Characterization of non-radiolabeled Thyroxine (T4) uptake in cryopreserved rat hepatocyte suspensions: Pharmacokinetic implications for PFOA and PFOS chemical exposure.

The alteration of thyroxine (T4) 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 T4 cellular uptake in cryopreserved rat hepatocyte suspensions. The environmental chemicals perfluorooctanoate (PFOA) and perfluorooctane sulfonate (PFOS) were evaluated for their effect on T4 hepatic uptake. At 37 °C, hepatic assays demonstrated saturable kinetics with increasing T4 concentrations, while a linear uptake rate consistent with passive diffusion was detected at 4 °C. Carrier-mediated (37-4 °C) transport of T4 was the predominant hepatic uptake process versus passive diffusion. Cyclosporin A (CsA) chemically inhibited T4 hepatic uptake, whereas PFOA/PFOS displayed no inhibition of T4 translocation. Increasing PFOA/PFOS concentration levels with the T4 serum carrier-protein transthyretin (TTR) present resulted in a dose-response increase in T4 hepatic uptake rates, correlating with increased T4 free fraction values. Hepatic assays conducted in the presence of PFOA/PFOS and TTR displayed an enhanced first-order T4 hepatic uptake rate consistent with carrier-mediated transport. These in vitro findings characterizing increased T4 hepatic uptake provides mechanistic insight regarding decreased T4 serum levels (hypothyroxinemia) previously observed within in vivo rodent studies following perfluorinated chemical exposure.