Myenteric neuronal antibodies in scleroderma: passive transfer evokes alterations in intestinal myoelectric activity in a rat model.

Although the mechanism for neuropathic gastrointestinal motility disturbances in scleroderma is unknown, we have previously described anti-myenteric antibodies in some patients with scleroderma. The aim of this study was to screen patients with scleroderma who had gastrointestinal symptoms for the presence of anti-myenteric neuronal antibodies and then purify the immunoglobulin G (IgG) fraction from serum samples for passive immunization into a rat model and observe for intestinal motility effects. Patients with scleroderma were screened, a serum sample from a patient with high titer anti-myenteric neuronal antibodies was obtained, and IgG was purified. Using a rat model with chronic indwelling intestinal electrodes to measure intestinal myoelectric activity, we passively transferred the IgG from either control subjects or this patient with scleroderma. We immunosuppressed the rats and intraperitoneally injected IgG from control subjects and this patient with scleroderma daily for 7 days. Recordings of myoelectric activity in control injected rats revealed no difference from baseline, but a prolongation in the activity front duration and interval and a disruption were seen after scleroderma IgG injections. IgG from a patient with scleroderma with antimyenteric neuronal antibodies, when passively immunized into a rat model, evokes intestinal myoelectric activity alterations. We hypothesize that these antibodies could account for the gastrointestinal neuropathic motility disturbances seen in scleroderma.     

Scl-86, a marker antigen for diffuse scleroderma.

More than 300 sera from patients with a connective tissue disease were analyzed with the immunoblotting technique. The presence of autoantibodies against an 86,000-mol wt marker antigen for diffuse scleroderma (Scl-86) was found in 14 out of 33 patients with scleroderma. The presence of anti-Scl-86 antibodies seemed to correlate with the diagnosis of diffuse scleroderma since they were found in 13 out of 22 diffuse scleroderma patients and in only one out of 11 patients with limited scleroderma. All scleroderma sera (33 patients' sera and 13 reference sera) were also tested for the presence of anti-Scl-70 antibodies. It was found that all anti-Scl-70 positive sera (n = 25) contained anti-Scl-86 antibody as well, suggesting a relationship between these two antigens. However, the Scl-86 antigen was shown to be an extremely insoluble nonchromosomal protein, resistant to boiling in sodium dodecyl sulfate. This contrasts with the Scl-70 antigen, which has been described as a thermolabile, soluble antigen present in the chromatin fraction. Together, our results are consistent with the idea that Scl-70 is a degradation product of Scl-86. The Scl-86 antigen is present in freshly prepared rabbit thymus, spleen, and liver nuclei as well as in nuclei from various cultured cell lines, but is not detectable in extractable nuclear antigen from rabbit thymus. In a limited retrospective study, the anti-Scl-86 antibodies were found in two sera from patients with Raynaud's phenomenon before the development of diffuse scleroderma. Therefore, it is possible that screening of patients' serum for this antibody might predict the development of diffuse scleroderma.     

Polynucleotide antibodies in connective tissue disease: viral markers or disease mediators?

Antibodies to polynucleotides are seen primarily in systemic lupus erythematosus (SLE), but also occur in a variety of other connective tissue diseases. We looked at the prevalence of antinucleotide antibodies (double- and single-stranded RNA and DNA [dRNA, sRNA, dDNA, and sDNA]) in the sera of patients with SLE (70), rheumatoid arthritis (RA) (31), juvenile rheumatoid arthritis (JRA) (68), osteoarthritis (12), and of 22 patients with a preceding viral illness. In comparison with sera from a control population, elevated mean antibody levels to sRNA were found in the sera of all the patients with connective tissue disease, as well as in the sera of patients with preceding RNA, but not DNA, viral infections. Elevated mean levels of antibodies to dRNA were seen in all groups with the exception of RA. Elevated mean antibody titers to sDNA were not seen in patients with JRA nor were they present in the sera of patients with preceding RNA viral infections. Elevated mean anti-dDNA titers were seen only in sera from patients with SLE. High correlation coefficients between the levels of antibodies to sRNA and dRNA in sera from SLE and RA, and between sDNA and dDNA in sera from SLE suggest cross-reactivity of the antibodies in these diseases. Immunization of an elderly population with influenza (RNA) viral vaccine induced antibodies to sRNA only. These studies further document the prevalence of antipolynucleotide antibodies in the sera of patients with connective tissue diseases.(ABSTRACT TRUNCATED AT 250 WORDS)     

抗核抗体谱在自身免疫性疾病中的临床应用

目的 探讨抗核抗体谱在自身免疫病中的应用价值.方法 ANA 检测采用间接免疫荧光法,ENA多肽抗体谱用免疫印迹法.结果 729例患者中,有58例自身免疫病患者,其中SLE 31例,混合性结缔组织病12例,干燥综合征13例,硬皮病和皮肌炎各1例.所有自身免疫病患者的ANA均为阳性,但SLE患者的ENA抗体谱中可显示抗nRNP、抗Sm、抗SSA、抗SSB、抗dsDNA等抗体阳性;而干燥综合征患者仅抗SSA或抗SSB抗体阳性;皮肌炎患者仅显示抗Jo-1抗体阳性;硬皮病患者仪显示抗Scl-70抗体阳性.结论 ENA抗体谱对种自身免疫病的诊断具有重要意义.     

Association of autoantibodies to different nuclear antigens with clinical patterns of rheumatic disease and responsiveness to therapy

Using a hemagglutination test which can detect antibodies to (a) native and denatured deoxyribonucleic acid (DNA) and (b) an extractable nuclear antigen (ENA), a comparative study of patterns of autoantibody formation has been done in systemic lupus erythematosus (SLE) and related rheumatic diseases. Antibody to native DNA was present in the serum in 96% of patients with active SLE and disappeared during remissions. Antibody to ENA was found in 86% of those patients with SLE nephritis who responded to treatment but in only 8% of those who did not. The highest titers of antibody to ENA were found in patients having a mixed connective tissue disease syndrome with features of SLE, scleroderma, and myositis. The latter syndrome was notable for the absence of renal disease and for a striking responsiveness to corticosteroid therapy. Hemagglutination testing of 277 sera from normal persons and patients with a wide variety of acute diseases other than SLE revealed the presence of antibody to native DNA in only 1.4% and antibody to ENA in only 0.4%.These results yield significant correlations among the pattern of autoimmune reactivity, the clinical form of the rheumatic disease, and responsiveness to treatment. They implicate the qualitative nature of the patient's immune response as a conditioning factor in the type of disease. Together with other correlations they may allow classification of rheumatic diseases into more biologically meaningful groups and lead to more selective methods of therapy.     

Antibodies to T- and L-isoforms of the cytoskeletal protein, fimbrin, in patients with systemic lupus erythematosus.

The cytoskeleton is a complex network of proteins that maintain cell shape, mobility, and organelle function. Its components can be divided into three distinct classes: microfilaments, microtubules, and intermediate filaments. Fimbrins are microfilament proteins, a family of cytoplasmic phosphoproteins. Expression of the L-fimbrin isoform is restricted to replicating blood cells and expression of the T-fimbrin isoform to replicating cells of solid tissues. Sera from normals and from patients with systemic lupus erythematosus (SLE), juvenile arthritis, rheumatoid arthritis, Sj?gren's syndrome, osteoarthritis, vasculitis, scleroderma, and mixed connective tissue disease were tested for the presence of antibodies to T- and L-fimbrin by ELISA, using purified recombinant fimbrin. The mean OD of sera from SLE patients was significantly higher than in normals (T-fimbrin, P less than 0.0001; L-fimbrin, P less than 0.001). 48 of 98 SLE sera had antibodies to T-fimbrin; 32 had antibodies to L-fimbrin; 20 had antibodies to both; 28 had only anti-T, and 12 had only anti-L-fimbrin. The mean OD for sera of the other rheumatic diseases was not significantly different from normals. The presence of either L- or T-fimbrin antibody was associated with pleuropericarditis (P = 0.015), photosensitivity (P = 0.011), and anti-Sm antibody (P = 0.010). Central nervous system SLE was associated with the presence of the L-fimbrin antibody alone (P = 0.016). There was a strong association between DR7 (but not other MHC alleles) and anti-L-fimbrin antibodies in SLE patients (chi square = 18; P less than 0.00002). No MHC association was observed with anti-T-fimbrin antibodies.     

Antibodies to human caudate nucleus neurons in Huntington''s chorea.

Antibodies reacting with neuronal cytoplasmic antigens present in normal human caudate and subthalamic nuclei were detected in 37 of 80 probands afflicted with Huntington's disease (HD). IgG antibodies were detected by immunofluorescence using frozen sections of unfixed normal human and rat brain. Specificity of IgG binding was confirmed using pepsin F(ab')2 fragments of IgG isolated from positive sera. In vitro complement fixation of IgG antibody was detected in 22 of 31 sera tested. Neuronal cytoplasmic antigens reacting with positive HD sera were diminished after trypsin or RNAase treatment of tissue sections but were not removed by DNAase, neuraminidase, EDTA, or dithiothreitol treatment. Antibody staining of neurons could be removed after absorption with isolated caudate nucleus neurons or by using perchloroacetic acid extracts of caudate nucleus. Prevalence of antibody reacting with neuronal cytoplasm was 3% in 60 normal controls and 6% among a wide variety of patients with diverse neurological disorders. However, one-third of 33 patients with Parkinson's disease showed presence of antineuronal antibody. Among patients with HD, a significant association was noted between duration of clinical disease greater than 7 yr and titers of antibody of 1:2 or greater (P less than 0.001). When 115 family members of HD probands were tested, 30% of unaffected spouses showed presence of antineuronal antibody. 23.2% of first-degree relatives at risk for developing HD was also positive (P less than 0.001). 10.5% of second-degree relatives showed presence of antineuronal antibody. These data may support an environmental or infectious factor somehow involved in the ultimate expression of HD.     

Ku抗原的提取及其抗体的检测

目的 测定本组自身免疫性结缔组织疾病（ＣＴＤｓ）患者中抗Ｋｕ抗体的阳性率。方法 制备兔腺丙酮粉,进而提取Ｋｕ抗原,采用免疫双扩散法检测４３８份ＣＴＤｓ和５０份正常对照血清中的抗Ｋｕ抗体。结果 自制的Ｋｕ抗原与标准Ｋｕ抗血清免疫双扩散出现清晰的沉淀线。４８８份血清中,抗Ｋｕ抗体阳性率２．５％（１２／４８８）,其中皮肌炎为２．２％（１／４５）,弥漫型系统性硬化症为３．４％（２／５８）,系统性硬化症并多     

Pharmacologic characterization of the changes in cholinergic sensitivity of rat jejunal circular muscle after myenteric plexus ablation

Neurons located in the myenteric plexus are generally believed responsible for motor control of intestinal circular muscle. The in vitro isometric responses of naive and myenterically denervated (MD) rat jejunal circular muscle to bethanechol and carbachol, alone and in the presence and absence of neuronal antagonists (hexamethonium bromide, tetrodotoxin and Botulinum toxin A) 15 and 30 days after myenteric plexus ablation, were determined. The responses to bethanechol indicated no differences in muscarinic sensitivity between naive and MD tissue. The relative potency of carbachol, which acts at both muscarinic and nicotinic receptors, in MD tissue 15 days after denervation was significantly higher than that in naive tissue. However, 30 days after denervation, the relative potencies of carbachol in naive and MD circular muscle were comparable. The presence of neuronal antagonists had no effect on the relative potency of carbachol 15 days after myenteric denervation, but altered significantly the responses 30 days after denervation. The effects produced by the neuronal antagonists 30 days after myenteric denervation were qualitatively and quantitatively different than those produced in naive tissue, suggesting that the nature of the innervation in these tissues was different. These results demonstrate that circular muscle was denervated initially after myenteric plexus ablation but reinnervation occurred within 30 days. The reinnervation observed is likely due to neurons located in the submucosal plexus.     

定量酶联免疫吸附试验检测各种结缔组织病患者血清中的抗可提取核抗原抗体

目的　定量检测各种结缔组织疾病 (CTD)患者血清中抗可提取核抗原 (ENA)自身抗体滴度 ,并探索其临床意义。方法　自制抗各种ENA单克隆抗体 ,以夹心酶联免疫吸附试验 (ELISA)定量检测 1 2 8例CTD患者、5 0例非CTD患者及 6 0名健康人血清的各种抗ENA自身抗体滴度。用健康人血清各ENA抗体滴度的均数加上 3倍标准差 ( x +3s)为cutoff值 ,界定各种ENA抗体检出率。结果　与健康对照组和非CTD患者组比较 ,抗Sm抗体在系统性红斑狼疮 (SLE)、抗ul RNP抗体在SLE和混合性结缔组织病 (MCTD)、抗SSA和SSB在干燥综合征 (SS)、抗Scl 70抗体在系统性硬化病 (PSS)、抗Jo 1抗体在多发性肌炎 /皮肤炎 (PM/DM)患者中的水平和阳性率都明显升高 (P <0 .0 1 )。结论　定量检测CTD患者血清中各种ENA抗体滴度 ,对CTD疾病的鉴别、诊断和分型具有重要价值     

Relationship between snRNA species contained in nuclear antigens recognized by autoantibodies and the clinical profile in systemic rheumatic diseases

Antibodies to extractable nuclear antigens (ENA) are generally used in the diagnosis of connective tissue diseases. Using a rapid, very sensitive method we have shown that extractable nuclear antigens, which are now well-characterized at the molecular level, differ by their RNA content. The method was applied to the sera of 17 patients suffering from different connective tissue diseases. The results show that mixed connective tissue disease (MCTD) and other mild connective tissue diseases are characterized by the presence in the antigen of U1 small nuclear RNA (U1 snRNA) only. On the other hand, antibodies from 6 out of 8 patients tested with Systemic Lupus Erythematosus (SLE) recognize antigens exhibiting a more complex RNA pattern. Three of them precipitated all five snRNAs U2, U1, U4, U5, U6 whereas some snRNAs were lacking or quantitatively less important in precipitates obtained with the three others.     

Elevated Levels of Antibodies to Epstein-Barr Virus Antigens in Sera and Synovial Fluids of Patients with Rheumatoid Arthritis

The frequencies and levels of antibodies to Epstein-Barr virus (EBV)-specific antigens were determined in paired sera and synovial fluids from patients with rheumatoid arthritis (RA) and in sera from patients with other connective tissue diseases; i.e., systemic lupus erythematosus, progressive systemic sclerosis, and osteoarthritis (OA). The specimens were also tested for the presence of antibodies to RA-associated nuclear antigen. Compared to healthy controls, the patients' sera showed increased frequencies of elevated antibody titers (≥320) to Epstein-Barr viral capsid antigen, a correspondingly enhanced (twofold to threefold) geometric mean titer, and an increased frequency of antibodies at elevated titers (≥10), usually to the restricted component and rarely the diffuse component of the early antigen complex. Levels of antibody to the EBV-associated nuclear antigen were within the normal range. Enhancement of antibody titers was more pronounced in seropositive RA patients (i.e., positive for rheumatoid factor) than in those who were not. Enhancement was also found in systemic lupus erythematosus and progressive systemic sclerosis. Antibody to RA-associated nuclear antigen was detected at an increased frequency only in the group of seropositive RA patients (90%), as compared to 8-15% in the other connective tissue diseases and 6-8% in healthy controls. The antibody titers in the synovial fluids equaled or were at most twofold higher or lower than those in the sera. In addition, levels of EBV-specific antibodies were studied serially over a period of 6-10 mo in patients with RA and OA. Parameters of disease activity were determined and compared to antibody levels. EBV-specific antibodies in sera of OA patients remained constant and within normal limits throughout the study. Although EBV-specific antibodies were often elevated in RA patients, they also remained constant, with the exception of three patients, who showed gradual increases in one of the four antibodies, which did not correlate with disease activity.     

Anti-(U1) small nuclear RNA antibodies in anti-small nuclear ribonucleoprotein sera from patients with connective tissue diseases.

Small nuclear ribonucleoprotein (snRNP) particles are a class of RNA-containing particles in the nucleus of eukaryotic cells. Sera from patients with connective tissue diseases often contain antibodies against the proteins present in these snRNPs. Antibodies against the RNA components of snRNPs, the U snRNAs, are thought to be rare. We tested 118 anti-snRNP sera for the presence of anti-snRNA antibodies and found them in 45 sera (38%). In all sera the antibodies (IgG and F(ab)2 fragments thereof) were exclusively directed against U1 snRNA. The anti-(U1) RNA antibodies were always accompanied by anti-(U1)RNP antibodies but were not found in sera which contain antibodies of the Sm serotype directed against all nucleoplasmic U snRNP particles. Like anti-RNP antibodies, anti-U1 RNA activity is confined to sera from patients with SLE or SLE overlap syndromes and is rarely found in patients with other connective tissue diseases. By analyzing binding to subfragments of U1 snRNA made in vitro, it was demonstrated that anti-(U1)RNA antibodies recognize epitopes distributed throughout the U1 RNA molecule. In most sera, however, either the second or the fourth hairpin loop is the main target of the antibody. The possible mechanisms that could lead to the production of this new type of autoantibody are discussed.     

Epidemiology of systemic lupus erythematosus and other connective tissue diseases in Rochester, Minnesota, 1950 through 1979

The incidence and prevalence rates of connective tissue disease syndromes in Rochester, Minnesota, from 1950 through 1979 are reported. The incidence of definite systemic lupus erythematosus (SLE) has not increased since 1960. The incidence of SLE in the elderly population was higher than that in previous reports. Rates of SLE and discoid lupus erythematosus were approximately equal. Other diagnoses (in decreasing order of frequency) were suspected lupus erythematosus, scleroderma, drug-induced lupus, and overlapping connective tissue disease syndromes. The 10-year survival of patients with definite SLE was decreased, and the survival of patients with suspected SLE was the same as that of the general population.     

联合检测抗核小体、抗组蛋白和抗双链DNA抗体在SLE中的意义

目的探讨抗核小体抗体、抗组蛋白抗体和抗双链DNA抗体联合检测在SLE中的意义。方法采用免疫斑点法对199例SLE患者,309例其他弥漫性结缔组织病（CTD）患者和72例正常组进行上述3个抗体的联合检测,并与SLE患者临床及实验室指标进行相关性分析。结果核小体抗体在SLE组、CTD组和正常组中的阳性率为47．24％、0．97％和0．00％;抗组蛋白抗体为41．71％、0．97％和0．00％;抗双链DNA抗体为33．17％、0．65％和0．00％,上述各组内阳性率差异均有统计学意义（P〈0．05）。抗核小体抗体、抗组蛋白抗体和抗双链DNA抗体在SLE组中的特异性分别为96．91％、96．51％和97．06％;任何两种或三种抗体同时阳性对SLE的特异性为100％。三种抗体在SLE活动组中的阳性率均明显高于非活动组（P〈0．01）;抗核小体抗体与WBC、HB、C3、C4、胸膜炎等指标间具有相关性（P〈0．05）。结论联合检测抗核小体抗体、抗组蛋白抗体和抗双链DNA抗体对SLE中有较好的敏感性和特异性,抗核小体抗体的敏感性最高且与SLE病情活动密切相关。     

全自动免疫分析法检测抗核抗体谱诊断系统性红斑狼疮的价值

目的评价全自动免疫分析法检测抗核抗体谱在系统性红斑狼疮（SLE）中的价值。 方法采用BioPlex^TM 2200全自动免疫分析仪和酶联免疫吸附法（ELISA）检测30例SLE患者血清中dsDNA、Sm、RNP等抗体;采用BioPlex^TM 2200全自动免疫分析仪检测109例SLE患者（SLE组）、109例结缔组织病患者（结缔组织病组）及33名健康对照者（对照组）血清中15项抗核抗体谱阳性率并比较3组的差异。比较2种检测方法的一致性。 结果SLE组抗dsDNA、抗核染色质、抗核糖体P蛋白、Sm、SmRNP、RNP、RNP68、SSA、RNP-A、Ro60、Ro52、SSB的阳性率与结缔组织病组、对照组比较,差异均具有统计学意义（P均<0.05）,CenB的阳性率与结缔组织病组、对照组比较,差异均无统计学意义（P均>0.05）,Any ANAs与对照组比较,差异具有统计学意义。全自动分析法和酶联免疫吸附法检测Sm的阳性率差异有统计学意义（66.67% vs 56.67%,χ²=4.17,P=0.03）,检测抗dsDNA、RNP、Ro60、Ro52、SSB、抗核糖体P蛋白的阳性率差异均无统计学意义（P均>0.05）。2种方法检测dsDNA、Ro60、SSB、抗核糖体P蛋白的一致性良好（Kappa值分别为0.79、0.87、0.76、0.80）,检测Sm、RNP、Ro52的一致性一般（Kappa值分别为0.61、0.60、0.50）。 结论BioPlex^TM 2200全自动免疫分析仪检测抗核抗体谱对SLE的诊断具有一定价值,其检测能力与酶联免疫吸附法相似。     

Clinical importance of antibodies against platelet activating factor in antiphospholipid syndrome manifestations

BACKGROUND: We assessed whether antibodies against platelet activating factor (PAF) are related to the presence of antiphospholipid syndrome (APS) clinical manifestations, in particular thrombosis, in patients with connective tissue diseases. MATERIALS AND METHODS: Anti-PAF, anticardiolipin (aCL), antibeta2 glycoprotein I (antibeta2GPI) and antiphosphatidylcholine (anti-PC) antibodies were determined in 52 patients with APS, 29 patients with systemic lupus erythematosus (SLE) aCL but without APS, 30 patients with SLE without aCL, and 30 patients with scleroderma. A new enzyme-linked immunosorbent assay (ELISA) was developed for determining anti-PAF antibodies in a bovine serum-free fashion. RESULTS: The ELISA showed high specificity. Homologous inhibition experiments showed 60-70% inhibition. Anti-PAF antibodies were found in 18/52 APS patients, 10/29 SLE/aCL+ patients, 9/30 SLE/aCL- patients and 3/30 scleroderma patients. Anti-PAF antibodies were significantly associated with anti-PC antibodies (odds ratio [OR] 12. 7, P < 0.01), and there was a modest association with immunoglobulin G (IgG) aCL (OR 3.1, P > 0.10), but not with IgM aCL or antibeta2GPI. Three SLE/aCL+ patients and five SLE/aCL- patients had clinical manifestations characteristic of APS. All these patients had anti-PAF antibodies, while none had high titres of aCL or antibeta2GPI antibodies and only one had anti-PC antibodies. Among the combined APS and SLE groups, the presence of anti-PAF antibodies was significantly associated with clinical manifestations which are characteristic of APS (OR 2.6, P = 0.02). The effect was independent of IgG aCL and antibeta2GPI antibodies. CONCLUSIONS: Anti-PAF antibodies are common in APS and SLE and comprise an independent factor for the development of thrombosis. Several patients experiencing thromboses have anti-PAF antibodies without other antiphospholipid specificities.     

T lymphocyte interaction with immunoglobulin G antibody in systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is an autoimmune disease with multiple immune disturbances whose mechanisms remain unclear. We examined the interaction of antilymphocyte antibodies with cultured normal T lymphocytes. T cells were prepared by E-rosetting after petri-dish removal of adherent cells and cultured for 2-7 d in the presence of SLE sera or normal human sera. Cultured T cells were washed and sonicated, and the amount of cell-associated IgG was quantitated by radioimmunoassay or enzyme-linked immunoassay (ELISA) methods. T cells cultured with 27 of 39 SLE sera showed marked increments of associated immunoglobulin G (IgG) although this was not observed with sera from mixed connective tissue disease patients containing high titers of ribonucleoprotein antibody or normal donors. The effective factors for IgG association in SLE sera were absorbed with normal peripheral blood lymphocytes or T cells. Anti-T cell IgG cytotoxic activity strongly correlated with T cell IgG association (P less than 0.01). T cell-associated IgG was not removed by stripping of cell membrane IgG from living cells by acid buffer treatment; indirect immunofluorescence of cells fixed after 2-4 d of culture revealed cytoplasmic IgG staining. IgG anti-T cell antibodies appeared to associate inside the cell membrane or to penetrate into the cytoplasm of cells. T cell Fc receptor blocking by heat-aggregated IgG or anti-beta 2-microglobulin antibody did not alter IgG cell association. Since pepsin-digested SLE sera showed no T cell association activity, whole IgG antibody molecules appeared to be necessary for interaction with cultured T cells. In addition, reduction and alkylation of active SLE sera completely nullified T cell reactivity. When normal T cells were cultured with SLE sera showing marked IgG T cell association, viability of cultured T cells decreased rapidly after 4 d, which suggests that IgG anti-T cell antibodies were associated with cell destruction. IgG cell-associating antilymphocyte antibodies present in SLE sera may cause T cell disturbances in vivo and may be related to the lymphocytopenia present in SLE patients.     

Use of monoclonal antibodies for the characterization of novel DNA-binding proteins recognized by human autoimmune sera

Autoantibodies to a DNA-binding heterodimer consisting of 70,000 and 80,000 dalton subunits were identified in 30-50% of human autoimmune sera from patients with systemic lupus erythematosus (SLE), mixed connective tissue disease (MCTD), and scleroderma. Three murine monoclonal antibodies (mAb) against the heterodimer were produced in BALB/c mice by immunizing with isolated human B cell nuclei. By immunofluorescence, the mAb and autoimmune sera demonstrated both speckled nucleoplasmic staining and diffuse nucleolar staining in all human cell types examined. The nucleoplasmic staining was sensitive to DNase but not RNase pretreatment, while the nucleolar staining was sensitive to RNase but not DNase pretreatment. Biochemical characterization of the 70,000 and 80,000 dalton proteins using the mAb indicated that two forms of the antigen, with different mobilities on sucrose gradients, are present in human B cells. A 10 S form consists of the physically associated 70,000 and 80,000 dalton proteins, while a larger, 10-20 S form probably represents the same two proteins bound to DNA. Binding of the proteins to nucleolar RNA could not be confirmed in biochemical studies. These studies indicate that non-histone, DNA-binding proteins may be more frequently recognized by autoantibodies in SLE, MCTD, and scleroderma than has been previously recognized. Along with previous studies on RNA-binding proteins such as Sm, RNP, Ro, and La, the present findings suggest that nucleic acid-binding proteins, as a class, may be particularly frequent targets of autoimmunity in SLE and related disorders.     

Paraneoplastic anti-neuronal nuclear IgG autoantibodies (type I) localize antigen in small cell lung carcinoma.

Type I anti-neuronal nuclear autoantibodies (ANNA-I, also known as "Hu") are a distinctive serologic marker of small cell lung carcinoma (SCLC) in patients who have peripheral neuropathies or encephalomyeloradiculopathies. A tumor antigen reactive with these antibodies has been identified by other investigators by Western blot analyses, but an antigen has not been localized in SCLC immunohistochemically. We therefore tested, by indirect immunofluorescence, the sera of 49 sequential ANNA-I-positive patients and 30 control subjects for IgG reactive with SCLC. Two tumor cell lines were tested, one established from the primary tumor of a patient with Lambert-Eaton syndrome and the second from the metastatic lesion of a patient without neurologic disease. IgG in all ANNA-I-positive sera bound to both tumors. In most instances, the pattern resembled that seen in neurons, with strong homogeneous nuclear staining, sparing of nucleoli, and faint cytoplasmic staining. A highly significant correlation was noted between endpoint dilutions obtained on SCLC substrates and on central and peripheral neurons (r = 0.863; P less than 0.001). IgG in 3 of 30 control sera bound in low titer to SCLC cells but not to neurons, and 9 control sera contained non-organ-specific anti-nuclear antibodies (ANA). The ANA IgG was absorbed equivalently by homogenates of SCLC or colonic adenocarcinoma cells. In contrast, the reactivity of ANNA-I IgG with cerebellar and myenteric plexus neurons was absorbed only by homogenized SCLC cells. These findings suggest that SCLC antigens account for all neuronal reactivity of ANNA-I. IgG of this specificity may serve as a useful reagent for identifying SCLC cells in surgical pathologic and cytologic specimens.