Multiple SNPs in intron 41 of thyroglobulin gene are associated with autoimmune thyroid disease in the Japanese population.

BACKGROUND: The etiology of the autoimmune thyroid diseases (AITDs), Graves' disease (GD) and Hashimoto's thyroiditis (HT), is largely unknown. However, genetic susceptibility is believed to play a major role. Two whole genome scans from Japan and from the US identified a locus on chromosome 8q24 that showed evidence for linkage with AITD and HT. Recent studies have demonstrated an association between thyroglobulin (Tg) polymorphisms and AITD in Caucasians, suggesting that Tg is a susceptibility gene on 8q24. OBJECTIVES: The objective of the study was to refine Tg association with AITD, by analyzing a panel of 25 SNPs across an extended 260 kb region of the Tg. METHODS: We studied 458 Japanese AITD patients (287 GD and 171 HT patients) and 221 matched Japanese control subjects in association studies. Case-control association studies were performed using 25 Tg single nucleotide polymorphisms (SNPs) chosen from a database of the Single Nucleotide Polymorphism Database (dbSNP). Haplotype analysis was undertaken using the computer program SNPAlyze version 7.0. PRINCIPAL FINDINGS AND CONCLUSIONS: In total, 5 SNPs revealed association with GD (P<0.05), with the strongest SNP associations at rs2256366 (P = 0.002) and rs2687836 (P = 0.0077), both located in intron 41 of the Tg gene. Because of the strong LD between these two strongest associated variants, we performed the haplotype analysis, and identified a major protective haplotype for GD (P = 0.001). These results suggested that the Tg gene is involved in susceptibility for GD and AITD in the Japanese.

Proteomic profiling of thyroid papillary carcinoma.

Papillary thyroid carcinoma (PTC) is the most common endocrine malignancy. We performed shotgun liquid chromatography (LC)/tandem mass spectrometry (MS/MS) analysis on pooled protein extracts from patients with PTC and compared the results with those from normal thyroid tissue validated by real-time (RT) PCR and immunohistochemistry (IHC). We detected 524 types of protein in PTC and 432 in normal thyroid gland. Among these proteins, 145 were specific to PTC and 53 were specific to normal thyroid gland. We have also identified two important new markers, nephronectin (NPNT) and malectin (MLEC). Reproducibility was confirmed with several known markers, but the one of two new candidate markers such as MLEC did not show large variations in expression levels. Furthermore, IHC confirmed the overexpression of both those markers in PTCs compared with normal surrounding tissues. Our protein data suggest that NPNT and MLEC could be a characteristic marker for PTC.

Association of the protein tyrosine phosphatase nonreceptor 22 haplotypes with autoimmune thyroid disease in the Japanese population.

BACKGROUND: A missence single-nucleotide polymorphism (SNP) in the protein tyrosine phosphatase nonreceptor 22 (PTPN22) gene known as R620W (rs2476601) was recently reported to be associated with several autoimmune diseases including Graves' disease (GD). The association was repeatedly confirmed in the populations of North European ancestry. However, this amino acid was reported to be nonpolymorphic in the Asian populations. Since the gene confers an impact on autoimmune diseases, we attempt to explore an association between the PTPN22 gene and autoimmune thyroid disease (AITD) in a Japanese population without restricting to rs2476601. Previous investigations have also demonstrated that two intronic SNPs (rs706778 and rs3118470) in the interleukin-2 receptor-alpha (IL2RA) gene were associated with type 1 diabetes in the Japanese population. PATIENTS AND METHODS: We genotyped the five SNPs (rs12760457, rs2797415, rs1310182, rs2476599, and rs3789604) of the PTPN22 and the two SNPs (rs706778 and rs3118470 in the IL2RA gene) in 456 Japanese patients with AITD (286 with GD, 170 with Hashimoto's thyroiditis) and 221 matched Japanese control subjects. Seven SNPs were analyzed by either the SNAPshot method or the high-resolution melting and unlabeled probe methods. Case-control association studies were performed using the chi(2) and Fisher's exact tests with Yates correction. Haplotype was conducted using the expectation-maximization algorithm. RESULTS: No association was found between any of the individual SNPs of the PTPN22 gene and AITD. Permutation analysis revealed that the distribution of one haplotype is significantly different between patients with AITD and controls (p = 0.0036). A novel protective effect of a haplotype containing five SNPs was observed (p < 0.0001 for AITD, p < 0.0001 for GD, and p < 0.0001 for Hashimoto's thyroiditis, respectively). The GG allele of rs3118470 in the IL2RA gene was significantly associated with GD (p = 0.03), although the association was weak. CONCLUSIONS: Significant difference in the distribution of the haplotype suggests that the PTPN22 gene rather than rs2476601 is involved in the development of AITD in the Japanese population.

Increased plasma urotensin-II and carotid atherosclerosis are associated with vascular dementia.

AIM: Human urotensin-II (UII) is a cyclic neuropeptide with potent vasoconstrictive activity in the vasculature. The expression of UII and its receptor (UT) mRNA is detected at high levels in the brain. We evaluated the relationship between plasma UII levels and vascular dementia (VaD) caused by stroke or atherosclerotic small vessel disease. METHODS: Carotid artery intima-media thickness (IMT), plaques, plasma levels of immunoreactive UII (IR-UII), and atherosclerotic biomarkers were determined in 42 patients with VaD, 197 with Alzheimer's disease (AD), and 47 non-demented elderly controls. RESULTS: Age, gender, body mass index, systolic blood pressure (SBP), fasting plasma glucose, insulin, triglycerides, high-density lipoprotein cholesterol, leptin, and plasminogen activator inhibitor-1 levels were not significantly different among these groups. IR-UII, low-density lipoprotein (LDL) cholesterol, lipoprotein(a), lipid peroxides, interleukin-6, and high-sensitive C-reactive protein (hs-CRP) levels, and maximum IMT were significantly higher in VaD than in AD patients or controls. IR-UII level showed a significantly positive correlation with SBP or maximum IMT. Multivariate logistic regression analysis revealed a significantly independent association between IR-UII levels or increased maximum IMT (> or =1.1 mm) and VaD as compared with SBP, LDL cholesterol, and interleukin-6 levels. CONCLUSION: Increased plasma IR-UII levels and carotid atherosclerosis may be involved in the pathogenesis and progression of VaD.

Association studies of the IL-23R gene in autoimmune thyroid disease in the Japanese population.

Autoimmune thyroid diseases (AITDs), including Graves' disease (GD) and Hashimoto's thyroiditis (HT), are caused by interplays of genetic factors and environmental triggers. Interleukin-23 and its receptor (IL-23R) guide T cells towards the Th17 phenotype. IL-23R single nucleotide polymorphisms (SNPs) have been shown to be associated with several autoimmune diseases, including Crohn's disease and rheumatoid arthritis, and Graves' ophthalmopathy (GO) in Caucasians. To determine whether variants in the IL-23R gene are associated with AITDs in Japanese, 464 Japanese AITD patients (290 with GD, 174 with HT) and 179 matched Japanese control subjects were genotyped for four SNPs spanning the IL-23R gene. SNPs rs11209026 and rs7530511 were genotyped using TaqMan allelic discrimination assays and SNPs rs2201841 and rs10889677 were genotyped using a fluorescent-based restriction fragment length polymorphism method. Case-control association studies were performed using the chi(2) and Fisher's exact tests with Yates correction. Of the four SNPs rs11209026 was non-polymorphic in our dataset. The other three SNPs were not associated with GD or GO or HT in our Japanese population. These results suggest that the IL-23R gene is associated with AITDs only in a specific ethnic group.

A fulminant case of renal vein thrombosis in a patient with autoimmune disorder and membranous nephropathy.

A previously healthy middle-aged woman noted a rapid onset of flank pain with gross hematuria. Enhanced CT scan showed thrombosis of the inferior vena cava and right renal vein. Laboratory findings revealed nephrotic proteinuria, Sjogren's syndrome (SjS), and Graves' disease (GD). A right nephrectomy was performed because of progressive and refractory renal necrosis. Renal specimens showed venous infarction with diffuse hemorrhagic and severe congestive renal necrosis, and membranous nephropathy (MN). The present case was diagnosed as acute renal necrosis due to catastrophic thrombosis in a patient with SjS, GD, and MN. It was thought that sudden development of thrombosis may have been caused by the status of the autoimmune disorders, and the associated MN.

A case of dual ectopic thyroid accompanied by positive antithyroid antibodies.

Ectopic thyroid tissue is an uncommon congenital aberration that is seldom present at two different sites simultaneously. The patient was a 32-year-old woman with dual ectopic thyroid accompanied by positive antithyroid antibodies. The simultaneous occurrence of dual ectopic thyroid and positive antithyroid antibodies has been documented in only two cases: the case discussed here and one previous case. The cervical ectopic thyroid was followed up by ultrasound, which showed an increase in the size of the lesion and an internal echo texture that became slightly heterogeneous after the patient had her second child. We speculated that these changes resulted from the changes in hormone demand brought on by pregnancy and parturition.

Cytotoxic T-lymphocyte associated antigen 4 gene polymorphisms and autoimmune thyroid disease: a meta-analysis.

CONTEXT: Cytotoxic T-lymphocyte associated antigen 4 (CTLA-4) polymorphisms have been widely examined for their associations with autoimmune thyroid diseases [Graves' disease (GD) and Hashimoto thyroiditis (HT)], but their relative population effect remains unclear. OBJECTIVE: The aim was to generate large-scale evidence on whether the CTLA-4 polymorphisms (A49G and CT60) and haplotypes thereof increase the susceptibility to GD and/or HT. DESIGN, SETTING, AND PARTICIPANTS: Meta-analyses of group-level data were reviewed from 32 (11,019 subjects) and 12 (4,479) published and unpublished studies for the association of the A49G polymorphism with GD and HT, respectively (PubMed and HuGeNet search until July 2006). There were 15 (n = 7246) and six (n = 3086) studies available for the CT60 polymorphism, respectively. Meta-analyses of individual-level data from 10 (4906 subjects) and five (2386) collaborating teams for GD and HT, respectively, were also reviewed. MAIN OUTCOME MEASURES: Association of gene variants and haplotypes with GD and HT was measured. RESULTS: Group-level data suggested significant associations with GD and HT for both A49G [odds ratios 1.49 (P = 6 x 10(-14)) and 1.29 (P = 0.001) per G allele, respectively] and CT60 [1.45 (P = 2 x 10(-9)) and 1.64 (P = 0.003) per G allele, respectively]. Results were consistent between Asian and Caucasian descent subjects. Individual-level data showed that compared with the AA haplotype, the risk conferred by the GG haplotype was 1.49 (95% confidence interval 1.31,1.70) and 1.36 (95% confidence interval 1.16,1.59) for GD and HT, respectively. Data were consistent with a dose-response effect for the G allele of CT60. CONCLUSION: The CT60 polymorphism of CTLA-4 maps an important genetic determinant for the risk of both GD and HT across diverse populations.

The regulatory T cell gene FOXP3 and genetic susceptibility to thyroid autoimmunity: an association analysis in Caucasian and Japanese cohorts.

FOXP3 is a key gene in the development of regulatory T cells (Treg). FOXP3 expression commits naïve T cells to become Treg cells. Indeed, mutations in the FOXP3 gene cause severe systemic autoimmune diseases in humans and in mice. Therefore, we hypothesized that the FOXP3 gene may be associated with thyroid autoimmunity which is among the typical autoimmune diseases that develop in individuals with FOXP3 mutations. Moreover, the FOXP3 gene is located within an X-chromosome locus (Xp11.23) previously shown to be linked with autoimmune thyroid diseases (AITD). We tested the FOXP3 gene locus for association with AITD in two large cohorts of US Caucasians and Japanese AITD patients. We analyzed 269 Caucasian AITD patients (52 males and 217 females) and 357 Caucasian controls (159 males and 198 females), as well as 377 female Japanese AITD patients and 179 female Japanese controls. The FOXP3 gene locus was analyzed using four microsatellite polymorphisms [(GT)n; (TC)n; DXS573; DXS1208] flanking the FOXP3 gene locus. Interestingly, while no association was found between FOXP3 polymorphisms and AITD in the Japanese cohort there was a significant association in the Caucasian cohort. There was a significant association of the (TC)n polymorphism with AITD in the Caucasian male AITD patients (p=0.011; 5 degrees of freedom [df]). Similarly, there was an association between the DXS573 microsatellite and AITD in the Caucasian female AITD patients (p=0.00023; 4 df). These results suggest that polymorphisms of the FOXP3 gene may play a role in the genetic susceptibility to AITD in Caucasians, perhaps by altering FOXP3 function and/or expression.

Impact of increased plasma serotonin levels and carotid atherosclerosis on vascular dementia.

Serotonin (5-HT), a potent vasoconstrictor in the large cerebral arteries, is considered to play a key role in atherothrombosis and to be implicated in ischemic cerebrovascular events followed by delayed neuronal death. The present study aims at evaluating the relationship between plasma levels of 5-HT and vascular dementia (VaD) caused by stroke or atherosclerotic small vessel disease. Carotid artery intima-media thickness (IMT), plaques, plasma 5-HT levels and atherosclerotic parameters were determined in 20 patients with VaD and 40 age-matched controls. Age, gender, body mass index, systolic and diastolic blood pressure, fasting plasma glucose levels and serum levels of insulin, triglycerides, high-density lipoprotein cholesterol, leptin, adiponectin and interleukin-6 and plasma levels of plasminogen activator inhibitor-1 were not significantly different between the two groups. Serum levels of insulin-like growth factor-1 (IGF-1) were significantly lower in VaD patients than in controls. Plasma 5-HT levels, serum levels of hepatocyte growth factor (HGF), low-density lipoprotein (LDL) cholesterol and high-sensitive C-reactive protein (hs-CRP), max IMT and plaque frequency were significantly greater in VaD patients than in controls. There was a significant positive correlation of max IMT with 5-HT or HGF levels. Multiple logistic regression analysis revealed that increased plasma levels of 5-HT and carotid plaque prevalence had significantly independent association with VaD as compared with serum levels of IGF-1, HGF, LDL cholesterol and hs-CRP. These results suggest that increased plasma levels of 5-HT and carotid atherosclerotic plaques may be involved in the pathogenesis and progression of VaD.

Association of a C/T single-nucleotide polymorphism in the 5' untranslated region of the CD40 gene with Graves' disease in Japanese.

CONTEXT: Graves' disease (GD) is caused by an interplay of genetic factors and environmental triggers. Recently, a susceptibility locus for GD was mapped to chromosome 20q11 (GD-2). Furthermore, a novel single nucleotide polymorphism (SNP) in the CD40 gene, which is located in 20q11, was found to be associated and linked with GD in Caucasians and in Koreans. OBJECTIVES: To examine a C/T SNP in the 5' untranslated region of the CD40 gene (CD40-E1SNP) for association with autoimmune thyroid diseases (AITDs) in a Japanese dataset. DESIGN, SETTING, AND PATIENTS: Case-control association studies were performed using the CD40-E1SNP. We studied 485 Japanese patients with AITD (301 with GD, 184 with Hashimoto's thyroiditis [HT]) and 177 matched Japanese control subjects in association studies. MAIN OUTCOME: Frequencies of genotypes and alleles of the CD40- E1SNP. RESULTS: The distribution of genotype frequencies differed significantly between patients with GD and controls in a dominant manner (p = 0.039). The CC+CT genotypes of the CD40-E1SNP were associated with the increased risk for GD (p = 0.015, odds ratio [OR] = 1.9). In contrast, no differences in genotype frequencies were observed between HT patients and controls for the CD40-E1SNP. CONCLUSION: These results suggested that the CD40 gene is involved in susceptibility for GD in the Japanese.

The codon 620 single nucleotide polymorphism of the protein tyrosine phosphatase-22 gene does not contribute to autoimmune thyroid disease susceptibility in the Japanese.

The etiology of the autoimmune thyroid diseases (AITDs), Graves' disease (GD), and Hashimoto's thyroiditis (HT) is largely unknown. However, genetic susceptibility is believed to play a major role. The lymphoid tyrosine phosphatase (LYP), encoded by the protein tyrosine phosphatase-22 (PTPN22) gene, is a powerful inhibitor of T cell activation. Recently, a single-nucleotide polymorphism (SNP), encoding a functional arginine to tryptophan residue change at PTPN22 codon 620 in Caucasians has been shown to be associated with GD and other autoimmune diseases. We have used a polymerase chain reaction (PCR)-restriction fragment (XcmI) assay to examine genotypes at the codon 620 polymorphism in 334 unrelated patients with AITD and 179 controls. None of the patients with AITD and controls had the tryptophan allele. These data suggest that the codon 620 polymorphism of the PTPN22 gene does not have a causal role for AITD in the Japanese. However, we cannot exclude the PTPN22 region as harboring another susceptibility locus for AITD in linkage disequilibrium with the Trp/Arg SNP.

Association of a CTLA-4 3' untranslated region (CT60) single nucleotide polymorphism with autoimmune thyroid disease in the Japanese population.

The etiology of the autoimmune thyroid diseases (AITDs), Graves' disease (GD) and Hashimoto's thyroiditis is largely unknown. However, genetic susceptibility is believed to play a major role. The cytotoxic T-lymphocyte antigen-4 (CTLA-4) gene, encoding a negative regulator of the T-lymphocyte immune response, had been reported to be associated and/or linked to AITD. Recently, AITD susceptibility in the Caucasians was mapped to the 6.1-kb 3'UTR of the CTLA-4 gene, in which the single-nucleotide polymorphism (SNP), CT60, was most strongly associated with AITD. In order to determine the association of the CTLA-4 gene with AITD in the Japanese, case-control association analysis for the CT60 of the CTLA-4 gene using 264 AITD patients and 179 healthy controls was done. The frequency of the disease-susceptible G allele of the CT60 of the Japanese control was higher than that of the Caucasians (72.6 vs. 52.3%). However, the G allele of the CT60 was associated with GD (84.0 vs. 72.6%, P=0.0008) and AITD (80.1 vs. 72.6%, P=0.009) in the Japanese. Furthermore, the G allele of the CT60 was associated with the increased risk for GD [P=0.004, odds ratio (OR)=2.0] and AITD (P=0.03,OR=1.6) in a recessive model. These results suggested that the CTLA-4 gene is involved in the susceptibility for GD and AITD in the Japanese.

Subtle 17alpha-hydroxylase/17,20-lyase deficiency with homozygous Y201N mutation in an infertile woman.

Steroid 17alpha-hydroxylase deficiency is characterized by failed sexual development and mineralocorticoid hypertension. Female patients usually exhibit primary amenorrhea. Some patients with partial deficiency are reported to have menses, yet they have hypertension and hypokalemia. We describe here a normotensive, infertile female patient with menses and minimal defects in secondary sex characteristics. The patient experienced menarche at age 13, and her menstrual cycles were regular until age 18 and irregular thereafter. Pubic hair was present (Tanner stage 3), and breast maturation was within normal range (Tanner stage 5). The patient's resting blood pressure was normal, and hypokalemia was not observed despite high blood corticosterone levels and reduced plasma renin activity. Analysis of the CYP17 gene revealed that the patient was homozygous for the Y201N mutation. In vitro expression of the mutated Y201N enzyme revealed reduced activities of both 17alpha-hydroxylase and 17,20-lyase; however, these reductions were less than those of the F53/54DEL mutation, which also shows mild clinical deficiency of 17alpha-hydroxylase/17,20-lyase. Thus, the 17alpha-hydroxylase/17,20-lyase deficiency in the present case is very mild both clinically and enzymatically. This case raises the possibility that there are infertile, menstruating women with undiagnosed 17alpha-hydroxylase deficiency.

Association of a thyroglobulin gene polymorphism with Hashimoto's thyroiditis in the Japanese population.

OBJECTIVE: The aetiology of the autoimmune thyroid diseases (AITDs), Graves' disease (GD) and Hashimoto's thyroiditis is largely unknown. However, genetic susceptibility is believed to play a major role. Two whole genome scans from Japan and from the USA identified a locus on chromosome 8q24 which showed evidence for linkage with AITD and HT. Recent studies have demonstrated an association between a Tg polymorphisms and AITD, suggesting that Tg is the susceptibility gene on 8q24. PATIENTS: We studied 308 Japanese AITD patients (194 GD and 114 HT patients) and 417 Japanese control subjects in association studies. DESIGN: Case-control association studies were performed using D8S284 and D8S272, microsatellite markers located in the 8q24 region, Tgms1 and Tgms2, microsatellite markers in introns 10 and 27, respectively, of Tg, and a SNP in exon 33 of Tg. RESULTS: No differences in allele frequencies were observed between AITD patients and controls for D8S284, D8S272 and Tgms1. Similarly, for Tgms2 and the exon 33 SNP no significant differences in allele frequency distribution were observed for all AITD patients. However, when analysing the HT patients alone we found a significant association between the 330 bp/352 bp genotype of Tgms2 and HT (HT = 16.7%, controls = 7.1%; corrected P-value = 0.01, OR = 2.6). CONCLUSION: Our results confirm the previous reports of an association between the Tg gene and AITD and suggest that Tg is an AITD susceptibility gene.

Antineutrophil cytoplasmic antibodies in patients with Graves' disease: association of antimyeloperoxidase autoantibodies with propylthiouracil therapy.

Abstract Antineutrophil cytoplasmic antibody (ANCA)-associated vasculitic disorders have been reported in patients with Graves' disease during propylthiouracil (PTU) therapy. To investigate whether ANCA are found in serum samples from patients with Graves' disease, and whether PTU therapy is associated with ANCA positivity, levels of serum ANCA were examined in Graves'-disease patients receiving either PTU (n = 49) or 1-methyl-2-mercaptoimidazole (methimazole, MMI) (n = 50), and in untreated Graves'-disease patients (n = 32) by enzyme-linked immunosorbent assay (ELISA). Serum samples from patients with Hashimoto's thyroiditis (n = 46) were also analyzed. Antimyeloperoxidase (MPO) autoantibodies (MPO-ANCA) were present in 10 (20.4%) of 49 Graves'-disease patients receiving PTU therapy, whereas MPO-ANCA were not detected in Graves'-disease patients receiving MMI, in untreated Graves'-disease patients, or in Hashimoto's thyroiditis patients. The MPO-ANCA-positive sera showed a perinuclear staining pattern which was detected by indirect immunofluorescence microscopy using a human polymorphonuclear leukocyte-cytospin preparation. Furthermore, Western blot analysis revealed that MPO-ANCA in the Graves'-disease patients, as well as MPO-ANCA in patients with idiopathic pauci-immune necrotizing and crescentic glomerulonephritis, recognize the 105-kD protein of native MPO. These results indicate that MPO-ANCA in Graves'-disease patients are strongly associated with PTU therapy, and not simply related to the autoimmune thyroid disease. This study also suggests that the presence of MPO-ANCA alone may not be sufficient for the development of vasculitic disorders.

Circulating nuclear matrix protein in Graves' disease.

The Fas/Fas ligand system induces apoptosis, while soluble Fas (sFas) blocks the system and soluble Fas ligand (sFasL) functions to induce apoptosis. The assay of nuclear matrix protein (NMP) released from dead or dying cells can be used to quantitate cell death. Therefore, we evaluated the relationship among serum levels of NMP, sFas, and sFasL in patients with Graves' disease. We measured serum levels of sFas, sFasL, NMP, thyroid hormones and TSH receptor antibody in 20 normal control subjects (5 men, 15 women; mean age, 44.3 years), 32 patients with untreated Graves' disease (4 men, 28 women; mean age, 44.1 years), and 10 patients with Graves' disease treated by methimazole (3 men, 7 women; mean age 39.2 years). Serum NMP was significantly lower (10.4 +/- 4.3 IU/ml, p < 0.02) in patients with untreated Graves' disease than in patients with treated Graves' disease (16.4 +/- 7.3 IU/ml) and control subjects (15.3 +/- 8.9 IU/ml). Serum sFas and sFasL were significantly higher in patients with untreated Graves' disease than in patients with treated Graves' disease and in control subjects. In the patient groups with Graves' disease, serum NMP was negatively correlated with sFas (r = -0.612, p < 0.001) and serum sFas was positively correlated with FT4 (r = 0.360, p < 0.05) and TRAb (r = 0.384, p < 0.05). Serum NMP was correlated with sFas. These results suggest that serum NMP is decreased in patients with untreated Graves' disease, and that cell death or apoptosis in patients with Graves' disease is affected by soluble Fas under the influence of thyroid function.

Accelerated production of nucleosome in cultured human mononuclear cells in untreated Graves' disease.

The apoptosis of lymphocytes, which occurs in autoimmune diseases, is usually induced by the Fas/Fas ligand system. As the assay of nucleosomes produced by apoptotic cells can be used to quantitate apoptosis, we evaluated nucleosome and soluble Fas ligand (sFasL) levels of cultured mononuclear cells to clarify the apoptosis of mononuclear cells in patients with autoimmune thyroid diseases by enzyme-linked immunosorbent assay. Nucleosome levels of cultured mononuclear cells in patients with untreated Graves' disease were significantly higher (3.27+/-2.90 U/ml) than those of control subjects (1.39+/-0.24 U/ml) and euthyroid patients with treated Graves' disease (1.53+/-0.33 U/ml). Nucleosome levels of cultured mononuclear cells were positively correlated with sFasL levels (r=0.544, p<0.01). It is therefore likely that increased sFasL levels elicit apoptosis of these cells in untreated Graves' disease.