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The diagnosis, treatment, and monitoring of the primary systemic vasculitides associated with circulating antineutrophil cytoplasm autoantibodies (ANCA) have formed the focus of a multicenter collaborative study. Consensus has been reached on criteria for classification, clinical subgroupings by extent and severity of disease, and ''standard'' and ''best alternative'' therapeutic regimens. Two series of randomized controlled trials have been designed; their aims are (1) to harmonize current approaches to treatment and (2) to test the value of newer therapeutic agents, In support of these trials, semiobjective scoring systems have been created and validated, and previous standardization of ANCA serologic and histologic analysis has been adopted. The systems of classification and clinical management described herein represent the recommendations of a multidisciplinary study group that hopes to improve the outcome of patients with primary systemic vasculitis by wide dissemination of the collective experience from interested centers.

Background. The predictive value of clinical and renal histological features for renal outcome in patients with anti-neutrophil cytoplasmic autoantibody (ANCA)-associated glomerulonephritis was investigated in a prospective analysis of 96 patients with ANCA-associated vasculitis, and moderate renal involvement (creatinine

Proteinase 3 (PR3), which is also called myeloblastin, the target autoantigen for antineutrophil cytoplasmic antibodies (ANCA) in Wegener's granulomatosis, is a serine proteinase stored in azurophil granules of human neutrophils. We have previously shown that, in contrast to elastase or myeloperoxidase, PR3 is also expressed at the plasma membrane of a subset of unactivated neutrophils and that a high proportion of neutrophils expressing membrane PR3 is a risk factor for vasculitis. The present study demonstrates that the association of PR3 with the plasma membrane is not an ionic interaction and seems to be covalent. Fractionation of neutrophils shows that, besides the azurophil granules, PR3 could be detected both in specific granules and in the plasma membrane-enriched fraction containing secretory vesicles, whereas elastase and myeloperoxidase were exclusively located in azurophil granules. Electron microscopy confirms that PR3 is present along with CR1 in secretory vesicles as well as in some specific granules. In neutrophils stimulated with an increasing dose of FMLP, membrane PR3 expression increased with the degranulation of secretory vesicles, followed by specific granules, and culminated after azurophil granules mobilization. The presence of a readily plasma membrane-mobilizable pool of PR3 contained in the secretory vesicles might play a relevant role in the pathophysiological mechanisms of ANCA-associated vasculitis.

We retrospectively analyzed the significance of persistent ANCA positivity after clinical remission in 53 consecutive patients with histologically and/or angiographically proven polyarteritis nodosa (PAN), or Churg-Strauss syndrome (CSS) followed between 1981 and 1993.ANCA were detected using an immunofluorescence assay and ELISA: Each patient met the American College of Rheumatology 1990 criteria for PAN or CSS. Clinical and biological evaluations were always essential factors in the decision to intensify therapy.ANCA were initially present in 15 patients (28.3%): 3/26 (11.5%) with HBV-related PAN, 6/18 (33.3%) with PAN of unknown etiology and 6/9 (66.7%) with CSS. Five patients remained ANCA-positive after clinical remission: 3 with PAN (one of them relapsed) and 2 with CSS who both relapsed. Among the 12 patients who died during follow-up, only 1 (8.3%) was initially ANCA-positive. Fifteen of the 41 survivors (29.2%) were ANCA-positive.Persistence of ANCA positivity in PAN and CSS may be a marker of an underlying disease process, but does not adequately reflect disease activity and, thus, in no case should be the only indication for therapeutic intensification.

We report on an unusual association between partial IgA deficiency and acute lymphoblastic leukemia in a young man. We also report results of the family study of immunoglobulin levels, sIgA B cells, in vitro IgA synthesis and molecular analysis of the structural C alpha genes. The IgA deficiency was present at diagnosis of leukemia prior to any therapy. The indirect arguments for a preexisting IgA deficiency are the absence of improvement of the IgA level during complete remission and especially the finding of a similar partial IgA deficiency in a sister who shared the same HLA haplotype, had a low percentage of sIgA B cells and decreased in vitro IgA production. The mother, who had a normal IgA serum level also had decreased in vitro IgA synthesis. No major structural C alpha gene defect was found. The relationship between acute lymphoblastic leukemia and IgA deficiency remains to be elucidated.

Resume De nombreux produits sont desormais connus pour leurs proprietes immunostimulantes. Ceux-ci peuvent etre classes en differents groupes: produits bacteriens, extraits leucocytaires (Facteur de Transfert, Interferon), hormones thymiques et drogues synthetiques (Levamisole). Pour differentes raisons, leurs indications au cours des maladies infectieuses sont encore peu frequentes. Premierement, les immunostimulants, en particulier les produits d'origine bacterienne ont des effets toxiques. Deuxiemement, ces produits ne sont generalement pas completement purifies et caracterises chimiquement. Troisiemement, les mecanismes d'action des immunostimulants sont encore incompletement analyses et ces substances peuvent egalement avoir des effects suppresseurs selon la voie et le moment de leur injection. Pour ces raisons, la generalisation de leur utilisation necessite l'obtention de produits purifies ou synthetiques, et l'analyse de leurs effets experimentaux et cliniques. Ces conditions sont deja remplies pour certains immunostimulants au cours d'infections virales en particulier chez des patients immunodeficients.

A reproducible immunocyto-adherence assay has been developed to study the modulation of Fc receptors for IgA (Fc alpha R), using a murine T cell hybridoma (T2D4), which expresses Fc receptors for all known isotypes of secreted immunoglobulins. By using sheep red blood cells coated with the hapten 2-4-6 trinitrophenyl (TNP), as indicator cells, and a murine monoclonal IgA (MOPC 315) antibody with anti-TNP activity, we were able to study the Fc alpha R on T2D4 cells. We found that: (a) murine Fc alpha R can bind human monoclonal IgA, and this binding is isotype specific since it was inhibited by human monoclonal IgA but not by human monoclonal IgG or IgM; (b) the expression of murine Fc alpha R is unducible by human monoclonal IgA, and this effect is isotype specific since it is not observed with human monoclonal IgM or IgG (c) sera from patients with IgA nephropathy can also induce Fc alpha R expression; by contrast, no induction was observed with normal human sera, (d) in one serum from an IgA-nephropathy patient, the inducer factor was characterized by affinity chromatography on anti-IgA-Sepharose and by gel filtration: high molecular weight IgA, probably IgA aggregates or immune complexes were recognized to be responsible for the induction of murine Fc alpha R expression.

The presence of DNA-anti-DNA immune complexes in sera from patients with systemic lupus erythematosus (SLE) was investigated by a new solid phase radioimmunoassay (RIA). This assay used murine monoclonal anti-double stranded DNA (dsDNA) antibody to recognize DNA present in the complexes and 125I-rabbit anti-human gamma globulin as a tracer. DNA-anti-DNA immune complexes were found in certain SLE sera but not in sera from patients with other immune complex diseases and from healthy blood donors. The presence of circulating DNA-anti-DNA complexes was associated with low C4 levels. It was not related to the presence of immune complexes detected by the polyethylene glycol assay suggesting either that the assay did not detect all DNA-anti-DNA complexes or that other antigen-antibody systems constitute the major immune complex components in SLE sera. The clinical significance of circulating DNA-anti-DNA complexes in SLE sera as well as the potential use of this solid phase RIA using various monoclonal antibodies to detect specific antigen-antibody systems is discussed.

In vitro CR1-dependent phagocytosis of C3b-coated erythrocytes, by monocytes and PMN, was found to be significantly decreased in SLE patients. This was in many cases related to a specific defect of CR1 receptors, since the FcR-ingestion of EIgG was normal. On the other hand, CR1 levels of PMN stimulated by FMLP were also found to be decreased in SLE patients, while both the expression of circulating PMN (cells isolated at 4 degrees C) and the total cellular CR1 content were normal. On the basis of the available data, we propose that the impaired phagocytosis is due to a functional defect of CR1 or a defective anchorage of the receptor to the plasma membrane, possibly related to the decreased capacity of CR1 to be up-regulated by FMLP. To study the importance of the genetic background in the CR1 abnormalities, the families of 22 young SLE patients, in which the onset of the disease had occurred before the age of 15, were studied. The expression of CR1 on erythrocytes, and the total CR1 content of PMN, in parents and siblings of these patients did not differ significantly from normal controls. By contrast, the ingestion of EIgGC3b by PMN from healthy relatives of these patients was decreased (65% of the normal mean of PI), while EIgG phagocytosis was normal. A relation between this CR1 functional defect and the familial occurrence of autoimmune disorders is therefore possible.

Three recent cases of inherited deficiency of the seventh component of complement (C7) associated with recurrent infectious meningitis are described. Two cases were associated with meningococcal meningitis, the third is the first case report of C7 inherited deficiency associated with Haemophilus parainfluenzae meningitis. Family studies are consistent with inheritance and non-HLA-linked, autosomal codominant trait of the C7 deficiency. The three patients have remained well, following antibiotic treatment.

DNA structural changes responsible for hereditary angioedema were sought in the C1-inhibitor gene, which contains unusually dense clusters of Alu repeats in various orientations. Among patients belonging to 45 unrelated families, eight partial C1-inhibitor gene deletions and a partial duplication were found. Four deletions had one of the boundaries within the gene and the other in extragenic regions--in three cases 5' of the gene and in one case 3' of the gene. The boundaries of the partial duplication and of the remaining four deletions mapped instead within a few kilobases of exon 4. The same element--Alu 1--the first of three tandem Alu repeats preceding exon 4, contained one of the breakpoints of each of these five rearrangements. Moreover, these recombination breakpoints spread over the entire length of Alu 1, in contrast with the tight clustering observed near the 5' end of Alu sequences rearranged in other human genes. Thus, two uncommon recombinational biases are observed in the Alu rearrangements of hereditary angioedema patients; one promotes the occurrence of intragenic breakpoints in a single Alu repeat, and the other allows the breaks to be distributed over the entire Alu structure rather than within the hot spot of the left Alu monomer. A region of potential Z-DNA structure, located 1.7 kb upstream of Alu 1, may contribute to both peculiarities.

Background. We reported previously that in renal disease in relation to antineutrophil cytoplasm auto-antibodies (ANCA)-associated vasculitis, renal outcome correlates better with the percentage of normal glomeruli than with separate active lesions. This may imply that glomeruli, once affected by necrotizing and crescentic lesions, are irreversibly damaged. We quantified and evaluated the course of renal lesions in the present study. Methods. We retrospectively analysed 31 patients with renal disease in relation to ANCA-associated vasculitis, all treated with immunosuppressive drugs. In all patients, a renal biopsy was performed at diagnosis. A follow-up biopsy was performed in all patients on the indication of a suspected renal relapse, after a mean interval of 31 months. Results. The mean percentage of normal glomeruli in the renal biopsy did not change over time (29% in the initial and 30% in the follow-up biopsy). The mean percentage of glomeruli with crescents, however, significantly decreased from 57 to 30% (P < 0.001). The percentage of glomerulosclerosis significantly increased from 12 to 39% (P < 0.001). The data were independent of diagnosis, gender, age, time interval between the biopsies, and treatment. Conclusions. This is the first study to quantify glomerular changes between two time points in patients with renal vasculitis. Our results suggest that, on average, no new glomeruli are recruited into the active disease process. The sum of the percentage of crescentic and sclerotic glomeruli in the initial biopsies is larger than the percentage of sclerotic glomeruli in the follow-up biopsies. Thus, therapy seems not only to prevent normal glomeruli from being recruited into the active disease process for a certain time, but seems also to allow part of the active lesions to revert into a normal phenotype, although another part of the active lesions will be transformed to a chronic phenotype

Diagnostic value of standardized assays for anti-neutrophil cytoplasmic antibodies in idiopathic systemic vasculitis. Anti-neutrophil cytoplasmic antibodies (ANCA) are widely used as diagnostic markers for Wegener’s granulomatosis (WG), microscopic polyangiitis (MPA), Churg-Strauss syndrome (CSS) and idiopathic rapidly progressive glomerulonephritis (iRPGN). The objective of this study was to evaluate the diagnostic value of ANCA measurement by the indirect immunofluorescence (IIF) test, and by anti-PR3 and anti-MPO ELISA performed in different locations, in patients with idiopathic small vessel vasculitis. Fourteen centers participated in a standardization study of ANCA assays, and entered a total number of 169 newly diagnosed and 189 historical patients with idiopathic systemic vasculitis or iRPGN. Patients were classified according to a pre-defined diagnostic classification system. Results were compared with those of 184 disease controls and 740 healthy controls. The IIF test was performed according to standard methodology; ELISAs had been standardized among the participants in a previous phase of the study. The sensitivities of assays in patients were as follows. The sensitivity in WG was: cANCA 64%, pANCA 21%, anti-PR3 66%, anti-MPO 24%. In MPA the sensitivity was: cANCA 23%, pANCA 58%, anti-PR3 26%, anti-MPO 58%. Sensitivity in iRPGN was: cANCA 36%, pANCA 45%, anti-PR3 50%, anti-MPO 64%. The specificity of assays (related to disease controls) was: cANCA 95%, pANCA 81%, anti-PR3 87%, anti-MPO 91%. When the results of the IIF test were combined with those of the ELISAs (cANCA/anti-PR3 positive, pANCA/anti-MPO positive), the diagnostic specificity increased to 99%. The sensitivity of the combination of cANCA + anti-PR3 or pANCA + anti-MPO for WG, MPA or iRPGN was 73%, 67% and 82%, respectively. From this study we conclude that the value of the IIF test for ANCA detection can be greatly increased by the addition of a well standardized antigen-specific ELISA. In a significant number of patients with idiopathic small vessel vasculitis, however, the ANCA test results (either in IIF or ELISA) are negative.

Anti-neutrophil cytoplasmic antibodies (ANCA) are diagnostic markers for systemic vasculitis. They are classically detected by an indirect immunofluorescence test using normal donor neutrophils as substrate. This assay lacks antigenic specificity and is not quantitative. The 'EC/BCR Project for ANCA Assay Standardization' is an international collaboration study with the aim to develop and standardize solid phase assays for ANCA detection. In this part of the study the isolation and characterization of proteinase-3 and myeloperoxidase, the two main target molecules for ANCA, and the development and standardization of ELISAs with these antigens are described. Six laboratories successfully isolated purified proteinase-3 preparations that could be used. Three of these preparations, together with one myeloperoxidase preparation, were subsequently used for ANCA testing by ELISA. The ELISA technique was standardized in two rounds of testing in the 14 participating laboratories. The coefficient of variation of these new assays decreased from values of approx. 50% in the first round to approx. 20% in the second round. We conclude that purified proteinase-3 and myeloperoxidase can be used in standardized ELISAs for ANCA detection. Whether such procedures offer advantages over the IIF test will be determined in a prospective clinical study.

This study describes the results of phase I of an international effort to develop and standardize assays for the detection of anti-neutrophil cytoplasmic antibodies (ANCA). 12 sera, four of which were selected for their potential to cause problems in the detection of various ANCA specificities, were analyzed in the standard indirect immunofluorescence (IIF) test and in ELISAs for ANCA routinely performed in the seven participating laboratories. The IIF methodology differed with respect to the dilution of the serum being screened and the concentration of the conjugate used. Results from sera with high ANCA titers were similar, although the quantitative values could not be compared. In sera containing rheumatoid factor and anti-nuclear antibodies (ANA), ANCA-unrelated staining patterns were observed. Six antigen preparations were used in ELISA for the detection of cANCA. In ELISA with purified proteinase-3 all three cANCA sera were positive, but not anti-myeloperoxidase (MPO) or anti-lactoferrin (LF) positive sera. The other assays were less sensitive or gave inconsistent results. Various preparations of purified MPO and LF used in ELISA were readily recognized by anti-MPO and anti-LF positive sera. From this study it can be concluded that the IIF test, although performed with different methods, shows comparable results using strongly positive sera. In general solid phase assays for cANCA detection are not well standardized and need improvement although the purified proteinase-3 ELISA is possibly an exception. MPO and LF can be used in ELISA procedures for the detection of pANCA-related antibodies.

Systemic lupus erythematosus (SLE) is a complex, multisystem autoimmune disorder, which often involves referral to multiple medical specialists. Lupus nephritis (LN) occurs in ~35% of adults with SLE and predicts poor survival. There is currently no consensus on how to manage patients with SLE or LN across specialties and across different European countries. The Lupus Nephritis Terminology Advisory Group was formed to address this issue as it impacts upon LN treatment. It has developed consensus statements based on opinions from expert panel meetings with nephrologists, nephropathologists, rheumatologists, clinical immunologists and internal medicine specialists from many European countries, after reviewing current guidelines from the European League Against Rheumatism, the American College of Rheumatology and the participants' experience. In this article, we report consensus statements that were developed in six important areas: classification of patients with LN, how classification affects the selection of treatment options and definitions of induction, response, flare and maintenance. We have also proposed a consensus for the terminology involved in the management of LN that is consistent with clinical opinion gathered from multidisciplinary expert meetings and with existing guidelines. We believe this consensus approach provides agreed expert opinion to clinicians and will form the basis for optimising LN treatment. © 2009 SAGE Publications