Detection in whole blood of autoantibodies for the diagnosis of connective tissue diseases in near patient testing condition.

A novel technology, photonic ring immunoassay (PRI), for detecting 12 autoantibodies simultaneously in whole blood in less than 15 minutes was evaluated by comparing results from 235 clinically diagnosed patients with standard laboratory tests. The overall agreement was greater than 91% for 10 of the 12 assays, with positive percent agreement greater than 89% for 9 of the assays and negative percent agreement greater than 91% for 10 of them. Thus, the clinical sensitivities and specificities were similar for the 2 methods. In addition, 199 normal blood donors were tested on the ANA 12 PRI, yielding specificities greater than 97.5% for all assays. This proof of concept study shows that this new system is suitable for point of care testing for clinically useful autoantibodies, allowing the doctor to have test results in minutes rather than days.

Photonic ring resonance is a versatile platform for performing multiplex immunoassays in real time.

Photonic ring resonance is a property of light where in certain circumstances specific wavelengths are trapped in a ring resonator. Sensors based on silicon photonic ring resonators function by detecting the interaction between light circulating inside the sensor and matter deposited on the sensor surface. Binding of biological material results in a localized change in refractive index on the sensor surface, which affects the circulating optical field extending beyond the sensor boundary. That is, the resonant wavelength will change when the refractive index of the medium around the ring resonator changes. Ring resonators can be fabricated onto small silicon chips, allowing development of a miniature multiplex array of ring based biosensors. This paper describes the properties of such a system when responding to the refractive index changed in a simple and precise way by changing the ionic strength of the surrounding media, and in a more useful way by the binding of macromolecules to the surface above the resonators. Specifically, a capture immunoassay is described that measures the change of resonant wavelength as a patient serum sample with anti-SS-A autoantibodies is flowed over a chip spotted with SS-A antigen and amplified with anti-IgG. The technology has been miniaturized and etched into a 4×6mm silicon chip that can measure 32 different reactions in quadruplicate simultaneously. The variability between 128 rings on a chip as measured by 2M salt assays averaged 0.6% CV. The output of the assays is the average shift per cluster of 4 rings, and the assays averaged 0.5% CV between clusters. The variability between chips averaged 1.8%. Running the same array on multiple instruments showed that after some improvements to the wavelength referencing system, the upper boundary of variation was 3% between 13 different instruments. The immunoassay displayed about 2% higher variability than the salt assays. There are several outstanding features of this system. The amount of antigen used on the chip for each test is around 200 picograms, only a few microliters of sample is necessary, and the assays take <10min.

Disability and quality-of-life are not influenced by the prevalence of autoantibodies in early rheumatoid arthritis patients--results of the Brasília Cohort.

INTRODUCTION: Although many studies have suggested that the presence of autoantibodies, such as rheumatoid factor (RF) and/or anti-cyclic citrullinated peptide (anti-CCP) in rheumatoid arthritis (RA) are predictors of joint damage, the association with disability and quality of life questionnaires are not known. OBJECTIVES: To evaluate the correlation between the Health Assessment Questionnaire (HAQ) and the Medical Outcomes Study 36-Item Short-Form Health Survey (SF-36) scores with serological markers, such as RF, anti-CCP, and anti-citrullinated vimentin (anti-Sa). PATIENTS AND METHODS: Sixty five patients with early RA (ERA) from the Brasília Cohort of ERA were evaluated. Serology tests (ELISA) for RF (IgM, IgG, and IgA), anti-CCP (CCP2, CCP3, and CCP3.1), and anti-Sa were performed, with the application of the HAQ and SF-36 questionnaires in the initial evaluation. RESULTS: The mean age was 45 years, with a female predominance (86%). At the initial evaluation, RF was positive in 32 individuals (49.23%), anti-CCP in 34 (52.3%), and anti-Sa in nine (13.8%). The initial HAQ score was 1.8. The SF-36 scores were as follow: role-emotional, 19.3; social functioning, 43.1; bodily pain, 25.43; general health, 57.6; mental health, 48.1; vitality, 49.5; role-physical, 4.6; and physical functioning, 24.7. The HAQ and SF-36 scores did not vary with autoantibody levels. CONCLUSION: In many patients, ERA has a major impact on physical ability and health-related quality of life. Although RF and anti-CCP tests have been related with joint destruction and worse clinical prognosis, there is no correlation with the results of questionnaires of quality of life and disability.

Incapacitação e qualidade de vida não são influenciadas pela prevalência de autoanticorpos em pacientes com artrite reumatoide inicial - resultados da Coorte Brasília / Disability and quality-of-life are not influenced by the prevalence of autoantibodies in early rheumatoid arthritis patients - results of the Brasília Cohort

INTRODUÇÃO: Embora muitos estudos sugiram que a presença de autoanticorpos, tais como fator reumatoide (FR) e/ou antipeptídeos citrulinados cíclicos (anti-CCP), sejam preditores de danos articulares na artrite reumatoide (AR), a associação entre os questionários de incapacidade e de qualidade de vida ainda são desconhecidos. OBJETIVOS: Avaliar a correlação entre os questionários Health Assessment Questionnaire (HAQ) e Medical Outcomes Study 36-Item Short-Form Health Survey (SF-36) com marcadores como FR, anti-CCP e antivimentina citrulinada (anti-Sa). PACIENTES E MÉTODOS: Foram avaliados no momento do diagnóstico 65 pacientes da Coorte Brasília com AR inicial. Foram realizadas sorologias (ELISA) para FR (IgM, IgG e IgA), anti-CCP (CCP2, CCP3 e CCP3.1) e anti-Sa, com a aplicação do HAQ e SF-36 na avaliação inicial. RESULTADOS: A idade média foi de 45 anos, predominando o gênero feminino (86%). Na avaliação inicial, o FR foi positivo em 32 indivíduos (49,23%); anti-CCP em 34 indivíduos (52,3%); e anti-Sa em nove indivíduos (13,8%). O escore inicial do HAQ foi de 1,8. Os escores dos domínios do SF-36 foram: emocional, 19,3; social, 43,1; dor, 25,43; estado geral, 57,6; saúde mental, 48,1; vitalidade, 49,5; físico, 4,6; e limitação por aspecto físico, 24,7. HAQ e escores do SF-36 não variaram com os níveis de autoanticorpos. CONCLUSÃO: Muitos pacientes com AR inicial apresentam comprometimento na qualidade de vida relacionada aos domínios da capacidade física e mental. Embora FR e anti-CCP tenham sido relacionados com dano articular e pior prognóstico clínico, não há correlação entre os questionários e as avaliações da qualidade de vida e incapacidade.

INTRODUCTION: Although many studies have suggested that the presence of autoantibodies, such as rheumatoid factor (RF) and/or anti-cyclic citrullinated peptide (anti-CCP) in rheumatoid arthritis (RA) are predictors of joint damage, the association with disability and quality of life questionnaires are not known. OBJECTIVES: To evaluate the correlation between the Health Assessment Questionnaire (HAQ) and the Medical Outcomes Study 36-Item Short-Form Health Survey (SF-36) scores with serological markers, such as RF, anti-CCP, and anti-citrullinated vimentin (anti-Sa). PATIENTS AND METHODS: Sixty five patients with early RA (ERA) from the Brasília Cohort of ERA were evaluated. Serology tests (ELISA) for RF (IgM, IgG, and IgA), anti-CCP (CCP2, CCP3, and CCP3.1), and anti-Sa were performed, with the application of the HAQ and SF-36 questionnaires in the initial evaluation. RESULTS: The mean age was 45 years, with a female predominance (86%). At the initial evaluation, RF was positive in 32 individuals (49.23%), anti-CCP in 34 (52.3%), and anti-Sa in nine (13.8%). The initial HAQ score was 1.8. The SF-36 scores were as follow: role-emotional, 19.3; social functioning, 43.1; bodily pain, 25.43; general health, 57.6; mental health, 48.1; vitality, 49.5; role-physical, 4.6; and physical functioning, 24.7. The HAQ and SF-36 scores did not vary with autoantibody levels. CONCLUSION: In many patients, ERA has a major impact on physical ability and health-related quality of life. Although RF and anti-CCP tests have been related with joint destruction and worse clinical prognosis, there is no correlation with the results of questionnaires of quality of life and disability.

Third generation anti-citrullinated peptide antibody assay is a sensitive marker in rheumatoid factor negative rheumatoid arthritis.

INTRODUCTION: We compared 2 anti-citrullinated protein antibody (ACPA) assays using a routine patient cohort. METHODS: Two-hundred ninety-five sera were collected from patients for whom ACPA was ordered and tested for ACPA by QUANTA Lite® CCP 3 (INOVA Diagnostics, Inc., San Diego) and EliA® CCP (CCP, Phadia, Germany). Rheumatoid factor (RF) was determined using Quantex RF(II) (Biokit, Spain). RESULTS: Acceptable qualitative (96.6%, kappa=0.93) and quantitative agreements (Spearman rho=0.77; p<0.0001) were observed between the two ACPA assays. Nine samples were CCP3+/CCP2- and one sample was CCP2+/CCP3-. Of the 9 CCP3+/CCP2- patients, 6 (66.7%) had RA, one patient had ankylosing spondylitis, one osteoarthritis and one psoriatic arthritis. The CCP3-/CCP2+ patient had juvenile RA. At the manufacturer's cut-offs, the sensitivities and specificities were 77.3%/98.1% (CCP2), 81.6%/96.8% (CCP3) and 65.2%/89.6% (RF), respectively. At 98.7% specificity level, the sensitivities in the total cohort were 59.6% (CCP2) and 69.5% (CCP3) while the sensitivities in the RF-negative group were 49.0% (CCP2) and 57.1% (CCP3). In the RF-negative group, sensitivities for patients with a disease duration of ≤ 5years were 38.7% (CCP2) and 51.6% (CCP3). CONCLUSION: Discrimination between RA and non-RA patients was better using CCP3, most pronounced in RF-negative RA.

IgM autoantibodies to distinct apoptosis-associated antigens correlate with protection from cardiovascular events and renal disease in patients with SLE.

Emerging evidence suggests that there are IgM-autoantibodies that may play protective roles in SLE. While IgM are often considered polyreactive, we postulate that there are distinct sets of IgM-autoantibodies of defined autoreactive specificities relevant to different features of SLE. We examined the relationships between levels of IgM natural autoantibodies (NAbs) to apoptosis-associated phosphorylcholine (PC) or malondialdehyde (MDA) antigens, with lupus-associated autoantibodies and features of disease, in 120 SLE patients. IgM anti-PC was significantly higher in patients with low disease activity and less organ damage determined by the SELENA-SLEDAI, the physician's evaluation and the SLICC damage score. Furthermore, IgM anti-PC was significantly higher in patients without cardiovascular events. In contrast, IgM anti-cardiolipin and IgM anti-dsDNA were significantly higher in patients without renal disease. These results support the hypothesis that some IgM autoantibodies are part of a natural immune repertoire that provide homeostatic functions and protection from certain clinical lupus features.

Levamisole-induced vasculopathy: a report of 2 cases and a novel histopathologic finding.

Although cocaine-induced pseudovasculitis and urticarial vasculitis have been reported in the past, levamisole-induced vasculopathy with ecchymosis and necrosis, termed here LIVEN, has only recently been described in association with cocaine use. Levamisole, a veterinary antihelminthic agent used previously as an immunomodulating agent, is present as a "cutting agent" in approximately two-thirds of the cocaine currently entering the United States. Levamisole is believed to potentiate the effects of cocaine and may also be used as a "signature" for tracing its market distribution. Herein, we report 2 cases of LIVEN in patients with histories of chronic cocaine use. In both the cases, a temporal association with neutropenia preceding the eruption was noted. A novel histopathologic finding present only in the second case was the presence of extensive interstitial and perivascular neovascularization. Our 2 cases reaffirm that neutropenia may precede the cutaneous eruption of LIVEN. Case 2 extends the spectrum of histopathologic findings to include the novel phenomenon of neovascularization-hitherto unreported in this entity.

Measurement of autoantibodies in pediatric-onset systemic lupus erythematosus and their relationship with disease-associated manifestations.

OBJECTIVE: To evaluate an autoantibody profile in pediatric-onset systemic lupus erythematosus (SLE) patients to determine clinical and statistical associations with disease-associated manifestations. METHODS: Sera from 53 SLE patients and 22 healthy individuals were collected. Antibodies to C1q, histone, chromatin, ribosomal P, dsDNA, and high-avidity dsDNA were measured by enzyme-linked immunosorbent assays. Patient records were evaluated for clinical and laboratory associations. RESULTS: The most prevalent autoantibodies found in the SLE cohort were anti-C1q antibodies (n = 32, 60%), which correlated significantly with proteinuria and decreased complement levels (P < 0.05). Anti-C1q and antihistone antibodies were significantly elevated in patients with class III/IV nephritis compared with class I/II/V nephritis (P = 0.041). SLE patients with active nephritis at the time of sample collection demonstrated significantly elevated levels of anti-C1q antibodies compared with those without active nephritis, also exhibiting 100% sensitivity for active nephritis, proteinuria, and urinary casts. Antibodies to C1q, dsDNA, histone, and chromatin were significantly elevated in patients with active disease (P < 0.01). Chart-documented anti-dsDNA antibodies were positive in 28 SLE patients, INOVA anti-dsDNA antibodies in 25 patients, and high-avidity anti-dsDNA antibodies in 8 patients. Antihistone correlated significantly with leukopenia and hemolytic anemia (P < 0.05). CONCLUSIONS: This study indicates the importance of measuring anti-C1q antibodies in pediatric-onset SLE patients because elevated anti-C1q antibodies may be more indicative of renal disease activity, showing significant correlation with proteinuria, urinary casts, and active nephritis. Antibodies to C1q, histone, chromatin, and dsDNA exhibited the strongest association with clinical features, indicating the importance of measuring all of these antibodies in pediatric-onset SLE patients.

Baseline HAQ and SF-36 questionnaire scores cannot predict clinical remission, radiographic progression or the need for biological therapy in a three-year prospective study of a Brazilian early rheumatoid arthritis cohort.

This study evaluates prospectively whether baseline scores [Health Assessment Questionnaire (HAQ) and SF-36] can predict clinical and radiographic evolution in a cohort of early rheumatoid arthritis (RA) during a 3-year follow-up. Forty consecutive early RA patients were followed for 3 years, while receiving standardized treatment according to a pre-established protocol. HAQ and SF-36 were administered at the initial evaluation and at 3, 6, 12, 18, 24 and 36 months. Hands and feet radiographs were obtained at the initial evaluation and at 12, 24 and 36 months. Preselected outcomes were the occurrence of radiographic erosions, the achievement of an EULAR remission, low disease activity status and the need for biological therapy. The mean age at onset was 45 years with a 90% female predominance. Erosions were found in 42% of patients at T0 and in 70% after 3 years (P < 0.001). At T0, the proportion of patients in remission, low, moderate or high disease activity was 0, 0, 7.5 and 92.5% and 22.5, 7.5, 32.5 and 37.5%, respectively, at 3 years. The mean baseline HAQ score was 1.89 and 0.77 by the third year (P < 0.0001). Most SF-36 domains showed significant improvement except for general state and vitality. Biological therapy was deemed necessary in 22.5% of patients. The initial HAQ and SF-36 scores were not associated with clinical remission, bone erosions or the need for biological therapy at 36 months. The HAQ and SF-36 scores measured at baseline could not predict at 3 years, the preselected outcomes in a Brazilian cohort.

Atypical clinical presentation of a subset of patients with anti-RNA polymerase III--non-scleroderma cases associated with dominant RNA polymerase I reactivity and nucleolar staining.

INTRODUCTION: Anti-RNA polymerase III (RNAP III) antibodies are highly specific markers of scleroderma (systemic sclerosis, SSc) and associated with a rapidly progressing subset of SSc. The clinical presentation of anti-RNAP III positive patients, onset of Raynaud's phenomenon (RP) and SSc in unselected patients in a rheumatology clinic were evaluated. METHODS: Autoantibodies in sera from 1,966 unselected patients (including 434 systemic lupus erythematosus (SLE), 119 SSc, 85 polymyositis/dermatomyositis (PM/DM)) in a rheumatology clinic were screened by radioimmunoprecipitation. Anti-RNAP III positive sera were also tested by immunofluorescence antinuclear antibodies and anti-RNAP III ELISA. Medical records of anti-RNAP III positive patients were reviewed. RESULTS: Among 21 anti-RNAP III positive patients, 16 met the American College of Rheumatology (ACR) SSc criteria at the initial visit but 5 did not; diagnoses were vasculitis, early polyarthritis, renal failure with RP, interstitial lung disease, and Sjögren's syndrome. The first two patients developed rapidly progressive diffuse SSc. An additional case presented with diffuse scleroderma without RP and RP developed two years later. Anti-RNAP III antibodies in these 6 cases of atypical clinical presentation were compared with those in 15 cases of typical (SSc with RP) cases. Anti-RNAP III levels by ELISA were lower in the former group (P = 0.04 by Mann-Whitney test) and 3 of 6 were negative versus only 1 of 15 negative in the latter (P < 0.05 by Fisher's exact test). Three cases of non-SSc anti-RNAP III positive patients had predominant reactivity with RNAP I with weak RNAP III reactivity and had a strong nucleolar staining. Three anti-RNAP III patients, who did not have RP at the initial visit, developed RP months later. Scleroderma developed prior to RP in 5 out of 16 (31%) in the anti-RNAP III group, but this was rare in patients with other autoantibodies. The interval between the onset of RP to scleroderma was short in anti-RNAP III positive patients. CONCLUSIONS: Anti-RNAP III antibodies are highly specific for SSc; however, a subset of anti-RNAP III positive patients do not present as typical SSc. The interval between RP and scleroderma in this group is short, and 31% of patients developed scleroderma prior to RP in this group. Anti-RNAP III positive patients may not present as typical SSc and detecting anti-RNAP III may have predictive value.

Cocaine-related retiform purpura: evidence to incriminate the adulterant, levamisole.

The term 'cocaine-induced pseudovasculitis' was coined to encompass a constellation of clinical and laboratory findings which mimics a systemic vasculitis but lacks confirmatory evidence of vasculitis on biopsy. Antineutrophil cytoplasmic antibodies reacting with human neutrophil elastase (HNE) have been reported to distinguish the cocaine-related syndrome from a true autoimmune vasculitis. Published cases of retiform purpura related to cocaine use are rare and an etiologic role for levamisole, a common adulterant of cocaine, has been postulated. We describe two female patients aged 39 and 49 years with cocaine-related retiform purpura, mainly affecting the legs. The initial clinical and serological profile in case 1 led to a suspicion of anti-phospholipid syndrome and in case 2 to Wegener's granulomatosis with an unexplained associated neutropenia. Skin biopsies revealed a mixed pattern of leukocytoclastic vasculitis and microvascular thrombosis in case 1 and pure microvascular thrombosis in case 2. Identification of anti-HNE antibodies in both patients linked their disease to cocaine. The mixed vasculopathic pattern in case 1 and the associated neutropenia in case 2, both known adverse effects of levamisole, point to this as the true etiologic agent. Urine toxicology shortly after a binge of cocaine use in each case was positive for levamisole.

'Actin-reactive' discriminated from 'non-actin-reactive' smooth muscle autoantibody by immunofluorescence reactivity with rat epithelial cell line.

AIMS: To compare smooth muscle antibody (SMA) patterns in tissue sections with patterns in an immunofluorescence assay (IFA) using a rat intestinal epithelial cell line and results from an F-actin IgG ELISA. METHODS: SMA positive sera (n = 188) were classified by immunofluorescence staining of rodent kidney, stomach and liver sections as SMA-T (tubules) (n = 124) or SMA-V (vessels) (n = 64). The F-actin pattern on the rat epithelial cell line was identified by immunofluorescence staining of actin cables that was confirmed by dual immunofluorescence co-localisation with phalloidin. RESULTS: Of 124 SMA-T positive sera, 123 reacted with the epithelial cell line and 120 with F-actin by ELISA, giving sensitivity for detection of anti-F-actin antibody of 99% and 97%, respectively. Of 64 SMA-V positive sera, four reacted with the epithelial cell line (6%) and 41 with F-actin by ELISA (64%). Tests of 493 normal blood donors and 100 disease controls yielded specificities of 584/593 (98.5%) and 562/593 (94.8%) for the cell line IFA and F-actin ELISA, respectively. CONCLUSIONS: The rat epithelial cell line IFA is a robust diagnostic assay for anti-F-actin antibody that can either replace the routine screening for actin-reactive SMA-T antibody in tissue sections or be used as a confirmatory assay for anti-F-actin antibody after screening by F-actin ELISA or on rodent tissue sections by IFA.

Prevalence of antibodies to Ro-52 in a serologically defined population of patients with systemic sclerosis.

BACKGROUND: Antibodies against Ro-52 have been described in patients with a broad spectrum of autoimmune disease, most commonly in association with anti-Ro-60 in systemic lupus erythematosus and Sjogrens syndrome. However, in inflammatory myositis anti-Ro-52 is frequently present without anti-Ro-60 and is closely linked to the presence of aminoacyl-tRNA synthetase (aats) antibodies. To date there have been no comprehensive reports on the frequency of anti-Ro-52 in systemic sclerosis (SSc), a disease characterised by hallmark autoantibodies that occur in non-overlapping subsets. Clinically, each antibody-defined group has a distinct pattern of organ involvement, some featuring myositis. OBJECTIVES: To determine the frequency of anti-Ro-52 in serologically defined groups of SSc patients and to investigate a possible link with myositis-associated autoantibodies. METHODS: Serum samples from 1010 patients with SSc and 55 and 32 patients with anti-aats and anti-Ku respectively were tested for the presence of anti-Ro-52 using a commercial ELISA. RESULTS: The prevalence of anti-Ro-52 was 15-38% in nine of the eleven sub-groups. There were no significant differences in mean anti-Ro-52 levels in these groups with the exception of that defined by the presence of anti-U1-RNP. In the remaining groups defined by anti-Ro-60 and anti-aats, anti-Ro-52 was present in 92% and 100% respectively. In sera from non-SSc patients with anti-aats, anti-Ro-52 was detected in 64%. CONCLUSION: Anti-Ro-52 is present throughout the SSc population. It is neither more prevalent in the myositis-associated antibody groups nor does it segregate with any other major SSc-specific autoantibodies. The co-existence of anti-Ro-52 with both anti-Ro-60 and anti-aats is confirmed.

Anti-C1q antibodies: association with nephritis and disease activity in systemic lupus erythematosus.

BACKGROUND: Anti-C1q antibodies have been described in systemic lupus erythematosus (SLE) as well as in other connective tissue diseases. They have been considered as a marker for disease activity and presence of nephritis. OBJECTIVE: The aim of this study was to determine the prevalence of anti-C1q antibodies in Brazilian lupus patients as well as analyze their association with different clinical and serologic parameters. METHODS: Sera from 81 SLE patients, based on the American College of Rheumatology (ACR) criteria, were collected from a lupus referral outpatient clinic in Salvador, Brazil. Antibodies to C1q were detected by an enzyme-linked immunoassay (ELISA) kit and antibodies to other cellular antigens identified by indirect immunofluorescence on HEp-2 cell substrate (ANA), or Crithidia luciliae (dsDNA), and to nucleosome by ELISA. A cutoff of 20 U was established for anti-C1q and anti nucleosome assays. RESULTS: Anti-C1q antibodies were detected in 39.5% (32/81) of SLE sera. The presence of anti-C1q antibodies was associated with proteinuria (P=0.028) but not with other laboratory or clinical features, such as anti nucleosome or anti-dsDNA antibodies, hematuria, urinary casts or renal failure, leukopenia, pericarditis, pleuritis, malar rash, seizures, and psychosis. There was a positive correlation between the titers of anti-C1q antibodies and the systemic lupuis erythematosus disease activity index (SLEDAI) score (r=0.370; P=0.001). CONCLUSION: This study in Brazilian SLE patients confirms previous findings of the association of anti-C1q antibodies with nephritis and disease activity.

Anti-chromatin (anti-nucleosome) antibodies: diagnostic and clinical value.

Anti-chromatin (nucleosome) autoantibodies were one of the first autoantibodies ever detected since they make up the majority of antibodies causing LE Cell formation. The prevalence of anti-chromatin antibodies in systemic lupus erythematosus (SLE) varies from 50% to 100%, being similar to that of the classical positive LE cell. The presence of these antibodies can be used, in conjunction with clinical findings and other laboratory tests, to help in the diagnosis of SLE and drug-induced lupus. Anti-chromatin antibodies have also been found in a lesser percentage of other autoimmune disorders such as primary Sjögren's syndrome and primary antiphospholipid syndrome. The presence of anti-chromatin antibodies has also been linked to glomerulonephritis and disease activity in SLE patients. Recent studies demonstrated the induction of anti-chromatin (anti-nucleosome) antibodies after an anti-tumour necrosis factor (TNF)-alpha agent treatment.

Antinucleosome antibodies in primary antiphospholipid syndrome: a hint at systemic autoimmunity?

BACKGROUND: Antinucleosome antibodies (anti-NCS) are reported to be highly sensitive and specific for systemic lupus erythematosus (SLE) and to correlate with disease activity. They may appear in early stages of the disease, in particular before anti-dsDNA antibodies, being a potential marker for identifying patients susceptible to SLE. Patients with primary antiphospholipid syndrome (PAPS) may develop full-blown SLE but there is no evidence for markers predictive for that. AIM: To evaluate whether anti-NCS may be predictors for full-blown or lupus like disease (LL) in a cohort of PAPS patients. METHODS: A multicentric cohort of 105 PAPS patients was tested for IgG/IgM anti-NCS by using a home made assay with H1-stripped chromatin as antigen. RESULTS: Eighty-one out of 105 (77%) of the patients were positive for anti-NCS; medium-high titre results were present only in 49/105 (46%). Anti-NCS were more frequently detected in PAPS+LL, but no relationship with clinical/serological features was found, except for a weak correlation with anti-dsDNA antibodies. Two PAPS patients evolved into full-blown SLE during the follow-up and displayed high titre anti-NCS many years before. CONCLUSIONS: Our findings suggest that anti-NCS might be added to the mosaic of autoimmune phenomena characterizing PAPS patients and in particular those with more chance to evolve to SLE.

Multiplex assays to diagnose celiac disease.

Patients with celiac disease are sensitive to the gluten fractions of wheat. Symptoms include gastrointestinal problems and a failure to thrive in children, but may range from headaches to general malaise in adults. Thus, it is difficult to diagnose celiac disease by symptoms alone. The standard diagnostic criteria include the presence of the characteristic anti-gliadin or anti-tissue transglutaminase antibodies (anti-tTG) in serum, flattened mucosa on intestinal biopsy, and improved symptoms on a gluten-free diet. Because of the ease of use of the tTG enzyme-linked immunosorbent assay (ELISA) compared to endomysial by indirect immunofluorescence assay, there has been much more screening for celiac disease in recent years. This increased screening showed that celiac disease was more prevalent than previously believed. We compared a new multiplex assay that includes a novel form of deamidated gliadin and recombinant human tTG as the antigens to other assays using standard antigens. In addition, the new assay detects the presence of selective IgA deficiency, which shows a 10-fold increase in prevalence in patients with celiac disease compared to the general population. The combination of sensitivity and specificity of the new multiplex assay was equal or better than those for standard assays. Thus the performance, ease of use, and ability to measure three clinically important parameters in a single test make the new multiplex assay a viable alternative to standard assays in a clinical lab.

Antibodies to RNA polymerase III in systemic sclerosis detected by ELISA.

OBJECTIVE: To determine serological and clinical variables associated with anti-RNA polymerase III (RNAP-III) antibodies in patients with systemic sclerosis (SSc) using a new ELISA method. METHODS: Sera from 242 patients with SSc were collected from 14 Canadian clinics. Control sera were from 287 blood donors, and 42 patients with infectious disease, 30 with rheumatoid arthritis (RA), and 30 with systemic lupus erythematosus (SLE). Antibodies to RNAP-III were detected by an ELISA kit and antibodies to other cellular antigens were identified by indirect immunofluorescence (IIF) on HEp-2 cell substrate, line immunoassay, immunoprecipitation of recombinant protein, and addressable laser bead immunoassay (ALBIA). RESULTS: Anti-RNAP-III antibodies were detected in 47/242 (19.4%) SSc sera, 0% RA and SLE sera, 1/287 blood donor sera, and 2/42 infectious disease sera. Diffuse disease (59.5%) was more common than limited disease (36.1%) in the anti-RNAP-III-positive patients (p = 0.006) and there was an association between the presence of anti-RNAP-III and kidney and joint/tendon involvement, but there was no association with a nucleolar IIF pattern, lung involvement, or other clinical indicators. There was a negative association between the presence of anti-RNAP-III antibodies and anticentromere by IIF (p = 0.00004) and anti-Scl-70 by ALBIA (p = 0.0005) and line immunoassay (p = 0.003), suggesting a virtually exclusive presence of these antibodies in SSc. CONCLUSION: Anti-RNAP-III autoantibodies were found in nearly 20% of SSc patients but in less than 1% of controls, thus detection of this antibody is a useful marker to help diagnose SSc. As well, this antibody has prognostic utility, since it is associated with scleroderma renal crisis and the diffuse cutaneous form of SSc.

Anti-chromatin (anti-nucleosome) autoantibodies.

Anti-chromatin autoantibodies were one of the first autoantibodies ever detected since they make up the majority of antibodies causing LE Cell formation. Anti-chromatin autoantibodies have had many names over the last few decades: LE cell factor; anti-nucleosome; anti-deoxyribonucleoprotein (DNP); and anti-(H2A-H2B-DNA). These autoantibodies are found in approximately 75% of people with systemic lupus erythematosus and up to 100% of people with drug-induced lupus. They are also found in 20-50% of patients with autoimmune hepatitis type I (lupoid hepatitis). Anti-chromatin are not generally found in any other disease, thus showing very good sensitivity and specificity for patients with lupus, drug-induced lupus and lupoid hepatitis. A number of studies have shown that in patients with lupus, anti-chromatin often correlates better with kidney disease than anti-DNA. Recent genetic analyses of murine models of lupus have identified at least three loci that work together to cause anti-chromatin antibodies and glomerulonephritis in mice. It will be an important breakthrough when the functions of the genes at these loci are identified.