Thyroid-stimulating Activity Research Articles

The effect of radioiodine and antithyroid drugs on serum long acting thyroid stimulator protector (IATS-P). A three year prospective study.

Over a three year period we have studied the effect of either a one year course of Carbimazole or a single dose of radioiodine in a group of 46 patients with Graves' disease. Initially, in untreated patients LATS-P was present in 39 (85%) but at the end of the study was only detectable in 19 (41%). The clinical outcome in 29 patients initially treated with carbimazole correlated well with changes in serum LATS-P which persisted in 18. Thirteen of these ultimately required radioiodine or sub-total thyroidectomy. With radioiodine two patterns of response were seen, in some LATS-P levels declined, whereas in others transient increases were seen usually during the first year but subsequently fell. There was no difference in clinical response between the two groups. Overall, the study indicates that serum LATS-P is related to the clinical course of Graves' disease but there remains a minority of patients in whom the activity cannot be detected.

Serum long acting thyroid stimulator (LATS) and LATS-protector (LATS-P) in Graves' disease associated with localized myxedema.

When localized myxedema occurs in Graves' disease, there is often very high serum long acting thyroid stimulator (LATS) activity. However, this association is not invariable and no pathogenetic role for this IgG associated activity is known. Serum LATS protector (LATS-P) is a closely related IgG activity which is present in the majority of cases of untreated Graves' disease. It usually coexists in LATS positive sera in a substantially greater concentration. Its association with localized myxedema has not been studied, nor have serial studies been performed on this activity during the clinical course of the disease. Fourteen patients (13 females, 1 male) with localized myxedema and a history of Graves' disease were examined. In 13 serum LATS was detectable with a wide range of activity from 2.4 to 1,000 units/ml. Serum LATS-P was detected in all including the LATS negative patient with a range of activity from 46 to 4,068 units/ml. Serial studies for at least 2 years were conducted in 8 patients. In two there was no change in either skin lesions or in serum LATS and LATS-P. In 6 the skin lesions partially or completely resolved. In 5 this was associated with statistically significant falls in serum LATS and LATS-P but in one no significant change occurred. The study demonstrated the high prevalence of LATS and LATS-P in localized myxedema. In the sole LATS negative patient there was a high concentration of LATS-P. The role of these activities in the pathogenesis of the disease remains unknown but in serial studies falls in activity were usually associated with clinical improvement.

Further studies on the response of a cytochemical bioassay to thyroid stimulators, using reference preparations of thyrotropin and long acting thyroid stimulator

The fundamental response of this assay is shown to be a shortening of a latency period prior to a peak of staining after tissue sections have been exposed to either TSH or LATS. The magnitude of the peak for a given tissue is both stimulator and dose independent. Simultaneous exposure to a combined dose of the two stimulator resulted in a further shortening of the latency period, and no evidence of two peaks was observed in the presence of both stimulators. Dose-response curves after exposure to reference preparations of the two stimulators for the same time period (180 s.) were parallel: this indicates that given incremental doses of either stimulator result in indistinguishable increases in the fundamental response of this assay.

Studies on the suppressor T cell function in patients with autoimmune thyroid diseases

Suppressor T cell function induced by concanavalin A (con A) was evaluated in patients with Graves' disease and Hashimoto's thyroiditis. Patients with Graves' disease were divided into the following two groups: (1) untreated, and (2) euthyroid during antithyroid drug (methylmercaptoimidazole) therapy. T cells (2 X 10(5)), activated by con A for 48 hours, were added to preincubated responder cells (2 X 10(5)) and re-incubated for 7 days in the presence of pokeweed mitogen (PWM). IgG produced in the culture medium was measured by radioimmunoassay and then % suppressions (IgG) were calculated. Thyroid stimulating activity (TSA) in serum was measured by McKenzie's method by means of normal human thyroid slices, and % suppressions (c-AMP) were calculated. IgG produced in lymphocyte culture medium was suppressed by added con A-activated cells in untreated and euthyroid groups of Graves' disease, Hashimoto's thyroiditis and normal controls. The value of % suppression (IgG) was reduced in each group of Graves' disease compared to normal controls. No significant relation was observed between TSA in serum and % suppression (IgG), but three cases with high serum TSA showed low % suppressions (IgG). In 12 cases of Graves' disease, % suppression (IgG) had a positive relation with % suppression (c-AMP) in same medium. The amount of c-AMP produced in thyroid slices incubated in medium, in which responder cells (8 X 10(5)), was elevated in all 7 untreated cases of Graves' disease, while not elevated in 7 euthyroid cases. The value of % suppression (c-AMP) in euthyroid cases with Graves' disease was significantly higher than that in untreated cases. The value of % suppression (IgG) was reduced and had a significant negative relation with logarithm of serum antimicrosomal antibody titer in patients with Hashimoto's thyroiditis. These results indicate that low activity of suppressor T cell had a role on antibody production, including thyroid stimulating antibody, and pathogenesis of autoimmune thyroid diseases.

TSH receptor binding and thyroid stimulation by sera from patients with Graves' disease.

An investigation of the ability of TSH receptor antibodies to bind to the TSH receptor and stimulate thyroid function is described. Binding studies were carried out using 125I-labelled TSH and detergent solubilised porcine TSH receptors, and the parameter of thyroid stimulation employed was 125I-organification in isolated porcine thyroid cells. Different Graves' sera were found to show different dose-response relationships and three types of antibody activity were evident in the six samples studied. In two samples receptor binding and thyroid stimulating activities were approximately parallel. In two receptor binding was detectable at lower doses than stimulation while in the last two stimulation was detectable at lower doses than binding. It is proposed that these characteristics are due to different populations of receptor antibodies which exhibit different degrees of TSH agonism. Furthermore one of the reasons for the relatively poor correlation between TSH receptor binding and thyroid stimulation observed in series of individual Graves' sera may be the presence of different populations of antibodies showing different dose-response relationships with regard to binding and stimulation.

Consistency and variability in the character of thyrotrophin receptor antibodies in Graves' disease.

At diagnosis there was no correlation between the uptake of pertechnetate by the thyroid and thyrotrophin receptor antibodies (TRAb) measured as TSH binding inhibitory immunoglobulins in a series of 27 patients with Graves' disease. TRAb were detectable initially in 19 patients, in 11 of these there was a significant positive correlation (P less than 0.05) between serial measurements of pertechnetate uptake and TRAb made during 2 years following diagnosis. In five patients pertechnetate uptake fell with time whilst TRAb levels were maintained or fluctuated. In the remaining three of the 19 patients both measurements were low and did not change during treatment. We conclude that TRAb in any individual patient are a mixture of immunoglobulins of variable effectiveness as thyroid stimulators. In a majority of patients the composition of this mixture remains constant during the course of the illness and the clinical state reflects the levels of TRAb in the blood. In a minority, however, the character of these antibodies may alter with time or there is a change in the responsiveness of the thyroid gland. The general lack of correlation between measurements of thyroid stimulating activity and TSH binding inhibitory immunoglobulins in groups of patients is due to differences between patients in the composition of TRAb.

Heterogeneity of the Graves' immunoglobulins directed toward the thyrotropin receptor-adenylate cyclase system.

The TSH-displacing and the thyroid-stimulating activities of Graves' immunoglobulins G (GIgG) have been accounted for by either homogeneous TSH receptor antibodies or heterogeneous antibodies directed toward the TSH receptor-adenylate cyclase system. To clarify this matter, the study of the interactions of GIgG preparations from 14 untreated Graves' patients with human thyroid membranes was undertaken. Dose-response curves of GIgG, diluted in IgG from normal subjects, were carried out in the [125I]TSH radioreceptor assay and the adenylate cyclase assay in the presence or absence of TSH. In the radioreceptor assay, GIgG were constantly negative in 3 cases (21%) and positive, depending of the dose, in 11 cases. In the adenylate cyclase assay, the dose-activity profiles in the absence of TSH were bell-shaped curves in 3 cases and sigmoid curves in 9 cases; in 2 cases (14%), GIgG preparations were devoid of any effect. Binding-isotherms and dose-activity profiles did not appear to share simple relationships. In the presence of TSH, GIgG preparations elicited a decrease in 6 cases, an increase in 3 cases, and no effect in 5 cases (36%) in the adenylate cyclase activity. The data obtained by radioreceptor assay and adenylate cyclase assay in the presence or absence of TSH were found statistically correlated (P less than 0.05 to P less than 0.001) but not linearly related, the points being scattered on specific parts of the diagrams. These observations could not be accounted for by TSH receptor antibodies in GIgG being an entity of constant properties, albeit varying in titer among patients. Rather, GIgG effects fit well the patterns of action of a heterogeneous ligand, as shown by computing a theoretical model for ligand heterogeneity with respect to binding equilibrium constant and intrinsic biological activity. Accordingly, GIgG activity in the TSH receptor-adenylate cyclase system could be attributed to heterogeneous antibodies varying with regard to binding constant and acting as stimulating or blocking antibodies of the adenylate cyclase.

Serum long acting thyroid stimulator protector in pregnancy complicated by Graves' disease.

Activities of serum long acting thyroid stimulator protector were measured in a series of nine pregnancies in eight mothers who had Graves' disease, one of whom had been successfully treated by surgery. In all but two instances the activities tended to decline as pregnancy progressed. After delivery activities rose in three out of five patients in whom these had disappeared in pregnancy and, as this occurred, the patients relapsed. In the two patients whose activities did not decline thyrotoxicosis persisted throughout pregnancy and after delivery. None of the nine babies in this study suffered from neonatal thyrotoxicosis because maternal activities of the thyroid stimulator protector, though high enough to induce Graves' disease in adults, were not above the threshold for the induction of thyroid overactivity in neonates.

Lack of refractoriness to stimulation with long acting thyroid stimulator of thyroid hormone synthesis and thyroid hormone secretion in mice in vivo.

The present study was undertaken to examine whether long acting thyroid stimulator (LATS) induces refractoriness of thyroid hormone synthesis and thyroid hormone secretion to the stimulator in vivo. Male DDY mice fed with a low iodine diet and given 5 micrograms/ml of triiodothyronine (T3) in drinking water and libitum for 4 days were injected with 0.025 ml of LATS positive serum (1000%/0.25 ml in rhe McKenzie bioassay) ip every 24 h for 9 days. Groups of 5 mice were sacrificed before and 1, 3, 5, 7 and 9 days after the first injection of LATS for the determinations of serum thyroxine (T4) concentrations, the 1 h thyroid 131I uptake and thyroid weight. Control mice were injected with LATS negative pooled normal sera. Serum T4 concentrations elevated significantly 24 h after the 3rd injection of LATS and remained elevated until the end of the experiment. One hour thyroid 131I uptake elevated about 3-fold 24 h after the first injection of LATS. It further increased 24 h after the 3rd injection of LATS to 10-fold of the value for control animals and stayed elevated at this same level for the remainder of the study. These results indicate that stimulating effects of LATS on thyroid hormone synthesis assessed by thyroid 131I uptake and thyroid hormone secretion assessed by serum T4 concentrations were not diminished by the prior administration of the stimulator. These findings suggest that LATS does not induce refractoriness of either thyroid hormone synthesis or thy roid hormone secretion to the stimulator in mide in vivo.

Pathogenesis of hyperthyroidism.

Graves' disease is the most frequent cause of hyperthyroidism in the United States. It occurs predominantly in a genetically predisposed population. Autoimmune abnormalities constitute a major component of the disease. Immunoglobulins of patients with Graves' disease manifest a number of thyroid-targeted activities, including a thyroid-stimulating activity that is related to interaction of immunoglobulin G (IgG) with the TSH receptor. That hyperthyroidism in Graves' disease is caused by thyroid-stimulating IgGs is an attractive hypothesis but the evidence in favor of the hypothesis is still insufficient. A better alternative hypothesis is not available, however.

Graves' IgG stimulation of continuously cultured rat thyroid cells: a sensitive and potentially useful clinical assay.

A continuously cultured line of normal rat thyroid (FRTL) cells can be stimulated by immunoglobulin preparations from patients with Graves' disease as measured by increases in intracellular cAMP levels. Responsiveness is concentration-dependent but is delayed in time relative to thyrotropin. Additionally, the cells respond to Graves' immunoglobulins which have no long-acting thyroid stimulator (LATS) activity and are negative when adenylate cyclase stimulation in human thyroid membrane preparations is assayed. No correlation exists between the stimulation activity and the ability of a Graves' immunoglobulin preparation to inhibit thyrotropin binding; cells are responsive even in the presence of such inhibitor activity.

Discrepancy between thyroid-stimulating and thyrotropin-binding inhibitory activities of Graves' immunoglobulin Gs assessed in the mouse.

TSH receptor binding-inhibiting (TBI) and thyroid-stimulating (TSI) activities of Graves' immunoglobulin Gs (IgGs) were measured using the murine TSH receptor (mTBI) and the McKenzie LATS bioassay. In contrast to the expectation that only the IgGs positive for LATS showed mTBI activity, mTBI activity was closely correlated with TBI activity measured at the human TSH receptor regardless of LATS potency (r = 0.79; P less than 0.01), indicating a lack of species specificity of IgG in this particular aspect. In 7 to 15 IgGs positive for both mTBI and LATS, there was a significant correlation between the 2 parameters (r = -0.89; P less than 0.01). However, 4 IgGs among the 15 were positive for mTBI but negative for LATS even when tested at 3- to 7-fold concentrations of IgG.

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

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 X 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.

Effect of methyldopa on thyroid stimulating immunoglobulins producted by peripheral lymphocytes from patients with juvenile Graves'disease

In an attempt to test effects of methyldopa (inhibitor for suppressor T-cell) on the production of thyroid stimulating immunoglobulins (TSI), peripheral lymphocytes were preincubated with or without the drug followed by cultures with human thyroid plasma membrane for 7 days. Thyroid stimulating activities (TSA) in the supernatants of cultures were measured as cAMP generated during incubation of ATP with bovine thyroid membrane, and then were expressed as a percentage of control level. When the cells were preincubated in the presence of methyldopa, IgG synthesis from both patients and normal subjects generally increased. The pretreatment of methyldopa enhanced mean TSA from a level of 120±48% to 192±57% in the patients (P 0.01), while the effect was not observed in normal subjects. All these activities producted by lymphocytes in vitro were well correlated with those of IgG purified from the sera through DEAE-cellulose column chromatography. These results demonstrated that methyldopa is of use for in vitro production of TSI. The findings suggest that the inhibitory effect of methyldopa on T-cell proliferation may enhance the TSI production.

Purification of a stimulator of microtubule assembly from bovine thyroid.

A protein from bovine thyroid which stimulated the assembly of microtubules from phosphocellulose-purified tubulin was isolated. The purification scheme involved fractionation of bovine thyroid homogenates successively on phosphocellulose, DEAE-cellulose, and Sephadex G-200. Activity was determined as ability to stimulate phosphocellulose-purified tubulin assembly measured turbidimetrically. Purification of stimulatory activity resulted in the selective enrichment to near homogeneity of a 40,000-45,000-dalton protein. The molecular weight was estimated by Sephadex G-200 gel filtration and polyacrylamide gel electrophoresis. This protein stimulated both the rate and the extent of assembly of thyroid and brain tubulin. In the turbidity assay which contained 4 M glycerol, the threshold for assembly of brain tubulin (at 1.7 mg/ml) was 0.01 mg/ml for the thyroid stimulator. Brain and thyroid microtubules assembled with thyroid stimulator appeared normal in diameter and shape by electron microscopy. Thyroid stimulator activity was destroyed by trypsinization or boiling. Activity was maximal at 1 mM GTP and 4 mM MgCl2 and was blocked by colchicine. No stimulatory protein of similar molecular weight could be isolated from bovine brain by similar techniques.

Absence of thyroid-stimulating antibody and long acting thyroid stimulator in relatives of Graves' disease patients.

None of the 76 euthyroid relatives of patients with Graves' disease had detectable LATS in their serum nor was thyroid-stimulating antibody (TSAb), assessed by an increment of cAMP in human thyroid slices, detected in any of the 60 sera tested. Seven of 41 were slightly positive for TSH binding-inhibiting immunoglobulin (TBII). Nineteen of 73 sera were positive (greater than 1:1600) for antibody to thyroid antimicrosomal antigen, of which 4 were also positive (greater than 1:400) for antibody to thyroglobulin; 3 of the 19 had a slightly elevated basal TSH which rose excessively after TRH. Thus, although these euthyroid relatives had evidence of thyroid immunological defects, a thyroid-stimulating antibody was not found.

Serum Long Acting Thyroid Stimulator (Lats) and Lats-Protector (LATS-P) in Graves' Disease Complicated by Localized Myxoedema

When localized myxedema occurs in Graves’ disease, there is often very high serum long acting thyroid stimulator (LATS) activity. However, this association is not invariable and no pathogenetic role for this IgG associated activity is known. Serum LATS protector (LATS-P) is a closely related IgG activity which is present in the majority of cases of untreated Graves’ disease. It usually coexists in LATS positive sera in a substantially greater concentration. Its association with localized myxedema has not been studied, nor have serial studies been performed on this activity during the clinical course of the disease. Fourteen patients (13 females, 1 male) with localized myxedema and a history of Graves’ disease were examined. In 13 serum LATS was detectable with a wide range of activity from 2.4 to 1,000 units/ml. Serum LATS-P was detected in all including the LATS negative patient with a range of activity from 46 to 4,068 units/ml. Serial studies for at least 2 years were conducted in 8 patients. In two there was no change in either skin lesions or in serum LATS and LATS-P. In 6 the skin lesions partially or completely resolved. In 5 this was associated with statistically significant falls in serum LATS and LATS-P but in one no significant change occurred. The study demonstrated the high prevalence of LATS and LATS-P in localized myxedema. I n the sole LATS negative patient there was a high concentration of LATS-P. The role of these activities in the pathogenesis of the disease remains unknown but in serial studies falls in activity were usually associated with clinical improvement.

Thyroid function in gestational trophoblastic neoplasia: Evidence that the thyrotropic activity of chorionic gonadotropin mediates the thyrotoxicosis of choriocarcinoma

An investigation was made of thyroid function in 20 patients with gestational trophoblastic neoplasia. Two patients were judged to be overtly thyrotoxic on the basis of the symptoms and physical findings; both patients had widely metastatic choriocarcinoma, markedly increased serum T4 levels (21.4 and 27.7 μg/100 ml), and extremely high levels of serum human chorionic gonadotropin (hCG) (3,220 and 6,720 IU/ml) relative to those of normal gestation (<100 IU/ml). Three other patients had moderately increased serum T4 levels (13 to 17.1 μg/100 ml), moderately increased serum hCG levels (110 to 310 IU/ml), and findings on clinical examination which suggested euthyroidism. Using the mouse thyroid bioassay, we found that the biologic characteristics of the thyroid-stimulating factor were those of purified hCG, and that the levels of thyroid-stimulating activity in both serum and urine correlated closely with the levels of hCG. These results provide evidence that the thyroid-stimulating activity intrinsic to the hCG molecule plays the central pathophysiologic role in choriocarcinoma-associated thyrotoxicosis.

Human thyroid stimulating activity and clinical state in antithyroid treatment of juvenile Graves' disease.

Abstract. Human thyroid stimulating activity in 37 patients with juvenile hyperthyroidism was studied during the course of antithyroid drug treatment. Observation periods varied from 6 months to 6 years (average 3 years). The existence of human thyroid stimulator (HTS) in their sera was determined by triiodothyronine (T3) release from resected thyroid adenomas. A measurement of T3 above the control value of 140% was judged to be positive. Before treatment, 25 of the 28 patients with Graves' disease had HTS in their sera, one of the 7 patients with Hashimoto's thyroiditis had HTS, and none of the 19 normal children had HTS in their sera. During the therapy, 7 of 18 Graves' patients had HTS. After cessation of the therapy, 9 of 22 patients remained HTS-positive, and 8 of the 9 patients with positive HTS relapsed within 6 months, whereas in 13 HTS-negative patients, 3 escaped and 10 remained in remission. There was no significant correlation between human thyroid stimulating activity (HTSA) and the following: serum T3 concentration, 131I-thyroidal uptake, TSH responsiveness to TRH and antithyroid antibody titre. These results indicate that (1) almost all patients with Graves' disease have HTS when they are in a state of hyperthyroidism, (2) HTS tends to disappear with the clinicial remission, but its persistence seems to predict relapse in the future, and (3) routine laboratory tests cannot serve as alternative to HTS.

Receptor-binding activity of highly purified bovine luteinizing hormone and thyrotropin, and their subunits.

Highly purified preparations of bovine TSH (bTSH) and LH (bLH) and their subunits have been obtained by affinity chromatography using immobilized antibodies directed against counterpart subunits. The purified preparations were assessed for biological activity in radioligand-receptor assays for TSH and LH. After affinity purification against bLH beta, a TSH preparation whose initial potency in the LH assay had been 0.15% that of LH, failed to compete with [125I]LH in amounts up to 100 microgram. Thus, it appears that bTSH does not bind to LH receptors in the rat testis and that interaction of less purified TSH with gonadotropin receptors is attributable to LH contamination. In contrast, LH, whose initial potency in the TSH receptor assay was 0.6% that of TSH, retained a potency of 0.004% of TSH (equivalent to 3.6 mU/mg) after immunoadsorption by anti-bTSH beta. The retention of TSH receptor-binding activity by affinity-purified LH indicates that the LH molecule (like hCG) has a low intrinsic thyroid-stimulating activity. Affinity-purified LH subunits have little or no demonstrable affinity for the LH receptor in vitro. Affinity-purified TSH subunits and affinity-purified LH, however, exhibit very weak receptor-binding activity in the TSH radioligand receptor assay. An evaluation of the capacity of the immunoadsorbents to remove TSH from artificial mixtures suggests that the residual binding does not result entirely from contamination, and therefore, that alpha-subunits as well as LH have some intrinsic TSH-binding activity.