Immunoglobulins of untreated Graves' patients with or without thyrotropin receptor antibody (determined by porcine thyrocytes) universally elicit potent thyroid hormone-releasing activity in cultured human thyroid follicles

Thyrotropin receptor antibody (TRAb), comprising thyrotropin binding inhibi tor immunoglobulin (TBII) and thyroid-stimulating antibody (TSAb), both of which are conventionally determined using porcine thyrocytes in Japan, is n ot always positive in patients with untreated Graves' disease. To elucidate whether immunoglobulin G (IgG) obtained from TBII/TSAb-positive (+) or neg ative (-) Graves' disease patients are responsible for hyperthyroidism, we investigated the thyroid hormone-releasing activity (THRA) of these IgGs in human thyroid follicles in suspension culture, in which bovine thyrotropin (bTSH) is detectable even at 0.1 mu U/mL. Human thyroid follicles, obtaine d from Graves' disease patients by subtotal thyroidectomy, were cultured in serum-free F-12/RPMI-1640 medium supplemented with bTSH or purified Graves ' IgGs. After preculturing for 3 days, I-125 was added, and after an additi onal 3 days of culture, 125I incorporated into the thyroid follicles and or ganic I-125 released into the culture medium (mainly I-125 - T-4 + I-125 - T-3) were counted. Seventy TBII(+)/TSAb(+)-, 3 TBII(+)/TSAb(-)-, and 3 TBII(-)/TSAb(+)- patien ts with untreated Graves' disease were all positive for THRA, which became undetectable in spontaneous remission obtained after several years of medic al treatment. The THRA was equivalent to 0.8-230 mu U/mL bTSH. Furthermore, 2 TBII(-)/TSAb(-) patients were significantly positive for THRA. This TBII (-)/TsAb(-)IgG stimulated human thyrocytes to produce cyclic adenosine mono phosphate (cAMP), and this was partially inhibited by antihuman IgG antibod y. The THRA induced by TBII(+)/TSAb(+) IgGs as well as TBII(-)/TsAb(-) IgG was inhibited by blocking-type TRAb obtained from TBII(+) patients with myx edema. There was a significant correlation between THRA and TSAb. These in vitro findings suggest that all IgGs obtained from untreated Graves' patien ts (n = 78) elicit potent THRA in human thyroid follicles in suspension cul ture. Because the TBII(-)/TSAb(-) IgGs can stimulate cAMP production in hum an but not in porcine thyrocytes, they probably recognize epitope(s) of TSH -binding sites specific to the human thyrotropin (hTSH) receptor. Furthermo re, we have demonstrated that the thyroid gland of hyperthyroid Graves' pat ients is stimulated by IgG(s) equivalent to at least 0.8 mu U/mL bTSH (abou t 5 mu U/mL hTSH) in vitro.