A mechanistically-anchored and human stem cell-based in vitro test battery for assessing liver steatogenic potential of chemicals

Abstract

Fatty liver disease, which can result from various factors including chemical exposure, is an increasing clinical concern. A key event in its development is steatosis, referring to the accumulation of lipids within hepatocytes. To enable early detection of chemical-induced liver steatosis, we developed a mechanistically-anchored, human-relevant new approach methodology (NAM). This NAM consists of a 2-tiered in vitro test battery, aligned with the adverse outcome pathway (AOP) network for steatosis, and utilizes human skin-derived precursor cells differentiated into hepatic cells (hSKP-HPC), previously shown to be responsive to steatogenic triggers. In total, 6 well-known steatogenic compounds, including 3 pharmaceuticals (sodium valproate, tetracycline hydrochloride, amiodarone hydrochloride), a pesticide (cyproconazole), and 2 plasticizers (tricresyl phosphate and perfluorohexanesulfonic acid) alongside 2 non-steatogenic chemicals (minocycline hydrochloride and tartaric acid), were tested over a 72-hour period. Tier 1 evaluated transcriptional changes in key lipid metabolism pathways, and modulations were observed in nuclear receptors (peroxisome proliferator-activated receptor), fatty acid uptake (fatty acid translocase), de novo lipogenesis (diacylglycerol acyl transferase 2, fatty acid synthase and stearoyl-CoA desaturase 1), as well as in VLDL secretion (apolipoprotein B100). Tier 2 assays assessed and confirmed downstream functional disruptions in fatty acid uptake and lipid accumulation as ultimate specific key events for steatogenic chemicals. Overall, this human stem cell-based NAM offers a promising tool for supporting early hazard identification of steatogenic chemicals across diverse sectors, bridging mechanistic insights to outcomes relevant to the initiation of fatty liver disease.