François Tron

Correlation of anti–signal recognition particle autoantibody levels with creatine kinase activity in patients with necrotizing myopathy

OBJECTIVEnAnti-signal recognition particle (anti-SRP) autoantibodies are associated with severe acquired necrotizing myopathies. The role of these autoantibodies remains elusive, and the evolution of anti-SRP levels over time is unknown. In this study, we developed an addressable laser bead immunoassay (ALBIA) technique to investigate a correlation between anti-SRP levels, serum creatine kinase (CK) levels, and muscle strength in patients with necrotizing myopathy.nnnMETHODSnThe diagnostic value of the ALBIA assay was determined by comparing serum levels of anti-SRP autoantibodies in 31 anti-SRP immunodot-positive patients to those in 190 healthy blood donors and 199 control patients with different inflammatory/autoimmune conditions or polyclonal hypergammaglobulinemia. Among the 31 anti-SRP-positive patients, serum samples from 8 patients were monitored over time for levels of anti-SRP autoantibodies and levels of CK (determined at least 3 times, consecutively, over a mean followup period of 783 days). The relationship between levels of anti-SRP autoantibodies and levels of CK was tested using a linear mixed model.nnnRESULTSnThe assay yielded positive results for anti-SRP in all anti-SRP immunodot-positive serum samples tested, while all control sera tested negative. The 8 anti-SRP-positive patients who were followed up longitudinally were found to have normalized CK levels and improved muscle strength. There was a striking correlation between the degree of myolysis, as measured by CK levels, in patients receiving therapy and the anti-SRP54 autoantibody levels in these same patients (P = 0.002).nnnCONCLUSIONnAnti-SRP-positive myositis appears to be one of the few autoimmune diseases in which specific autoantibody levels are correlated with surrogate disease activity markers. These results reveal the usefulness of monitoring anti-SRP autoantibody levels in patients receiving therapy, and may also suggest a possible pathogenic role for anti-SRP autoantibodies in the necrotizing myopathies.

Candidate autoantigens identified by mass spectrometry in early rheumatoid arthritis are chaperones and citrullinated glycolytic enzymes

IntroductionThe aim of our study was to identify new early rheumatoid arthritis (RA) autoantibodies.MethodsSera obtained from 110 early untreated RA patients (<6 months) were analyzed by western blot using HL-60 cell extract, separated on one-dimensional and two-dimensional gel electrophoresis (1-DE, 2-DE). Sera from 50 healthy blood donors and 20 patients with non-RA rheumatisms were used as controls for 1-DE and 2-DE, respectively. The immunoreactive proteins were identified by MALDI-TOF mass spectrometric analysis and the presence of potential sites of citrullination in each of these proteins was evaluated. FT-ICR mass spectrometry was used to verify experimentally the effect of citrullination upon the mass profile observed by MALDI-TOF analysis.ResultsThe 110 1-DE patterns allowed detection of 10 recurrent immunoreactive bands of 33, 39, 43, 46, 51, 54, 58, 62, 67 and 70 kDa, which were further characterized by 2-DE and proteomic analysis. Six proteins were already described RA antigens: heterogeneous nuclear ribonucleoprotein A2/B1, aldolase, α-enolase, calreticulin, 60 kDa heat shock protein (HSP60) and BiP. Phosphoglycerate kinase 1 (PGK1), stress-induced phosphoprotein 1 and the far upstream element-binding proteins (FUSE-BP) 1 and 2 were identified as new antigens. Post-translational protein modifications were analyzed and potentially deiminated peptides were found on aldolase, α-enolase, PGK1, calreticulin, HSP60 and the FUSE-BPs. We compared the reactivity of RA sera with citrullinated and noncitrullinated α-enolase and FUSE-BP linear peptides, and showed that antigenicity of the FUSE-BP peptide was highly dependent on citrullination. Interestingly, the anti-cyclic citrullinated peptide antibody (anti-CCP2) status in RA serum at inclusion was not correlated to the reactivity directed against FUSE-BP citrullinated peptide.ConclusionsTwo categories of antigens, enzymes of the glycolytic family and molecular chaperones are also targeted by the early untreated RA autoantibody response. For some of them, and notably the FUSE-BPs, citrullination is involved in the immunological tolerance breakdown observed earlier in RA patients. Autoantibodies recognizing a citrullinated peptide from FUSE-BP may enhance the sensibility for RA of the currently available anti-CCP2 test.

Critical Role of TLR2 and TLR4 in Autoantibody Production and Glomerulonephritis in lpr Mutation-Induced Mouse Lupus

Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by pathogenic autoantibodies directed against nuclear Ags and immune complex deposits in damaged organs. Environmental factors have been thought to play a role in the onset of the disease. The recognition of these factors is mediated by TLRs, in particular TLR2 and TLR4 which bind pathogen-associated molecular patterns of Gram+ and Gram− bacteria, respectively. We attempted to determine the role of these TLRs in SLE by creating TLR2- or TLR4-deficient C57BL/6lpr/lpr mice. These mice developed a less severe disease and fewer immunological alterations. Indeed, in C57BL/6lpr/lpr-TLR2 or -TLR4-deficient mice, glomerular IgG deposits and mesangial cell proliferation were dramatically decreased and antinuclear, anti-dsDNA, and anti-cardiolipin autoantibody titers were significantly reduced. However, the response against nucleosome remained unaffected, indicating a role of TLR2 and TLR4 in the production of Abs directed against only certain categories of SLE-related autoantigens. Analysis of B cell phenotype showed a significant reduction of marginal zone B cells, particularly in C57BL/6lpr/lpr-TLR4-deficient mice, suggesting an important role of TLR4 in the sustained activation of these cells likely involved in autoantibody production. Interestingly, the lack of TLR4 also affected the production of cytokines involved in the development of lupus disease.

ELISA testing of anti-desmoglein 1 and 3 antibodies in the management of pemphigus.

OBJECTIVEnTo assess the predictive value of anti-desmoglein (Dsg) 1 and anti-Dsg3 antibody (Ab) enzyme-linked immunosorbent assay (ELISA) values for the occurrence of relapses in pemphigus.nnnDESIGNnRetrospective study.nnnSETTINGnDermatology departments from 13 university hospitals in France. Patients The study population comprised 26 patients with typical clinical, histologic, and immunofluorescence findings of pemphigus, who were followed up over a 17-month period.nnnMAIN OUTCOME MEASURESnSerial anti-Dsg1 and anti-Dsg3 Ab ELISA values were recorded during the patients follow-up examinations and correlated with the occurrence of skin and/or mucosal relapses.nnnRESULTSnA significant reduction of anti-Dsg1 (P < .001) and anti-Dsg3 (P < .001) Ab ELISA values was observed in serum samples from patients with pemphigus foliaceus or pemphigus vulgaris after the initial treatment. During the long-term follow-up, anti-Dsg1 Ab ELISA values correlated with the course of skin lesions (P = .03); the 20 U/mL cutoff for the anti-Dsg1 Ab ELISA value provided a 79% positive and an 84% negative predictive value for the occurrence of cutaneous relapses. No correlation was observed between anti-Dsg3 Ab ELISA values and the course of mucosal lesions (P = .13). Anti-Dsg3 Ab ELISA values higher than the 14-U/mL cutoff were observed in 5 of the 5 patients with relapse and in 10 of the 13 patients with ongoing mucosal remission, providing a 100% sensitivity but a poor specificity of 23%. A cutoff value of 130 U/mL for anti-Dsg3 Abs was calculated based on the receiver operating characteristics curve and provided an 84% positive and an 81% negative predictive value.nnnCONCLUSIONSnAnti-Dsg1 Ab ELISA values are more closely correlated than anti-Dsg3 Ab ELISA values with the course of the disease in patients with pemphigus vulgaris or pemphigus foliaceus. This should be taken into account for the management of patients with pemphigus.

Clinical significance of autoantibodies recognizing Sjögren's syndrome A (SSA), SSB, calpastatin and alpha-fodrin in primary Sjögren's syndrome

The aim of our study was (i) to compare the clinical and biological characteristics of 148 (137 women, 11 men) primary Sjögrens syndrome (pSS) patients at diagnosis as a function of their sex and (ii) to assess the prognostic value of anti‐calpastatin and anti‐alpha‐fodrin autoantibodies. In addition, the presence of anti‐nuclear antibodies (ANA), anti‐52‐ and 60‐kDa Sjögrens syndrome A (SSA), anti‐Sjögrens syndrome B (SSB), anti‐cyclic citrullinated peptide (CCP) antibodies and rheumatoid factors (RF) of IgA, IgG and IgM isotypes was sought in sera collected at pSS onset. Raynauds syndrome, significantly more frequent in women, was the only systemic manifestation of pSS whose frequency differed significantly as a function of the patients sex (Pu2003=u20030·02). ANA (Pu2003=u20030·001) and anti‐60‐kDa SSA autoantibodies (Pu2003=u20030·03) were significantly more common in women, while men never synthesized detectable levels of anti‐SSB, anti‐calpastatin or IgG anti‐alpha‐fodrin autoantibodies. In addition, anti‐CCP autoantibodies were found in low percentages of pSS patients (4% F/18% M). The absence of autoantibodies does not exclude the diagnosis of pSS in men that will be based mainly on the anatomopathological findings of a minor salivary gland biopsy. Positivity of anti‐60‐kDa SSA, anti‐SSB, anti‐calpastatin, IgA and IgG anti‐alpha‐fodrin antibodies is not associated with pSS clinical and biological severity.

Impact of Three Anti-TNFα Biologics on Existing and Emergent Autoimmunity in Rheumatoid Arthritis and Spondylarthropathy Patients

ObjectiveThe objective of this study was to analyze the effects of 3 anti-TNFα agents on markers of autoimmunity in rheumatoid arthritis (RA) and spondylarthropathy (SPA) patients.MethodsFirst-time anti-TNFα biologics (infliximab, etanercept, or adalimumab) were prescribed to 156 RA and 95 SPA (58 ankylosing spondylarthritides, 37 psoriatic arthritides). During 1–2xa0years of follow-up, clinical, biological [antinuclear (ANA) and anti-double-stranded (dsDNA) antibodies, rheumatoid factors (RF), and anti-cyclic citrullinated peptide (CCP) for RA], and therapeutic data were collected biannually.ResultsANA appeared or ANA and anti-dsDNA titers increased significantly (Pu2009<u20090.001) more under infliximab than etanercept in both rheumatisms and than adalimumab in RA patients. During the 2-year follow-up, ANA appeared more in RA patients taking adalimumab than etanercept (Pu2009=u20090.003), but independently of the anti-TNFα used; anti-dsDNA titers rarely became positive. Under etanercept or infliximab, ANA and anti-dsDNA were not influenced by the underlying pathology nor were they affected by infliximab intensification over 18xa0months. Only one case of cutaneous lupus was observed in a patient having IgG anti-dsDNA. The therapeutic responses were independent of ANA and anti-dsDNA titers for all rheumatisms and biologics. In RA patients, RF titers, but not anti-CCP levels, declined with the therapeutic response for all biologics.ConclusionThis is the first study that has evaluated the impact of three TNFα blockers on ANA and anti-dsDNA antibodies in RA and SPA patients. Autoimmunity was more induced with infliximab than etanercept and to a lesser degree to adalimumab but, more importantly, this emergent autoimmunity was exceptionally associated to clinical manifestations of lupus.

Taenia crassiceps invasive cysticercosis: a new human pathogen in acquired immunodeficiency syndrome?

A fluctuant, painful, subcutaneous, and intermuscular tumor developed in a 38-year-old man with severe acquired immunodeficiency syndrome (AIDS) in which immunodeficiency was severe. Surgery revealed lesions that formed a multilocular pouch embedded in deep tissues in the forearm filled with tapiocalike material containing a viscous fluid, granules, and cysticercilike small vesicles. Pathologic and parasitologic evaluation showed cysticerci embedded in a fibrocollagen reaction with inflammatory granulomatous reaction. Each cysticercus contained an invaginated scolex with two rows of small (i.e., 80 microm) and large (i.e., 114 microm) rostellar hooks, identical to larva of Taenia crassiceps. All clinical, parasitologic, and pathologic features of these cysticerci were very different from those of all other larval cestode (i.e., Taenia solium cysticercosis, coenurosis, sparganosis, cysticercosis due to Taenia saginata [Cysticercus bovis], primary and secondary hydatidosis [Echinococcus species]). T crassiceps cysticerci usually develop in subcutis and pleuroperitoneal cavities of rodents, whereas the adult tapeworm is commonly found in the digestive tract of foxes. Biologic properties of T crassiceps cysticerci and epidemiologic characteristics of pandemic human immunodeficiency virus (HIV) could eventually indicate new potential cases of T crassiceps cysticercosis in humans.

Contribution of PTPN22 1858T, TNFRII 196R and HLA-shared epitope alleles with rheumatoid factor and anti-citrullinated protein antibodies to very early rheumatoid arthritis diagnosis

OBJECTIVESnTo evaluate the predictive value of TNFRII 196R, PTPN22 1858T and HLA-shared epitope (SE) alleles, RFs and anti-citrullinated protein antibodies (ACPAs) for RA diagnosis in a cohort of patients with very early arthritis.nnnMETHODSnWe followed up 284 patients who had swelling of at least two joints that had persisted for longer than 4 weeks but had been evolving for <6 months. At 2 yrs, patients were classified as having RA or non-RA rheumatic diseases according to the ACR criteria. Patients were genotyped with respect to TNFRII 196M/R and PTPN22 1858C/T polymorphisms and HLA-SE. The presence of IgA, IgG and IgM RF isotypes and ACPA was sought in sera collected at disease onset.nnnRESULTSnHLA-SE alleles alone, concomitant presence of TNFRII 196R and PTPN22 1858T alleles, IgA, IgG and IgM RF alone and ACPA were found to be significantly associated with RA diagnosis. Using logistic regression analysis, the concomitant presence of RF and ACPA at disease onset was the best association to predict RA diagnosis. In patients (n = 34) who did not fulfil the ACR criteria for RA at inclusion but who progressed to ACR positivity, the study of the genetic risk markers did not contribute to predict RA diagnosis at 2 yrs.nnnCONCLUSIONSnPTPN22 1858T, TNFRII 196R and HLA-SE alleles do not improve the predictive value of RF and ACPA for RA diagnosis in our cohort, and do not contribute to an earlier diagnosis in undifferentiated patients initially negative for RF and ACPA.

Can rheumatoid arthritis responsiveness to methotrexate and biologics be predicted

This review briefly recapitulates the existing markers predictive of RA responsiveness to treatment, focusing on MTX alone or combined with a biologic. In addition to the demographic and clinical factors, an update is provided of the predictive biomarkers identified by large-scale gene and protein analyses that generated new insights into the ability of high-throughput analysis of biological systems to select new potential indicators. Among the large-scale analysis tools now available, pharmacogenetics and pharmacogenomics (including transcriptomic and proteomic approaches) have been shown to provide such new putative biomarkers of therapeutic responses.

Binding of autoantibodies is not restricted to desmosomes in pemphigus vulgaris

Pemphigus vulgaris (PV) and pemphigus foliaceus (PF) are autoimmune blistering diseases characterized by a loss of cell-cell adhesion and by autoantibodies directed against epidermal cadherins. PF antigen has been established as desmoglein I which is located strictly on the desmosome, whereas the precise ultrastructural localization of PV antigen remains unclear and controversial to date. To further investigate this question, we compared the location of immune deposits in 14 patients with PV and 10 patients with PF by both direct and indirect immunoelectron microscopy (IEM). Inclusion criteria were based upon clinical features, histological level of cleavage and characterization of circulating antibodies by Western blot on epithelial bovine tongue extracts. IEM was performed on unfixed 0.7-mm slices of skin for the direct technique or on normal skin for the indirect technique using peroxidase labelling. In PF, by both direct and indirect IEM, immune deposits were located on the extracellular part of desmosomes (desmoglea) in all the samples studied. In PV, by both direct and indirect IEM, deposits were situated on the desmoglea and along large portions of the keratinocyte membrane without desmosomal structures in 15 of the 18 samples studied and only on the desmoglea in 3 samples. These results suggest that, in contrast to PF, the target antigen in PV is not always restricted to desmosomes. As various types of adherens junctions have been reported to mediate cell adhesion in the epidermis, the PV antigen could be a component of desmosomes and of other focal adhesions.