécurité
dc.title - en: A rapid assay of human thyroid peroxidase activity
dc.type - en: Article
dc.type - fr: Article

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