scholarly journals Human immune response to group A streptococcal carbohydrate (A-CHO). I. Quantitative and qualitative analysis of the A-CHO-specific B cell population responding in vitro to polyclonal and specific activation.

1985 ◽  
Vol 161 (3) ◽  
pp. 547-562 ◽  
Author(s):  
F Emmrich ◽  
B Schilling ◽  
K Eichmann
Keyword(s):  
Immune Response ◽  
B Cells ◽  
B Cell ◽  
Peripheral Blood ◽  
Pokeweed Mitogen ◽  
Specific Antibodies ◽  
Group A ◽  
Precursor B Cells

The immune response to the group-specific carbohydrate of group A streptococci (A-CHO) provides an informative in vitro model for the investigation of several aspects of human anticarbohydrate immune responses. A-CHO-specific B cells can be polyclonally activated by pokeweed mitogen (PWM), and, specifically, by in vitro immunization with streptococcal vaccine. High levels of A-CHO-specific antibodies, mainly directed to the immunodominant side chain N-acetyl-D-glucosamine (GlcNAc), occur in healthy adult individuals. Serum antibody levels are reflected in high frequencies of precursor B cells among peripheral blood lymphocytes. In one particular case, greater than 15% of all B cells activated by PWM for IgM production were found to produce IgM anti-A-CHO antibodies, as determined in limiting dilution experiments, as well as by analyzing Ig concentrations in bulk culture experiments. The case with the lowest proportion observed had 0.3% A-CHO-specific B cells among IgM-producing B cells. Preferential PWM activation of anti-A-CHO-producing B cells could be excluded. The comparison of the proportions of anti-A-CHO IgM produced in vivo, and of B cells producing antibodies of this specificity in peripheral blood, suggests a similar distribution of specific precursor B cells in the antibody-producing lymphoid tissue compartments and in peripheral blood. However, nearly all specific antibodies produced in vitro belong to the IgM isotype, whereas IgG anti-A-CHO in high amounts, mostly exceeding the specific IgM, was found only among anti-A-CHO antibodies produced in vivo. Low anti-A-CHO IgG production was seen in polyclonally activated as well as in antigen-activated cultures, whereas, in contrast, total IgG was produced in considerable amounts after polyclonal activation. This suggests a different distribution pattern, and/or diverse differentiation requirements for anti-A-CHO-producing B cells, compared with other B cell species.

Extensive in vivo self-renewal, long-term reconstitution capacity, and hematopoietic multipotency of Pax5-deficient precursor B-cell clones

Blood ◽  
2002 ◽  
Vol 99 (8) ◽  
pp. 2760-2766 ◽  
Author(s):  
Christoph Schaniel ◽  
Marie Gottar ◽  
Eddy Roosnek ◽  
Fritz Melchers ◽  
Antonius G. Rolink
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
B Cell ◽  
Telomerase Activity ◽  
Self Renewal ◽  
Cell Clones ◽  
Precursor B Cells

Abstract Self-renewal, pluripotency, and long-term reconstitution are defining characteristics of single hematopoietic stem cells.Pax5−/− precursor B cells apparently possess similar characteristics. Here, using serial transplantations, with in vitro recloning and growth of the bone marrow–homed donor cells occurring after all transplantations, we analyzed the extent of self-renewal and hematopoietic multipotency ofPax5−/− precursor B-cell clones. Moreover, telomere length and telomerase activity in these clones was analyzed at various time points. Thus far, 5 successive transplantations have been performed. Clones transplanted for the fifth time, which have proliferated for more than 150 cell divisions in vitro, still repopulate the bone marrow with precursor B cells and reconstitute these recipients with lymphoid and myeloid cells. During this extensive proliferation, Pax5−/− precursor B cells shorten their telomeres at 70 to 90 base pairs per division. Their telomerase activity remains at 3% of that of HEK293 cancer cells during all serial in vivo transplantations/in vitro expansions. Together, these data show thatPax5−/− precursor B-cell clones possess extensive in vivo self-renewal capacity, long-term reconstitution capacity, and hematopoietic multipotency, with their telomeres shortening at the normal rate.

Differential effects of epratuzumab on peripheral blood B cells of patients with systemic lupus erythematosus versus normal controls

2007 ◽  
Vol 67 (4) ◽  
pp. 450-457 ◽  
Author(s):  
A M Jacobi ◽  
D M Goldenberg ◽  
F Hiepe ◽  
A Radbruch ◽  
G R Burmester ◽  
...  
Keyword(s):  
Systemic Lupus Erythematosus ◽  
B Cells ◽  
B Cell ◽  
Lupus Erythematosus ◽  
Peripheral Blood ◽  
Therapeutic Option ◽  
Normal Subjects ◽  
Systemic Lupus

Objective:B lymphocytes have been implicated in the pathogenesis of lupus and other autoimmune diseases, resulting in the introduction of B cell-directed therapies. Epratuzumab, a humanised anti-CD22 monoclonal antibody, is currently in clinical trials, although its effects on patients’ B cells are not completely understood.Methods:This study analysed the in vivo effect of epratuzumab on peripheral B cell subsets in 12 patients with systemic lupus erythematosus, and also addressed the in vitro effects of the drug by analysing anti-immunoglobulin-induced proliferation of isolated B cells obtained from the peripheral blood of 11 additional patients with lupus and seven normal subjects.Results:Upon treatment, a pronounced reduction of CD27– B cells and CD22 surface expression on CD27– B cells was observed, suggesting that these cells, which mainly comprise naïve and transitional B cells, are preferentially targeted by epratuzumab in vivo. The results of in vitro studies indicate additional regulatory effects of the drug by reducing the enhanced activation and proliferation of anti-immunoglobulin-stimulated lupus B cells after co-incubation with CD40L or CpG. Epratuzumab inhibited the proliferation of B cells from patients with systemic lupus erythematosus but not normal B cells under all culture conditions.Conclusions:Epratuzumab preferentially modulates the exaggerated activation and proliferation of B cells from patients with lupus in contrast to normal subjects, thus suggesting that epratuzumab might offer a new therapeutic option for patients with systemic lupus erythematosus, as enhanced B cell activation is a hallmark of this disease.

scholarly journals Regulation of memory B-cell survival by the BH3-only protein Puma

Blood ◽  
2011 ◽  
Vol 118 (15) ◽  
pp. 4120-4128 ◽  
Author(s):  
Cyril Clybouw ◽  
Silke Fischer ◽  
Marie Thérèse Auffredou ◽  
Patricia Hugues ◽  
Catherine Alexia ◽  
...  
Keyword(s):  
Immune Response ◽  
B Cells ◽  
B Cell ◽  
Cell Survival ◽  
Family Member ◽  
Cell Immune Response ◽  
Memory B Cell ◽  
B Cell Survival

Abstract Apoptosis is crucial for immune system homeostasis, including selection and survival of long-lived antibody-forming cells and memory cells. The interactions between proapoptotic and pro-survival proteins of the Bcl-2 family are critical for this process. In this report, we show that expression of the proapoptotic BH3-only Bcl-2 family member Puma was selectively up-regulated on in vitro activation with antigens or mitogens of both human and mouse B cells. Puma expression coincided in vivo, with the prosurvival Bcl-2 family member Mcl-1 within the germinal centers and its expression correlates with the germinal center like phenotype of Burkitt lymphoma. Experiments performed in Puma-deficient mice revealed that Puma is essential for apoptosis of mitogen-activated B cells in vitro and for the control of memory B-cell survival. In conclusion, using both human and murine models, our data show that Puma has a major role in the T cell– dependent B-cell immune response. These data demonstrate that Puma is a major regulator of memory B lymphocyte survival and therefore a key molecule in the control of the immune response.

scholarly journals Production of colony-stimulating activity by normal and neoplastic human B lymphocytes

Blood ◽  
1987 ◽  
Vol 69 (5) ◽  
pp. 1340-1347 ◽  
Author(s):  
V Pistoia ◽  
R Ghio ◽  
S Roncella ◽  
F Cozzolino ◽  
S Zupo ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
B Lymphocytes ◽  
Peripheral Blood ◽  
Lymphocytic Leukemia ◽  
Target Cells ◽  
Sephadex Column ◽  
Colony Stimulating Activity

Abstract Normal human B cells were purified from peripheral blood or tonsils and tested for their ability to release colony-stimulating activity (CSA) in short-term cultures. The target cells used in the CSA assays were from peripheral blood or bone marrow. Unstimulated B cells produced CSA in amounts similar to those present in the GCT-conditioned medium used as a positive control. The B cell-derived CSA predominantly promoted the growth of colonies that contained macrophages alone or macrophages and granulocytes. CSA eluted in a single peak from a G-75 Sephadex column with an approximate molecular weight (mw) of 65 to 70 kilodaltons (kd). Fractionation of tonsil B lymphocytes on Percoll density gradients showed that large B cells, probably already activated in vivo, were the main source of CSA. By contrast, small, resting B cells recovered from a different fraction of the Percoll gradient released minimum amounts or no CSA. However, these B cells became CSA producers following stimulation with Staphylococcus aureus Cowan (SAC) in vitro. B cells purified from the peripheral blood of nine out of 12 patients with B-cell chronic lymphocytic leukemia (B-CLL) also released CSA in vitro in the absence of stimuli. These findings suggest that by releasing CSA, B cells may have a role in the regulation of hematopoiesis and in the control of the inflammatory process.

Rescue of B Cells Development in an Animal Model of X-Linked Agammaglobulinemia(XLA) Via B Lineage-Specific Lentiviral Gene Therapy.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1287-1287
Author(s):  
S. Humblet-Baron ◽  
W. Zhang ◽  
K. Kipp ◽  
S. Khim ◽  
J. Jarjour ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Peripheral Blood ◽  
Lentiviral Vector ◽  
Cell Development ◽  
B Cell Development ◽  
Gfp Expression ◽  
B Lineage

Abstract X-linked agammaglobulinemia (XLA) is a human immunodeficiency caused by mutations in Bruton’s tyrosine kinase (Btk) and characterized by an arrest in early B-cell development, absence of serum immunoglobulin, and recurrent bacterial infections. Using Btk and Tec double deficient (Btk/Tec−/ −) mice as a model for XLA, we recently showed that onco-retroviral-mediated Btk gene transfer into hematopoietic stem cells (HSC) reconstituted in vivo Btk-dependent B-cell development and function (Yu et al. Blood 104(5):1281–90). In order to increase the safety of this approach, we developed a SIN-lentiviral vector with a B cell specific enhancer/promoter element, Eμ B29. Using SIN-lentiviral vectors expressing GFP, we observed that Eμ B29 consistently promoted 3–5 fold higher GFP expression in human B lineage cells derived from transduced HSC in vitro and in vivo (ASGT 2002 abstract #1302). We also evaluated this vector, CSOM-Eμ B29-GFP-WPRE, in lentiviral transgenic mice where it exhibited the highest GFP expression in peripheral B cells compared with all other hematopoietic lineages. Specifically, in more than 8 independent founder strains the MFI for GFP expression in B cells was > 3 fold higher than that in T cells (p=0.0002). Based upon these findings we developed Eμ B29-huBtk SIN-lentiviral vectors with or without the insulator element derived from the chicken β-globulin insulator (HS4). Using both vectors to transduce Btk −/ − DT40 B cells, followed by cloning by limiting dilution, we demonstrate Btk protein expression by intracellular staining and western blotting and full rescue of Btk-dependent, B cell receptor (BCR)-mediated Ca2+ signaling in all clones evaluated including those exhibiting a single viral integration. Next we tested the capacity of these vectors to reconstitute Btk-dependent B-cell development and function in a cohort of Btk/Tec−/ − mice. Marrow from 5-FU treated Btk/Tec −/ − mice was harvested, cultured on fibronectin coated plates with growth factors (mIL-3,mIL-6, mSCF, mTPO and mFLT3ligand) and concentrated lentivirus (2.3x107pg/106 cells measured by p24 level). After 48h of in vitro culture, cells were transplanted into lethally irradiated animals and transplanted animals were serially evaluated for presence of B cells in the peripheral blood. B-cell numbers progressively increased with a significant difference as early as within 6 weeks in mice receiving transduced (16–18% B220+ cells) vs. control marrow (8–9%; mock transduced). Further, mature B cells (B220+IgMlowIgDhi) represented 14–20% of total B cells in treated compared to <5% in control mice. Finally, mice receiving transduced cells exhibited a rescue of total serum IgM and IgG3 levels and responses to TI-II dependent immunization. Results of two additional animal cohorts will be presented. In summary, our data demonstrate that Eμ B29-Btk SIN-lentiviral vector specifically promotes Btk expression in B lineage cells, and correction of the Btk-deficient phenotype in vitro and in vivo. Peripheral blood B cells were analyzed for relative IgM and IgD expression at 6 weeks post reconstitution. Representative data from animals receiving mock-vs/ EμB29-Btk transduced marrow are shown. Upper left quadrant shows percentage of circulating mature B cells. Peripheral blood B cells were analyzed for relative IgM and IgD expression at 6 weeks post reconstitution. Representative data from animals receiving mock-vs/ EμB29-Btk transduced marrow are shown. Upper left quadrant shows percentage of circulating mature B cells.

scholarly journals Production of colony-stimulating activity by normal and neoplastic human B lymphocytes

Blood ◽  
1987 ◽  
Vol 69 (5) ◽  
pp. 1340-1347
Author(s):  
V Pistoia ◽  
R Ghio ◽  
S Roncella ◽  
F Cozzolino ◽  
S Zupo ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
B Lymphocytes ◽  
Peripheral Blood ◽  
Lymphocytic Leukemia ◽  
Target Cells ◽  
Sephadex Column ◽  
Colony Stimulating Activity

Normal human B cells were purified from peripheral blood or tonsils and tested for their ability to release colony-stimulating activity (CSA) in short-term cultures. The target cells used in the CSA assays were from peripheral blood or bone marrow. Unstimulated B cells produced CSA in amounts similar to those present in the GCT-conditioned medium used as a positive control. The B cell-derived CSA predominantly promoted the growth of colonies that contained macrophages alone or macrophages and granulocytes. CSA eluted in a single peak from a G-75 Sephadex column with an approximate molecular weight (mw) of 65 to 70 kilodaltons (kd). Fractionation of tonsil B lymphocytes on Percoll density gradients showed that large B cells, probably already activated in vivo, were the main source of CSA. By contrast, small, resting B cells recovered from a different fraction of the Percoll gradient released minimum amounts or no CSA. However, these B cells became CSA producers following stimulation with Staphylococcus aureus Cowan (SAC) in vitro. B cells purified from the peripheral blood of nine out of 12 patients with B-cell chronic lymphocytic leukemia (B-CLL) also released CSA in vitro in the absence of stimuli. These findings suggest that by releasing CSA, B cells may have a role in the regulation of hematopoiesis and in the control of the inflammatory process.

scholarly journals Radiosensitivity of peripheral blood B cells in pigs

2009 ◽  
Vol 54 (No. 5) ◽  
pp. 223-235 ◽  
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 <I>in vitro</I> experiments with the <I>in vivo</I> 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 <I>in vitro</I> 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 <I>in vitro</I> and lymphocyte dynamics in peripheral blood of irradiated animals clearly document the limits of <I>in vitro</I> 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.

scholarly journals Differential Regulation of Mouse B Cell Development by Transforming Growth Factor β1

2002 ◽  
Vol 9 (2) ◽  
pp. 86-95 ◽  
Author(s):  
Denise A. Kaminski ◽  
John J. Letterio ◽  
Peter D. Burrows
Keyword(s):  
B Cells ◽  
Growth Factor ◽  
B Cell ◽  
Transforming Growth Factor ◽  
Plasma Cells ◽  
Population Analysis ◽  
Cell Development ◽  
B Cell Development

Transforming growth factor β (TGFβ) can inhibit thein vitroproliferation, survival and differentiation of B cell progenitors, mature B lymphocytes and plasma cells. Here we demonstrate unexpected, age-dependent reductions in the bone marrow (BM) B cell progenitors and immature B cells in TGFβ1-/-mice. To evaluate TGFβ responsiveness during normal B lineage development, cells were cultured in interleukin 7 (IL7)±TGFβ. Picomolar doses of TGFβ1 reduced pro-B cell recoveries at every timepoint. By contrast, the pre-B cells were initially reduced in number, but subsequently increased compared to IL7 alone, resulting in a 4-fold increase in the growth rate for the pre-B cell population. Analysis of purified BM sub-populations indicated that pro-B cells and the earliest BP1-pre-B cells were sensitive to the inhibitory effects of TGFβ1. However, the large BP1+pre-B cells, although initially reduced, were increased in number at days 5 and 7 of culture. These results indicate that TGFβ1 is important for normal B cell developmentin vivo, and that B cell progenitors are differentially affected by the cytokine according to their stage of differentiation.

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

Blood ◽  
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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