The vast majority of ACPA-IgG harbour N-glycans in their variable domains. As N-linked glycosylation requires glycosylation consensus sites in the protein sequence and as these are lacking in the 'germline-counterparts' of identified variable domains, our data indicate that the N-glycosylation sites in ACPA variable domains have been introduced by somatic hypermutation. This finding also suggests that ACPA-hyperglycosylation confers a selective advantage to ACPA-producing B cells. This unique and completely novel feature of the citrulline-specific immune response in RA elucidates our understanding of the underlying B cell response.

We developed a technology to identify and isolate citrullinated antigen-specific B cells from peripheral blood of patients with RA. Most cells have a memory phenotype, express IgA or IgG and are present in relatively high frequencies. These data pave the path for a direct and detailed molecular characterisation of ACPA-expressing B cells and could lead to the identification of novel therapeutic targets.

ACPA-specific peripheral blood B cells are not confined to the CD20 positive memory pool, as circulating plasmablasts/cells spontaneously producing ACPA are also readily detectable. The latter points to an ongoing B cell immune response against citrullinated proteins and contrasts conventional immune responses against, for example, vaccines, where antigen-specific plasmablasts appear in peripheral blood only shortly after vaccination. These circulating, ACPA-specific plasmablasts/cells might represent targets for novel therapeutic interventions.

Autoreactive B cells mediate autoimmune pathology, but exactly how remains unknown. A hallmark of rheumatoid arthritis (RA), a common autoimmune disease, is the presence of disease-specific anticitrullinated protein antibodies (ACPAs). Here, we showed that ACPA-positive B cells in patients with RA strongly expressed T cell–stimulating ligands, produced abundant proinflammatory cytokines, and were proliferative while escaping inhibitory signals. This activated state was found at different degrees in different stages of disease: highest in patients with recent-onset RA, moderate in patients with established RA, and far less pronounced in ACPA-positive individuals “at risk” for developing disease. The activated autoreactive B cell response persisted in patients who achieved clinical remission with conventional treatment. ACPA-positive B cells in blood and synovial fluid secreted increased amounts of the chemoattractant interleukin-8, which attracted neutrophils, the most abundant immune cell in arthritic joints. Tetanus toxoid–specific B cells from the same patients exhibited properties of memory B cells without the activation and proliferation phenotype, but these cells transiently acquired a similar proliferative phenotype upon booster vaccination. Together, these data indicated that continuous antigenic triggering of autoreactive B cells occurs in human autoimmune disease and support the emerging concept of immunological activity that persists under treatment even in clinical remission, which may revise our current concept of treatment targets for future therapeutic interventions. In addition, our data pointed to a pathogenic role of ACPA-positive B cells in the inflammatory disease process underlying RA and favor approaches that aim at their antigen-specific inactivation or depletion.

Objective Anti–citrullinated protein antibodies (ACPAs) are a hallmark of rheumatoid arthritis (RA). Aside from autoantibody production, the function of autoantigen‐specific B cells remains poorly understood in the context of this disease. This study set out to elucidate autoantigen‐specific B cell functions through the isolation and immortalization of unique citrullinated protein/peptide (CP)–reactive B cell clones from RA patients. Methods B cell clones from either the blood or synovial fluid of cyclic citrullinated peptide 2 (CCP2) antibody–positive RA patients were immortalized by genetic reprogramming with Bcl‐6 and Bcl‐xL. Enzyme‐linked immunosorbent assay and flow cytometry were used to identify CCP2‐reactive clones and to further characterize surface marker and cytokine expression as well as B cell receptor signaling competence. Global gene expression profiles were interrogated by RNA sequencing. Results Three unique CP‐reactive memory B cell clones were generated from the blood or synovial fluid of 2 RA patients. CP‐reactive memory B cells did not appear to be broadly cross‐reactive, but rather had a fairly restricted epitope recognition profile. These clones were able to secrete both pro‐ and antiinflammatory cytokines and had a unique surface profile of costimulatory molecules and receptors, including CD40 and C5a receptor type 1, when compared to non‐CP–reactive clones from the same patient. In addition, CP‐reactive clones bound citrullinated protein, but not native protein, and could mobilize calcium in response to antigen binding. Conclusion CP‐reactive memory B cells comprise a rare, seemingly oligoclonal population with restricted epitope specificity and distinct phenotypic and molecular characteristics suggestive of antigen‐presenting cells. Cloning by genetic reprogramming opens new avenues to study the function of autoreactive memory B cells, especially in terms of antigen processing, presentation, and subsequent T cell polarization.

This study demonstrates a high frequency of differentiated, spontaneously ACPA-IgG-secreting cells in SF. These cells are supported by SFMC for prolonged survival and autoantibody secretion, demonstrating that the synovial compartment is equipped to function as inflammatory niche for PC survival.

Highlights d MAIT cells are activated in individuals with hemorrhagic fever with renal syndrome (HFRS) d MAIT cell activation correlates with HFRS severity markers during hantavirus infection d MAIT cell blood levels decline during acute HFRS d Hantavirus-mediated MAIT cell activation is type I IFN dependent

Human hantavirus infections can cause hemorrhagic fever with renal syndrome (HFRS), major signs of the disease being thrombocytopenia and transient kidney dysfunction. By a comprehensive and longitudinal study of circulating B cells, we demonstrate that these two pathologies associate with distinct effects on the humoral immune system during HFRS. Low thrombocyte counts strongly associated with an abnormal frequency of plasmablasts in circulation, whereas kidney dysfunction was indicative of an accumulation of CD27 -B cells and plasmablasts. Finally, we provide evidence that high levels of extracellular ATP in circulation during HFRS correlates with shedding of surface CD27 on B cells via a metallomatrix proteinase-8-mediated mechanism. Since extracellular ATP is known to regulate kidney function, our study reveals a link between kidney dysfunction and the generation of CD27 -IgD -B cells, and a potential molecular target for treatment of the symptomatic phase of HFRS. Dobrava, Seol and Puumala (PUUV) strains, where PUUV is endemic to Scandinavia. To date, no efficacious treatment or vaccination regimen exists to protect against severe hantavirus infections.Well controlled human studies have shown that hantavirus infections cause aberrant activation of both innate and adaptive immunity (5)(6)(7)(8). A potent antiviral IgG-response is associated with protection from severe disease during both HPS and HFRS (9-12) and passive transfer of serum antibodies could reduce case fatality rate in a small cohort of HPS patients (13). These findings clearly demonstrate that activation of the humoral immune system and subsequent elicitation of antiviral antibodies play a central role in the control of viremia and/or pathogenesis during hantavirus infections.A recent study of HPS demonstrated that very high levels of plasmablasts (PBs) and CD27 -IgD -B cells were detected in circulation of patients (14). The rapid and extensive PB-response is similar to that reported during acute dengue virus infection and contrasts to the comparably moderate levels of PBs that are found in circulation during influenza infection or after influenza vaccination (15). An expansion of the CD27 -IgD -B cell subset has previously been shown for numerous inflammatory and infectious diseases, as well as during ageing and cancer (16-21), yet their functional role in humoral immunity remains undetermined. The CD27 -IgD -B cells resemble memory B cells, have isotype switched and hypermutated B cell receptors and therefore likely originate from T cell-dependent germinal center reactions in secondary lymphoid organs (16, 20). In systemic lupus erythematosus (SLE), an expanded population of CD27 -IgD -B cells was associated with the manifestation of nephritis in patients (16). This suggests that their detection in blood may be linked to reduced kidney function, but the cause for their expansion remains to be determined.During HFRS-causing hantavirus infections, reduced kidney function occurs independently of the induced thrombocytopenia (22, 23). We therefor...