This review considers recent developments in our understanding of the properties of TRAb, particularly measurement of the antibodies and their sites of action and synthesis. Two new assay methods have allowed considerable improvements in the sensitivity, specificity, precision, and ease of measuring TRAb. In particular: 1) receptor assays based on inhibition of receptor-purified labeled TSH binding to detergent-solubilized TSH receptors and 2) bioassays based on stimulation of cAMP release from monolayer cultures of isolated thyroid cells. Detailed studies with the two assays indicate that TSH receptor antibodies nearly always act as TSH agonists in patients with a history of Graves' hyperthyroidism. Studies in areas of dietary iodine sufficiency suggest that measurement of the antibodies at various stages in the course of treating Graves' disease can be of value in predicting the outcome of therapy. However, in areas of iodine deficiency, difficulties in the ability of patients' thyroid tissue to recover from the effects of antithyroid drugs may prevent the receptor antibodies from causing a relapse of thyrotoxicosis. Consequently, the predictive value of receptor antibody measurements would be expected to be lower in these geographical areas. Although patients with a history of Graves' hyperthyroidism nearly always have TRAb which act as TSH agonists, about 20% of patients with frank hypothyroidism due to autoimmune destruction of the thyroid have TRAb which act as TSH antagonists (blocking antibodies). There is some evidence that these blocking antibodies can cause hypothyroidism particularly in the neonate. With regard to the site of synthesis of TRAb, there is now direct evidence that they are synthesized by thyroid lymphocytes, particularly the lymphocytes in close proximity to thyroid follicular cells. This is consistent with the well established effects of antithyroid treatment (drugs, radioiodine, or surgery) on TRAb levels in addition to their effects on thyroid hormone synthesis. Recent studies using affinity labeling with 125I-labeled TSH have enabled elucidation of the structure of the TSH receptor. TSH receptors in human, porcine, and guinea pig thyroid tissue have a two-chain structure in which the TSH binding site is formed on the outside surface of the cell membrane by a water-soluble A subunit (Mr approximately 50 K). The A subunit is linked by a disulfide bridge and weak noncovalent bonds to the amphiphilic B subunit (Mr approximately 30 K). This subunit, which penetrates the lipid bilayer, probably forms the site for interaction of the receptor with the regulatory subunits of adenylate cyclase.(ABSTRACT TRUNCATED AT 400 WORDS)

Autoantibodies to the thyrotropin receptor.

I. Introduction II. TSHR Ectodomain Structure A. Primary amino acid structure B. Subunit structure C. Disulfide bonds in the TSHR ectodomain D. Carbohydrate moieties in the TSHR ectodomain E. TSHR dimerization III. Recombinant TSHR Expression A. Approaches to TSHR expression B. TSHR expression in prokaryotes C. TSHR expression as cell-free translates or as synthetic peptides D. TSHR expression in eukaryotic insect cells E. TSHR expression in mammalian cells IV. TSH and Autoantibody Binding to the TSHR Ectodomain A. Holoreceptor vs. ectodomain B. Conformational nature of binding sites C. TSHR carbohydrate: part of the binding site? D. TSHR amino acid residues in TSH-binding site E. Autoantibody epitopes V. TSHR Autoantibody Assays A. Historical background B. In vivo and in vitro bioassays C. Indirect TBI assays D. Direct assays for TSHR autoantibodies E. Confusion on the terminology and types of TSHR autoantibodies VI. Monoclonal Autoantibodies to the TSHR Ectodomain A. Introduction B. Mouse TSHR mAb C. Hu...

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The Thyrotropin (TSH)-Releasing Hormone Receptor: Interaction with TSH and Autoantibodies

Publisher Summary This chapter discusses the action of thyrotropin on thyroid metabolism. Growth and function of the thyroid are mainly controlled by the level of thyrotropin (TSH) concentration in the plasma. The principal element of this control, the action of TSH on its target, the thyroid follicular cell, has been investigated intensively during recent years. TSH activates rapidly the steps of thyroidal iodine metabolism—that is, the formation of thyroid hormones from iodide and tyrosines and the secretion of these hormones, and induces, after a delay, the growth of thyroid tissue. Among the various systems available for the study of the interaction of a tropic hormone with its target cell, the thyroid and TSH present several advantages and disadvantages. The specialized metabolism of the follicular cell, the metabolism of iodine, is easily studied with the various radioisotopes of iodine. Thyroid tissue is very resistant to adverse conditions, which explains the great stability and the great variety of in vitro thyroid systems.

The action of thyrotropin on thyroid metabolism.

Ribonuclease inhibitor from human placenta. Purification and properties.

Thyroid Follicular Cell Carcinogenesis

In view of the similarity of TSH and LATS effects on the thyroid, we compared the effects of TSH (NIH-TSH-B4 and Thytropar) and purified LATS-IgG on lipolysis and the metabolism of U-14C-glucose by slices of guinea pig epididymal adipose tissue. Neither TSH nor LATS consistently affected glucose oxidation or incorporation of glucose carbon into total glycerides but both substances stimulated lipolysis and the incorporation of glucose carbon into glyceride-glycerol. Lipolysis was enhanced by TSH in concentrations of 0.010 mU/ml or more and this was inhibited by the addition of antiserum to TSH but not by that to human IgG. LATS-IgG in concentrations of 0.166 mg/ml (an amount per flask that had no significant effect in the mouse bioassay) enhanced lipolysis; a peak effect was seen with 0.66 mg/ml. This stimulation of lipolysis was inhibited by antiserum to human IgG, but not by anti-TSH, and also was inhibited by preincubation of the LATSIgG with 10 gEq of thyroid microsomes, whereas liver microsomes had no...

Comparison of the effects of thyrotropin and the long-acting thyroid stimulator on guinea pig adipose tissue.

In an attempt to purify the thyroid receptor for TSH and to study interaction with the thyroid-stimulating antibody (TSAb) of Graves' disease, we used both bovine and human thyroid glands. With either tissue, the 10,000 × g pellet of homogenized material was solubilized with 0.5% Triton N-101; excess Triton was removed by Amberlite XAD-2, and purification was effected by TSH affinity chromatography, followed by gel filtration on Sepharose 6B. Greatest purification was achieved with bovine tissue; this receptor preparation was 183-fold concentrated over the starting material, but contained only 6% of the original TSH-binding activity, due in part to spontaneous loss over the 4 days required for processing. On polyacrylamide gel electrophoresis, there were at least three protein bands, one of which was probably a subunit of thyroglobulin. Purified immunoglobulin G with thyroid-stimulating antibody activity inhibited the binding of TSH at all stages of purification of the receptor.

Solubilization, purification, and partial characterization of thyrotropin receptor from bovine and human thyroid glands.

The in vitro incorporation of [3H]uridine into RNA and [3H]leucine into protein in slices of porcine thyroid was studied. Thyrotropin (10-500 mU/ml of medium), when added with [3H]uridine, inhibited incorporation into RNA, but as little as 10 mU of thyrotropin per ml stimulated incorporation of [3H]orotic acid into RNA. Uridine kinase (EC 2.7.1.48) was found to be inhibited in slices incubated with thyrotropin whereas UMP 5′ nucleotidase (EC 2.1.3.5) was not. Preincubation of slices with thyrotropin (5-50 mU/ml) led to enhanced incorporation of subsequently added [3H]uridine and [3H]leucine. When slices were preincubated with long-acting thyroid stimulator-IgG (2.5 or 5 mg per ml of medium) incorporation of [3H]uridine and [3H]leucine was similarly enhanced, with the smaller concentration being more effective. Without preincubation these stimulatory effects were mimicked by 1 mM dibutyryl 3′,5′-AMP and, to a lesser extent, 1 mM 3′,5′-AMP. AMP and ATP also stimulated [3H]uridine incorporation in this system but only after more prolonged periods of incubation than were required for the other nucleotides. RNA polymerase (EC 2.7.7.6) activity measured in isolated thyroid nuclei had two components, one Mg2+-stimulated and the other requ ring Mn2+ and high salt content [0.4 M (NH4)2SO4]. These activities, and particularly the former, were enhanced if thyroid slices were incubated with thyrotropin (5-100 mU/ml of medium), 2.5 mg or 5.0 mg of long-acting thyroid stimulator-IgG per ml, or 1 mM dibutyryl 3′,5′-AMP, before isolatior of the nuclei and measurement of enzyme activities; 1 mM AMP, ADP, or 2′,3′-GMP had no influence. Added directly to the nuclei, thyrotropin, long-acting thyroid stimulator-IgG, and dibutyryl 3′,5′-AMP had no effect on RNA polymerase activities. These data are seen as affording evidence for mediation by 3′,5′-AMP of effects of thyrotropin and long-acting thyroid stimulator on thyroid RNA and protein synthesis, at least in part through an indirect stimulation of nuclear RNA polymerase activities.

Stimulation by thyrotropin, long-acting thyroid stimulator, and dibutyryl 3',5'-adenosine monophosphate of protein and ribonucleic acid synthesis and ribonucleic acid polymerase activities in porcine thyroid in vitro

Thyroid hydrolysis of cyclic AMP as influenced by thyroid gland activity

The long-acting thyroid stimulator (LATS) is now accepted as a feature of Graves' disease, and a great deal is known about its biochemical characteristics and its mode of action on the thyroid. Briefly, it is an IgG that directly influences thyroid gland function; its action involves stimulation of adenyl cyclase and enhanced production of 3′, 5′ cyclic adenosine monophosphate in the thyroid. It has an effect on at least one other extrathyroid tissue, adipose tissue, as shown by increased lipolysis in guinea pig epididymal fat that is incubated with LATS. While these facts are all well established, little is known about factors controlling the production of LATS, and its precise role in Graves' disease is much debated. The relationship of LATS to features of Graves' disease, apart from hyperthyroidism, is probably quite indirect; indeed, there may be in the syndrome one or more other immunoglobulins that have more significance than LATS has for pretibial myxedema (dermopathy) and ophthalmopathy. Although LATS is said to occur in a majority of patients with hyperthyroidism of Graves' disease when concentrates of IgG are tested, even tenfold concentration has failed to show 100% positive results. This fact has raised doubts about the colse pathogenetic relationship of LATS to hyperthyroidism. Moreover, a few patients have been reported to have LATS in blood but to have suppressible (with thyroid hormone) thyroid function. Therefore, LATS as the sole cause of hyperthyroidism is a theory that is now difficult to uphold. The existence of a permissive factor that facilitates its action on the thyroid is one postulate that may reconcile the apparent discrepancies.

J. M. Mckenzie

Papers

Comparison of the effects of thyrotropin and the long-acting thyroid stimulator on guinea pig adipose tissue.

Solubilization, purification, and partial characterization of thyrotropin receptor from bovine and human thyroid glands.

Stimulation by thyrotropin, long-acting thyroid stimulator, and dibutyryl 3',5'-adenosine monophosphate of protein and ribonucleic acid synthesis and ribonucleic acid polymerase activities in porcine thyroid in vitro

Thyroid hydrolysis of cyclic AMP as influenced by thyroid gland activity

Does LATS cause hyperthyroidism in Graves' disease? (A review biased toward the affirmative).