CD27+ peripheral blood B-cells are a useful biodosimetric marker in vitro

The CD8+ natural killer (NK) subpopulation has recently been identified as a fast and reliable biodosimetric indicator within human peripheral blood mononuclear cells (PBMC) in vitro. In irradiated and subsequently cultivated PBMC, a decrease of the relative number of intact CD3- CD8+ lymphocytes 16 and 48 h after treatment has allowed for estimating the received dose in the range of 0 - 10 Gy and lethal/sublethal dose discrimination, respectively. Here we show that suitable biodosimeters can also be found in the peripheral blood B-cell compartment. Multiparameter flow cytometric analysis of irradiated and subsequently cultivated human PBMC revealed that both the CD27+ and CD21- B-cell subpopulations can be used as biodosimeters and the CD19+CD27+ lymphocytes have proved useful for retrospective determination of the received dose in the range of 0 - 6 Gy. In addition, several CD19+ lymphocyte subsets characterized by co-expression of CD21, CD27 and CD38 have been shown to bear biodosimetric potential, too. However, when important parameters like the original size within the CD19+ compartment, its radiation-induced changes and data variation had been taken into account, the CD27+ subpopulation proved superior to the other B-cell subpopulations and subsets. It appears that, in the dose range of 0 - 6 Gy, the relative decrease of CD27+ B lymphocytes provides more sensitive and reliable data than that of CD8+ NK-cells due mainly to lower data variation. In contrast to CD27+ B-cells, the proportions of CD27+ subpopulations of T-cells were not affected by irradiation. We have also proposed a simple experimental protocol based on full blood cultivation and three-color CD27/CD3/CD19 immunophenotyping as a time-saving and inexpensive approach for practical biodosimetric evaluations on simple, three-to-four color flow cytometers.

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Radiosensitivity of peripheral blood B cells in pigs

Veterinární Medicína ◽

10.17221/59/2009-vetmed ◽

2009 ◽

Vol 54 (No. 5) ◽

pp. 223-235 ◽

Cited By ~ 4

Author(s):

Z. Sinkorova ◽

J. Sinkora ◽

L. Zarybnicka ◽

Z. Vilasova ◽

J. Pejchal

Keyword(s):

B Cells ◽

B Cell ◽

Peripheral Blood ◽

Dose Range ◽

Whole Body ◽

Large Animal Model ◽

Large Animal ◽

Cell Subpopulations

: Swine are here introduced to biodosimetry in an attempt to develop a large animal model allowing for comparison of in vitro experiments with the in vivo processes occurring after exposure to gamma radiation. This work investigates the radiosensitivity of the B cell compartment in peripheral blood. Four-week-old piglets were irradiated using the whole body protocol or full blood samples were irradiated in vitro in the dose range of 0–10 Gy. Relative radioresistance of B cell subpopulations and subsets was determined by measuring their relative numbers in leukocyte preparations at selected time intervals after irradiation using two color immunophenotyping and flow cytometry. Porcine B cells represent the most radiosensitive lymphocyte population in peripheral blood. Among B cell subpopulations and subsets investigated, the CD21+SWC7+ and CD21+CD1+ cells are highly radiosensitive and possess biodosimetric potential, at least in the range of low doses. Differences between cultures irradiated in vitro and lymphocyte dynamics in peripheral blood of irradiated animals clearly document the limits of in vitro data extrapolation in biodosimetry. We have shown that pigs can successfully be used in radiobiology and experimental biodosimetry due mainly to their availability, size and a relatively broad spectrum of available immunoreagents for lymphocyte classification.

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THU0030 DISTINCT EFFECTS OF FIVE JAK INHIBITORS IN THE MODULATION OF HUMAN B CELL ACTIVATION

Annals of the Rheumatic Diseases ◽

10.1136/annrheumdis-2020-eular.5763 ◽

2020 ◽

Vol 79 (Suppl 1) ◽

pp. 228.2-229

Author(s):

N. Frede ◽

J. Hueppe ◽

R. Lorenzetti ◽

A. Troilo ◽

M. T. Schleyer ◽

...

Keyword(s):

B Cells ◽

B Cell ◽

Cell Activation ◽

Flow Cytometric Analysis ◽

Cytokine Secretion ◽

B Cell Activation ◽

Jak Inhibitors ◽

Cell Subpopulations ◽

Inhibitor Treatment

Background:JAK inhibitors have been successfully introduced in the treatment of rheumatoid arthritis (RA) and psoriatic arthritis and are in clinical trials for numerous other autoimmune diseases. JAK inhibition effectively reduces cytokine-mediated activation and survival of pathology-driving immune cells by targeting signaling downstream of cytokine receptors. The outcome of such immunomodulation hence will largely depend on the intrinsic expression of the four different JAKs, the cytokine environment and the targeted cell type. Comparative studies investigating the effect on B cells are lacking. In light of the use of JAK inhibitor treatment in autoantibody mediated diseases, the study of the B cell compartment represents a milestone to assess their potential.Objectives:We thus aimed to study the B cell compartment as well as B cell function under JAK inhibition in RA patients and to compare the specific effect the JAK inhibitors tofacitinib (pan-JAK), baricitinib (JAK1/2), ruxolitinib (JAK1/2), upadacitinib and filgotinib (selective JAK1) on in vitro B cell activation, differentiation, proliferation, and class switch.Methods:B cell subpopulations in RA patients treated with baricitinib or tofacitinib was assessed by flow cytometric analysis of peripheral blood mononuclear cells. For in vitro studies, magnetically isolated total B cells from healthy donors were stimulated T-cell -independently with CpG and treated with scalar doses of the JAK inhibitors tofacitinib, baricitinib, ruxolitinib, upadacitinib and filgotinib. Flow cytometric analysis was performed on days 0, 3 and 6. Cytokine secretion was measured by Cytokine Multiplex Assay.Results:B cell phenotyping of RA patients treated with JAK inhibitors baricitinib or tofacitinib showed an increase in marginal zone (MZ) B cells. To investigate this further, we turned to an in vitro model of T-cell-independent B cell activation with CpG via TLR9, known to support MZ B cell expansion. Here, JAK1/2 and selective JAK1 inhibitor treatment led to a dose-dependent decrease of total B cell numbers. When assessing B cell-subpopulations, we observed an altered B cell differentiation with a significant increase in MZ-like B cells under JAK inhibition, which led to a subsequent increase in plasmablast differentiation in the first days. This effect was more pronounced upon pan-JAK inhibitor treatment than JAK1 or JAK1/2 inhibition, indicating that broader JAK inhibition is associated with a stronger effect (tofa > ruxo > bari > upa > filgo).Notably, we further detected a significant dose-dependent reduction of switched memory formation, strongest with JAK1/2 inhibition (upa > ruxo > bari > tofa > filgo). Consistent with this finding, we observed decreased AID expression under JAK inhibition. Concomitantly, induction of STAT3 expression and STAT3 phosphorylation were reduced under JAK inhibition, suggesting that downstream signalling was abrogated.To assess the role of autocrine signaling in this system, we measured cytokine secretion upon JAK inhibition and found that JAK2 inhibition led to reduced IL10 secretion. This in turn resulted in an increase of inflammatory cytokines such as IL6, TNF, highlighting the importance of B cell as cytokine-secreting cell type.Conclusion:In a T-independent in vitro B cell model JAK inhibition led to a reduced total B cell number as well as reduced switched memory development, whereas MZ-like B cells were increased. Especially JAK2 inhibition strongly impaired switched memory formation. JAK inhibition does not only impact cytokine signalling but also leads to changes in cytokine secretion dynamics and amounts, potentially impacting other cell types.In conclusion, JAK inhibition has a major effect on B cell activation and maturation, with differential outcomes between JAK inhibitors hinting towards distinct and unique effects on B cell homeostasis.Disclosure of Interests:None declared

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POS0387 ACPA STATUS CORRELATES WITH DIFFERENTIAL IMMUNE PROFILE OF RHEUMATOID ARTHRITIS PATIENTS

Annals of the Rheumatic Diseases ◽

10.1136/annrheumdis-2021-eular.1885 ◽

2021 ◽

Vol 80 (Suppl 1) ◽

pp. 423.2-424

Author(s):

A. Floudas ◽

M. Canavan ◽

T. McGarry ◽

V. Krishna ◽

S. Nagpal ◽

...

Keyword(s):

T Cell ◽

B Cells ◽

B Cell ◽

Synovial Tissue ◽

Peripheral Blood ◽

Flow Cytometric Analysis ◽

Algorithm Analysis ◽

Treat To Target ◽

Tnf Α ◽

Acpa Status

Background:Rheumatoid arthritis (RA) is a progressive erosive autoimmune disease that affects 1% of the world population. Anti-citrullinated protein autoantibodies (ACPA) are routinely used for the diagnosis of RA, however 20-30% of patients are ACPA negative. ACPA status is a delineator of RA disease endotypes with similar clinical manifestation but potentially different pathophysiology. Elucidating the underlying mechanisms of disease pathogenesis could inform a treat to target approach for both ACPA-positive and ACPA-negative RA patients.Objectives:To identify peripheral blood and synovial tissue immune population differences that associate with RA disease endotype.To identify unique RA patient synovial tissue gene signatures and enriched pathways that correlate with ACPA status.Methods:Detailed high dimensionality flow cytometric analysis with supervised and unsupervised algorithm analysis of ACPApos and ACPAneg RA patient peripheral blood and synovial tissue single cell suspensions. Ex vivo peripheral blood and synovial tissue T cell stimulation and cytokine production characterisation. RNAseq analysis with specific pathway enrichment analysis of APCApos and ACPAneg RA patient synovial tissue biopsies.Results:Detailed profiling based on high dimensionality flow cytometric analysis of key peripheral blood and synovial tissue immune populations including B cells, T follicular helper (Tfh) cells, T peripheral helper cells (Tph) and CD4 T cell proinflammatory cytokine responses with supervised and unsupervised algorithm analysis revealed unique RA patient peripheral blood B cell and Tfh cell profiles. ACPApos RA patients were characterised by significantly (*P=0.03) increased frequency of Tfh (CXCR5+CD4+) cells and distinct clustering influenced by increased switched (IgD-CD27+) and DN (IgD-CD27-) memory B cells compared to APCAneg RA patients. Surprisingly synovial tissue B cell subpopulation distribution was similar between ACPAneg and ACPApos RA patients, with significant accumulation of switched and double negative memory B cells, highlighting a key role for specific B cell subsets in both disease endotypes. Interestingly, synovial tissue CD4 T cell proinflammatory cytokine (TNF-α, IFN-γ, IL-2, GM-CSF, IL-17A, IL-22, IL-4) production was markedly different between ACPAneg and APCApos RA patients with hierarchical clustering and PCA analysis revealing endotype specific cytokine profiles with ACPAneg RA patient synovial T cells showing increased TNF-α (P=0.01) expression. RNAseq analysis of RA patient synovial tissue revealed significant disease endotype specific gene signatures with specific enrichment for B cell receptor signalling and T cell specific pathways in ACPApos compared to ACPAneg RA patients. Additionally, significantly different chemokine receptor expression based on RA patient ACPA status was observed with increased CXCR3 (P<0.001), CCR7 (P=0.002), and CCR2 (P=0.004) but decreased CXCR7 (P=0.007) expression in APCApos compared to ACPAneg RA patient synovial biopsies.Conclusion:ACPA status associates with unique synovial tissue immune cell and gene profile signatures highlighting differences in the underlying immunological mechanisms involved, therefore reinforcing the need for a treat to target approach for both endotypes of RA.Figure 1.RNAseq analysis of synovial tissue biopsies revealed specific T cell related pathway enrichment in ACPA positive compared to ACPA negative RA patients (n=50, analysis performed with the DESq2 and pathfindeR pipelines in R).Disclosure of Interests:Achilleas Floudas: None declared, Mary Canavan: None declared, Trudy McGarry Employee of: Novartis, Vinod Krishna Employee of: Janssen, Sunil Nagpal Employee of: Janssen, GSK, Douglas Veale Speakers bureau: Abbvie, Janssen, Novartis, MSD, Pfizer, UCB, Consultant of: Abbvie, Janssen, Novartis, MSD, Pfizer, UCB, Grant/research support from: Janssen, Abbvie, Pfizer, UCB, Ursula Fearon Speakers bureau: Abbvie, Grant/research support from: Janssen, Abbvie, Pfizer, UCB

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Monoclonal antibodies to the 140,000 mol wt glycoprotein of B lymphocyte membranes (CR2 receptor) initiates proliferation of B cells in vitro

Blood ◽

10.1182/blood.v66.4.824.bloodjournal664824 ◽

1985 ◽

Vol 66 (4) ◽

pp. 824-829

Author(s):

BS Wilson ◽

JL Platt ◽

NE Kay

Keyword(s):

Monoclonal Antibodies ◽

B Cells ◽

B Cell ◽

Peripheral Blood ◽

B Lymphocyte ◽

Human Peripheral Blood ◽

Raji Cell ◽

Lymphoid Cells ◽

Humoral Immune

Several mouse monoclonal IgG antibodies (AB1, AB2, AB3, and AB5) were developed that reacted with a 140,000 mol wt glycoprotein on the surface of cultured RAJI B lymphoid cells. The antibodies reacted with purified normal human peripheral blood B cells and CLL Ig+ B cells and showed specific germinal center and mantle zone staining in tissue sections of secondary lymphoid organs. Immunodepletion studies using 125I surface-labeled Raji cell membrane antigens demonstrated that the antigen identified by AB5 is the same 140,000 mol wt glycoprotein detected by anti-B2 that has recently been shown to react with the C3d fragment or CR2 receptor. (Iida et al: J Exp Med 158:1021, 1983). Addition of the AB series and anti-B2 monoclonal antibodies to cultures of purified human peripheral blood B cells resulted in the uptake of 3H- thymidine at two to six times background control levels provided that irradiated autologous T cells were added to the culture. Stimulation was not evoked by other monoclonal antibodies to B cell surface molecules (ie, B1, BA-1, BA-2, and HLA-DR). Pepsin-generated F(ab')2 fragments of anti-CR2 antibodies were essentially as effective as the intact IgG molecule in stimulating B cells. Induction of B cell proliferation by antibody binding to CR2 suggests that the C3d receptor may have an integral role in regulation of humoral immune response.

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Interleukin-13 in Combination With CD40 Ligand Potently Inhibits Apoptosis in Human B Lymphocytes: Upregulation of Bcl-xL and Mcl-1

Blood ◽

10.1182/blood.v89.12.4415 ◽

1997 ◽

Vol 89 (12) ◽

pp. 4415-4424 ◽

Cited By ~ 54

Author(s):

Jon Lømo ◽

Heidi Kiil Blomhoff ◽

Sten Eirik Jacobsen ◽

Stanislaw Krajewski ◽

John C. Reed ◽

...

Keyword(s):

B Cells ◽

B Cell ◽

B Lymphocytes ◽

Peripheral Blood ◽

Cell Apoptosis ◽

Cell Types ◽

Cd40 Ligand ◽

Interleukin 13

Abstract Interleukin-13 (IL-13) is a novel T-cell–derived cytokine with IL-4–like effects on many cell types. In human B lymphocytes, IL-13 induces activation, stimulates proliferation in combination with anti-IgM or anti-CD40 antibodies, and directs Ig isotype switching towards IgE and IgG4 isotypes. We show here that IL-13 also regulates human B-cell apoptosis. IL-13 reduced spontaneous apoptosis of peripheral blood B cells in vitro, as shown by measurement of DNA fragmentation using the TUNEL and Nicoletti assays. The inhibition of cell death by IL-13 alone was significant but modest, but was potently enhanced in combination with CD40 ligand (CD40L), a survival stimulus for B cells by itself. Interestingly, IL-13 increased the expression of CD40 on peripheral blood B cells, providing a possible mechanism for the observed synergy. IL-13 alone was a less potent inhibitor of apoptosis than IL-4. Moreover, there was no additive effect of combining IL-4 and IL-13 at supraoptimal concentrations, which is consistent with the notion that the IL-4 and IL-13 binding sites share a common signaling subunit. The combination of IL-13 with CD40L augmented the expression of the Bcl-2 homologues Bcl-xL and Mcl-1, suggesting this as a possible intracellular mechanism of induced survival. By contrast, levels of Bcl-2, and two other Bcl-2 family members, Bax and Bak, remained unaltered. Given the importance of the CD40-CD40L interaction in B-cell responses, these results suggest a significant role of IL-13 in the regulation of B-cell apoptosis.

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In VitroMeasles Virus Infection of Human Lymphocyte Subsets Demonstrates High Susceptibility and Permissiveness of both Naive and Memory B Cells

Journal of Virology ◽

10.1128/jvi.00131-18 ◽

2018 ◽

Vol 92 (8) ◽

pp. e00131-18 ◽

Cited By ~ 19

Author(s):

Brigitta M. Laksono ◽

Christina Grosserichter-Wagener ◽

Rory D. de Vries ◽

Simone A. G. Langeveld ◽

Maarten D. Brem ◽

...

Keyword(s):

T Cells ◽

B Cells ◽

B Cell ◽

Immune Suppression ◽

Peripheral Blood ◽

Measles Virus ◽

Memory B Cells ◽

T And B Cells ◽

B Cell Subsets

ABSTRACTMeasles is characterized by a transient immune suppression, leading to an increased risk of opportunistic infections. Measles virus (MV) infection of immune cells is mediated by the cellular receptor CD150, expressed by subsets of lymphocytes, dendritic cells, macrophages, and thymocytes. Previous studies showed that human and nonhuman primate memory T cells express higher levels of CD150 than naive cells and are more susceptible to MV infection. However, limited information is available about the CD150 expression and relative susceptibility to MV infection of B-cell subsets. In this study, we assessed the susceptibility and permissiveness of naive and memory T- and B-cell subsets from human peripheral blood or tonsils toin vitroMV infection. Our study demonstrates that naive and memory B cells express CD150, but at lower frequencies than memory T cells. Nevertheless, both naive and memory B cells proved to be highly permissive to MV infection. Furthermore, we assessed the susceptibility and permissiveness of various functionally distinct T and B cells, such as helper T (TH) cell subsets and IgG- and IgA-positive memory B cells, in peripheral blood and tonsils. We demonstrated that TH1TH17 cells and plasma and germinal center B cells were the subsets most susceptible and permissive to MV infection. Our study suggests that both naive and memory B cells, along with several other antigen-experienced lymphocytes, are important target cells of MV infection. Depletion of these cells potentially contributes to the pathogenesis of measles immune suppression.IMPORTANCEMeasles is associated with immune suppression and is often complicated by bacterial pneumonia, otitis media, or gastroenteritis. Measles virus infects antigen-presenting cells and T and B cells, and depletion of these cells may contribute to lymphopenia and immune suppression. Measles has been associated with follicular exhaustion in lymphoid tissues in humans and nonhuman primates, emphasizing the importance of MV infection of B cellsin vivo. However, information on the relative susceptibility of B-cell subsets is scarce. Here, we compared the susceptibility and permissiveness toin vitroMV infection of human naive and memory T- and B-cell subsets isolated from peripheral blood or tonsils. Our results demonstrate that both naive and memory B cells are more permissive to MV infection than T cells. The highest infection levels were detected in plasma cells and germinal center B cells, suggesting that infection and depletion of these populations contribute to reduced host resistance.

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Telomerase Activity Is Induced in Human Peripheral B Lymphocytes by the Stimulation to Antigen Receptor

Blood ◽

10.1182/blood.v89.4.1299 ◽

1997 ◽

Vol 89 (4) ◽

pp. 1299-1307 ◽

Cited By ~ 73

Author(s):

Hideya Igarashi ◽

Nobuo Sakaguchi

Keyword(s):

B Cells ◽

B Cell ◽

Peripheral Blood ◽

Cell Activation ◽

Interleukin 2 ◽

Antigen Receptor ◽

Telomerase Activity ◽

B Cell Activation ◽

Molecular Events

Abstract To understand the molecular events for the proliferation of B cells, we studied the induction of telomerase activity in vitro after stimulation to B-cell antigen receptor (BCR) on human peripheral B cells. Although unstimulated purified B cells of tonsils and peripheral blood from healthy volunteers do not express detectable telomerase activity, anti-IgM beads induce telomerase activity in these B cells. Soluble anti-IgM antibody (Ab) alone does not induce telomerase activity, but the second signal, given by either one of the cytokines of interleukin-2 (IL-2), IL-4, and IL-13 or by anti-CD40 monoclonal Ab (MoAb), is effective as the costimulation for the induction of the activity. Stimulation with antiIgM Ab and anti-CD40 MoAb induces telomerase activity in most mature B cells of the tonsils and peripheral blood. The stimuli to both IgM and IgD receptors similarly induce the activity. Induction of telomerase activity is accompanied with the proliferation of B cells, but is not absolutely correlated with the extent of B-cell growth. Phorbol dibutylate (PDB) plus calcium (Ca) ionophore (PDB/Ca), which replace the activation through BCR and the costimulatory molecules, also induce telomerase activity. Moreover, it is suggested that phosphoinositide (PI) 3-kinase plays a role for the induction of telomerase activity in B cells stimulated with anti-IgM Ab and anti-CD40 MoAb. These results suggest that telomerase activity is induced in the B-cell activation of the antigen specific immune response.

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Non-Cognate Cytotoxic T Cells Can Be Retargeted Against CD20 Positive Tumour Cells Using [Fab' × MHC Class I/Peptide] Conjugates.

Blood ◽

10.1182/blood.v116.21.2838.2838 ◽

2010 ◽

Vol 116 (21) ◽

pp. 2838-2838

Author(s):

Angela D Hamblin ◽

Ben CR King ◽

Ruth R French ◽

Claude H Chan ◽

Alison L Tutt ◽

...

Keyword(s):

T Cells ◽

B Cells ◽

B Cell ◽

Mhc Class I ◽

Peripheral Blood ◽

Class I ◽

B Cell Depletion ◽

Cytokine Release ◽

Anti Cd20

Abstract Abstract 2838 To circumvent cytotoxic T lymphocyte (CTL) tolerance of tumour-associated antigens, the concept of redirecting CTLs against non-cognate targets has developed. One way of doing this is to use bispecific antibodies comprising anti-CD3 and anti-tumour antigen moieties. Unfortunately, this is frequently associated with unacceptable toxicity due to inflammatory cytokine release. As an alternative our approach has been to use a bivalent conjugate recognising a tumour antigen (through an antibody fragment) and a defined population of CTLs (specific for a single antigenic peptide e.g. viral epitope) through peptide presented in the context of recombinant MHC class I. We have produced a conjugate consisting of an anti-human CD20 Fab' fragment joined via a chemical crosslinker (succinimidyl-4-(N-maleimidomethyl) cyclohexane-1-carboxylate) to murine MHC class I/peptide (Kbα1-α3 domains/β2microglobulin presenting the ovalbumin-derived peptide SIINFEKL; expressed bacterially as a continuous polypeptide single chain trimer after Yu et al, J Immunol 2002). Size exclusion chromatography allowed purification of conjugates with [Fab':MHC class I/peptide] ratios of 1:1 and 2:1 (F2 and F3 respectively). In vitro both constructs were able to redirect the transgenic murine CTL line OT-1 (specific for KbSIINFEKL) to lyse human CD20+ tumour cells (lymphoblastoid Daudi cell line) at effector: target ratios of 10:1. This lysis could be blocked by the addition of 100 fold excess of either anti-CD20 F(ab')2 or the Kb/SIINFEKL-specific antibody 25D1. The constructs were also able to cause in vitro proliferation of naïve OT-1 cells (but not irrelevant CD8+ T cells) in the presence of human CD20+ cells in both thymidine incorporation and CFSE dilution assays. Using a human CD20 transgenic mouse model (Ahuja et al, J Immunol 2007) we have evaluated both constructs in vivo for their ability to redirect adoptively transferred OT-1 cells to deplete B cells from the peripheral blood. A single dose of 1 nmole F3 and 2 nmole F2 caused respectively up to 95% and 85% B cell depletion at day 7. The efficacy of lower doses suggested a dose: response relationship. As a marker of toxicity, we have measured cytokine levels at 2, 8 and 24 hours following a dose of 1 nmole F3 and compared them to those seen after administration of an [anti-CD3 × anti-CD20] bispecific F(ab')2 at a dose (0.5 nmole) which produced similar day 7 peripheral blood B cell depletion: phosphate-buffered saline was given as a negative control. Maximal cytokine release was seen at 2 hours with the levels of IL-4, IL-5, KC, IL-2 and IL-10 being lower after administration of the F3 than after the bispecific F(ab')2. However, interestingly, the F3 resulted in greater IL-12 release. Overall these data suggest that [Fab' × MHC class I/peptide] constructs have the potential to redirect non-cognate CTLs to deplete CD20+ malignant B cells from the peripheral blood and that this is associated with a lower level of cytokine release than a similarly efficacious dose of an anti-CD3-containing bispecific F(ab')2. Furthermore, the ability of [Fab' × MHC class I/peptide] constructs to cause proliferation of OT-1 cells in vitro suggests it may be possible to use a single molecule to both generate a secondary cytotoxic T cell response and subsequently to retarget it, increasing the viability of the approach if adopted in the clinic. Disclosures: No relevant conflicts of interest to declare.

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Steady-state generation of mucosal IgA+ plasmablasts is not abrogated by B-cell depletion therapy with rituximab

Blood ◽

10.1182/blood-2010-01-266536 ◽

2010 ◽

Vol 116 (24) ◽

pp. 5181-5190 ◽

Cited By ~ 57

Author(s):

Henrik E. Mei ◽

Daniela Frölich ◽

Claudia Giesecke ◽

Christoph Loddenkemper ◽

Karin Reiter ◽

...

Keyword(s):

Rheumatoid Arthritis ◽

Steady State ◽

B Cells ◽

B Cell ◽

Peripheral Blood ◽

Plasma Cells ◽

Cell Depletion ◽

B Cell Depletion ◽

Ki 67

AbstractThe anti-CD20 antibody rituximab depletes human B cells from peripheral blood, but it remains controversial to what extent tissue-resident B cells are affected. In representative patients with rheumatoid arthritis, we here demonstrate that recently activated presumably short-lived plasmablasts expressing HLA-DRhigh and Ki-67 continuously circulate in peripheral blood after B-cell depletion by rituximab at 26%-119% of their initial numbers. They circulate independent of splenectomy, express immunoglobulin A (IgA), β7 integrin, and C-C motif receptor 10 (CCR10) and migrate along CCL28 gradients in vitro, suggesting their mucosal origin. These plasmablasts express somatically hypermutated VH gene rearrangements and spontaneously secrete IgA, exhibiting binding to microbial antigens. Notably, IgA+ plasmablasts and plasma cells were identified in the lamina propria of patients treated with rituximab during peripheral B-cell depletion. Although a relation of these “steady state”–like plasmablasts with rheumatoid arthritis activity could not be found, their persistence during B-cell depletion indicates that their precursors, that is, B cells resident in the mucosa are not deleted by this treatment. These data suggest that a population of mucosal B cells is self-sufficient in adult humans and not replenished by CD20+ B cells immigrating from blood, lymphoid tissue, or bone marrow, that is, B cells depleted by rituximab.

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Arhgap25 deficiency leads to decreased numbers of peripheral blood B cells and defective germinal center reactions

10.1101/2020.03.05.965228 ◽

2020 ◽

Author(s):

Silke E. Lindner ◽

Colt A. Egelston ◽

Stephanie M. Huard ◽

Peter P. Lee ◽

Leo D. Wang

Keyword(s):

B Cells ◽

B Cell ◽

Germinal Center ◽

Peripheral Blood ◽

Cell Development ◽

B Cell Development ◽

Dependent Manner ◽

B Cell Differentiation ◽

Nucleotide Exchange

ABSTRACTRho family GTPases are critical for normal B cell development and function and their activity is regulated by a large and complex network of guanine nucleotide exchange factors (GEFs) and GTPase activating proteins (GAPs). However, the role of GAPs in B cell development is poorly understood. Here we show that the novel Rac-GAP ARHGAP25 is important for B cell development in mice in a CXCR4-dependent manner. We show that Arhgap25 deficiency leads to a significant decrease in peripheral blood B cell numbers, as well as defects in mature B cell differentiation. Arhgap25-/- B cells respond to antigen stimulation in vitro and in vivo but have impaired germinal center formation and decreased IgG1 class switching. Additionally, Arhgap25-/- B cells exhibit increased chemotaxis to CXCL12. Taken together, these studies demonstrate an important role for Arhgap25 in peripheral B cell development and antigen response.

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