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The predominant risk factor of metabolic syndrome is intra-abdominal fat accumulation, which is determined by waist circumference, waist-hip ratio measurements and visceral fat area (VFA); the latter can be accurately measured by performing computed tomography (CT). In addition to environmental factors, genetic factors play an important role in obesity and fat distribution. New genetic loci associated with body mass index (BMI) and adiposity have been identified by genome-wide association studies (GWASs). This study utilized CT to investigate whether single nucleotide polymorphisms (SNPs) that confer susceptibility to higher BMI are associated with VFA, subcutaneous fat area (SFA), and the ratio of VFA to SFA (V/S ratio). We measured the VFA and SFA of 1424 obese Japanese subjects (BMI ≥ 25 kg/m(2), 635 men and 789 women) who were genotyped for 13 single nucleotide polymorphisms (SNPs) reported by recent GWASs, namely, TNNI3K rs1514175, PTBP2 rs1555543, ADCY3 rs713586, IRS1 rs2943650, POC5 rs2112347, NUDT3 rs206936, LINGO2 rs10968576, STK33 rs4929949, MTIF3 rs4771122, SPRY2 rs534870, MAP2K5 rs2241423, QPCTL rs2287019, and ZC3H4 rs3810291. The G-allele of NUDT3 rs206936 was significantly associated with increased BMI (P = 5.3 × 10(-5)) and SFA (P = 0.00039) in the obese Japanese women. After adjustment with BMI, the association between rs206936 and SFA was not observed. This significant association was not observed in the men. The other SNPs analyzed were not significantly associated with BMI, VFA, SFA, or V/S ratio. Our results suggest that NUDT3 rs206936 is associated with BMI in Japanese women.

AIM Visceral fat accumulation plays an integral role in morbidity and mortality rates by increasing the risk of developing metabolic disorders such as type 2 diabetes, dyslipidemia, and hypertension. New genetic loci associated with fat distribution, measured by waist-hip ratios and computed tomography (CT), have recently been identified by genome-wide association studies in European-descent populations. This study used CT to investigate whether single nucleotide polymorphisms (SNPs) that confer susceptibility to fat distribution are associated with visceral fat area (VFA) and subcutaneous fat area (SFA) in the Japanese population. METHODS We measured the VFAs and SFAs of 1424 obese Japanese subjects (BMI≥25 kg/m(2), 635 men and 789 women) that were genotyped at 15 SNPs, namely, TBX15 rs984222, DNM3 rs1011731, LYPLAL1 rs4846567, GRB14 rs10195252, NISCH rs6784615, ADAMTS9 rs6795735, CPEB4 rs6861681, LY86 rs1294421, VEGFA rs6905288, RSPO3 rs9491696, NFE2L3 rs1055144, ITPR2 rs718314, HOXC13 rs1443512, ZNRF3 rs4823006 and THNSL2 rs1659258. RESULTS The G-allele of LYPLAL1 rs4846567 was borderline associated with an increased ratio of VFA to SFA (V/S ratio; p= 0.0020). LYPLAL1 rs4846567 had a stronger effect on the V/S ratio in women (p= 0.0078) than in men (p= 0.12); however, neither result was significant after Bonferroni correction for multiple comparisons. NISCH rs6784615 was nominally associated with increased VFA (p=0.040) and V/S ratio (p= 0.020). The other SNPs analyzed were not significantly associated with body mass index (BMI), VFA, or SFA. CONCLUSION Our results suggest that LYPLAL1 rs4846567 and NISCH rs6784615 may influence fat distribution in the Japanese population.

The predominant risk factor of metabolic syndrome is intra-abdominal fat accumulation, which is determined by waist circumference, waist-hip ratio measurements and visceral fat area (VFA); the latter can be accurately measured by performing computed tomography (CT). In addition to environmental factors, genetic factors play an important role in obesity and fat distribution. New genetic loci associated with body mass index (BMI) and adiposity have been identified by genome-wide association studies (GWASs). This study utilized CT to investigate whether single nucleotide polymorphisms (SNPs) that confer susceptibility to higher BMI are associated with VFA, subcutaneous fat area (SFA), and the ratio of VFA to SFA (V/S ratio). We measured the VFA and SFA of 1424 obese Japanese subjects (BMI ≥ 25 kg/m(2), 635 men and 789 women) who were genotyped for 13 single nucleotide polymorphisms (SNPs) reported by recent GWASs, namely, TNNI3K rs1514175, PTBP2 rs1555543, ADCY3 rs713586, IRS1 rs2943650, POC5 rs2112347, NUDT3 rs206936, LINGO2 rs10968576, STK33 rs4929949, MTIF3 rs4771122, SPRY2 rs534870, MAP2K5 rs2241423, QPCTL rs2287019, and ZC3H4 rs3810291. The G-allele of NUDT3 rs206936 was significantly associated with increased BMI (P = 5.3 × 10(-5)) and SFA (P = 0.00039) in the obese Japanese women. After adjustment with BMI, the association between rs206936 and SFA was not observed. This significant association was not observed in the men. The other SNPs analyzed were not significantly associated with BMI, VFA, SFA, or V/S ratio. Our results suggest that NUDT3 rs206936 is associated with BMI in Japanese women.

Visceral fat accumulation has an important role in increasing the morbidity and mortality rates, by increasing the risk of developing several metabolic disorders, such as type 2 diabetes, dyslipidemia and hypertension. New genetic loci that are associated with increased systolic and diastolic blood pressures have been identified by genome-wide association studies in Caucasian populations. This study investigates whether single nucleotide polymorphisms (SNPs) that confer susceptibility to high blood pressure are also associated with visceral fat obesity. We genotyped 1279 Japanese subjects (556 men and 723 women) who underwent computed tomography for measuring the visceral fat area (VFA) and subcutaneous fat area (SFA) at the following SNPs: FGF5 rs16998073, CACNB2 rs11014166, C10orf107 rs1530440, CYP17A1 rs1004467, NT5C2 rs11191548, PLEKHA7 rs381815, ATP2B1 rs2681472 and rs2681492, ARID3B rs6495112, CSK rs1378942, PLCD3 rs12946454, and ZNF652 rs16948048. In an additive model, risk alleles of the CYP17A1 rs1004467 and NT5C2 rs11191548 were found to be significantly associated with reduced SFA (P=0.00011 and 0.0016, respectively). When the analysis was performed separately in men and women, significant associations of rs1004467 (additive model) and rs11191548 (recessive model) with reduced VFA (P=0.0018 and 0.0022, respectively) and SFA (P=0.00039 and 0.00059, respectively) were observed in women, but not in men. Our results suggest that polymorphisms in the CYP17A1 and NT5C2 genes influence a reduction in both visceral and subcutaneous fat mass in Japanese women.

The predominant risk factor of metabolic syndrome is intra-abdominal fat accumulation, which is determined by waist circumference and waist-hip ratio measurements and visceral fat area (VFA) that is measured by computed tomography (CT). There is evidence that waist circumference and waist-hip ratio in the Caucasian population are associated with variations in several genes, including neurexin 3 (NRXN3), transcription factor AP-2β (TFAP2B), methionine sulfoxide reductase A (MSRA), lysophospholipase-like-1 (LYPLAL1), fat mass and obesity associated (FTO) and melanocortin 4 receptor (MC4R) genes. To investigate the relationship between VFA and subcutaneous fat area (SFA) and these genes in the recruited Japanese population, we genotyped 8 single-nucleotide polymorphisms (SNPs) in these 6 genes from 1228 subjects. Multiple regression analysis revealed that gender, age, and rs1558902 and rs1421085 genotypes (additive model) in FTO were significantly associated with body mass index (BMI; P=0.0039 and 0.0039, respectively), SFA (P=0.0027 and 0.0023, respectively) and VFA (P=0.045 and 0.040, respectively). However, SNPs in other genes, namely, NRXN3, TFAP2B, MSRA, LYPLAL1 and MC4R were not significantly associated with BMI, SFA or VFA. Our data suggest that some SNPs, which were identified in genome-wide studies in the Caucasians, also confer susceptibility to fat distribution in the Japanese subjects.

There is evidence that the obesity phenotype in the Caucasian populations is associated with variations in several genes, including neuronal growth regulator 1 (NEGR1), SEC16 homolog B (SCE16B), transmembrane protein 18 (TMEM18), ets variant 5 (ETV5), glucosamine-6-phosphate deaminase 2 (GNPDA2), prolactin (PRL), brain-derived neurotrophic factor (BDNF), mitochondrial carrier homolog 2 (MTCH2), Fas apoptotic inhibitory molecule 2 (FAIM2), SH2B adaptor protein 1 (SH2B1), v-maf musculoaponeurotic fibrosarcoma oncogene homolog (MAF), Niemann-Pick disease, type C1 (NPC1), melanocortin 4 receptor (MC4R) and potassium channel tetramerisation domain containing 15 (KCTD15). To investigate the relationship between obesity and these genes in the Japanese population, we genotyped 27 single-nucleotide polymorphisms (SNPs) in 14 genes from obese subjects (n=1129, body mass index (BMI) > or =30 kg m(-2)) and normal-weight control subjects (n=1736, BMI

Genetic factors are important in the development of metabolic syndrome. However, the genetic background of metabolic syndrome remains unclear. We screened polymorphisms in 85 obesity-related genes to determine which may be associated with metabolic syndrome. A total of 336 single-nucleotide polymorphisms (SNPs) in 85 genes selected from the JSNP database were genotyped. We conducted case-control association analyses using patients with metabolic syndrome (n=1080) and control individuals (n=528) who had no risk of the metabolic syndrome. Three SNPs in the McKusick-Kaufman syndrome (MKKS) gene were significantly related to metabolic syndrome by case-control association study; rs1545 (odds ratio (OR) adjusted for age and gender, 1.45; 95% confidence interval (CI), 1.21-1.74; P=0.000043 (additive model)); rs1547 (OR, 1.45; 95% CI, 1.21-1.74; P=0.000041); and rs2294901 (OR, 1.46; 95% CI, 1.22-1.75; P=0.000033). We selected five tag SNPs (rs2294901, rs221667, rs6133922, rs6077785 and rs6108572) in the MKKS gene. They were in one linkage disequilibrium (LD) block and rs6133922 (P=0.00042), rs6077785 (P=0.000013) and rs6108572 (P=0.000019) as well as rs2294901 were significantly associated with metabolic syndrome. TGAAA haplotype was protective against the metabolic syndrome (P=0.0074), and CCGTT haplotype was susceptible (P=0.00070) to the metabolic syndrome. Our data suggest that genetic variations at MKKS gene influence the risk of metabolic syndrome.

The single nucleotide polymorphism (SNP) rs7566605 in the upstream region of the insulin-induced gene 2 (INSIG2) is associated with the obesity phenotype in many Caucasian populations. In Japanese, this association with the obesity phenotype is not clear. To investigate the relationship between rs7566605 and obesity in Japanese, we genotyped rs7566605 from severely obese subjects [n = 908, body mass index (BMI) ≥ 30 kg/m2] and normal-weight control subjects (n = 1495, BMI < 25 kg/m2). A case–control association analysis revealed that rs7566605 was significantly associated with obesity in Japanese. The P value in the minor allele recessive mode was 0.00020, and the odds ratio (OR) adjusted for gender and age was 1.61 [95% confidential interval (CI) = 1.24–2.09]. Obesity-associated phenotypes, which included the level of BMI, plasma glucose, hemoglobin A1c, total cholesterol, triglycerides, high-density lipoprotein (HDL) cholesterol, and blood pressure, were not associated with the rs7566605 genotype. Thus, rs7566605 in the upstream region of the INSIG2 gene was found to be associated with obesity, i.e., severe obesity, in Japanese.

Context: Genetic factors are important for the development of obesity. However, the genetic background of obesity still remains unclear. Objective: Our objective was to search for obesity-related genes using a large number of gene-based single-nucleotide polymorphisms (SNPs). Design and Setting: We conducted case-control association analyses using 94 obese patients and 658 controls with 62,663 SNPs selected from the SNP database. SNPs that possessed P ≤ 0.02 were further analyzed using 796 obese and 711 control subjects. One SNP (rs3764220) in the secretogranin III (SCG3) gene showed the lowest P value (P = 0.0000019). We sequenced an approximately 300-kb genomic region around rs3764220 and discovered SNPs for haplotype analyses. SCG3 was the only gene within a haplotype block that contained rs3764220. The functions of SCG3 were studied. Patients: Obese subjects (body mass index ≥ 30 kg/m2, n = 890) and control subjects (general population; n = 658, body mass index ≤ 25kg/m2; n = 711) were recruited for this study. Results: Twelve SNPs in the SCG3 gene including rs3764220 were in almost complete linkage disequilibrium and significantly associated with an obesity phenotype. Two SNPs (rs16964465, rs16964476) affected the transcriptional activity of SCG3, and subjects with the minor allele seemed to be resistant to obesity (odds ratio, 9.23; 95% confidence interval, 2.77–30.80; χ2 = 19.2; P = 0.0000067). SCG3 mRNA and immunoreactivity were detected in the paraventricular nucleus, lateral hypothalamic area, and arcuate nucleus, and the protein coexisted with orexin, melanin-concentrating hormone, neuropeptide Y, and proopiomelanocortin. SCG3 formed a granule-like structure together with these neuropeptides. Conclusions: Genetic variations in the SCG3 gene may influence the risk of obesity through possible regulation of hypothalamic neuropeptide secretion.

Intestinal intraepithelial lymphocytes (i-IEL) expressing CD8α are located in the intestine and may confer protection against invasion of intestinal microflora. We found that mice rendered deficient in CD8α molecules by homologous recombination were susceptible to 5-fluorouracil (5-FU)-induced lethality accompanied by translocation of members of the enterobacteria. The number of i-IEL was greatly reduced on day 6 after 5-FU administration in both CD8α+/−mice and CD8α−/−mice, whereas the recovery of the level of i-IEL thereafter was significantly impaired in CD8α−/−mice compared with that in CD8α+/−mice. The ability of i-IEL to produce gamma interferon in response to immobilized T-cell receptor (TCR) αβ or TCR γδ monoclonal antibodies was significantly lower in CD8α−/−mice than in CD8α+/−mice. Transfer of CD8+i-IEL conferred significant protection against 5-FU-induced lethality in CD8α−/−mice. The results suggest that CD8+i-IEL play an important role in protection against 5-FU-induced lethality with translocation ofEnterobacteriaceae.

To better understand the pathogenesis of type 1 diabetes, we have developed pancreatic biopsy under laparoscope for recent-onset type 1 diabetic patients. The patients included 29 acute-onset type 1 diabetic patients, 5 latent-onset type 1 diabetic patients, and 1 type 2 diabetic patient. Their median age was 28 years, and the duration of diabetes at the time of biopsy was ∼3 months. In 31 of 35 patients, we could obtain the pancreas tissue by punching. No serious complications, such as heavy bleeding, peritonitis, or pancreatitis, have been experienced. Pneumoderma was observed in two patients, and abdominal dull pain had continued for 2 days in two patients. However, special treatment was not necessary for these complications. T-cell–predominant infiltration to islets (insulitis) and hyperexpression of major histocompatibility complex class I antigens on islet cells were the two major findings and were observed in 17 of 29 recent-onset type 1 diabetic patients. These findings could be regarded as evidence of immune attack against β-cells, and their presence was closely correlated with the presence of either anti-GAD or anti–IA-2 antibodies (P = 0.02). In conclusion, pancreatic biopsy under laparoscope is a safe procedure without serious complications, according to our findings, for detecting in situ autoimmune phenomenon in recent-onset type 1 diabetic patients.

Aims/hypothesis. Type I (insulin-dependent) diabetes results mainly from T-cell-mediated autoimmune destruction of pancreatic beta cells. Cytotoxic T lymphocytes destroy target cells via a perforin-based or Fas-based mechanism. Our previous study indicated that the Fas-Fas ligand (FasL) pathway is required for the development of autoimmune diabetes in the NOD mouse. We now investigated whether or not the Fas-FasL system is involved in the beta-cell destruction in human Type I diabetes. Methods. We immunohistochemically analysed pancreas biopsy specimens of 13 recent-onset patients. Results. Pancreatic islets were identified but showed various degrees of reduction in beta-cell volume in all patients. Out of 13 patients 6 had insulitis. In these 6 patients Fas was expressed in both the islets and infiltrating cells but not in either cell type in the 7 other patients without insulitis. Double immunostaining showed that Fas was positive in 92.2 to 97.7 % of beta cells but only in 17.6 to 46.7 % of alpha cells in Fas-positive, insulin-remaining islets. We found FasL was expressed exclusively in islet-infiltrating cells in patients with insulitis. Double immunostaining revealed that the most prevalent phenotype of FasL-positive cells was CD8, which was followed by macrophages and CD4. Conclusion/interpretation. The interaction between Fas on beta cells and FasL on infiltrating cells might trigger selective apoptotic beta-cell death in inflamed islets, leading to immune-mediated Type I diabetes. [Diabetologia (1999) 42: 1332–1340]

Aims/hypothesis. To determine whether the clinical heterogeneity observed in the development of Type I (insulin-dependent) diabetes mellitus correlates with immunohistochemical differences observed at diagnosis. Methods. Patients (n = 17) with recent-onset diabetes clinically considered to be insulin dependent (Type I), underwent pancreatic biopsy for immunohistological analysis. These patients were divided into two groups based on the presence or absence of islet immunological abnormalities (insulitis or hyperexpression of MHC class I antigens or both). The patients were also HLA typed and tested for islet cell antibodies and antibodies to glutamic acid decarboxylase (GAD-Ab). All patients were followed monthly for 2 years and their fasting plasma glucose, haemoglobin A1C and daily insulin doses were recorded. The clinical course of patients with islet immunological abnormalities was compared with that of patients without those abnormalities. Results. Patients with and without islet immunological abnormalities did not differ with regard to HLA type or islet cell antibodies. Antibodies to glutamic acid decarboxylase correlated with the presence of insulitis and MHC class I hyperexpression. These local immunological abnormalities were also associated with higher haemoglobin A1C values (p < 0.05) and a trend towards greater insulin requirements. Further, patients with the islet abnormalities had higher fasting plasma glucose concentrations 2 years after the biopsy than at the time of the biopsy (p < 0.05). Conclusion/interpretation. The heterogeneous clinical course observed following diagnosis in patients with Type I diabetes correlates with islet immunological abnormalities. Insulitis and hyperexpression of MHC class I correlate with deteriorating glycaemic control. [Diabetologia (1999) 42: 574–578]

(1999). Emerging therapeutic strategies in autoimmune diabetes: aetiology, prediction, prevention and cure. Emerging Therapeutic Targets: Vol. 3, No. 1, pp. 177-193.

To clarify the mechanism of regenerative processes of pancreatic β-cells, we constructed a new diabetic model of mice and investigated their pancreatic endocrine cells by electron microscopy. Male ICR mice (8 weeks old) were partially and chemically depancreatized by perfusing alloxan (100 mg/kg body weight) via the caudal vein after clamping the cranial mesenteric artery. By this method, we could render the mice diabetic by partial reduction of β-cells localized in the splenic, gastric, and parabiliary segment. Glucose intolerance gradually ameliorated without any treatment. In the perfused segments, pancreatic β-cells showed pyknosis and the mitochondria were swollen 6h after the treatment, while non-β-cells including α-cells remained intact. At 5 days, β-cells were few and the islets became smaller in size. At 20 weeks, small islet cell clusters (ICCs) were observed budding from interlobular and intralobular ductal epithelial cells. β-cells scattering in the exocrine pancreas were also frequently observed. In the alloxan-nonperfused segment, β-cells with thin rough endoplasmic reticulum and immature secretory granules without an electron-opaque halo were observed, and the number of mitochondria increased in some β-cells at 1 day and 5 days after the treatment. At 20 weeks, β-cells that contained only mature granules were observed in hypertrophic islets. In this model, both proliferation of residual β-cells and differentiation of pancreatic endocrine cells from the ductal epithelial cells were recognized.

To clarify the regeneration process of pancreatic beta-cells, we established a new mouse model of diabetes induced by selective perfusion of alloxan after clamping the superior mesenteric artery. In this model, diabetes could be induced by the destruction of beta-cells in alloxan-perfused segments, while beta-cells in nonperfused segments were spared. Intraperitoneal glucose tolerance tests showed glucose intolerance, which gradually ameliorated and was completely normalized in 1 year with a concomitant increase of insulin content in the pancreas. Histological examination showed neo-islet formation in the alloxan-perfused segment and the proliferation of spared beta-cells in the nonperfused segment. In the alloxan-perfused segment, despite a marked reduction of islets in size and number at an early stage, both the number of islets, including islet-like cell clusters (ICCs), and the relative islet area significantly increased at a later stage. Increased single beta-cells and ICCs were located in close contact with duct cell lining, suggesting that they differentiated from duct cells and that such extra-islet precursor cells may be important for beta-cell regeneration in beta-cell-depleted segment. In addition to beta-cells, some nonhormone cells in ICCs were positive for nuclear insulin promoter factor 1, which indicated that most, if not all, nonhormone cells positive for this factor were beta-cell precursors. In the nonperfused segment, the islet area increased significantly, and the highest 5-bromo-2-deoxyuridine-labeling index in beta-cells was observed at day 5, while the number of islets did not increase significantly. This indicated that the regeneration of islet endocrine cells occurs mostly through the proliferation of preexisting intra-islet beta-cells in the nonperfused segment. In conclusion, the regeneration process of beta-cells varied by circumstance. Our mouse model is useful for studying the mechanism of regeneration, since differentiation and proliferation could be analyzed separately in one pancreas.

Insulin-dependent diabetes mellitus (IDDM) is assumed to be a T cell–mediated autoimmune disease. To investigate the role of Fas-mediated cytotoxicity in pancreatic β cell destruction, we established nonobese diabetic (NOD)-lymphoproliferation (lpr)/lpr mice lacking Fas. Out of three genotypes, female NOD-+/+ and NOD-+/lpr developed spontaneous diabetes by the age of 10 mo with the incidence of 68 and 62%, respectively. In contrast, NOD-lpr/lpr did not develop diabetes or insulitis. To further explore the role of Fas, adoptive transfer experiments were performed. When splenocytes were transferred from diabetic NOD, male NOD-+/+ and NOD-+/lpr developed diabetes with the incidence of 89 and 83%, respectively, whereas NOD-lpr/lpr did not show glycosuria by 12 wk after transfer. Severe mononuclear cell infiltration was revealed in islets of NOD-+/+ and NOD-+/lpr, whereas islet morphology remained intact in NOD-lpr/lpr. These results suggest that Fas-mediated cytotoxicity is required to initiate β cell autoimmunity in NOD mice. Fas–Fas ligand system might be critical for autoimmune β cell destruction leading to IDDM.

We examined at autopsy 47 cases (22 males and 25 females) of insulin-dependent diabetes mellitus (IDDM) from 21 hospitals in Japan to clarify the pathological changes that occur in the pancreas vs. those in control patients. The mean age was 39.7 +/- 13.9 (mean +/- SD) years, and the duration of IDDM from clinical onset was 13.1 +/- 6.5 years. Causes of death included renal complications, infections, acute diabetic complications such as ketoacidosis or hyper- or hypoglycemic coma, and atherosclerotic disease. This study revealed noticeable decreases in the islet area and beta cell area, and a slight decreases in the alpha cell area and preservation of the number of islets. Insulitis was found in only 1 case, representing 25% of the cases with a duration of IDDM of one year or less. Lymphocytic infiltration of the exocrine gland was seen in 22 cases (46.8%). Predominant phenotypes of the lymphocytes were T lymphocytes and macrophages. Fibrosis, fatty change and atrophy were also found. Although this is not a strictly age- and sex-matched study, the high incidence of lymphocytic infiltration of the exocrine pancreas indicates that the exocrine tissue as well as beta cells is the target of immune reactions in Japanese patients with IDDM.

In order to clarify the nature of T lymphocytes infiltrating the pancreatic islets of patients with insulin-dependent diabetes mellitus (IDDM), we analysed T cell receptor (TCR) gene transcripts expressed in pancreatic biopsy specimens of patients with recent-onset IDDM. We also investigated the expression of cytokines (interferon-gamma: IFN-gamma; tumour necrosis factor-alpha: TNF-alpha; interleukin-4: IL-4; interleukin-6: IL-6) in the same specimens. The TCR V beta repertoire was not restricted either in the pancreas or the peripheral lymphocytes of IDDM patients. In contrast, the TCR V alpha repertoire was restricted in the pancreas, but not in the peripheral blood lymphocytes, of IDDM patients. The sequence analysis of the complementarity-determining region 3 (CDR3) of the TCR alpha revealed the presence of dominant clonality in alpha chains of T cells in the patients. IFN-gamma mRNA was highly expressed in the pancreas of IDDM patients, while IL-4 mRNA was deficient. A lower level of expression of IL-6 mRNA was detected in the IDDM pancreas than in the control tissue. These results indicate that T cells bearing a distinct TCR alpha chain are selectively retained and activated within the pancreas of recent-onset IDDM.

Insulin-dependent diabetes mellitus (IDDM) results mainly from T cell mediated pancreatic beta cell destruction. To fully activate antigen specific T cells, current evidence suggests that two signals are required. One signal is delivered via the antigen specific T cell receptor (TCR) when engaged by major histocompatibility complex presented antigen (MHC:Ag), the other via the T cell's CD28 when engaged by CD80/86. Recent studies have demonstrated that transgenic mice expressing CD80 on their pancreatic beta cells are susceptible to autoimmune beta cell destruction. To further explore whether CD80/86 expression plays a role in IDDM pathogenesis, we analysed pancreatic biopsy specimens from 16 recent-onset IDDM patients (13 men and 3 women; age 29.7 +/- 8.8 years) for CD80/86 expression. While no biopsy revealed any islet cell specific CD80 or CD86 expression, biopsies from six of the nine patients with insulitis revealed both CD80 and CD86 expression on the islet infiltrating cells. Triple immunofluorescent staining for CD80/86, CD3, and glucagon revealed that the CD80/86-positive cells were also CD3-positive. Of the CD3-positive cells, 19.4% expressed CD80 and 21.7% expressed CD86. CD80 and CD86-positive cells were similarly distributed throughout the inflamed islets. These data suggest that CD28 engagement with CD80/86 may play a pathogenic role in the beta cell destruction underlying IDDM.

Cytokines are thought to contribute to the induction of pancreatic beta-cell destruction in insulin-dependent diabetes mellitus. The molecular mechanisms that underlie beta-cell death were investigated by studying cytokine-induced cell death in beta-cell lines. A combination of three cytokines (interleukin-1 beta, tumour necrosis factor-alpha, and interferon-gamma) induced apoptotic cell death in the mouse pancreatic beta-cell line beta TC1, as judged from the appearance of cells with hypodiploid nuclei and oligonucleosomal DNA fragmentation. The same treatment also induced apoptosis in the mouse pancreatic alpha-cell line alpha TC1 and the NOD/Lt mouse beta-cell line NIT-1, although to a lesser extent than in beta TC1 cells. The abundance of endogenous Bcl-2 in beta TC1 cells was lower than that in the other two cell lines. Overexpression of human Bcl-2 in beta TC1 cells partially protected them from cytokine-induced cell death. These results suggest that apoptosis may be responsible, at least in part, for cytokine-induced beta-cell destruction and that Bcl-2 prevents apoptosis in pancreatic islet cells.

Viral infection is assumed to trigger or exacerbate autoimmune responses against pancreatic beta cells leading to the development of insulin-dependent diabetes mellitus (IDDM). We therefore examined by polymerase chain reaction the presence of two candidate viruses, cytomegalovirus and Epstein-Barr virus, in IDDM pancreases. Pancreas tissues were obtained by biopsy under laparoscopy from 16 recent-onset IDDM patients: age 17-53 years; disease duration 0-7 months; six had flu-like symptoms before onset. Frozen sections were made and subjected to DNA amplification. DNA samples were prepared from the frozen sections and polymerase chain reaction was performed using primers specific to cytomegalovirus, Epstein-Barr virus and control gene for HLA-DP. Cytomegalovirus- and Epstein-Barr virus-infected cells were used for positive control. Southern blot analysis could detect cytomegalovirus DNA from as few as 2 x 10(-1) cytomegalovirus-infected cells and Epstein-Barr virus DNA from two Epstein-Barr virus-infected cells. This highly sensitive analysis, however, could not detect cytomegalovirus or Epstein-Barr virus genomes in pancreases of recent-onset IDDM. A single copy human gene (HLA-DP) was amplified from all IDDM pancreases indicating that DNA amplification was performed without inhibition. We conclude that cytomegalovirus or Epstein-Barr virus genomes are unlikely to exist in pancreas biopsy specimens of recent-onset IDDM patients.

To elucidate the role played by interferon-alpha (IFN alpha) in the pathogenesis of autoimmune endocrine disease, we determined the autoantibody status, thyroid function test results, hemoglobin-A1c levels, and clinical symptoms of 58 patients who received IFN alpha for treatment of chronic active type C hepatitis. Each patient was treated for 6 months with a total dose of 391 +/- 140 x 10(6) U (mean +/- SD). Thyroid microsomal and/or thyroglobulin antibodies newly appeared or were increased in titer in 6 patients, 2 of whom developed hypothyroidism during IFN alpha therapy. Neither islet cell antibodies nor insulin-dependent diabetes mellitus developed during IFN alpha therapy, although hemoglobin-A1c levels were increased in 2 patients. One patient became positive for antimitochondrial antibodies, and another patient with preexisting antimitochondrial antibodies also manifested deterioration in liver function test results. Parietal cell antibodies and smooth muscle cell antibodies were the most frequent newly developed antibodies in 7 patients. Adrenal medullary cell antibodies and nuclear antibodies newly developed in 2 and 1 patients, respectively. At least 1 of 8 autoantibodies newly appeared in 19 patients (32.8%) and hypothyroidism developed in 2 patients (3.4%) during IFN alpha therapy. On the other hand, in 19 age- and sex-matched patients who did not receive IFN alpha, no autoantibody appeared, and no autoimmune disease developed during a follow-up period of 3 months. These findings suggest that IFN alpha acts as an immunomodulatory agent, inducing autoantibody production and the development of autoimmune disease in susceptible patients. Special attention should be paid to the development of hypothyroidism during IFN alpha therapy.

Visceral fat accumulation has an important role in the development of several metabolic disorders, such as type 2 diabetes, dyslipidemia and hypertension. New genetic loci that contribute to the development of type 2 diabetes have been identified by genome-wide association studies. To examine the association of type 2 diabetes susceptibility loci and visceral fat accumulation, we genotyped 1279 Japanese subjects (556 men and 723 women), who underwent computed tomography for measurements of visceral fat area (VFA) and subcutaneous fat area (SFA) for the following single-nucleotide polymorphisms (SNPs): NOTCH2 rs10923931, THADA rs7578597, PPARG rs1801282, ADAMTS9 rs4607103, IGF2BP2 rs1470579, VEGFA rs9472138, JAZF1 rs864745, CDKN2A/CDKN2B rs564398 and rs10811661, HHEX rs1111875 and rs5015480, TCF7L2 rs7901695, KCNQ1 rs2237892, KCNJ11 rs5215 and rs5219, EXT2 rs1113132, rs11037909, and rs3740878, MTNR1B rs10830963, DCD rs1153188, TSPAN8/LGR5 rs7961581, and FTO rs8050136 and rs9939609. None of the above SNPs were significantly associated with VFA. The FTO rs8050136 and rs9939609 risk alleles exhibited significant associations with body mass index (BMI; P=0.00088 and P=0.0010, respectively) and SFA (P=0.00013 and P=0.00017, respectively). No other SNPs were significantly associated with BMI or SFA. Our results suggest that two SNPs in the FTO gene are associated with subcutaneous fat accumulation. The contributions of other SNPs are inconclusive because of a limitation of the sample power.

We report a case of IDDM which occurred during interferon therapy for chronic hepatitis. A 31-year-old man intermittently received 2.5 x 10(8) units of alpha-IFN and 1 x 10(8) units of beta-IFN for treatment of chronic viral hepatitis type B. Four years after the beginning of IFN therapy, he acutely developed moderate hyperglycemia and severe ketonuria with positive islet cell antibody, and then 28 units/day of insulin injection was started. After the start of insulin therapy, there was a remission period for about 3 years but insulin-dependency recurred thereafter. The clinical course of this case indicates that IFN therapy precedes IDDM. During and after IFN therapy we should consider the possibility of occurrence of IDDM as well as other autoimmune diseases and observe the clinical course carefully.

Metabolic syndrome is defined as a cluster of multiple risk factors, including central obesity, dyslipidemia, hypertension and impaired glucose tolerance, that increase cardiovascular disease morbidity and mortality. Genetic factors are important in the development of metabolic syndrome, as are environmental factors. However, the genetic background of metabolic syndrome is not yet fully clarified. There is evidence that obesity and obesity-related phenotypes are associated with variations in several genes, including NEGR1, SEC16B, TMEM18, ETV5, GNPDA2, BDNF, MTCH2, SH2B1, FTO, MAF, MC4R, KCTD15, SCG3, MTMR9, TFAP2B, MSRA, LYPLAL1, GCKR and FADS1. To investigate the relationship between metabolic syndrome and variations in these genes in the Japanese population, we genotyped 33 single-nucleotide polymorphisms (SNPs) in 19 genes from 1096 patients with metabolic syndrome and 581 control individuals who had no risk factors for metabolic syndrome. Four SNPs in the FTO gene were significantly related to metabolic syndrome: rs9939609 (P=0.00013), rs8050136 (P=0.00011), rs1558902 (P=6.6 × 10(-5)) and rs1421085 (P=7.4 × 10(-5)). rs3764220 in the SCG3 gene (P=0.0010) and rs2293855 in the MTMR9 gene (P=0.0015) were also significantly associated with metabolic syndrome. SNPs in the FTO, SCG3 and MTMR9 genes had no SNP × SNP epistatic effects on metabolic syndrome. Our data suggest that genetic variations in the FTO, SCG3 and MTMR9 genes independently influence the risk of metabolic syndrome.

Visceral fat accumulation has an important role in increasing morbidity and mortality rate by increasing the risk of developing several metabolic disorders, such as type 2 diabetes, dyslipidemia and hypertension. New genetic loci that contribute to the development of obesity have been identified by genome-wide association studies in Caucasian populations. We genotyped 1279 Japanese subjects (556 men and 723 women), who underwent computed tomography (CT) for measuring visceral fat area (VFA) and subcutaneous fat area (SFA), for the following single-nucleotide polymorphisms (SNPs): NEGR1 rs2815752, SEC16B rs10913469, TMEM18 rs6548238, ETV5 rs7647305, GNPDA2 rs10938397, BDNF rs6265 and rs925946, MTCH2 rs10838738, SH2B1 rs7498665, MAF rs1424233, and KCTD15 rs29941 and rs11084753. In the additive model, none of the SNPs were significantly associated with body mass index (BMI). The SH2B1 rs7498665 risk allele was found to be significantly associated with VFA (P=0.00047) but not with BMI or SFA. When the analysis was performed in men and women separately, no significant associations with VFA were observed (P=0.0099 in men and P=0.022 in women). None of the other SNPs were significantly associated with SFA. Our results suggest that there is a VFA-specific genetic factor and that a polymorphism in the SH2B1 gene influences the risk of visceral fat accumulation.

We examined pancreas biopsy specimens from 18 newly diagnosed insulin-dependent diabetes mellitus (IDDM) patients to elucidate the mechanism underlying beta cell destruction. Pancreas islets were seen in all patients and insulitis in eight patients. Infiltrating mononuclear cells consisted of CD4+T, CD8+T, B lymphocytes, and macrophages. Among them, CD8+T lymphocytes were predominant and macrophages followed. The expression of MHC class I antigens was increased in islet and endothelial cells in nine patients. MHC class II expression was increased in endothelial cells of the same patients. The expression of intercellular adhesion molecule-1 was increased in endothelial cells in two of the nine patients with MHC hyperexpression; in one of them, lymphocyte function-associated antigen-3 expression was also increased. Out of the eight patients with insulitis, seven showed MHC class I hyper-expression, whereas 2 of the 10 patients without insulitis showed the phenomenon (P < 0.05). The relation between insulitis and the hyperexpression of adhesion molecules was not evident. In conclusion, we revealed the close relation between CD8+T lymphocyte-predominant insulitis and MHC class I hyperexpression in islet cells. This suggests that infiltrating CD8+T lymphocytes recognize islet autoantigens in association with increased MHC class I molecules and act as major effector cells in autoimmune response against islet cells in IDDM pancreases. The role of adhesion molecules in the pathogenesis of IDDM still remains to be elucidated.

Chromosomal genes for two mouse homologous beta subunits (AIC2A and AIC2B) of the interleukin-3, granulocyte/macrophage colony-stimulating factor, and interleukin-5 receptors were characterized. Both AIC2A and AIC2B genes were present on a 250-kilobase MluI restriction fragment and were mapped on murine chromosome 15 (these loci were provisionally designated as Il3rb-1 (AIC2A) and Il3rb-2 (AIC2B)), closely linked to the c-sis locus. Both genes consist of 14 exons and span about 28 kb each. The major transcription initiation sites of both genes were mapped at 194 bp from the initiation codon. These genes are 95% identical up to 700 bp from the transcription initiation sites. Potential recognition sequences for hemopoietic transcription factors including GATA-1 and PU.1 in addition to a TATA-like sequence are present in the 5'-flanking region. A stretch of 20 bp including the initiation site is homologous to the corresponding region of the erythropoietin receptor and the interleukin-7 receptor genes and to the initiator sequence of the adeno-associated virus P5 promoter, suggesting a possible role in transcription initiation. Comparison of the exon/intron boundaries of AIC2A and AIC2B genes with those of other members of the cytokine receptor superfamily reveals a conserved evolutionary structure. Isolation of various forms of AIC2 cDNAs reveals differential splicing of the transcripts.

We investigated transthyretin (TTR) in the pancreases and sera of 10 newly diagnosed type I diabetic patients by immunohistochemistry and nephelometry. In the type I diabetic pancreases, glucagon-positive A-cells showed strong immunoreactivity for TTR, the intensity and distribution pattern of which corresponded to those in normal subjects. Morphometric analysis revealed that the amount of strongly TTR-positive A-cells was not significantly different from that in normal subjects. On the contrary, insulin-positive B-cells, which normally show uneven and weak TTR immunoreactivity, decreased in number, and only a few residual B-cells showed faint immunoreactivity. Neither somatostatin cells nor pancreatic polypeptide cells were positive for TTR. The serum TTR concentration showed a significant decrease in type I diabetic patients compared with that in normal subjects (P less than 0.005). These data suggest that the synthesis or storage of TTR in A-cells is not affected, but that in B-cells is impaired in type I diabetes. The decrease in serum TTR might be one of the features of metabolic disorders in type I diabetes.

Complement-fixing adrenal medullary antibodies were examined in sera from 170 (114 Type 1 and 56 Type 2) diabetic patients and normal subjects by indirect immunofluorescence methods. Two types of antibodies were detected; one showed a homogeneous immunofluorescence pattern (homogeneous-type) and the other a spotty pattern (spotty-type) in the cytoplasm of adrenal medullary cells. Both antibodies were IgG class and adrenal medulla-specific. The prevalence of the homogeneous-type was significantly higher in Type 1 diabetic patients with disease duration under 1 year (36%) than in those with duration of 1 year or more (1.1%), in Type 2 diabetic patients (1.8%) or in normal subjects (0%; P less than 0.01). Conversely, the prevalence of the spotty-type was not significantly different among all subjects examined (3.6-4.5%). The epitope for the homogeneous-type is likely to be a glycoconjugate since binding of this antibody was abolished after periodate oxidation. The epitope for spotty-type antibody is considered to be a peptide since it was trypsin sensitive. Patients who were positive for the homogeneous-type were also positive for islet cell antibodies, although their antibody titers were not correlated. We conclude that (1) adrenal medullary antibodies are of homogeneous-type or spotty-type and the antigenic determinants of these antibodies are different, and (2) the prevalence of the homogeneous-type is significantly higher in newly diagnosed Type 1 diabetic patients and its presence is associated with that of islet cell antibodies.

A case of cellular schwannoma originating from the retroperitoneum is reported. The histological features of the tumor resembled those of a smooth muscle tumor. However, the tumor cells were positive for S-100 protein, which is almost exclusively identified within nerve sheath tumor cells. This finding confirmed the diagnosis of cellular schwannoma. This type of tumor is generally considered to be benign but is sometimes mistaken for a malignant tumor. These findings indicate that it is important to prove whether or not S-100 protein is present in the cells of a tumor of unknown origin especially arising in the mediastinum or retroperitoneum.

A 68-year-old woman underwent distal gastrectomy for gastric cancer in February 2003. The Histological type was poorly differentiated adenocarcinoma, and the final finding was T3 (se, INF gamma, ly1, v0), pN1, sH0, sP0, pCY0; fStage IIIA. Two years and 5 months after the operation, a painless tumor was noticed on the upper abdomen. Biopsy specimen showed adenocarcinoma. A CT scan detected abdominal tumors, and there was no other distant metastasis. Fourteen courses of systemic chemotherapy (IRIS) were performed for sixteen months. We detected several abdominal metastases, but no distant metastasis was occurred during IRIS.

Interleukin-3 (IL-3) binds to its receptor with high and low affinities, induces tyrosine phosphorylation, and promotes the proliferation and differentiation of hematopoietic cells. A binding component of the IL-3 receptor was cloned. Fibroblasts transfected with the complementary DNA bound IL-3 with a low affinity [dissociation constant (Kd) of 17.9 +/- 3.6 nM]. No consensus sequence for a tyrosine kinase was present in the cytoplasmic domain. Thus, additional components are required for a functional high affinity IL-3 receptor. A sequence comparison of the IL-3 receptor with other cytokine receptors (erythropoietin, IL-4, IL-6, and the beta chain IL-2 receptor) revealed a common motif of a distinct receptor gene family.

Genetic factors are clearly involved in the development of obesity, but the genetic background of obesity remains largely unclear. Starting from 62 663 gene-based single-nucleotide polymorphisms (SNPs) in three sequential case-control association studies, we identified a replicated association between the obesity phenotype (BMI > or =30 kg/m(2)) and a SNP (rs2293855) located in the myotublarin-related protein 9 (MTMR9) gene in the chromosomal segment 8p23-p22. P-values (minor allele dominant model) of the first set (93 cases versus 649 controls) and the second set (564 cases versus 562 controls) were 0.008 and 0.0002, respectively. The association was replicated in the third set [394 cases versus 958 controls, P = 0.005, odds ratio (95% CI) =1.40 (1.11-1.78)]. The global P-value was 0.0000005. A multiple regression analysis revealed that gender, age BMI and rs2293855 genotype (minor allele dominant model) were significantly associated with both systolic and diastolic blood pressures. MTMR9 was shown to be the only gene within the haplotype block that contained SNPs associated with obesity. Both the transcript and protein of MTMR9 were detected in the rodent lateral hypothalamic area as well as in the arcuate nucleus, and the protein co-existed with orexin, melanin concentrating hormone, neuropeptide Y and proopiomelanocortin. The levels of MTMR9 transcript in the murine hypothalamic region increased after fasting and were decreased by a high-fat diet. Our data suggested that genetic variations in MTMR9 may confer a predisposition towards obesity and hypertension through regulation of hypothalamic neuropeptides.

A 63-year-old woman was admitted to the hospital for investigation of abdominal discomfort. Further examination revealed that she had sigmoid colon cancer. The serum CEA level was 20.1 ng/ml, and the CA19-9 level was 8.9 U/ml. Laparotomy findings showed H0, P0, and lymph-nodes swelling observed at LN253-LN16b2. Massive tumor thrombosis was present in IMV, the splenic vein, and the advanced region in the portal vein. Metastatic tumor was present in the pancreas body. An anterior resection of the colon combined with resection of lymph nodes, pancreas body and tail and spleen were performed. Resection of IMV and splenic vein including tumor thrombosis was performed prudently. Histological examination revealed that the pancreas tumor and tumor thrombosis were of the same origin (moderately differentiated from adenorarcinoma), and massive LN metastases existed. Post operative chemotherapy, an oral administration of UFT/UZEL, was performed. After 24 months of operation, there has been no sign of recurrence detected.

Type I human diabetes (IDDM), the BB rat, and the NOD mouse appear to result from autoimmune B-cell destruction in a setting of genetic predisposition.1,2 The NOD/Shi mouse was established in 1980 at the Shionogi Aburahi Laboratories.3 The NOD mouse develops IDDM secondary to B-cell destruction by infiltrating immune cells (insulitis) (Fig. 1; Table 1). Insulitis appears as early as 5 wk of age. To develop overt diabetes, at least 90% of the total islets need to be damaged by infiltrating immune cells. The immune cells invade islets and are often seen adjacent to the ducts and blood vessels. The incidence of insulitis is 90% in NOD/Shi females and 70% in NOD/Shi males at 9 wk of age, and 100% in both sexes at 5 months of age. The incidence and degree of insulitis increase with age, resulting in a lack of intrinsic insulin secretion, hyperglycemia, loss of body weight, and ketosis, as seen in human IDDM. The NOD/Shi mouse develops diabetes usually by 7 months of age.

We investigated the contribution of mitochondrial DNA mutations to the pathogenesis of IDDM by analyzing mitochondrial DNA in pancreatic biopsy specimens and peripheral blood cells from 18 patients with newly-diagnosed IDDM. All patients presented with typical abrupt onset of diabetes and ketosis on initial examination. Point mutations at nucleotides 3243, 3271 and 8344 were assayed by polymerase chain reaction and restriction fragment length polymorphism analysis or by mismatch-primer analysis. A common large deletion from nucleotides 8483-13459 was analyzed by a primer shift method. All of these mutations are known to be pathogenic mutations. However, none of the mitochondrial DNA mutations were detected in any of 18 IDDM patients. Several types of mitochondrial DNA mutation have been identified in the peripheral blood cells in some patients with non-insulin-dependent diabetes mellitus as well as in some with IDDM, however, our results suggest that abrupt-onset IDDM does not correlate with any of the known mitochondrial DNA mutations.

DURING mammalian development, many cells are programmed to die1,2 most mediated by apoptosis3. The Fas antigen4 coded by the structural gene for mouse lymphoproliferation mutation (lpr)5,6, is a cell surface protein belonging to the tumour necrosis factor/nerve growth factor receptor family7,8, and mediates apoptosis7. The Fas antigen messenger RNA is expressed in the thymus, liver, heart, lung and ovary8. We prepared a monoclonal antibody against mouse Fas antigen, which immunoprecipitated the antigen (Mr 45K) and had cytolytic activity against cell lines expressing mouse Fas antigen. We report here that staining of mouse thymocytes with the antibody indicated that thymocytes from the wild-type and lprcg mice expressed the Fas antigen, whereas little expression of the Fas antigen was found in lpr mice. Intraperitoneal administration of the anti-Fas antibody into mice rapidly killed the wild-type mice but neither lpr nor lprcg mice. Biochemical, histological and electron microscope analyses indicated severe damage of the liver by apoptosis. These findings suggest that the Fas antigen is important in programmed cell death in the liver, and may be involved in fulminant hepatitis in some cases.

The highly purified yeast mRNA capping enzyme is composed of two separate chains of 52 (alpha) and 80 kDa (beta), responsible for the activities of mRNA guanylyltransferase and RNA 5'-triphosphatase, respectively (Itoh, N., Yamada, H., Kaziro, Y., and Mizumoto, K. (1987) J. Biol. Chem. 262, 1989-1995). The gene encoding the mRNA guanylyltransferase subunit (alpha subunit), CEG1, has been isolated by immunological screening of a yeast genomic expression library in lambda gt11 with polyclonal antibodies directed against purified yeast capping enzyme. The identity of CEG1 was confirmed by epitope selection and by expressing the gene in Escherichia coli to give a catalytically active mRNA guanylyltransferase. The gene is present in one copy per haploid genome, and encodes a polypeptide of 459 amino acid residues. From its primary structure as well as its mRNA size, it was concluded that the alpha and the beta subunits of yeast mRNA capping enzyme are encoded by two separate genes, not as a fused protein. CEG1 is located on the chromosome VII by a pulse-field gel electrophoresis. Gene disruption experiment indicated that CEG1 is essential for the growth of yeast. We have also found another open reading frame (ORF2) which lies in close proximity to CEG1 in our clones and encodes a 450 amino acid-polypeptide of yet unknown function.

Mouse anti-Fas monoclonal antibody has a cytolytic activity on human cells that express the antigen. Complementary DNAs encoding the cell surface antigen Fas were isolated from a cDNA library of human T cell lymphoma KT-3 cells. The nucleotide sequence of the cDNAs revealed that the molecule coding for the Fas antigen determinant is a 319 amino acid polypeptide (M r 36,000) with a single transmembrane domain. The extracellular domain is rich in cysteine residue, and shows a similarity to that of human tumor necrosis factor receptors, human nerve growth factor receptor, and human B cell antigen CD40. Murine WR19L cells or L929 cells transformed with the human Fas antigen cDNA were killed by the anti-Fas antibody in the process known as apoptosis.

The effects of interferon-gamma (IFN gamma) on the morphology of thyroid follicles and the expression of major histocompatibility complex (MHC) class II antigens were examined. The thyroid follicles were suspended in RPMI-1640 containing 10% fetal calf serum with or without IFN gamma (200 U/ml). After culture for 5 days, follicles were incubated in the presence of TSH (10 mU/ml) for 1 h and fixed for electron microscopic and immunohistochemical examination. Regardless of the presence of IFN gamma, suspended follicles became inverted within 5 days. However, MHC class II antigens were expressed only in inverted follicles cultured with IFN gamma. In inverted follicles cultured without IFN gamma, TSH stimulation induced remarkable morphological changes, such as elongation of microvilli and an appearance of pseudopods. On the other hand, the follicles cultured with IFN gamma showed poor response to TSH. Thus, IFN gamma induced the expression of MHC class II antigens of cultured thyroid follicles and inhibited TSH-induced morphological changes in the cells.

Using a monoclonal antibody to the interleukin 3 (IL-3) receptor (anti-Aic2), we isolated a cDNA (AIC2B) from a mouse mast cell line which is homologous to the previously characterized gene for the IL-3 receptor (AIC2A). This cDNA encodes a polypeptide of 896 amino acid residues and has 91% amino acid sequence identity with the IL-3 receptor. A consensus sequence defining an additional cytokine receptor family is present in this clone. Compared to the AIC2A clone, the AIC2B cDNA encodes a protein with amino acid substitutions, insertions, and deletions dispersed throughout the entire protein. Oligonucleotide probes specific for each cDNA hybridized with different genomic fragments, indicating that the AIC2A and AIC2B proteins are encoded by two distinct genes. Fibroblasts transfected with the AIC2B cDNA expressed the protein at the cell surface as determined by binding with the anti-Aic2 antibody but did not bind IL-3 or other cytokines, including IL-2, IL-4, granulocyte-macrophage colony-stimulating factor, erythropoietin, and IL-9 (p40) at concentrations between 1 and 10 nM. An S1 nuclease protection assay was used to discriminate between the AIC2A and AIC2B transcripts. We found that the AIC2B gene was coexpressed with the AIC2A gene. These results suggest a potential involvement of AIC2B in cytokine signal transduction.

Interleukin 4 (IL-4) is a potent mediator of growth and differentiation for various lymphoid and myeloid cells. To isolate a cDNA encoding the murine IL-4 receptor, we have developed an expression cloning method that uses biotinylated ligand as a probe and that may be generally applicable to cloning of receptor genes. COS-7 cells transiently transfected with the cloned full-length cDNA bind murine IL-4 specifically with a Kd = 165 pM. Crosslinking of 125I-labeled IL-4 to COS-7 cells transfected with the cDNA reveals binding to proteins of 120-140 kDa. IL-4-responsive cells also express IL-4-binding proteins of 120-140 kDa but show additional bands at 60-70 kDa; the relationship of the smaller proteins to the larger ones is unclear. The nucleotide sequence indicates that the full-length cDNA encodes 810 amino acids including the signal sequence. While no consensus sequence for protein kinases is present in the cytoplasmic domain, a sequence comparison with the erythropoietin receptor, the IL-6 receptor, and the beta chain of the IL-2 receptor reveals a significant homology in the extracellular domain, indicating that the IL-4 receptor is a member of a cytokine receptor family.

Klinefelter’s syndrome is a sex chromosomal aberration of male infertility. Most diabetes of this syndrome shows insulin resistance. Chronic proinflammation is involved in the pathogenesis of diabetes, and cytokines play a role in insulin resistance (1,2,3). Interleukin (IL)-8 is a cytokine secreted by monocytes and endothelial cells (6). IL-6 is associated with insulin resistance and is an inducer of C-reactive protein (CRP) (1,2,7). IL-8, IL-6, and CRP are higher in diabetes (1,2,3,4,5). We report a 30-year-old …