for the Japan Thyroid Association Background: Thyroid storm (TS) is life threatening. Its incidence is poorly defined, few series are available, and population-based diagnostic criteria have not been established. We surveyed TS in Japan, defined its characteristics, and formulated diagnostic criteria, FINAL-CRITERIA1 and FINAL-CRITERIA2, for two grades of TS, TS1, and TS2 respectively. Methods: We first developed diagnostic criteria based on 99 patients in the literature and 7 of our patients (LIT-CRITERIA1 for TS1 and LIT-CRITERIA2 for TS2). Thyrotoxicosis was a prerequisite for TS1 and TS2 as well as for combinations of the central nervous system manifestations, fever, tachycardia, congestive heart failure (CHF), and gastrointestinal (GI)/hepatic disturbances. We then conducted initial and follow-up surveys from 2004 through 2008, targeting all hospitals in Japan, with an eight-layered random extraction selection process to obtain and verify information on patients who met LIT-CRITERIA1 and LIT-CRITERIA2. Results: We identified 282 patients with TS1 and 74 patients with TS2. Based on these data and information from the Ministry of Health, Labor, and Welfare of Japan, we estimated the incidence of TS in hospitalized patients in Japan to be 0.20 per 100,000 per year. Serum-free thyroxine and free triiodothyroine concentrations were similar among patients with TS in the literature, Japanese patients with TS1 or TS2, and a group of patients with thyrotoxicosis without TS (Tox-NoTS). The mortality rate was 11.0% in TS1, 9.5% in TS2, and 0% in Tox-NoTS patients. Multiple organ failure was the most common cause of death in TS1 and TS2, followed by CHF, respiratory failure, arrhythmia, disseminated intravascular coagulation, GI perforation, hypoxic brain syndrome, and sepsis. Glasgow Coma Scale results and blood urea nitrogen (BUN) were associated with irreversible damages in 22 survivors. The only change in our final diagnostic criteria for TS as compared with our initial criteria related to serum bilirubin concentration > 3 mg/dL. Conclusions: TS is still a life-threatening disorder with more than 10% mortality in Japan. We present newly formulated diagnostic criteria for TS and clarify its clinical features, prognosis, and incidence based on nationwide surveys in Japan. This information will help diagnose TS and in understanding the factors contributing to mortality and irreversible complications. 1 The First Department of Medicine, Wakayama Medical University, Wakayama, Japan. 2 Department of Medicine and Molecular Science, Gunma University Graduate School of Medicine, Maebashi, Japan. 3 Department of Medicine 2, Tokyo Women's Medical University, Tokyo, Japan. 4 Division of Endocrinology and Metabolism, Department of Internal Medicine, Fujita Health University, Toyoake, Japan. 5 Department of Internal Medicine, Keio University, Tokyo, Japan. 6 Department of Clinical and Molecular Endocrinology, Graduate School, Tokyo Medical and Dental University, Tokyo, Japan. 7 Division of Diabetes, Metabolism, and Endocrinolog...

Activation of T cells requires at least two signals transduced by the Ag-specific TCR and a costimulatory ligand such as CD28. CTLA-4, expressed on activated T cells, binds to B7 present on APCs and functions as a negative regulator of T cell activation. Our laboratory previously reported the association of Graves’ disease (GD) with a specific CTLA-4 gene polymorphism. In theory, reduced expression or function of CTLA-4 might augment autoimmunity. In the present study, we categorized autoimmune thyroid disease patients and normal controls (NC) by genotyping a CTLA-4 exon 1 polymorphism and investigated the function of CTLA-4 in all subjects. PBMCs and DNA were prepared from GD (n = 45), Hashimoto’s thyroiditis (HT) (n = 18), and NC (n = 43). There were more GD patients with the G/G or A/G alleles (82.2% vs 65.1% in NC), and significantly fewer patients with the A/A allele (17.8% vs 34.9% in NC). In the presence of soluble blocking anti-human CTLA-4 mAb, T cell proliferation following incubation with allogeneic EBV-transformed B cells was augmented in a dose-dependent manner. Augmentation induced by CTLA-4 mAb was similar in GD and NC (GD, HT, NC = 156%, 164%, 175%, respectively). We related CTLA-4 polymorphism to mAb augmentation of T cell proliferation in each subgroup (GD, HT, NC). Although PBMC from individuals with the G/G alleles showed 132% augmentation, those with the A/A alleles showed 193% augmentation (p = 0.019). CTLA-4 polymorphism affects the inhibitory function of CTLA-4. The G allele is associated with reduced control of T cell proliferation and thus contributes to the pathogenesis of GD and presumably of other autoimmune diseases.

Graves' disease is an autoimmune disease believed to be caused by a combination of environmental and genetic factors. One of the candidate genes is CTLA-4, a negative regulator of T cell activation. Three polymorphisms of the gene have been described, in the promoter at position -318, at position 49 in exon 1, and an (AT)n repeat within the 3'-untranslated region of exon 4. Many studies describe the association between a polymorphism of the CTLA-4 gene and autoimmune disease. To investigate the association of these CTLA-4 gene polymorphisms with each other, we analyzed the combined frequencies of each polymorphism and calculated the disequilibrium coefficients. We studied DNA samples from 120 Graves' disease (GD) patients and 80 healthy donors (NC). The exon 1 position 49 A/G polymorphism and promoter polymorphism at position -318, were typed using a PCR-restriction fragment length polymorphism method (PCR-RFLP). The polymorphic (AT)n repeat in exon 4 was determined by PCR amplification of genomic DNA, resolution of the amplified products on sequencing gels, and detection by autoradiography. There was a significant difference between GD and NC patients and occurrence of the polymorphism in exon 1 and exon 3, but not for the polymorphism in the promoter region. Furthermore, we found that the genotype with both the G allele in exon 1 and the 106 bp allele of the AT repeat in exon 4 occurred with much higher frequency in GD than NC (p<0.01), and that these polymorphisms are in linkage disequilibrium with each other. These results support the concept that CTLA-4 plays a critical role in the autoimmune process in GD, and that GD depends on multiple genetic susceptibility factors. Because the exon 1 and exon 4 polymorphisms are in strong linkage disequilibrium. It is not possible at this time to determine their unique relation to CTLA-4 function. Studies relating each polymorphism to CTLA4 function are required to determine whether one, or both, polymorphism(s) promote autoimmune disease.

Objective: Programmed cell death-1 (PD-1) and its ligands (PD-L1 and PD-L2) inhibit T-cell proliferation and activation. This inhibition down-regulates the immune responses. The association of a PD-L1 polymorphism with Graves' disease (GD) was studied. Design: The association of an A/C polymorphism at position 8923 in PD-L1 intron 4 with GD was studied. Patients: The study included 327 GD patients and 192 controls, of which 252 GD patients were followed over 5-10 years. Measurements: PD-L1 intron 4 position 8923 A/C polymorphism was typed using the PCR-restriction fragment length polymorphism method. Results: The A/C genotype frequencies were significantly different between GD patients and controls. The A/C and C/C frequencies were higher in GD patients than in controls. The A/A frequencies were lower in GD patients than in controls. C-allele frequency was higher in GD patients than in controls. A total of 252 GD patients were followed over 5-10 years; 200 had discontinued antithyroid drugs (ATD) while 52 continued to take ATD. Of these 200, 176 continued to be in remission and 24 had relapsed into hyperthyroidism. Significant differences in the duration of positive TBII, positive thyroid-stimulating antibodies, and ATD treatment were noted between the patients in remission and those that had relapsed. Significant differences in the A-and C-allele frequencies were noted between the two. The C-allele frequency was higher in GD patients who did not achieve remission than in those who achieved remission.

Cyclic adenosine monophosphate (cAMP) production during a 5-hour incubation using porcine thyroid cells (PTC) was stimulated significantly more by polyethylene glycol (PEG) 22.5% precipitated fractions (ppt frs) than by PEG 12.5% ppt frs from almost all Graves' sera. However, the thyrotropin (TSH) binding inhibition (TBI) activities of the PEG 12.5% and 22.5% ppt frs using porcine thyroid membranes were similar, and did not change in the 5-hour incubation. When the PEG 12.5% ppt fr from Graves' serum and the PEG 22.5% ppt fr from normal human serum (NHS) were coincubated, cAMP production was also stimulated as much as by the PEG 22.5% ppt fr from Graves' serum. When purified thyroid stimulating antibody (TSAb)-immunoglobulin G (IgG) and the PEG 22.5% ppt fr from NHS were coincubated, increased cAMP production was also observed, whereas bovine thyrotropin (bTSH) did not produce this effect. When purified TSAb-IgG and PEG solutions were coincubated, maximum increases in cAMP production (approximately 10-fold) with 5% PEG were found, whereas no increase was observed using bTSH. The stimulatory effect of high PEG concentrations on thyroid stimulating activity was observed by TSAb-IgG in salt-free or salt-containing medium (<0.15 mol/L NaCl concentration) but not by either TSAb-IgG conjugated to protein A-sepharose 4B or the inactivated TSAb-IgG by the treatment of 70 degrees C for 10 minutes. No stimulatory action by PEG was found with the thyroid stimulating substances such as GTPgammaS, forskolin, or pituitary adenylate-cyclase activating polypeptide (PACAP). The increased thyroid stimulating activity of Graves (IgG) at high PEG concentrations suggests the existence of some factors influencing the ability of TSAb to stimulate thyroid cells, although the exact mechanism remains to be clarified.

Graves' disease (GD) is thought to be an autoimmune disease with a strong genetic component. Candidate genes include human leukocyte antigen (HLA) class II genes and CTLA-4. The CTLA-4 gene has a variable length AT-repeat polymorphism in the 3'-untranslated region. We previously found that the AT-repeat of 104 bp or longer was associated with GD. In this study, we categorized patients with GD and normal controls (NC) by genotyping the CTLA-4 AT-repeat and investigated the function of CTLA-4. Peripheral blood mononuclear cells (PBMC) and DNA were prepared from adult Caucasians (NC = 34, GD = 37). Genotypes of the AT-repeat polymorphism were divided into three groups according to their alleles. We related the CTLA-4 polymorphism in each genotype to augmentation of T-cell proliferation induced by a soluble anti-CTLA-4 antibody during incubation with irradiated Epstein-Barr virus (EBV)-transformed B cells. Proliferation of T cells from subjects with the 86/86 bp (shorter) allele was less than T cells from patients with longer alleles. The length of the AT-repeat allele correlated inversely with augmentation of proliferation after CTLA-4 blockade in subjects with GD. The CTLA-4 AT-repeat polymorphism affects the inhibitory function of CTLA-4. The long AT-repeat allele is associated with reduced control of T-cell proliferation and thus contributes to the pathogenesis of GD.

Cytotoxic T-lymphocyte antigen-4 (CTLA-4) decreases the immune response of T cells by inactivating the signal that occurs with interaction between CD28 on T cells and B7 on antigen-presenting cells. Gene polymorphisms involving CTLA-4 promoter (-318 C/T), exon 1 (49 A/G), and exon 4 (microsatellite (AT)n) have been linked to Hashimoto's thyroiditis (HT) and other autoimmune diseases. HT also has a reported association with human T-cell lymphotrophic virus-1 (HTLV-1) infection. We investigated the occurrence of CTLA-4 polymorphisms in Japanese patients with HT with and without anti-HTLV-1 antibodies (HTLV-1 Ab). DNA samples from 143 patients with HT and 199 controls were subjected to polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis using the restriction enzymes, Bbv 1, Tse 1, and Mse 1. In the HTLV-1 Ab-positive group the exon 1 G allele was more frequent in patients with HT than in controls (67% vs. 53%, p = 0.0377), and in HTLV-1 Ab-negative group it was also frequent in patients with HT than in controls (68% vs. 53%, p = 0.0041). Frequency of the G allele in HT with HTLV-1 Ab was comparable to those without HTLV-1 Ab. Frequency of polymorphism in the promoter did not differ between patients with HT and controls, nor between controls with and without HTLV-1 Ab. HTLV-1 infection is not associated with CTLA-4 polymorphisms in either HT or controls. HTLV-1 infection is not regulated by genetic factor such as CTLA-4, and may affect occurrence of HT as an independent purely environmental factor.

It is important to choose the optimal treatment for DM in myotonic dystrophy, because the patients have hyperinsulinemia caused by specific mechanism and could not reduce the insulin resistance. Metformin improved hyperglycemia through increased insulin-independent glucose uptake in peripheral muscle. We believe metformin is the optimal agent for these patients.