Anti-signal recognition particle antibody in patients without inflammatory myopathy: a survey of 6180 patients with connective tissue diseases.

OBJECTIVES: To clarify the prevalence of anti-signal recognition particle (anti-SRP) antibody in connective tissue diseases (CTDs) and investigate the clinical characteristics of patients without inflammatory myopathy. METHOD: Sera from 6180 patients with CTD were examined by immunoprecipitation (IPP) assays, and the records of patients positive for anti-SRP antibody were reviewed retrospectively. The antibody against the 54-kDa protein of SRP (SRP54) was quantified by enzyme-linked immunosorbent assay (ELISA) in patients with anti-SRP antibody. RESULTS: Of the 28 patients positive for anti-SRP antibody, nine (32.1%) did not have inflammatory myopathy. The clinical diagnoses and characteristics of those patients varied considerably. In patients with inflammatory myopathy, the index of anti-SRP54 was much higher than in those without myopathy (1.15 vs. 0.46; p = 0.036). CONCLUSIONS: The prevalence of anti-SRP antibody was 0.5% in a cohort of Japanese patients with CTD, and one-third of them did not have inflammatory myopathy. Sera from patients with inflammatory myopathy recognized SRP54 more strongly than in those without myopathy.

Clinical associations with autoantibody reactivities to individual components of U1 small nuclear ribonucleoprotein.

The reactivities to individual U1 small nuclear ribonucleoprotein (snRNP) components and their relationship to clinical features in patients with anti-U1 snRNP antibodies were examined. We evaluated 114 patients with connective tissue disease whose sera were positive for anti-U1 snRNP antibodies, but negative for anti-Sm antibodies. Antibodies to the U1 snRNP polypeptides 70K, A, and C were detected using subunit-specific enzyme-linked immunosorbent assays and antibodies to U1 small nuclear RNA (snRNA) were identified by an immunoprecipitation assay using deproteinized HeLa cell extracts. The clinical features were retrospectively obtained by chart review and prospectively collected after study entry. The pattern of antibody reactivities to U1 snRNP components varied among patients. The frequency of anti-70K, anti-A, anti-C, and anti-U1 snRNA antibodies was 60%, 86%, 74%, and 46%, respectively. There was no relationship between each reactivity and the clinical findings, but the presence of reactivities to increasing numbers of U1 snRNP components was correlated with sclerodactyly, shortness of the sublingual frenulum, esophageal dysfunction, and a lack of persistent proteinurea (p < 0.05 for all comparisons). The detection of autoantibody reactivities to individual components of the U1 snRNP particle is potentially useful for predicting the clinical course in patients with connective tissue disease and anti-U1 snRNP antibodies.

Clinical characteristics of cytomegalovirus infection in rheumatic diseases: multicentre survey in a large patient population.

OBJECTIVE: To survey and elucidate the clinical characteristics of CMV infection in rheumatic disease patients. METHODS: A detailed questionnaire survey on CMV infection was carried out against rheumatic disease patients hospitalized in member hospitals, and the obtained clinical and/or laboratory data were analysed. RESULTS: Out of 7377 patients, 151 were diagnosed as having CMV infection. The underlying diseases ranged broadly, but SLE, microscopic polyangiitis, and dermatomyositis were the most common. Four were diagnosed histopathologically, and the others via positive CMV antigenaemia. In addition to oral corticosteroid for all but one patient, 81 were treated with pulsed methylprednisolone (MPSL), 64 with cyclophosphamide (CYC) and 36 with other immunosuppressants. Forty-four had a fatal outcome, for which presence of clinical symptoms, other infectious complications, lymphopenia, an older age (>59.3 yrs) and the use of pulsed MPSL were significant risk factors (P < 0.05) by univariate analysis. Multivariate analysis retained the first three (P < 0.05). The CMV antigenaemia count was significantly higher for the symptomatic than asymptomatic [10.1 (0.0-2998.0) vs 4.0 (1.3-1144.4)/10(5) PMNs, respectively, P < 0.05; threshold count: 5.6/10(5) PMNs]. No treatment benefit by anti-viral agent was observed as for survival. CONCLUSION: CMV infection was mostly diagnosed by antigenaemia, and occurred among patients under strong immunosuppressive therapy using pulsed MPSL and/or immunosuppressants. Lymphopenia, presence of symptoms and other infections are significant risk factors for a poor outcome and pulsed MPSL and an older age may predict it. Patients were prone to be symptomatic with anti-genaemia count over 5.6/10(5) PMNs.

Autoantibodies to RNA polymerase II are common in systemic lupus erythematosus and overlap syndrome. Specific recognition of the phosphorylated (IIO) form by a subset of human sera.

Autoantibodies to RNA polymerases (RNAP) I, II, and III are reported to be highly specific for the diagnosis of scleroderma (systemic sclerosis, SSc). In the present study, the specificity of autoantibodies to RNAP I and III for SSc was confirmed by immunoprecipitation of 35S-labeled proteins. However, we report here the previously unrecognized production of anti-RNAP II autoantibodies by 9-14% of patients with SLE and mixed connective tissue disease/overlap syndrome. 12 out of 32 anti-RNAP II positive sera (group 1) immunoprecipitated a diffuse 220-240-kD band identified as the largest subunit of RNAP II whereas the remaining 20 (group 2) immunoprecipitated preferentially the 240-kD phosphorylated (IIo) form of the large subunit. After pulse labeling, group 1 sera immunoprecipitated only the 220-kD (IIa) RNAP II subunit, whereas the diffuse IIa/IIo band plus the 145-kD second largest RNAP II subunit (IIc) were immunoprecipitated after several hours of cold chase, suggesting that these sera recognized primarily the largest subunit of RNAP II. Group 2 sera recognized the IIc subunit after pulse labeling, and immunoprecipitated the IIc and IIo, but not the IIa, subunits after cold chase. Although it has been suggested that autoantibodies to RNAP II are usually accompanied by anti-RNAP I/III in SSc, all but one of the anti-RNAP II positive sera from SLE or mixed connective tissue disease/overlap syndrome patients, as well as most of the SSc sera, were negative for anti-RNAP I/III. Moreover, in contrast to previous reports suggesting that anti-RNAP antibodies rarely coexist with other SSc subset marker antibodies, anti-RNAP II antibodies were often accompanied by anti-Ku, anti-nRNP, or anti-topoisomerase I autoantibodies in the present study. We conclude that autoantibodies to RNAP II are not a specific marker for SSc, whereas autoantibodies to RNAP I/III are associated primarily with SSc. In addition, we have identified two distinctive patterns of RNAP II antigen recognition by autoantibodies, one of them characterized by specific recognition of the transcriptionally active (phosphorylated) form of RNAP II. The clinical significance of these different patterns remains to be determined.

Identification of autoantibodies to RNA polymerase II. Occurrence in systemic sclerosis and association with autoantibodies to RNA polymerases I and III.

In this study, autoantibodies to RNA polymerase II from sera of patients with systemic sclerosis have been identified and characterized. These antibodies immunoprecipitated polypeptides of 220 kD (IIA) and 145 kD (IIC), the two largest subunits of RNA polymerase II, and bound both subunits in immunoblots. These polypeptides were immunoprecipitated by the anti-RNA polymerase II monoclonal antibody 8WG16, which recognizes the carboxyl-terminal domain of the 220-kD subunit, and their identity to the proteins bound by human sera was confirmed in immunodepletion studies. Sera with anti-RNA polymerase II antibodies also immunoprecipitated proteins that were consistent with components of RNA polymerases I and III. In vitro transcription experiments showed that the human antibodies were an effective inhibitor of RNA polymerase II activity. In indirect immunofluorescence studies, anti-RNA polymerase II autoantibodies stained the nucleoplasm, as expected from the known location of RNA polymerase II, and colocalized with the anti-RNA polymerase II monoclonal antibody. The human sera also stained the nucleolus, the location of RNA polymerase I. From a clinical perspective, these antibodies were found in 13 of 278 patients with systemic sclerosis, including 10 with diffuse and three with limited cutaneous disease, but were not detected in sera from patients with other connective tissue diseases and from normal controls. We conclude that anti-RNA polymerase II antibodies are specific to patients with systemic sclerosis, and that they are apparently associated with antibodies to RNA polymerases I and III. These autoantibodies may be useful diagnostically and as a probe for further studies of the biological function of RNA polymerases.

Autoantibodies to DNA-dependent protein kinase. Probes for the catalytic subunit.

DNA-dependent protein kinase (DNA-PK) is an important nuclear enzyme which consists of a catalytic subunit known as DNA-PKcs and a regulatory component identified as the Ku autoantigen. In the present study, we surveyed 312 patients in a search for this specificity. 10 sera immunoprecipitated a large polypeptide which exactly comigrated with DNA-PKcs in SDS-PAGE. Immunoblot analysis demonstrated that this polypeptide was recognizable by a rabbit antiserum specific for DNA-PKcs. Although the patient sera did not bind to biochemically purified DNA-PKcs in immunoblots or ELISA, they were able to deplete DNA-PK catalytic activity from extracts of HeLa cells in a dose-dependent manner. We conclude that these antibodies should be useful probes for studies which aim to define the role of DNA-PK in cells. Since six sera simultaneously contained antibodies to the Ku protein, these studies suggest that relatively intact forms of DNA-PK complex act as autoantigenic particles in selected patients.

Identification of cDNA encoding rat eosinophil cationic protein/eosinophil-associated ribonuclease.

Using the rapid amplification of cDNA ends (RACE) procedure, we have determined the complete nucleotide sequence for the cDNA encoding rat eosinophil cationic protein (ECP)/eosinophil-associated ribonuclease (EAR). The deduced amino acid sequence revealed that the molecular weight of rat preECP/EAR is 18.0 kDa and the isoelectric point is 9.85, indicating that rat ECP/EAR is highly cationic. The homology of amino acid sequence between rat ECP/EAR and human ECP is 54%, and that between rat ECP/EAR and human eosinophil-derived neurotoxin (EDN) is 51%. Rat ECP/EAR is also homologous to human ribonuclease k6 (homology 47%).

Clinical characteristics of Japanese patients with anti-PL-7 (anti-threonyl-tRNA synthetase) autoantibodies.

OBJECTIVE: The clinical and laboratory features of seven Japanese patients with anti-aminoacyl-tRNA synthetase (ARS) autoantibodies against PL-7 (anti-threonyl-tRNA synthetase) were analyzed and compared with previously published findings. METHODS: Serum samples from 1,135 Japanese patients with various autoimmune diseases were screened for anti-PL-7 antibodies using RNA and protein immunoprecipitation assays. The patients whose sera contained anti-PL-7 antibodies were assessed regarding clinical symptoms and clinical course. RESULTS: Sera from seven patients were found to have anti-PL-7 antibodies. These autoantibodies were associated with polymyositis/dermatomyositis (PM/DM) and/or interstitial lung disease (ILD). The clinical diagnoses of these seven patients were PM - systemic sclerosis (SSc) overlap (5 patients), DM (1 patient) and idiopathic pulmonary fibrosis (IPF) (1 patient). All patients had ILD with a chronic course and six also had arthritis (85%) and five sclerodactyly (71%). CONCLUSIONS: These results indicate that anti-PL-7 autoantibodies are closely associated with PM-SSc overlap as well as ILD, arthritis and sclerodactyly in our series of Japanese patients.

Successful treatment of recurrent intracardiac thrombus in Behçet's disease with immunosuppressive therapy.

Behçet's disease (BD) is a chronic multisystem inflammatory disorder characterized by recurrent oral and genital ulcers, skin eruptions and uveitis. Neurological, gastrointestinal, and musculoskeletal systems are also involved. Although venous and arterial vasculitis occur in up to one-third of patients, intracardiac thrombus is a very rare complication. We herein report the case of a 46-year-old man with BD who presented with a large right atrial thrombus. Within a month after surgical removal, the thrombus recurred and was successfully treated with immunosuppressants that included prednisolone and cyclophosphamide.

Human anti-asparaginyl-tRNA synthetase autoantibodies (anti-KS) increase the affinity of the enzyme for its tRNA substrate.

Autoantibodies directed against specific human aminoacyl-tRNA synthetases have been associated with a clinical picture including myositis, arthritis, interstitial lung disease and other features that has been referred to as the "anti-synthetase syndrome". Anti-asparaginyl-tRNA synthetase autoantibodies (anti-KS), the most recently described anti-synthetase autoantibodies, are directed against human cytosolic asparaginyl-tRNA synthetase and neutralize specifically its activity. Here we show that these antibodies recognize two epitopes on the human enzyme, an N-terminal epitope reactive in immunoblot experiments and a heat-labile epitope in the catalytic domain. In contrast to the well studied anti-Jo-1 autoantibodies anti-KS when bound to the synthetase increase the affinity of the synthetase for its tRNA substrate and prevent aminoacylation without interfering with the amino acid activation step.

Tyrosine-kinase dependent TGF-beta and extracellular matrix expression by mechanical stretch in vascular smooth muscle cells.

Vascular hypertrophy, which is characterized by proliferation of vascular smooth muscle cells (VSMC) and accumulation of extracellular matrix (ECM), is a major pathological change in blood vessels after chronic exposure to hypertension. Blood pressure is transmitted to the arterial walls and counterbalanced by mechanical stress, leading to stretching of circumferentially oriented VSMC, which may play some role in the pathogenesis of vascular hypertrophy. The present study was designed, therefore, to investigate the effect of mechanical stretch on the expression of ECM components and transforming growth factor-beta (TGF-beta), a potent stimulator for ECM production, and to examine the signal transduction mechanisms of the induction of TGF-beta in cultured rat VSMC. VSMC were subjected to cyclic stretch to provide a maximal elongation of 20% at a rate of 60 cycles per minute for up to 24 h. Mechanical stretch stimulated TGF-beta1 mRNA expression in a time- and elongation-dependent manner. Indeed, the secretion of TGF-beta proteins into the culture media was increased after stretch. Stretch also stimulated mRNA expression of the ECM components, type I and type IV collagen, and fibronectin, which was largely inhibited by addition of neutralizing antibody against TGF-beta. The tyrosine kinase inhibitors genistein and herbimycin A blocked the induction of TGF-beta1 and type I collagen by stretch, while protein kinase C inhibitors, the calcium channel blockers nitrendipine and gadolinium, or Ca removal from the media had no effect. These results suggest that stretch-induced, tyrosine kinase-mediated autocrine/paracrine production of TGF-8 may play a critical role in the progression of vascular remodeling associated with high blood pressure.

Thrombocytopenia responsive to warfarin in a patient with systemic sclerosis--systemic lupus erythematosus overlap.

We report a patient with overlapping systemic sclerosis-systemic lupus erythematosus who developed refractory thrombocytopenia and recurrent thromboses. Coagulation and platelet labeling studies revealed that platelets were being consumed by the thrombus formation, and anticoagulation with warfarin dramatically increased the platelet count. This case report suggests that platelet consumption due to thrombus formation is one of the causes of thrombocytopenia in patients with rheumatic disease.

A case of polymyositis associated with hepatitis B infection.

This report describes the case of a 47-year old man who developed myositis in association with hepatitis B surface antigen-positive hepatitis. Interestingly, the myositis repeatedly worsened 2 months after the exacerbation of hepatitis in this case, suggesting a close association between hepatitis B infection and myositis. The dose of prednisolone was increased twice in order to treat the exacerbating myositis, resulting in improvement of the muscle symptoms, but the patient eventually died of liver failure. Only 5 other myositis patients with hepatitis B antigenemia have been reported in the literature. We review these cases of the association between hepatitis B infection and myositis.

Rheumatoid arthritis associated with methotrexate-induced pneumonitis: improvement with i.v. cyclophosphamide therapy.

Pneumonitis is one of the most serious adverse effects associated with low-dose weekly methotrexate (MTX) therapy. Immediate cessation of MTX, and the introduction of oxygen therapy and glucocorticoids usually results in a dramatic improvement in the pulmonary toxicity. We report here a case of MTX-induced pneumonitis in a patient with rheumatoid arthritis (RA). Severe hypoxemia and interstitial infiltration in both lung fields did not respond to the withdrawal of MTX and the administration of oxygen and steroid pulse therapy. When intravenous cyclophosphamide (CYC) pulse therapy was initiated, however, rapid physiologic and radiographic improvement was seen. Our case suggests that CYC treatment may have a beneficial effect on MTX-induced pneumonitis that is resistant to steroid therapy.

A case of systemic lupus erythematosus presenting with rectal ulcers as the initial clinical manifestation of disease.

Gastrointestinal involvement is often seen in patients with systemic lupus erythematosus (SLE). All parts of the gastrointestinal tract may be affected. However, rectal involvement at onset is rare. We describe here a case of SLE in which rectal ulcers due to vasculitis occurred as the initial manifestation of the disease without involvement of any other organ. The ulcers worsened, along with the appearance of lupus nephritis 5 years later When steroid therapy was initiated, there was rapid clinical and radiographic improvement. Our case suggests that rectal ulcer is a rare but important complication of SLE and can represent the initial and sole clinical manifestation of the disease.

Systemic lupus erythmatosus associated with autoimmune hepatitis two cases with novel autoantibodies to transfer RNA-related antigens.

Two cases of systemic lupus erythematosus (SLE) with autoimmune hepatitis are reported. Both patients had a mild form of SLE without central nervous system or renal involvement and showed a rapid response to corticosteroid therapy. Neither of them had antibodies to mitochondria, smooth muscle, and liver/kidney microsome-1 related to autoimmune hepatitis. Instead, novel autoantibodies which react with transfer RNA-related antigen were detected. These autoantibodies could be a useful marker for classification of SLE associated with autoimmune hepatitis.

Pyloric stenosis in a patient with progressive systemic sclerosis.

A 64-year-old Japanese woman with progressive systemic sclerosis (PSS) who developed severe pyloric stenosis is described. The conservative treatments brought only the temporary symptomatic relief, and pyloroplasty became necessary. No ulcerative lesions or tumors were found in the resected stomach or duodenum specimens implicated for stenosis. The histological examinations revealed edema and replacement fibrosis in the pyloric ring. The possible mechanisms of pyloric stenosis are discussed.

Association of human leukocyte antigen class II genes with autoantibody profiles, but not with disease susceptibility in Japanese patients with systemic sclerosis.

OBJECT: To examine the role of human leukocyte antigen (HLA) class II genes in the development of systemic sclerosis (SSc) as well as in the clinical and serologic expression of SSc in patients. METHODS: HLA-DRB1, DRB3, DRB4, DQB1, and DPB1 alleles were determined by genotyping; and serum antinuclear antibodies were identified using indirect immunofluorescence, double immunodiffusion and immunoprecipitation. PATIENTS: One hundred and five Japanese patients with SSc and 104 race-matched healthy controls. RESULTS: Frequencies of DRB1 and DQB1 alleles were not different between SSc patients and healthy controls, while DPB1*0901 was marginally increased in SSc patients. In contrast, SSc-related autoantibodies were closely associated with the clinical features. HLA class II genes were detected as follows: anti-DNA topoisomerase I antibody with diffuse cutaneous involvement, pulmonary fibrosis, and DRB1*1502-DQB1*0601-DPB1*0901; anti-U1RNP antibody with overlapping features of lupus and/or myositis and DRB1*0401/*0802-DQB1*0302; and anticentromere antibody with limited cutaneous involvement and DRB1*0101-DQB1*0501-DPB1*0402. In the analysis of the association of HLA class II and the clinical features in SSc patients significant differences were obtained only for the increased frequencies of arthritis and rheumatoid factor in patients with DRB1*0405 compared to those without. CONCLUSION: HLA class II genes strongly influence the production of SSc-related autoantibodies rather than the development of SSc. In addition, SSc is a composite disease of distinctive subsets defined by serum autoantibodies, which have specific clinical and HLA class II associations.

[Frequency of anti-centromere antibodies in the anti-mitochondrial antibody positive sera].

Autoantibodies give us useful informations for the understanding of the co-existence of multiple autoimmune diseases in an individual patient. This study was undertaken to clarify the frequency of anti-centromere antibodies in the sera which were positive for anti-mitochondrial antibodies. Serum samples with anti-mitochondrial antibodies were examined on the frozen rat liver section and HEp-2 cells by indirect immunofluorescence. 57 out of 239 anti-mitochondrial antibody positive sera were found to contain specific autoantibodies reactive with centromeric antigens of the HEp-2 chromosomal spreads. The co-existence of anticentromere antibodies with anti-mitochondrial antibodies were significantly more frequent than with other defined autoantibodies. Clinical observation which shows frequent association of CREST syndrome and primary biliary cirrhosis appear to be explained by the frequent occurrence of anti-centromere antibodies and anti-mitochondrial antibodies.

[Analysis of autoantigens and clinical significance of antinuclear antibodies].

A highly sensitive RNA-immunoprecipitation assay (Lerner-Steitz assay) is a unique and useful method of identifying autoantibodies to RNA-associated antigens. In this study, we identified novel autoantibodies to tRNAs using RNA-immunoprecipitation assay. In screening of 1472 sera by RNA-immunoprecipitation using HeLa cell extracts as an antigen source, 41 sera were found to immunoprecipitate tRNAs. Fifteen of these 41 sera also immunoprecipitated deproteinized tRNAs, indicating that these 15 sera contained anti-tRNA antibodies. Three sera immunoprecipitated naked tRNA from E. coli. When in vitro transcripts from cDNAs encoding E. coli tRNAs and their synthesized mutants were used as antigens, it was demonstrated that the representative serum recognized the conformational epitope of the "L"-shape structure which was conserved in all tRNAs of all species. This finding suggests the role of bacterial infection in the development of autoantibodies and autoimmune diseases. Two of 15 sera containing anti-tRNA antibodies were identified as anti-PL-12 (alanyl-tRNA synthetase) antibodies. Eleven of the remaining 13 patients were diagnosed as either SLE, Sjögren's syndrome or their overlap. In addition, fever, Raynaud's phenomenon, polyarthritis, leukocytopenia and characteristic annular erythema were frequently found in these patients. Novel autoantibodies to tRNAs appeared to be associated with common clinical features but were distinct from previously described anti-aminoacyl-tRNA synthetases.