PLAUR splicing pattern in hereditary angioedema patients' monocytes and macrophages.

BACKGROUND: The PLAUR gene encodes the urokinase-like plasminogen activator receptor (uPAR) and may undergo alternative splicing. Excluding cassette exons 3, 5 and 6 from the transcript results in truncated protein variants whose precise functions have not been elucidated yet. The PLAUR gene is one of several expressed in myeloid cells, where uPAR participates in different cellular processes, including the contact activation system and kallikrein-kinin system, which play an important role in hereditary angioedema (HAE) pathogenesis. A hypothesis about the PLAUR splicing pattern impact on HAE severity was tested. METHODS AND RESULTS: The RT-PCR quantified by capillary electrophoresis was used. Although no significant difference in alternative transcript frequency was observed between healthy volunteers and HAE patients, a significant increase in all cassette exon inclusion variants was revealed during monocyte-to-macrophage differentiation. CONCLUSIONS: PLAUR alternative splicing in monocytes and macrophages neither was different between HAE patients and healthy controls, nor reflected disease severity. However, the results showed an PLAUR splicing pattern was changing during monocyte-to-macrophage differentiation, but the significance of these changes is unknown and awaits future clarification.

TNFR2 expression is a hallmark of human memory B cells with suppressive function.

Tumor Necrosis Factor Receptor 2 (TNFR2) expression is increasingly being linked to tolerogenic immune reactions and cells with suppressor function including a subset of T-regulatory cells. B-regulatory cells play an important role in control of T-cell responses and inflammation. Recently, we described TNFR2 as a marker for IL-10-producing B cells, a hallmark of this cell subset. Here, we demonstrate that proliferation of T cells is reduced in the presence of TNFR2 positive human memory B cells generated with TLR9 ligand, while TNFR2- and TNFR2+CD27- B cells display costimulatory activity. Our data further reveal that IL-10 secretion is characteristic of IgM+ naïve and memory B cells but suppressive activity is not restricted to IL-10: (i) the inhibitory effect of TNFR2+ switched memory B cells was comparable to that exerted by TNFR2+ IgM+ memory B cells although IL-10 secretion levels in the cocultures were lower; (ii) supernatants from TNFR2+ memory B cells failed to suppress T-cell proliferation. Based on our findings, we propose that formation of Breg is a specific characteristic of human memory B cells undergoing terminal differentiation. Our data further corroborate that TNFR2 represents a viable marker for identification of memory B cells with regulatory function.

Neutrophil and Granulocytic Myeloid-Derived Suppressor Cell-Mediated T Cell Suppression Significantly Contributes to Immune Dysregulation in Common Variable Immunodeficiency Disorders.

Common variable immunodeficiency disorders (CVID) represent a group of primary immunodeficiency diseases characterized by hypogammaglobulinemia and impaired specific Ab response, resulting in recurrent infections due to dysfunctional immune response. The specific mechanisms mediating immune deficiency in CVID remain to be determined. Previous studies indicated that immune dysregulation in CVID patients is associated with chronic microbial translocation, systemic immune activation, and altered homeostasis of lymphocytic and myeloid lineages. A detailed phenotypic, functional characterization of plasma markers and immune cell populations was performed in 46 CVID patients and 44 healthy donors. CVID patients displayed significantly elevated plasma levels of a marker of neutrophil activation neutrophil gelatinase-associated lipocalin. Neutrophils from CVID patients exhibited elevated surface levels of CD11b and PD-L1 and decreased levels of CD62L, CD16, and CD80, consistent with a phenotype of activated neutrophils with suppressive properties. Neutrophils from CVID patients actively suppressed T cell activation and release of IFN-γ via the production of reactive oxygen species. Furthermore, CVID was associated with an increased frequency of low-density neutrophils (LDNs)/granulocytic myeloid-derived suppressor cells. LDN/granulocytic myeloid-derived suppressor cell frequency in CVID patients correlated with reduced T cell responsiveness. Exogenous stimulation of whole blood with bacterial LPS emulated some but not all of the phenotypic changes observed on neutrophils from CVID patients and induced neutrophil population with LDN phenotype. The presented data demonstrate that neutrophils in the blood of CVID patients acquire an activated phenotype and exert potent T cell suppressive activity. Specific targeting of myeloid cell-derived suppressor activity represents a novel potential therapeutic strategy for CVID.

Absence of Surface IgD Does Not Impair Naive B Cell Homeostasis or Memory B Cell Formation in IGHD Haploinsufficient Humans.

Surface IgD is coexpressed with IgM on naive mature B cells. Still, the role of surface IgD remains enigmatic even 50 y after its initial discovery. In this study, we examined the in vivo role of surface IgD in human B cell homeostasis and Ab responses in four individuals with heterozygous nonsense mutations in IGHD All IGHD heterozygous individuals had normal numbers of B cells and serum Igs and did not show signs of immunodeficiency or immune dysregulation. IgD+ and IgD- naive mature B cells were present in equal numbers and showed similar immunophenotypes, except for decreased expression of CD79b in the IgD- subset. Furthermore, both IgD+ and IgD- naive mature B cells had normal replication histories and similar capacities to differentiate into plasma cells upon in vitro stimulation, and Ig class-switched memory B cells showed similar levels of somatic hypermutations. Thus, human B cells lacking IgD expression develop normally and generate immunological memory in vivo, suggesting that surface IgD might function more restrictedly in regulating of B cell activation to specific antigenic structures.

Defects in memory B-cell and plasma cell subsets expressing different immunoglobulin-subclasses in patients with CVID and immunoglobulin subclass deficiencies.

BACKGROUND: Predominantly antibody deficiencies (PADs) are the most prevalent primary immunodeficiencies, but their B-cell defects and underlying genetic alterations remain largely unknown. OBJECTIVE: We investigated patients with PADs for the distribution of 41 blood B-cell and plasma cell (PC) subsets, including subsets defined by expression of distinct immunoglobulin heavy chain subclasses. METHODS: Blood samples from 139 patients with PADs, 61 patients with common variable immunodeficiency (CVID), 68 patients with selective IgA deficiency (IgAdef), 10 patients with IgG subclass deficiency with IgA deficiency, and 223 age-matched control subjects were studied by using flow cytometry with EuroFlow immunoglobulin isotype staining. Patients were classified according to their B-cell and PC immune profile, and the obtained patient clusters were correlated with clinical manifestations of PADs. RESULTS: Decreased counts of blood PCs, memory B cells (MBCs), or both expressing distinct IgA and IgG subclasses were identified in all patients with PADs. In patients with IgAdef, B-cell defects were mainly restricted to surface membrane (sm)IgA+ PCs and MBCs, with 2 clear subgroups showing strongly decreased numbers of smIgA+ PCs with mild versus severe smIgA+ MBC defects and higher frequencies of nonrespiratory tract infections, autoimmunity, and affected family members. Patients with IgG subclass deficiency with IgA deficiency and those with CVID showed defects in both smIgA+ and smIgG+ MBCs and PCs. Reduced numbers of switched PCs were systematically found in patients with CVID (absent in 98%), with 6 different defective MBC (and clinical) profiles: (1) profound decrease in MBC numbers; (2) defective CD27+ MBCs with almost normal IgG3+ MBCs; (3) absence of switched MBCs; and (4) presence of both unswitched and switched MBCs without and; (5) with IgG2+ MBCs; and (6) with IgA1+ MBCs. CONCLUSION: Distinct PAD defective B-cell patterns were identified that are associated with unique clinical profiles.

Neutrophils Are Dysregulated in Patients with Hereditary Angioedema Types I and II in a Symptom-Free Period.

Neutrophils impact on processes preceding the formation of bradykinin, a major swelling mediator in hereditary angioedema (HAE), yet their potential role in HAE pathogenesis has not been sufficiently studied. We assessed the relative mRNA expression of 10 genes related to neutrophil activation using RNA extracted from the peripheral blood neutrophils of 23 HAE patients in a symptom-free period and 39 healthy donors. Increased relative mRNA expression levels of CD274, IL1B, IL1RN, IL8, MMP9, and TLR4, together with a lack in their mutual correlations detected in HAE patients compared to healthy controls, suggested a preactivated state and dysregulation of patients' neutrophils. Patients' neutrophil-alerted state was further supported by increased CD11b, decreased CD16 plasma membrane deposition, and increased relative CD274+ and CD87+ neutrophil counts, but not by increased neutrophil elastase or myeloperoxidase plasma levels. As CD274 mediates inhibitory signals to different immune cells, neutrophils were cocultured with T-cells/PBMC. The decrease in CD25+ and IFN-γ + T-cell/PBMC ratio in patients indicated the patients' neutrophil suppressive functions. In summary, the results showed neutrophils' alerted state and dysregulation at the transcript level in patients with HAE types I and II even in a symptom-free period, which might make them more susceptible to edema formation. Neutrophils' T-cell suppressive capacity in HAE patients needs to be further investigated.

Treatment of Hereditary Angioedema Attacks with Icatibant and Recombinant C1 Inhibitor During Pregnancy.

PURPOSE: Hereditary angioedema (HAE) is a rare disease caused by a C1 inhibitor (C1-INH) deficit. Clinically, HAE is manifested by repeated episodes of localized subcutaneous or submucosal oedema attacks. Managing HAE patients in pregnancy is challenging, since there are only limited data on the safety and efficacy of various therapeutic approaches. METHODS: We present our clinical experience treating acute HAE attacks during pregnancy in six consecutive patients. RESULTS: During the pregnancies, 79 HAE attacks occurred. The most frequent were abdominal 53 (67.1%) followed by peripheral 21 (26.6%), facial 10 (12.7%), and laryngeal 10 (12.7%) oedemas; 13 (16.5%) attacks were combined. Fifty (63.3%) attacks were treated with recombinant human C1-INH (rhC1-INH); 17 (21.5%) with plasma-derived, pasteurized, nanofiltered C1-INH (pnfC1-INH); 13 (16.5%) with icatibant; and 1 (1.3%) with plasma-derived, nanofiltered C1-INH (nfC1-INH). Treatment had to be repeated in 5 attacks (6.3%). All six deliveries (one caesarean section and five spontaneous vaginal deliveries) were complication free. All pregnancies went to the full term and the patients delivered healthy babies with a birth weight ranging from 2850 to 3690 g. No congenital abnormalities were detected in the neonates. No abortions occurred. CONCLUSIONS: Our results show good C1-INH or icatibant treatment efficacy for HAE attacks in pregnancy. The treatment by the first drug used was effective in 93.7% of all attacks. In 6.3% of attacks, a second treatment had to be used. No adverse effects were observed.

Selective IgM Deficiency: Clinical and Laboratory Features of 17 Patients and a Review of the Literature.

PURPOSE: Primary selective IgM deficiency (sIgMD) is a primary immunodeficiency with unclear pathogenesis and a low number of published cases. METHODS: We reviewed clinical and laboratory manifestations of 17 sIgMD patients. Serum IgM, IgG, and its subclasses, IgA, IgE, antibodies against tetanus toxoid, pneumococcal polysaccharides and Haemophilus influenzae type b, isohemagglutinins, and T and B lymphocyte subsets, expressions of IgM on B cells and B lymphocyte production of IgM were compared with previously reported case reports and a small series of patients, which included 81 subjects in total. RESULTS: We found that some patients in our cohort (OC) and published cases (PC) had increased IgE levels (OC 7/15; PC 21/37), decreased IgG4 levels (OC 5/14), very low titers of isohemagglutinins (OC 8/8; PC 18/21), increased transitional B cell counts (OC 8/9), decreased marginal zone B cell counts (OC 8/9), and increased 21low B cell counts (OC 7/9). Compared with the PC (20/20), only two of five OC patients showed very low or undetectable production of IgM after stimulation. A majority of the patients had normal antibody production to protein and polysaccharide antigens, basic lymphocyte subset counts, and expression of surface IgM molecules on B cells. CONCLUSIONS: Low IgM levels are associated with various immunopathological disorders; however, pathogenic mechanisms leading to decreased IgM serum level in selective IgM deficiency remain unclear. Moreover, it is difficult to elucidate how strong these associations are and if these immunopathological conditions are primary or secondary.

Terminally differentiated memory T cells are increased in patients with common variable immunodeficiency and selective IgA deficiency.

INTRODUCTION: Previous studies showed that several lymphocyte abnormalities seen in the most frequent symptomatic immunoglobulin deficiency, common variable immunodeficiency (CVID), were also observed in a genetically related asymptomatic disorder - selective IgA deficiency (IgAD). In this study we searched for abnormalities in the differentiation stages of T cells as well as for similarities of these abnormalities in CVID and IgAD patients. MATERIAL AND METHODS: Using flow cytometry in 80 patients with IgAD, 48 patients with CVID, and 80 control persons we determined T-lymphocyte subsets: both CD4 and CD8 were divided into the naïve CD45RO-CD27+, early differentiated CD45RO+CD27+, late differentiated CD45RO+CD27- and fully differentiated effector CD45RO-CD27- memory T cells, as well as Treg cells, defined as CD4+CD25highCD127low T cells. RESULTS: An increase of CD4+ and CD8+ late differentiated memory cells was observed comparing CVID patients to controls, as well as comparing IgAD patients to controls. In CVID patients an increase of CD4+ early differentiated memory cells, a decrease of CD8+ intermediate memory cells, and CD4+ and CD8+ naïve cells were found as well. The abnormalities in IgAD patients might be explained by higher CMV seropositivity observed in our IgAD. We confirmed the repeatedly published decrease of Treg cells in CVID patients, while Treg cells in IgAD patients were increased compared to controls. CONCLUSIONS: Our results show T-cell activation not only in CVID, but also in IgAD patients. The increase in IgAD patients may be influenced by a more frequent CMV infection in our group of IgAD patients.

Regulatory B cells in CVID patients fail to suppress multifunctional IFN-γ+ TNF-α+ CD4+ T cells differentiation.

Common variable immunodeficiency (CVID) refers to primary hypogammaglobulinemia with unknown pathogenesis. Although there is evidence for intrinsic B cell defects in some CVID patient groups, various abnormalities in cytokine production by T cells in CVID patients are frequently observed. Here, we demonstrate a relationship in the production of pro-inflammatory Th1 cytokines and regulatory B cells producing IL-10 between CVID patients and healthy controls. We describe CD19(+)CD24(hi)CD38(hi)IL-10(+) regulatory B cells generated after T cell stimulation of human peripheral blood lymphocytes ex vivo are able to suppress IFN-γ(+)TNF-α(+) producing CD4(+) T cells. This process is impaired in CVID patients, who present with both low numbers of CD19(+)CD24(hi)CD38(hi)IL-10(+) B cells and increased numbers of IFN-γ(+)TNF-α(+)CD4(+) T cells. Disruption of the regulatory B cell response to T cell stimulation explains the excessive T cell activation regarded as an immunoregulatory abnormality that is a frequent finding in CVID patients.

Pregnancy Outcome in Patients with Common Variable Immunodeficiency.

PURPOSE: The aim of our retrospective study was to clarify fertility, pregnancy complications and outcomes in common variable immunodeficiency (CVID) females. METHODS: Retrospective data were obtained from three Czech referral centres. The data were compared with data obtained from the Czech National Registry of Reproduction Health. RESULTS: Our cohort of patients comprised 54 women with 115 pregnancies; 88 pregnancies in 50 females were finished with live births (77 %). In only 8 women (15%) was the diagnosis of CVID established before the first pregnancy. Replacement immunoglobulin therapy was performed in 10 patients without any moderate or severe adverse effects. Compared with the Czech population, the CVID patients suffered significantly more frequently from the threat of preterm labour (p < 0.0001), vaginal bleeding (p = 0.0001), eclampsia/preeclampsia (p = 0.009) and a higher number of stillbirths (p < 0.0001). Furthermore, the frequency of babies with low birth weight (less than 2500 g) born to the CVID patients was increased compared with the normal population (p < 0.0001). Serum IgG, IgA and IgM determination was done in 57 children of 50 mothers showing 13 cases of IgA deficiency (23%). There was no significant difference among the non-symptomatic, symptomatic untreated and symptomatic treated females in any of the determined gynaecological complications. The number of unsuccessful pregnancies was higher in the symptomatic untreated women. CONCLUSIONS: Fertility in CVID patients is not decreased, and their pregnancies could be considered more risky compared with those of the general population.

Successful renal transplantation in a patient with a Wiskott-Aldrich syndrome protein (WASP) gene mutation.

Wiskott-Aldrich syndrome (WAS) is a rare primary immunodeficiency disorder caused by mutations in the WAS protein (WASP) gene. Renal disease progressing to renal failure is a well-recognized complication in patients with WAS. Only a few case reports of renal transplantation have been reported to date. Here, we present a patient with a WASP mutation who suffered from severe atopic eczema, mild thrombocytopenia and only a slightly increased frequency of infections, who then developed IgA nephropathy and consequently underwent renal transplantation, which was successful. This study demonstrates that renal transplantation is possible in patients with WAS, regardless of conceivable complications.

Altered serum cytokine signature in common variable immunodeficiency.

PURPOSE: Common variable immunodeficiency (CVID) is the most frequent form of primary symptomatic hypogammaglobulinemia. CVID patients display a number of abnormalities in lymphocyte subpopulations including chronic T-cell activation and decreased numbers of circulating CD4(+) T cells and NK cells. We and others have recently shown that CVID is associated with increased concentration of soluble CD14 (sCD14) and other factors indicating limited microbial translocation. METHODS: To address the mechanisms of chronic immune activation in CVID, we performed a detailed analysis of cytokine serum levels in 36 patients with CVID, 52 patients with selective IgA deficiency (IgAD), and 56 healthy volunteers. RESULTS: We show that CVID is associated with elevated serum levels of CXCL-10/IP-10, IL-1R antagonist, TNF-α, IL-10, IL-12 (p40), CCL-2/MCP-1, G-CSF, and CCL-11/eotaxin. The detected cytokine signature is consistent with an ongoing activation of cells of myeloid lineage. In contrast, the levels of cytokines typically produced by CD4(+) T helper cells of Th1 (IFN-γ, IL-2), Th2 (IL-9, IL-13), and Th17 (IL-17) subtypes were suppressed in CVID patients compared to healthy donors. CONCLUSIONS: Presented data suggest that the altered cytokine profile observed in patients with CVID may be attributed to the activation of monocyte-macrophage and granulocyte lineages, possibly driven by the translocation of bacterial components across the gastrointestinal or respiratory tracts mucosal barrier.

Phenotype and oxidative burst of low-density neutrophil subpopulations are altered in common variable immunodeficiency patients.

Common variable immunodeficiency disorder (CVID) is the most common form of primary antibody immunodeficiency. Due to low antibody levels, CVID patients receive intravenous or subcutaneous immunoglobulin replacement therapy as treatment. CVID is associated with the chronic activation of granulocytes, including an increased percentage of low-density neutrophils (LDNs). In this study, we examined changes in the percentage of LDNs and the expression of their surface markers in 25 patients with CVID and 27 healthy donors (HD) after in vitro stimulation of whole blood using IVIg. An oxidative burst assay was used to assess the functionality of LDNs. CVID patients had increased both relative and absolute LDN counts with a higher proportion of mLDNs compared to iLDNs, distinguished based on the expression of CD10 and CD16. Immature LDNs in the CVID and HD groups had significantly reduced oxidative burst capacity compared to mature LDNs. Interestingly we observed reduced oxidative burst capacity, reduced expression of CD10 after stimulation of WB, and higher expression of PD-L1 in mature LDNs in CVID patients compared to HD cells. Our data indicate that that the functional characteristics of LDNs are closely linked to their developmental stage. The observed reduction in oxidative burst capacity in mLDNs in CVID patients could contribute to an increased susceptibility to recurrent bacterial infections among CVID patients.

B cell subsets reconstitution and immunoglobulin levels in children and adolescents with B non-Hodgkin lymphoma after treatment with single anti CD20 agent dose included in chemotherapeutic protocols: single center experience and review of the literature.

BACKGROUND: RTX, an anti-CD20 monoclonal antibody, added to chemotherapy has proven to be effective in children and adolescents with high-grade, high-risk and matured non-Hodgkin lymphoma. RTX leads to prompt CD19+ B lymphocyte depletion. However, despite preserved immunoglobulin production by long-lived plasmablasts after treatment, patients remain at risk of prolonged hypogammaglobulinemia. Further, there are few general guidelines for immunology laboratories and clinical feature monitoring after B cell-targeted therapies. The aim of this paper is to describe B cell reconstitution and immunoglobulin levels after pediatric B-NHL protocols, that included a single RTX dose and to review the literature. METHODS: A retrospective single-center study on the impact of a single RTX dose included in a chemotherapeutic pediatric B Non-Hodgkin Lymphoma (B-NHL) treatment protocols. Immunology laboratory and clinical features were evaluated over an eight hundred days follow-up (FU) period, after completing B-NHL treatment. RESULTS: Nineteen patients (fifteen Burkitt lymphoma, three Diffuse large B cell lymphoma, and one Marginal zone B cell lymphoma) fulfilled the inclusion criteria. Initiation of B cell subset reconstitution occurred a median of three months after B-NHL treatment. Naïve and transitional B cells declined over the FU in contrast to the marginal zone and the switched memory B cell increase. The percentage of patients with IgG, IgA, and IgM hypogammaglobulinemia declined consistently over the FU. Prolonged IgG hypogammaglobulinemia was detectable in 9%, IgM in 13%, and IgA in 25%. All revaccinated patients responded to protein-based vaccines by specific IgG antibody production increase. Following antibiotic prophylaxes, none of the patients with hypogammaglobulinemia manifested with either a severe or opportunistic infection course. CONCLUSION: The addition of a single RTX dose to the chemotherapeutic treatment protocols was not shown to increase the risk of developing secondary antibody deficiency in B-NHL pediatric patients. Observed prolonged hypogammaglobulinemia remained clinically silent. However interdisciplinary agreement on regular long-term immunology FU after anti-CD20 agent treatment is required.

Hepcidin and ferritin levels as markers of immune cell activation during septic shock, severe COVID-19 and sterile inflammation.

Introduction: Major clinically relevant inflammatory events such as septic shock and severe COVID-19 trigger dynamic changes in the host immune system, presenting promising candidates for new biomarkers to improve precision diagnostics and patient stratification. Hepcidin, a master regulator of iron metabolism, has been intensively studied in many pathologies associated with immune system activation, however these data have never been compared to other clinical settings. Thus, we aimed to reveal the dynamics of iron regulation in various clinical settings and to determine the suitability of hepcidin and/or ferritin levels as biomarkers of inflammatory disease severity. Cohorts: To investigate the overall predictive ability of hepcidin and ferritin, we enrolled the patients suffering with three different diagnoses - in detail 40 patients with COVID-19, 29 patients in septic shock and eight orthopedic patients who were compared to nine healthy donors and all cohorts to each other. Results: We showed that increased hepcidin levels reflect overall immune cell activation driven by intrinsic stimuli, without requiring direct involvement of infection vectors. Contrary to hepcidin, ferritin levels were more strongly boosted by pathogen-induced inflammation - in septic shock more than four-fold and in COVID-19 six-fold in comparison to sterile inflammation. We also defined the predictive capacity of hepcidin-to-ferritin ratio with AUC=0.79 and P = 0.03. Discussion: Our findings confirm that hepcidin is a potent marker of septic shock and other acute inflammation-associated pathologies and demonstrate the utility of the hepcidin-to-ferritin ratio as a predictor of mortality in septic shock, but not in COVID-19.

Myeloid lineage cells evince distinct steady-state level of certain gene groups in dependence on hereditary angioedema severity.

Hereditary angioedema (HAE) is a rare genetic disorder with variable expressivity even in carriers of the same underlying genetic defect, suggesting other genetic and epigenetic factors participate in modifying HAE severity. Recent knowledge indicates the role of immune cells in several aspects of HAE pathogenesis, which makes monocytes and macrophages candidates to mediate these effects. Here we combined a search for HAE phenotype modifying gene variants with the characterization of selected genes' mRNA levels in monocyte and macrophages in a symptom-free period. While no such gene variant was found to be associated with a more severe or milder disease, patients revealed a higher number of dysregulated genes and their expression profile was significantly altered, which was typically manifested by changes in individual gene expression or by strengthened or weakened relations in mutually co-expressed gene groups, depending on HAE severity. SERPING1 showed decreased expression in HAE-C1INH patients, but this effect was significant only in patients carrying mutations supposedly activating nonsense-mediated decay. Pro-inflammatory CXC chemokine superfamily members CXCL8, 10 and 11 were downregulated, while other genes such as FCGR1A, or long non-coding RNA NEAT1 were upregulated in patients. Co-expression within some gene groups (such as an NF-kappaB function related group) was strengthened in patients with a severe and/or mild course compared to controls. All these findings show that transcript levels in myeloid cells achieve different activation or depression levels in HAE-C1INH patients than in healthy controls and/or based on disease severity and could participate in determining the HAE phenotype.

B-lymphocyte subpopulations in patients with selective IgA deficiency.

INTRODUCTION: Selective deficiency IgA (IgAD) is the most common primary abnormality of immunoglobulin production with unknown pathophysiology. It is genetically related to common variable immunodeficiency (CVID), where besides IgA also IgG and frequently IgM serum levels are decreased. In this study we focused on determination of B-lymphocyte developmental stages and searching for similarities between CVID and IgAD. MATERIALS AND METHODS: Using flow cytometry we determined major lymphocyte subpopulations and B-lymphocyte subsets: naïve (CD27(-)IgD(+)), marginal zone cells (CD27(+)IgD(+)), class-switched memory cells (CD27(+)IgD(-)), "double-negative" B cells (CD27(-)IgD(-)), transitional cells (IgM(++)CD38(++)), plasmablasts (CD38(+++)IgM(+) or IgM(-)), and CD21(low)CD38(low) cells in 80 patients with IgAD, 48 patients with CVID, and 80 control persons. RESULTS: Compared to healthy controls, a decrease in the absolute number and frequency of CD4+ cells (both < 0.001) was observed in IgAD patients. A decrease in the frequency of switched memory cells (P < 0.001), transitional cells (P = 0.035) as well as plasmablasts (P < 0.001) and an increase in the CD21(low)CD38(low) subset (P = 0.007) was observed in IgAD patients compared to control persons. No significant differences were observed in the remaining B-cell developmental subsets. A decrease in CD27(+)IgD(-) (<0.4% of peripheral blood lymphocytes), frequently observed in CVID patients and also previously reported in IgAD, was found in only five patients (6%) with IgAD, two of them being first-degree relatives of CVID patients. CONCLUSION: Our results show a decrease of terminally differentiated B-lymphocyte subsets in patients with IgAD, similar as previously found in patients with CVID, but these results are less expressed than in CVID patients.

Characterization of lymphocyte subsets in patients with common variable immunodeficiency reveals subsets of naive human B cells marked by CD24 expression.

Increased proportions of naive B cell subset and B cells defined as CD27(neg)CD21(neg)CD38(neg) are frequently found in patients with common variable immunodeficiency (CVID) syndrome. Current methods of polychromatic flow cytometry and PCR-based detection of κ deletion excision circles allow for fine definitions and replication history mapping of infrequent B cell subsets. We have analyzed B cells from 48 patients with CVID and 49 healthy controls to examine phenotype, frequency, and proliferation history of naive B cell subsets. Consistent with previous studies, we have described two groups of patients with normal (CVID-21norm) or increased (CVID-21lo) proportions of CD27(neg)CD21(neg)CD38(neg) B cells. Upon further analyses, we found two discrete subpopulations of this subset based on the expression of CD24. The B cell subsets showed a markedly increased proliferation in CVID-21lo patients as compared with healthy controls, suggesting developmental arrest rather than increased bone marrow output. Furthermore, when we analyzed CD21(pos) naive B cells, we found two different subpopulations based on IgM and CD24 expression. They correspond to follicular (FO) I and FO II cells previously described in mice. FO I subset is significantly underrepresented in CVID-21lo patients. A comparison of the replication history of naive B cell subsets in CVID patients and healthy controls implies refined naive B cell developmental scheme, in which human transitional B cells develop into FO II and FO I. We propose that the CD27(neg)CD21(neg)CD38(neg) B cells increased in some of the CVID patients originate from the two FO subsets after loss of CD21 expression.

Immunosuppression Affects Neutrophil Functions: Does Calcineurin-NFAT Signaling Matter?

Neutrophils are innate immune cells with important roles in antimicrobial defense. However, impaired or dysregulated neutrophil function can result in host tissue damage, loss of homeostasis, hyperinflammation or pathological immunosuppression. A central link between neutrophil activation and immune outcomes is emerging to be the calcineurin-nuclear factor of activated T cells (NFAT) signaling pathway, which is activated by neutrophil detection of a microbial threat via pattern recognition receptors and results in inflammatory cytokine production. This potent pro-inflammatory pathway is also the target of several immunosuppressive drugs used for the treatment of autoimmune disorders, during solid organ and hematopoietic cell transplantations, and as a part of anti-cancer therapy: but what effects these drugs have on neutrophil function, and their broader consequences for immune homeostasis and microbial defense are not yet known. Here, we bring together the emerging literature describing pathology- and drug- induced neutrophil impairment, with particular focus on their effects on calcineurin-NFAT signaling in the innate immune compartment.