Preparation and characterization of monoclonal antithyrotropin receptor antibodies obtained from peripheral lymphocytes of hypothyroid patients with primary myxedema.

Anti-TSH receptor antibodies (TSH-R Ab), which have been detected in the serum of some patients with primary myxedema, are themselves considered to induce hypothyroidism. These are termed blocking-type TSH-R Ab (TSH-R BAb), because they inhibit adenylate cyclase stimulation by TSH on thyrocytes or nonthyroidal cells transfected with TSH-R complementary DNA. We prepared monoclonal TSH-R BAb and characterized them. Peripheral lymphocytes from three patients with primary hypothyroidism and potent TSH-R BAb were transformed by Epstein-Barr virus, and the culture supernatants were screened by TSH binding inhibitor immunoglobulin (TBII) assay. Twenty positive and 7 negative lymphocyte clones were obtained; their monoclonality was confirmed by Southern blot analysis, using an immunoglobulin (Ig) JH probe. These monoclonal antibodies were then tested for TSH-R BAb activity. TSH-R BAb activity ranged from 24.1-58.5% (normal range, < 24%) in all 20 TBII-positive clones and in 2 of 7 TBII-negative clones. An enzyme-linked immunosorbent assay showed that the Ig isotypes of these clones with TBII and/or TSH-R BAb activity were IgG in 8 and IgM in 14. Another enzyme-linked immunosorbent assay and Southern blot analysis of the light chains revealed that 13 clones had kappa-chains, whereas the light chains could not be determined in the other 9 clones. To summarize, 1) we obtained 22 clones that produced monoclonal TSH-R BAb, including 8 IgG-type clones. 2) The clones exhibited dominant usage of the kappa-chain. 3) Although all TBII clones had TSH-R BAb activity, their TBII and TSH-R BAb activities were not significantly correlated, and two TSH-R BAb clones did not show TBII activity.

[1]  T. Akamizu,et al.  Recent progress in TSH receptor studies with a new concept of "autoimmune TSH receptor disease". , 1994, Endocrine journal.

[2]  B. Prabhakar,et al.  Dual mechanism of perturbation of thyrotropin-mediated activation of thyroid cells by antibodies to the thyrotropin receptor (TSHR) and TSHR-derived peptides. , 1993, The Journal of clinical endocrinology and metabolism.

[3]  M. Pauza,et al.  Unusual patterns of immunoglobulin gene rearrangement and expression during human B cell ontogeny: human B cells can simultaneously express cell surface kappa and lambda light chains , 1993, The Journal of experimental medicine.

[4]  M. Ohmori,et al.  Immunization with human thyrotrophin receptor peptide induces an increase in thyroid hormone in rabbits. , 1992, The Journal of endocrinology.

[5]  B. Rapoport,et al.  Thyroid stimulatory autoantibodies in different patients with autoimmune thyroid disease do not all recognize the same components of the human thyrotropin receptor: selective role of receptor amino acids Ser25-Glu30. , 1992, The Journal of clinical endocrinology and metabolism.

[6]  M. Atassi,et al.  Biological activities of rabbit antibodies against synthetic human thyrotropin receptor peptides representing thyrotropin binding regions. , 1992, Biochemical and biophysical research communications.

[7]  T. Akamizu,et al.  Identification of separate determinants on the thyrotropin receptor reactive with Graves' thyroid-stimulating antibodies and with thyroid-stimulating blocking antibodies in idiopathic myxedema: these determinants have no homologous sequence on gonadotropin receptors. , 1992, Molecular endocrinology.

[8]  T. Akamizu,et al.  Site-directed mutagenesis of a portion of the extracellular domain of the rat thyrotropin receptor important in autoimmune thyroid disease and nonhomologous with gonadotropin receptors. Relationship of functional and immunogenic domains. , 1991, The Journal of biological chemistry.

[9]  M. Béné,et al.  Alternative rearrangements of immunoglobulin light chain genes in human leukemia. , 1991, Leukemia.

[10]  M. Ohmori,et al.  Role of N-terminal region of the thyrotropin (TSH) receptor in signal transduction for TSH or thyroid stimulating antibody. , 1991, Biochemical and biophysical research communications.

[11]  H. Imura,et al.  A synthetic oligopeptide derived from human thyrotropin receptor sequence binds to Graves' immunoglobulin and inhibits thyroid stimulating antibody activity but lacks interactions with TSH. , 1991, Biochemical and biophysical research communications.

[12]  M. Ohmori,et al.  Thyroid stimulating activity of rabbit antibodies toward the human thyrotropin receptor peptide. , 1991, Biochemical and biophysical research communications.

[13]  T. Akamizu,et al.  Increases in cytosolic Ca++ down regulate thyrotropin receptor gene expression by a mechanism different from the cAMP signal. , 1991, Biochemical and biophysical research communications.

[14]  H. Imura,et al.  Possible binding site of thyrotropin binding inhibitor immunoglobulin (TBII) on the thyrotropin (TSH) receptor, which is different from TSH binding site. , 1990, Biochemical and biophysical research communications.

[15]  C. Kozak,et al.  Cloning, chromosomal assignment, and regulation of the rat thyrotropin receptor: expression of the gene is regulated by thyrotropin, agents that increase cAMP levels, and thyroid autoantibodies. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[16]  Y. Ichikawa,et al.  Monoclonal antibodies to the thyrotropin receptor bind to a 56-kDa subunit of the thyrotropin receptor and show heterogeneous bioactivities. , 1988, The Journal of biological chemistry.

[17]  K. Burman,et al.  Development of a human monoclonal antibody from a Graves' disease patient that identifies a novel thyroid membrane antigen. , 1988, Journal of immunology.

[18]  Thomas L. Rothstein,et al.  The role of clonal selection and somatic mutation in autoimmunity , 1987, Nature.

[19]  R. Smallridge,et al.  Partial characterization and clinical correlation of circulating human immunoglobulins directed against thyrotrophin binding sites in guinea pig fat cell membranes. Development of a direct enzyme immunoassay. , 1983, The Journal of clinical investigation.

[20]  J. Konishi,et al.  Inhibition of thyrotropin-induced adenosine 3'5'-monophosphate increase by immunoglobulins from patients with primary myxedema. , 1983, The Journal of clinical endocrinology and metabolism.

[21]  P. Laccetti,et al.  Multicomponent structure of the thyrotropin receptor: relationship to Graves' disease. , 1983, Life sciences.

[22]  P. Vitti,et al.  Monoclonal antibodies to the thyrotropin receptor: stimulating and blocking antibodies derived from the lymphocytes of patients with Graves disease. , 1982, Proceedings of the National Academy of Sciences of the United States of America.

[23]  G. Miller,et al.  Release of infectious Epstein-Barr virus by transformed marmoset leukocytes. , 1973, Proceedings of the National Academy of Sciences of the United States of America.

[24]  J. Fierer,et al.  Immunoglobulins (E,M,G) and complement in the connective tissues of the thyroid in Graves's disease. , 1972, The New England journal of medicine.

[25]  T. Tomasi.  Human immunoglobulin A. , 1968, The New England journal of medicine.