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Background and purpose There is no general consensus as to whether autoimmune myasthenia gravis ( MG) is associated with heart diseases, despite the fact that myocarditis, a serious cardiac involvement treatable by immunotherapy, is a complication of MG. It has been observed previously that MG patients with clinically suspected myocarditis had anti-Kv1.4 antibodies. The purpose of this study was to disclose the association between anti-Kv1.4 antibodies and cardiac involvements in MG patients. Methods Anti-Kv1.4 antibody was detected by an immunoprecipitation assay using 35S-labeled rhabdomyosarcome cellular extract as the antigen source. Cardiac findings including electrocardiography ( ECG) and clinical features of clinically suspected myocarditis in MG patients with anti-Kv1.4 antibodies were investigated. Ultrasound echocardiography ( UCG) of ex vivo chick embryos was performed to determine the suppressive effects of sera with or without anti-Kv1.4 antibodies on heart muscle functions. Results Seventy (10.8%) of 650 MG patients had anti-Kv1.4 antibodies and 60% of them had abnormal ECG findings with high frequencies of T-wave abnormality and QT prolongation. Clinically suspected myocarditis was found in eight MG patients with anti-Kv1.4 antibodies but in none of the MG patients without anti-Kv1.4 antibodies. Most patients showed rapid deterioration with lethal arrhythmias such as ventricular tachycardia, sick sinus syndrome, or complete atrial ventricular block and severe heart failure. It was concluded using UCG of ex vivo chick embryos that MG serum with anti-Kv1.4 antibodies suppressed heart muscle functions. Conclusion It has been demonstrated that anti-Kv1.4 antibodies are possible markers for cardiac involvements in MG patients. [ABSTRACT FROM AUTHOR]

Immune thrombocytopenia ( ITP) is an autoimmune disease mediated by IgG antiplatelet autoantibodies, resulting in an isolated thrombocytopenia. The mechanism for ongoing antiplatelet antibody production is explained by a 'pathogenic loop' model consisting of macrophages in the reticuloendothelial system, platelet-reactive CD4+ T cells and B cells producing IgG antiplatelet antibodies. In ITP patients, a variety of negative immune regulators including CD4+ T regulatory cells, B regulatory cells and tolerogenic dendritic cells are dysfunctional, resulting in failure to efficiently suppress the pathogenic loop. In addition, Helicobacter pylori infection leads to defective inhibitory Fc γ RIIB signalling in macrophages and thereby increases susceptibility to ITP. In ITP patients, dysregulation of these negative immune regulators is associated with each other in the impaired immune regulatory network. Thus, strategies that enhance functions of these intrinsic negative immune regulators would be promising future approaches for treating ITP. [ABSTRACT FROM AUTHOR]

Active oxygens and free radicals are involved in the metabolism and clinical conditions of tissues; however, little is known about the localisation and expression levels of associated enzymes. The expressions of active oxygens, free radicals and associated enzymes are reported to be site-specific; therefore, the expression states of free radical enzymes differ between sites, even within the same cell. In particular, there has been no report concerning the catabolic enzymes of active oxygens in osteoblasts of the maxilla, other than normal osteoblasts that were weakly positive by immunohistochemical staining. We conducted this study to elucidate the relationship between osteodystrophy and radical-associated enzymes by investigating mRNAs of enzymes associated with active oxygens and free radicals using osteoblasts from the maxilla of normal and osteopetrotic model (op/op) mice. In op/op mouse maxilla osteoblasts, mRNAs of Mn-SOD, i-NOS and e-NOS were strongly positive. Mn-SOD and iNOS enzymes were considered to be highly expressed in osteoblasts of the narrowed medullary cavity of this bone. [ABSTRACT FROM AUTHOR]

Although the HLA region contributes to one-third of the genetic factors affecting rheumatoid arthritis (RA), there are few reports on the association of the disease with any of the HLA loci other than the DRB1. In this study we examined the association between RA and the alleles of the six classical HLA loci including DRB1. Six HLA loci ( HLA-A, -B, -C, -DRB1, -DQB1 and -DPB1) of 1659 Japanese subjects (622 cases; 488 anti-cyclic citrullinated peptides (CCP) antibody (Ab) positive (82.6%); 103 anti-CCP Ab negative (17.4%); 31 not known and 1037 controls) were genotyped. Disease types and positivity/negativity for CCP autoantibodies were used to stratify the cases. Statistical and genetic assessments were performed by Fisher's exact tests, odds ratio, trend tests and haplotype estimation. None of the HLA loci were significantly associated with CCP sero-negative cases after Bonferroni correction and we therefore limited further analyses to using only the anti CCP-positive RA cases and both anti-CCP positive and anti-CCP negative controls. Some alleles of the non- DRB1 HLA loci showed significant association with RA, which could be explained by linkage disequilibrium with DRB1 alleles. However, DPB1*02:01, DPB1*04:01 and DPB1*09:01 conferred RA risk/protection independently from DRB1. DPB1*02:01 was significantly associated with the highly erosive disease type. The odds ratio of the four HLA-loci haplotypes with DRB1*04:05 and DQB1*04:01, which were the high-risk HLA alleles in Japanese, varied from 1.01 to 5.58. C*07:04, and B*15:18 showed similar P-values and odds ratios to DRB1*04:01, which was located on the same haplotype. This haplotype analysis showed that the DRB1 gene as well as five other HLA loci is required for a more comprehensive understanding of the genetic association between HLA and RA than analyzing DRB1 alone. [ABSTRACT FROM AUTHOR]

OBJECTIVE: To identify the autoantigen recognized by the autoantibody that is associated with clinically amyopathic dermatomyositis (C-ADM) and rapidly progressive interstitial lung disease (ILD). METHODS: An anti-CADM-140 antibody-positive prototype serum sample was used to screen a HeLa cell-derived complementary DNA (cDNA) library. Selected cDNA clones were further evaluated by immunoprecipitation of their in vitro-transcribed and in vitro-translated products using anti-CADM-140 antibody-positive and anti-CADM-140 antibody-negative sera. The lysates of COS-7 cells transfected with the putative antigen were similarly tested. An enzyme-linked immunosorbent assay (ELISA) to detect the anti-CADM-140 antibody was established using a recombinant CADM-140 antigen, and its specificity and sensitivity for C-ADM and rapidly progressive ILD were assessed in 294 patients with various connective tissue diseases. RESULTS: By cDNA library screening and immunoprecipitation of in vitro-transcribed and in vitro-translated products, we obtained a cDNA clone encoding melanoma differentiation-associated gene 5 (MDA-5). The anti-CADM-140 antibodies in patients' sera specifically reacted with MDA-5 protein expressed in cells transfected with full-length MDA-5 cDNA, confirming the identity of MDA-5 as the CADM-140 autoantigen. The ELISA, using recombinant MDA-5 protein as the antigen, showed an analytical sensitivity of 85% and analytical specificity of 100%, in comparison with the 'gold standard' immunoprecipitation assay, and was useful for identifying patients with C-ADM and/or rapidly progressive ILD. CONCLUSION: Given that RNA helicase encoded by MDA-5 is a critical molecule involved in the innate immune defense against viruses, viral infection may play an important role in the pathogenesis of C-ADM and rapidly progressive ILD. Moreover, our ELISA using recombinant MDA-5 protein makes detection of the anti-CADM-140 antibody routinely available. [ABSTRACT FROM AUTHOR]

To examine the role of genetic factors in development of immune thrombocytopenic purpura (ITP) in association with Helicobacter pylori infection, gene polymorphisms within the loci for human leukocyte antigen class II, interleukin (IL)-1β (−511), tumor necrosis factor-β (+252), immunoglobulin (Ig)G1 heavy chain (+643), and Igκ light chain (+573) were determined in 164 adults with ITP and 75 healthy controls. Of these gene polymorphisms, the IL-1β (−511) T allele was less frequently detected in H. pylori-infected than in H. pylori-uninfected (58% vs 81%, P = 0.01, odds ratio = 0.31) ITP patients diagnosed before age 50. These findings suggest that a single nucleotide polymorphism within the IL-1β (−511) may affect susceptibility to early-onset ITP associated with H. pylori infection. [ABSTRACT FROM AUTHOR]

OBJECTIVE: To examine the role of an interaction between fibroblasts and mononuclear infiltrates through CD40-CD154 engagement in the development of tissue fibrosis. METHODS: Cultured dermal fibroblasts derived from healthy skin were induced to express CD40 by adenoviral gene transfer, stimulated with soluble CD154, and evaluated for proliferation, gene expression, and protein expression in vitro. The skin of mice with bleomycin-induced skin sclerosis, a model for systemic sclerosis (SSc), was assessed for CD40 and CD154 expression, in vivo fibroblast proliferation, and the expression of specific genes. The effects of an anti-CD154 monoclonal antibody on bleomycin-induced skin sclerosis were also examined. RESULTS: Upon stimulation with soluble CD154, cultured fibroblasts induced to express CD40 by adenoviral gene transfer proliferated and showed up-regulation of the genes for intercellular adhesion molecule 1, interleukin-6 (IL-6), IL-8, monocyte chemoattractant protein 1 (MCP-1), and RANTES, as well as up-regulation of their proteins. In the skin from bleomycin-treated mice, dermal fibroblasts expressed CD40, and mast cells and CD4+ T cells expressed CD154. Electron microscopic analysis revealed fibroblasts attached to mast cells and T cells with primitive contacts. The proliferation of fibroblasts and the up-regulated MCP-1 gene expression preceded thickening of the dermis. Finally, the anti-CD154 antibody inhibited the bleomycin-induced skin sclerosis by suppressing fibroblast proliferation and down-regulating MCP-1 expression. CONCLUSION: The interaction between fibroblasts and mast cells or T cells through CD40-CD154 signaling is critical for fibroblast activation early in the course of fibrosis. Blockade of the CD40-CD154 signal may be a novel therapeutic strategy for human fibrotic diseases, such as SSc. [ABSTRACT FROM AUTHOR]

OBJECTIVE: To evaluate whether atorvastatin can increase bone marrow-derived circulating endothelial precursors (CEPs) and improve the vascular symptoms in patients with systemic sclerosis (SSc; scleroderma). METHODS: The study was designed as an open-label, prospective study involving 14 patients with SSc who received 10 mg/day of atorvastatin for 12 weeks and were followed up for the subsequent 4 weeks. CEPs were quantified at weeks 0 (pretreatment), 4, 8, 12 (during treatment), and 16 (posttreatment) by cell sorting followed by 3-color flow cytometry. Raynaud's phenomenon variables, global measures, and psychological scales as well as circulating angiogenic factors and endothelial activation/injury markers were serially assessed. The potential of CEPs to differentiate into mature endothelial cells was examined in cultures with angiogenic stimuli. RESULTS: None of the patients experienced an adverse event, but 1 dropped out because of an excessive decrease in serum total cholesterol. Atorvastatin treatment resulted in a 1.7- to 8.0-fold increase in CEPs from baseline levels (P < 0.0001), but the numbers returned to within baseline levels at posttreatment. However, 8 patients (62%) experienced a gradual decrease in the number of CEPs, even while taking atorvastatin. Variables indicating the extent of Raynaud's phenomenon improved significantly, and up-regulated levels of angiogenic factors and vascular endothelial activation/injury markers decreased significantly during atorvastatin treatment. These variables returned to within baseline levels after discontinuation of the drug. In contrast, atorvastatin failed to improve the in vitro maturation potential of CEPs. CONCLUSION: The results of this pilot study suggest that atorvastatin treatment can increase CEPs and may be effective in improving Raynaud's phenomenon, even in SSc patients who have CEP dysfunction intrinsically. [ABSTRACT FROM AUTHOR]

OBJECTIVE: We have recently developed an enzyme-linked immunosorbent assay (ELISA) for detection of anti-RNA polymerase III (anti-RNAP III) antibody, using a recombinant fragment containing the immunodominant epitope as the antigen source. This study was conducted to assess the analytical accuracy and clinical associations of the anti-RNAP III ELISA in patients with systemic sclerosis (SSc). METHODS: To evaluate analytical sensitivity and specificity of the ELISA, both immunoprecipitation tests and ELISA were used to detect anti-RNAP III antibody in 534 SSc sera from patients at 3 medical centers. Sera from 522 SSc patients and 516 controls, including patients with other connective tissue diseases and blood bank donors, were also evaluated to assess the clinical sensitivity and specificity of the ELISA. Clinical findings in anti-RNAP III antibody-positive SSc patients were compared between patient groups stratified according to anti-RNAP III antibody levels determined by the ELISA. RESULTS: In SSc patients, our ELISA showed analytical sensitivity of 91% and analytical specificity of 99% compared with the immunoprecipitation assay (a gold standard for detection of anti-RNAP III antibody). The additional analysis using a large series of SSc and control sera showed that clinical sensitivity and specificity of the ELISA with respect to the diagnosis of SSc were 17% and 98%, respectively. A high level of anti-RNAP III antibody was associated with diffuse cutaneous SSc, higher maximum total skin score, and increased frequency of tendon friction rubs. CONCLUSION: The anti-RNAP III ELISA is analytically accurate and clinically specific. With this assay, testing for anti-RNAP III antibody can be made routinely available. [ABSTRACT FROM AUTHOR]

The purpose of this study was to evaluate the clinical characteristics of patients who had both myasthenia gravis (MG) and alopecia areata (AA). Clinical information was retrospectively collected for 159 Japanese patients with MG. Human leukocyte antigen (HLA)-DQB1 and DRB1 alleles were determined by genotyping. Of 159 MG patients, six (3.7%) developed AA after the onset of MG and thymectomy. The prevalence of AA in MG patients was higher than that reported in Caucasians. The frequencies of bulbar involvement, myasthenic crisis, and thymoma were significantly higher in MG patients with AA than in those without ( P = 0.007, 0.004, and 0.006, respectively). All but one patient with AA had advanced stage thymoma. Three patients with a severe form of AA (alopecia totalis) had additional autoimmune diseases: myocarditis, myositis, and pure red cell aplasia. DRB1*0901 and DQB1*0303 tended to be more frequently detected in the six MG patients with AA than in the 82 patients without it. In conclusion, a subset of MG patients who have severe neuromuscular symptoms and thymoma develop AA several years after thymectomy. [ABSTRACT FROM AUTHOR]

Sato N, Kamata T, Akiyama N, Kuwana M, Kanda T (Tokyo Metropolitan Bokutoh Hospital, Tokyo; Keio University School of Medicine, Tokyo; and Yamaguchi University School of Medicine, Ube; Japan). Acute inflammatory sensorimotor polyradiculoneuropathy associated with immune thrombocytopenic purpura (Case Report).J Intern Med2005;257:473–477.Although acute inflammatory polyneuropathy (AIP) and immune thrombocytopenic purpura (ITP) are both believed to be immune-mediated disorders, only a few cases have been reported in which these two diseases co-existed. We describe a case of a 67-year-old patient who developed quadriparesis, ophthalmoplegia and severe sensory impairment along with thrombocytopenia. Detailed examinations, including the measurement of anti-ganglioside antibodies and anti-glycoprotein-IIb-IIIa-IgG-producing B-cells, revealed that he developed AIP and ITP. By reviewing past similar reports, we noticed that AIP associated with ITP tends to manifest severe sensory impairment and is often preceded by upper respiratory tract infection, but not by gastrointestinal infection. [ABSTRACT FROM AUTHOR]

OBJECTIVE: To identify novel autoantibodies specific for dermatomyositis (DM), especially those specific for clinically amyopathic DM (C-ADM). METHODS: Autoantibodies were analyzed by immunoprecipitation in 298 serum samples from patients with various connective tissue diseases (CTDs) or idiopathic pulmonary fibrosis (IPF). Antigen specificity of the sera was further examined by immunoblotting and indirect immunofluorescence (IF). The disease specificity and clinical features associated with the antibody of interest were determined. RESULTS: Eight sera recognized a polypeptide of approximately 140 kd (CADM-140 autoantigen) by immunoprecipitation and immunoblotting. Immunoreactivity was detected in the cytoplasm, and indirect IF revealed a granular or reticular pattern. Anti-CADM-140 antibodies were detected in 8 of 42 patients with DM, but not in patients with other CTDs or IPF. Interestingly, all 8 patients with anti-CADM-140 antibodies had C-ADM. Among 42 patients with DM, those with anti-CADM-140 autoantibodies had significantly more rapidly progressive interstitial lung disease (ILD) when compared with patients without anti-CADM-140 autoantibodies (50% versus 6%; P = 0.008). CONCLUSION: These results indicate that the presence of anti-CADM-140 autoantibodies may be a novel marker for C-ADM. Further attention should be directed to the detection of rapidly progressive ILD in those patients with anti-CADM-140 autoantibodies. [ABSTRACT FROM AUTHOR]

HLA class II alleles and immunoglobulin allotypes were determined in 83 Japanese patients with immune thrombocytopenic purpura (ITP) and 114 race-matched healthy controls. Distribution of DRB and DQB1 alleles as well as G1M(f, z), G2M(n¹,n...), and KM(1, (1,2), 3) were not different between ITP patients and healthy controls, while DPB1*0201 was marginally increased in ITP patients vs. healthy controls (51% vs. 28%, Pc5 0.04, OR52.6 [1.4--4.8]). In contrast, strong associations between anti-glyco-protein autoantibodies and HLA class II genes were found as follows: anti-GPIIb- IIIa antibody with DRB1*0405 and DQB1*0401; and anti-GPIb-IX antibody with DRB1*0803 and DQB1*0601. When factors influencing therapeutic responses to splenectomy were examined, a poor response was correlated with the presence of DRB1*0405, DQB1*0401 and anti-GPIIb-IIIa antibody (P50.01, 0.002, and 0.03, respectively). Our results indicate that HLA class II genes influence the production of anti-glycoprotein antibody specificities rather than the development of ITP. In addition, HLA class II genotyping could be useful in predicting therapeutic responses to splenectomy in Japanese patients with ITP. [ABSTRACT FROM AUTHOR]

Fostamatinib, a spleen tyrosine kinase inhibitor, has been approved for the treatment of chronic primary immune thrombocytopenia (ITP) in the United States, Canada and some European countries. We conducted a phase 3, placebo-controlled, double-blind, parallel-group study to evaluate the efficacy and safety of fostamatinib in Japanese patients with primary ITP. Thirty-four patients were randomised to fostamatinib (n = 22) or placebo (n = 12) at 100-150 mg twice a day for 24 weeks. Stable responses (platelet ≥50 000/μl at ≥4 of the 6 visits from weeks 14 to 24) were observed in eight (36%) patients on fostamatinib and in none of the patients on placebo (p = 0.030). Overall responses (platelet ≥50 000/μl at ≥1 of the 6 visits from weeks 2 to 12) were seen in 10 (45%) patients on fostamatinib and in none of the patients on placebo (p = 0.006). Patients on fostamatinib required rescue medication less often and experienced fewer bleeding symptoms than patients on placebo. Adverse events observed were mild or moderate and were manageable. No new safety signals were identified in Japanese patients with ITP., (© 2022 The Authors. British Journal of Haematology published by British Society for Haematology and John Wiley & Sons Ltd.)

Several effective drugs have been identified for the treatment of systemic sclerosis (SSc). However, in advanced cases, not only their effectiveness is reduced but they may be also harmful due to their side-effects. Therefore, early diagnosis and early treatment is most important for the treatment of SSc. We established diagnostic criteria for SSc in 2003 and early diagnostic criteria for SSc in 2011, for the purpose of developing evaluation of each organ in SSc. Moreover, in November 2013, the American College of Rheumatology and the European Rheumatology Association jointly developed new diagnostic criteria for increasing their sensitivity and specificity, so we revised our diagnostic criteria and severity classification of SSc. Furthermore, we have revised the clinical guideline based on the newest evidence. In particular, the clinical guideline was established by clinical questions based on evidence-based medicine according to the New Minds Clinical Practice Guideline Creation Manual (version 1.0). We aimed to make the guideline easy to use and reliable based on the newest evidence, and to present guidance as specific as possible for various clinical problems in treatment of SSc., (© 2018 Japanese Dermatological Association.)

A multicenter, open-label study was performed to investigate the safety and tolerability of bosentan in Japanese patients with systemic sclerosis (SSc) and secondary digital ulcers. Twenty-eight patients were enrolled. The safety and tolerability of bosentan was monitored over 52 weeks of study treatment (primary end-point), while incidence and healing of digital ulcers were also assessed up to week 16. The following adverse events occurred in 5% or more of patients during the 52-week treatment period: upper respiratory tract infection (50.0%), abnormal liver function tests (42.9%), digital ulcers (25.0%), anemia (17.9%), peripheral edema (14.3%), diarrhea (10.7%), urinary tract infection (7.1%), arthralgia (7.1%), constipation (7.1%) and herpes zoster (7.1%). Eight patients experienced at least one serious adverse event, including drug-related serious adverse events in two patients, which were abnormal liver function tests and fluid retention (pericardial effusion) in one patient each. During the 16-week observation period, seven out of 28 patients (25%) developed new digital ulcers. In this study, adverse events were comparable with those previously reported with bosentan. Approximately half of the patients had adverse events associated with abnormal liver function tests, thus we conclude that liver function should be monitored regularly during treatment with bosentan., (© 2016 Japanese Dermatological Association.)

Objective: Systemic sclerosis (SSc) is an autoimmune disease for which multiple susceptibility genes have been reported. Genome-wide association studies have shown that large numbers of susceptibility genes are shared among autoimmune diseases. Recently, our group identified 9 novel susceptibility genes associated with rheumatoid arthritis (RA) in a Japanese population. The aim of this study was to elucidate whether the 18 genes that displayed associations or suggestive associations for RA in our previous study are associated with SSc in Japanese., Methods: We performed an association study that included 415 patients with SSc and 16,891 control subjects, followed by a replication study that included 315 patients and 21,054 control subjects. The 18 markers reported to display association with RA were analyzed for their associations with SSc in the first study, and 5 markers were further analyzed in the replication study. The inverse variance method was used to evaluate the associations of these markers with SSc in a combined study., Results: In the phospholipase D4 gene (PLD4), rs2841277 displayed a significant association with SSc in Japanese patients (P = 0.00017). We observed that rs2841280 in exon 2 of PLD4 was in strong linkage disequilibrium with rs2841277 and introduced an amino acid alteration. We also observed associations between SSc and rs6932056 in TNFAIP3 and rs2280381 in IRF8 (P = 0.0000095 and P = 0.0030, respectively), both of which displayed associations with SSc in a European population., Conclusion: We determined that PLD4 is a novel susceptibility gene for SSc in Japanese, thus confirming the involvement of PLD4 in autoimmunity. Associations between SSc and TNFAIP3 or IRF8 were also detected in our Japanese population. SSc and RA appear to share relatively large proportions of their genetic backgrounds., (Copyright © 2013 by the American College of Rheumatology.)

Objective: The complication of interstitial lung disease (ILD) in polymyositis/dermatomyositis (PM/DM) is associated with anti-aminoacyl-transfer RNA synthetase (anti-aaRS) antibody or anti-melanoma differentiation-associated gene 5 (anti-MDA-5) antibody positivity. Anti-MDA-5 antibody is associated with clinically amyopathic DM and fatal outcome due to rapidly progressive ILD in Asian populations. The association between genetic factors and anti-MDA-5 antibody-positive DM is unclear. This study was undertaken to investigate the HLA-DRB1 genotype in patients with anti-MDA-5 antibody-positive DM., Methods: We examined genetic differences among 17 patients with anti-MDA-5 antibody-positive DM, 33 patients with anti-aaRS antibody-positive PM/DM, 33 patients with PM/DM without anti-aaRS antibody or ILD, and 265 healthy controls., Results: The frequencies of HLA-DRB1*0101 and DRB1*0405 were 29% and 71%, respectively, in patients with anti-MDA-5 antibody-positive DM, which were higher than the frequencies in healthy controls (10% and 25%, respectively). Among the 17 patients with anti-MDA-5 antibody-positive DM, 16 (94%) harbored either the DRB1*0101 or DRB1*0405 allele. The combined frequency of the DRB1*0101 allele and the DRB1*0405 allele was significantly higher in patients with anti-MDA-5 antibody-positive DM than in patients with PM/DM without anti-aaRS antibody or ILD, with an odds ratio (OR) of 42.7 (95% confidence interval [95% CI] 4.9-370.2) (P = 1.1 × 10(-5)), or in patients with anti-aaRS antibody-positive PM/DM (OR 13.3 [95% CI 1.6-112.6], P = 4.5 × 10(-3))., Conclusion: Our findings indicate that HLA-DRB1*0101/*0405 is associated with susceptibility to anti-MDA-5 antibody-positive DM in the Japanese population., (Copyright © 2012 by the American College of Rheumatology.)

Dermatomyositis (DM) is an idiopathic systemic inflammatory disease that is often accompanied by interstitial lung disease (ILD) or internal malignancy. New autoantibodies, anti-clinically amyopathic dermatomyositis 140 (anti-CADM-140) antibody (Ab) and anti-155/140 Ab, as well as anti-aminoacyl-tRNA synthetase (anti-ARS) Ab and anti-Mi-2 Ab, have been discovered and their utility indicated. However, the association between these autoantibodies and the clinical characteristics of DM is not fully understood, and it is unclear whether anti-155/140 Ab is "specific" to DM patients with internal malignancy. Therefore, we analyzed 55 DM patients and 18 non-DM patients with malignancy to evaluate the clinical characteristics, especially skin manifestations, in association with DM-specific autoantibodies detected by immunoprecipitation. Six patients (11%) had anti-CADM-140 Ab, nine (16%) had anti-155/140 Ab, eight (15%) had anti-ARS Ab and six (11%) had anti-Mi-2 Ab. The frequency of DM patients positive for any type of autoantibody was 53%. Among the 20 DM patients with ILD, three (15%) had both anti-CADM-140 Ab and rapidly progressive ILD, and required intensive therapy (P < 0.05). ILD found in anti-ARS Ab-positive patients did not progress rapidly. The prevalence of muscle involvement in patients with anti-CADM-140 Ab was 83%. Among the 18 DM patients with internal malignancy, four (22%) had anti-155/140 Ab, and internal malignancy was found in four cases (44%) of nine anti-155/140 Ab-positive patients. None of the non-DM patients with malignancy were positive for anti-155/140 Ab. In conclusion, the results of the present study indicate that anti-155/140 Ab is specific to DM patients with internal malignancy and that we may be able to predict prognosis of ILD and the presence of malignancy to some extent, suggesting that examination of autoantibodies in DM patients is clinically very useful. However, further investigation is needed because several findings differ from those of previous reports., (© 2011 Japanese Dermatological Association.)

Objective: To examine associations of single-nucleotide polymorphisms (SNPs) within genes for hepatocyte growth factor (HGF) and its receptor c-met with disease susceptibility and organ involvement in Japanese patients with systemic sclerosis (SSc)., Methods: Four SNPs (HGF -1652 C/T, +44222 C/T, and +63555 G/T, and c-met -980 T/A) were analyzed in 159 SSc patients and 103 healthy control subjects with the use of a polymerase chain reaction-based assay. The influence of the HGF -1652 SNP on transcription activity was evaluated with a luciferase reporter assay and an electrophoretic mobility shift assay (EMSA)., Results: There was no difference in the distribution of HGF/c-met SNPs between SSc patients and controls. HGF -1652 TT was found much more frequently in SSc patients with end-stage lung disease (ESLD) than in those without (41% versus 8%; P = 0.0004). This association was confirmed by a replication study involving a separate cohort of 155 SSc patients. Kaplan-Meyer analysis revealed that HGF -1652 TT carriers had a higher probability of developing ESLD than did CT or CC carriers. The HGF promoter carrying the HGF -1652 T allele had lower transcription activity than did the promoter carrying the C allele. EMSA showed the presence of a potential negative transcription regulator that binds specifically to the HGF promoter carrying a T allele at position -1652. Finally, TT carriers had a relative inability to increase circulating HGF levels even in the presence of advanced interstitial lung disease., Conclusion: A SNP in the HGF promoter region may modulate the severity of interstitial lung disease by controlling the transcriptional efficiency of the HGF gene., (Copyright © 2011 by the American College of Rheumatology.)

Objective: To detect and characterize the autoreactive CD8+ T cells to major histocompatibility complex class I chain-related gene A (MICA), a stress-inducible antigen preferentially expressed on the epithelium and endothelium, in patients with Behcet's disease (BD)., Methods: A candidate for the antigenic MICA peptide was selected based on its predicted binding affinity for HLA-B51 and proteasomal cleavage sites. Peripheral blood T cells from 14 patients with BD and 15 healthy controls were repeatedly stimulated with the MICA peptide, and the specific T cell response was measured by peptide-induced interferon-gamma. Cytotoxic T lymphocyte activity was examined by chromium-51 release from an HLA-B51-transfected B cell line in the presence of the MICA peptide., Results: A 9-mer peptide AAAAAIFVI (termed MICA transmembrane [MICA-TM]) was selected as a candidate for the antigenic peptide presented by HLA-B51. A specific T cell response to MICA-TM was detected in 4 patients with BD (29%) but in none of the 15 healthy donors. All 4 responders had HLA-B51 and active disease, and the specific T cell response was lost after the BD-related symptoms disappeared. The MICA-induced T cell response was specifically inhibited by anti-HLA class I antibody or by CD8+ cell depletion. MICA-reactive T cells recognized an HLA-B51-transfected B cell line pulsed with MICA-TM or a B cell line transfected with both HLA-B51 and MICA in the absence of exogenous peptides. Finally, MICA-stimulated T cell lines lysed the HLA-B51-expressing B cell line in the presence of MICA-TM., Conclusion: HLA-B51-restricted cytotoxic T lymphocytes autoreactive to MICA may be involved in the pathogenesis of BD.

Single nucleotide polymorphisms (SNPs) of inflammatory cytokine genes were examined in 84 adult Japanese patients with chronic immune thrombocytopenic purpura (ITP) and 56 race-matched healthy controls. The SNPs examined were within the genes encoding tumour necrosis factor (TNF)-alpha (-238 G/A and -308 G/A), TNF-beta (+252 G/A), and interleukin (IL)-1beta (-511 C/T and +3953 T/C). Of these SNPs, the frequency of the TNF-beta (+252) G/G phenotype was significantly higher in ITP patients than in healthy controls (21% vs. 7%, P = 0.04, odds ratio = 3.6, 95% confidence interval 1.1-11.1), while no significant association was detected for the other SNPs. The distribution of the TNF-beta (+252) phenotype was not associated with human leucocyte antigen class II alleles or the therapeutic response in ITP patients. The frequency of circulating anti-glycoprotein IIb/IIIa antibody-producing B cells was significantly higher in ITP patients with the TNF-beta (+252) G/G phenotype than in those with the G/A or A/A phenotype (11.9 +/- 4.9 vs. 6.8 +/- 4.9 and 3.7 +/- 2.8 per 10(5) peripheral blood mononuclear cells; P = 0.02 and P < 0.001, respectively). These findings suggest that the SNP located at TNF-beta (+252) contributes to susceptibility to chronic ITP by controlling the autoreactive B-cell responses to platelet membrane glycoproteins.

Amegakaryocytic thrombocytopenia (AMT) associated with systemic sclerosis (SSc) has been described in several case reports, but the underlying mechanisms have not been identified. Here we describe a rare case of SSc accompanied by thrombocytopenia and megakaryocytic hypoplasia, in which autoantibody against thrombopoietin receptor (c-Mpl) was detected. A 61-year-old woman with limited SSc was admitted to our hospital because of severe thrombocytopenia (platelet count 0.2 x 10(4)/mm(3)) with gingival bleeding. Her bone marrow was hypocellular with absent megakaryocytes, consistent with AMT. Treatment with corticosteroids and intravenous immunoglobulin infusions resulted in an increased platelet count, and she sustained a remission over a 1-year period, with a platelet count averaging 10.0 x 10(4)/mm(3). Her serum was strongly positive for anti-c-Mpl antibody, and IgG fraction purified from her serum inhibited thrombopoietin-dependent cell proliferation in vitro. Our case report suggests that AMT in patients with SSc could be mediated by the anti-c-Mpl antibody, which functionally blocks an interaction between thrombopoietin and c-Mpl.

Objective: To characterize an immunodominant epitope on RNA polymerase III (RNAP III) recognized by systemic sclerosis (SSc) sera and to develop an enzyme-linked immunosorbent assay (ELISA) for the detection of serum anti-RNAP I/III antibodies., Methods: RNAP III-specific subunits RPC62 and RPC155 were generated in a bacterial expression system as a series of recombinant fragments. Reactivities to these recombinant fragments were examined by immunoblots and/or ELISA in 16 SSc sera containing anti-RNAP I/III antibodies, 89 SSc sera lacking anti-RNAP I/III antibodies, 61 systemic lupus erythematosus (SLE) sera, and 61 healthy control sera., Results: Anti-RNAP I/III-positive SSc sera recognized several distinct epitopes on RPC62 and RPC155 in various combinations, but the fragment encoding amino acids at positions 732-1166 of RPC155 was recognized by all 11 anti-RNAP I/III-positive SSc sera tested. Carboxyl- and amino-terminal deletion studies showed that at least 130 amino acids at positions 891-1020 of RPC155 were necessary for the antibody binding, but strong reactivity required an additional amino-terminal extension. When a purified recombinant fragment containing the immunodominant epitope was used as the antigen source in an ELISA, elevated antibody reactivity was detected in all 16 anti-RNAP I/III-positive SSc sera, but in no anti-RNAP I/III-negative SSc, SLE, or healthy control sera, representing a sensitivity of 100% and a specificity of 100%., Conclusion: A major epitope commonly recognized by SSc sera containing anti-RNAP I/III antibodies was identified on RPC155. The ELISA using a recombinant fragment expressing the immunodominant epitope should be a valuable tool for routine screening for anti-RNAP I/III antibodies in clinical diagnostic laboratories.

Objective: To examine the prevalence, clinical associations, and pathogenic role of autoantibodies to c-Mpl, the thrombopoietin (TPO) receptor, in patients with systemic lupus erythematosus (SLE)., Methods: Sera from 69 SLE patients, 84 patients with idiopathic thrombocytopenic purpura (ITP), and 60 healthy individuals were screened for anti-c-Mpl antibodies by enzyme-linked immunosorbent assay using recombinant c-Mpl as an antigen. Clinical findings, autoantibody profiles, and serum TPO levels were compared between SLE patients with and without anti-c-Mpl antibodies. A pathogenic role for the anti-c-Mpl antibody was evaluated by examining its inhibitory effect on TPO-dependent cell proliferation and megakaryocyte colony formation., Results: Serum anti-c-Mpl antibody was detected in 8 SLE patients (11.6%) and 7 ITP patients (8.3%), but in none of the healthy controls. Anti-c-Mpl antibody was associated with thrombocytopenia (P = 0.0002) and a decrease in bone marrow megakaryocytes (P = 0.02) in SLE patients. Serum TPO levels in thrombocytopenic SLE patients with anti-c-Mpl antibodies were significantly elevated compared with levels in those without the antibodies (P = 0.007). IgG fractions purified from anti-c-Mpl antibody-positive sera bound to c-Mpl expressed on the cell surface and inhibited TPO-dependent cell proliferation and megakaryocyte colony formation., Conclusion: Autoantibody to c-Mpl is present in a subset of SLE patients with thrombocytopenia and megakaryocytic hypoplasia. It is likely that the impaired thrombopoiesis in these patients is mediated by the anti-c-Mpl antibody, which functionally blocks an interaction between TPO and c-Mpl.

Objective: To examine T and B cell responses to topoisomerase I (topo I) in a monozygotic twin pair discordant for systemic sclerosis (SSc)., Methods: The peripheral blood T cell proliferative responses induced by topo I and in vitro anti-topo I antibody production in cultures of T and B cells were examined in an SSc patient with serum anti-topo I antibody and in her healthy monozygotic twin. Topo I-reactive T cell lines were generated from the twin pair and analyzed for antigenic specificity, major histocompatibility complex class II restriction, and T cell receptor (TCR) gene usage., Results: T cell proliferative responses to topo I were detected in both the SSc patient and her healthy twin, although the kinetics of the T cell response were accelerated in the patient compared with the healthy twin. The estimated frequency of circulating topo I-reactive T cells was 1/6,700 in the SSc patient and 1/39,000 in the healthy twin. Anti-topo I antibody production was observed in cultures of T and B cells from the SSc patient, but not in those from the healthy twin. When the cells from the twins were mixed in different combinations, T cells from the healthy twin did stimulate the SSc patient's B cells to produce anti-topo I antibody through a CD40-dependent mechanism. Topo I-reactive T cell lines generated from the twins had similar characteristics, including a CD4+ phenotype, restriction by HLA-DR, recognition of epitopes within amino acid residues 209-386 of topo I, and dominant usage of the TCR Vbeta20 gene segment., Conclusion: These results indicate that topo I-reactive T cells were activated and clonally expanded in the SSc patient. However, there were no substantial differences in either phenotypic or functional properties of topo I-reactive T cells obtained from the SSc patient and those obtained from her healthy identical twin. It is likely, therefore, that the anti-topo I antibody response in the SSc patient is induced by in vivo activation of topo I-reactive T cells derived from the normal T cell repertoire.

Objective: Previously, we demonstrated with the use of microsatellite markers that a 2-cM haplotype on chromosome 15q containing the fibrillin 1 gene (FBN1) was strongly associated with systemic sclerosis (SSc) in the Choctaw, a population with high SSc prevalence. In this study, all 69 known FBN1 exons were sequenced to ascertain the presence of changes that might show associations with SSc in the Choctaw and Japanese SSc patients and controls., Methods: Screening of FBN1 exons was accomplished by polymerase chain reaction-based fluorescence sequencing of genomic DNA using single-nucleotide polymorphism (SNP) haplotypes, and their frequencies were determined with a new algorithm that recognizes past recombination events between sites. Haplotype phylogenies were inferred using the median-joining network analysis., Results: Five SNPs were identified in FBN1. They are located in the 5'-untranslated region (SNP-1), exon 15 (SNP-2), intron 17 (SNP-3), exon 27 (SNP-4), and intron 27 (SNP-5). Only SNP-1 (T-->C) demonstrated an association with SSc in the Choctaw. Eleven FBN1 SNP haplotypes were ascertained in the Choctaw population, 2 of which (SNPs 5 and 6) were found only in the SSc patients. These same FBN1 SNP haplotypes were associated with SSc in the Japanese., Conclusion: A SNP in the 5'-untranslated region of FBN1 (SNP-1, C allele) was strongly associated with SSc in the Choctaw. Furthermore, this polymorphism is present on 2 unique FBN1 haplotypes found only in Choctaw SSc patients. The same 2 haplotypes demonstrate associations with SSc in the Japanese. These data extend the earlier microsatellite studies and are consistent with the hypothesis that FBN1 or a nearby gene on chromosome 15q is involved in SSc susceptibility in the Choctaw and the Japanese.

Objective: To examine serial changes in serum anti-topoisomerase I (anti-topo I) antibody levels in patients with systemic sclerosis (SSc), as well as associations with clinical features and the in vivo activation status of circulating topo I-reactive T and B cells., Methods: Serum anti-topo I antibody levels were serially measured at different time points in 28 SSc patients who were positive for anti-topo I antibody at their first visit (range of followup 6-29 years). The patients were subgrouped according to the disappearance (group 1) or persistence (group 2) of anti-topo I antibody. Clinical findings as well as T and B cell responses to topo I were compared between these 2 groups., Results: Serum anti-topo I antibody disappeared during the period of followup in 6 patients (group 1), but persisted in 22 patients (group 2). Loss of anti-topo I antibody occurred within 10 years after the first visit and independently of treatment. Group 1 patients had less extensive skin and lung involvement and better survival rates than did group 2 patients. Complete loss of anti-topo I antibody followed a reduction in isotype expression and epitope reactivities. The kinetics of in vitro T cell proliferation induced by topo I were delayed and circulating topo I-reactive T cells were less frequently detected in group 1 versus group 2 patients, suggesting that the disappearance of anti-topo I antibody was due to loss of activation of topo I-reactive T cells. In vitro production of anti-topo I antibody in peripheral blood mononuclear cell cultures in response to antigenic stimulation in both group 1 and group 2 patients indicated persistence of anti-topo I antibody-producing "memory" B cells even after the loss of serum anti-topo I antibody., Conclusion: Our results indicate that there is a distinct subset of anti-topo I-positive SSc patients who lose anti-topo I antibody during the disease course and have a favorable outcome. In vivo production of anti-topo I autoantibody may require antigenic stimulation that activates topo I-reactive T and B cells.

Objective: To identify the T cells responsive to beta2-glycoprotein I (beta2GPI) that mediate antiphospholipid antibody production in patients with antiphospholipid syndrome (APS)., Methods: In vitro proliferative responses and anti-beta2GPI antibody production induced by beta2GPI were examined in peripheral blood mononuclear cell (PBMC) cultures from 12 APS patients, 13 systemic lupus erythematosus patients without APS, and 12 healthy donors., Results: Peripheral blood T cells from all subjects failed to respond to beta2GPI in its native form. In contrast, reduced beta2GPI was able to stimulate T cells not only from all 12 patients with anti-beta2GPI antibodies, but also from 10 of 25 individuals without anti-beta2GPI antibodies. The specificity of the responses to beta2GPI was confirmed by activation of the reduced beta2GPI-primed T cells by recombinant beta2GPI in secondary cultures. Characterization of the T cell response induced by beta2GPI revealed that the response was associated with the presence of the DR53-associated alleles, the responding T cells were CD4+ and restricted by HLA class II, and antigenic peptides were located in domains IV and/or V. Anti-beta2GPI antibody production was induced specifically in anti-beta2GPI antibody-positive patients, in PBMC cultures with reduced beta2GPI. Anti-beta2GPI antibodies produced in vitro recognized beta2GPI immobilized with cardiolipin or beta2GPI coated on "high-binding" polystyrene plates., Conclusion: These results strongly suggest that CD4+ and HLA class II-restricted T cells responsive to beta2GPI are involved in the production of antiphospholipid antibodies in APS patients.

Objective: To characterize an immunodominant epitope recognized by anti-DNA topoisomerase I (topo I) antibody, a major autoantibody in sera of patients with systemic sclerosis (SSc)., Methods: Topo I fragments were generated as fusion proteins using a bacterial expression system as well as polypeptides translated in vitro using a eukaryotic expression system. Reactivities to the 2 preparations of recombinant topo I polypeptides in anti-topo I-positive sera from SSc patients of varied ethnic backgrounds were examined by immunoblotting, immunoprecipitation, and/or enzyme-linked immunosorbent assay., Results: The fragment encoding amino acids 489-573 of topo I was recognized by 98 of 100 anti-topo I-positive SSc sera. Both carboxyl- and amino-terminal deletion studies as well as competitive inhibition assays using topo I synthetic peptides showed that a region of > or =52 amino acids (512-563) was necessary for recognition by anti-topo I antibodies. The minimum epitope region and conformation required for this reactivity were variable among sera from Caucasian, African American, Japanese, and Choctaw SSc patients., Conclusion: An immunodominant epitope recognized by anti-topo I autoantibody is located in the region of amino acids 489-573 of the topo I protein and is largely conformational in nature. The recognition pattern of this region by anti-topo I-positive sera is heterogeneous and is influenced by ethnic background.

Objective: To investigate the effect of ethnicity on clinical and serologic expression in patients with systemic sclerosis (SSc) and anti-DNA topoisomerase I (anti-topo I) antibody., Methods: Clinical and serologic features, as well as HLA class II allele frequencies, were compared among 47 North American white, 15 North American black, 43 Japanese, and 12 Choctaw Native American SSc patients with anti-topo I antibody., Results: The frequency of progressive pulmonary interstitial fibrosis was lower, and cumulative survival rates were better in white compared with black and Japanese patients. Sera of white and black patients frequently recognized the portion adjacent to the carboxyl terminus of topo I, sera of Japanese patients preferentially recognized the portion adjacent to the amino terminus of topo I, and sera of Choctaw patients recognized both portions of topo I. Anti-RNA polymerase II and anti-SSA/Ro antibodies were present together with anti-topo I antibody more frequently in sera of Japanese patients than in sera of white patients. The HLA-DRB1 alleles associated with anti-topo I antibody differed; i.e., DRB1*1101-*1104 in whites and blacks, DRB1*1502 in Japanese, and DRB1*1602 in Choctaws. Multivariate analysis showed that ethnic background was an independent determinant affecting development of severe lung disease as well as survival., Conclusion: Clinical and serologic features in SSc patients were strongly influenced by ethnic background. The variability of disease expression in the 4 ethnic groups suggests that multiple factors linked to ethnicity, including genetic and environmental factors, modulate clinical manifestations, disease course, and autoantibody status in SSc.

Objective: To determine the subunit specificity of autoantibody directed to RNA polymerases (RNAP) I, II, and III, which is one of the major autoantibody responses in patients with systemic sclerosis (SSc)., Methods: Thirty-two SSc sera with anti-RNAP antibodies (23 with anti-RNAP I/III, 5 with anti-RNAP I/III and II, and 4 with anti-RNAP II alone) were analyzed by immunoblotting using affinity-purified RNAP and by immunoprecipitation using 35S-labeled cell extracts in which RNAP complexes were dissociated. Antibodies bound to individual RNAP subunits were eluted from preparative immunoblots and were further analyzed by immunoblotting and immunoprecipitation., Results: At least 15 different proteins were bound by antibodies in anti-RNAP-positive SSc sera in various combinations. All 9 sera immunoprecipitating RNAP II and all 28 sera immunoprecipitating RNAP I/III recognized the large subunit proteins of RNAP II and III, respectively. Reactivity to RNAP I large subunits was strongly associated with bright nucleolar staining by indirect immunofluorescence. Affinity-purified antibodies that recognized a 62-kd subunit protein cross-reacted with a 43-kd subunit protein and immunoprecipitated both RNAP I and RNAP III. Antibodies that recognized a 21-kd subunit protein obtained from sera that were positive for anti-RNAP I/III and II antibodies immunoprecipitated both RNAP II and RNAP III., Conclusion: Anti-RNAP antibodies recognize multiple subunits of RNAP I, II, and III. Moreover, the results of this study provide the first direct evidence that antibodies that recognize shared subunits of human RNAPs or epitopes present on different human RNAP subunits are responsible for the recognition of multiple RNAPs by SSc sera.

Annexin V has a calcium-dependent binding affinity for anionic phospholipids and activated platelets, and prevents prothrombinase activity. We investigated the clinical significance of IgG anti-annexin V antibodies in patients with SLE. The study population consisted of 140 patients with SLE. Sera were examined for IgG anti-annexin V antibodies by ELISA. IgG anti-annexin V antibodies were detected in 27 of 140 patients (19%). Significantly higher incidences of arterial or venous thrombosis, intrauterine fetal loss, and prolonged activated partial thromboplastin time were found in patients with anti-annexin V antibodies than in those without anti-annexin V antibodies. Three patients with thrombosis were found not to have anticardiolipin antibodies, but to show sustained serological reactions for anti-annexin V antibodies, irrespective of prednisolone administration. These results indicated the clinical characteristics of SLE patients with anti-annexin V antibodies, and that these antibodies may be associated with the pathogenesis of thrombotic events.

Objective: To elucidate the roles of HLA genes in the clinical presentation of patients with connective tissue disease and serum anti-U1 small nuclear RNP antibody., Methods: HLA class I antigens and HLA class II alleles were determined in 43 Japanese patients with anti-U1 RNP antibody alone, by microcytotoxicity testing and DNA typing, respectively. Prospectively recorded clinical and laboratory features were analyzed in relation to HLA class I and class II types., Results: DQB1*0303 was associated with lupus-related symptoms including fever, malar rash, oral ulcers, hypocomplementemia, and high-titer anti-double-stranded DNA antibody. Other HLA-clinical associations included DR2 with pleuritis, DR4 with hand swelling, and DRB1*0405 with arthritis., Conclusion: These HLA-clinical associations explain, in part, the heterogeneous clinical presentation in patients with anti-U1 RNP antibody.

Anti-DNA topoisomerase I (anti-topo I) antibody profiles were compared before and after lung cancer in 2 patients with systemic sclerosis (SSc). Both patients developed adenocarcinoma of the lung late in the course of SSc and died of the cancer. Anti-topo I antibody levels, determined by double immunodiffusion and enzyme-linked immunosorbent assay, increased markedly at the time of diagnosis of lung cancer. Furthermore, patients' sera obtained after lung cancer reacted with multiple epitopes on the entire topo I molecule, some of which had not previously been recognized. These results further support the concept that anti-topo I antibody production in SSc patients is due to an antigen-driven process.

Objective: To better define immunogenetic associations with the anti-U1 ribonucleoprotein (U1 RNP) autoantibody response., Methods: HLA class II alleles were determined by genotyping in 49 Japanese rheumatic disease patients with anti-U1 RNP antibody and 43 race-matched healthy controls. Immunoreactivities to U1 RNP constituent proteins (70K, A, B/B', and C) were detected by immunoblots using purified HeLa cell Sm antigen, and antibody titer was determined by passive hemagglutination assay., Results: DQB1*0302 was significantly more frequent in anti-U1 RNP-positive patients than in controls (43% versus 14%; odds ratio [OR] = 4.6, corrected P = 0.03). All anti-U1 RNP-positive patients had either a DQB1*0601, *0602, *0301, *0302, or *0303 allele, which share tyrosine at position 30, and the amino acid sequence Thr, Arg, Ala, Glu, Leu, Asp, and Thr at positions 71-77 in the DQB1 beta 1 domain. In contrast, one of these alleles was found in 81% of the controls (OR = 24, P = 0.002). In addition, anti-U1 RNP antibody was associated with unique DQB1*0302; DRB1*0401 haplotype. Anti-70K reactivity and antibody titer were positively associated with a basic amino acid residue, arginine or histidine, at position 13 (DR2 or DR4) and were negatively associated with the amino acid sequence Ile, Leu, Glu, and Asp at positions 67-70, which was present in some of the DR5-, DR6-, and DR8-associated alleles, in the DRB1 beta 1 domain. Anti-C reactivity was strongly associated with DR2, particularly with DRB1*1502., Conclusion: The several shared epitopes located on HLA-DRB1 and DQB1 genes control the anti-U1 RNP autoantibody response.

Objective: To determine racial differences in the frequencies of systemic sclerosis (SSc)-related serum antinuclear antibodies (ANA)., Methods: We tested serum samples from 275 Japanese, 416 North American Caucasian, and 24 North American black SSc patients for 8 SSc-related serum ANA, using indirect immunofluorescence, double immunodiffusion, and radioimmunoprecipitation assays., Results: In comparing the 3 racial groups, we found that anti-U1 RNP, anti-RNA polymerase I, II, and III, and anti-U3 RNP antibodies were the most frequently detected antibodies in Japanese, Caucasian, and black patients, respectively. Anti-PM-Scl antibody was found exclusively in Caucasians., Conclusion: The production of SSc-related serum ANA is related to immunogenetic background.

Objective: To clarify the clinical features and prognosis of systemic sclerosis (SSc) based on serum antinuclear antibodies (ANA)., Methods: We studied 275 consecutive Japanese patients newly diagnosed as having SSc, who were first evaluated during the period 1971-1990. Eight SSc-related ANA were identified using indirect immunofluorescence, double immunodiffusion, or immunoprecipitation assays. Clinical and prognostic features were retrospectively analyzed in patient groups, categorized by their serum ANA., Results: Cumulative survival rates at 10 years after diagnosis of SSc were 93% in patients with anticentromere antibodies (ACA), 72% in those with anti-U1 RNP, 66% in those with anti-DNA topoisomerase I (anti-topo I), and 30% in those with anti-RNA polymerases I, II, and III (anti-RNAP). Major organ involvement linked to cause of death included biliary cirrhosis in patients with ACA, isolated pulmonary arterial hypertension and cerebral hemorrhage in those with anti-U1 RNP, pulmonary interstitial fibrosis in those with anti-topo I, and cardiac and renal involvement in those with anti-RNAP., Conclusion: Determinations of serum ANA in SSc patients are useful in predicting organ involvement and long-term outcome.

Objective: To elucidate the clinical and immunogenetic associations with reactivity to autoantigenic epitopes on DNA topoisomerase I (topo I) recognized by sera from patients with systemic sclerosis (SSc)., Methods: Autoantigenic epitopes on topo I were identified by screening an epitope library constructed from topo I complementary DNA restriction fragments using autoimmune anti-topo I-positive sera as a probe. Epitope reactivities of sera from 43 anti-topo I-positive SSc patients were surveyed by immunoblotting, and associations with clinical symptoms and HLA-DR types were examined., Results: Four different epitope regions were identified on the topo I molecule. Immunoreactivity to the region encompassing amino acid residues 658-700, termed ER4, was found to be associated with diffuse cutaneous SSc, progressive pulmonary interstitial fibrosis, and poor prognosis for 15-year survival. SSc patients with ER4 reactivity frequently displayed the DR2/DRw52 phenotype., Conclusion: Molecular analysis of precise antigenic epitopes on topo I is helpful in classifying clinical subsets of SSc.