Flemish network on rare connective tissue diseases (CTD): patient pathways in systemic sclerosis. First steps taken.

Despite the low prevalence of each rare disease, the total burden is high. Patients with rare diseases encounter numerous barriers, including delayed diagnosis and limited access to high-quality treatments. In order to tackle these challenges, the European Commission launched the European Reference Networks (ERNs), cross-border networks of healthcare providers and patients representatives. In parallel, the aims and structure of these ERNs were translated at the federal and regional levels, resulting in the creation of the Flemish Network of Rare Diseases. In line with the mission of the ERNs and to ensure equal access to care, we describe as first patient pathways for systemic sclerosis (SSc), as a pilot model for other rare connective and musculoskeletal diseases. Consensus was reached on following key messages: 1. Patients with SSc should have multidisciplinary clinical and investigational evaluations in a tertiary reference expert centre at baseline, and subsequently every three to 5 years. Intermediately, a yearly clinical evaluation should be provided in the reference centre, whilst SSc technical evaluations are permissionably executed in a centre that follows SSc-specific clinical practice guidelines. In between, monitoring can take place in secondary care units, under the condition that qualitative examinations and care including interactive multidisciplinary consultations can be provided. 2. Patients with early diffuse cutaneous SSc, (progressive) interstitial lung disease and/or pulmonary arterial hypertension should undergo regular evaluations in specialised tertiary care reference institutions. 3. Monitoring of patients with progressive interstitial lung disease and/or pulmonary (arterial) hypertension will be done in agreement with experts of ERN LUNG.

The role of endothelial cells in the vasculopathy of systemic sclerosis: A systematic review.

INTRODUCTION: Systemic sclerosis (SSc) is an autoimmune connective tissue disorder characterized by fibroproliferative vasculopathy, immunological abnormalities and progressive fibrosis of multiple organs including the skin. In this study, all English speaking articles concerning the role of endothelial cells (ECs) in SSc vasculopathy and representing biomarkers are systematically reviewed and categorized according to endothelial cell (EC) (dys)function in SSc. METHODS: A sensitive search on behalf of the EULAR study group on microcirculation in Rheumatic Diseases was developed in Pubmed, The Cochrane Library and Web of Science to identify articles on SSc vasculopathy and the role of ECs using the following Mesh terms: (systemic sclerosis OR scleroderma) AND pathogenesis AND (endothelial cells OR marker). All selected papers were read and discussed by two independent reviewers. The selection process was based on title, abstract and full text level. Additionally, both reviewers further searched the reference lists of the articles selected for reading on full text level for supplementary papers. These additional articles went through the same selection process. RESULTS: In total 193 resulting articles were selected and the identified biomarkers were categorized according to description of EC (dys)function in SSc. The most representing and reliable biomarkers described by the selected articles were adhesion molecules for EC activation, anti-endothelial cell antibodies for EC apoptosis, vascular endothelial growth factor (VEGF), its receptor VEGFR-2 and endostatin for disturbed angiogenesis, endothelial progenitors cells for defective vasculogenesis, endothelin-1 for disturbed vascular tone control, Von Willebrand factor for coagulopathy and interleukin (IL)-33 for EC-immune system communication. Emerging, relatively new discovered biomarkers described in the selected articles, are VEGF165b, IL-17A and the adipocytokines. Finally, myofibroblasts involved in tissue fibrosis in SSc can derive from ECs or epithelial cells through a process known as endothelial-to-mesenchymal transition. CONCLUSION: This systematic review emphasizes the growing evidence that SSc is primarily a vascular disease where EC dysfunction is present and prominent in different aspects of cell survival (activation and apoptosis), angiogenesis and vasculogenesis and where disturbed interactions between ECs and various other cells contribute to SSc vasculopathy.

Lupus anticoagulant-hypoprothrombinemia syndrome: report of two cases and review of the literature.

Lupus anticoagulant-hypoprothrombinemia syndrome (LA-HPS) is a rare acquired disorder caused by prothrombin antibodies. The disease is most common in the pediatric age group (<16 years), and more prevalent in women. There are well-established clinical diseases associated with LA-HPS, most notably systemic lupus erythematosus (SLE) and viral infections. The clinical manifestation of LA-HPS varies greatly in severity and it may cause severe life-threatening bleeding diathesis. LA-HPS is to be suspected when a patient presents with bleeding and a prolonged activated partial thromboplastin and prothrombin time, in combination with a lupus anticoagulant. The diagnosis is confirmed in the laboratory by identification of reduced prothrombin levels. There are no standardized recommendations for treatment of the hemorrhage associated with the syndrome; corticosteroids are used as first-line treatment. This review summarizes what is currently known about the pathogenesis, clinical features, diagnosis, treatment and prognosis of LA-HPS, and presents two case reports.

Work participation in patients with systemic sclerosis: a systematic review.

OBJECTIVES: With this systematic review an overview is given of what is known about work participation in patients with systemic sclerosis (SSc). METHODS: The databases Pubmed, Cinahl, Nursing and Allied Health and PsychARTICLES have been checked from 1980 onwards. The search string consisted of all combinations of key words for work participation and SSc. Two investigators evaluated the eligibility for the articles. Reference lists were searched for other studies. RESULTS: Eight quantitative and one qualitative study were scrutinised in depth. The percentage of patients not working ranges from 18% to 61%. A meta-analysis of the percentage patients not working was performed and a weight mean of 37% was found. The following parameters are associated with the work variable in multivariate analysis (number of studies in which the variable was independently associated with the work variable/number of studies in which the variable was multivariately assessed): global disability (4/5), health (3/5), educational level (2/4), disease duration (3/3), skin/lung involvement (1/3), age/fatigue/muscle involvement/hand function (1/2) and having a decreased income/race/social support/physically demanding job (1/1). In the qualitative study, management of the work situation, disclosure of limitations at the work force and adaptation of resources in daily life are discussed. CONCLUSIONS: Most studies concerning work participation are at this very moment quantitative and cross-sectionally designed. Longitudinal studies are needed to assess causality and qualitative research may be opportune to have a more comprehensive view on the topic of work participation in patients with SSc.

Tuberculin skin test versus interferon-gamma release assays for the diagnosis of tuberculosis infection.

OBJECTIVE: Accurate detection of latent tuberculosis infection (LTBI) is becoming increasingly important due to the increasing use of immunosuppressive medications and the human immunodeficiency epidemic, which have increased the risk for reactivation to active tuberculosis (TB) infection. LTBI is detected by tuberculin skin test (TST) and interferon-gamma release assays (IGRAs). The latter include T-SPOT(®).TB (Oxford Immunotec) and QuantiFERON(®)-TB Gold In-Tube (QFT-GIT; Cellestis). We examined the value of TST versus IGRAs in the diagnosis of TB infection by meta-analysis based on data derived from a systematic literature review. METHODS: PubMed was searched for articles in English published between January 2010 and July 2012 in which TST and IGRA were performed simultaneously in individuals with and without active TB infection. A random effect model meta-analysis was performed to determine pooled sensitivity and specificity values for TST, T-SPOT.TB, and QFT-GIT. Owing to the absence of a gold standard for the diagnosis of LTBI, active TB infection was used as a surrogate for LTBI. RESULTS: Nineteen studies were included. T-SPOT.TB was significantly more sensitive [90% (95% confidence interval: 85-95) versus 64% (46-81)] than TST. The specificity of T-SPOT.TB was higher than the specificity of TST, but there was overlap between confidence intervals [77% (68-85) versus 57% (41-72)]. QFT-GIT seemed to be more sensitive than TST [75% (61-86) versus 64% (48-78)] but similarly specific [71% (62-86) versus 70% (57-81)]. CONCLUSIONS: IGRAs, especially T-SPOT.TB, are more effective at detecting TB infection than TST. Despite their higher cost, they have added value and can be requested in addition to TST.

Optimization and diagnostic performance of a single multiparameter lineblot in the serological workup of systemic sclerosis.

INTRODUCTION: Detection of systemic sclerosis-associated antibodies (SSc-Ab) in routine clinical practice is mostly restricted to anti-centromere and anti-topoisomerase-I antibodies. However, also other SSc-Ab (e.g. anti-RNA-polymerase-III, anti-PM/Scl, anti-fibrillarin and anti-Th/To) have been shown to be valuable diagnostic and prognostic markers for the disease, but testing methodologies for their detection are laborious and time-consuming. This study aimed to optimize interpretational criteria of a multiparameter lineblot (LB) for the parallel detection of SSc-Ab. We also assessed its global diagnostic value as an alternative for combined conventional techniques (CCT) in the serological workup of systemic sclerosis (SSc) patients. METHODS: The presence of SSc-Ab (anti-centromere, anti-topoisomerase-I, anti-RNA-polymerase-III, anti-PM/Scl, anti-fibrillarin and anti-Th/To) was identified by LB on 145 consecutive SSc patients and on 277 disease controls. Diagnostic sensitivity and specificity were calculated for both individual reactivities and the global LB. Cohen's kappa coefficient was used to examine agreement between LB and CCT and guided the definition of final interpretational criteria for LB. RESULTS: Applying the optimal cut-off values and interpretational criteria, LB identified SSc-Ab in 110 SSc patients (sensitivity=76%) and in 19 disease controls (specificity=93%). Globally, there was a substantial agreement between CCT and LB (κ=0.787, concordance 92.4%). LB and CCT showed a very good correlation (κ>0.800) for most SSc-Ab (anti-centromere, anti-topoisomerase-I, anti-RNA-polymerase-III and anti-PM/Scl). The best agreement for anti-RNA-polymerase-III and anti-PM/Scl was achieved when positivity for both components was taken as a criterion. CONCLUSIONS: LB is a reliable alternative for the laborious and time-consuming conventional techniques in the diagnostic workup of SSc, especially for the detection of anti-centromere, anti-topoisomerase-I, anti-RNA-polymerase-III and anti-PM/Scl.

The autoimmune disease-associated IL2RA locus is involved in the clinical manifestations of systemic sclerosis.

Regulatory T cells (T(regs)) are crucial in the maintenance of the immune tolerance and seem to have an important role in systemic sclerosis (SSc). The interleukin 2 receptor α (IL2RA) is an important T(reg) marker, and polymorphisms of IL2RA gene are associated with a number of autoimmune diseases. Therefore, we aimed to investigate for the first time the association of the IL2RA locus in SSc. For this purpose, a total of 3023 SSc patients and 2735 matched healthy controls, from six European Caucasian cohorts, were genotyped for the IL2RA gene variants rs11594656, rs2104286 and rs12722495 using the TaqMan allelic discrimination technology. The overall meta-analysis reached statistical significance when the three polymorphisms were tested for association with SSc, the limited subtype (lcSSc) and anti-centromere auto-antibodies (ACAs). However, no significant P-values were obtained when the ACA-positive patients were removed from the SSc and lcSSc groups, suggesting that these associations rely on ACA positivity. The strongest association signal with ACA production was detected for rs2104286 (P(FDR)=2.07 × 10(-4), odds ratio=1.30 (1.14-1.47)). The associations of rs11594656 and rs12722495 were lost after conditioning to rs2104286, and allelic combination tests did not evidence a combined effect, indicating that rs2104286 best described the association between IL2RA and ACA presence in SSc.

Microscopic polyangiitis involving the breast.

We report a case of a 76-year-old woman, presenting with a persistent dry cough, subfebrility, arthralgia and myalgia, weight loss and a breast lesion. She has elevated inflammatory parameters, impaired renal function with proteinuria, bilateral lung nodules on computed tomography scan (CT scan) and a suspect lesion on mammography. A diagnosis of microscopic polyangiitis with involvement of the breast is made based on clinical and radiographic findings, with positive auto-immune serology and histological confirmation. Although vasculitis of the breast is uncommon, this case illustrates that when a breast lesion is found, in combination with constitutional symptoms, we should think about the possibility of an anti-Neutrophil cytoplasmic antibodies (ANCA)-associated vasculitis. Therefore it may be important to perform auto-immune serology in these cases before proceeding to major surgery.

Analysis of the influence of PTPN22 gene polymorphisms in systemic sclerosis.

OBJECTIVE: Two functional single nucleotide polymorphisms (SNP) in the PTPN22 gene (rs24746601 and rs33996649) have been associated with autoimmunity. The aim of this study was to investigate the role of the R263Q SNP for the first time and to re-evaluate the role of the R620W SNP in the genetic predisposition to systemic sclerosis (SSc) susceptibility and clinical phenotypes. METHODS: 3422 SSc patients (2020 with limited cutaneous SSc and 1208 with diffuse cutaneous SSc) and 3638 healthy controls of Caucasian ancestry from an initial case--control set of Spain and seven additional independent replication cohorts were included in our study. Both rs33996649 and rs2476601 PTPN22 polymorphisms were genotyped by TaqMan allelic discrimination assay. A meta-analysis was performed to test the overall effect of these PTPN22 polymorphisms in SSc. RESULTS: The meta-analysis revealed evidence of association of the rs2476601 T allele with SSc susceptibility (p(FDRcorrected)=0.03 pooled, OR 1.15, 95% CI 1.03 to 1.28). In addition, the rs2476601 T allele was significantly associated with anticentromere-positive status (p(FDRcorrected)=0.02 pooled, OR 1.22, 95% CI 1.05 to 1.42). Although the rs33996649 A allele was significantly associated with SSc in the Spanish population (p(FDRcorrected)=0.04, OR 0.58, 95% CI 0.36 to 0.92), this association was not confirmed in the meta-analysis (p=0.36 pooled, OR 0.89, 95% CI 0.72 to 1.1). CONCLUSION: The study suggests that the PTPN22 R620W polymorphism influences SSc genetic susceptibility but the novel R263Q genetic variant does not. These data strengthen evidence that the R620W mutation is a common risk factor in autoimmune diseases.

A red face in a lupus patient: thinking beyond lupus rash.

A 61-year-old woman with a history of systemic lupus erythematosus presented with a facial rash followed by fever. The diagnosis of lupus flare was made by the general practioner who prescribed corticosteroids. The evolution was unfavorable and the patient was hospitalized. On admission, she presented with high fever and a sharply demarcated papular erythema overlying her cheeks. Biology was inflammatory with elevated C-reactive protein, sedimentation rate and leucocyte count. The diagnosis of erysipelas of the face was made on the basis of the malar and non pruritic papular erythema, fever, the high C-reactive protein levels and the absence of response to corticosteroids. The evolution was favorable under intravenous antibiotics and rapid tapering of the corticosteroids. To our knowledge, this is the first report of facial erysipelas in a patient with a history of systemic lupus erythematosus. This coincidence is a rare condition which may lead to erroneous diagnosis and inappropriate therapy.

Rituximab in diffuse cutaneous systemic sclerosis: an open-label clinical and histopathological study.

OBJECTIVES: The safety and potential efficacy of rituximab was examined in diffuse cutaneous systemic sclerosis (dc-SSc). METHODS: A 24 week open-label study in which eight patients with dc-SSc received an infusion of 1000 mg rituximab administered at baseline and day 15, together with 100 mg methylprednisolone at each infusion. Assessment included CD19+ peripheral blood lymphocyte number, skin sclerosis score, indices of internal organ functioning, the health assessment questionnaire disability index, the 36-item Short Form health survey and histopathological evaluation of the skin. RESULTS: Ritixumab induced effective B-cell depletion in all patients (<5 CD19+ cells/microl blood). There was a significant change in skin score at week 24 (p<0.001). Also, significant improvements were measured in the dermal hyalinised collagen content (p = 0.014) and dermal myofibroblast numbers (p = 0.011). Two serious adverse events occurred, which were thought to be unrelated to the rituximab treatment. CONCLUSIONS: Rituximab appears to be well tolerated and may have potential efficacy for skin disease in dc-SSc.

Classification models for disease: the effect of associations between markers on calculating the risk for disease by likelihood ratio products.

OBJECTIVES: The risk for disease or a bad prognosis can be calculated by means of prediction or classification models that take into account multiple variables. Different methods exist to construct such models. Some of those methods, including the likelihood ratio (LR) product method neglect dependency between variables. We aimed to evaluate the effect of neglecting dependency between variables in prediction or classification models. PATIENTS AND METHODS: Population I consisted of 1003 consecutive patients with a new diagnostic problem for which RA was included in the differential diagnosis and final diagnoses (RA or non-RA) were established after 1 year. The baseline variables included in the model are rheumatoid factor, anti-citrullinated protein/peptide antibodies and the HLA-shared epitope. Population II consisted of 847 patients with definite ankylosing spondylitis (AS). Six variables (psoriasis, inflammatory bowel disease, uveitis, HLA-B27 status and latest available CRP) were evaluated. Here, specificities of the features were derived from literature and different scenarios of association between variables in controls and diseased are estimated. RESULTS: When two features are similarly associated in cases and controls, risks for disease will be overestimated by neglecting dependency between variables. In the presented datasets, this resulted in a up to 12% overestimation of the risk. CONCLUSIONS: We showed how the height of over- or underestimation of risks can be evaluated when dependencies between two variables are neglected. This is important to evaluate the predictive value of combinations of features in cases where no data are available on associations in controls.

Reversible changes in serum immunoglobulin galactosylation during the immune response and treatment of inflammatory autoimmune arthritis.

OBJECTIVES: Improved DNA sequencer-aided fluorophore-assisted carbohydrate electrophoresis (DSA-FACE) technology was used to monitor the changes in the galactosylation status of serum immunoglobulins during the immune response and therapy of autoimmune arthritis. METHODS: Collagen-induced arthritis (CIA) was induced in susceptible DBA/1 mice and the undergalactosylation status (UGS) of serum immunoglobulins was determined using the improved DSA-FACE technology. Prophylactic intravenous tolerisation with type II collagen as well as semitherapeutic treatment with dexamethasone (DEX) were performed and UGS was analysed. Next, the serum immunoglobulin glycosylation profiles of patients with rheumatoid arthritis (RA) and spondyloarthropathy (SpA) were studied and changes in the UGS scores during anti-tumour necrosis factor (TNF)alpha therapy followed. RESULTS: In the longitudinal CIA study, the undergalactosylation state of immunoglobulins was found to be significantly correlated with the clinical arthritis scores. Upon collagen-specific tolerisation as well as glucocorticoid semitherapeutic treatment, improvement of the clinical arthritis scores correlated with decreased levels of UGS. It was also demonstrated that withdrawal of DEX was associated with an increased UGS score. Interestingly, reversibility in the UGS was also shown during treatment of patients with RA and SpA with anti-TNFalpha. CONCLUSIONS: These findings demonstrate that the UGS of serum immunoglobulins changes during the disease course of CIA and that this UGS is inhibited by antigen-specific and antigen-independent treatment procedures. The observation that Ig galactosylation is a reversible process is also documented during treatment of patients with RA and SpA with anti-TNFalpha.

Juvenile-onset systemic lupus erythematosus: different clinical and serological pattern than adult-onset systemic lupus erythematosus.

OBJECTIVE: To investigate differences in clinical signs and symptoms, and in antinuclear antibodies (ANA), between patients with juvenile-onset and adult-onset systemic lupus erythematosus (SLE). METHODS: Clinical and serological data of 56 patients with juvenile-onset SLE were compared with data of 194 patients with adult-onset SLE. ANA were determined by line immunoassay and by indirect immunofluorescence on Crithidia luciliae. RESULTS: Renal involvement, encephalopathy and haemolytic anaemia were seen, and anti-dsDNA, anti-ribosomal P and antihistone antibodies found, significantly more often in juvenile-onset SLE. Anti-dsDNA antibodies were directly associated, and anti-ribosomal P antibodies inversely associated, with renal involvement in juvenile-onset SLE. In juvenile patients with SLE and anti-dsDNA and without anti-ribosomal P antibodies the odds ratio for glomerulonephritis was 9.00; no patients with anti-ribosomal P but without anti-dsDNA had renal involvement. CONCLUSION: Patients with juvenile-onset SLE more often have renal involvement and encephalopathy than patients with adult-onset SLE. Anti-ribosomal P, anti-dsDNA and antihistone antibodies are more often found in patients with juvenile-onset SLE.

Melanoma inhibitory activity, a biomarker related to chondrocyte anabolism, is reversibly suppressed by proinflammatory cytokines in rheumatoid arthritis.

OBJECTIVE: In mice, melanoma inhibitory activity (MIA) is a chondrocyte-specific molecule with similar regulation to collagen type II. As MIA is a small secreted protein, its value as cartilage biomarker in human inflammatory arthritis was assessed. METHODS: MIA tissue distribution was studied by quantitative PCR and immunohistochemistry. The regulation of MIA production was studied in vivo in rheumatoid arthritis (RA) (n = 37) and spondyloarthritis (SpA) (n = 30) synovial fluid (SF), and in vitro in alginate embedded human chondrocytes. Therapeutic modulation of serum MIA was evaluated during tumour necrosis factor (TNF)alpha and interleukin (IL)1 blockade in RA. RESULTS: MIA was primarily expressed by chondrocytes in the human joint. SF MIA levels were lower in RA than in SpA despite similar levels of overall synovial inflammation. Further analysis indicated that these levels were inversely correlated with the degree of joint inflammation in RA, but not in SpA, and that the levels of TNFalpha and IL1beta were significantly increased in RA versus SpA. Accordingly, these proinflammatory cytokines suppressed MIA mRNA and protein in cultured chondrocytes. This suppression was paralleled by suppression of cartilage anabolism as assessed by collagen type 2 and aggrecan mRNA. Treatment of patients with RA with TNF blockade or IL1 blockade induced an increase of serum MIA levels. CONCLUSION: The decreased levels of MIA in the inflamed RA joint and the coregulation of MIA and cartilage matrix molecules by proinflammatory cytokines indicate that joint inflammation in RA not only drives accelerated cartilage degradation but also suppresses cartilage anabolism. This inflammation-driven suppression is reversible in vivo.

Validation of a new screening strategy for anti-extractable nuclear antigen antibodies.

OBJECTIVE: Although indirect immunofluorescence (IIF) is the most widely applied screening test for antinuclear antibodies (ANA), it lacks specificity for the identification of specific diseases or antigen reactivities. The aim of the present study was to validate an anti-extractable nuclear antigen (ENA) screening strategy encompassing a three-step cascade whereby an ELISA with pooled specific ENA is positioned between the IIF and the final anti-ENA identification. METHODS: Sera from 4 populations were tested for anti-ENA using an automated ELISA (EliA Symphony) and a line immunoassay (INNO-LIA ANA update). RESULTS: At the manufacturer's cut-off, a 96% sensitivity (95% CI 94%-98%) and 96% specificity (95% CI 94%-98%) of EliA Symphony for anti-ENA was obtained in a consecutive selection of 328 IIF positive serum samples referred for ANA testing. In addition, a high sensitivity was demonstrated for anti-ENA reactivities in patients with SLE (99%, 95% CI 97%-101%) and SSc (100%), and for anti-ENA monoreactivities. CONCLUSION: The EliA Symphony test was shown to be a sensitive second-line screening test for anti-ENA antibodies. In the context of a high clinical suspicion of connective tissue disease or autoreactivities not included in the EliA Symphony assay, third-line testing may be useful, even if the anti-ENA screening is negative.

Over-representation of construction-related occupations in male patients with systemic sclerosis.

AIMS: Based on preliminary observations, we tested the hypothesis that construction-related occupations are associated with systemic sclerosis (SSc). METHODS: The professional occupation of 91 patients with SSc (71 females and 20 males) was recorded. Categorisation into construction-related and other professions was performed. A double definition was used for construction-related occupations. The first (limited) definition was based upon categories of the Belgian National Institute of Statistics (NIS) occupational list. The following occupations were considered construction-related: electricians, joiners, masons and tilers, plumbers and pipefitters. The use of this list also allows us to compare the distribution of professions in these patients with that in the general population. As the NIS occupational list is limitative and leaves out some "real-life" construction-related occupations, a second and broader interpretation was given to the concept of construction-related occupations. RESULTS: The prevalence of construction-related professions in males with SSc, according to the limited definition, was 10-fold higher than in the general working population (50% vs 5%; p<0.001). Interestingly, most of the patients with construction-related occupations were electricians. In the broader interpretation, 75% of the men with SSc fell into the category of construction-related occupations. CONCLUSIONS: The data show an association between SSc and professional occupation.

Synovial detection and autoantibody reactivity of processed citrullinated isoforms of vimentin in inflammatory arthritides.

OBJECTIVES: To investigate the presence and characteristics of citrullinated vimentin in protein extracts of inflamed synovial tissue. METHODS: Cytosolic protein extracts obtained from RA (n = 14) and SpA patients (n = 14) were analysed by gel electrophoresis and western blotting. Citrullinated vimentin isoforms were visualized by a combination of anti-modified citrulline (AMC) staining and anti-vimentin detections (V9, H-84). This was subsequently confirmed by immunoprecipitation. Autoantibody detection was verified using sera obtained form RA (n = 6) and SpA (n = 6) patients. RESULTS: A specific cluster of spots displayed on the 2D gel images of cytosolic synovial tissue extracts, was identified by mass spectrometry as vimentin. Interestingly, our results suggested that these isoforms could be the result of caspase cleavage. In addition, these cleaved forms of vimentin were found to be citrullinated in synovial cytosolic protein extracts of inflammatory arthritides, mainly in RA patients. Caspase-3 is able to cleave vimentin at amino acid 85. Western blot analysis with a specific antibody against amino acids 1-84 of vimentin (H-84) confirmed that the citrullinated isoforms of vimentin were lacking this part of the protein. These results were also confirmed by immunoprecipitation of vimentin derived from cytosolic protein extracts of RA and SpA patients. Furthermore, the presence of autoantibodies against these citrullinated processed forms of vimentin was found to be predominantly associated with RA patients. CONCLUSIONS: These findings show the presence of processed citrullinated vimentin in inflammatory arthritides, mainly in RA and suggest a possible origin of the ACPA immune response in RA.

Do all anti-citrullinated protein/peptide antibody tests measure the same? Evaluation of discrepancy between anti-citrullinated protein/peptide antibody tests in patients with and without rheumatoid arthritis.

BACKGROUND: Different methods exist to demonstrate anti-citrullinated protein/peptide antibodies (ACPA). AIMS: To evaluate discrepancy between four ACPA tests. PATIENTS AND METHODS: Population 1 consisted of patients with a new diagnostic problem, including 86 patients with rheumatoid arthritis (RA) and 450 patients without RA. Population 2 consisted of 155 patients with RA who had long-standing disease. Population 3 consisted of 188 patients with psoriatic arthritis and in population 4 there were 192 patients with systemic lupus erythematosus. Populations 1 and 2 were tested with the anti-human fibrinogen antibody (AhfibA) test, anti-CCP2 from Eurodiagnostica (CCP2-euro), anti-CCP2 from Pharmacia (CCP2-phar) and anti-CCP3 test by Inova (CCP3). Samples were annotated as discrepant if positive in one and negative in at least one other test. Each discrepant sample was re-analysed in a different run. Populations 3 and 4 were analysed in the CCP2-euro and AhFibA test. RESULTS: In population 1, ACPA positivity was found in 17 of 450 (3.8%) patients without RA; 14 (82%) of these 17 samples were discrepant. In contrast, 61 of 86 (70.9%) patients with RA were ACPA positive of whom 18 of 61 (29.5%) were discrepant (70.9% vs. 29.5%, p<0.001). The discrepancies between tests could be partly attributed to borderline results, inter-assay discrepancy and inter-test variability. They were more prevalent in patients with systemic lupus erythematosus who were ACPA positive than in those with psoriatic arthritis who were ACPA positive. CONCLUSIONS: Discrepancy between different ACPA tests was observed attributable to the occurrence of borderline results, inter-assay variability and mainly to inter-test variability. The lowest inter-test discrepancy is observed between tests that use the same substrate.

Polymorphisms in the ficolin 1 gene (FCN1) are associated with susceptibility to the development of rheumatoid arthritis.

OBJECTIVES: We investigated the possible association of rheumatoid arthritis (RA) with single nucleotide polymorphisms (SNP) within the ficolin (FCN) genes. Two SNPs in the FCN1 gene, four SNPs in the FCN2 gene and one SNP in the FCN3 gene were studied. METHODS: The SNPs within the FCN genes were detected by an experimental INNO-LiPA methodology (Innogenetics, Belgium) in a population consisting of 338 RA patients and 595 controls. The significant SNPs were further evaluated in two subpopulations and related to carriage of the human leukocyte antigen-shared epitope (HLA-SE), rheumatoid factor (RF) and the presence of anti-citrullinated protein/peptide antibodies (ACPA). RESULTS: Two SNPs in the FCN1 gene were significantly associated with RA: the A allele rs2989727 was significantly increased in RA patients (67%) compared with controls (60%) (P = 0.002). Also, the frequency of the G allele of rs1071583 was increased in RA patients (68%) compared with controls (61%) (P = 0.003). Analysis of agreement between SNPs suggested strong linkage between rs2989727 and rs1071583. Carriage of a FCN1 SNP was independent of carriage of the HLA-SE, RF status and ACPA positivity. CONCLUSIONS: We describe two linked SNPs in the FCN1 gene that are associated with the development of RA.