Antibodies to membrane antigens in autoimmune thyroid disease

1987 ◽  
Vol 116 (1) ◽  
pp. 13-20 ◽  
Author(s):  
J. Furmaniak ◽  
J. Bradbury ◽  
B. Rees Smith
Keyword(s):  
Solid Phase ◽  
Protein A ◽  
Human Thyroid ◽  
Intermediate Filament Protein ◽  
Autoimmune Thyroid Diseases ◽  
Autoimmune Thyroid ◽  
Microsomal Antibody ◽  
Membrane Antigens ◽  
Solid Phase Assay ◽  
Microsomal Antigen

Abstract. The possibility that sera from patients with autoimmune thyroid diseases contain autoantibodies to thyroid membrane proteins distinct from microsomal antigen and the TSH receptor has been investigated using (a) solid phase assay system based on human thyroid membranes and 125I-labelled protein A and (b) immunoprecipitation of detergent solubilized 125I-labelled thyroid membranes followed by gel electrophoresis and autoradiography. In the solid phase assay binding to membranes showed a highly significant correlation with binding to microsomes (r = 0.82; P < 0.001; N = 82) indicating that the interaction between the serum and the membranes was due principally to microsomal antibody binding to microsomal antigen contaminating the membrane preparations. However, there were some discrepancies suggesting that an additional antigen-antibody system was involved. This possibility was then investigated using immunoprecipitation of 125I-labelled thyroid membranes. A labelled protein with mol wt 54 K was specifically immunoprecipitated (relative to normal pool serum) by 3 out of 4 sera from patients with Graves' disease who showed high binding to thyroid membranes. A further 4 sera from such patients with low membrane binding affinity failed to immunoprecipitate the 54 K protein. Sera from some patients with Hashimoto's disease and some patients with rheumatoid arthritis and one patient with Addison's disease also immunoprecipitated the 54 K protein from solubilized thyroid membranes. These studies suggested that antibodies interacting with the 54 K protein contributed to the discrepancies between thyroid membrane and microsome binding. However, the 54 K protein was also immunoprecipitated from detergent solubilized membranes prepared from human placenta, skeletal muscle and adrenal tissue. Immunoprecipitation studies with antisera to cytoskeleton proteins suggested that the 54 K band was the intermediate filament protein desmin. Consequently, thyroid specific antibody-antigen systems distinct from those involving microsomal antibody (or thyroglobulin antibody) could not be detected in thyroid membranes by immunoprecipitation.

Simple and sensitive method for estimation of antithyroid plasma membrane antibodies in the serum of patients with autoimmune thyroid diseases; comparison with other assays

1984 ◽  
Vol 105 (4) ◽  
pp. 492-499 ◽  
Author(s):  
Andrzej Gardas ◽  
Barbara Czarnocka ◽  
Marta Faryna ◽  
Grazyna Adler ◽  
Alexandra Lewartowska ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Plasma Membranes ◽  
Blood Donors ◽  
Protein A ◽  
Mean Value ◽  
Adenyl Cyclase ◽  
Graves Disease ◽  
Autoimmune Thyroid Diseases ◽  
Autoimmune Thyroid ◽  
Index Estimation

Abstract. The ability of protein A from Staphylococcus aureus to interact with Fc fragments of IgG was used to estimate the antithyroid plasma membrane antibodies in sera of patients with Graves' disease. The results were expressed as an antithyroid plasma membrane antibodies (ATMA) index. The ATMA index estimated in 60 healthy blood donors varied from 0.57 to 1.28, with a mean value of 0.99, sd Ø 0.20. The ATMA index in hyperthyroid untreated Graves' disease varied from 1.80 to 8.0, with a mean value of 4.7. Autoantibody binding to thyroid plasma membranes could be inhibited by (Fab)2 fragments obtained from the serum of patients with Graves' disease but not by (Fab)2 fragments obtained from the serum of healthy blood donors. The influence of rabbit antithyroglobulin and antimicrosomal antibodies on the ATMA index estimation has been evaluated. The ATMA index estimation was compared with the thyrotrophin binding inhibiting immunoglobulins (TBII) index and with the adenyl cyclase stimulating activity of immunoglobulins obtained from 92 hyperthyroid Graves' patients. The ATMA index was positive in 97%, the TBII index in 62% and TSI in 35% of cases. This method using protein A could also be used for estimation of ATMA in other autoimmune thyroid disorders.

scholarly journals B-cell autoepitopes on the acetylcholinesterase-homologous region of human thyroglobulin: association with Graves' disease and thyroid eye disease

2001 ◽  
pp. 119-127 ◽  
Author(s):  
A Thrasyvoulides ◽  
M Sakarellos-Daitsiotis ◽  
G Philippou ◽  
A Souvatzoglou ◽  
C Sakarellos ◽  
...  
Keyword(s):  
Solid Phase ◽  
Significant Proportion ◽  
Terminal Region ◽  
Thyroid Eye Disease ◽  
Eye Disease ◽  
Homologous Region ◽  
Graves Disease ◽  
Healthy Controls ◽  
Autoimmune Thyroid Diseases ◽  
Autoimmune Thyroid

OBJECTIVE: Thyroglobulin (Tg) is a large autoantigen involved in autoimmune thyroid diseases. Tg epitopes have, so far, been identified within large peptides. In the present study, we used small synthetic peptides to finely map serological epitopes on the highly immunogenic C-terminal region of Tg. Homology of this region to acetylcholinesterase (AChE) has been implicated in the pathogenesis of thyroid eye disease (TED) through cross-reactive antibodies. METHODS: We tested total IgG purified from four pilot Graves' disease (GD) sera reactive with both Tg and AChE and from three healthy controls, for reactivity against overlapping 20mer peptides (pin synthesis) covering the sequence 2171-2748 of human Tg. Antibody-reactive peptides were subsequently synthesized by a solid-phase technique for confirmation with a large number of sera: 99 GD, 32 Hashimoto's thyroiditis (HT) and 45 healthy controls. RESULTS: Peptides TgP15, TgP26 and TgP41 (amino acids 2339-2358, 2471-2490 and 2651-2670 respectively) were found to be targets of autoantibodies on intact Tg, recognized by a statistically significant proportion of GD sera (22.2%, 35.4% and 30.3% respectively), compared with HT (0%, 15.6% and 6.3% respectively) and healthy controls (0%, 4.4% and 4.4% respectively). The majority of GD sera (56.6%) were positive for at least one of the three peptides. In GD, TgP26 reactivity was found to be associated with TED (48.6% with TED versus 25.5% without TED, P<0.05). CONCLUSION: Some epitopes on the C-terminal region of Tg are associated with GD. A subset of Tg-reactive autoantibodies, directed to this region, is associated with TED and may be involved in the development of the disease.

scholarly journals Multicenter study on TGPO autoantibody prevalence in various thyroid and non-thyroid diseases; relationships with thyroglobulin and thyroperoxidase autoantibody parameters

1999 ◽  
pp. 563-569 ◽  
Author(s):  
V Estienne ◽  
C Duthoit ◽  
VD Costanzo ◽  
PJ Lejeune ◽  
M Rotondi ◽  
...  
Keyword(s):  
Thyroid Nodules ◽  
Solid Phase ◽  
Post Partum ◽  
Large Population ◽  
Collaborative Study ◽  
Normal Subjects ◽  
Thyroid Diseases ◽  
Serum Samples ◽  
Autoimmune Thyroid Diseases ◽  
Autoimmune Thyroid

OBJECTIVE: TGPO autoantibodies (aAbs) that bind simultaneously to thyroglobulin (Tg) and thyroperoxidase (TPO) are present in the serum of patients with autoimmune thyroid diseases (AITD) and have been found to differ from monospecific Tg and TPO aAbs. To obtain further insights on the prevalence defined as the rate of occurrence and significance of TGPO aAbs in a large population, we carried out a collaborative study involving 15 European teams. METHODS: Serum samples from 3122 patients with various thyroid and non-thyroid diseases and normal subjects were assayed using a novel TGPO aAb detection kit. This test was designed so that TGPO aAbs are trapped between the Tg-coated solid phase and the soluble TPO labeled with a radioiodinated monoclonal antibody. RESULTS: Only three out of the 220 normal subjects (prevalence of 1.4%) were found to have positive TGPO aAb levels, which were mainly observed in the patients with AITD: the group of patients suffering from Hashimoto's thyroiditis had a TGPO aAb prevalence of 40.5% (n=437 patients), those with Graves' disease, a prevalence of 34.6% (n=645) and those with post-partum thyroiditis, 16.0% (n=243). Among the non-AITD patients with positive TGPO aAb levels, the TGPO aAb prevalence ranged from 20.7% among those with thyroid cancer (n=246) to 0% among those with toxic thyroid nodules (n=47). Among the patients with non-thyroid diseases, the TGPO aAb prevalence ranged from 9.8% in the case of Biermer's pernicious anemia (n=78) to 0% in that of premature ovarian failure (n=44). It is worth noting that the groups showing the highest TGPO aAb prevalence also contained the patients with the highest TGPO aAb titers. Statistical comparisons between the TGPO aAb prevalences in the various groups showed that TGPO aAb could be used as a parameter to distinguish between the groups of Hashimoto's and Graves' patients and between the women with post-partum thyroiditis and the post-partum women with only Tg and/or TPO aAb established during early pregnancy. Unexpectedly, the correlations between TGPO aAbs and Tg and TPO aAbs were found to depend mainly on the assay kit used. CONCLUSION: High TGPO aAb titers are consistently associated with AITD but the reverse was not found to be true. TGPO aAbs are a potentially useful tool, however, for establishing Hashimoto's diagnosis, and would be worth testing in this respect with a view to using them for routine AITD investigations.

scholarly journals Microarray Analysis of Cytokine Activation of Apoptosis Pathways in the Thyroid

Endocrinology ◽  
2007 ◽  
Vol 148 (10) ◽  
pp. 4844-4852 ◽  
Author(s):  
Su He Wang ◽  
Mary Van Antwerp ◽  
Rork Kuick ◽  
Paul G. Gauger ◽  
Gerard M. Doherty ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
P38 Mapk ◽  
Human Thyroid ◽  
Autoimmune Thyroid Diseases ◽  
Thyroid Cells ◽  
Mapk Phosphorylation ◽  
Autoimmune Thyroid ◽  
Cytokine Activation ◽  
Ifn Γ ◽  
Sb 203580

It has been suggested that Fas-mediated apoptosis plays an important role in the pathogenesis of autoimmune thyroid diseases. Our previous studies have demonstrated that normal primary thyroid epithelial cells are resistant to Fas-mediated apoptosis, but the resistance can be overcome by pretreatment with a combination of interferon-γ (IFN-γ) and IL-1β. To understand the molecular mechanism responsible for the IFN-γ/IL-1β effects, we profiled changes in the transcription induced by these two cytokines in normal human thyroid cells, using cDNA microarrays. We found that IFN-γ/IL-1β showed a significant increase in apoptosis-related genes such as inducible nitric oxide synthase (iNOS), receptor-interacting protein 2 (RIP2), and caspases 10. These increases were confirmed by other methods, including real-time PCR and Western blot. Furthermore, the sensitization of primary thyroid epithelial cells to Fas-mediated apoptosis by IFN-γ/IL-1β was significantly blocked by a general caspase inhibitor, z-VAD, or by the combination of two specific individual caspase inhibitors. In addition, our results showed that IFN-γ/IL-1β enhance p38 MAPK phosphorylation and that SB 203580, a p38 MAPK inhibitor, can inhibit IFN-γ/IL-1β-induced p38 MAPK phosphorylation. SB 203580 also significantly prevented cytokine-induced iNOS expression and caspase activation and thus blocked Fas-mediated apoptosis of thyroid cells sensitized by IFN-γ/IL-1β. In conclusion, our data suggest that both p38 MAPK and iNOS are involved in IFN-γ/IL-1β-induced sensitization of the thyroid cells to Fas-mediated apoptosis via the activation of caspases 3, 7, and 10 and that this pathway may be further activated by BID. This hints that inflammatory cytokines regulate death-receptor-mediated apoptosis at multiple points in the process.

scholarly journals Third generation radioimmunoassay (RIA) for TSH receptor autoantibodies (TRAb) – one step less, similar results?

2021 ◽  
Vol 60 (01) ◽  
pp. 38-46
Author(s):  
Johannes J. Roggenbuck ◽  
Grit Zarske ◽  
Peter Schierack ◽  
Gerd Wunderlich ◽  
Karsten Conrad ◽  
...  
Keyword(s):  
Solid Phase ◽  
Tsh Receptor ◽  
Operating Characteristics ◽  
Step Method ◽  
Autoimmune Thyroid Diseases ◽  
Autoimmune Thyroid ◽  
Assay Procedure ◽  
Assay Performance ◽  
One Step ◽  
The One

Abstract Aim TSH-receptor (TSHR)-autoantibody (TRAb) is the serological hallmark of Graves’ disease (GD). Recently, 3rd-generation radioimmunoassays (RIA) employing monoclonal TRAb such as M22 or T7 instead of TSH for the inhibition of human TRAb binding with solid-phase TSHR (coated tubes) have been introduced into laboratory routine. Methods As current assays typically employ a consecutive incubation of patient serum and labelled monoclonal TRAb, automation of TRAb RIA is a challenge. Thus, the assay procedure using human TSHR-coated tubes and the mouse monoclonal TRAb T7 was modified by combining both steps. The novel one-step method was compared with its corresponding consecutive 3rd-generation RIA by investigating 304 individuals encompassing 102 patients with active GD (GDa), 43 patients with GD after successful therapy (GDt), 31 with Hashimoto’s disease (HD), 28 with non-autoimmune thyroid diseases (NAITD) and 100 healthy subjects (HS). Results With the new method, the incubation time was shortened by approximately one hour. Both 3rd-generation RIAs did not reveal a significantly different assay performance by comparing areas under the curve (AUC) with receiver operating characteristics curve analysis (AUC one-step: 0.94, AUC two-step: 0.96, p > 0.05, respectively). The two-step TRAb RIA demonstrated sensitivity and specificity values of 87.5 % and 96.2 %, respectively, whereas the one-step revealed 84.6 % and 96.2 %, respectively. Conclusion One-step 3rd-generation RIA may be used for the reliable detection of TRAb. The shorter and easier assay design may improve its use and enable automation in routine nuclear medicine laboratories.

A solid-phase assay for antiactin antibody and actin using protein-A

1979 ◽  
Vol 94 (1) ◽  
pp. 140-149 ◽  
Author(s):  
James L. Lessard ◽  
Dawn Carlton ◽  
Diane C. Rein ◽  
Richard Akeson
Keyword(s):  
Solid Phase ◽  
Protein A ◽  
Solid Phase Assay

scholarly journals Analytical and clinical performance goals for testing autoantibodies to thyroperoxidase, thyroglobulin, and thyrotropin receptor

1996 ◽  
Vol 42 (1) ◽  
pp. 160-163 ◽  
Author(s):  
U Feldt-Rasmussen
Keyword(s):  
Medical Research Council ◽  
Clinical Performance ◽  
Binding Capacity ◽  
Thyroid Autoimmunity ◽  
Human Thyroid ◽  
Thyroid Autoantibodies ◽  
Thyrotropin Receptor ◽  
Autoimmune Thyroid Diseases ◽  
Autoimmune Thyroid ◽  
Antigen Epitope

Abstract Methods for measuring thyroid autoantibodies--to thyroperoxidase (TPOAb), thyroglobulin (TgAb), and thyrotropin receptor (TRAb)--have improved over the last decade, but increasingly, accurate and sensitive methods are needed for identifying patients with autoimmune thyroid diseases and individuals at high risk for onset of thyroid autoimmunity. With the increased quality requirements for these methods, it becomes more important to look at the functional sensitivities and precision profiles of the various methods. International standardization in this field is also needed. Because most sera containing human thyroid autoantibodies display a variety of antigen-specific immunoglobulins of different classes and subclasses with different affinity and avidity in their epitope reaction, investigators must decide whether the autoantibodies should be quantified in terms of immunoglobulin content, antigen/epitope reactivity, or binding capacity. Until these problems are solved, the best means for standardization are the Medical Research Council calibrators for TPOAb and TgAb, whereas no standardization exists for TRAb.

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