Anti-citrullinated Protein Antibodies Antibodies Research Articles

Autoantibodies against citrullinated and native proteins and prediction of rheumatoid arthritis-associated interstitial lung disease: A nested case-control study.

To identify fine specificity anti-citrullinated protein antibodies (ACPA) associated with incident rheumatoid arthritis-associated interstitial lung disease (RA-ILD). This nested case-control study within the Brigham RA Sequential Study matched incident RA-ILD cases to RA-noILD controls on time of blood collection, age, sex, RA duration, and rheumatoid factor status. A multiplex assay measured ACPA and anti-native protein antibodies from stored serum prior to RA-ILD onset. Logistic regression models calculated odds ratios (OR) with 95% confidence intervals (CI) for RA-ILD, adjusting for prospectively-collected covariates. We estimated optimism-corrected area under the curves (AUC) using internal validation. Model coefficients generated a risk score for RA-ILD. We analyzed 84 incident RA-ILD cases (mean age 67 years, 77% female, 90% White) and 233 RA-noILD controls (mean age 66 years, 80% female, 94% White). We identified six fine specificity antibodies that were associated with RA-ILD. The antibody isotypes and targeted proteins were: IgA2 to citrullinated histone 4 (OR 0.08 per log-transformed unit, 95% CI 0.03-0.22), IgA2 to citrullinated histone 2A (OR 4.03, 95% CI 2.03-8.00), IgG to cyclic citrullinated filaggrin (OR 3.47, 95% CI 1.71-7.01), IgA2 to native cyclic histone 2A (OR 5.52, 95% CI 2.38-12.78), IgA2 to native histone 2A (OR 4.60, 95% CI 2.18-9.74), and IgG to native cyclic filaggrin (OR 2.53, 95% CI 1.47-4.34). These six antibodies predicted RA-ILD risk better than all clinical factors combined (optimism-corrected AUC=0·84 versus 0·73). We developed a risk score for RA-ILD combining these antibodies with the clinical factors (smoking, disease activity, glucocorticoid use, obesity). At 50% predicted RA-ILD probability, the risk scores both without (score=2·6) and with (score=5·9) biomarkers achieved specificity ≥93% for RA-ILD. Specific ACPA and anti-native protein antibodies improve RA-ILD prediction. These findings implicate synovial protein antibodies in the pathogenesis of RA-ILD and suggest clinical utility in predicting RA-ILD once validated in external studies. National Institutes of Health.

Rheumatoid arthritis-associated antibodies in healthy first-degree relatives of RA patients.

Rheumatoid arthritis (RA) affects approximately 1.5% of the population worldwide and 0.5-3.3% of the Mexican population. The presence of rheumatoid factor (RF), anti-citrullinated protein antibodies (ACPA) and anti-carbamylated protein (anti-CarP) antibodies has been described in populations at risk of RA development, such as first-degree relatives (FDR). Anti-CarP antibodies are present in RA patients (44%), FDR of RA patients (18%) and healthy controls (4.7%). Anti-CarP antibodies have not been described in FDR of the Mexican population. The objective of this study was to determine the prevalence of Rheumatoid Factors (RF) isotypes, ACPA and anti-CarP antibodies isotypes in FDR of RA patients.An observational, cross-sectional study, in an FDR of RA cohort, was performed. We measured IgA, IgG and IgM isotypes of RF, ACPA and anti-CarP antibodies. A total of 144 FDRs from 99 RA patients were enrolled. The prevalence of anti-CarP antibodies was 2.8% for IgA, 4.2% for IgG, whereas IgM was not detected. The serologic association was for RF/ACPA 4.48%, RF/anti-CarP 2.7%, FR 64.5%, ACPA 1.3%, ACPA/anti-CarP 0.69%, anti-CarP 3.4%, and no RF/ACPA/anti-CarP was observed. We found a low prevalence of anti-CarP antibodies in our cohort of FDR of RA patients, but the prevalence of ACPA and RF were higher than other cohorts previously reported.

AB0668 THE ONSET OF RHEUMATOID ARTHRITIS AFTER COVID-19 – COINCIDENCE OR CONNECTED?

Background:COVID-19, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has been suggested to induce autoimmune phenomena. Multiple studies have reported the presence of autoantibodies in patients with COVID-19. Also the presence of anti-citrullinated protein antibodies (ACPA) and flaring of rheumatoid arthritis (RA) after COVID-19 has been described.[1, 2] Furthermore, in rheumatology clinics patients may present with polyarthritis compatible with RA shortly after SARS-CoV-2 infection. However, it is unclear how often ACPA occur after COVID-19 and whether preceding COVID-19 impacts on disease presentation of RA and phenotype of the ACPA response.Objectives:This study aims to determine the seroprevalence of ACPA after COVID-19 and to investigate the association between preceding COVID-19 infection and disease presentation of new-onset RA, including clinical phenotype and autoantibody response.Methods:To estimate the prevalence of ACPA after COVID-19 we measured ACPA IgG in samples from 61 patients visiting the specialized post-COVID outpatient clinic of the LUMC 5 weeks after hospitalization, using routine tests or in-house ELISA. Furthermore, we identified 5 patients presenting with polyarthritis compatible with RA after SARS-CoV-2 infection. To study the impact of COVID-19 on disease presentation, we examined clinical phenotype, autoantibody isotype positivity and ACPA IgG variable domain (V-domain) glycosylation of these patients and compared these features to regular RA patients. Autoantibody isotypes, including rheumatoid factor (RF) IgM/IgA, anti-CCP2 IgG/ IgM/IgA and anti-carbamylated protein antibodies (anti-CarP) IgG were measured using in-house ELISA’s. The percentage of V-domain glycosylation of purified ACPA IgG was measured with UHPLC.Results:None of the 61 post-COVID patients tested positive for ACPA 5 weeks after hospitalization, except two patients previously diagnosed with ACPA-positive RA. Thus, we could not observe an increase in ACPA-positivity shortly after COVID-19. Of the 5 patients who developed polyarthritis compatible with RA after SARS-CoV-2 infection, the average age was 63.6 years and 2/5 were female. 4/5 patients had been hospitalized due to severe COVID-19. On average, joint complaints started 6.6 weeks after infection, although two patients reported symptoms before infection. 4/5 patients fulfilled the ACR 2010 criteria for RA. Three patients (patient 1, 4, 5) were phenotypically very similar to regular new-onset RA patients. Patient 3 had a history of seronegative RA and had been in DMARD-free remission for 5 years. She flared 6 weeks after SARS-CoV-2 infection. Patient 2 had a remarkably different presentation. He was admitted with a suspected septic polyarthritis or pneumonia with reactive polyarthritis 6 weeks after COVID-19. ACPA level was low positive. The patient died unexpectedly after two days and autopsy revealed dilating myocarditis of unclear underlying cause. No causative pathogen could be identified.Previous studies have shown that RA-patients are most often either seronegative or triple-positive for RF, ACPA and anti-CarP antibodies. Autoantibody measurements on sera of the post-COVID polyarthritis patients revealed a similar pattern (Figure 1A) with two patients being completely seronegative, and three patients positive for a range of autoantibodies. In all post-COVID samples, the percentage of ACPA IgG V-domain glycosylation was significantly increased compared to total IgG (Figure 1B), similar as in regular RA.Conclusion:In conclusion, we found that the seroprevalence of ACPA is not increased post-COVID and that most patients presenting with polyarthritis after COVID-19 resemble regular RA patients, both regarding clinical phenotype and autoantibody characteristics. Although sample size and follow-up was limited, it appears that RA post-COVID may be coincidence rather than connected.

Peptidyl Arginine Deiminase Type 4 Gene Promoter Hypo-Methylation in Rheumatoid Arthritis.

Protein citrullination is carried out by peptidylarginine deiminase type 4 (PAD4) enzyme. As a consequence of this process, post-translationally modified proteins are formed that become antigens for anti-citrullinated protein antibodies (ACPA). The study aimed at identifying whether the PADI4 gene is subject to epigenetic regulation through methylation of its promoter region, whether the degree of methylation differs in healthy individuals vs. rheumatoid arthritis (RA) patients and changes in correlation with ACPA, anti-PAD4 and disease activity. A total of 125 RA patients and 30 healthy controls were enrolled. Quantitative real-time methylation-specific PCR was used to analyze the methylation status. ACPA and anti-PAD4 antibodies were determined in serum by enzyme-linked immunosorbent immunoassay. The differences were observed in the degree of PADI4 gene promoter methylation between RA patients and HC, along with an upward trend for the methylation in RA, which was inversely proportional to the disease activity. A weak or modest negative correlation between the degree of PADI4 gene methylation and anti-PAD4, disease activity score (DAS28) and ACPA level has been found. The elevated methylation is associated with lower disease activity, lower levels of ACPA and aPAD4. The methylation degree in this area is growing up during effective treatment and might play a role in the RA pathophysiology and therefore could be a future therapeutic target.

The prevalence of ACPA is lower in rheumatoid arthritis patients with an older age of onset but the composition of the ACPA response appears identical

BackgroundRheumatoid arthritis (RA) consists of two syndromes, one autoantibody-positive and one autoantibody-negative. Existing data on the relation between age of onset and prevalence of autoantibodies were conflicting. Therefore this multicohort study assessed the age of onset in relation to the presence of autoantibodies. The association with characteristics of the anti-citrullinated protein antibodies (ACPA) response was also explored.MethodsThe 1987 criteria-positive RA patients included in the Leiden EAC, BARFOT, ESPOIR, Umeå and Lund cohorts (n = 3321) were studied at presentation for age of onset and the presence of ACPA, rheumatoid factor (RF) and anti-carbamylated protein (anti-CarP) antibodies. Logistic regression analyses were performed; effect sizes were summarized in inverse-weighted meta-analyses. Within ACPA-positive RA, ACPA level was studied in all cohorts; ACPA isotypes, ACPA fine specificity and ACPA avidity index and clinical characteristics were studied in the Leiden EAC.ResultsFrom the age of 50 onward, the proportion of ACPA-negative RA patients increased with age in the five cohorts. Similar observations were made for RF and anti-CarP. The composition of the ACPA response did not change with increasing age of onset with respect to titer, isotype distribution, fine specificity and avidity index. With increasing age of onset, RA patients smoked less often, had higher acute phase reactants and more often had a sub(acute) symptom onset.ConclusionsData of five cohorts revealed that with older age of onset ACPA-negative RA is more frequent than ACPA-positive RA, while characteristics of ACPA-positive RA as judged by the composition of the ACPA response appeared not age dependent. Further biologic studies are needed to characterize the pathogenesis of ACPA-negative polyarthritis at older age and to promote personalized treatment decisions in ACPA-negative patients in daily practice.

Smoking is associated with the concurrent presence of multiple autoantibodies in rheumatoid arthritis rather than with anti-citrullinated protein antibodies per se: a multicenter cohort study.

BackgroundThe contribution of smoking to rheumatoid arthritis (RA) is hypothesized to be mediated through formation of anti-citrullinated protein antibodies (ACPA). In RA, however, autoantibodies such as ACPA, rheumatoid factor (RF), and anti-carbamylated protein antibodies (anti-CarP) often occur together, and it is thus unclear whether smoking is specifically associated with some autoantibodies rather than others. We therefore investigated whether smoking is only associated with ACPA or with the presence of multiple RA-related autoantibodies.MethodsA population-based Japanese cohort (n = 9575) was used to investigate the association of smoking with RF and anti-cyclic citrullinated peptide antibodies (anti-CCP2) in individuals without RA. Furthermore, RA patients fulfilling the 1987 criteria from three early arthritis cohorts from the Netherlands (n = 678), the United Kingdom (n = 761), and Sweden (n = 795) were used. Data on smoking, RF, anti-CCP2, and anti-CarP were available. A total score of autoantibodies was calculated, and odds ratios (ORs) and 95% confidence intervals (95% CIs) were calculated by logistic regression.ResultsIn the population-based non-RA cohort, no association was found between smoking and one autoantibody (RF or anti-CCP2), but smoking was associated with double-autoantibody positivity (OR 2.95, 95% CI 1.32–6.58).In RA patients, there was no association between smoking and the presence of one autoantibody (OR 0.99, 95% CI 0.78–1.26), but smoking was associated with double-autoantibody positivity (OR 1.32, 95% CI 1.04–1.68) and triple-autoantibody positivity (OR 2.05, 95% CI 1.53–2.73).ConclusionsSmoking is associated with the concurrent presence of multiple RA-associated autoantibodies rather than just ACPA. This indicates that smoking is a risk factor for breaking tolerance to multiple autoantigens in RA.Electronic supplementary materialThe online version of this article (doi:10.1186/s13075-016-1177-9) contains supplementary material, which is available to authorized users.

MRI-detected osteitis is not associated with the presence or level of ACPA alone, but with the combined presence of ACPA and RF.

BackgroundIn rheumatoid arthritis (RA) bone marrow edema (BME, osteitis) and anti-citrullinated protein antibodies (ACPA) are associated with radiographic progression. ACPA have been associated with BME, but it is unknown if this association is confined to ACPA and BME. We performed cross-sectional analysis of the association of ACPA, rheumatoid factor (RF) and anti-carbamylated protein (anti-CarP) antibodies with BME and other types of inflammation (synovitis, tenosynovitis) detected by magnetic resonance imaging (MRI).MethodsDisease-modifying antirheumatic drug (DMARD)-naïve patients with early arthritis (n = 589), included in the Leiden Early Arthritis Clinic cohort, underwent contrast-enhanced 1.5 T MRI of unilateral wrist, metacarpophalangeal and metatarsophalangeal-joints at baseline. BME, synovitis and tenosynovitis were scored by two readers. ACPA, rheumatoid factor (RF) and anti-CarP were determined at baseline.ResultsIn univariable analyses ACPA-positive patients had higher BME scores than ACPA-negative patients (median 4.5 vs. 2.0, p < 0.001), but not more synovitis and tenosynovitis. Also RF (median 3.75 vs. 2.0, p < 0.001) and anti-CarP antibodies (median 3.5 vs. 2.5, p = 0.012) were associated with higher BME scores. Because the autoantibodies were concomitantly present, analyses were stratified for the presence of different autoantibody combinations. ACPA-positive (ACPA+), RF-negative (RF-), anti-CarP-negative (anti-CarP-) patients did not have higher BME-scores than ACPA-negative (ACPA-), RF-, anti-CarP- patients. However ACPA+, RF-positive (RF+), anti-CarP- patients and ACPA+, RF+, anti-CarP-positive (anti-CarP+) patients had higher BME scores than ACPA-, RF-, anti-CarP- patients (median 5.0 and 4.5 vs. 2.0, p < 0.001 and p < 0.001). ACPA levels were not associated with BME scores. Analyses within RA- and UA-patients revealed similar results.ConclusionsThe presence of ACPA alone or ACPA level was not statistically significantly associated with BME scores, but the combined presence of ACPA and RF was associated with more BME. This suggests an additive role of RF to ACPA in mediating osteitis.Electronic supplementary materialThe online version of this article (doi:10.1186/s13075-016-1076-0) contains supplementary material, which is available to authorized users.

Antibodies to carbamylated α-enolase epitopes in rheumatoid arthritis also bind citrullinated epitopes and are largely indistinct from anti-citrullinated protein antibodies.

BackgroundIn addition to anti-citrullinated protein antibodies (ACPAs), antibodies targeting carbamylated (i.e., homocitrullinated) proteins (anti-CarP antibodies) have been described in rheumatoid arthritis (RA). However, the extent to which anti-CarP antibodies are truly distinct from ACPA remains unclear, and few studies have focused on specific autoantigens. Here, we examine cross-reactivity between ACPA and anti-CarP antibodies, in the context of the candidate autoantigen α-enolase.MethodsCross-reactivity was examined by immunoblotting of citrullinated and carbamylated proteins using purified ACPA; and by peptide absorption experiments, using the citrullinated α-enolase peptide CEP-1 and a homocitrulline-containing version (carb-CEP-1) in ELISA. The population-based case-control cohort EIRA (n = 2836 RA; 373 controls) was screened for reactivity with CEP-1 and carb-CEP-1, using the ISAC multiplex array. Associations between anti-CarP antibodies, smoking and genetic risk factors were analysed using unconditional logistic regression models. Differences in antibody levels were investigated using the Mann-Whitney U test.ResultsAffinity-purified ACPA was found to bind carbamylated proteins and homocitrulline-containing peptides, demonstrating definitive cross-reactivity between ACPA and anti-CarP antibodies. Anti-carb-CEP-1 reactivity in EIRA was almost exclusively confined to the CEP-1-positive subset, and this group of RA patients (21 %) displayed a particularly strong ACPA response with marked epitope spreading. The small RA subset (3 %) with homocitrulline reactivity in the absence of citrulline reactivity did not associate with smoking or risk genes, and importantly had significantly lower anti-carb-CEP-1 antibody levels.ConclusionOur data presented herein cast doubt on the specificity of anti-CarP antibodies in RA, which we posit may be a subset of cross-reactive ACPA.Electronic supplementary materialThe online version of this article (doi:10.1186/s13075-016-1001-6) contains supplementary material, which is available to authorized users.

Anticarbamylated protein antibodies are associated with long-term disability and increased disease activity in patients with early inflammatory arthritis: results from the Norfolk Arthritis Register

ObjectivesAnticarbamylated protein (anti-CarP) antibodies are a novel family of autoantibodies recently identified in patients with inflammatory arthritis. The aim of this study was to investigate their association with long-term outcomes...

Genetic control of antibody production during collagen induced arthritis development in heterogeneous stock mice

ObjectivesThe genetic factors that drive pathogenic autoantibody formation in rheumatoid arthritis (RA) and its experimental models need to be elucidated in order to understand the aetiology of the disease and...