Anti-citrullinated Protein Antibodies Levels Research Articles

Unexpected finding of anticitrullinated protein antibodies in cerebrospinal fluid of RA patients with intact blood brain barrier

BackgroundThe authors have recently shown upregulation of inflammatory mediators in cerebrospinal fluid (CSF) from patients with rheumatoid arthritis (RA), with correlations to fatigue mechanisms. The authors have now sought to...

Atacicept in patients with rheumatoid arthritis and an inadequate response to tumor necrosis factor antagonist therapy: Results of a phase II, randomized, placebo-controlled, dose-finding trial

To assess the efficacy, safety, and biologic activity of atacicept in patients with rheumatoid arthritis (RA) in whom the response to treatment with tumor necrosis factor antagonists was inadequate. The Atacicept for Reduction of Signs and Symptoms in Rheumatoid Arthritis Trial (AUGUST I) was a multicenter, phase II, double-blind, placebo-controlled dose-finding study involving 256 patients randomized 1:1:1:1 to receive atacicept (25 mg, 75 mg, or 150 mg) or placebo twice weekly for 4 weeks, then weekly for 21 weeks, with a 13-week treatment-free followup period (week 38). The primary end point was a response at week 26 according to the American College of Rheumatology criteria for 20% improvement in disease severity, using the C-reactive protein level. No statistically significant differences were observed in the efficacy end points at week 26 (P = 0.410 for overall treatment effect). However, atacicept significantly reduced immunoglobulin and rheumatoid factor (RF) levels, but not anti-citrullinated protein antibody levels, in a dose-dependent manner, with levels returning toward baseline values during followup. The effects of treatment on IgG-RF and IgA-RF were more pronounced than the effects on total IgG and IgA. Adverse events (AEs), including serious AEs, leading to withdrawal were more common among patients treated with atacicept compared with placebo. AEs were variable in nature, and no dose-dependent trends were observed. The frequency of infection-related AEs was similar across treatments. No notable effect of treatment on immunization status (protective versus nonprotective titer) was observed after initiation of treatment. This study did not meet the primary efficacy end point. However, clear biologic activity consistent with the proposed mechanism of action was observed. The results suggest that decreasing the expression of RF may not be sufficient to induce clinical improvement in RA. The safety of atacicept was considered acceptable in this patient population.

Specific expression of PAD4 and citrullinated proteins in lung cancer is not associated with anti-CCP antibody production

Anti-citrullinated protein antibodies (ACPAs), produced against citrullinated proteins, are diagnostic and prognostic markers of rheumatoid arthritis (RA). The underlying mechanism that explains the connection of smoking, citrullination [catalyzed by peptidyl arginine deiminases (PADs)] and ACPAs is still unclarified in RA. Thus, we searched for a non-arthritic model in which an increased cell death allows the formation of autoantibodies. Data supporting that lung cancer might be a good candidate are as follows: (i) smoking plays a role in its pathogenesis, (ii) the disease is frequently accompanied by paraneoplastic syndrome, (iii) smoking increases citrullination in the lung, (iv) various types of malignancies are associated with increased citrullination and (v) lung cancer tissue shows similarities with RA synovium. Serum PAD4, rheumatoid factor (RF) and ACPA levels were measured in 42 lung cancer patients; expression of cytokeratin 7 (CK7), PAD4 and citrullinated proteins was visualized in 113 lung cancer tissues. All parameters were analyzed in correlation with smoking history. None of the patients had polyarthritis or autoimmune disease. Significantly increased RF levels were associated with higher PAD4 levels in smoker lung cancer patients compared with non-smokers. Both PAD4 and citrullination immunostaining strongly correlated with that of CK7 in lung cancer, however, did not differ according to smoking history. Two of 30 smoker lung cancer patients had high anti-cyclic citrullinated peptide levels. In conclusion, PAD4 and citrullination may be helpful in distinguishing lung cancer from healthy tissue. Smoking, abnormal serum PAD4 and RF levels may not be sufficient for the production of ACPAs and development of autoimmunity.

Toward a data‐driven evaluation of the 2010 American College of Rheumatology/European League Against Rheumatism criteria for rheumatoid arthritis: Is it sensible to look at levels of rheumatoid factor?

Recently, new classification criteria for rheumatoid arthritis (RA) have been devised by methodology that used first a quantitative approach (data from databases), then a qualitative approach (consensus; based on paper patients), and finally a common sense-based approach (evaluation of the former phases). Now the individual items that make up these criteria are being evaluated. This study was undertaken to analyze the item "autoantibodies," in particular rheumatoid factor (RF) level. Three separate cohorts comprising a total of 972 patients with undifferentiated arthritis were studied for RA development (according to the 1987 American College of Rheumatology criteria) and arthritis persistence. The positive predictive value (PPV), negative predictive value (NPV), and likelihood ratios (LRs) were compared between different levels of RF and the presence of anti-citrullinated protein antibody (ACPA). A similar comparison was made in 686 RA patients for the rate of joint destruction and achievement of sustained disease-modifying antirheumatic drug-free remission during 7 years of followup. The variation in RF levels obtained by different measurement methods in the same RF-positive sera was explored. Compared to high RF levels, presence of ACPA had a better balance between positive LR and negative LR and between PPV and NPV for RA development. The additive value of ACPA assessment after testing for RF level was higher than vice versa. The association between high RF level and RA severity was not as strong as that between ACPA antibodies and RA severity. The RF level obtained by different methods in the same patients' sera varied considerably. Our findings indicate that determination of RF level is subject to large variation; high RF level has limited additive prognostic value compared to ACPA positivity. Thus, omitting RF level and using RF presence, ACPA presence, and ACPA level may improve the 2010 criteria for RA.

Pathogenesis of rheumatoid arthritis in a fibrinogen-induced mouse model of autoimmunity (101.8)

Abstract Citrullinated proteins, derived from the conversion of peptidyl-arginine to peptidyl-citrulline, are present in the joints of rheumatoid arthritis (RA) patients who also produce high levels of anti-citrullinated protein antibodies (ACPAs). T cells also have a role in RA and are thought to be involved in initiating, controlling and driving antigen-specific immune responses. We and others found that mice immunized with human fibrinogen (HF) or human citrullinated fibrinogen (HCF) develop mild inflammatory arthritis and peripheral T cells proliferate in response to HCF and HF. To determine if HCF-reactive T cells contribute to disease, DBA1/J mice were immunized with HCF and at various time points, T cells were assessed directly ex vivo or cultured in vitro. T cells were evaluated ex vivo for the presence of activation markers and cytokine production after antigen stimulation; T cell lines were cultured in vitro with antigen and IL-2 and evaluated in the same manner. Following two immunizations, mice produced CD4 T cells with an activated phenotype (CD44hi+CD62L-); TNFα and IL-17 were produced by CD4 T cells in response to HCF. T cell lines produced high levels of TNFα, IL-17, and IL-6 when stimulated by HCF, and one line also produced IFNγ. In sum, CD4 T cells are activated during disease progression and exhibit an inflammatory phenotype that may be pathogenic. T cell lines are being evaluated in vivo to determine if they can precipitate or accelerate disease.

Increased serum PAD4 and RF in lung cancer is not associated with anti CCP antibody production

BackgroundRheumatoid arthritis (RA) is characterised by anticitrullinated protein antibodies (ACPAs), produced against citrullinated proteins. In the pathogenesis of the disease the interaction of environment and genetics have a role as...

Levels of anti-citrullinated protein antibodies and IgM rheumatoid factor are not associated with outcome in early arthritis patients: a cohort study

IntroductionTo investigate whether baseline levels of anti-citrullinated protein antibody (ACPA) or IgM rheumatoid factor (IgM-RF) and changes in the year thereafter are associated with disease activity, functional and radiographic outcome in early arthritis patients, and provide additional information over baseline autoantibody status.MethodsIn 545 early arthritis patients ACPA and IgM-RF levels, disease activity (DAS28), the Health Assessment Questionnaire (HAQ) and Sharp/Van der Heijde Score (SHS) were assessed annually. Baseline status, levels and first-year changes of the autoantibodies were associated with these measures at the two-year follow-up and sub-analysed according to autoantibody status.ResultsThe mean age was 52.7 years, 69% was female, at baseline 56% was ACPA positive, 47% IgM-RF positive. At the two-year follow-up the mean DAS28 was 2.88, and the median HAQ and SHS were 0.38 and 1, respectively. At one year, ACPA and IgM-RF levels had decreased by 31% and 56%, respectively. A switch from negative to positive occurred in 2% for ACPA and 3% for IgM-RF. Positive ACPA and RF status were both associated with SHS at two years (P < 0.001), but baseline levels only showed a minor correlation of ACPA with DAS28 and HAQ at two years. Level changes were not associated with the outcome parameters.ConclusionsBaseline levels and first-year changes of ACPA and IgM-RF are hardly associated with outcome after two years. Seroconversion seldom occurs. Therefore, it does not appear useful to repeat ACPA or IgM-RF measurements.

Antibodies to several citrullinated antigens are enriched in the joints of rheumatoid arthritis patients

High titers of specific anti-citrullinated protein antibodies (ACPAs) are frequently present in the serum of rheumatoid arthritis (RA) patients, but their presence in synovial fluid is less well characterized. The purpose of this study was to compare the levels of antibody to 4 well-defined citrullinated candidate RA autoantigens in serum and synovial fluid and to determine whether antibodies to one citrullinated antigen are dominant over another. Furthermore, we studied their relationships with mutated citrullinated vimentin (MCV), a newly identified RA-specific serum assay, and the classic cyclic citrullinated peptide (CCP) in the synovial fluid of well-defined HLA-DR groups. Paired serum and synovial fluid samples from 290 RA patients and serum samples from 100 age- and sex-matched healthy controls were analyzed for the presence of anti-MCV and anti-CCP antibodies and for reactivity to citrullinated fibrinogen, alpha-enolase, type II collagen, and vimentin. A total of 219 of the 290 patients were genotyped for the HLA-DR shared epitope alleles. Significantly higher proportions of antibodies against all RA-associated citrullinated antigens were found in synovial fluid as compared with serum. This was also true for the MCV and CCP responses but not for non-RA-associated anti-tetanus toxoid antibodies. As expected, we found a high correlation between citrullinated vimentin and MCV responses. All synovial fluid ACPAs were predominantly associated with HLA-DRB1*04 alleles and were confined to the CCP+/MCV+ subset of patients. MCV and CCP positivity represent a similar subset of RA patients, whereas ACPAs with different fine specificities fall into subgroups of anti-CCP+/anti-MCV+ patients. The levels of all specific ACPAs were elevated in synovial fluid, suggesting that there is local antibody production and/or retention of ACPAs at the site of inflammation governed by RA-predisposing genes.

Development of inflammatory arthritis in DR4 transgenic and wild type DBA/1J mice immunized with human fibrinogen (49.9)

Abstract The human MHC allele DR4 is associated with susceptibility for rheumatoid arthritis (RA). Citrullinated proteins, derived from the conversion of peptidyl-arginine to peptidyl-citrulline, are present in the joints of RA patients, who also uniquely produce high levels of anti-citrullinated protein antibodies (ACPAs). ACPAs enhance damage in animal models of autoimmune arthritis. DR4 molecules have been shown to preferentially bind citrullinated peptides over arginine controls. Recently, the DR4 allele was shown to be essential to develop arthritis in C57Bl/6 mice immunized with human citrullinated fibrinogen, but these mice demonstrated delayed arthritis at relatively low incidence and severity as compared to other arthritis models. To potentially obtain a more susceptible strain, we backcrossed the DR4 gene onto the DBA1/J strain. Following immunization with either human citrullinated or noncitrullinated fibrinogen, these mice developed inflammatory arthritis around day 24 regardless of whether they were DR4+ or DR4- littermates. Serum from both strains demonstrated ACPA generation as defined by reactivity to a citrullinated but not control peptide, as well as a splenic proliferative response to both citrullinated and noncitrullinated human, but not mouse, fibrinogen. The DR4 DBA/1J transgenic strain may provide a useful tool for the analysis of ACPA-related arthritis. Grant U19 AI146374

Plasma asymmetric dimethylarginine is related to anticitrullinated protein antibodies in rheumatoid arthritis of short duration

We have recently demonstrated elevated plasma levels of an endogenous nitric oxide synthase inhibitor, asymmetric dimethyl- l-arginine (ADMA), and its association with carotid atherosclerosis in rheumatoid arthritis (RA). Both an elevated risk of myocardial infarction and increased levels of anticitrullinated protein antibodies (ACPAs), specific for RA, had been shown to precede the onset of clinical RA symptoms. Therefore, our aim was to verify the hypothesis that ADMA accumulation might accompany raised ACPAs titers in RA of short duration (≤3 years). Twenty patients (16 women, 4 men; mean age, 45 ± 12 years; mean disease duration, 2.3 ± 0.5 years) with active RA despite chronic disease-modifying antirheumatic medication, free of cardiovascular disease or atherosclerotic risk factors, were studied. Plasma levels of ADMA and its stereoisomer, symmetric dimethyl- l-arginine (SDMA), were assayed by liquid chromatography/tandem mass spectrometry. The ACPAs were measured by a second-generation enzyme-linked immunosorbent assay. In addition to routine biochemical assays, plasma concentrations of tumor necrosis factor α, monocyte chemoattractant protein–1, and vascular cell adhesion molecule–1 soluble form were analyzed with respective enzyme-linked immunosorbent assays. A significant positive correlation between levels of ACPAs and ADMA ( r = .60, P = .005), but not SDMA ( r = −.02, P = .9), was found. Neither ADMA nor SDMA was correlated to any of the clinical or biochemical parameters reflecting disease activity and inflammatory activation. Thus, excessive ADMA accumulation accompanies elevated ACPAs levels in patients with RA of short duration free of cardiovascular disease or risk factors.

Marked differences in fine specificity and isotype usage of the anti–citrullinated protein antibody in health and disease

Anti-citrullinated protein antibodies (ACPAs) display high association with rheumatoid arthritis (RA) and are implicated in its pathogenesis. The presence of ACPAs is known to precede the onset of RA. In order to identify the features that could confer its pathogenicity, we extensively characterized this antibody response in a unique North American native population of patients with RA and their unaffected relatives. The levels of IgA, IgM, and IgG ACPAs, as well as IgM and IgA rheumatoid factor (RF), were measured in serum samples obtained from 81 patients with RA and 195 of their unaffected relatives. The isotype distribution, the fine specificity of the ACPA response, and its association with RF were compared in health and disease. ACPA positivity was observed in 19% of the healthy relatives and approximately 91% of the patients with RA. ACPA isotype usage was strikingly lower in unaffected relatives than in patients with RA (1-2 versus 5-6 isotypes). Fine specificity studies showed that reactivity to citrullinated fibrinogen and vimentin was present in sera from patients with RA, while it was virtually absent in their unaffected relatives. Finally, the ACPA and RF responses were associated in patients with RA but were discordant in their healthy relatives. Extended analyses revealed that the presence of ACPAs was associated with RA irrespective of RF status, while the association of RF with disease relied on its interaction with ACPAs. The fine specificity and isotype usage of the ACPA response are qualitatively different in health and disease. Epitope spreading and expansion of the isotype repertoire might be necessary for development of RA, and this could be facilitated by the presence of RF antibodies.

Preferential decrease in IgG4 anti-citrullinated protein antibodies during treatment with tumour necrosis factor blocking agents in patients with rheumatoid arthritis

Objective:To investigate the dynamics of IgG1 and IgG4 anti-citrullinated protein antibody (ACPA) subclasses during anti-tumour necrosis factor (TNF) treatment in patients with rheumatoid arthritis (RA).Methods:IgG, IgG1 and IgG4 ACPA levels...

An unexpectedly high frequency of MEFV mutations in patients with anti–citrullinated protein antibody–negative palindromic rheumatism

To investigate whether the MEFV gene, which is involved in the regulation of the inflammatory response and has been associated with familial Mediterranean fever (FMF) and intermittent hydrarthrosis, is implicated in the pathogenesis of palindromic rheumatism (PR) and to examine its clinical presentation and its evolution in a Spanish cohort of PR patients. Family histories, demographic clinical data, and laboratory characteristics of 75 patients diagnosed as having PR were collected from medical records and personal interviews. The healthy control group included 325 blood bank donors. The FMF control group was made up of 84 Spanish FMF patients. Genomic DNA was isolated, and MEFV gene mutation analysis was performed by polymerase chain reaction amplification and sequence analysis. Sixty-five unrelated PR patients were finally included in the study. MEFV gene mutation analysis identified 8 of these 65 patients (12.3%) as carriers of at least 1 mutated MEFV allele. Patients with MEFV mutations had higher mean age and age at disease onset, but lower mean serum levels of anti-citrullinated protein antibodies (ACPAs). No other significant differences were observed between patients with and those without mutations. The frequency of MEFV mutations in ACPA-negative PR patients was 22.2%, compared with 5.3% in ACPA-positive PR patients (P = 0.058). This study shows a previously unreported high prevalence of mutations of the MEFV gene in patients with ACPA-negative PR. This supports the hypothesis that it might be a susceptibility gene. Our findings also support the hypothesis that the MEFV gene might participate in the pathogenesis of other undifferentiated relapsing inflammatory rheumatic disorders.

Synovial intracellular citrullinated proteins colocalizing with peptidyl arginine deiminase as pathophysiologically relevant antigenic determinants of rheumatoid arthritis–specific humoral autoimmunity

To address the ongoing debate concerning the specificity of synovial citrullinated proteins for rheumatoid arthritis (RA) and to analyze their pathophysiologic relevance to the induction or perpetuation of the RA-specific anti-citrullinated protein antibodies (ACPAs). Synovium of 19 RA patients and 19 non-RA controls was immunostained for the presence of citrullinated proteins with a mouse monoclonal antibody (F95), for the citrullinating enzyme peptidyl arginine deiminase type 2 (PAD-2), and for the free citrulline-producing enzyme inducible nitric oxide synthase (iNOS). Extending the RA cohort to 61 patients, the findings of anticitrulline staining in synovium were related to serum and synovial fluid ACPA levels, as measured by enzyme-linked immunosorbent assay. F95 staining indicated the presence of synovial intracellular citrullinated proteins in 53% of RA samples versus 5% of control samples, whereas extracellular staining was not RA specific. Immunoblotting and inhibition experiments confirmed that the antibody recognized citrullinated proteins but not free citrulline. Accordingly, iNOS was equally found in RA and control synovium and in intracellular citrullinated protein-positive and intracellular citrullinated protein-negative samples. In contrast, intracellular citrullinated proteins colocalized with PAD-2, which was found in 59% of RA samples versus 17% of control samples. Independent of local disease activity, the presence of the RA-specific synovial intracellular citrullinated proteins was associated with significantly higher systemic and local ACPA levels and with local ACPA production in the joint. These data confirm the presence of RA-specific intracellular citrullinated proteins in synovium. The link with PAD-2 and local and systemic ACPA levels emphasizes their pathophysiologic relevance for RA-specific humoral autoimmunity.