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

2018 ◽  
Vol 92 (8) ◽  
pp. e00131-18 ◽  
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.

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

Blood ◽  
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.

scholarly journals Selective Targeting of T and B Cell Populations by Alemtuzumab in the Treatment of Multiple Sclerosis

2017 ◽  
Vol 12 (02) ◽  
pp. 78
Author(s):  
Nikolaos C Grigoriadis ◽  
Keyword(s):  
Multiple Sclerosis ◽  
T Cells ◽  
B Cells ◽  
B Cell ◽  
Cell Types ◽  
Tolerability Profile ◽  
T And B Cells ◽  
Brain Volume Loss ◽  
Inflammatory Phenotype ◽  
B Cell Subsets

Upstream targeting of both T and B cells is a rational therapeutic approach in multiple sclerosis (MS) in view of how both cell types and the interaction between them contribute to MS pathophysiology. This article will discuss this new way of thinking in MS: the targeting of both T and B cells, with a focus on the recently developed therapy, alemtuzumab (Lemtrada®, Genzyme, UK). Alemtuzumab depletes T and B lymphocytes, mainly via complement-dependent cytolysis and antibody-dependent cytolysis; depletion of B cells is not an enduring effect compared with the depletion of T cells. After dosing, CD4+ and CD8+ T cells and CD19 B cells decrease initially but increase over the following 11 months. During repopulation after alemtuzumab treatment, there is a shift in the relative proportions of T cell and B cell subsets whereby proportions of regulatory T cells and memory-phenotype T cells are increased and the proportion of naive T cells is decreased. A switch from a pro- to an anti-inflammatory phenotype and cytokine profile caused by alemtuzumab may underpin the long-lasting suppression of MS activity that has been observed in clinical trials. Alemtuzumab treatment is also associated with a consistently good safety and tolerability profile. Further, alemtuzumab appears to promote neurorehabilitation by improving measures of physical functioning, disability, measures of quality of life, and brain volume loss. Alemtuzumab therefore has the potential to reduce disease burden and improve substantially the prognosis for patients with MS.

CD19+,CD27+, IgM+ B Cells of Patients with Persistent Polyclonal B Cell (PPBL) Proliferate in a Low Intensity CD40-CD154 Interaction and Show Evidence of Immunoglobulin Isotype Switching

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 4920-4920
Author(s):  
Robert Delage ◽  
Emmanuelle Dugas-Bourdages ◽  
Annie Roy ◽  
Sonia Neron ◽  
Andre Darveau
Keyword(s):  
T Cells ◽  
B Cells ◽  
B Cell ◽  
B Lymphocytes ◽  
Cell Population ◽  
Genetic Defect ◽  
Memory B Cells ◽  
Isotype Switching ◽  
Low Intensity

Abstract Persistent polyclonal B cell lymphocytosis (PPBL) is a rare disorder characterized by an expansion of memory B cells CD19+, CD27+, IgM+. PPBL occurs mainly in female, is associated with HLA DR7, an increased level of serum IgM and the lymphocytes frequently show a bi-nucleated morphology. The patients have in most cases smoking habits and the clinical evolution is usually benign but we have previously described one case of lymphoma 19 years after a diagnosis of PPBL. Although the pathophysiology remains unknown, a familial occurrence is at the basis of this disorder suggesting a genetic defect. Moreover, multiple bcl-2\Ig gene rearrangements are present in all patients and an extra isochromosome 3 (i3)(q10) is frequently shown in the B cell population. The binding of CD40 to CD154 expressed on activated T cells plays a central role in B cell activation, proliferation and Ig isotype switching. We have previously shown that PPBL B lymphocytes were unable to respond to the proliferative signal delivered in vitro by CD40 in the CD40-CD154 system, indicating a possible defect in the CD40 pathway although CD40 expression, sequencing and tyrosine phosphorylation appeared normal. However, it has been shown recently that a reduced intensity of CD40-CD154 interaction in the presence of IL-2, IL-4 and IL-10 results in the proliferation, expansion and immunoglobulin secretion of normal memory CD19+,CD27+, IgM+ B cells. PPBL B lymphocytes sharing the same phenotype as normal memory B cells, we design a study to investigate the response of B lymphocytes from patient with PPBL in culture in high and low CD154 interaction. Proliferation and flow cytometry analysis of B lymphocytes from 6 patients with PPBL were closely monitored through a 14 day culture period and the Ig secretion was determined by Elisa. Our results show that a low intensity CD40- CD154 interaction in the presence of IL-2, IL-4 and IL-10 induces proliferation of the CD19+,CD27+,IgM+ PPBL population 6 to 20 times higher compared to high CD154 interaction. Interestingly, the CD19+, IgG+ cell population that constitutes less than 5% of the cell population at the beginning of the culture, increased over 25% on day 14. As for normal controls, we observed the emergence of a CD19+,CD27− cell population and the disappearance of surface IgD. Culture of B cells from patients with PPBL resulted in high Ig secretion. Moreover on day 14, Ig isotype analysis showed higher IgG levels compared to IgM. We conclude that PPBL B lymphocytes could proliferate in the CD40- CD154 system under proper condition and that proliferation also results in IgM and IgG secretion indicating an adequate CD40 signalling pathway. Moreover, this report provides the first evidence of in vitro Ig isotype switching of CD19+,CD27+,IgM+ B lymphocytes from PPBL. These results also suggest a possible defect in the interaction with T cells as observed in the hyper-IgM syndrome or alternatively, other cells from the microenvironment.

Mesenchymal Stromal Cells Suppress T-and B-Cells via Galectin-9 in a Donor Dependent Manner

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 1248-1248
Author(s):  
Christopher Ungerer ◽  
Patricia Quade-Lyssy ◽  
Reinhard Henschler ◽  
Erhard Seifried ◽  
Heinfried Radeke ◽  
...  
Keyword(s):  
T Cells ◽  
B Cells ◽  
Immune Suppression ◽  
Immune Cells ◽  
Stromal Cells ◽  
Coagulation Factor ◽  
T And B Cells ◽  
Ifn Γ

Abstract Abstract 1248 Therapeutic approaches using multipotent mesenchymal stromal cells (MSCs) are advancing in regenerative medicine, transplantation and autoimmune diseases. Until now the way of action for MSC-mediated immune suppression is still controversial and relies most probably on a multifactorial mechanism. MSCs have been demonstrated to produce the suppressive molecules hepatocyte growth factor (HGF), tumor growth factor-β (TGF-β), prostaglandin E2 (PGE2) and indoleamine 2,3-dioxygenase (IDO). Furthermore, it has been described that immunosuppression by MSCs is enhanced via stimulation with interferon-γ (IFN-γ). Recently, galectin-1, a β-galactoside binding lectin with immune modulatory properties, has been added to the group of immune modulatory molecules that are responsible for MSC-mediated immune suppression. Here, we identified galectin-9 (Gal-9) as a new molecule involved in MSC-mediated immune modulation. First, we isolated MSCs from bone marrow of randomly selected donors and performed several in vitro experiments regarding their immune modulatory potential (e.g proliferation and IgG production). Interestingly, Gal-9 was the only investigated protein, which was strongly upregulated in MSCs upon activation with IFN-γ. We moreover demonstrate that Gal-9 is a major mediator of the anti-proliferative effect of MSCs on T-cells. Although a B-cell suppressive function of Gal-9 has previously not been reported, we were surprised to detect the same inhibitory effect on isolated B-cells. Proliferation of immune cells was triggered upon either stimulation with either PHA and LPS, or CD40L and PHA. Activation of MSCs with IFN-γ resulted in a major decrease of proliferation of both T-cells and B-cells. In addition, Gal-9 and activated MSCs contribute to the suppression of VZV triggered immunoglobulin release as well. Again activation of MSCs with IFN-γ decreased the IgG release, whereas blocking Gal-9 with lactose, a well characterized inhibitor of Gal-9 function, reversed the effect almost completely. Further, we determined that Gal-9 expression levels (mRNA and protein) distinguish between MSC cultures from different donors after activation. Among donors, we could differentiate between individuals with high Gal-9 levels and higher immune modulatory potential and such with low Gal-9 expression and lower immune modulatory potential. Compared to untreated MSCs we demonstrated a three- to fifty-fold rise in Gal-9 levels after prior activation with IFN-γ. In addition, we demonstrated the upregulation of Gal-9 in MSCs by cell-cell contacts with either T-or B-cells. The upregulation was additionally at least two fold increased by previeously activating MSCs with IFN-γ. Because our group is interested in the therapy of hemophilia A and because of the unxpected suppressive effect of Gal-9 on B-cells and B-cell function, we next tested the effect of MSCs and Gal-9 on the induction of inhibitory antibodies to coagulation factor VIII (FVIII). Mice were immunized with human coagulation factor VIII (FVIII) in the presence or absence of either human MSCs, anti-murine Gal-9 or human Gal-9. As predicted, MSCs suppressed and anti-Gal-9 antibodies anhanced antibody formation. However in contrary to the expected, human Gal-9 co-treatment enhanced the anti-FVIII antibody response. A set of additional experiments revealed, that human Gal-9 suppresses murine regulatory T-cells in vivo. Further, in contrast to human immune cells, murine-derived T- and B-cells did not respond to human recombinant Gal-9 in vitro, but human IFN-γ activated MSCs were able to suppress proliferation of murine immune cells. Because of only 60% homology of murine and human Gal-9 we assume that the murine model cannot predict the function of human Gal-9 and that MSC-mediated immune modulatory functions are exerted via alternative pathways in this setting. Experiments with murine Gal-9 to demonstrate the in vivo function of Gal-9 are ongoing. In conclusion, Gal-9 is novel mediator of MSC immunomodulatory functions and affectsmultiple immune cell types including B-cells. Gal-9 is differentially expressed in MSCs from different donors and may therefore serve as a predictive indicator for clinical MSC functionality. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Selective B cell depletion upon intravenous infusion of replication-incompetent anti-CD19 CAR lentivirus

2020 ◽  
Author(s):  
Craig M. Rive ◽  
Eric Yung ◽  
Lisa Dreolini ◽  
Daniel J. Woodsworth ◽  
Robert A. Holt
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Peripheral Blood ◽  
Immune Cell ◽  
Wild Type ◽  
Immune Cell Subsets ◽  
Car T

AbstractAnti-CD19 CAR-T therapy for B cell malignancies has shown clinical success, but a major limitation is the logistical complexity and high cost of manufacturing autologous cell products. Direct infusion of viral gene transfer vectors to initiate in vivo CAR-T transduction, expansion and anti-tumor activity could provide an alternative, universal approach for CAR-T and related immune effector cell therapies that circumvents ex vivo cell manufacturing. To explore the potential of this approach we first evaluated human and murine CD8+ T cells transduced with VSV-G pseudotyped lentivectors carrying an anti-CD19CAR-2A-GFP transgene comprising either an FMC63 (human) or 1D3 (murine) anti-CD19 binding domain. To evaluate CD19 antigen-driven CAR-T proliferation in vitro we co-cultured transduced murine T cells with an excess of irradiated splenocytes and observed robust expansion over a 9 week period relative to control T cells transduced with a GFP transgene (mean fold expansion +/- SD: ID3-CD19CAR-GFP modified T cells, 12.2 +/- 0.09 (p < 0.001); FMC63-CD19CAR-GFP modified T cells 8.8 +/- 0.03 (p < 0.001). CAR-T cells isolated at the end of the expansion period showed potent B cell directed cytolytic activity in vitro. Next, we administered approximately 20 million replication-incompetent lentiviral particles carrying either ID3-CD19CAR-GFP, FMC63-CD19CAR-GFP, or GFP-only transgene to to wild-type C57BL/6 mice by tail vein infusion and monitored the dynamics of immune cell subsets isolated from peripheral blood at weekly intervals. We saw emergence of a persistent CAR-transduced CD3+ T cell population beginning week 3-4 that reaching a maximum of 13.5 +/- 0.58 % (mean +/- SD) and 7.8 +/- 0.76% of the peripheral blood CD3+ T cell population in mice infused with ID3-CD19CAR-GFP lentivector or FMC63-CD19CAR-GFP lentivector, respectively, followed by a rapid decline, in each case of, the B cell content of peripheral blood. Complete B cell aplasia was apparent by week 5 and was sustained until the end of the protocol (week 8). None of these changes were observed in mice infused with GFP-only control lentivector, and significant CAR positive populations were not observed within other immune cell subsets, including macrophage, natural killer, or B cells. Within the T cell compartment, CD8+ effector memory cells were the predominant CAR-positive subset. Modest weight loss of 5.5 +/- 2.97 % (mean +/- SD) observed in some animals receiving an anti-CD19CAR-GFP transgene during the protocol. These results indicate that direct IV infusion of lentiviral particles carrying an anti-CD19 CAR transgene can transduce T cells that then fully ablate endogenous B cells in wild type mice. Based on these results it may be useful to further explore, using currently available vectors, the feasibility of systemic gene therapy as a modality for CAR-T intervention.

EBV Can Induce Somatic Hypermutation in Naïve B Cells In Vitro but Ig Class Switching Requires T Cell Help.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 2370-2370
Author(s):  
Sridhar Chaganti ◽  
Noelia Begue Pastor ◽  
Gouri Baldwin ◽  
Claire Shannon-Lowe ◽  
Regina Feederle ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Germ Line ◽  
Somatic Hypermutation ◽  
Memory B Cells ◽  
Class Switching ◽  
B Cell Subsets ◽  
T Cell Help

Abstract Following primary infection, Epstein-Barr virus (EBV) establishes life long persistence in the host IgD− CD27+ memory B cell compartment rather than the IgD+ CD27+ marginal zone (MZ)-like or the IgD+ CD27− naïve B cell compartments. One possible explanation for such exclusive persistence in memory B cells is that EBV preferentially infects memory B cells. Alternatively, the virus may infect all B cell subsets but then drive MZ and naïve B cells to acquire the Ig isotype-switched phenotype and hypermutated Ig genotype of memory cells. Here we ask whether there is any evidence for one or other hypothesis from in vitro experiments. B cells from healthy donor blood samples were FACS sorted on the basis of IgD/CD27 expression into naïve, MZ, and memory B cell subsets with purities of >99%, >97% and >98% respectively. Analysis of the IgVH sequence further confirmed purity of the FACS sorted B cell subsets. Accordingly, 102 of 105 IgVH sequences amplified from purified naïve B cells were germ-line where as the vast majority of sequences amplified from MZ and memory B cells were mutated. All three B cell subsets expressed equal amounts of CD21 (EBV receptor on B cells), bound similar amounts of virus, and transformed with equal efficiency to establish B lymphoblastoid cell lines (LCLs) in vitro. Naïve B cell transformants upregulated CD27 expression but retained the IgM+, IgD+ phenotype as determined by FACS analysis and RT-PCR; MZ-B derived LCLs likewise were IgM+, IgD+, CD27+; and memory-B derived LCLs were consistently CD27+, IgD− and expressed either IgG, IgA or in some cases IgM. Therefore, EBV infection per se did not induce class switching. However, both naïve and MZ-B derived LCLs could still be induced to switch to IgG in the presence of CD40 ligand and IL-4; signals that are normally provided by T cells in vivo. To assess if EBV infection might drive Ig hypermutation, we carried out IgVH sequence analysis on the naïve-B derived LCL clones. Interestingly, 42 of 114 clonal IgVH sequences amplified from naïve-B derived LCLs had 3 or more mutations and the patterns of mutation seen were consistent with that produced by somatic hypermutation (SHM). Furthermore, within some naïve-B cell derived LCL clones, there were both germ-line and mutated sequences all sharing the same VDJ rearrangement (CDR3 sequence), again implying sequence diversification following EBV transformation of a single naïve B cell. Some intraclonal variation of the already hypermutated IgVH sequence was also noted in memory and MZ-B derived LCLs further suggesting ongoing mutational activity. Consistent with this, activation-induced cytidine deaminase (AID) expression was upregulated in transformants as assessed by real time RT-PCR. Our in vitro data is therefore compatible with a model of EBV persistence where the virus infects all mature B cell subsets but then drives infected naïve B cells to acquire a memory genotype by inducing SHM. In addition, EBV infected naïve and MZ-B cells may undergo Ig class switching to acquire the IgD− CD27+ memory phenotype in the presence of T cell help in vivo. EBV’s ability to induce SHM may also contribute to the lymphomagenic potential of the virus in addition to its B cell transforming and growth promoting properties.

B Cell With Regulatory Function Are Enriched Within Transitional and IgM Memory B Cell Subsets In Healthy Donors But Are Reduced and Functionally Impaired In Patients With Chronic Graft-Versus-Host Disease

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 4478-4478
Author(s):  
Anushruti Sarvaria ◽  
Ahmad Khoder ◽  
Abdullah Alsuliman ◽  
Claude Chew ◽  
Takuya Sekine ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Cd4 T Cells ◽  
Memory B Cells ◽  
Regulatory Function ◽  
Healthy Controls ◽  
Transitional B Cells ◽  
B Cell Subsets

The immunosuppressive function of IL10 producing regulatory B cells (Bregs) has been shown in several murine models of inflammation and autoimmune disease. However, there is a paucity of data regarding the existence of an equivalent regulatory B cell subset in healthy individuals and their potential role in the pathogenesis of chronic graft-versus-host disease (cGVHD) remains unknown. Here, we examined the functional regulatory properties of peripheral blood (PB)-derived human B cell subsets from healthy individuals. In addition, we carried out studies to explore their role in cGVHD, using B cells from patients following allogeneic stem cell transplantation (HSCT). We first determined whether human IL-10 producing B cells are enriched within any othe previously described human B cell subsets: CD19+IgM+CD27+ IgM memory, CD19+IgM-CD27+ switched memory, CD19+IgM+CD27- naive, and and transitional CD19+CD24hiCD38hi. Following in vitro stimulation with CD40 ligand, the majority of IL-10 producing B cells were found within the CD24hiCD38hi transitional and CD19+IgM+CD27+B cell subsets. We next assessed the regulatory properties of the PB-derived B cell subsets, by sort-purifying IgM memory (CD19+IgM+CD27+), switched memory (CD19+IgM-CD27+), naïve (CD19+IgM+CD27-) and transitional (CD19+CD24hiCD38hi) B cells from healthy controls, and cultured them 1:1 with autologous magnetic-bead purified CD4+ T cells. CD3/CD28 stimulated CD4+ T cells cultured with either CD19+IgM+CD27- naïve or CD19+IgM-CD27+ switched memory B cells proliferated to the same extent and produced equivalent amounts of IFN-γ to cultures containing CD4+ T cells alone. In contrast, culture of CD4+ T cells with IgM memory and transitional B cells significantly suppressed CD4+ T cell proliferation [median percent proliferating CD4+ T cells 52.5%; (33%-75%)] and 51% (25%-63%)], respectively when compared with CD3/CD28 stimulated CD4+ T cells (positive control) [89.5% (75%-92%], p=0.0001. The inhibitory effect of IgM memory and transitional B cells on CD4+ T cell proliferation was cell dose dependent with the highest suppression observed at a ratio of 1:1. These data suggest that human PB transitional and IgM memory B cells are endowed with regulatory function. We next examined if the in vitro suppressive effect of transitional and IgM memory B cells is mediated by regulatory T cells (Tregs). For this purpose, CD4+ T cells were depleted of CD127lo CD25hi CD4+ T cells by magnetic cell purification. B cell subsets were cultured with CD3/CD28 stimulated CD4+ CD25- T cells at a ratio of 1:1. IgM memory and transitional B cells were able to significantly suppress the proliferation and Th1 cytokine response by CD4+ CD25- T cells compared to cultures containing CD4+ CD25-T cells alone, indicating that the suppressive activity of Bregs is independent of Tregs. To further understand the underlying mechanims though which Bregs exert T-cell suppression, we used antibody blockade experiments and showed that this suppressive effect was mediated partially via the provision of IL-10, but not TGF-ß. Using transwell experiments, we further determined that the suppressive function of Bregs is also partly dependent on direct T cell/B cell contact. We next assessed whether the activity of Breg cells might be altered in patients with cGVHD. B cells from patients with cGVHD were refractory to CD40 stimulation and produced less IL-10 when compared to patients without cGVHD post-SCT and healthy controls, [1.02% (0.22-2.26) vs.1.72% (0.8-5.52) vs. 2.16 (1.3- 5.6), p=0.001]. Likewise, the absolute number of IL-10 producing B cells was significantly lower in cGvHD patients compared to patients without cGVHD and healthy controls (p=0.007), supporting both a qualitative and quantitative defect in IL-10 producing B cells in cGvHD. Our combined studies provide important new data defining the phenotype of B cell populations enriched in regulatory B cells in healthy humans and provide evidence for a defect in the activity of such cells in patients with cGVHD post-SCT. In association with previous reports showing defects in Treg cell activity in GVHD, our results suggest the existence of a broad range of deficiencies in immune regulatory cell function in cGvHD patients. * Both Anushruti Sarvaria and Ahmad K contributed equally. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Cell-to-cell interaction controlled by immunoglobulin genes. Role of Thy-1-, Lyt-1+, Ig+ (B') cell in allotype-restricted antibody production.

1982 ◽  
Vol 156 (2) ◽  
pp. 443-453 ◽  
Author(s):  
K Okumura ◽  
K Hayakawa ◽  
T Tada
Keyword(s):  
T Cells ◽  
B Cells ◽  
Antibody Response ◽  
B Cell ◽  
Antibody Production ◽  
Helper T Cells ◽  
Secondary Antibody ◽  
T And B Cells ◽  
Athymic Nude Mice

A novel lymphocyte subpopulation, designated "B' cell" because of its characteristic dual expression of Ig and Lyt-1 antigen, is described in relation to its ability to augment the in vitro secondary antibody response. The cells are found in the spleens of normal unprimed mice as well as those of athymic nude mice and represent a small of normal unprimed mice as well as those of athymic nude mice and represent a small number (2-3%) of immunoglobulin-positive cells. No other distinguishing surface markers of conventional T and B cells, such as Thy-1, Lyt-2, Ia, and ThB antigens, are detected on the B' cell. In the in vitro anti-hapten secondary antibody response, the addition of a small number of B' cells from unprimed mice to the mixture of T and B cells greatly augmented the anti-hapten antibody formation when the number of carrier-specific helper T cells was limited. This augmentation was observed only when B and B' cells shared the same set of IgVH genes. The identity of the immunoglobulin gene between T cells and B or B' cells was not necessary for optimum antibody production. The results indicate that the presence of B' cells is necessary for the induction of an optimum antibody response when helper T cells are limited. It is suggested that B' cells deliver an additional signal to the B cell network to magnify the antibody response.

scholarly journals Collaboration of allogeneic T and B lymphocytes in the primary antibody response to sheep erythrocytes in vitro.

1975 ◽  
Vol 142 (4) ◽  
pp. 928-935 ◽  
Author(s):  
E Heber-Katz ◽  
D B Wilson
Keyword(s):  
T Cells ◽  
B Cells ◽  
B Cell ◽  
Dose Response ◽  
Cell Activation ◽  
B Cell Activation ◽  
T And B Cells ◽  
Response Curves ◽  
Major Histocompatibility Complex Haplotype

This study provides a direct quantitative comparison of the helper effects of allogeneic and syngeneic rat T cells in the production of direct SRBC plaque-forming cell (PFC) responses by B cells in culture. In syngeneic T-B combinations, log-log plots of the number of PFC generated after 5.5 days in culture vs. the number of T cells employed as helpers showed a linear response between 10(4) and 2.5 times 10(5) T cells added. Allogeneic T-B combinations, in which the T cells possess the capacity for reactivity to major alloantigens of the B-cell donor, showed a different dose/response relationship in which PFC responses were decreased at high T/B ratios and augmented at low T/B rations. In this system responses were detected with as few as 10(3) allogeneic T cells. Use of negatively selected allogeneic T populations, specifically depleted of mixed lymphocyte interaction (MLI) and graft-vs-host reactivity for B-cell alloantigens, as helpers gave dose/response curves quantitatively identical to responses with syngeneic T-B combinations and also with F1 T-cell parental B-cell combinations. These data indicate that rat T and B cells need not share a major histocompatibility complex haplotype in order to collaborate effectively in a primary direct PFC response to SRBC in culture. In addition, the PFC response required the combinaed presence of T and B cells as well as antigen in the cultures, a finding consistent with the two signal model of B-cell activation. Finally, the dose/response data obtained suggest the possibility that although SRBC antigen is required in the cultures helper activity with low numbers of normal allogeneic T cells may not depend on T cells having specificity for this antigen.

Pharmacodynamic Effects of Administration of Maytansine Conjugated Anti-CD22 Monoclonal Antibodies to Cynomolgus Monkeys

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 4996-4996
Author(s):  
Franklin Fuh ◽  
Reina Fuji ◽  
Kirsten A Poon ◽  
Dongwei Li ◽  
Clarissa David ◽  
...  
Keyword(s):  
T Cells ◽  
B Cells ◽  
B Cell ◽  
Peripheral Blood ◽  
Lymphoid Tissues ◽  
Marginal Zone B Cells ◽  
Cynomolgus Monkeys ◽  
Pharmacodynamic Effects ◽  
Pharmacodynamic Profile ◽  
B Cell Subsets

Abstract CD22 is a B cell-specific glycoprotein expressed on the cell surface of all mature B cells. A candidate therapeutic anti-CD22 antibody, 10F4v3, was conjugated to the anti-mitotic agent maytansine (10F4v3-DM1). DM1 disrupts cellular mitosis through inhibition of tubulin polymerization when internalized into cells. The anti-CD22 DM1 conjugate was shown to have significant potency in preclinical efficacy models of NHL. In order to further characterize this antibody-drug conjugate in preclinical studies, we first evaluated the binding characteristics of the 10F4v3 to peripheral blood B cells from various geographical sources of cynomolgus monkeys. 10F4v3 bound to peripheral blood B cells from all cynomolgus monkeys of Indonesian and Mauritian origins, but displayed only limited binding to cynomolgus monkeys of Chinese and Cambodian origins. Therefore, further pre-clinical evaluation of 10F4v3-DM1 was conducted in Indonesian cynomolgus monkeys to examine the safety, pharmacokinetic, and pharmacodynamic effects in monkeys dosed at 10, 20, and 30 mg/kg (2000, 4000, and 6000 mg/m2 DM1). Pharmacodynamic assessments of peripheral blood and lymphoid tissues included examination of B cells, B cell subsets, CD4+ T cells, CD8+ T cells, and CD3−CD20− (NK) cells. B cell subsets included CD20+, CD20+CD21+, CD20+CD21−, CD20+CD21+CD27+, CD20+CD21+CD27−, and CD20+CD21high lymphocytes which are phenotypically similar to human B cells, mature B cells, germinal center B cells, memory B cells, naïve B cells, and marginal zone B cells, respectively. B cells and B cell subsets were substantially depleted in peripheral blood at all doses, with no apparent dose-dependent effects. In lymphoid tissue, B cells were also depleted, with substantial depletion of CD20+CD21− and CD20+CD21high B cell subsets in spleen and bone marrow. Based on the nonclinical data, 10F4v3-DM1 exhibits an encouraging pharmacodynamic profile that supports clinical development for the potential treatment of non-Hodgkin’s lymphoma.

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