Label-free electrochemical impedimetric immunosensor for sensitive detection of IgM rheumatoid factor in human serum.

The authors report a label-free and direct detection of rheumatoid factor- Immunoglobulin M (IgM-RF) based on an impedimetric-interdigitated wave type microelectrode array (IDWµE). IDWµE was functionalized with a self-assembled monolayer (SAM) of thioctic acid for antigen immobilization. The SAM functionalized IDWµE were characterized by atomic force microscopy, Energy Dispersive X-Ray Spectroscopy, and X-ray photoelectron spectroscopy. The covalent immobilization of IgG-Fc onto the SAM modified electrode arrays was characterized morphologically via AFM and electrochemically via cyclic voltammetry and electrochemical impedance spectroscopy. Impedimetric measurements in the presence of redox probe (Potassium ferrocyanide and potassium ferricyanide) showed a significant change in both the impedance spectrum and charge transfer resistance upon IgM-RF binding. Impedance measurements were target specific and linear with an increase in IgM-RF concentrations between 1 and 200 IU mL-1 in redox probe and human serum, respectively. The sensor showed detection limits of 0.6 IU mL-1 in the presence of redox probe and 0.22 IU mL-1 in the human serum. The biosensor exhibited good reproducibility (relative standard deviation (RSD), 4.96%) and repeatability (RSD, 2.31%) with an acceptable selectivity towards IgM-RF detection. The sensor also showed a good stability for 3 weeks at 4 °C in 1X PBS. Therefore, the sensitive and stable immunosensor exhibiting IDWµE features can be integrated with a miniaturized potentiostat to develop a sensing system that detects IgM-RF for point-of-care applications.

Falange de marfil: un hallazgo específico y poco conocido en artritis psoriásica / Ivory phalanx: a specific and not widely known finding in psoriatic arthritis

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IgM and IgA rheumatoid factors purified from rheumatoid arthritis sera boost the Fc receptor- and complement-dependent effector functions of the disease-specific anti-citrullinated protein autoantibodies.

Rheumatoid factors (RF) and the disease-specific anti-citrullinated protein autoantibodies (ACPA) coexist in the joints of rheumatoid arthritis (RA) patients where they probably contribute to synovitis. We investigated the influence of IgM and IgA RF on the FcR- and complement-dependent effects of ACPA immune complexes (ACPA-IC). When stimulated by ACPA-IC formed in the presence of IgM RF or IgA RF fractions purified from RA serum pools, M-CSF-generated macrophages skewed their cytokine response toward inflammation, with increases in the TNF-α/IL-10 ratio and in IL-6 and IL-8 secretion, and decreases in the IL-1Ra/IL-1ß ratio. In the IgM RF-mediated amplification of the inflammatory response of macrophages, the participation of an IgM receptor was excluded, notably by showing that they did not express any established receptor for IgM. Rather, this amplification depended on the IgM RF-mediated recruitment of more IgG into the ACPA-IC. However, the macrophages expressed FcαRI and blocking its interaction with IgA inhibited the IgA RF-mediated amplification of TNF-α secretion induced by ACPA-IC, showing its major implication in the effects of RF of the IgA class. LPS further amplified the TNF-α response of macrophages to RF-containing ACPA-IC. Lastly, the presence of IgM or IgA RF increased the capacity of ACPA-IC to activate the complement cascade. Therefore, specifically using autoantibodies from RA patients, the strong FcR-mediated or complement-dependent pathogenic potential of IC including both ACPA and IgM or IgA RF was established. Simultaneous FcR triggering by these RF-containing ACPA-IC and TLR4 ligation possibly makes a major contribution to RA synovitis.

False decrease of HBsAg S/CO values in serum with high-concentration rheumatoid factors.

OBJECTIVES: To investigate whether high-concentration RFs cause false decrease of S/CO values of serum HBsAg. DESIGN AND METHODS: Serum HBsAg was determined in 100 RF-positive sera using one-step ELISA. Twenty-three HBsAg-negative sera with high-concentration RFs were selected randomly to perform dilution with mixed normal sera. Serum models consisting of HBsAg and high-concentration RFs were made by blending high-concentration RFs and HBsAg-positive sera at the ratio of 9:1. RESULTS: In the 23 samples, one-step ELISA showed that HBsAg-positive rate was 69.57% at dilution of 1:2, 60.87% at dilution of 1:4 and 40.00% at dilution of 1:8, whereas two-step ELISA showed that it was 100% without any dilution. Thirty serum models were made and divided into six groups. Median S/CO value of HBsAg was 3.00 in control, whereas it ranged from 0.13 to 1.78 in the six groups. CONCLUSIONS: High-concentration RFs cause false decrease of S/CO values of HBsAg using one-step ELISA.

Pannus reumatoide: una nomenclatura equívoca / Rheumatoid pannus: an ambiguous nomenclature

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Respuesta: Pannus reumatoide: una nomenclatura equívoca / Reply: Rheumatoid pannus: an ambiguous nomenclature

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Dietary fish and fish oil and the risk of rheumatoid arthritis.

BACKGROUND: Intake of fish oil and oily fish has been reported to improve clinical symptoms in people who have rheumatoid arthritis. Whether the intake of oily fish and fish oil might also protect against the development of rheumatoid arthritis is not known. OBJECTIVE: We investigated the association between intake of oily fish and fish oil supplements and the risk of rheumatoid arthritis in a population-based case-control study. METHODS: The study comprised 1889 incident cases of rheumatoid arthritis and 2145 randomly selected controls recruited from a geographically defined area of Sweden during 1996-2005. Data on the consumption of oily fish and fish oil supplements 5 years preceding enrollment had been obtained through a questionnaire. We calculated odds ratios (ORs) for the development of rheumatoid arthritis, using logistic regression to adjust for age, residential area, body mass index, smoking, and alcohol consumption. RESULTS: Compared with subjects who never or seldom consumed oily fish, the OR for developing rheumatoid arthritis was 0.8 (95% confidence interval = 0.6-1.0) for subjects who consumed oily fish 1-7 times a week. The results did not change notably when stratifying the cases for rheumatoid factor or for antibodies to citrullinated peptide antigens. Similar results were seen for subjects consuming oily fish 1-3 times a month. Cases and controls did not differ in their consumption of fish oil supplements. CONCLUSION: Intake of oily fish was associated with a modestly decreased risk of developing rheumatoid arthritis.

Removal of autoantibodies by 4-mercaptoethylpyridine-based adsorbent.

Extracorporeal immunoadsorption (ECI) therapy using Staphylococcal Protein A columns has proven effective for removing autoantibodies and circulating immune complexes from patients selectively, providing a promising treatment for autoimmune diseases. However, due to the drawbacks of Protein A in terms of cost and stability, the widespread use of Protein A based ECI is limited. In this study, we investigated the feasibility of 4-mercaptoethylpyridine (MEP, MW 139 Da), a simple and inexpensive synthetic compound, as an alternative to Protein A for autoantibody removal therapy. MEP-based adsorbents were prepared by coupling MEP to Sepharose CL-6B. We found that ligand density was an adjustable parameter for the synthesis of adsorbents aiming at different pathogenic factors, depending on the class of antibody. MEP-Sepharose with a ligand density of 98.8 micromol/ml could remove 80% of the anti-double-stranded DNA antibodies from human serum, whereas a ligand density of 64.5 micromol/ml was enough to remove 96% of the rheumatoid factor (RF) in the serum. Moreover, MEP-based adsorbents showed a lower degree of individual differences compared to Protein A-Sepharose. RF removal of 90% was achieved for all 12 serum samples from different individuals. Among the 14 serum samples derived from systemic lupus erythematosus patients, 11 samples had markedly reduced antinuclear antibody titers. In addition, non-specific adsorption of plasma components to MEP-Sepharose was limited, and the binding capacity of the absorbent for IgG was still about 20 mg/ml of gel after 10 cycles. The results indicated that MEP-based adsorbent could offer a new type of adsorber for the treatment of autoimmune diseases.

Molecular studies of rheumatoid factor using pseudobioaffinity membrane chromatography.

Using pseudobioaffinity ligand L-histidine immobilized to poly(ethylene vinyl alcohol) hollow fiber membrane is an interesting approach for the purification of total IgG and its subclasses from untreated serum in a single step. Gentle adsorption and elution conditions of this chromatography system allow efficient recovery of the protein in its native form. This approach was employed for the recovery and molecular study of rheumatoid factor (RF), an anti-IgG autoantibody (AAb) that form immune complexes with autologous IgG Abs in the sera. The purity of the recovered molecule was analyzed on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), revealed a 150-kDa IgG band and an additional approximately 300 kDa band which may be RF bound IgG complex. Since RF is an AAb, the purified protein was studied for its catalytic functions like peptide, DNA, and RNA hydrolyzing activities. The substrate Pro-Phe-Arg-4-methyl-coumaryl-7-amide (PFR-MCA) hydrolyzing activity by total IgG from different patient sera was found to be greater than healthy controls. In an effort to identify the subclass specificity for the proteolytic function, the pre-purified total IgG fractions from rheumatoid arthritis (RA) sera were subjected to rechromatography using a discriminating buffer. In this experiment, the activity was found in the non-retained fractions suggesting IgG2 specificity for the catalytic function. A comparative study between different catalytic functions was performed for IgG separated from individual patient.

Neumotórax bilateral en un paciente con nódulos reumatoideos colonizados por Aspergillus fumigatus / No disponible

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¿Cómo evaluamos una artritis psoriásica? ¿Cuál es el mejor método? / No disponible

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Miocardiopatía y artritis reumatoide. Una paciente en lista de trasplante cardíaco / Rheumatoid arthritis and myocardiopathy. A patient awaiting a heart transplant

La enfermedad cardiovascular es común en la artritis reumatoide. No obstante, la afección miocárdica es inusual y suele cursar de forma asintomática. Presentamos un caso de miocardiopatía dilatada en una paciente con artritis reumatoide e insuficiencia cardíaca progresiva que requirió su inclusión en lista de trasplante cardíaco (AU)

Cardiovascular pathology is common in rheumatoid arthritis. However, myocardial affection is unusual and clinical disease is rare. We report a case of dilated cardiomyopathy in a patient with rheumatoid arthritis and progressive heart failure that required inclusion into a heart transplantation list (AU)

Anticuerpos anticitrulina en la artritis reumatoide / Antibodies to citrullinated peptides in rheumathoid arthritis

La artitris reumatoide es una enfermedad autoinmune caracterizada por la inflamación de las articulaciones sinoviales produciendo destrucción articular. En el suero de estos pacientes pueden aparecer muchos autoanticuerpos, sobre todo el factor reumatoide, que está incluido dentro de los criterios de clasificación de la enfermedad del American College of Rheumatology (ACR), a pesar de que tiene sólo un especificidad moderada para la enfermedad. Los anticuerpos antipéptidos citrulinados han proporcionado al clínico una prueba de gran ayuda para el diagnóstico precoz. Se ha demostrado que pueden preceder en años al comienzo de la enfermedad y son útiles para el diagnóstico y el pronóstico, debido a su buena sensibilidad y especificidad y su relación con el daño estructural. La respuesta inmunológica frente a antígenos citrulinados define un subtipo de artritis reumatoide inmunogenéticamente característico, en el que se ha podido establecer una relación entre factores ambientales y genéticos, y en el que la reacción inmunológica es una parte fundamental de la patogenia de la enfermedad. Se propone una teoría que relaciona su producción con el desarrollo de la inflamación sinovial crónica

Rheumatoid arthritis is a systemic autoimmune disease characterized by chronic inflammation of the synovial joints leading to progressive joint destruction. The serum of these patients contains a large repertoire of autoantibodies, mainly rheumatoid factor, which is part of the ACR classification criteria in spite of having only moderate specificity. Antibodies directed to citrullinated proteins provide clinicians with a valuable tool for early diagnosis. It has been shown that these antibodies can be detected years before presentation of the first symptom and are very useful for diagnosis and prognosis, due to good sensitivity and specificity and prediction of development of erosive disease. The immune response against citrullinated antigens is characteristic of an immuno-genetic subtype of disease, in which the combined role of genes, environmental factors and autoimmunity has become the prime suspected for disease pathogenesis. A model is proposed of how these antibodies are produced and lead to chronic synovial inflammation

Rheumatoid sinovial fluid T cells are sensitive to Apo2L/Trail Regulation

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Medida de Anticuerpos Anti-CCP en suero mediante un nuevo procedimiento de Fluoroenzimoinmunoanálisis / Measure of Antibodies anti-CCP by a new Fluoroenzymoimmunoanalysis in serum

Comparar los resultados obtenidos con un nuevo método automatizado de fluoroenzimoinmunoanálisis de segunda generación con los encontrados con un procedimiento ELISA bien establecido, así como comprobar el valor diagnóstico de este nuevo marcador en los casos de artritis reumatoide presentes en nuestra población. MÉTODOS El número total de pacientes seleccionados para el estudio fue de 284: 105 pacientes con artritis reumatoide y los 179 restantes pertenecían a pacientes del grupo control, que incluyó donantes de sangre, pacientes sin historia reumatológica conocida y pacientes con enfermedad reumática no artritis reumatoide. Los anticuerpos anti-CCP de isotipo IgG fueron medidos mediante el nuevo método automatizado de fluoroenzimoinmunoanálisis de segunda generación (EliATM CCP) desarrollado por Pharmacia. Así mismo, la medida también se llevó a cabo con un método semicuantitativo de enzimoinmunoanálisis (QUANTA LiteTM CCP IgG ELISA. INOVA Diagnostics, Inc.). RESULTADOS Para el valor discriminante seleccionado mediante curva de rendimiento diagnóstico de 12 μg/l, la sensibilidad diagnóstica conseguida para los anticuerpos anti-péptido cíclico citrulinado fue del 74.3% y la especificidad diagnóstica fue del 94%. El área bajo la curva de rendimiento diagnóstico fue 0.845. La correlación entre los dos métodos empleados para la determinación de anticuerpos anti-CCP resultó ser buena (r = 0.936), no existiendo errores sistemáticos estadísticamente significativos entre ellos (p 0.05). CONCLUSIONES La determinación de anticuerpos anti-CCP tiene gran interés en el diagnóstico de la artritis reumatoide, interés que aumenta cuando para la medida de estos autoanticuerpos empleamos nuevos métodos totalmente automatizados como el analizado en este trabajo

OBJECTIVE The aim of the present study was to compare the results obtained with a new automated second generation method of fluoroenzymoimmunoanalysis with the results obtained with a well established ELISA procedure, as well as to study the diagnostic value of this new marker in the present cases of rheumatoid arthritis in our population. METHOD Patients selected for the study was 284: 105 patients with rheumatoid arthritis and the others 179 belonged to patients of the group control, that included blood donors, patients without well-known reumatologic history and patients with rheumatic disease (no rheumatoid arthritis). Anti-CCP antibodies were measured by the new automated method (EliA‘ CCP) developed by Pharmacia and, with a semiquantitative method of enzymoimmunoanalisys (QUANTA Lite‘ CCP IgG ELISA. INOVA Diagnostics, Inc.). RESULTS At a cut-off value of 12 μg/l, diagnostic sensitivity was 74,3% and diagnostic specificity was 94% for the Pharmacia procedure, being the area under the ROC curve of 0,845. The correlation between methods was good (r = 0,936), not existing statistically significant bias among them (p 0,05). CONCLUSIONS The determination of anti-CCP antibodies is usefull in the diagnosis of rheumatoid arthritis, and the procedure is improved with the use of a new automated fluoroimmunoassay

Rheumatoid factors induce signaling from B cells, leading to Epstein-Barr virus and B-cell activation.

B-cell antigen receptor signaling is initiated upon binding of the antigen to membrane-bound immunoblobulin (Ig), and the anti-Ig antibody (Ab) mimics this signaling. In B cells latently infected with Epstein-Barr virus (EBV), the same signals induce virus activation. We examine here whether rheumatoid factors (RFs), autoantibodies directed against the Fc portion of IgG, induce EBV and B-cell activation. As a source of RFs, RF-producing lymphoblastoid cell line (LCL) clones were isolated from peripheral blood mononuclear cells (PBMC) and synovial cells from patients with rheumatoid arthritis (RA) by EBV transformation. Burkitt's lymphoma-derived Akata cells, which are highly responsive to EBV activation by anti-Ig Abs, were used for the assay of EBV activation. Akata cells expressed IgG3 as membrane-bound Ig. RFs from a synovium-derived LCL were directed to IgG3 and induced EBV activation in 16 to 18% of Akata cells, whereas RFs from another synovium-derived LCL were directed to IgG1 and did not induce EBV activation. Pretreatment of RFs with the purified Fc fragment of human IgG completely abolished EBV activation. Furthermore, B-cell activation was assessed by incorporation of [3H]thymidine. RFs from synovium-derived LCLs efficiently induced B-cell activation, and the addition of CD40 ligand had a synergistic effect. On the other hand, RFs from PBMC-derived LCLs were polyreactive, had a lower affinity to IgG, and did not induce EBV and B-cell activation. The present findings imply a possible role for RFs as EBV and B-cell activators.

Interference by rheumatoid factor activity in the detection of antiavian antibodies in pigeon breeders disease.

The assessment of antiavian antibodies is relevant for the study of pigeon breeder's disease; nevertheless, different factors may hamper their accurate detection. The objective of this study was to determine whether an endogenous interfering effect in pigeon breeder's disease might explain the simultaneous presence of IgM, IgG, and IgA antiavian antibodies in high titers as assessed by ELISA. Fifty-nine patients with pigeon breeder's disease, 80 healthy controls, and 47 asymptomatic breeders were studied. To assess possible interfering effects by endogenous immunoglobulins, serum IgG was separated through protein A-Sepharose CL-4B chromatography. Antiavian antibodies were measured in whole and separated samples by ELISA. Since a decline of IgM antiavian antibodies following IgG removal was consistent with a false-positive effect, the causes were studied. High values of IgM, IgG, and IgA antiavian antibodies were found in 47.4% [corrected] of patients with pigeon breeder's disease. An IgM rheumatoid factor activity against IgG was found through ELISA in sera with false-positive IgM antiavian antibodies. Rheumatoid factor binding was confirmed by Western blot. Experimental addition of purified rheumatoid factor to sera with IgG antiavian antibodies replicated the interfering effect. A control group of rheumatoid arthritis with high rheumatoid factor values did not show positive antiavian antibodies tests. No IgG with anti-IgM or anti-IgA activity was found, and the detection of IgA against IgM and IgG was negative. In conclusion, the study of antiavian antibodies might be affected by different immunoassay conditions. An endogenous rheumatoid factor activity produced false-positive IgM results. Other similar interferences warrant a careful evaluation during the serological assessment of pigeon breeder's disease.