Characterization of non-radiolabeled Thyroxine (T) uptake in cryopreserved rat hepatocyte suspensions: Pharmacokinetic implications for PFOA and PFOS chemical exposure.

The alteration of thyroxine (T) cellular uptake by an environmental chemical can serve as a contributing factor in thyroid hormone (TH) disruption. Herein, we describe a non-radiolabeled (LC-MS/MS) oil-filtration technique designed to characterize the mechanism(s) responsible for T cellular uptake in cryopreserved rat hepatocyte suspensions. The environmental chemicals perfluorooctanoate (PFOA) and perfluorooctane sulfonate (PFOS) were evaluated for their effect on T hepatic uptake. At 37 °C, hepatic assays demonstrated saturable kinetics with increasing T concentrations, while a linear uptake rate consistent with passive diffusion was detected at 4 °C. Carrier-mediated (37-4 °C) transport of T was the predominant hepatic uptake process versus passive diffusion. Cyclosporin A (CsA) chemically inhibited T hepatic uptake, whereas PFOA/PFOS displayed no inhibition of T translocation. Increasing PFOA/PFOS concentration levels with the T serum carrier-protein transthyretin (TTR) present resulted in a dose-response increase in T hepatic uptake rates, correlating with increased T free fraction values. Hepatic assays conducted in the presence of PFOA/PFOS and TTR displayed an enhanced first-order T hepatic uptake rate consistent with carrier-mediated transport. These in vitro findings characterizing increased T hepatic uptake provides mechanistic insight regarding decreased T serum levels (hypothyroxinemia) previously observed within in vivo rodent studies following perfluorinated chemical exposure.