Developing adverse outcome pathways underlying CAR activation-induced liver injuries using HepG2 spheroid model.

Area of Science:

• Hepatology
• Toxicology
• Cell Biology

Background:

• Constitutive androstane receptor (CAR) is crucial for processing foreign compounds in the liver.
• Understanding CAR's role in chemical-induced liver injury is limited by current models.
• A 3D HepG2 spheroid model offers a more advanced platform for studying liver toxicity.

Purpose of the Study:

• To investigate chemical-induced liver injuries mediated by CAR activation using a 3D HepG2 spheroid model.
• To compare the efficacy of 3D versus 2D cell cultures in assessing CAR-mediated toxicity.
• To elucidate the mechanisms of CAR-induced hepatotoxicity, DNA damage, and dyslipidemia.

Main Methods:

• Utilized a 3D HepG2 spheroid model for enhanced liver toxicity studies.
• Employed immunofluorescence to track CAR localization.
• Conducted transcriptomics, knockdown experiments, DNA damage assays, and lipidomics.

Main Results:

• The 3D HepG2 model demonstrated superiority over 2D cultures for studying metabolizing enzymes and hepatic pathways.
• CAR activation was confirmed to mediate hepatotoxicity induced by acetaminophen and tetrabromodiphenyl ether.
• CAR was found to mediate DNA damage from aristolochic acids and lipid accumulation from perfluorooctanoic acid and nonylphenol.

Conclusions:

• The 3D HepG2 spheroid model provides a robust platform for studying CAR-mediated liver toxicity.
• CAR activation is a key mediator of chemical-induced hepatotoxicity, DNA damage, and dyslipidemia.
• Adverse outcome pathways linking CAR activation to specific toxicities were established, offering insights into exogenous chemical effects on the liver.