Blondeau J P, Osty J, Francon J
Unité de Recherche sur la Glande Thyroide et la Régulation Hormonale, Unité 96, l'Institut National de la Santé et de la Recherche Médicale, Le Kremlin-Bicêtre, France.
J Biol Chem. 1988 Feb 25;263(6):2685-92.
Transport of 3,5-[3'-125I]triiodo-L-thyronine ([125I]T3) was studied in isolated rat liver hepatocytes. T3 transport was temperature-sensitive, the initial velocity of uptake, at low substrate concentration, was 60 times higher at 25 degrees C than at 0 degrees C. The activation energy of cellular uptake (26 kcal/mol) was different from that of binding to cytosolic proteins (6 kcal/mol), indicating that the latter was not the rate-limiting step. Uptake obeyed simple Michaelis-Menten kinetics, with an apparent Km of 0.34 microM and a Vmax of 0.15 fmol/min/cell at 25 degrees C. No simple diffusion occurred. Unlabeled T3, L-thyroxine (T4), 3,5,3'-triiodo-D-thyronine, and triiodothyroacetic acid inhibited T3 uptake with Kl values of 0.32, 1.4, 4.1, and 5.4 microM, respectively, indicating specificity of uptake which was different from specificity of intracellular binding sites. [125I]T4 was also taken up by a saturable process (Km = 0.65 microM and Vmax = 0.16 fmol/min/cell at 25 degrees C). T3 was a better competitor than T4 for the uptake of [125I]T4, indicating that both hormones were taken up by the same carrier system. Metabolic inhibitors had either no effect on T3 uptake or inhibitory effects unrelated to cellular ATP depletion. Uptake was not affected by modification of the membrane potential or the sodium ion gradient. T3 and T4 uptake was pH-dependent. It is suggested that the un-ionized 4'-OH form of the hormones was preferentially taken up. Inhibition of uptake by various compounds was compared to inhibition of thyroid hormone binding to transthyretin, nuclear receptor, and cytosolic-binding proteins. We conclude that, in freshly isolated hepatocytes, thyroid hormones are taken up by a saturable, stereospecific, Na+-independent carrier system.
在分离的大鼠肝脏肝细胞中研究了3,5-[3'-125I]三碘-L-甲状腺原氨酸（[125I]T3）的转运。T3转运对温度敏感，在低底物浓度下，摄取的初始速度在25℃时比在0℃时高60倍。细胞摄取的活化能（26千卡/摩尔）与与胞质蛋白结合的活化能（6千卡/摩尔）不同，这表明后者不是限速步骤。摄取符合简单的米氏动力学，在25℃时表观Km为0.34微摩尔，Vmax为0.15飞摩尔/分钟/细胞。未发生简单扩散。未标记的T3、L-甲状腺素（T4）、3,5,3'-三碘-D-甲状腺原氨酸和三碘甲状腺乙酸分别以0.32、1.4、4.1和5.4微摩尔的Kl值抑制T3摄取，表明摄取具有特异性，这与细胞内结合位点的特异性不同。[125I]T4也通过可饱和过程摄取（在25℃时Km = 0.65微摩尔，Vmax = 0.16飞摩尔/分钟/细胞）。对于[125I]T4的摄取，T3比T4是更好的竞争者，这表明两种激素通过相同的载体系统摄取。代谢抑制剂对T3摄取要么没有影响，要么具有与细胞ATP消耗无关的抑制作用。摄取不受膜电位或钠离子梯度改变的影响。T3和T4摄取依赖于pH。提示激素的未电离4'-OH形式优先被摄取。将各种化合物对摄取的抑制作用与甲状腺激素与转甲状腺素蛋白、核受体和胞质结合蛋白结合的抑制作用进行了比较。我们得出结论，在新鲜分离的肝细胞中，甲状腺激素通过可饱和、立体特异性、不依赖钠离子的载体系统摄取。
