High-Throughput Transcriptomic Analysis of Human Primary Hepatocyte Spheroids Exposed to Per- and Polyfluoroalkyl Substances as a Platform for Relative Potency Characterization

Abstract

Per- and poly-fluoroalkyl substances (PFAS) are widely found in the environment because of their extensive use and persistence. Although several PFAS are well studied, most lack toxicity data to inform human health hazard and risk assessment. This study focussed on four model PFAS: perfluorooctanoic acid (PFOA; 8 carbon), perfluorobutane sulfonate (PFBS; 4 carbon), perfluorooctane sulfonate (PFOS; 8 carbon), and perfluorodecane sulfonate (PFDS; 10 carbon). Human primary liver cell spheroids (pooled from 10 donors) were exposed to 10 concentrations of each PFAS and analyzed at four time-points. The approach aimed to: (1) identify gene expression changes mediated by the PFAS; (2) identify similarities in biological responses; (3) compare PFAS potency through benchmark concentration analysis; and (4) derive bioactivity exposure ratios (ratio of the concentration at which biological responses occur, relative to daily human exposure). All PFAS induced transcriptional changes in cholesterol biosynthesis and lipid metabolism pathways, and predicted PPARα activation. PFOS exhibited the most transcriptional activity and had a highly similar gene expression profile to PFDS. PFBS induced the least transcriptional changes and the highest benchmark concentration (i.e., was the least potent). The data indicate that these PFAS may have common molecular targets and toxicities, but that PFOS and PFDS are the most similar. The transcriptomic bioactivity exposure ratios derived here for PFOA and PFOS were comparable to those derived using rodent apical endpoints in risk assessments. These data provide a baseline level of toxicity for comparison with other known PFAS using this testing strategy.

Integration of Toxicogenomics and Physiologically Based Pharmacokinetic Modeling in Human Health Risk Assessment of Perfluorooctane Sulfonate

“…Gene profiling was conducted using Affymetrix 430_2 microarrays. In GSE144775, the human primary hepatocytes and Kupffer cells from 10 different human liver donors were cultured in vitro . The cells were exposed to PFOS for 1, 4, 10, or 14 days at doses of 0, 0.02, 0.1, 0.2, 1, 2, 10, and 20 μM.…”

Adverse Effects of Perfluorooctane Sulfonate on the Liver and Relevant Mechanisms

“…PFOS increased the expression of PPARα target genes Acox1 and CYP4A10 in mouse liver, thereby activating PPARα and causing pathological changes in the liver tissue [ 136 ]. Recently, in a study using a primary human hepatocyte spheroid model, PPARα activity was found to be increased by PFAS exposure in human liver spheroids, particularly by PFOS exposure [ 137 ]. Previous studies have shown that PFOS can differentially activate PPARα in mice and humans [ 138 , 139 ].…”

The use of in vitro methods in assessing human health risks associated with short‐chain perfluoroalkyl and polyfluoroalkyl substances (PFAS)

“…The use of well‐characterized cell lines offers a wide array of possibilities for manipulating experimental variables to obtain a multitude of endpoints in a timely manner. The utility of screening methods employing HTS methods has already been demonstrated for PFAS chemicals (Reardon et al, 2020; Rowan‐Carroll et al, 2021; Singh & Hsieh, 2021). While not inclusive of all assays, endpoints, and cell types, Table 2 provides some examples of approaches that can be and are being used for assessing the human health effects of short‐chain PFAS.…”