Comparison of mechanisms mediating uptake and efflux of thyroid hormones in the human choriocarcinoma cell line, JAR.

We compared the specificities of transport mechanisms for uptake and efflux of thyroid hormones in cells of the human choriocarcinoma cell line, JAR, to determine whether triiodothyronine (T3), thyroxine (T4) and reverse T3 (rT3) are carried by the same transport mechanism. Uptake of 125I-T3, 125I-T4 and 125I-rT3 was saturable and stereospecific, but not specific for T3, T4 and rT3, as unlabelled L-stereoisomers of the thyroid hormones inhibited uptake of each of the radiolabelled hormones. Efflux of 125I-T3 was also saturable and stereospecific and was inhibited by T4 and rT3. Efflux of 125I-T4 or 125I-rT3 was, in contrast, not significantly inhibited by any of the unlabelled thyroid hormones tested. A range of compounds known to interfere with receptor-mediated thyroid hormone uptake in cells inhibited uptake of 125I-T3 and 125I-rT3, but not 125I-T4. We conclude that in JAR cells uptake and efflux of 125I-T3 are mediated by saturable and stereospecific membrane transport processes. In contrast, the uptake, but not the efflux, of 125I-T4 and 125I-rT3 is saturable and stereospecific, indicating that uptake and efflux of T4 and rT3 in JAR cells occur by different mechanisms. These results suggest that in JAR cells thyroid hormones may be transported by at least two types of transporters: a low affinity iodothyronine transporter (Michaelis constant, Km, around 1 microM) which interacts with T3, T4 and rT3, but not amino acids, and an amino acid transporter which takes up T3, but not T4 or rT3. Efflux of T4 and rT3 appears to occur by passive diffusion in these cells.

Uptake of reverse T3 in the human choriocarcinoma cell line, JAr.

The uptake and efflux of reverse triiodothyronine (rT3) in JAr cells were investigated. Uptake of 125I-rT3 was time dependent and reversible with a saturable component of around 70 per cent of total uptake after 30 min of incubation. Efflux was not saturable. Kinetic analysis of the initial specific uptake rates revealed an uptake process with a Michaelis constant of 3.04+/-0.53 microM (mean+/-SEM, n=15) and a corresponding maximum velocity of 9.65+/-2.49 pmol/min/mg protein (n=15). Uptake of rT3 was stereospecific, but not specific for rT3, as unlabelled L stereoisomers of thyroid hormone analogues were more effective as inhibitors of 125I-rT3 uptake than rT3. Unlabelled T3 and thyroxine (T4) (10 microM) reduced cellular uptake of 125I-rT3 by around 82 and 74 per cent, respectively. The calculated inhibition constants Ki were 1.23+/-0.29 microM (n=4) and 0.66+/-0.19 microM (n=4) for T3 and T4, respectively. Similarly, rT3 reduced cellular uptake of 125I-T3 and 125I-T4 by 34 and 23 per cent, respectively. The calculated inhibition constants Ki were 1.75+/-0.55 microM (n=8) and 1.08+/-0.36 microM (n=8) for the inhibition of 125I-T3 and 125I-T4 uptake, respectively. Reverse T3 inhibited efflux of 125I-T3 from the cells by around 20 per cent, but did not inhibit efflux of 125I-T4. These results suggest that uptake of rT3 in JAr cells may occur via a single, saturable membrane carrier, which also interacts with T3 and T4, while efflux of rT3 may occur by passive diffusion.