Patrick Verschueren

Detection of antinuclear antibodies by indirect immunofluorescence and by solid phase assay

Testing for antinuclear antibodies is useful for the diagnosis of systemic rheumatic diseases. Solid phase assays are increasingly replacing indirect immunofluorescence for detection of antinuclear antibodies. In the most recent generation of solid phase assays, manufacturers attempt to improve the performance of the assays by adding extra antigens. Solid phase assay (EliA CTD Screen, Phadia, in which antibodies to 17 antigens are detected) was compared to indirect immunofluorescence for the detection of antinuclear antibodies in diagnostic samples of 236 patients with autoimmune connective tissue diseases, in 149 healthy blood donors, 139 patients with chronic fatigue syndrome, and 134 diseased controls. The sensitivity of EliA CTD Screen for systemic lupus erythematosus, systemic sclerosis, primary Sjögrens syndrome, mixed connective tissue disease, and inflammatory myopathy was 74%, 72%, 89%, 100%, and 39%, respectively. The reactivity in blood donors, in patients with chronic fatigue syndrome, and in diseased controls was <4%. Likelihood ratios increased with increasing antibody concentrations. Generally, a positive test result by EliA CTD Screen had a higher likelihood ratio for systemic rheumatic disease than a positive test result by indirect immunofluorescence. A negative test result by indirect immunofluorescence, however, had a lower likelihood ratio than a negative test result by EliA CTD Screen, indicating that the negative predictive value was higher for indirect immunofluorescence than for EliA CTD screen.

Antinuclear antibody detection by automated multiplex immunoassay in untreated patients at the time of diagnosis

Fully automated multiplex immunoassays are increasingly used as first line screening for antinuclear antibodies. The diagnostic performance of such multiplex assays in untreated patients at the time of diagnosis has not been reported. Antinuclear antibodies were measured by indirect immunofluorescence (IIF) (dilution 1:160) and by BioPlex 2200 ANA screen (antibodies to dsDNA, chromatin, ribosomal protein, SSA-52, SSA-60, SSB, Sm, SmRNP, RNP-A, RNP-68, Scl-70, Jo-1, and centromere B) in 236 patients with a systemic rheumatic disease at the time of diagnosis, 149 blood donors, 139 patients with chronic fatigue syndrome (CFS), and 134 diseased controls. BioPlex ANA screen and IIF were positive in, respectively, 79% and 90% of patients with systemic lupus erythematosus (SLE), 60% and 60% with cutaneous lupus, 72% and 93% with systemic sclerosis (SSc), 100% and 100% with mixed connective tissue disease (MCTD), 89% and 56% with primary Sjögrens (SS) syndrome, 36% and 36% with polymyositis/dermatomyositis, 5.4% and 6% of blood donors, 7.2% and 3.6% of patients with CFS, and 11% and 18% of diseased controls. BioPlex test result interval specific likelihood ratios increased with increasing antibody concentration. The simultaneous presence of at least three antibodies by BioPlex was found in 35% of patients with SLE, 4% with SSc, 100% with MCTD, 64% with SS, 7% with inflammatory myopathy, 0.7% of CFS and diseased controls, and none of the blood donors. In conclusion, test result specific likelihood ratios and the presence of multiple autoantibodies help with the interpretation of data generated by multiplex immunoassays.

Methotrexate in combination with other DMARDs is not superior to methotrexate alone for remission induction with moderate-to-high-dose glucocorticoid bridging in early rheumatoid arthritis after 16 weeks of treatment: the CareRA trial

Objectives To compare the efficacy and safety of intensive combination strategies with glucocorticoids (GCs) in the first 16 weeks (W) of early rheumatoid arthritis (eRA) treatment, focusing on high-risk patients, in the Care in early RA trial. Methods 400 disease-modifying antirheumatic drugs (DMARD)-naive patients with eRA were recruited and stratified into high risk or low risk according to classical prognostic markers. High-risk patients (n=290) were randomised to 1/3 treatment strategies: combination therapy for early rheumatoid arthritis (COBRA) Classic (methotrexate (MTX)+ sulfasalazine+60 mg prednisone tapered to 7.5 mg daily from W7), COBRA Slim (MTX+30 mg prednisone tapered to 5 mg from W6) and COBRA Avant-Garde (MTX+leflunomide+30 mg prednisone tapered to 5 mg from W6). Treatment modifications to target low-disease activity were mandatory from W8, if desirable and feasible according to the rheumatologist. The primary outcome was remission (28 joint disease activity score calculated with C-reactive protein <2.6) at W16 (intention-to-treat analysis). Secondary endpoints were good European League Against Rheumatism response, clinically meaningful health assessment questionnaire (HAQ) response and HAQ equal to zero. Adverse events (AEs) were registered. Results Data from 98 Classic, 98 Slim and 94 Avant-Garde patients were analysed. At W16, remission was reached in 70.4% Classic, 73.6% Slim and 68.1% Avant-Garde patients (p=0.713). Likewise, no significant differences were shown in other secondary endpoints. However, therapy-related AEs were reported in 61.2% of Classic, in 46.9% of Slim and in 69.1% of Avant-Garde patients (p=0.006). Conclusions For high-risk eRA, MTX associated with a moderate step-down dose of GCs was as effective in inducing remission at W16 as DMARD combination therapies with moderate or high step-down GC doses and it showed a more favourable short-term safety profile. EudraCT number: 2008-007225-39.

CCR5 blockade in rheumatoid arthritis: a randomised, double-blind, placebo-controlled clinical trial

Objective C-C chemokine receptor type 5 (CCR5), a chemokine receptor expressed on T cells and macrophages, and its ligands are found in inflamed synovial tissue (ST) of patients with rheumatoid arthritis (RA). The rationale for testing CCR5 blockade in patients with RA was supported by the effects of a CCR5 antagonist in collagen-induced arthritis in rhesus monkeys. The effects of CCR5 blockade in patients with active RA were explored. Methods In this phase Ib randomised, placebo-controlled trial, treatment with an oral CCR5 inhibitor (SCH351125) in patients with active RA was evaluated. Clinical efficacy was assessed using European League Against Rheumatism and American College of Rheumatology response criteria. ST biopsies were taken before and after 28 days of treatment, and analysed for CCR5+ cells. In a subset of patients, MRIs of an inflamed joint were obtained before and after treatment. Results In all, 32 patients were included; 20 received SCH351125 and 12 placebo. Three patients who received SCH351125 did not complete the study due to adverse events; none of these were serious. No improvement was observed in the active treatment group compared to placebo. Results were consistent for clinical evaluation, ST analysis and MRI. Conclusion This proof of concept study does not support the use of CCR5 blockade as a therapeutic strategy in patients with active RA.

Detection of antinuclear antibodies by automated indirect immunofluorescence analysis

BACKGROUND Testing for antinuclear antibodies is useful for the diagnosis of systemic rheumatic diseases. Automated systems for image acquisition and interpretation of indirect immunofluorescence-based tests are increasingly used. The diagnostic performance of such automated approach in untreated patients has not been reported. METHODS Antinuclear antibodies were measured by automated indirect immunofluorescence using Zenit G. Sight on HEp2 and HEp2000 substrate in 268 consecutive samples submitted to the laboratory for antinuclear antibody testing, and in 231 patients with a systemic rheumatic disease at the time of diagnosis, 143 blood donors, 134 patients with chronic fatigue syndrome, and 133 diseased controls. RESULTS Image acquisition by G-Sight was of high quality. The accuracy of pattern assignment was limited. There was a significant correlation between automated estimation of fluorescence intensity (probability index of positivity) and end-point titer. Probability index interval specific likelihood ratios for systemic rheumatic disease increased with increasing level of positivity probability. With the HEp-2 substrate, the likelihood ratio for systemic lupus erythematosus was 0.06, 0.4, 6.8, 12.1, and 43.9 for a probability measure of positivity of ≤10, 11-≤30, 31-≤50, 51-≤85, and >85, respectively. CONCLUSION Quantitative data generated by automated image acquisition facilitates standardized interpretation.

Predictors of remission, normalized physical function, and changes in the working situation during follow‐up of patients with early rheumatoid arthritis: an observational study

Objectives: To evaluate possible predictors of remission, normalized physical function, and work change in rheumatoid arthritis (RA). Methods: We determined in our early RA cohort the proportion of patients in remission [Disease Activity Score (DAS28)<2.6], with normalized function [Health Assessment Questionnaire (HAQ) = 0], and with changed working situation since disease onset. Candidate predictors of remission, normalized function, and work change were studied by subgroup comparison and logistic regression analysis, including demographics, education, rheumatoid factor (RF), anti‐cyclic citrullinated peptide (anti‐CCP) antibodies, treatment, and DAS28, HAQ, pain and fatigue scores (on the visual analogue scale, VAS). Results: Median (interquartile range, IQR) disease duration was 18 (29) months. Of 89 patients included, 69% were in remission. DAS28, HAQ, pain and fatigue scores of these patients were lower throughout year 1, although similar at baseline, compared to patients not in remission. At month 4, more of these patients were already good responders. Remission at month 4 independently predicted remission at follow‐up. Thirty‐eight per cent had no functional limitations; compared to patients with limitations, they had a lower baseline HAQ and lower DAS28, HAQ, pain and fatigue scores during year 1. At month 4, more achieved remission or HAQ = 0. Male sex, baseline HAQ, and month 4 good European League Against Rheumatism (EULAR) response predicted long‐term HAQ = 0, but month 4 HAQ = 0 was the strongest independent predictor. Of the 40% with a paid job at baseline, 43.8% reported changes in their work situation; they had higher DAS28, HAQ, pain and fatigue scores during year 1. Failing a month 4 good EULAR response independently predicted work change. Conclusion: Month 4 disease response predicts later remission, normalized physical function, and work change in RA.

Benefit of anti-TNFalpha treatment for nephrotic syndrome in a patient with juvenile inflammatory bowel disease associated spondyloarthropathy complicated with amyloidosis and glomerulonephritis

Historically, AA amyloidosis accounts for almost half of the deaths among patients with juvenile chronic arthritis, mainly due to complications of end stage renal failure.1 Improved survival has been reported in patients whose underlying inflammatory disorder was brought to remission.2 Tumour necrosis factor (TNFα) blocking agents have been used successfully in the treatment of inflammatory disorders complicated with AA amyloidosis.3–5 We report the effect of TNFα in a case of AA amyloidosis secondary to juvenile spondyloarthropathy. A 26 year old man with juvenile, inflammatory bowel disease associated spondyloarthropathy (HLA-B27+) was admitted to our hospital with proteinuria and ankle oedema. He received combination therapy with methotrexate, sulfasalazine, methylprednisolone, and naproxen. His blood pressure was 130/80 mmHg. Table 1 shows the results of laboratory tests. View this table: Table 1 Laboratory results at the time of admission Ultrasound examination showed an increased bipolar size (130 mm) of both kidneys. A chest x ray examination was normal. A renal …

Effectiveness of an integrated outpatient care programme compared with present-day standard care in early rheumatoid arthritis

OBJECTIVES To investigate the effectiveness of an integrated care programme in daily practice compared with present-day standard care for ambulatory early rheumatoid arthritis patients. METHODS In this cross-sectional study, group A received programmed multidisciplinary outpatient care and group B standard rheumatologist-centred care. Demographics, disease duration, initial and actual treatment, disease activity (Disease Activity-28 Score), general health (Short Form-36 [SF-36]), functionality (Health Assessment Questionnaire [HAQ]), coping style (Utrechts Coping List), illness perception (Dutch-Revised Illness Perception Questionnaire) and satisfaction about care were recorded. RESULTS Eight-nine patients were included in group A and 102 in group B. Demographics, rheumatoid factor, antibodies against cyclic citrullinated peptides and disease duration were comparable. More patients in group A received initial combination therapy (35% versus 3%). Actual treatment regimens were comparable. More patients were in remission (69% versus 39%) or had low disease activity (80% versus 60%), mean HAQ-scores were lower (0.52 versus 0.80), more patients had no functional impairment (38% versus 15%) and SF-36 scores were higher in group A. Coping style and illness perception were similar, except for illness coherence. Satisfaction differed only for aspects typically favouring a care programme. Participation in a care programme independently predicted remission and absence of disability in a regression model, including gender and initial treatment as other predictors. CONCLUSION Disease activity was better controlled and functionality and general health better preserved in patients following an outpatient care programme. This was partly due to the easier implementation of an intensive initial treatment strategy but apparently also to other aspects of organized pharmacological and non-pharmacological care, to be defined in randomized, controlled studies.

Pyoderma gangrenosum developing during therapy with TNF-alpha antagonists in a patient with rheumatoid arthritis

Dear Editor, Tumour necrosis factor-alpha (TNF) antagonists are effective for the treatment of rheumatoid arthritis (RA), inflammatory bowel disease (IBD), psoriatic arthritis and ankylosing spondylitis. They are sometimes used off label in various inflammatory disorders with cutaneous manifestations such as sarcoidosis [1], multi-centric reticulohistiocytosis [2], cicatricial pemphigoid [3] and pyoderma gangrenosum (PG) [4]. There have been a number of reports [4–9] and one randomised trial of PG [10] responding to TNF blockade. One case report showed a beneficial effect switching from one TNF blocker to another [11]. We present a case of a patient with RA who developed pyoderma gangrenosum under infliximab, and persisting cutaneous inflammation when switching to etanercept. A 53-year-old woman with a 5-year history of rheumatoid factor, anti-CCP positive, erosive RA developed an erythematous induration leading to progressive ulcers on her left foot. For the preceding 6 months, she had been treated with methotrexate 20 mg once weekly, methylprednisolone 4 mg/day and infliximab 3 mg/kg once every 8 weeks. Infection was ruled out by cultures on biopsymaterial that was consistent with PG. Methotrexate and infliximab were stopped, and the patient was treated with cyclosporine (3 mg/kg/day) while 4 mg/day of methylprednisolone was continued. The lesions resolved completely. However, she developed a flare of polysynovitis, which was successfully controlled with methotrexate (20 mg weekly) and methylprednisolone 8 mg daily, but when tapering steroids to 4 mg/day the cutaneous lesions of the left foot reappeared together with a flare of joint symptoms. Etanercept, 25 mg twice a week in combination with cyclosporine was effective for the joint symptoms but failed to control cutaneous inflammation and ulceration. When oral minocycline at a daily dose of 100 mg was started by her general practitioner for a non-related infection, this resulted in almost complete disappearance of the PG while continuing methotrexate, steroids and etanercept for the joint symptoms. PG is a neutrophilic dermatosis of unknown cause, often associated with underlying systemic disease such as IBD, RA, monoclonal gammapathy or malignancies. No uniform pattern of immune response has been described although TNF-alpha [12] is considered to be a major cytokine in the pathogenesis. On the other hand, bacterial infection or bacterial superantigens are also suspected in the ethiopathogenesis [13], which might question the use of TNF antagonists. Our case appears to be unique: PG appeared under infliximab for RA and stopping the drug resulted in the complete resolution of the lesion. Reinitiating TNF blockade with etanercept for active arthritis did not affect the skin lesions that had relapsed. Ultimately, this patient’s PG responded to minocycline therapy while continuing etanercept. This response to minocycline relies most likely on an immunomodulating role as described [14] in patients with RA. An explanation for our case might be that TNF blockade facilitates a bacterial infection responsible for the genesis or exacerbation of cutaneous lesions. The paradox was recently discussed, reviewing multiple reports of developClin Rheumatol (2007) 26:2205–2206 DOI 10.1007/s10067-007-0733-8

Inflammatory factors in the circulation of patients with active rheumatoid arthritis stimulate osteoclastogenesis via endogenous cytokine production by osteoblasts

SummaryThe combination of cytokines present in the circulation of patients with active rheumatoid arthritis might contribute to the generalized bone loss that commonly occurs in these patients, by directly inhibiting osteoblast proliferation and differentiation, but especially by enhancing endogenous cytokine (i.e., receptor activator of nuclear factor-kappa B ligand (RANKL) and interleukin-6 (IL)-6) production by osteoblasts, thereby stimulating osteoclastogenesis.IntroductionGeneralized bone loss, as occurs in patients with rheumatoid arthritis (RA), is related to elevated levels of circulating cytokines. Individual cytokines have deleterious effects on proliferation and differentiation of osteoblast cell lines, but little is known about the effect of the interaction between inflammatory factors in the circulation of patients with active RA on human osteoblast function, including their communication towards other bone cells. We investigated whether serum from patients with active RA enhances cytokine production by osteoblasts, thereby effectively altering osteoblast-stimulated osteoclastogenesis.MethodsSerum was obtained from 20 patients with active RA (active RA sera) and from the same patients in clinical remission (remission RA sera). To determine osteoclastogenesis, RA serum-pretreated primary human osteoblast cultures were established in direct contact with human osteoclast precursors in the presence or absence of osteoprotegerin (OPG) or IL-6 inhibitor.ResultsCompared to remission RA sera, active RA sera inhibited osteoblast proliferation and differentiation in vitro as demonstrated by a reduced DNA content and gene expression of KI-67, collagen type 1, osteopontin, and osteocalcin. Active RA sera inhibited OPG expression and enhanced RANKL and IL-6 expression but did not alter IL-8 expression in osteoblasts. IL-1β, IL-17, and tumor necrosis factor-α (TNF-α) expression were undetectable. In coculture, active RA sera treatment of osteoblasts stimulated while addition of OPG or IL-6 inhibitory antibodies significantly reduced the number of osteoclasts.ConclusionActive RA sera contain circulating factors, likely cytokines and chemokines, that might contribute to bone loss by directly inhibiting osteoblast proliferation and differentiation, but especially, these factors modulate endogenous cytokine production by osteoblasts, thereby affecting osteoclastogenesis.