In vitro binding of human and rat PPAR alpha, beta/delta, and gamma receptors to PFAS, fatty acids, and clofibric acid
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The molecular mechanism by which some per- and polyfluorinated alkyl substances (PFAS) exert toxicity has largely been attributed to activation of peroxisome proliferator-activated receptor alpha (PPARα). Less attention has been given to PPAR beta/delta (PPARβ/δ) or PPAR gamma (PPARγ). Further, it is unknown how, or if, PFAS activation of PPAR paralogs differs between rats and humans. We utilized in vitro luciferase reporter assays with either human or rat PPARα, β/δ, or γ ligand binding domains hybridized with a Gal4 DNA binding domain. We also tested for estrogen (ER), androgen (AR), and glucocorticoid receptor (GR) transcriptional activation using previously established stable transfection (ER) and viral transduction (AR, GR) luciferase reporter assays. Nine PFAS (hexafluoropropylene oxide-dimer acid ammonium salt (GenXAS) and free acid (GenXFA), nafion byproduct 2 (NBP2), perfluorooctanoic acid (PFOA), perfluorooctane sulfonate (PFOS), perfluorobutane sulfonate (PFBS), perfluorononanoic acid (PFNA), 6:2 fluorotelomer alcohol (6:2 FTOH), and perfluoromethoxyacetic acid (PFMOAA)), 3 endogenous fatty acids (oleic, linoleic, and octanoic), and the drug metabolite clofibric acid were assayed at 30 µM -1 mM. Thus far all compounds, except 6:2 FTOH, have activated both PPARα and PPARγ in both human and rat. Based on EC20 values, oleic and linoleic acid were the most potent human PPARα activators, while PFOS was the least potent. For rat PPARα, GenX was the most potent activator (no difference between GenXFA and GenXAS), while NBP2 and PFMOAA were the least with low fold induction (<10% of max). For PPARγ, all compounds had similar potency in the human and rat receptor assays (log EC20 ranges: -3.14 – -3.74 versus -3.05 – -3.53), except for PFMOAA which only reached 3% and 1% of max fold induction, respectively. In contrast, the only compounds that activated both human and rat PPARβ/δ were oleic and linoleic acid; however, NBP2 weakly activated (~10% of max) rat PPARβ/δ at 300 µM. No compounds displayed in vitro AR or GR transcriptional activation. Only 6:2 FTOH exhibited weak ER agonism. Overall, all PFAS studied, except 6:2 FTOH, displayed both human and rat PPARα and γ activity, with some having similar potency to endogenous free fatty acids. Thus, it is likely that both PPARα and γ activation contribute to the adverse in vivo effects observed for PFAS. Abstract does not necessarily reflect USEPA policy.