The Experts below are selected from a list of 3339 Experts worldwide ranked by ideXlab platform

George J Kahaly - One of the best experts on this subject based on the ideXlab platform.

  • comparison of a novel homogeneous cyclic amp assay and a luciferase assay for measuring stimulating Thyrotropin Receptor autoantibodies
    European thyroid journal, 2020
    Co-Authors: Tanja Diana, Paul D Olivo, Yiehwa Chang, C Wuster, Michael Kanitz, George J Kahaly
    Abstract:

    Objective: Stimulating Thyrotropin-Receptor antibodies (TSAb) cause Graves' disease (GD). We tested a novel homogeneous fluorescent 3',5' cyclic adenine monophosphate (cAMP) assay for the detection of TSAb in a bioassay. Methods: Chinese hamster ovary (CHO) cell lines expressing either a chimeric (MC4) or wild-type (WT) TSH-R were incubated with the adenyl cyclase activator forskolin, a human TSAb monoclonal antibody (M22), and with sera from GD patients. Intracellular cAMP levels were measured using a Bridge-It® cAMP assay, and the results were compared with a luciferase-based bioassay. Results: Both cell lines were stimulated with forskolin concentrations (0.006-200 µM) in a dose-dependent manner. The linear range in the MC4 and WT cells was 0.8-25 and 3.1-50 µM, respectively. Levels of cAMP and luciferase in forskolin-treated MC4 and WT cells were positively correlated (r = 0.91 and 0.84, both p < 0.001). The 50% maximum stimulatory concentration of forskolin was more than 16-fold higher for the CHO-WT cells than the CHO-MC4 cells in the cAMP assay and 4-fold higher in the luciferase assay. Incubation of both cell lines with M22 (0.006-50 ng/mL) resulted in a dose-dependent increase in cAMP levels with linear ranges for the MC4 and WT cells of 0.8-12.5 and 0.2-3.125 ng/mL, respectively. Comparison of cAMP and luciferase levels in M22-treated MC4 and WT cells also showed a positive correlation (r = 0.88, p < 0.001 and 0.75, p = 0.002). A positive correlation was also noted when using patient samples (r = 0.96, p < 0.001) that were all TSH-R-Ab binding assay positive. Conclusion: The novel, rapid, simple-to-perform cAMP assay provides TSAb-mediated stimulatory results comparable to a luciferase-based bioassay.

  • comparison of a bridge immunoassay with two bioassays for Thyrotropin Receptor antibody detection and differentiation
    Hormone and Metabolic Research, 2019
    Co-Authors: Stephanie Allelein, Tanja Diana, Michael Kanitz, Margret Ehlers, Derik Hermsen, Matthias Schott, George J Kahaly
    Abstract:

    A rapid and fully automated chemiluminescent immunoassay for the detection of Thyrotropin Receptor autoantibodies (TSHR-Ab) based on a bridge technology was compared with two bioassays that measure either stimulating (TSAb) or blocking (TBAb) antibodies for the detection and differentiation of TSHR-Ab. A total of 229 patients with various thyroid disorders [151 with Graves’ disease (GD), 35 with Hashimoto’s thyroiditis (HT), 32 with nodular goiter, and 11 with thyroid cancer] were included. The bridge immunoassay was performed according to the manufacturer’s instructions (cut-off>0.55 IU/l). TSAb and TBAb were measured with reporter bioassays. Blocking activity was defined as percent inhibition of luciferase expression relative to induction with bovine TSH alone (cut-off>34% inhibition). TSAb was reported as percentage of specimen-to-reference ratio (> 140 SRR%). The 3 TSHR-Ab assays were negative in all patients with benign euthyroid nodular goiter and differentiated thyroid cancer. In contrast, in all patients with GD, irrespective of the disease duration, TSHR-Ab positivity was present in 127 of 151 (84%) and 140 (93%) for the bridge assay and TSAb bioassay, respectively (p

  • standardization of a bioassay for Thyrotropin Receptor stimulating autoantibodies
    Thyroid, 2015
    Co-Authors: Tanja Diana, Paul D Olivo, Michael Kanitz, Markus Lehmann, George J Kahaly
    Abstract:

    Background: Cell-based bioassays for functional thyroid stimulating autoantibodies (TSAb) are sensitive diagnostic tools. However, there is no bioassay available that is standardized with international reference material. We aimed to promote the standardization of the test results among laboratories that perform TSAb bioassays and calibrate TSAb levels against the second international standard (IS) 08/204 from the National Institute for Biological Standards and Control (NIBSC). Methods: Serum TSAb activity was measured with a FDA-cleared bioassay that utilizes CHO cells expressing a chimeric Thyrotropin Receptor (TSHR) and a c-AMP response-element-dependent luciferase. The IS was applied for calibration. TSAb results were reported as percentage of specimen-to-reference ratio (SRR%) and converted into mIU/L. Results: The IS dose–response curve was obtained using concentrations from 0.3125 to 200 mIU/L. Mean TSAb SRR%±standard deviation (SD) values for the IS concentrations 0.3125, 0.625, 1.25, 2.5, 5, 10, ...

Gilbert Vassart - One of the best experts on this subject based on the ideXlab platform.

  • oncogenic mutations in the Thyrotropin Receptor of autonomously functioning thyroid nodules in the japanese population
    European Journal of Endocrinology, 2002
    Co-Authors: V Vanvooren, Jacques Emile Dumont, Jasmine Parma, Laurence Duprez, Gilbert Vassart, J Van Sande, S Uchino, Maria Jose Costa, Julie Vandekerckhove, S Noguchi
    Abstract:

    Objective: Constitutively activating mutations of the Thyrotropin Receptor (TSHR) have been found in the majority of autonomously functioning thyroid nodules (AFTNs) in European patients. The reported frequency of these mutations varies among reports but amounts to 50 ‐ 80%. To date, only one such mutation responsible for AFTNs has been identified in the Japanese population and the pathogenic role of such mutations in Japanese AFTNs has been questioned. In the present study, we evaluated the frequency of activating mutations in the TSHR and Gas in 10 Japanese AFTNs. Design: Genomic DNA was extracted from fresh frozen tissue. The TSHR and the almost entire sequence of the gene coding for the a subunit of Gs have been amplified and sequenced. Results: In sequence analysis, four mutations in the TSHR (T632A, I486M, M453T and L512R) were found. To complete our analysis, we searched mutations in the gene coding for the a subunit of Gs, in the samples negative for TSHR mutations. In one case a mutation (R201H) affecting GTPase activity was found. Conclusions: If we focus on the solitary nodules, we obtain the same mutation proportion as in European patients (70%). The absence of TSHR and Gas mutations in a significant proportion of autonomous adenomas in multinodular goiters suggests that other causes may also play a role in the genesis of these lesions.

  • genetic immunization of outbred mice with Thyrotropin Receptor cdna provides a model of graves disease
    Journal of Clinical Investigation, 2000
    Co-Authors: Sabine Costagliola, Mariechristine Many, Jeanfrancois Denef, Joachim Pohlenz, Samuel Refetoff, Gilbert Vassart
    Abstract:

    We performed genetic immunization of outbred NMRI mice, using a cDNA encoding the human Thyrotropin Receptor (TSHr). All mice produced antibodies capable of recognizing the recombinant Receptor expressed at the surface of stably transfected Chinese hamster ovary (CHO) cells, and sera from most of the immunized mice blocked TSH-dependent stimulation of cAMP accumulation in cells expressing the TSHr. Five out of 29 female mice showed sign of hyperthyroidism including elevated total T4 and suppressed TSH levels. The serum of these mice contained thyroid-stimulating activity, as measured in a classic assay using CHO cells expressing recombinant TSHr. In contrast, only 1 male out of 30 had moderately elevated serum total T4 with undetectable TSH values. The hyperthyroid animals had goiters with extensive lymphocytic infiltration, characteristic of a Th2 immune response. In addition, these animals displayed ocular signs reminiscent of Graves' ophthalmopathy, including edema, deposit of amorphous material, and cellular infiltration of their extraocular muscles. Our results demonstrate that genetic immunization of outbred NMRI mice with the human TSHr provides the most convincing murine model of Graves' disease available to date.

  • familial gestational hyperthyroidism caused by a mutant Thyrotropin Receptor hypersensitive to human chorionic gonadotropin
    The New England Journal of Medicine, 1998
    Co-Authors: P Rodien, Jasmine Parma, Sabine Costagliola, Gilbert Vassart, Catherine Bremont, Marielaure Raffin Sanson, Jacqueline Van Sande, Jeanpierre Luton, Laurence Duprez
    Abstract:

    Some degree of stimulation of the thyroid gland by human chorionic gonadotropin is common during early pregnancy.1–3 When serum chorionic gonadotropin concentrations are abnormally high — for example, in women with molar pregnancies — overt hyperthyroidism may ensue. The pathophysiologic mechanism is believed to be promiscuous stimulation of the Thyrotropin Receptor by the excess chorionic gonadotropin.4,5 The explanation for this stimulation is the close structural relations between chorionic gonadotropin and Thyrotropin and between their Receptors.6 Hyperemesis gravidarum is characterized by excessive vomiting in early pregnancy, leading to the loss of 5 percent or more of body weight.4 It . . .

  • genetic immunization against the human Thyrotropin Receptor causes thyroiditis and allows production of monoclonal antibodies recognizing the native Receptor
    Journal of Immunology, 1998
    Co-Authors: Sabine Costagliola, Mariechristine Many, P Rodien, Marian Ludgate, Gilbert Vassart
    Abstract:

    The generation of Abs recognizing the native structure of the human Thyrotropin Receptor (hTSHR) has been difficult because there is currently no method allowing the purification of correctly folded Ag in the amounts required by classical immunization protocols. The majority of Abs made against the hTSHR react preferentially with denatured molecules. We report that a humoral response against the native hTSHR, compatible with mAb production, is elicited in mice by immunization with a DNA construct encoding the Receptor. BALB/c mice were inoculated in the anterior tibialis muscle with 100 microg of plasmid DNA harboring the hTSHR cDNA. Eleven weeks after the first injection, 10 mice of 14 showed by FACS analysis a strong IgG response against the hTSHR expressed at the surface of Chinese hamster ovary cells. A clear TSH-binding inhibiting Ig and Thyrotropin-blocking Ab activity (competition with TSH binding and TSH activity, respectively) was demonstrated in the majority of sera tested. One serum exhibited a clear stimulating activity. Despite the maintenance of normal circulating free T4 levels in all mice, these bioactivities persisted until 18 wk, in which mice were sacrificed, their thyroids were examined histologically, and spleens from two animals were used for mAb production. All mice displayed a severe lymphocytic infiltration of their thyroids, composed mostly of activated B cells. Three mAbs were produced against conformational epitopes of the hTSHR. We conclude that genetic immunization is an efficient method of generating Abs recognizing the native structure of the hTSHR and a new way of inducing thyroiditis in mice murine.

  • familial congenital hypothyroidism due to inactivating mutation of the Thyrotropin Receptor causing profound hypoplasia of the thyroid gland
    Journal of Clinical Investigation, 1997
    Co-Authors: Marc Abramowicz, Jasmine Parma, Laurence Duprez, Gilbert Vassart, Claudine Heinrichs
    Abstract:

    Thyroid gland agenesis is the most common cause of congenital hypothyroidism and is usually sporadic. We investigated a brother and sister from consanguineous parents, ascertained through systematic newborn screening, and initially diagnosed with thyroid agenesis. Careful cervical ultrasonography in both patients revealed a very hypoplastic thyroid gland. By direct sequencing of the Thyrotropin Receptor gene, we identified the substitution of threonine in place of a highly conserved alanine at position 553, in the fourth predicted transmembrane domain. The mutation was found homozygous in the affected siblings, and heterozygous in both parents and two unaffected siblings. Functional analysis in transfected COS-7 cells showed that it resulted in extremely low expression at the cell surface as compared with the wild-type Receptor, in spite of an apparently normal intracellular synthesis. The small amount of mutated Receptor expressed at the surface of transfected cells bound Thyrotropin with normal affinity and responded in terms of cAMP production, but the in vivo significance of these data from overexpressed Receptor in transfected cells is unclear. Of note, blood thyroglobulin was unexpectedly elevated in the patients at the time of diagnosis, a finding that might prove useful in refining etiologies of congenital hypothyroidism.

Lewis E Braverman - One of the best experts on this subject based on the ideXlab platform.

Sridevi Devaraj - One of the best experts on this subject based on the ideXlab platform.

  • 140 DOI: 10.1309/AJCPX5VNAUYN8MUB © American Society for Clinical Pathology AJCP / Editorial Thyrotropin Receptor Antibody Assays Clinical Utility
    2016
    Co-Authors: William E. Winter, Ishwarlal Jialal, Sridevi Devaraj
    Abstract:

    In this issue of the Journal, Leschik and colleagues1 com-pare the analytical performance and the clinical utility of a commercially available thyroid-stimulating immunoglobulin (TSI) reporter bioassay with an automated Thyrotropin-bind-ing inhibitory immunoglobulin (TBII) electrochemilumines-cent immunoassay. The TSI reporter bioassay is a cell-based assay (or “bioassay”) that uses a genetically engineered cell line capable of specifically detecting serum TSI using cyclic AMP–dependent luciferase activity as the readout. The TBII assay is an anti–Thyrotropin Receptor (TSHR) electrochemilu-minescent immunoassay that uses a porcine TSHR and human anti-TSHR autoantibody M22. The authors report that the TSI bioassay is able to detect lower levels of anti-TSHR autoan-tibodies than the TBII assay and exhibits better precision, but they do not present data indicating whether either assa

  • Thyrotropin Receptor antibody assays clinical utility
    American Journal of Clinical Pathology, 2013
    Co-Authors: William E. Winter, Ishwarlal Jialal, Sridevi Devaraj
    Abstract:

    In this issue of the Journal , Leschik and colleagues1 compare the analytical performance and the clinical utility of a commercially available thyroid-stimulating immunoglobulin (TSI) reporter bioassay with an automated Thyrotropin-binding inhibitory immunoglobulin (TBII) electrochemiluminescent immunoassay. The TSI reporter bioassay is a cell-based assay (or “bioassay”) that uses a genetically engineered cell line capable of specifically detecting serum TSI using cyclic AMP–dependent luciferase activity as the readout. The TBII assay is an anti–Thyrotropin Receptor (TSHR) electrochemiluminescent immunoassay that uses a porcine TSHR and human anti-TSHR autoantibody M22. The authors report that the TSI bioassay is able to detect lower levels of anti-TSHR autoantibodies than the TBII assay and exhibits better precision, but they do not present data indicating whether either assay is more predictive than the other in determining patients’ responses to antithyroid treatment. However, they clearly establish the sensitivity of both assays. TSHR autoantibody (TRAb) measurements (as either TSI or TBII) generally have few potential clinical applications: confirming the diagnosis of Graves disease, the prediction of risk for Graves disease in a neonate born to a mother with Graves disease in pregnancy or in the past, and the prediction of relapse in the medical treatment of Graves disease. In most patients, the diagnosis of Graves disease can be made in the clinical setting of thyrotoxicosis, diffuse goiter, ophthalmopathy (exophthalmos), and dermopathy (pretibial myxedema).2,3 The diagnosis is easily confirmed in most patients with a low thyroid-stimulating hormone (TSH) concentration and an elevated FT4 concentration. In early Graves disease, if T4 levels are normal the …

Rebecca S Bahn - One of the best experts on this subject based on the ideXlab platform.