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Background: Children with systemic lupus erythematosus (SLE) frequently have kidney involvement. Lupus nephritis sometimes presents alone, without systemic SLE features, representing the so-called full-house nephropathy (FHN). Distinguishing patients with SLE or FHN has therapeutic and prognostic implications., Methods: In this retrospective observational study, we determined the presence of IgM on the surface of T cells (T cell IgM) by flow cytometry and characterized its ability in distinguishing SLE and FHN patients in a large pediatric cohort (n = 84). Fifty-seven patients with SLE (≥ 4 SLICC criteria at disease onset or during the follow-up) and 27 patients with FHN (3 or less SLICC criteria) were enrolled., Results: Elevated T cell IgM levels were found in 24/25 SLE patients in active phase of disease and in 29/45 SLE patients in remission. In contrast, among FHN patients, only 1/9 presented this characteristic in active phase of disease and 0/20 in remission. Compared with standardized SLICC laboratory parameters, i.e., autoantibody titers and hypocomplementemia, T cell IgM positivity showed an extremely high sensitivity and specificity for the diagnosis of SLE, with the highest area under the curve (0.97, p < 0.001) by receiver operating characteristic analysis, similar to ANA (0.96, p < 0.001) and anti-dsDNA (0.90, p < 0.001) autoantibodies., Conclusions: Altogether, our data indicate that T cell IgM intensity may be a useful tool to correctly classify patients with lupus nephritis as SLE or FHN since disease onset.

SARS-CoV-2 is a novel coronavirus, not encountered before by humans. The wide spectrum of clinical expression of SARS-CoV-2 illness suggests that individual immune responses to SARS-CoV-2 play a crucial role in determining the clinical course after first infection. Immunological studies have focused on patients with moderate to severe disease, demonstrating excessive inflammation in tissues and organ damage. In order to understand the basis of the protective immune response in COVID-19, we performed a longitudinal follow-up, flow-cytometric and serological analysis of innate and adaptive immunity in 64 adults with a spectrum of clinical presentations: 28 healthy SARS-CoV-2-negative contacts of COVID-19 cases; 20 asymptomatic SARS-CoV-2-infected cases; eight patients with Mild COVID-19 disease and eight cases of Severe COVID-19 disease. Our data show that high frequency of NK cells and early and transient increase of specific IgA, IgM and, to a lower extent, IgG are associated with asymptomatic SARS-CoV-2 infection. By contrast, monocyte expansion and high and persistent levels of IgA and IgG, produced relatively late in the course of the infection, characterize severe disease. Modest increase of monocytes and different kinetics of antibodies are detected in mild COVID-19. The importance of innate NK cells and the short-lived antibody response of asymptomatic individuals and patients with mild disease suggest that only severe COVID-19 may result in protective memory established by the adaptive immune response., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2020 Carsetti, Zaffina, Piano Mortari, Terreri, Corrente, Capponi, Palomba, Mirabella, Cascioli, Palange, Cuccaro, Milito, Zumla, Maeurer, Camisa, Vinci, Santoro, Cimini, Marchioni, Nicastri, Palmieri, Agrati, Ippolito, Porzio, Concato, Onetti Muda, Raponi, Quintarelli, Quinti and Locatelli.)

The clinical heterogeneity of idiopathic nephrotic syndrome in childhood may reflect different mechanisms of disease that are as yet unclear. Here, we evaluated the association between an atypical presence of IgM on the surface of T cells (T-cell IgM) and the response to steroid therapy in a total of 153 pediatric patients with idiopathic nephrotic syndrome in different phases of disease. At disease onset, T-cell IgM median levels were significantly elevated and predictive of risk of relapse in 47 patients. They were also significantly increased comparing 58 steroid-dependent to 8 infrequently relapsing and 14 frequently relapsing patients, especially during relapse, whereas they were within the normal range in 7 genetic steroid-resistant patients. T-cell IgM in vivo was not affected by the amount of total circulating IgM, nor by concomitant acute infections or oral immunosuppression. However, it was affected by rituximab treatment in 21 steroid-dependent patients. By in vitro experiments, elevated T-cell IgM was not influenced by total circulating IgM levels or by the presence of other circulating factors, and there was no distinctive antigen-specificity or atypical IgM polymerization. Rather, we found that increased T-cell IgM correlates with reduced IgM sialylation, which influences T-cell response to steroid inhibition and T-cell production of podocyte-damaging factors. Thus, the atypical presence of IgM on the surface of T cells may predispose a subset of steroid-sensitive pediatric patients with idiopathic nephrotic syndrome to a poor response to steroid therapy since disease onset., (Copyright © 2019 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

Children with Down syndrome (DS) have increased susceptibility to infections and a high frequency of leukemia and autoimmune disorders, suggesting that immunodeficiency and immune dysfunction are integral parts of the syndrome. A reduction in B-cell numbers has been reported, associated with moderate immunodeficiency and normal immunoglobulin levels. Here, we compared B-cell populations of 19 children with DS with those in healthy age-matched controls. We found that all steps of peripheral B-cell development are altered in DS, with a more severe defect during the later stages of B-cell development. Transitional and mature-naïve B-cell numbers are reduced by 50% whereas switched memory B cells represent 10-15% of the numbers in age-matched controls. Serum IgM levels were slightly reduced, but all other immunoglobulin isotypes were in the normal range. The frequency of switched memory B cells specific for vaccine antigens was significantly lower in affected children than in their equivalently vaccinated siblings. In vitro switched memory B cells of patients with DS have an increased ability to differentiate into antibody-forming cells in response to TLR9 signals. Tailored vaccination schedules increasing the number of switched memory B cells may improve protection and reduce the risk of death from infection in DS., (© 2014 The Authors. European Journal of Immunology published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Immunological memory is our reservoir of ready-to-use antibodies and memory B cells. Because of immunological memory a secondary infection will be very light or not occur at all. Antibodies and cells, generated in the germinal center in response to the first encounter with antigen, are highly specific, remain in the organism virtually forever and are mostly of IgG isotype. Long lived plasma cells homing to the bone marrow ensure the constant production of protective antibodies, whereas switched memory B cells proliferate and differentiate in response to secondary challenge. IgM memory B cells represent our first-line defense against infections. They are generated by a T-cell independent mechanism probably triggered by Toll-like receptor-9. They produce natural antibodies with anti-bacterial specificity and the spleen is indispensable for their maintenance. We will review the characteristics and functions of IgM memory B cells that explain their importance in the immediate protection from pathogens. IgM memory B cells, similar to mouse B-1a B cells, may be a remnant of a primitive immune system that developed in the spleen of cartilaginous fish and persisted throughout evolution notwithstanding the sophisticated tools of the adaptive immune system., (Copyright © 2013 Elsevier B.V. All rights reserved.)

A young woman presenting respiratory infections, polyarthritis, severe neutropenia, and increased serum IgM was treated with intravenous immunoglobulin (IVIG) with good clinical and laboratory outcome followed by a loss of efficacy. The increased serum IgM associated to recurrent infections and autoimmune manifestations suggested the diagnosis of a hyper-IgM syndrome (HIGMs). The frequency of peripheral T cells, the expression of CD40 on the patients' B cells and CD40L on T cells and the activation-induced cytidine deaminase (AID) and uracil-DNA glycosylase (UNG) at mRNA level was comparable to controls. In contrast, the frequency of B cells was one half of the healthy control and all cells showed an atypical phenotype. Although AID and UNG were normal, class-switch recombination was not very efficient because circulating switched memory were reduced and, once stimulated with CpG, generated less antibody-secreting cells than controls. An increase in serum B Lymphocytes stimulator (BLyS) was also found. The patient presented a peculiar clinical and immunological phenotype fitting for many aspects of both HIGM4 and Common Variable Immunodeficiency (CVID). These findings underline the need to better explore the complex link between these two diseases.

TLR9 activation by unmethylated CpG provides a homeostatic mechanism to maintain B cell memory in the absence of Ag. In this study, we demonstrate that CpG also triggers the generation of somatically mutated memory B cells from immature transitional B cells. In response to CpG, a fraction of transitional B cells proliferates and introduces somatic hypermutations in the H chain V regions. The nonproliferating pool of transitional B cells mostly maintains germline configurations. Mutations are VH specific: VH5 is the least mutated family, whereas VH1 and VH4/6 are the most mutated families. CpG stimulation also results in upregulation of VH5 transcripts in proliferating cells. Therefore, early recognition of bacterial DNA preferentially expands VH5-expressing B cells while inducing somatic hypermutations in other families. The mutation frequency, range, and type of substitutions observed in vitro are comparable to those found in memory B cells from the peripheral blood of Hyper IgM type 1 patients and the spleen of normal infants. The process triggered by TLRs may represent a first step leading to additional diversification of the germline repertoire and to the generation of memory B cells that will further refine their repertoire and specificity in the germinal centers.

Background: Under experimental chronic inflammation, tumor necrosis factor (TNF)-alpha plays a role in damaging spleen marginal zone. This latter has a crucial function in mounting B cell-dependent immune responses against infections by encapsulated bacteria. In Crohn's disease (CD), a chronic inflammatory disorder where TNF-alpha is centrally involved, impaired splenic function may increase the susceptibility to bacterial infections. On this basis, we aimed to investigate the influence of anti-TNF therapy on splenic function in CD patients., Methods: Peripheral blood samples were obtained from 15 CD patients before and after treatment with infliximab administered at weeks 0, 2, and 6 at a dose of 5 mg/kg. Counting of erythrocytes with membrane abnormalities (pitted red cells) was used as an indicator of splenic function. Multicolor flow cytometry was performed to analyze circulating B cells., Results: A substantial clinical improvement in 10 of the 15 CD patients was associated with a significant reduction of pitted red cells (from median 6.0% to 3.6%; P < 0.01) after 10 weeks of treatment. In responder patients the improvement of splenic function was accompanied by a parallel increase of circulating IgM-memory B cells (from median 6.9% to 13.3%; P < 0.005). Splenic function was not ameliorated in nonresponder patients., Conclusions: Splenic function improved in CD patients who responded to infliximab and was accompanied by a concomitant restoration of the IgM-memory B cell pool responsible for the protection against encapsulated bacteria. Restoration of splenic function after infliximab treatment is intriguing and requires further investigation.

Background: The hemagglutinin (HA) glycoprotein is the principal target of protective humoral immune responses to influenza virus infections but such antibody responses only provide efficient protection against a narrow spectrum of HA antigenic variants within a given virus subtype. Avian influenza viruses such as H5N1 are currently panzootic and pose a pandemic threat. These viruses are antigenically diverse and protective strategies need to cross protect against diverse viral clades. Furthermore, there are 16 different HA subtypes and no certainty the next pandemic will be caused by an H5 subtype, thus it is important to develop prophylactic and therapeutic interventions that provide heterosubtypic protection., Methods and Findings: Here we describe a panel of 13 monoclonal antibodies (mAbs) recovered from combinatorial display libraries that were constructed from human IgM(+) memory B cells of recent (seasonal) influenza vaccinees. The mAbs have broad heterosubtypic neutralizing activity against antigenically diverse H1, H2, H5, H6, H8 and H9 influenza subtypes. Restriction to variable heavy chain gene IGHV1-69 in the high affinity mAb panel was associated with binding to a conserved hydrophobic pocket in the stem domain of HA. The most potent antibody (CR6261) was protective in mice when given before and after lethal H5N1 or H1N1 challenge., Conclusions: The human monoclonal CR6261 described in this study could be developed for use as a broad spectrum agent for prophylaxis or treatment of human or avian influenza infections without prior strain characterization. Moreover, the CR6261 epitope could be applied in targeted vaccine strategies or in the design of novel antivirals. Finally our approach of screening the IgM(+) memory repertoire could be applied to identify conserved and functionally relevant targets on other rapidly evolving pathogens.

HyperIgM syndrome is a heterogenous immunodeficiency characterized by impaired class-switch recombination due to different molecular abnormalities. We report on two female patients affected by a novel syndrome associating HIGM, growth and pubertal disturbances, and severe lymphoid hyperplasia with eventual development into lymphomas, suggesting a DNA repair defect.

Objectives: IgM memory B cells that are responsible for the protection against infections by encapsulated bacteria, require the spleen for their generation and/or survival. Since the association between inflammatory bowel disease and functional hyposplenism is well described, our aim was to verify whether IgM memory B cells mirror the reduced splenic function in Crohn's disease and ulcerative colitis patients., Methods: Peripheral blood samples were obtained from 32 Crohn's disease and 29 ulcerative colitis patients, 33 healthy controls, and 27 splenectomized patients. Perendoscopic intestinal biopsies were also collected from 15 of 32 Crohn's disease patients, 14 of 29 ulcerative colitis patients and 13 of 33 control subjects. Counting of erythrocytes with membrane abnormalities (pitted red cells) was used as an indicator of splenic function and flow cytometry was performed to analyze both peripheral and mucosal B cells., Results: Twelve of 32 Crohn's disease patients and 13 of 29 ulcerative colitis patients had pitted red cell values >4% and were considered to be hyposplenic. In inflammatory bowel disease patients circulating IgM memory B cells were significantly lower than in control subjects. We observed a significant inverse correlation between the frequency of circulating IgM memory B cell and the pitted red cell values in inflammatory bowel disease patients with hyposplenism. To exclude the possibility that the reduction of circulating IgM memory B cells reflected their recruitment in the inflamed bowel mucosa, lamina propria B-cell populations were also characterized. We found that the frequency of IgM memory B cells was similar in the blood and in the lamina propria of the same patient., Conclusions: Our findings show that peripheral IgM memory B cells are reduced in inflammatory bowel disease patients and this defect seems to be related to the impairment of splenic function.

Background: Recurrent lower respiratory tract infections caused by encapsulated bacteria might cause permanent organ damage in patients with common variable immunodeficiency (CVID). Despite the profound hypogammaglobulinemia, some patients do not experience bacterial pneumonia. We have shown that IgM memory B cells and natural antibodies play an important role in the defense against encapsulated bacteria., Objective: In this study we addressed the question of whether the apparent paradox of patients with severe hypogammaglobulinemia but no increased frequency of respiratory infections can be explained by the presence of IgM memory B cells and anti-pneumococcal polysaccharide (anti-PnPS) IgM., Methods: We measured the frequency of memory B cells and the levels of anti-PnPS IgM antibodies in 26 patients with CVID with recurrent bacterial pneumonia and bronchiectasis (group 1) and 22 who never had pneumonia and showed no lung lesions (group 2). An additional 6 patients had a clinical history of recurrent pneumonia without lung abnormalities at computed tomographic scanning., Results: Patients of group 1 lacked IgM memory B cells and failed to produce anti-PnPS IgM antibodies, and those of group 2 had a normal frequency of IgM memory B cells and produced anti-PnPS IgM antibodies., Conclusions: IgM memory B cells and anti-PnPS IgM antibodies protect patients with CVID from bacterial pneumonia. Evaluation of these 2 parameters discriminates patients with low or high risk of recurrent infections caused by encapsulated bacteria and low or high risk of bronchiectasis. Identification of high-risk individuals at diagnosis might help in the planning of a more effective therapeutic strategy and prevent permanent organ damage.

Background & Objectives: Memory B cells represent 30-60% of the B cell pool and can be subdivided in IgM memory and switched memory. IgM memory B cells differ from switched because they express IgM and their frequency may vary from 20-50% of the total memory pool. Switched memory express IgG, IgA or IgE and lack surface expression of IgM and IgD. Switched memory B cells derive from the germinal centres, whereas IgM memory B cells, which require the spleen for their survival and/or generation, are involved in the immune response to encapsulated bacteria. Since infections are one of the most frequent comorbid conditions in inflammatory bowel disease, we aimed to verify whether IgM memory B cell pool was decreased in Crohn's disease and ulcerative colitis patients., Patients & Methods: Peripheral blood samples were obtained from 22 Crohn's disease patients, 20 ulcerative colitis patients, 22 healthy controls and 18 splenectomized patients. To analyse peripheral blood lymphocytes, flow cytometry was performed using anti-CD19, anti-CD22, anti-CD27, anti-IgM, anti-IgD and anti-CD38 monoclonal antibodies., Results: Circulating IgM memory B cells were significantly lower in Crohn's disease (median 7.1%, range 1.8-20.7) and ulcerative colitis patients (median 8.1%, range 2.1-18.8) in comparison to control subjects (median 14.0%, range 6.8-31.1). As expected, there was a highly significant difference in the proportion of IgM memory B cells between splenectomized patients (median 2.4%, range 0.9-6.9) and healthy controls. Crohn's disease patients with abscesses showed the lowest frequency of IgM memory B cells., Discussion: Our findings show that peripheral IgM memory B cells are reduced in inflammatory bowel disease patients. Further studies are necessary to answer the question of whether high risk of infection (abscess development) is promoted by the reduction/depletion of IgM memory B-cell pool in inflammatory bowel disease.

Splenectomized and asplenic patients have a high incidence of infections by encapsulated bacteria and do not respond to polysaccharide vaccines. To understand whether the absence of the spleen is associated with a defined B cell defect, we analyzed B cell subsets in the peripheral blood. We found that a population of B cells known as immunoglobulin (Ig)M memory is lacking in patients without spleen. The absence of IgM memory B cells correlates with an impaired immune response to encapsulated bacteria not only in splenectomized patients, but also in individuals with an intact spleen. We show that the physiological and transient predisposition to pneumococcal infections of young children (0-2 yr) is associated with the lack of circulating IgM memory B cells and of serum antipolysaccharide IgM. We also demonstrate that IgM memory B cells are undetectable in a fraction of patients with common variable immunodeficiency, who have recurrent and invasive infections by encapsulated bacteria. IgM memory B cells, therefore, require the spleen for their generation and/or survival and are responsible for the protection against encapsulated bacteria.

CD45 expression is essential for immunoglobulin (Ig)-mediated B cell activation. Treatments with either anti-Ig or anti-CD45 suggest that CD45 may facilitate early signaling events such as calcium mobilization, and phosphoinositide hydrolyis as well as later events leading to transcription of genes such as c-myc. To examine the role of CD45 more extensively, CD45-deficient mice were generated by disruption of exon 6. Although normal numbers of B cells were found in peripheral lymphoid tissues, CD45-deficient cells failed to proliferate upon IgM crosslinking. In the present study, we demonstrate that the fraction of high buoyant density B cells is reduced while low buoyant density cells are increased. Moreover, there is a significant decline in the number of splenic B cells of the mature IgDhi, IgMlo phenotype. Although both the basal and anti-Ig-induced levels of phosphorylation of Ig-alpha and phospholipase C gamma 2 are indistinguishable from that observed in CD45+ control B cells, a major distinction was found in Ca2+ mobilization. While anti-Ig-induced mobilization of intracellular Ca2+ stores was normal, influx from extracellular sources was abrogated. This finding reveals a novel pathway of regulating B cell responses mediated by CD45.

The B cell-specific transmembrane protein RP-105 belongs to the family of Drosophila toll-like proteins which are likely to trigger innate immune responses in mice and man. Here we demonstrate that the Src-family protein tyrosine kinase Lyn, protein kinase C beta I/II (PKCbetaI/II), and Erk2-specific mitogen-activated protein (MAP) kinase kinase (MEK) are essential and probably functionally connected elements of the RP-105-mediated signaling cascade in B cells. We also find that negative regulation of RP-105-mediated activation of MAP kinases by membrane immunoglobulin may account for the phenomenon of antigen receptor-mediated arrest of RP-105-mediated B cell proliferation.