ABSTRACT
We have studied responses in thymoma patients to interferon-alpha and to the acetylcholine receptor (AChR) in early-onset myasthenia gravis (EOMG), seeking clues to autoimmunizing mechanisms. Our new evidence implicates a two-step process: (step 1) professional antigen-presenting cells and thymic epithelial cells prime AChR-specific T cells; then (step 2) thymic myoid cells subsequently provoke germinal center formation in EOMG. Our unifying hypothesis proposes that AChR epitopes expressed by neoplastic or hyperplastic thymic epithelial cells aberrantly prime helper T cells, whether generated locally or infiltrating from the circulation. These helper T cells then induce antibody responses against linear epitopes that cross-react with whole AChR and attack myoid cells in the EOMG thymus. The resulting antigen-antibody complexes and the recruitment of professional antigen-presenting cells increase the exposure of thymic cells to the infiltrates and provoke local germinal center formation and determinant spreading. Both these and the consequently enhanced heterogeneity and pathogenicity of the autoantibodies should be minimized by early thymectomy.
Subject(s)
Autoimmunity , B-Lymphocytes/immunology , Myasthenia Gravis/immunology , T-Lymphocytes/immunology , Age of Onset , Animals , Autoantibodies , Bungarotoxins/metabolism , Enzyme-Linked Immunosorbent Assay , Epithelial Cells/physiology , Epitopes/immunology , Fluorescent Antibody Technique , Germinal Center , Histocompatibility Antigens Class II/metabolism , Humans , Insulin/metabolism , Interferon-alpha/immunology , Interleukin-2/immunology , Keratins/metabolism , Models, Immunological , Mutation , Myasthenia Gravis/metabolism , Receptors, Cholinergic/immunology , Stromal Cells , T-Lymphocytes/classification , Thymoma/immunology , Thymus Gland/cytology , Thymus Gland/physiology , Thymus Neoplasms , Troponin I/metabolismABSTRACT
A recurring epitope in the human acetylcholine receptor (AChR) alpha subunit (alpha146-160) is presented to specific T cells from myasthenia gravis patients by HLA-DRB3*0101-"DR52a"-or by DR4. Here we first map residues critical for DR52a in this epitope by serial Ala substitution. For two somewhat similar T cells, this confirms the recently deduced importance of hydrophobic "anchor" residues at peptide p1 and p9; also of Asp at p4, which complements this allele's distinctive Arg74 in DRbeta. Surprisingly, despite the 9 sequence differences in DRbeta between DR52a and DR3, merely reducing the bulk of the peptide's p1 anchor residue (Trp149-->Phe) allowed maximal cross-presentation to both T cells by DR3 (which has Val86 instead of Gly). The shared K71G73R74N77 motif in the alpha helices of DR52a and DR3 thus outweighs the five differences in the floor of the peptide-binding groove. A second issue is that T cells selected in vitro with synthetic AChR peptides rarely respond to longer Ag preparations, whereas those raised with recombinant subunits consistently recognize epitopes processed naturally even from whole AChR. Here we compared one T cell of each kind, which both respond to many overlapping alpha140-160 region peptides (in proliferation assays). Even though both use Vbeta2 to recognize peptides bound to the same HLA-DR52a in the same register, the peptide-selected line nevertheless proved to depend on a recurring synthetic artifact-a widely underestimated problem. Unlike these contaminant-responsive T cells, those that are truly specific for natural AChR epitopes appear less heterogeneous and therefore more suitable targets for selective immunotherapy.
Subject(s)
Antigen Presentation , Epitopes, B-Lymphocyte/metabolism , Epitopes, T-Lymphocyte/metabolism , HLA-DR Antigens/immunology , Receptors, Cholinergic/immunology , Alleles , Amino Acid Sequence , Amino Acid Substitution/immunology , Animals , Cell Line , Epitopes, B-Lymphocyte/chemistry , Epitopes, B-Lymphocyte/immunology , Epitopes, T-Lymphocyte/chemistry , Epitopes, T-Lymphocyte/immunology , HLA-DR Antigens/chemistry , HLA-DR Antigens/metabolism , Humans , Mice , Molecular Sequence Data , Myasthenia Gravis/immunology , Peptides/chemical synthesis , Peptides/immunology , Peptides/metabolism , Protein Binding/genetics , Protein Binding/immunology , Receptors, Cholinergic/chemistry , Receptors, Cholinergic/genetics , TorpedoSubject(s)
Myasthenia Gravis/immunology , Myasthenia Gravis/pathology , Receptors, Cholinergic/immunology , T-Lymphocytes/immunology , Thymoma/immunology , Thymus Gland/pathology , Thymus Neoplasms/immunology , Amino Acid Sequence , Animals , Genetic Variation , HLA-DP Antigens/immunology , Humans , Molecular Sequence Data , Peptide Fragments/immunology , Receptors, Cholinergic/chemistry , Sequence Alignment , Sequence Homology, Amino Acid , Thymectomy , Thymoma/surgery , Thymus Gland/immunology , Thymus Neoplasms/pathologyABSTRACT
Most thymic epithelial tumours that associate with MG express an epitope that resembles the sequence alpha373-380 from the cytoplasmic loop of the acetylcholine receptor (AChR). It has been proposed that sensitization to this linear epitope initiates autoimmunity to the AChR in thymoma-associated MG. We therefore tested whether MG/thymoma patients have T cell responses or antibodies to this region of the AChR. We found no significant recognition of the alpha309-417 region by their thymoma or peripheral blood T cells, or by their serum anti-AChR antibodies. Instead, the T cell epitopes that were recognized, like the previously characterized B cell epitopes, were in the extracellular AChR domain.
