| Research Abstract |
Monoclonal antibodies were established by immunizing mice with a synthetic peptide (P354-14) corresponding to "the immunogenic peptide" of human thyrotropin receptor (hTSH-R). Obtained antibodies showed various biological activities, and gave an evidence of B-cell epitope spreading. These antibodies were radiolabeled and used for binding assay to hTSH-R-expressed CHO cell. To this system, many sera from patients with Graves' disease showed inhibitory action. But the clinical features did not correlate with their inhibitory profiles. Simultaneously, the spleen cells were subjected to antigen-specific ELISPOT assay in co-culture with various synthetic peptides of THS-R.Used cytokines were Interferon gamma (IFN-gamma ) for Th1, and Interleukin 4 (IL-4) for Th2. Results indicated that immunization with C-terminal TSH-R-specific insert peptide causes fluctuation in the Th2 cell population but not Th1 cells in the autoimmune system against the TSH-R, and the recognition repertoire of Th2 was expanded by the peptide. This supported the results of former B-cell epitope spreading at the level of helper T cells. Then, we established stable antigen-specific assay system of ELISPOT to measure activated helper T cells at their subclass levels. We applied this assay to examine the activation of T cell from Graves' patients. Basal activity of Th2 cells and their responsibility to TSH-R were increased in Graves' patients than healthy controls. No difference of Th1 activity was observed in Graves' patients. In conclusion, antigen-specific ELISPOT assay using synthetic peptides corresponding to human TSH-R is very useful to diagnose and research for autoimmune thyroid disease (Graves' disease).
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