Yasushi Kawaguchi

Association of STAT4 with susceptibility to rheumatoid arthritis and systemic lupus erythematosus in the Japanese population.

OBJECTIVEnSTAT4 encodes a transcriptional factor that transmits signals induced by several key cytokines, and it might be a key molecule in the development of autoimmune diseases. Recently, a STAT4 haplotype was reported to be associated with rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE) in Caucasian populations. This was replicated in a Korean RA population. Interestingly, the degree of risk of RA susceptibility with the STAT4 haplotype was similar in the Caucasian and Korean populations. The present study was undertaken to investigate the effect of STAT4 on susceptibility to RA and SLE in the Japanese.nnnMETHODSnWe performed an association study using 3 independent Japanese RA case-control populations (total 3,567 cases and 2,199 controls) and 3 independent Japanese SLE populations (total 591 cases). All samples were genotyped using the TaqMan fluorogenic 5 nuclease assay for single-nucleotide polymorphism (SNP) rs7574865, which tags the susceptibility haplotype. The association of the SNP with disease susceptibility in each case-control study was calculated using Fishers exact test, and the results were combined, using the Mantel-Haenszel method, to obtain combined odds ratios (ORs).nnnRESULTSnWe observed a significant association of the STAT4 polymorphism with susceptibility to both RA and SLE. The combined ORs for RA and SLE, respectively, were 1.27 (P = 8.4 x 10(-9)) and 1.61 (P = 2.1 x 10(-11)) for allele frequency distribution; these ORs were quite similar to those previously observed in the Caucasian population.nnnCONCLUSIONnWe conclude that STAT4 is associated with RA and SLE in the Japanese. Our results indicate that STAT4 is a common genetic risk factor for autoimmune diseases, with similar strength across major racial groups.

Association of a functional polymorphism in the IRF5 region with systemic sclerosis in a Japanese population

OBJECTIVEnInterferon regulatory factor 5, an established susceptibility factor for systemic lupus erythematosus (SLE), plays a role in type I interferon and proinflammatory cytokine induction. A recent study showed association of a functional single-nucleotide polymorphism (SNP) in intron 1 of IRF5, rs2004640, with systemic sclerosis (SSc) in a European French population. We undertook the present study to determine whether IRF5 polymorphisms are also associated with a predisposition to SSc in Japanese.nnnMETHODSnA case-control association study was performed for rs2004640 as well as for rs10954213 and rs2280714, all of which were previously reported to be associated with SLE, in 281 SSc patients and 477 healthy controls. Patients with SSc complicated by SLE or Sjögrens syndrome were excluded. Association of the rs2280714 genotype with messenger RNA (mRNA) levels of IRF5 and adjacently located transportin 3 (TNPO3) was examined using the GENEVAR database.nnnRESULTSnAll 3 SNPs were significantly associated with SSc, with the rs2280714 A allele having the strongest association (allele frequency P=0.0012, odds ratio 1.42 [95% confidence interval 1.15-1.75]). Association was preferentially observed in subsets of patients with diffuse cutaneous SSc (dcSSc) and anti-topoisomerase I antibody positivity. Conditional analysis revealed that rs2280714 could account for most of the association of these SNPs, while an additional contribution of rs2004640 was also suggested for dcSSc. The genotype of rs2280714 was strongly associated with IRF5 mRNA expression, while only marginal association was detected with TNPO3 mRNA expression.nnnCONCLUSIONnAssociation of IRF5 with SSc was replicated in a Japanese population. Whether the causal SNP is different among populations requires further investigation.

Anti-MDA5 antibody, ferritin and IL-18 are useful for the evaluation of response to treatment in interstitial lung disease with anti-MDA5 antibody-positive dermatomyositis

OBJECTIVEnThe aim of this study was to investigate the precise clinical characteristics and to analyse the association between the anti-MDA5 antibody (anti-MDA5ab) titre and disease status in patients with anti-MDA5ab-positive DM.nnnMETHODSnTwenty-seven patients who presented with DM and were positive for the anti-MDA5ab were enrolled. The association between the clinical manifestations and the clinical parameters, including the anti-MDA5ab, was analysed.nnnRESULTSnThe complication of rapidly progressive interstitial lung disease (RP-ILD) occurred in 20 (74%) patients. The frequencies of fatal outcome, relapse and malignancy were 33, 4 and 4%, respectively. Remarkably, a fatal outcome occurred within the first 6 months. Compared with six non-RP-ILD patients, elderly age at onset, severely involved pulmonary function and high levels of serum ferritin were present in 20 RP-ILD patients with anti-MDA5ab. Alveolar-arterial oxygen difference (AaDO(2)) ≥32u2009mmHg and ferritin ≥828u2009ng/ml at admission were poor prognostic factors in RP-ILD patients with anti-MDA5ab-positive DM. The median value of the anti-MDA5ab titre on admission was higher in patients who later died than in those who survived. The efficacy of treatment was indicated by the anti-MDA5ab, ferritin and IL-18 concentrations. The decline index of the anti-MDA5ab titre after treatment was lower in the subset of patients who died than in the subset of patients who lived. Sustained high levels of anti-MDA5ab, ferritin and IL-18 were present in the patients who died.nnnCONCLUSIONnAnti-MDA5ab titre and ferritin and IL-18 concentrations are useful for the evaluation of the response to treatment and the status of ILD in patients with anti-MAD5ab-positive DM.

Association of STAT4 polymorphism with systemic sclerosis in a Japanese population

Some susceptibility genes are shared by multiple autoimmune diseases. For example, the interferon regulatory factor 5 ( IRF5 ) gene is associated with various autoimmune diseases including systemic lupus erythematosus (SLE) and systemic sclerosis (SSc).1–3 The signal transducer and activator of transcription 4 ( STAT4 ) gene has been shown to be a susceptibility gene for rheumatoid arthritis (RA) and SLE.4 5 STAT4 is a transcription factor involved in interleukin (IL)12 signalling in induction of interferon (IFN)γ production and T helper type 1 (Th1) cell differentiation, type I IFN signalling and IL23-induced IL17 production.6 An association of rs7574864 with SSc was recently reported by a large-scale case-control study in European populations.7 In the present study, we examined the association of a STAT4 intron 3 single nucleotide …

NOS2 polymorphisms associated with the susceptibility to pulmonary arterial hypertension with systemic sclerosis: contribution to the transcriptional activity.

Systemic sclerosis (SSc) is a connective tissue disease characterized by tissue fibrosis. One of several complications of SSc, pulmonary arterial hypertension (PAH) can be refractory to treatment, both novel and established. In the present study we investigated the ratio of circulating nitric oxide to endothelin-1 in patients with both SSc and PAH, and determined whether polymorphisms in NOS2 (the nitric oxide synthase 2 gene) are associated with susceptibility to PAH. Endothelin-1 in plasma and nitric oxide metabolites (nitrate and nitrite) in serum were measured. The nitric oxide/endothelin-1 ratio was significantly lower in patients with both SSc and PAH than in patients with SSc only or in healthy control individuals. We confirmed the presence of two single nucleotide polymorphisms at positions -1,026 and -277 and a pentanucleotide repeat (CCTTT) at -2.5 kilobases. There were significant differences in single nucleotide polymorphisms between patients with SSc who had PAH and those who did not, and between patients with both SSc and PAH and healthy control individuals. The CCTTT repeat was significantly shorter in patients with both SSc and PAH than in patients with SSc only or in healthy control individuals. Transcriptional activity were analyzed using the luciferase reporter assay. The transcriptional activity of NOS2 was much greater in fibroblasts transfected by a vector with a long allele of the CCTTT repeat than in those transfected by a vector with a short allele. Polymorphisms in the NOS2 gene are associated with transcriptional activity of the NOS2 gene and with susceptibility to SSc-related PAH.

Muscarinic-3 acetylcholine receptor autoantibody in patients with systemic sclerosis: contribution to severe gastrointestinal tract dysmotility

Objective: Patients with systemic sclerosis (SSc) complicated by severe gastrointestinal tract (GIT) dysmotility at an early stage are difficult to treat and mortality is high. To clarify the pathogenesis of GIT involvement, the occurrence of autoantibody was investigated for muscarinic-3 acetylcholine receptor (M3R) in patients with SSc. Methods: Fourteen patients with severe GIT involvement (malabsorption syndrome and/or pseudo-obstruction) within 2 years of SSc onset (group 1) were enrolled in the present study. Sixty-two patients with SSc without severe GIT involvement within 2 years of onset (group 2) were also recruited, along with 70 healthy control subjects. Using an established enzyme immunoassay (EIA) system detecting autoantibody against the second loop domain of M3R, the presence of an anti-M3R antibody was examined in SSc patients. Results: The mean optical density (OD) titres of group 1 were significantly higher than those of group 2 (0.65 (SD 0.58) vs 0.066 (SD 0.13), p<0.001). The positivity of anti-M3R antibody was significantly higher in group 1 than in group 2 (9/14 vs 3/62, pu200a=u200a2.5 × 10−6 by Fisher’s exact test). The cutoff OD was calculated from the EIA reaction of the 70 healthy controls (the mean value plus 2 SD was 0.295). Conclusion: The findings indicated that anti-M3R antibody very frequently appears in patients with SSc, which is accompanied by severe GIT involvement, suggesting that M3R-mediated enteric cholinergic neurotransmission may provide a pathogenic mechanism for GIT dysmotility in SSc.

Association study of TRAF1-C5 polymorphisms with susceptibility to rheumatoid arthritis and systemic lupus erythematosus in Japanese

Objective: The primary aim of this study was to investigate the association of polymorphisms of TRAF1-C5, a newly identified rheumatoid arthritis (RA) risk locus in Caucasians, with susceptibility to RA and systemic lupus erythematosus (SLE) in Japanese populations. Gene expression levels of TRAF1 and C5 to assess the functional significance of genotypes were also analysed. Methods: A multicentre association study consisting of 4 RA case-control series (4397 cases and 2857 controls) and 3 SLE case-control series (591 cases and 2199 shared controls) was conducted. Genotyping was performed using TaqMan genotyping assay for two single nucleotide polymorphisms (SNPs) that showed the best evidence of association in the previous Caucasian studies. Quantifications of TRAF1 and C5 expression were performed with TaqMan expression assay. Results: Significant differences in allele frequency for both SNPs were observed between RA and control subjects (combined odds ratio u200a=u200a1.09), while no significant difference was detected between patients with SLE and controls. Interestingly, alleles rs3761847 A and rs10818488 G had increased the risk for RA in the present study, while they decreased the risk in the original studies. A significant difference was found between risk allele carriers and non-carriers of rs10818488 for the expression level of TRAF1 in phorbol myristate acetate-stimulated lymphoblastoid cell lines (pu200a=u200a0.04). Conclusion: Association of TRAF1-C5 locus with RA susceptibility was detected in the Japanese populations with modest magnitude, while no significant association was observed for SLE. Significant positive effect of genotype on the expression of TRAF1 might support the genetic association between TRAF1 and RA.

Clinical usefulness of anti-RNA polymerase III antibody measurement by enzyme-linked immunosorbent assay

OBJECTIVEnTo evaluate the clinical usefulness of measuring anti-RNA polymerase (RNAP) III antibody with a commercially available ELISA in Japanese patients with SSc.nnnMETHODSnThis multicentre study involved 354 patients with SSc, 245 with non-SSc CTDs and 102 healthy controls. ELISAs were used to detect anti-RNAP III antibody, anti-topo I antibody and ACA. The presence of anti-RNAP III antibody in selected serum samples was confirmed by immunoprecipitation (IP) assay.nnnRESULTSnBy ELISA, anti-RNAP III antibody was detected in 38 (10.7%) patients with SSc, 3 (1.2%) with non-SSc CTD and no healthy controls. The clinical specificity for SSc was excellent (98.8%), although a small number of false positives occurred. The sensitivity of the anti-topo I and ACA ELISAs for SSc was 59.9%, which increased to 68.2% without a reduction in specificity when the anti-RNAP III measurement was added. Clinical features associated with positivity for the anti-RNAP III antibody include dcSSc, a high total skin score and a trend towards high prevalence of renal crisis, consistent with previous studies that used an IP assay. Furthermore, on clinical severity scales, SSc patients with anti-RNAP III antibody scored highest for skin and renal involvement among patients subgrouped by the presence of individual SSc-related antibodies.nnnCONCLUSIONSnThe measurement of anti-RNAP III antibody by ELISA is useful in routine clinical practice, because it helps diagnose SSc and identify a disease subset with severe skin and renal involvement.

Association study of a polymorphism of the CTGF gene and susceptibility to systemic sclerosis in the Japanese population

Objectives: To validate the association of a single nucleotide polymorphism (SNP) of the connective tissue growth factor gene (CTGF) with susceptibility to systemic sclerosis (SSc) in the Japanese population. Methods: 395 Japanese patients with SSc, 115 patients with rheumatoid arthritis and 269 healthy Japanese volunteers were enrolled in the study. An SNP (rs6918698) at –945 bp from the start codon in the promoter region of the CTGF gene was determined by allelic discrimination with the use of a specific TaqMan probe. Results: The G allele showed a significantly higher frequency in patients with SSc than in controls (p<0.001; odds ratio 1.5; 95% confidence interval 1.2 to 1.9). In particular, the clinical subsets of SSc showed a more significant association between the G allele and diffuse cutaneous SSc (p<0.001) and the presence of interstitial lung disease (p<0.001), the presence of anti-topoisomerase I antibody (p<0.001) and anti-U1RNP antibody (pu200a=u200a0.010). Association analyses using the genotype of the SNP yielded results similar to those of analyses using the allele. Conclusions: This study confirms the association between an SNP in the CTGF gene and susceptibility to SSc, especially in the presence of diffuse cutaneous SSc, interstitial lung disease and anti-topoisomerase I antibody. The results strongly suggest that this SNP may be a powerful indicator of severe skin and lung involvement in patients with SSc.

Identification of a significant association of a single nucleotide polymorphism in TNXB with systemic lupus erythematosus in a Japanese population

AbstractSystemic lupus erythematosus (SLE) is one of the common autoimmune diseases, with complex genetic components. Here, we report on a case–control association study of 178 SLE patients and 899 control subjects, using genome-wide gene-based single nucleotide polymorphism (SNP) markers. An SNP, rs3130342, in a 5′ flanking region of the TNXB gene revealed a significant association with SLE [P = 0.000000930, odds ratio (OR) 3.11, with 95% confidence interval (95%CI) of 1.89–5.28] in a Japanese population. This association was replicated independently with 203 cases and 294 controls (P = 0.0440, OR 1.52, with 95%CI of 1.01–2.78). Although a copy number variation (CNV) of the C4 gene adjacent to the TNXB gene was reported to be associated with SLE, our analysis on this CNV revealed that the association of CNV of the C4 gene was weaker than the SNP in the TNXB gene and likely to reflect the linkage disequilibrium between C4 CNV and this particular SNP. Stratified analysis also revealed that the association of SNP rs3130342 with SLE was independent of the HLA-DRB1*1501 allele that has been shown to be associated with SLE. Our findings strongly imply that the TNXB gene is a candidate gene susceptible to SLE in the Japanese population.