Development of a human stem cell-based in vitro model to assess toxicity hazards induced by thyroid hormone system disruptors during the early phases of liver development

Thyroid hormone system disruptors (THSDs) pose a significant threat by disturbing the intricate processes of thyroid hormone (TH) production, distribution, metabolization, and secretion, crucial for fetal development. Their interference extends to the liver, a key player in TH transport and metabolism. Despite some elucidated mechanisms of THSD action, many remain poorly understood, compounded by the reliance on rodent models, which do not fully mirror human physiology. To address this gap, a novel in vitro toxicity model leveraging human stem cells is created, focusing on alterations caused by THSDs during the early phases of liver development. As such, multipotent human skin derived precursor cells (hSKPs) were differentiated into “hepatic progenitor cells” (hSKP-HPCs) to mimic in vivo liver development in vitro and enable studying ‘human foetal’ hepatic TH signalling. Triiodthyronin and thyroxin - essential THs to regulate the basal metabolic rate of all cells including hepatocytes - were incorporated into the culture throughout differentiation at levels comparable to systemic in vivo concentrations. Current culture methodology involves 2D monolayer formation, which is known to fall short in capturing the full complexity of the in vivo microenvironment and induce structural and functional alterations to the cells. Hence, a transition to a 3D setup was additionally studied to better mimic natural conditions and functional alterations. Characterization efforts are ongoing and encompass gene expression and protein profiling to elucidate hepatic aspects and TH signalling pathways in the presence or absence of THs and in relation to the cells’ spatial configuration. Additionally, metabolic function evaluation involves quantifying TH metabolite concentrations in the supernatant via LC/MS/MS analysis. Once the optimization and characterization of the hSKP-HPC cell system is completed, it will be evaluated for its potency to confirm known and identify new potential THSDs. This in vitro system aims to support
next generation animal-free identification of THSDs.