title: Targeted expression of the human thyrotropin receptor A-subunit to the mouse thyroid: insight into overcoming the lack of response to A-subunit adenovirus immunization.
abstract: The thyrotropin receptor (TSHR), the major autoantigen in Graves' disease, is posttranslationally modified by intramolecular cleavage to form disulfide-linked A- and B-subunits. Because Graves' hyperthyroidism is preferentially induced in BALB/c mice using adenovirus encoding the free A-subunit rather than full-length human TSHR, the shed A-subunit appears to drive the disease-associated autoimmune response. To further investigate this phenomenon, we generated transgenic mice with the human A-subunit targeted to the thyroid. Founder transgenic mice had normal thyroid function and were backcrossed to BALB/c. The A-subunit mRNA expression was confirmed in thyroid tissue. Unlike wild-type littermates, transgenic mice immunized with low-dose A-subunit adenovirus failed to develop TSHR Abs, hyperthyroidism, or splenocyte responses to TSHR Ag. Conventional immunization with A-subunit protein and adjuvants induced TSHR Abs lacking the characteristics of human autoantibodies. Unresponsiveness was partially overcome using high-dose, full-length human TSHR adenovirus. Although of low titer, these induced Abs recognized the N terminus of the A-subunit, and splenocytes responded to A-subunit peptides. Therefore, "non-self" regions in the B-subunit did not contribute to inducing responses. Indeed, transgenic mice immunized with high-dose A-subunit adenovirus developed TSHR Abs with thyrotropin-binding inhibitory activity, although at lower titers than wild-type littermates, suggesting down-regulation in the transgenic mice. In conclusion, in mice expressing a human A-subunit transgene in the thyroid, non-self human B-subunit epitopes are not necessary to induce responses to the A-subunit. Our findings raise the possibility that autoimmunity to the TSHR in humans may not involve epitopes on a cross-reacting protein, but rather, strong adjuvant signals provided in bystander immune responses.
authors.affiliation: Autoimmune Disease Unit, Cedars-Sinai Research Institute, and University of California School of Medicine, Los Angeles, CA 90095, USA.
authors.forename: Pavel N
authors.initials: PN
authors.lastname: Pichurin
authors.forename: Chun-Rong
authors.initials: CR
authors.lastname: Chen
authors.forename: Gregorio D
authors.initials: GD
authors.lastname: Chazenbalk
authors.forename: Holly
authors.initials: H
authors.lastname: Aliesky
authors.forename: Nancy
authors.initials: N
authors.lastname: Pham
authors.forename: Basil
authors.initials: B
authors.lastname: Rapoport
authors.forename: Sandra M
authors.initials: SM
authors.lastname: McLachlan
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D001323:Autoantibodies / Q000276:immunology; D001324:Autoantigens / Q000276:immunology; D015551:Autoimmunity; D004797:Enzyme-Linked Immunosorbent Assay; D005822:Genetic Vectors; D006111:Graves Disease / Q000276:immunology; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D021122:Protein Subunits / Q000276:immunology*; D011989:Receptors, Thyrotropin / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology*; D017386:Sequence Homology, Amino Acid; D013961:Thyroid Gland / Q000276:immunology* / Q000473:pathology
chemical_list: D001323:Autoantibodies; D001324:Autoantigens; D021122:Protein Subunits; D011989:Receptors, Thyrotropin