Impact of understudied PFAS on Lipid Metabolism in Hepatocyte 2D and 3D in vitro model

Perfluoroalkyl substances (PFAS) are industrial chemicals with several applications, including heat and water resistance. These chemicals are ubiquitous in our environment, and human exposure to them has been linked to liver injury and disruption of lipid metabolism. This study investigated the effects of PFAS (fluorotelomer sulfonic acids 6:2 FTSA and 8:2 FTSA, and perfluorooctane sulfonamide (PFOSA)) on hepatic lipid metabolism using in vitro human cell models. Initially, we assessed the effects of 10 μM PFOSA, 100 μM 6:2 FTSA, and 100 μM 8:2 FTSA in HepG2 monolayer (2D) cultures, focusing on their impact on lipid metabolism. Subsequently, HepG2 spheroids (3D) were exposed to PFOSA (10 μM), the most active compound tested, to elucidate its effects on lipid metabolism further. In the 3D culture, PFOS (10 μM) was used as a reference compound. The results showed that PFOSA was the most disruptive to gene expression and lipid profiles in both 2D and 3D HepG2 cell cultures among the tested PFAS. PFOSA repressed several genes involved in cholesterol and fatty acid synthesis in both HepG2 monolayer and spheroids (3D). Except for triacylglycerides(TAGs) and ceramide (Cer), most lipid species were significantly elevated in PFOSA-exposed HepG2 2D, compared to control. Notably, the effects of PFOSA were less pronounced in HepG2 spheroids. In HepG2 spheroids, the PFOSA toxicity profile resembled that of PFOS, suggesting a similar mechanism of action. Our findings highlight the need for further research on understudied PFAS.