Yoshinari Takasaki

Sex-specific association of X-linked Toll-like receptor 7 (TLR7) with male systemic lupus erythematosus

Systemic lupus erythematosus (SLE) is a multisystem, autoimmune disease that predominantly affects women. Previous findings that duplicated Toll-like receptor 7 (Tlr7) promotes lupus-like disease in male BXSB mice prompted us to evaluate TLR7 in human SLE. By using a candidate gene approach, we identified and replicated association of a TLR7 3′UTR SNP, rs3853839 (G/C), with SLE in 9,274 Eastern Asians (Pcombined = 6.5 × 10−10), with a stronger effect in male than female subjects [odds ratio, male vs. female = 2.33 (95% CI = 1.64–3.30) vs. 1.24 (95% CI = 1.14–1.34); P = 4.1 × 10−4]. G-allele carriers had increased TLR7 transcripts and more pronounced IFN signature than C-allele carriers; heterozygotes had 2.7-fold higher transcripts of G-allele than C-allele. These data established a functional polymorphism in type I IFN pathway gene TLR7 predisposing to SLE, especially in Chinese and Japanese male subjects.

High diagnostic performance of ELISA detection of antibodies to citrullinated antigens in rheumatoid arthritis.

Objective: We investigated the rheumatoid arthritis (RA) diagnostic performances of anti‐cyclic citrullinated peptide antibody (anti‐CCP) and antifilaggrin antibody (AFA) in comparison with RF and matrix metalloproteinase‐3 (MMP‐3). Methods: We used a second generation enzyme‐linked immunosorbent assay (ELISA) kit for the detection of anti‐CCP. We constructed recombinant human filaggrin, which was citrullinated in vitro by human peptidylarginine deiminase, and subsequently used it as the coating antigen for AFA‐ELISA. A total of 549 RA patients and 208 other rheumatic disease patients were included in the study. Results: The specificities of anti‐CCP (88.9%) and AFA (94.7%) were superior to those of RF (81.7%) and MMP‐3 (49.5%). The sensitivity of anti‐CCP (87.6%) was superior to all others. However, the sensitivity of AFA (68.7%) was inferior to those of RF (69.8%) and MMP‐3 (75.7%). Furthermore, receiver operating characteristic curves of anti‐CCP and AFA passed closer to the upper left corner than those of RF and MMP‐3, and the areas under the curves (AUC) of AFA and anti‐CCP were significantly larger. In addition, the AUC of anti‐CCP was significantly larger than that of AFA. Conclusion: ELISA detection of antibodies to citrullinated antigens, especially a second generation anti‐CCP, showed higher discriminative ability than other assays, including RF, and would be useful to aid the diagnosis of RA in clinical practice.

Levels of IL-12 in the sera of patients with systemic lupus erythematosus (SLE)— relation to Th1- and Th2-derived cytokines

IL‐12 is a cytokine that induces Th1‐derived cytokines (interferon‐gamma (IFN‐γ) and IL‐2). The significance of IL‐12 in human autoimmunity is no clear, and the serum levels of IL‐12 in SLE are not clearly established. Therefore, we examined the levels of IL‐12 in 39 patients with active SLE, with sandwich ELISA. The levels of IL‐12 in patients were significantly higher than in normal subjects. Patients with high levels of IL‐12 also had high levels of IFN‐γ, while their levels of IL‐13 were significantly lower than in patients with normal levels of IL‐12. Patients with pulmonary involvement had high levels of IL‐12, and steroid therapy decreased the IL‐12 level in three patients. In a retrospective study of seven patients, various changes of IL‐12 and IL‐13 were recognized before disease flare. Thus, in SLE patients, the level of IL‐12 was increased and this increase was related to the change of Th1‐ or Th2‐derived cytokines with some organ involvement.

Role of STAT4 polymorphisms in systemic lupus erythematosus in a Japanese population: a case-control association study of the STAT1-STAT4 region

IntroductionRecent studies identified STAT4 (signal transducers and activators of transcription-4) as a susceptibility gene for systemic lupus erythematosus (SLE). STAT1 is encoded adjacently to STAT4 on 2q32.2-q32.3, upregulated in peripheral blood mononuclear cells from SLE patients, and functionally relevant to SLE. This study was conducted to test whether STAT4 is associated with SLE in a Japanese population also, to identify the risk haplotype, and to examine the potential genetic contribution of STAT1. To accomplish these aims, we carried out a comprehensive association analysis of 52 tag single nucleotide polymorphisms (SNPs) encompassing the STAT1-STAT4 region.MethodsIn the first screening, 52 tag SNPs were selected based on HapMap Phase II JPT (Japanese in Tokyo, Japan) data, and case-control association analysis was carried out on 105 Japanese female patients with SLE and 102 female controls. For associated SNPs, additional cases and controls were genotyped and association was analyzed using 308 SLE patients and 306 controls. Estimation of haplotype frequencies and an association study using the permutation test were performed with Haploview version 4.0 software. Population attributable risk percentage was estimated to compare the epidemiological significance of the risk genotype among populations.ResultsIn the first screening, rs7574865, rs11889341, and rs10168266 in STAT4 were most significantly associated (P < 0.01). Significant association was not observed for STAT1. Subsequent association studies of the three SNPs using 308 SLE patients and 306 controls confirmed a strong association of the rs7574865T allele (SLE patients: 46.3%, controls: 33.5%, P = 4.9 × 10-6, odds ratio 1.71) as well as TTT haplotype (rs10168266/rs11889341/rs7574865) (P = 1.5 × 10-6). The association was stronger in subgroups of SLE with nephritis and anti-double-stranded DNA antibodies. Population attributable risk percentage was estimated to be higher in the Japanese population (40.2%) than in Americans of European descent (19.5%).ConclusionsThe same STAT4 risk allele is associated with SLE in Caucasian and Japanese populations. Evidence for a role of STAT1 in genetic susceptibility to SLE was not detected. The contribution of STAT4 for the genetic background of SLE may be greater in the Japanese population than in Americans of European descent.

A Genome-Wide Association Study Identified AFF1 as a Susceptibility Locus for Systemic Lupus Eyrthematosus in Japanese

Systemic lupus erythematosus (SLE) is an autoimmune disease that causes multiple organ damage. Although recent genome-wide association studies (GWAS) have contributed to discovery of SLE susceptibility genes, few studies has been performed in Asian populations. Here, we report a GWAS for SLE examining 891 SLE cases and 3,384 controls and multi-stage replication studies examining 1,387 SLE cases and 28,564 controls in Japanese subjects. Considering that expression quantitative trait loci (eQTLs) have been implicated in genetic risks for autoimmune diseases, we integrated an eQTL study into the results of the GWAS. We observed enrichments of cis-eQTL positive loci among the known SLE susceptibility loci (30.8%) compared to the genome-wide SNPs (6.9%). In addition, we identified a novel association of a variant in the AF4/FMR2 family, member 1 (AFF1) gene at 4q21 with SLE susceptibility (rs340630; P = 8.3×10−9, odds ratio = 1.21). The risk A allele of rs340630 demonstrated a cis-eQTL effect on the AFF1 transcript with enhanced expression levels (P<0.05). As AFF1 transcripts were prominently expressed in CD4+ and CD19+ peripheral blood lymphocytes, up-regulation of AFF1 may cause the abnormality in these lymphocytes, leading to disease onset.

Cloning and nucleotide sequence of cDNA for Ki antigen, a highly conserved nuclear protein detected with sera from patients with systemic lupus erythematosus

Patients with systemic lupus erythematosus (SLE) produce autoantibodies against a variety of nuclear antigens including Ki antigen. Although anti‐Ki autoantibodies were found in a significant number of SLE patients, the nature of Ki antigen is poorly characterized. By using anti‐Ki serum as a probe we have cloned a bovine cDNA directing the synthesis in Escherichia coli of a polypeptide immunologically indistinguishable from the authentic Ki antigen. A homologous human cDNA was also cloned and its nucleotide sequence predicted the entire primary structure of a novel nuclear protein with a molecular weight of 29 508 and with highly hydrophilic and weakly acidic character. The gene is highly conserved not only in the coding region but also in the 3′‐untranslated region. The bacterially produced Ki antigen would be valuable for diagnosis of SLE.

The association of a nonsynonymous single-nucleotide polymorphism in TNFAIP3 with systemic lupus erythematosus and rheumatoid arthritis in the Japanese population.

OBJECTIVE Genome-wide association (GWA) studies in systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA) in Caucasian populations have independently identified risk variants in and near the tumor necrosis factor alpha (TNFalpha)-induced protein 3 gene (TNFAIP3), which is crucial for the regulation of TNF-mediated signaling and Toll-like receptor signaling. The aim of this study was to assess the role of TNFAIP3 in the development of SLE and RA in Japanese subjects. METHODS We selected 2 single-nucleotide polymorphisms (SNPs) from previous GWA studies. Rs2230926 is a nonsynonymous SNP in TNFAIP3 and is associated with SLE, while rs10499194 is an intergenic SNP associated with RA. We then performed 2 independent sets of SLE case-control comparisons (717 patients and 1,362 control subjects) and 3 sets of RA case-control comparisons (3,446 patients and 2,344 control subjects) using Japanese subjects. We genotyped SNPs using TaqMan assays. RESULTS We observed a significant association between rs2230926 and an increased risk of SLE and RA in the Japanese population (for SLE, odds ratio [OR] 1.92, 95% confidence interval [95% CI] 1.53-2.41, P = 1.9 x 10(-8); for RA, OR 1.35, 95% CI 1.18-1.56, P = 2.6 x 10(-5)). The intergenic SNP rs10499194 was also associated with SLE and RA, while the risk allele for RA in Caucasians was protective against the diseases in our population. CONCLUSION We demonstrated a significant association between the nonsynonymous variant in TNFAIP3 and the risk for SLE and RA in the Japanese population. TNFAIP3, similar to STAT4 and IRF5, may be a common genetic risk factor for SLE and RA that is shared between the Caucasian and Japanese populations.

International multicenter evaluation of autoantibodies to ribosomal P proteins.

ABSTRACT Autoantibodies to the ribosomal phosphoproteins (Rib-P) are a serological feature of patients with systemic lupus erythematosus (SLE). The reported prevalence of anti-Rib-P antibodies in SLE ranges from 10 to 40%, being higher in Asian patients. The variation in the observed frequency may be related to a number of factors but is dependent in large part on the test system used to detect the autoantibodies. An association of anti-Rib-P with central nervous system involvement and neuropsychiatric manifestations of SLE has been controversial. In the present international multicenter study, we evaluated the clinical accuracy of a new sensitive Rib-P-specific enzyme-linked immunosorbent assay based on recombinant Rib-P polypeptides. The results showed that 21.3% of 947 SLE patients, but only 0.7% of 1,113 control patients, had a positive test result (P < 0.0001). The sensitivity, specificity, positive and negative predictive values, and diagnostic efficiency were determined to be 21.3%, 99.3%, 95.6%, 62.2%, and 65.3%, respectively. When evaluated in the context of participating centers, the prevalence of anti-Rib-P antibodies was found in descending frequency, as follows: China (35%) > Poland (34%) > Japan (28%) > United States (26%) > Germany (Freiburg; 23.3%) > Denmark (20.5%) > Germany (Berlin; 19%) > Mexico (15.7%) > Israel (11.7%) > Brazil (10%) > Canada (8%). The substantial data from this study indicate that the prevalence of anti-Rib-P antibodies may not be restricted to the genetic background of the patients or to the detection system but may depend on regional practice differences and patient selection. We confirm previously reported associations of antiribosomal antibodies with clinical symptoms and serological findings. Remarkably, we found a lower occurrence of serositis in Rib-P-positive lupus patients.

Replication of the association between the C8orf13–BLK region and systemic lupus erythematosus in a Japanese population

OBJECTIVE Recent genome-wide association studies identified an association between single-nucleotide polymorphisms (SNPs) in the C8orf13 region of BLK, the B lymphoid tyrosine kinase gene, with systemic lupus erythematosus (SLE) in Caucasians. The purpose of this study was to evaluate the significance of this region in the genetic background of Japanese patients with SLE. METHODS Fourteen tag SNPs in the C8orf13-BLK region were genotyped in 327 Japanese patients with SLE and 322 healthy Japanese controls. The population-attributable risk percentage (PAR%) of rs13277113 in Japanese was compared with that in Caucasians as well as with that of other SLE susceptibility genes in Japanese. RESULTS As in Caucasians, rs13277113A demonstrated the strongest association in Japanese (P = 1.73 x 10(-6) for the genotype frequency, P = 4.75 x 10(-7) for the allele frequency, odds ratio [OR] 2.44 [95% confidence interval (95% CI) 1.43-4.16]). The association in Japanese was consistent with a recessive model (P = 2.74 x 10(-7), OR 2.27 [95% CI 1.66-3.11]). In contrast to the Caucasian population, this risk allele was the major allele in the Japanese population. Because both the risk allele frequency and the OR were higher in Japanese than in Caucasians, the PAR% of rs13277113 was estimated to be much higher in Japanese (35.4%) than in Caucasians (16.2%), and the second highest among the 6 confirmed SLE susceptibility genes in Japanese. CONCLUSION The association of the C8orf13-BLK region with SLE was replicated in a Japanese population. Contribution of this region to the genetic predisposition to SLE appeared to be greater in Japanese than in Caucasians.

DNA methylation: its contribution to systemic lupus erythematosus.

AbstractRecent studies on epigenetics, including the methylation of DNA and the enzymes regulating methylation, seem likely to contribute to understanding the pathogenesis of autoimmune diseases such as systemic lupus erythematosus (SLE). In fact, the relationship between DNA methylation and SLE has long been the subject of investigation. To obtain a deeper understanding of the role of DNA methylation in the onset of SLE, we reviewed the findings reported in the literature and our own data about DNA methylation and SLE. Various studies have indicated the possible importance of DNA methylation, especially hypomethylation, in the aetiology of SLE. Epigenetic studies may provide clues for elucidating the pathogenesis of SLE and for developing new strategies to treat this disease.