Frequency Of Bregs Research Articles

Enhanced frequency and potential mechanism of B regulatory cells in patients with lung cancer.

BackgroundRegulatory T cells (Tregs) and B cells (Bregs) play an important role in the development of lung cancer. The present study aimed to investigate the phenotype of circulating Tregs and Bregs in patients with lung cancer and explore potential mechanism by which lung cancer cells act on the development of both.MethodsPatients with lung cancer (n = 268) and healthy donors (n = 65) were enrolled in the study. Frequencies of Tregs and Bregs were measured by flow cytometry with antibodies against CD4, CD25, CD127, CD45RA, CD19, CD24, CD27 and IL-10 before and after co-cultures. qRT-PCR was performed to evaluate the mRNA levels of RANTES, MIP-1α, TGF-β, IFN-γ and IL-4.ResultsWe found a lower frequency of Tregs and a higher frequency of Bregs in patients with lung cancer compared to healthy donors. Co-culture of lung cancer cells with peripheral blood mononuclear cells could polarize the lymphocyte phenotype in the similar pattern. Lipopolysaccharide (LPS)-stimulated lung cancer cells significantly modulated regulatory cell number and function in an in vitro model.ConclusionWe provide initial evidence that frequencies of peripheral Tregs decreased or Bregs increased in patients with lung cancer, which may be modulated directly by lung cancer cells. It seems cancer cells per se plays a crucial role in the development of tumor immunity.Electronic supplementary materialThe online version of this article (doi:10.1186/s12967-014-0304-0) contains supplementary material, which is available to authorized users.

Regulatory B cells correlate with HIV disease progression.

A rare subset of IL-10-producing B cells, named Breg, was recently identified in mice and humans. Currently, there are no unified cell surface markers to identify Breg, and the relationship between the frequency of Breg and HIV disease progression in chronic HIV infection is unclear. In the present study, we determined whether the cell surface markers of Breg reported for other diseases are suitable for identifying Breg in HIV-infected patients. In addition, we examined the relationship between Breg and HIV disease progression. We found that Breg frequency correlated positively with viral load and negatively with CD4 count in chronic HIV infection. Following antiretroviral treatment, the CD4 count increased and the frequency of Breg decreased stepwise. There was no difference in IL-10 expression of CD1d(hi) or CD1d(lo) cells isolated from HIV-infected patients. Therefore, CD1d may not be a marker of Breg in HIV-infected patients.

Serum prevalence of B cell stimulatory TLR ligands, elevated B cell TLR2 expression and regulatory B cell frequency precede the genesis of HIV-non-Hodgkin’s lymphoma (NHL) in ART treated HIV+ subjects (TUM7P.951)

Abstract TLR2-signaling may contribute to B-cell activation during HIV infection leading to HIV-NHL genesis; a common malignancy in anti-retroviral therapy (ART) treated patients. TLR-signaling and anti-tumor immunity inhibitory regulatory B cells (Bregs) contribute to lymphomagenesis but their roles in HIV-NHL genesis remain unexplored. We investigated, if in HIV- donors, TLR-activated B cells mimic an activated/germinal center (GC)-like phenotype aberrantly expressed in B-cell lymphomas. Further, in HIV+ART subjects: (cases = <1 year pre-NHL, controls = HIV+ART but don’t develop NHL), we assessed: 1) B-cell TLR2 and TLR2 modulatory miRNA expression 2) Breg frequency and 3) serum levels of TLR-ligands. In B cells from HIV- donors only TLR2- (unlike TLR4 nor 9) stimulation (p=0.03), leads to an aberrant phenotype (CD10+CD71+CD86+AID+) identified in lymphomas. Compared to controls, cases exhibit significantly elevated: 1) frequency of CD10+CD71+CD86+AID+ B cells (p=0.01) and Bregs (p=0.0071), 2) serum levels of TLR2 and TLR9 ligands (lipoteichoic acid, p=0.03 and 16s DNA, p=0.03 respectively), 3) B-cell TLR2 expression (p=0.0024). Finally, in cases, the activated/GC-like cells exhibit dysregulation of miRNA regulating TLR2-expression including miR-9, miR-19a and miR146a. Our data suggest that HIV-NHL genesis is associated with aberrant GC-like B cells, elevated Bregs and a possible role for TLR2-stimulation.

Altered B cell balance, but unaffected B cell capacity to limit monocyte activation in anti-neutrophil cytoplasmic antibody-associated vasculitis in remission.

Regulatory B cells (Bregs) constitute a subset of B cells with immunomodulatory properties. Numerical and functional alterations in the Breg compartment have been associated with autoimmunity. The aim of this study was to assess the frequency and function of Bregs in patients with ANCA-associated vasculitis (AAV). B cell subsets were determined in the peripheral blood of 48 AAV patients (12 active, 36 in remission) and 41 healthy controls (HCs) by flow cytometry. Bregs were defined within the CD19(+) population as CD24(hi)CD38(hi) or CD24(hi)CD27(+) cells. The percentage of IL-10-positive B cells in circulation was analysed by flow cytometry. Sorted CD19(+) B cells were co-cultured with monocytes to evaluate their capacity to inhibit monocyte TNF-α production upon lipopolysaccharide stimulation. The frequency of circulating CD19(+)CD24(hi)CD38(hi) cells was not different in AAV patients in remission compared with HCs, but was decreased in patients with active disease [mean in HCs 5.5% (s.d. 1.6) vs active 3.8% (s.d. 2.8), P = 0.0104]. Furthermore, the percentage of CD19(+)CD24(hi)CD27(+) cells was significantly decreased in both remission and active patients when compared with HCs [HCs 15.0% (s.d. 9.3) vs remission 6.6% (s.d. 4.4) (P < 0.0001) vs active 6.4% (s.d. 6.2) (P = 0.0006)]. The frequency of IL-10-positive B cells was comparable between patients and HCs. B cells from AAV patients suppressed monocyte TNF-α production to a similar extent to cells from HCs. Based on immunophenotypic classification, Bregs are numerically diminished in AAV patients. However, B cell function in terms of IL-10 production and their capacity to suppress monocyte activation is not compromised in AAV patients in remission.

Regulatory B cell frequency correlates with markers of HIV disease progression and attenuates anti-HIV CD8+ T cell function in vitro

HIV infection is associated with elevated expression of IL-10 and PD-L1, contributing to impairment of T cell effector functions. In autoimmunity, tumor immunology, and some viral infections, Bregs modulate T cell function via IL-10 production. In this study, we tested the hypothesis that during HIV infection, Bregs attenuate CD8(+) T cell effector function, contributing to immune dysfunction. We determined that in vitro, TLR2-, TLR9-, and CD40L-costimulated Bregs from HIV(-) individuals exhibited a high frequency of cells expressing IL-10 and PD-L1. Compared with Bregs from HIV(-) individuals, a significantly higher percentage of Bregs from HIV(+) individuals spontaneously expressed IL-10 (P=0.0218). After in vitro stimulation with HIV peptides, Breg-depleted PBMCs from HIV(+) individuals exhibited a heightened frequency of cytotoxic (CD107a(+); P=0.0171) and HIV-specific CD8(+) T cells compared with total PBMCs. Furthermore, Breg depletion led to enhanced proliferation of total CD8(+) and CD107a(+)CD8(+) T cells (P=0.0280, and P=0.0102, respectively). In addition, augmented CD8(+) T cell effector function in vitro was reflected in a 67% increased clearance of infected CD4(+) T cells. The observed Breg suppression of CD8(+) T cell proliferation was IL-10-dependent. In HIV(+) individuals, Breg frequency correlated positively with viral load (r=0.4324; P=0.0095), immune activation (r=0.5978; P=0.0005), and CD8(+) T cell exhaustion (CD8(+)PD-1(+); r=0.5893; P=0.0101). Finally, the frequency of PD-L1-expressing Bregs correlated positively with CD8(+)PD-1(+) T cells (r=0.4791; P=0.0443). Our data indicate that Bregs contribute to HIV-infection associated immune dysfunction by T cell impairment, via IL-10 and possibly PD-L1 expression.

PI3K-Akt pathway modulates IL-10 producing regulatory B cell

Introduction: IL-10 producing regulatory B (Breg) cells negatively regulate autoimmune disease and inflammation, such as contact hypersensitivity (CHS). CD19 expression is critical for Breg cells development, since CD19 loss results in decreased number of Breg cell and increased reactivity of CHS, which is ameliorated by the adoptive transfer of Breg cells from wild-type mice. However, molecular mechanism of Breg cell development remains poorly understood. CD19 downstream signal is depend on the activation of phosphoinositide 3-kinases (PI3 K)-Akt pathways, while phosphatase and tensin homologue (PTEN) attenuates PI3K-Akt pathways. In this study, we investigated the role of PI3K-Akt pathway in the development of Breg cells.Method: The effect of PI3K-Akt pathway inhibitors on Breg cell development was examined in vitro. B cell-specific PTEN deficient mice, which exhibit aberrant activation of PI3K-Akt pathway in B cell, were generated using Cre-loxp system. CHS severity in B cell-specific PTEN deficient mice was investigated. Result: The current studies demonstrate that PI3K-Akt pathway inhibitors reduced Breg cells in vitro in dose-dependent manner. However, PTEN inhibitor had no effect on Breg cells. Breg cell number and frequency were significantly increased in various organs of B cell-specific PTEN deficient mice when compared with wild-type mice. Furthermore, CHS response was significantly diminished in B cell-specific PTEN deficient mice when compared with wild-type mice. Thus, PI3K-Akt pathway positively regulates Breg cell development, while PTEN negatively regulates it. Conclusion: PI3K-Akt pathway in B cell is critical for Breg cell development and could be a potent therapeutical target. JSID AbstractsJournal of Dermatological ScienceVol. 69Issue 2Preview Full-Text PDF

Regulatory B cells are induced in untreated HIV-1 infection and suppress HIV-1 specific T cell responses

Background Regulatory B cells (Breg), the B cells producing interleukin 10 (IL-10), have been identified in mice and humans. Mouse Breg can suppress innate and T cell responses and are implicated in pathogenesis of some autoimmune diseases and immune evasion of some pathogens. However, the role of Breg in humans is less clear. Methods PBMC and gut biopsy samples were obtained from healthy donors and HIV infected individuals. Flow cytometry and Luminex were used to quantify cytokine production. Flow cytometry were used to analyze Breg’s phenotype. Results Breg were elevated in both peripheral blood and gut tissue of untreated HIV-1 infected individuals and the elevation correlated with viral load in early HIV-1 infection. Breg from HIV-1 infected individuals were CD19 + TIM-1 + .Antiretroviral therapy could reduce elevated Breg frequency. Treatment of B cells from healthy donors with microbial translocation products could differentiate them toward a Breg phenotype. Ex vivo Bregs from HIV-1 infected individuals suppressed cytokine production /degranulation of HIV-1 specific T cells that was in part IL-10 dependent. Conclusion

Selective Depletion of CD19+CD24hiCD38hiIL-10high Regulatory B cells (Bregs) in HIV-infected Aviremic Individuals (76.2)

Abstract HIV-infection associated upregulation of IL-10 contributes to impaired antiviral effector T-cell functions promoting viral persistence. Bregs (CD19+CD24hiCD38hi) IL-10-mediated modulation of T-cell function has been reported in autoimmune disease but yet undefined during HIV infection. In this study, we confirmed high expression of IL-10 by Bregs in healthy controls (HC, n=16) and determined Breg frequency, T-cell immune activation (CD8+HLA-DR+CD38+, CD4+HLA-DR+CD38) and T-cell exhaustion (CD8+PD-1+) in aviremic (HIVavir, n=10) and viremic (HIVvir, n=15) HIV-infected individuals. We found a significant positive correlation between Breg frequency and viral load (r2=0.5166, p=0.0097); HIVavir-individuals exhibited a significant Bregs loss compared to HC (p=0.0004) and HIVvir-individuals (p=0.0122), despite no significant differences in total B-cell frequency. A significant positive correlation was observed between Breg frequency and immune activation (CD8+HLA-DR+CD38+, r2=0.6262, p=0.0041; CD4+HLA-DR+CD38+, r2=0.6298, p=0.0039) as well as T-cell exhaustion (r2=0.8853, p=0.0333). Additionally, we found a significant difference in Breg CD95 (Fas) expression between HIVavir and HIVvir-individuals (p=0.0119) yet no such difference between in HIVavir-individuals and HC. We hypothesize that, Bregs loss during HIV-infection may be due to Fas mediated apoptosis and that activated IL-10-secreting Bregs contribute to T cell impairment hence viral persistence in HIVvir individuals.

Perioperative dynamic alterations in peripheral regulatory T and B cells in patients with hepatocellular carcinoma

BackgroundIntratumoral and circulating regulatory T cells (Tregs) have been shown to be critical in the pathogenesis of hepatocellular carcinoma (HCC). However there is limited knowledge on the alterations of regulatory B cells (Bregs). We here investigated perioperative dynamic alterations of peripheral circulating Tregs and Bregs in HCC patients to reveal the relationship between regulatory lymphocytes and its clinical implications.Methods36 patients with HCC, 6 with chronic hepatitis B infection and 10 healthy donors were enrolled for this study. Frequencies of peripheral Tregs and Bregs were measured by flow cytometry with antibodies against CD4, CD25, CD127, CD19 and IL-10 before, and after radical surgery. Then, clinical informatics of HCC patients was achieved through Digital Evaluation Score System (DESS) for the assessment of disease severity. Finally, we analysed correlations between digitalized clinical features and kinetics of circulating regulatory lymphocytes.ResultsLevel of circulating CD4+CD25+CD127- Tregs in HCC patients was significantly lower than that in healthy donors and patients with chronic hepatitis B infection before surgery, but was increased after surgery. Preoperative level of CD19+ IL-10+ Bregs in HCC patients was also significantly lower than the other groups. However it dramatically was elevated right after surgery and remained elevated compared to controls (about 7 days after surgery, P = 0.04). Frequency of circulating Tregs was correlated with circulating leukocytes, ferritin, and clinical features suggesting tumor aggressiveness including portal vein thrombosis, hepatic vein involvement and advanced clinical stages. Frequency of circulating Bregs was associated with Hepatitis B e Antigen (HBeAg) and Hepatitis B virus (HBV) DNA copy number. In addition, DESS was significantly and positively correlated with other staging systems.ConclusionFrequencies of peripheral Tregs and Bregs in HCC patients increased after surgery. These results suggest that a postoperative combination of therapies against Tregs and Bregs may be beneficial for better outcome of HCC patients after resection.

Regulatory B Cell Defects In Patients with Immune Thrombocytopenia (ITP)

AbstractAbstract 379B lymphocytes participate in immune responses through production of antibodies, antigen presentation to T cells, and cytokine secretion. In ITP, B cells that produce platelet-specific autoantibodies play a major role in the pathogenesis of disease. Recent data from mouse models of autoimmune and inflammatory diseases suggest that B cells have regulatory functions mediated by the production of regulatory cytokines such as IL-10 and/or through inhibitory interactions with effector T cells. In humans, CD19+CD24hiCD38hi B cells, originally identified as immature transitional B cells, were recently shown to possess regulatory capacity mediated in part by IL-10. Given that ITP pathogenesis is in part related to defective T helper functions and since Bregs are important for controlling CD4+ T cell responses, we initiated studies to characterize the CD19+CD24hiCD38hi B cell compartment in ITP patients. Phenotypic analysis in patients with ITP showed statistically significant increases in the frequency of CD19+CD24hiCD38hi B cells (13.2± 1.5% versus 7.7±0.6%, p=0.0015) as well as in the percentage of CD19+CD24hiCD38int mature B cells (72.0± 1.7% versus 62.3±1.6%, p=0.0003). In contrast there was a pronounced decrease in CD19+CD24hiCD38− memory B cells (9.3± 2% versus 26.2±2%, p<0.0001 as confirmed by expression of CD27 memory marker) in patients (n=23) compared to healthy controls (n=23). The suppressive capacity of human CD19+CD24hiCD38hi Bregs involves CD80. We found a statistically significant decrease in the overall frequency of CD80 expressing CD19+ B cells in ITP patients compared to healthy controls (12.40± 1.3% versus 21.8±2.1%, p=0.002), indicating that despite the increase in the Breg frequency, their regulatory capacity may be impaired. To determine the functional activity of the Bregs, PBMCs from 4 patients with ITP were depleted of CD19+CD24hiCD38hi B cells by cell sorting and levels of cytokine production in the CD4+ T cell population following stimulation with plate-bound anti-CD3 for 72h was evaluated. As has been reported previously, we observed a >30% increase in the frequency of CD4+IFN-g+ in CD19+CD24hiCD38hi B cell-depleted compared with non-depleted PBMCs from healthy controls. However, depletion of CD19+CD24hiCD38hi B cells from PBMCs of ITP patients did not alter CD4+IFN-g+ production in 3 /4 patients, indicating a possible defect in the suppressive activity of Bregs in those 3 patients. IL-10 secretion following CD40 engagement in human Bregs is pivotal in mediating their suppressive activity. Given that platelets express CD40L, we tested the ability of Bregs to respond to platelets. We found two-fold lower IL-10 production in CD19+CD24hiCD38hi Bregs (1.9±2.1%, versus 5.0± 1.0%, p<0.05), but not in CD19+CD24hiCD38− B cells (p=0.9) from patients (n=3, different from above) compared to controls (n=5) when PBMCs were stimulated with acid-treated platelets (to remove HLA molecules) from normal controls, indicating a defect in IL-10 production in the Breg population in patients with ITP. Altogether, our in vitro studies of circulating Bregs suggest that these cells may be functionally compromised in some patients with ITP patients as indicated by reduced IL-10 production and indirectly by Breg depletion studies showing inability to dampen effector T cell responses. Given the important role of Bregs in controlling CD4+T responses, the data implicates defective Bregs as an additional mechanism to explain increased T cell responses in patients with ITP. The impaired Breg activity may also explain the variability in response to treatment with anti-CD20 in patients with ITP. It may be that B cell depletion therapy in patients who have defective Breg activity will result in the removal of primarily pathogenic B cells and therefore these patients will have a good response to rituximab treatment. However, in patients with intact Bregs, the same treatment will deplete both pathogenic and regulatory B cells, causing a less effective response. Ongoing studies to test patients' platelets as well as CD40-specific Breg responsiveness are currently underway to further explore the role of Bregs in the pathogenesis of ITP. Disclosures:Bussel:Portola: Consultancy; Amgen: Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Research Funding; GlaxoSmithKline: Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Research Funding; Ligand: Membership on an entity's Board of Directors or advisory committees, Research Funding; Shionogi: Membership on an entity's Board of Directors or advisory committees, Research Funding; Eisai, Inc.: Membership on an entity's Board of Directors or advisory committees; Cangene: Research Funding; Genzyme: Research Funding.