Characterization of Triiodothyronine Transport and Accumulation in Rat Erythrocytes*

Abstract

The transport of L-T3 was studied in washed rat erythrocytes. L-T3 uptake was temperature sensitive: the initial velocity of uptake at low substrate concentration was 40 times higher at 37 C than at 0C whereas, at equilibrium, the ratio of cell-associated to extracellular L-T3 was about 7 times lower at 37 C than at 0 C. When [125I]L-T3-loaded erythrocytes were diluted into a serum albumin-containing medium, the efflux of L-T3 proceeded at a rate similar to that of influx. A large excess of unlabeled L-T3 in the medium blocked influx and efflux of labeled L-T3, indicating a saturable carrier-mediated transport process across the plasma membrane. the transport obeyed simple Michaelis-Menten kinetics with an apparent Km of 53 nM and a Vmax of 4.3 pmol/min.10(8) cells at 0 C. The Km increased only slightly with temperature whereas the Vmax was 100 times higher at 37 than at 0 C. The Arrhenius activation energy of uptake was 21 Cal/mol. The nonsaturable adsorption of L-T3 to the cells did not exceed 1% of the equilibrium levels at 0 C and 10% at 37 C. Uptake of L-T3 was very specific: unlabeled L-T4, D-T3, triiodothyroacetic acid, rT3, and DL-thyronine inhibited uptake with inhibition constant (Ki) values which were 35, 60, 65, 110, and 250 times, respectively, greater than the Km of L-T3. [125I]L-T4 uptake was negligible. L-T3 uptake and L-T4 inhibition of L-T3 uptake were pH dependent. It is suggested that only the unionized 4'-OH forms of the hormones were recognized by the transport system. At equilibrium, L-T3 was accumulated within the cell (apparent intracellular concentration approximately 50 times higher than that in the medium at 37 C). However, uptake was not dependent on the transmembrane Na+ gradient, suggesting facilitated rather than active transport. Analysis of L-T3 binding to erythrocyte cytosolic proteins suggested that they were implicated in the intracellular trapping of L-T3. At a concentration of 5 x 10(9) erythrocytes/ml (approximately the blood concentration), the amount of L-T3 accumulated in the cells was 13.5 times higher than the extracellular amount. We conclude that L-T3 is solely transported by a saturable, stereospecific, and Na+-independent carrier system. The intracellular accumulation and the rapid transmembrane movements of L-T3 suggest that erythrocytes might play a role in the interorgan transport of L-T3.

Molecular Characterization and Tissue Distribution of a New Organic Anion Transporter Subtype (oatp3) That Transports Thyroid Hormones and Taurocholate and Comparison with oatp2

“…The transport of thyroid hormones across the plasma membrane determines the intracellular concentration of these hormones and hence the activation of the nuclear T 3 receptor. The existence of mechanisms regulating the transport of thyroid hormones has been suggested in cerebrocortical neurons (17), astrocytes (18), glial cells (19), hepatocytes (20,21), erythrocytes (22), and skeletal muscle (23). The following are comparisons of the pharmacological characteristics among oatp2, oatp3, and the known native thyroid hormone transporters.…”

Transport of Thyroid Hormones by Human Erythrocytes: Kinetic Characterization in Adults and Newborns*

“…We have carried out a detailed kinetic characterization of the T 3 transport system in the rat erythrocytes and shown that T 3 , but not T 4 , is transported across the plasma membrane by a stereospecific Na + -independent carrier-mediated process and that T 3 is accumulated by intracellular trapping (21). We have also shown that T 3 transport exhibits typical trarcs-inhibition kinetics (22).…”

Membrane transporters for thyroid hormone

“…Also, T3 is taken up in erythrocytes by a low-affinity (Km in mM range) carrier-mediated mechanism [26][27][28][29][30].…”

“…Furthermore, uptake of both Trp and T3 is inhibited by treatment of erythrocytes with the Cys-specific reagent N-ethylmaleimide, and the inactivation of Trp transport by N-ethylmaleimide is prevented in the presence of T3 and vice versa. These data strongly suggest that thyroid hormone uptake in red blood cells is importantly mediated by a T-type amino acid transporter, that is, a transporter specific for aromatic amino acids [26][27][28][29][30].…”