Alain Saraux

BAFF Overexpression Is Associated with Autoantibody Production in Autoimmune Diseases

Abstract: The B‐cell activity factor (BAFF) acts as a positive regulator of B‐cell function. To gain further insight into the understanding of B‐cell hyperactivity in autoimmune diseases, the serum level of BAFF was determined in 43 systemic lupus erythematosus (SLE) patients, 58 primary Sjögrens syndrome (pSS) patients, 28 rheumatoid arthritis (RA) patients, and 68 normal control subjects using an in‐house sandwich ELISA. A commercial kit was used to detect soluble CD23 (sCD23) reflecting B‐cell activation. In‐house assays for the detection of autoantibodies also were used. We found an increased level of BAFF in SLE, pSS, and RA sera compared with normal subjects (respectively, 10.6 ± 8.5, 15.8 ± 12.9, 9.7 ± 1.5 ng/mL vs. 4.6 ± 2.9/ng/mL, P < .001). sCD23 released on B‐cell activation also was found to be elevated in SLE, pSS, and RA compared with normal sera. However, no correlation was found between the circulating BAFF and the level of sCD23. By contrast, we observed that high levels of BAFF were associated with the presence of autoantibodies (anti‐double‐stranded DNA antibodies in SLE, anti‐SSA antibodies in pSS, and rheumatoid factors in RA). Our data suggest that BAFF is influential in driving antibody production rather than activation of the B lymphocytes in autoimmune diseases.

The environment, geo-epidemiology, and autoimmune disease: Rheumatoid arthritis.

Rheumatoid arthritis (RA) is a chronic inflammatory joint disease characterized by a distinctive pattern of bone and joint destruction. RA patients have an increased risk of death. The incidence and prevalence of RA vary across populations, statistical methods, and disease definitions. In North America and Northern Europe, the incidence of RA is estimated at 20-50 cases per 100,000 population and the prevalence at 0.5-1.1%. Lower incidences and prevalences have been reported in Southern Europe, and few data are available for developing countries. Some studies showed declining incidences and prevalences after the 1960s. RA is a multifactorial disease that results from interactions between genetic and environmental factors. The main genetic factors are HLA-DRB1 and the tyrosine-phosphatase gene PTPN22. Among environmental factors implicated in the development of RA, smoking shows the strongest association with RA susceptibility and is also linked to worse outcomes. The aim of this review is to discuss the available data on the incidence and prevalence of RA, as well as the genetic and environmental risk factors associated with RA.

Prevalence of spondyloarthropathies in France: 2001

Objective: To estimate the prevalence of spondyloarthropathies (SpAs) in France in a multiregional representative sample in the year 2001. Methods: A two stage random sample was constituted in seven areas from the national telephone directory and the next birthday method in each household. Interviewers were patient-members of self help groups trained to administer telephone surveys using a validated questionnaire for detecting inflammatory joint disease. Quality of data collection was controlled periodically. SpA was confirmed by the patient’s rheumatologist or by clinical examination. Prevalence estimates after probability sampling correction were standardised for age and sex (1999 national census). Results: Among the 15 219 anonymous telephone numbers selected, 3.6% were places of work or secondary residences and were excluded. The phone interview participation rate ranged across regions from 55.1 to 69.9%. 3554 men and 5841 women were included in the study. Twenty nine cases of SpA were confirmed. All but one fulfilled ESSG criteria. Mean age was 47 years (range 21–78). The overall prevalence standardised for age and sex was 0.30% (95% confidence interval (CI) 0.17 to 0.46). Prevalence was similar in women (0.29% (95% CI 0.14 to 0.49)) and men (0.31 % (95% CI 0.12 to 0.60)). Geographical analysis by department clustering found no significant differences. The prevalence of SpA was as high as that of rheumatoid arthritis. Conclusion: Prevalence of SpA in France was 0.30% in 2001, with no difference between women and men. Ankylosing spondylitis and psoriatic arthritis were the most common SpA subsets.

Prevalence of rheumatoid arthritis in France: 2001

Background: Prevalence estimates of rheumatoid arthritis (RA) vary across Europe. Recent estimates in southern European countries showed a lower prevalence than in northern countries. Objectives: To estimate the prevalence of RA in France in a multiregional representative sample in the year 2001. Methods: A two stage random sample was constituted in seven areas (20 counties) from the national telephone directory of households and by the next birthday method in each household. Patient-interviewers, member of self help groups, were trained to administer telephone surveys using a validated questionnaire for case detection of inflammatory rheumatism, and conducted the survey under quality control. All suspected cases of RA were confirmed by their rheumatologist or by clinical examination. Prevalence estimates after probability sampling correction were standardised for age and sex (national census 1999). Results: An average response rate of 64.7% (two stages combined) led to a total of 9395 respondents. Standardised prevalence was 0.31% (95% confidence interval 0.18 to 0.48) for RA, 0.51% in women and 0.09% in men, with a higher age-specific prevalence in the 65–74 year age band. A geographical analysis of county clustering showed significant variation across the country. Conclusion: This national multiregional cooperative study demonstrates the usefulness of working in association with patients of self help groups. It showed a similar prevalence of RA to that of the spondyloarthropathies estimated concomitantly during the survey. It provides a reliable basis for definition of population targets for healthcare delivery and drug treatments.

Treatment of primary Sjögren syndrome with rituximab: a randomized trial.

Context Few trials have examined treatments for primary Sjgren syndrome (pSS). Contribution This multicenter, double-blind, placebo-controlled, randomized trial found that rituximab (given in 2 infusions over 2 weeks) alleviated some symptoms at week 6 but did not alleviate symptoms or improve global activity score at month 6 in adults with recent-onset or systemic pSS. More infusion reactions occurred with rituximab than placebo. Caution Outcome measurements may have been insensitive for detecting improvement. Implication Rituximab infusions did not produce sustained or substantial alleviation of symptoms or improvement in disease activity in adults with recent-onset or systemic pSS. The Editors Primary Sjgren syndrome (pSS) is a chronic autoimmune disorder characterized by dryness of the eyes and mouth and systemic involvement in up to 50% of cases (1). Histopathology shows lymphocytic infiltration and destruction of the lachrymal and salivary glands (2). To date, no systemic treatment has been proved to significantly affect the course of pSS (3), but clinicians may prescribe hydroxychloroquine to patients having fatigue or arthralgia and corticosteroids, methotrexate, or immunosuppressants to patients with systemic involvement. Because mounting evidence points to a central pathophysiologic role for B cells (47), B-cell depletion is being evaluated as a treatment of pSS (811). The most widely studied target for achieving B-cell depletion is the CD20 antigen, a transmembrane protein found on pre-B and mature B cells. It is neither shed from the cell surface nor internalized on antibody binding (1214). In open-label studies, the anti-CD20 antibody rituximab had a good safety profile, induced rapid B-cell depletion in blood and salivary glands, and seemed beneficial in early active pSS and in pSS with active extraglandular involvement (8, 9, 15). Two small, double-blind, randomized trials have been published (10, 11). The first included 18 patients and suggested an effect on the visual analogue scale (VAS) fatigue score after 6 months, although the primary end point, a 20% or greater decrease in the VAS fatigue score, was not achieved (10). The second trial included 30 patients with recent active pSS and showed improvements in the VAS dryness score and stimulated total salivary flow rate after 6 months (11). The purpose of the randomized, placebo-controlled TEARS (Tolerance and Efficacy of Rituximab in Primary Sjgrens Syndrome) trial reported here was to evaluate the efficacy and adverse effects of rituximab in pSS. Methods Design Overview This randomized, placebo-controlled, parallel-group trial evaluated global disease, pain, fatigue, and dryness. French rheumatologists and internists recruited the patients between 6 March 2008 and 5 January 2011. Patients were randomly assigned in a 1:1 ratio to blinded treatment with intravenous infusions of rituximab (1 g) or placebo at weeks 0 and 2. All study personnel, investigators, and patients remained blinded to the treatment group throughout the study. This study was approved by the appropriate ethics committee (CPP Ouest VI), and all patients gave written informed consent before study enrollment. The protocol was registered on ClinicalTrials.gov (NCT00740948). Setting and Participants Patients were recruited at 14 university hospitals in France if they fulfilled the AmericanEuropean Consensus Group criteria for pSS (16) and had active disease, defined as scores of at least 50 mm on at least 2 of 4 VASs (scores range from 0 [none] to 100 mm [worst]) for global disease, pain, fatigue, and dryness. Additional requirements were onset of pSS symptoms (first visit for any sign) in the past 10 years and biologically active pSS (defined as autoantibodies [anti-Ro/SSA antibodies or rheumatoid factor], cryoglobulinemia, hypergammaglobulinemia, 2-microglobulin elevation, or hypocomplementemia) or systemic pSS with at least 1 extraglandular manifestation or current parotid gland enlargement. The other inclusion criteria were informed consent, being aged 18 to 80 years, stable nonsteroidal anti-inflammatory drug regimen, no use of immunosuppressive agents for at least 4 weeks before inclusion, and use of an effective contraceptive method for patients able to conceive. Exclusion criteria were secondary Sjgren syndrome; cytotoxic drug therapy in the past 4 months; severe renal or hematologic failure; history of cancer, hepatitis B or C, HIV infection, tuberculosis, severe diabetes, or any other chronic disease; evidence of infection; history of severe allergic or anaphylactic reactions to humanized or murine monoclonal antibodies; and an inability to understand the study protocol. Randomization and Interventions Randomization was stratified by site. A computer-generated random allocation sequence was prepared by our statistics department in Brest, France. The infusions were prepared after a telephone call to the statistics department by pharmacists who were not involved in any other study procedure and were instructed not to disclose the treatment group to the investigators. All patients received the same volume, but the infusion contained the solvent only (normal saline or 5% glucose) in the placebo group and the solvent plus rituximab in the rituximab group. Before each rituximab or placebo infusion, the patients received 100 mg of methylprednisolone intravenously and 500 mg of acetaminophen orally. Outcomes and Follow-up Efficacy was evaluated at weeks 6, 16, and 24. The primary outcome, chosen a priori on the basis of expert opinion, was a 30-mm or greater improvement at week 24 versus baseline on at least 2 of the 4 VAS scores. Secondary outcomes included variations from baseline in the individual VAS scores at weeks 6 and 16; disease activity, systemic manifestations, and treatment activity assessed by the investigator as present or absent and by using both a physician VAS for disease activity and the European League Against Rheumatism Sjgrens Syndrome Disease Activity Index (ESSDAI), a clinical index that is designed to measure disease activity in patients with pSS (12 domains with a total score ranging from 2 to 47) (17, 18); basal salivary flow rate; Schirmer test and van Bijsterveld scores and Chisholm grade (19, 20); C-reactive protein level and erythrocyte sedimentation rate; rheumatoid factor; antinuclear antibodies; serum IgG, IgA, and IgM levels; serum complement; cryoglobulinemia; and serum level of B-cellactivating factor (BAFF) (21). Clinicians collected open-ended adverse events and assessed severity and potential causality at each visit from baseline to week 24. At study completion, the chief investigator categorized the adverse events according to the Medical Dictionary for Regulatory Activities, which is required by European regulations. We used Lower-Level Terms in the System Organ Class system. Statistical Analysis Our power calculation was based on our primary end point assessed at week 24. To detect a difference of 30 percentage points between groups in the proportion of patients achieving the primary end point, with a 2-sided of 0.05 and 80% power, we needed 49 patients per group. We planned to enroll 120 patients to allow for withdrawals and missing data. All randomly assigned patients who did not withdraw before the first study-drug infusion were included in the efficacy analyses. They were analyzed in the group to which they had been randomly assigned, even when a protocol deviation was reported (intention-to-treat principle). We used a fully conditional specification method to do multiple imputation and to handle missing data, which were assumed to be missing at random. We used the MICE function in R, version 2.14 (R Foundation for Statistical Computing, Vienna, Austria), to generate 20 imputed data sets. The initial data set to impute contained all outcomes of interest, baseline characteristics, and center and random assignments. To build the imputation model, we used the quickpred function in R to include all predictors with an absolute correlation of at least 0.2 with the target or the response indicator. Study center and treatment group were forced to be included in the imputation model. Continuous variables that were clearly nonnormal were transformed before imputation then back-transformed to create the final imputed data set. We analyzed the primary outcome at week 24 using a generalized linear model with binomial distribution, identity link, and exchangeable correlation structure to account for study center. Although identity is not the usual link for a binary response, it can be used in the present situation (22, 23) to estimate a risk difference with the CI, as recommended by the CONSORT (Consolidated Standards of Reporting Trials) statement. Secondary outcomes were analyzed by using the same statistical method, except a normal distribution was used for continuous data. All efficacy analyses were first done for each week by using the imputed data, except for the serum BAFF level, which was not collected in all study centers and was analyzed by using the observed data. Reductions in BAFF levels were compared between the rituximab and placebo groups by using the Wilcoxon test. For the 4 VASs used to define the primary end point, longitudinal analyses were then done on the observed data by using a mixed model. In these analyses, we used treatment group, visit, and the visittreatment group interaction term as independent variables; study center as a random-effects factor; a compound symmetry covariance structure to account for repeated measures among visits; and age, sex, baseline antibody values, and recent-onset or systemic pSS information as covariates. We used SAS, version 9.3 (SAS Institute, Cary, North Carolina), for all analyses except for multiple imputation, for which we used R, version 2.14. P values less than 0.05 were considered statistically significant. We used the MICE function in R to generate imputed data sets, PROC MIANAL

The DESIR cohort: A 10-year follow-up of early inflammatory back pain in France: Study design and baseline characteristics of the 708 recruited patients

OBJECTIVES The French Society of Rheumatology has initiated a large national multicenter, longitudinal, prospective follow-up of patients presenting with early inflammatory back pain in order to set up a database to facilitate several investigations on diagnosis, prognosis, epidemiology, pathogenesis and medico-economics in the field of early inflammatory back pain and spondyloarthritis. METHODS Patients were recruited if they had inflammatory back pain of more than 3 months and less than 3 years. Patients will be followed every 6 months during the first 2 years then every year during at least 5years. Apart from information collected on a Case Report Form (demographics, disease activity, severity, co-morbidities, socio-economics, treatments, radiological and MRI evaluation of the spine and the pelvis according to the local investigators, and for some centers bone densitometry and ultrasonography of entheses), the digital X-rays and MRI of the spine and pelvis are stored using a specific software (Carestream) and the biological samples (DNA, RNA, sera, urines) are centralized at the Biological Resources Center (Bichat Hospital). RESULTS The recruitment period of the 708 patients (mean age: 34±9years, female 54%, HLA-B27 positive: 57%) in the 25 centers was 26 months (from December 2007 to April 2010). The modified New York criteria, Amor criteria, ESSG criteria and axial ASAS criteria were fulfilled by 26%, 77%, 76% and 67% of the patients at entry, respectively. A history or current symptoms suggestive of peripheral arthritis, acute anterior uveitis and inflammatory bowel disease were observed in 21%, 9% and 4% of the patients, respectively. The disease was active (BASDAI: 45±20) despite an NSAID intake in 66% of the patients. CONCLUSION This large cohort should facilitate the conduct of researches in different areas (clinical, medico-economics, translational) in order to improve our knowledge on the pathogenesis and natural history of axial spondyloarthritis.

Evaluation of several ultrasonography scoring systems for synovitis and comparison to clinical examination: results from a prospective multicentre study of rheumatoid arthritis

Objectives To evaluate different global ultrasonographic (US) synovitis scoring systems as potential outcome measures of rheumatoid arthritis (RA) according to the Outcome Measures in Rheumatoid Arthritis Clinical Trials (OMERACT) filter. Methods To study selected global scoring systems, for the clinical, B mode and power Doppler techniques, the following joints were evaluated: 28 joints (28-joint Disease Activity Score (DAS28)), 20 joints (metacarpophalangeals (MCPs) + metatarsophalangeals (MTPs)) and 38 joints (28 joints + MTPs) using either a binary (yes/no) or a 0–3 grade. The study was a prospective, 4-month duration follow-up of 76 patients with RA requiring anti-tumour necrosis factor (TNF) therapy (complete follow-up data: 66 patients). Intraobserver reliability was evaluated using the intraclass correlation coefficient (ICC), construct validity was evaluated using the Cronbach α test and external validity was evaluated using level of correlation between scoring system and C reactive protein (CRP). Sensitivity to change was evaluated using the standardised response mean. Discriminating capacity was evaluated using the standardised mean differences in patients considered by the doctor as significantly improved or not at the end of the study. Results Different clinimetric properties of various US scoring systems were at least as good as the clinical scores with, for example, intraobserver reliability ranging from 0.61 to 0.97 versus from 0.53 to 0.82, construct validity ranging from 0.76 to 0.89 versus from 0.76 to 0.88, correlation with CRP ranging from 0.28 to 0.34 versus from 0.28 to 0.35 and sensitivity to change ranging from 0.60 to 1.21 versus from 0.96 to 1.36 for US versus clinical scoring systems, respectively. Conclusion This study suggests that US evaluation of synovitis is an outcome measure at least as relevant as physical examination. Further studies are required in order to achieve optimal US scoring systems for monitoring patients with RA in clinical trials and in clinical practice.

IL-6 Modulates CD5 Expression in B Cells from Patients with Lupus by Regulating DNA Methylation

B lymphocytes from patients with systemic lupus erythematosus (SLE) are characterized by reduced expression levels of membrane CD5. Recent studies from our laboratory have revealed that the level of membrane CD5 is determined by the relative level of two alternative CD5 isoforms; CD5-E1A, which is expressed on the membrane, and CD5-E1B, which is retained in the cytoplasm. Using bisulfite sequencing and methylation-sensitive endonuclease assays we show that the promoter for the alternative CD5-E1B isoform is demethylated in B cells from patients with SLE but not in healthy controls. We go on to show that differential methylation is more pronounced following BCR engagement. As a result of this demethylation, CD5-E1B mRNA is transcribed at the expense of CD5-E1A mRNA transcription. We provide further evidence that production of high IL-6 levels by SLE B cells abrogates the ability of SLE B cells to induce DNA methyl transferase (DNMT1) and then to methylate DNA, an effect that is reversed in the presence of a blocking Ab to the IL-6 receptor. The pattern of demethylation of CpG islands in the CD5-E1B promoter in SLE B cells is similar to those in B cells from healthy controls stimulated in the presence of IL-6, or treated with the methylation inhibitor PD98059. The study reveals that engagement of the BCR with constitutive IL-6 down-regulates the level of membrane CD5, which negatively regulates BCR signaling, in SLE B cells. This altered signaling could, in turn, promote the activation and expansion of autoreactive B cells in SLE patients.

Ectopic Germinal Centers Are Rare in Sjögren’s Syndrome Salivary Glands and Do Not Exclude Autoreactive B Cells

This study reports on the characterization of B cells of germinal center (GC)-like structures infiltrating the salivary glands (SGs) of patients with Sjögren’s syndrome. Eight two-color combinations were devised to characterize the phenotype of these B cells in 11 SG specimens selected from biopsies obtained from 40 Sjögren’s syndrome patients and three normal tonsils. The 9G4 mAb, which recognizes V4.34-encoded autoAbs, enabled us to identify autoreactive B cells. Quantitative RT-PCR was used to determine the level of mRNAs for activation-induced cytidine deaminase (AICDA), repressors and transcription factors. CD20+IgD−CD38+CD21+CD24− B cells, similar to those identified in tonsil GCs, were seen in the SGs of four patients and, and since they expressed AICDA, they were termed “real GCs”. CD20+IgD+CD38−CD21+CD24+ B cells, seen in aggregates from the remaining seven samples, were characteristically type 2 transitional B cells and marginal zone-type B cells. They lacked AICDA mRNAs and were termed “aggregates”. Real GCs from SGs contained mRNAs for Pax-5 and Bcl-6, like tonsil GC cells, whereas aggregates contained mRNAs for Notch-2, Blimp-1, IRF-4, and BR3, similar to marginal zone B cells. Further experimental data in support of this dichotomy included the restriction of CXCR5 expression to real GC cells, while sphingosine 1-phosphate receptor 1 was expressed only in aggregates. In contrast, both types of B cell clusters expressed the idiotype recognized by the 9G4 mAb. Our data indicate that, in SGs, a minority of B cell clusters represent genuine GC cells, while the majority manifest features of being type 2 transitional B cells and marginal zone cells. Interestingly, both types of B cell aggregates include autoreactive B cells.

The environment, geo-epidemiology, and autoimmune disease: Rheumatoid arthritis

Rheumatoid arthritis (RA) is a chronic inflammatory joint disease characterized by a distinctive pattern of bone and joint destruction. RA patients have an increased risk of death. The incidence and prevalence of RA vary across populations, statistical methods, and disease definitions. In North America and Northern Europe, the incidence of RA is estimated at 20 to 50 cases per 100,000 population and the prevalence at 0.5% to 1.1%. Lower incidences and prevalences have been reported in Southern Europe, and few data are available for developing countries. Some studies showed declining incidences and prevalences after the 1960s. RA is a multifactorial disease that results from interactions between genetic and environmental factors. The main genetic factors are HLA-DRB1 and the tyrosine-phosphatase gene PTPN22. Among environmental factors implicated in the development of RA, smoking shows the strongest association with RA susceptibility and is also linked to worse outcomes. The aim of this review is to discuss the available data on the incidence and prevalence of RA, as well as the genetic and environmental risk factors associated with RA.