Delineation of Five Thyroglobulin T Cell Epitopes with Pathogenic Potential in Experimental Autoimmune Thyroiditis

Abstract

Experimental autoimmune thyroiditis (EAT) is a T cell-mediated disease that can be induced in mice after challenge with thyroglobulin (Tg) or Tg peptides. To date, five pathogenic Tg peptides have been identified, four of which are clustered toward the C-terminal end. Because susceptibility to EAT is under control of H-2Ak genes, we have used an algorithm-based approach to identify Ak-binding peptides with pathogenic potential within mouse Tg. Eight candidate synthetic peptides, varying in size from 9 to 15 aa, were tested and five of those (p306, p1579, p1826, p2102, and p2596) were found to induce EAT in CBA/J (H-2k) mice either after direct challenge with peptide in adjuvant or by adoptive transfer of peptide-sensitized lymph node cells (LNCs) into naive hosts. These pathogenic peptides were immunogenic at the T cell level, eliciting specific LNC proliferative responses and IL-2 and/or IFN-γ secretion in recall assays in vitro, but contained nondominant epitopes. All immunogenic peptides were confirmed as Ak binders because peptide-specific LNC proliferation was blocked by an Ak-specific mAb, but not by a control mAb. Peptide-specific serum IgG was induced only by p2102 and p2596, but these Abs did not bind to intact mouse Tg. This study reaffirms the predictive value of Ak-binding motifs in epitope mapping and doubles the number of known pathogenic T cell determinants in Tg that are now found scattered throughout the length of this large autoantigen. This knowledge may contribute toward our understanding of the pathogenesis of autoimmune thyroiditis.

Iodine content of thyroglobulin in NOD.H2h4 mice developing iodine-accelerated autoimmune thyroiditis

“…Lastly, we have tested T-cell responses of NOD.H2 h4 mice with ISAT against Tg, I-Tg and a panel of known A k -restricted, iodinated and non-iodinated peptides of Tg, most of which are pathogenic. 17,[19][20][21] Our results do not corroborate the view that ISAT in NOD-H2 h4 mice is associated with I-Tg formation or a response to neoantigenic Tg determinants containing iodine. Instead, they suggest that iodine may accelerate disease by amplifying dysregulation at other levels of the immune response.…”

Tolerogenic Semimature Dendritic Cells Suppress Experimental Autoimmune Thyroiditis by Activation of Thyroglobulin-Specific CD4+CD25+ T Cells

“…of outbred ICR mice (Harlan Bioproducts for Science), as previously described (25). OVA was purchased from Sigma-Aldrich.…”

“…Thyroid glands were removed 18 -21 days after Tg challenge to be assessed for thyroid pathology. Fixation, embedding, and sectioning of thyroids were performed, as previously described (25). Histological sections were stained with H&E, and the mononuclear cell infiltration index (I.I.)…”

“…Eighteen hours before harvesting, 1 Ci of [ 3 H]thymidine (6.7 Ci/mol; DuPont Pharmaceuticals) was added to each well in 25 l of complete medium. Cell harvesting and radioactivity measurements were performed, as previously described (25). Stimulation index is defined as: (cpm in the presence of Ag)/(cpm in the absence of Ag).…”

Identification of Pathogenic T Cell Epitopes Near Cathepsin Cleavage Sites in Thyroglobulin

“…Since its discovery as an autoantigen in 1956 (13), 25 Tg pathogenic epitopes have been identified using a variety of approaches (14). A common strategy involved the use of computer algorithms to delineate putative MHC-binding Tg peptides (15)(16)(17)(18)(19), whereas other methods used cloned T cell hybridomas as screening tools (20,21), elution of human MHC-bound peptides (22,23), or focused on searching for T cell epitopes shared between Tg and thyroid peroxidase (24). All known Tg pathogenic peptides have fallen under three categories: 1) iodinated peptides containing iodotyrosyls; 2) peptides encompassing hormonogenic sites; and 3) noniodinated peptides scattered throughout the Tg molecule (14).…”