Human monoclonal thyroglobulin autoantibodies: epitopes and immunoglobulin genes.
01 Oct 2004-The Journal of Clinical Endocrinology and Metabolism (Endocrine Society)-Vol. 89, Iss: 10, pp 5116-5123
TL;DR: The TgAb panel provides novel information regarding the repertoire of H chain genes encoding human TgAbs as well as the relationship between the H chains and the epitopes recognized on this major thyroid autoantigen.
Abstract: Autoantibodies to thyroglobulin (TgAbs) are common markers of thyroid autoimmunity, but relatively few human monoclonal TgAbs have been described. From a panel of 64 human monoclonal TgAbs (isolated from a thyroid-disease derived combinatorial Ig gene library), we selected seven with unique genetic features for detailed characterization. These TgAbs preferentially recognize native (not denatured) Tg, like serum autoantibodies. Most have high affinities for Tg (dissociation constant 10(-10) to 10(-9) m). Their light (L) chain Ig genes are not unusual, but four of the five heavy (H) chain genes are new. Moreover, one H chain belongs to the small VH2 family, not previously reported for autoantibodies to Tg or thyroid peroxidase. The TgAbs inhibit the binding to Tg of the thyroid donor's serum autoantibodies, indicating epitopic overlap. Competition analysis (surface plasmon resonance) shows that the TgAbs recognize overlapping epitopes in an immunodominant region on the Tg dimer ( approximately 660 kDa). Two major and several minor epitopic regions were defined, each associated with a particular H + L chain combination. In conclusion, our TgAb panel provides novel information regarding the repertoire of H chain genes encoding human TgAbs as well as the relationship between the H chains and the epitopes recognized on this major thyroid autoantigen.
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...[111–289], antibody–antigen [290–411] and receptor–...
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...research performed using Biacore instruments [111–931]....
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TL;DR: The goal of inducing self-tolerance to prevent autoimmune thyroid disease will require accurate prediction of at-risk individuals together with an antigen-specific, not blanket, therapeutic approach.
Abstract: Thyroid autoimmunity involves loss of tolerance to thyroid proteins in genetically susceptible individuals in association with environmental factors. In central tolerance, intrathymic autoantigen presentation deletes immature T cells with high affinity for autoantigen-derived peptides. Regulatory T cells provide an alternative mechanism to silence autoimmune T cells in the periphery. The TSH receptor (TSHR), thyroid peroxidase (TPO), and thyroglobulin (Tg) have unusual properties ("immunogenicity") that contribute to breaking tolerance, including size, abundance, membrane association, glycosylation, and polymorphisms. Insight into loss of tolerance to thyroid proteins comes from spontaneous and induced animal models: 1) intrathymic expression controls self-tolerance to the TSHR, not TPO or Tg; 2) regulatory T cells are not involved in TSHR self-tolerance and instead control the balance between Graves' disease and thyroiditis; 3) breaking TSHR tolerance involves contributions from major histocompatibility complex molecules (humans and induced mouse models), TSHR polymorphism(s) (humans), and alternative splicing (mice); 4) loss of tolerance to Tg before TPO indicates that greater Tg immunogenicity vs TPO dominates central tolerance expectations; 5) tolerance is induced by thyroid autoantigen administration before autoimmunity is established; 6) interferon-α therapy for hepatitis C infection enhances thyroid autoimmunity in patients with intact immunity; Graves' disease developing after T-cell depletion reflects reconstitution autoimmunity; and 7) most environmental factors (including excess iodine) "reveal," but do not induce, thyroid autoimmunity. Micro-organisms likely exert their effects via bystander stimulation. Finally, no single mechanism explains the loss of tolerance to thyroid proteins. The goal of inducing self-tolerance to prevent autoimmune thyroid disease will require accurate prediction of at-risk individuals together with an antigen-specific, not blanket, therapeutic approach.
160 citations
TL;DR: The pattern of Tg recognition is similar when HT patients are compared to GD and NTMG to PTC patients and differs when AITD are compared with non-AITD patients.
Abstract: Context: Thyroglobulin (Tg) epitopes of serum Tg autoantibodies (TgAb) have been characterized using inhibition of Tg binding by human monoclonal TgAb in autoimmune thyroid diseases (AITD) [Hashimoto’s thyroiditis (HT) and Graves’ disease (GD)] but not in non-AITD [nontoxic multinodular goiter (NTMG) and papillary thyroid carcinoma (PTC)]. Objective: Our objective was to compare Tg epitopes of serum TgAb from patients with AITD, non-AITD, and PTC associated with histological thyroiditis (PTC-T) using inhibition of Tg binding by four recombinant human TgAb-Fab (epitopic regions A–D). Design: Inhibition of Tg binding of 24 HT, 25 GD, 19 NTMG, 15 PTC, and 25 PTC-T TgAb-positive sera by each TgAb-Fab was evaluated in ELISA. Inhibition by the pool of the four TgAb-Fab was evaluated using labeled Tg. Results: Levels of inhibition were different for TgAb-Fab regions A (P = 0.001), B (0.007), and D (0.011). Inhibition by region A TgAb-Fab was significantly higher in HT, GD, and PTC-T than in NTMG and PTC patients...
71 citations
TL;DR: Iodine-induced thyroid autoimmunity is related to TgAb and the unmasking of a cryptic epitope on Tg contributes to this relationship in humans.
Abstract: Context: The mechanisms linking thyroid autoimmunity and iodine use in humans are unknown. Objective: Our aim was to correlate iodine intake, thyroid autoimmunity, and recognition of thyroglobulin (Tg) epitopes after implementation of iodine prophylaxis. Setting: The general community living in an Italian village was evaluated. Main Outcome Measures: Thyroglobulin autoantibodies (TgAb), thyroperoxidase autoantibodies (TPOAb), and urinary iodine excretion were assessed in 906 iodized salt users (IS-users) and 389 nonusers (IS-nonusers). Ultrasound (US) was performed to identify thyroid hypoechogenicity, suggestive of Hashimoto thyroiditis (HT). TgAb epitope pattern in 16 IS-users and 17 IS-nonusers was evaluated by an inhibition binding assay to Tg, using human monoclonal TgAb-Fab directed to A, B, C, and D epitopes on Tg. Results: Median urinary iodine excretion was slightly higher in IS-users than in IS-nonusers (112.0 μg/L vs 86.5 μg/L; P < .01). TgAb, and not TPOAb, was more frequent in IS-users (18.9%...
67 citations
TL;DR: In papillary thyroid carcinoma, LT on histology must be carefully searched for because it is frequently associated with TgAb and therefore mistakenly low or undetectable Tg, which can be underestimated in patients with T g autoantibodies (TgAb).
Abstract: Context: Serum thyroglobulin (Tg), the marker of residual tumor in papillary thyroid carcinoma, can be underestimated in patients with Tg autoantibodies (TgAb) TgAb are due to a coexistent lymphocytic thyroiditis (LT) or the papillary thyroid carcinoma per se TgAb assays are highly discordant Design: We evaluated 141 patients with a clinical diagnosis of nodular thyroid disease, 32 of Hashimoto's thyroiditis, and four of Graves' disease, who underwent total thyroidectomy for an associated papillary thyroid carcinoma Patients were classified as papillary thyroid carcinoma-lymphocytic thyroiditis (PTC-T) and papillary thyroid carcinoma (PTC) according to the presence or absence of LT on histology Tg was measured before thyroid remnant ablation, when it is expectedly detectable, by an immunometric assay (IMA) and TgAb by three noncompetitive IMA and three competitive radioimmunoassays (RIA) The number of lymphocytes was compared with TgAb concentration Results: Seventy-two of 177 patients (407%) were
59 citations
References
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TL;DR: Thyroglobulin autoantibodies in the globulin fractions from six human sera showed between 70 and 100% cross-reaction with one another, indicating that the majority of antibodies are directed against the same epitopes, consistent with two distinct major epitopic specificities with occasional sera having antibodies directed against a third site.
Abstract: Fragments of thyroglobulin containing the autoreactive epitopes were prepared by an existing procedure, plus an ion-exchange chromatography step which increased their purity. Before purification, 39% of the fragmental material was bound by rabbit anti-human thyroglobulin whereas only 8% was bound by autoantibody, thus confirming earlier reports of more limited epitopes for human autoantibodies than heterologous antibody. Thyroglobulin autoantibodies in the globulin fractions from six human sera showed between 70 and 100% cross-reaction with one another, indicating that the majority of antibodies are directed against the same epitopes. The data are consistent with an estimated of two distinct major epitopic specificities with occasional sera having antibodies directed against a third site. Chemical modification of tyrosine residues by iodination did not inhibit the binding of thyroglobulin to either human autoantibodies or rabbit anti-human thyroglobulin; thus tyrosine residues do not contribute significantly to the epitopes for either auto- or heteroantibodies.
82 citations
TL;DR: The finding of identical fine specificities for anti‐hTgb Ab in normal and pathological conditions implies that autoantibodies are produced in normal subjects and held to a low level by regulatory processes which fail with respect to selected epitopes in autoimmune diseases.
Abstract: Polyclonal anti-human thyroglobulin (hTgb) antibodies (Ab) were purified from sera of rabbits immunized with human thyroglobulin, normal humans and patients suffering from Graves' disease, Hashimoto's thyroiditis and thyroid carcinoma. The avidity of the various Ab preparations for hTgb ranged from 0.3 X 10(10) -2.2 X 10(10) M-1. By using well characterized mouse monoclonal antibodies (mAb) directed against hTgb, it was shown that the fine specificities of induced anti-hTgb Ab in rabbits, natural Ab in normal subjects and autoantibodies in diseased patients were similar; however, they differed from that of rabbit anti-bovine and anti-porcine thyroglobulin Ab which were able to inhibit the hTgb binding of only a few of the mAb. Anti-hTgb in rabbits and in patients with thyroid carcinoma varied from those in normal subjects only by uniformly elevated serum titers. In contrast, patients with Graves' disease and Hashimoto's thyroiditis showed an increased concentration essentially restricted to Ab reacting with few of the antigenic determinants recognized by the mAb. Our data suggest that the repertoire of anti-hTgb Ab is similar in mouse, rabbit and human. Furthermore, the finding of identical fine specificities for anti-hTgb Ab in normal and pathological conditions implies that autoantibodies are produced in normal subjects and held to a low level by regulatory processes which fail with respect to selected epitopes in autoimmune diseases.
75 citations
TL;DR: The regions on the human thyroglobulin molecule recognized by anti-hTg autoantibodies in the sera of patients with Hashimoto's thyroiditis, Graves' disease, and thyroid carcinoma were determined and the natural aAbs present in the serum at low concentrations were isolated.
74 citations
TL;DR: It is found that the 783 rearranged VH1 gene is likely to be the unmutated form of a germline VH gene that may encode the heavy chains of RF Bor and Kas and several other human monoclonal RF.
Abstract: We isolated a VH1 germline gene (Humhv 1263) that is closely related to the heavy chains of 2 human rheumatoid factors (RF), Bor and Kas. We also found that the 783 rearranged VH1 gene is actually 91–93% homologous to Bor and Kas, and it differs from a VH1 complementary DNA (51P1) by only a single base. Thus, 783 is likely to be the unmutated form of a germline VH gene that may encode the heavy chains of RF Bor and Kas and several other human monoclonal RF.
67 citations
TL;DR: The identification and cloning of TPO (thyroid microsomal antigen) provided the critical tool for analyzing B and T cell reactivity to this major thyroid autoantigen as discussed by the authors.
Abstract: Autoimmunity to thyroid peroxidase (TPO), manifest as high affinity IgG class auto-antibodies, is the common denominator of human thyroid autoimmunity, encompassing patients with overt hyper-or hypothyroidism as well as euthyroid individuals with subclinical disease. The identification and cloning of TPO (the “thyroid microsomal antigen”) provided the critical tool for analyzing B and T cell reactivity to this major thyroid autoantigen. In particular, the availability of immunoreactive TPO permitted the isolation of essentially the entire repertoire of human monoclonal antibodies, a feat unparalled in an organ-specific autoimmune disease. These recombinant autoantibodies (expressed as Fab) provide insight into the genes encoding their H and L chains as well as the conformational epitopes on TPO with which serum autoantibodies interact. Analyses of TPO autoantibody epitopic “fingerprints” indicate a lack of epitope spreading as well as a genetic basis for their inheritance. Limited data are available for t...
66 citations