T. Nguyen-Jones1, J. McKenna2, W. Chen2, K. K. Wolf3, T. Stone3, E. Bianchi2, J. R. LaRocca2, J. Chen1, E. LeCluyse3, J. Klein1, E. Rogers1
1.LifeNet Health LifeSciences, Virginia Beach, VA, 2. Corteva Agriscience, Indianapolis, IN; 3LifeNet Health LifeSciences, Research Triangle Park, NC
Background and Purpose:
To streamline efforts in identifying and evaluating chemicals that may interfere with the human endocrine system while chemical production and use continue to rise in the 21st century, the U.S. EPA’s Endocrine Disruptor Screening Program (EDSP) has established a tiered testing strategy to screen the extensive list of chemicals. Tier 1 tests focus on screening to identify chemicals that potentially interact with the endocrine system while Tier 2 involves long-term studies of the adverse effects. Cell lysates, microsomes, cell lines, or engineered cells are often employed as in vitro methods for chemical screening. However, these approaches lack human physiological relevance as the native thyroid phenotype and ability to synthesize thyroid hormones are absent. Emerging organoid models and organ-on-a-chip technologies often offer better recapitulation of biological processes. This study evaluated the use of cryopreserved primary human thyrocytes (P1) as an in vitro model for testing thyroid-disrupting chemicals (TDCs). Both ELISAs and LC-MS/MS were employed as quantitative methods to determine the levels of T4 and T3 synthesized. Results indicated that all donor lots formed microtissues and T4 levels met the quality benchmark for TDC applications (>1.0 ng/mL)1. Dose response curves and IC50 values were obtained for all donor lots.
Conclusions:
- Microtissue formation were achieved in all 3 donor lots with measured T4 levels that met the current TDC applications criteria on day 14 (>1.0 ng/mL) as defined in Foley et al. publication.
- Dose-response curves and IC50 values for T4 were consistent across all donors as well as quantifying methods (ELISA and LC- MS/MS) indicating assay reliability.
- Vehicle controls’ T4/T3 ratios for two out of the three tested donor lots were > 2.5 (2218233 and 2318558) and the ratio for the third donor lot was >2.0 (2319211).
- Cryopreserved primary human thyrocytes offer a promising approach for evaluating potential thyroid disrupting chemicals and molecules.