scholarly journals Oct2 enhances antibody-secreting cell differentiation through regulation of IL-5 receptor α chain expression on activated B cells

2008 ◽  
Vol 205 (2) ◽  
pp. 409-421 ◽  
Author(s):  
Dianne Emslie ◽  
Kathy D'Costa ◽  
Jhagvaral Hasbold ◽  
Donald Metcalf ◽  
Kiyoshi Takatsu ◽  
...  
Keyword(s):  
Cell Differentiation ◽  
B Cells ◽  
Target Genes ◽  
B Lymphocyte ◽  
Plasma Cells ◽  
Ectopic Expression ◽  
Cell Receptor ◽  
Α Chain ◽  
Activated B Cells

Mice lacking a functional gene for the Oct2 transcriptional activator display several developmental and functional deficiencies in the B lymphocyte lineage. These include defective B cell receptor (BCR) and Toll-like receptor 4 signaling, an absence of B-1 and marginal zone populations, and globally reduced levels of serum immunoglobulin (Ig) in naive and immunized animals. Oct2 was originally identified through its ability to bind to regulatory regions in the Ig loci, but genetic evidence has not supported an essential role for Oct2 in the expression of Ig genes. We describe a new Oct2-mediated role in B cells. Oct2 augments the ability of activated B cells to differentiate to antibody-secreting plasma cells (ASCs) under T cell–dependent conditions through direct regulation of the gene encoding the α chain of the interleukin (IL) 5 receptor. Ectopic expression of IL-5Rα in oct2-deficient B cells largely restores their ability to differentiate to functional ASCs in vitro but does not correct other phenotypic defects in the mutants, such as the maturation and specialization of peripheral B cells, which must therefore rely on distinct Oct2 target genes. IL-5 augments ASC differentiation in vitro, and we show that IL-5 directly activates the plasma cell differentiation program by enhancing blimp1 expression.

scholarly journals Optimized Protocols for in Vitro T Cell-Dependent and T Cell-Independent Activation for B Cell Differentiation Studies Using Limited Cells

2021 ◽  
Author(s):  
Casper Marsman ◽  
Dorit Verhoeven
Keyword(s):  
Cell Differentiation ◽  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Plasma Cells ◽  
Cell Formation ◽  
B Cell Differentiation ◽  
Differentiation Potential ◽  
Human B Cell

Background/methods: For mechanistic studies, in vitro human B cell differentiation and generation of plasma cells are invaluable techniques. However, the heterogeneity of both T cell-dependent (TD) and T cell-independent (TI) stimuli and the disparity of culture conditions used in existing protocols makes interpretation of results challenging. The aim of the present study was to achieve the most optimal B cell differentiation conditions using isolated CD19+ B cells and PBMC cultures. We addressed multiple seeding densities, different durations of culturing and various combinations of TD stimuli and TI stimuli including B cell receptor (BCR) triggering. B cell expansion, proliferation and differentiation was analyzed after 6 and 9 days by measuring B cell proliferation and expansion, plasmablast and plasma cell formation and immunoglobulin (Ig) secretion. In addition, these conditions were extrapolated using cryopreserved cells and differentiation potential was compared. Results: This study demonstrates improved differentiation efficiency after 9 days of culturing for both B cell and PBMC cultures using CD40L and IL-21 as TD stimuli and 6 days for CpG and IL-2 as TI stimuli. We arrived at optimized protocols requiring 2500 and 25.000 B cells per culture well for TD and TI assays, respectively. The results of the PBMC cultures were highly comparable to the B cell cultures, which allows dismissal of additional B cell isolation steps prior to culturing. In these optimized TD conditions, the addition of anti-BCR showed little effect on phenotypic B cell differentiation, however it interferes with Ig secretion measurements. Addition of IL-4 to the TD stimuli showed significantly lower Ig secretion. The addition of BAFF to optimized TI conditions showed enhanced B cell differentiation and Ig secretion in B cell but not in PBMC cultures. With this approach, efficient B cell differentiation and Ig secretion was accomplished when starting from fresh or cryopreserved samples. Conclusion: Our methodology demonstrates optimized TD and TI stimulation protocols for more indepth analysis of B cell differentiation in primary human B cell and PBMC cultures while requiring low amounts of B cells, making them ideally suited for future clinical and research studies on B cell differentiation of patient samples from different cohorts of B cell-mediated diseases.

scholarly journals Modulation of spontaneous B-cell differentiation in macroglobulinemia by retinoic acid

Blood ◽  
1994 ◽  
Vol 83 (8) ◽  
pp. 2206-2210 ◽  
Author(s):  
Y Levy ◽  
S Labaume ◽  
MC Gendron ◽  
JC Brouet
Keyword(s):  
Retinoic Acid ◽  
Cell Differentiation ◽  
B Cells ◽  
B Cell ◽  
Plasma Cells ◽  
B Cell Differentiation ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid ◽  
Culture Supernatants

Abstract We previously showed that clonal blood B cells from patients with macroglobulinemia spontaneously differentiate in vitro to plasma cells. This process is dependent on an interleukin (IL)-6 autocrine pathway. We investigate here whether all-trans-retinoic acid (RA) interferes with B-cell differentiation either in patients with IgM gammapathy of undetermined significance (MGUS) or Waldenstrom's macroglobulinemia (WM). RA at a concentration of 10(-5) to 10(-8) mol/L inhibited by 50% to 80% the in vitro differentiation of purified B cells from four of five patients with MGUS and from one of five patients with WM as assessed by the IgM content of day 7 culture supernatants. We next determined whether this effect could be related to an inhibition of IL- 6 secretion by cultured B cells and/or a downregulation of the IL-6 receptor (IL-6R), which was constitutively expressed on patients' blood B cells. A 50% to 100% (mean, 80%) inhibition of IL-6 production was found in seven of 10 patients (five with MGUS and two with WM). The IL- 6R was no more detectable on cells from patients with MGUS after 2 days of treatment with RA and slightly downregulated in patients with WM. It was of interest that B cells susceptible to the action of RA belonged mostly to patients with IgM MGUS, which reinforces our previous data showing distinct requirements for IL-6-dependent differentiation of blood B cells from patients with VM or IgM MGUS.

scholarly journals B Lymphocyte Chemotaxis Regulated in Association with Microanatomic Localization, Differentiation State, and B Cell Receptor Engagement

1998 ◽  
Vol 187 (5) ◽  
pp. 753-762 ◽  
Author(s):  
Conrad C. Bleul ◽  
Joachim L. Schultze ◽  
Timothy A. Springer
Keyword(s):  
B Cells ◽  
B Cell ◽  
B Lymphocytes ◽  
Messenger Rna ◽  
B Lymphocyte ◽  
Somatic Hypermutation ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Cxc Chemokine

Migration of mature B lymphocytes within secondary lymphoid organs and recirculation between these sites are thought to allow B cells to obtain T cell help, to undergo somatic hypermutation, to differentiate into effector cells, and to home to sites of antibody production. The mechanisms that direct migration of B lymphocytes are unknown, but there is evidence that G protein–coupled receptors, and possibly chemokine receptors, may be involved. Stromal cell– derived factor (SDF)-1α is a CXC chemokine previously characterized as an efficacious chemoattractant for T lymphocytes and monocytes in peripheral blood. Here we show with purified tonsillar B cells that SDF-1α also attracts naive and memory, but not germinal center (GC) B lymphocytes. Furthermore, GC B cells could be converted to respond to SDF-1α by in vitro differentiation into memory B lymphocytes. Conversely, the migratory response in naive and memory B cells was significantly reduced after B cell receptor engagement and CD40 signaling. The receptor for SDF-1, CXC chemokine receptor 4 (CXCR4), was found to be expressed on responsive as well as unresponsive B cell subsets, but was more rapidly downregulated on responsive cells by ligand. Finally, messenger RNA for SDF-1 was detected by in situ hybridization in a layer of cells surrounding the GC. These findings show that responsiveness to the chemoattractant SDF-1α is regulated during B lymphocyte activation, and correlates with positioning of B lymphocytes within a secondary lymphoid organ.

scholarly journals An in vitro model of differentiation of memory B cells into plasmablasts and plasma cells including detailed phenotypic and molecular characterization

Blood ◽  
2009 ◽  
Vol 114 (25) ◽  
pp. 5173-5181 ◽  
Author(s):  
Michel Jourdan ◽  
Anouk Caraux ◽  
John De Vos ◽  
Geneviève Fiol ◽  
Marion Larroque ◽  
...  
Keyword(s):  
B Cells ◽  
In Vitro Model ◽  
Plasma Cells ◽  
Expression Profiles ◽  
Gene Expression Profiles ◽  
Memory B Cells ◽  
Vitro Model ◽  
Intermediate Cells ◽  
Activated B Cells

Abstract Human plasma cells (PCs) and their precursors play an essential role in humoral immune response but are rare and difficult to harvest. We report the generation of human syndecan-1+ and immunoglobulin secreting PCs starting from memory B cells in a 3-step and 10-day (D) culture, including a 6-fold cell amplification. We report the detailed phenotypic and Affymetrix gene expression profiles of these in vitro PCs as well as of intermediate cells (activated B cells and plasmablasts) compared with memory B cells and bone marrow PCs, which is accessible through an open web ATLAS (http://amazonia.transcriptome.eu/). We show this B cell–to-PC differentiation to involve IRF4 and AICDA expressions in D4 activated B cells, decrease of PAX5 and BCL6 expressions, and increase in PRDM1 and XBP1 expressions in D7 plasmablasts and D10 PCs. It involves down-regulation of genes controlled by Pax5 and induction of genes controlled by Blimp-1 and XBP1 (unfold protein response). The detailed phenotype of D10 PCs resembles that of peripheral blood PCs detected after immunization of healthy donors. This in vitro model will facilitate further studies in PC biology. It will likewise be helpful to study PC dyscrasias, including multiple myeloma.

scholarly journals The B29 (Immunoglobulin β-Chain) Gene Is a Genetic Target for Early B-Cell Factor

1999 ◽  
Vol 19 (1) ◽  
pp. 392-401 ◽  
Author(s):  
Peter Åkerblad ◽  
Maria Rosberg ◽  
Tomas Leanderson ◽  
Mikael Sigvardsson
Keyword(s):  
Transcription Factor ◽  
B Cells ◽  
B Cell ◽  
B Lymphocyte ◽  
Ectopic Expression ◽  
Cell Receptor ◽  
Cell Development ◽  
B Cell Development ◽  
Chain Gene ◽  
Early B Cell Factor

ABSTRACT Early B-cell factor (EBF) is a transcription factor suggested as essential for early B-lymphocyte development by findings in mice where the coding gene has been inactivated by homologous disruption. This makes the identification of genetic targets for this transcription factor pertinent for the understanding of early B-cell development. The lack of B29 transcripts, coding for the β subunit of the B-cell receptor complex, in pro-B cells from EBF-deficient mice suggested that B29 might be a genetic target for EBF. We here present data suggesting that EBF interacts with three independent sites within the mouse B29 promoter. Furthermore, ectopic expression of EBF in HeLa cells activated a B29promoter-controlled reporter construct 13-fold and induced a low level of expression from the endogenous B29 gene. Finally, mutations in the EBF binding sites diminished B29 promoter activity in pre-B cells while the same mutations did not have as striking an effect on the promoter function in B-cell lines of later differentiation stages. These data suggest that the B29gene is a genetic target for EBF in early B-cell development.

scholarly journals Ikaros is absolutely required for pre-B cell differentiation by attenuating IL-7 signals

2013 ◽  
Vol 210 (13) ◽  
pp. 2823-2832 ◽  
Author(s):  
Beate Heizmann ◽  
Philippe Kastner ◽  
Susan Chan
Keyword(s):  
Cell Differentiation ◽  
B Cells ◽  
B Cell ◽  
Cell Receptor ◽  
B Cell Differentiation ◽  
B Cell Signaling ◽  
And Migration ◽  
And Function

Pre-B cell receptor (pre-BCR) signaling and migration from IL-7–rich environments cooperate to drive pre-B cell differentiation via transcriptional programs that remain unclear. We show that the Ikaros transcription factor is required for the differentiation of large pre-B to small pre-B cells. Mice deleted for Ikaros in pro/pre-B cells show a complete block of differentiation at the fraction C′ stage, and Ikaros-null pre-B cells cannot differentiate upon withdrawal of IL-7 in vitro. Restoration of Ikaros function rescues pre-B cell differentiation in vitro and in vivo and depends on DNA binding. Ikaros is required for the down-regulation of the pre-BCR, Igκ germline transcription, and Ig L chain recombination. Furthermore, Ikaros antagonizes the IL-7–dependent regulation of >3,000 genes, many of which are up- or down-regulated between fractions C′ and D. Affected genes include those important for survival, metabolism, B cell signaling, and function, as well as transcriptional regulators like Ebf1, Pax5, and the Foxo1 family. Our data thus identify Ikaros as a central regulator of IL-7 signaling and pre-B cell development.

scholarly journals The quantity of CD40 signaling determines the differentiation of B cells into functionally distinct memory cell subsets

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Takuya Koike ◽  
Koshi Harada ◽  
Shu Horiuchi ◽  
Daisuke Kitamura
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Fate ◽  
Plasma Cells ◽  
Memory Cell ◽  
Cell Receptor ◽  
Primary Immune Response ◽  
Cell Fate Decisions ◽  
Cd40 Stimulation

In mice, memory B (Bmem) cells can be divided into two subpopulations: CD80hi Bmem cells, which preferentially differentiate into plasma cells; and CD80lo Bmem cells, which become germinal center (GC) B cells during a recall response. We demonstrate that these distinct responses can be B-cell-intrinsic and essentially independent of B-cell receptor (BCR) isotypes. Furthermore, we find that the development of CD80hi Bmem cells in the primary immune response requires follicular helper T cells, a relatively strong CD40 signal and a high-affinity BCR on B cells, whereas the development of CD80lo Bmem cells does not. Quantitative differences in CD40 stimulation were enough to recapitulate the distinct B cell fate decisions in an in vitro culture system. The quantity of CD40 signaling appears to be translated into NF-κB activation, followed by BATF upregulation that promotes Bmem cell differentiation from GC B cells.

scholarly journals Memory B Cells Are Biased Towards Terminal Differentiation: A Strategy That May Prevent Repertoire Freezing

1997 ◽  
Vol 186 (6) ◽  
pp. 931-940 ◽  
Author(s):  
Christophe Arpin ◽  
Jacques Banchereau ◽  
Yong-Jun Liu
Keyword(s):  
Cell Differentiation ◽  
B Cells ◽  
B Cell ◽  
Plasma Cell ◽  
Plasma Cells ◽  
Cell Clone ◽  
Terminal Differentiation ◽  
Memory B Cells ◽  
B Cell Differentiation

Isolation of large numbers of surface IgD+CD38− naive and surface IgD−CD38− memory B cells allowed us to study the intrinsic differences between these two populations. Upon in vitro culture with IL-2 and IL-10, human CD40–activated memory B cells undergo terminal differentiation into plasma cells more readily than do naive B cells, as they give rise to five- to eightfold more plasma cells and three- to fourfold more secreted immunoglobulins. By contrast, naive B cells give rise to a larger number of nondifferentiated B blasts. Saturating concentrations of CD40 ligand, which fully inhibit naive B cell differentiation, only partially affect that of memory B cells. The propensity of memory B cells to undergo terminal plasma cell differentiation may explain the extensive extra follicular plasma cell reaction and the limited germinal center reaction observed in vivo after secondary immunizations, which contrast with primary responses in carrier-primed animals. This unique feature of memory B cells may confer two important capacities to the immune system: (a) the rapid generation of a large number of effector cells to efficiently eliminate the pathogens; and (b) the prevention of the overexpansion and chronic accumulation of one particular memory B cell clone that would freeze the available peripheral repertoire.

Interleukin-6 is a growth factor for nonmalignant human plasmablasts

Blood ◽  
2001 ◽  
Vol 97 (6) ◽  
pp. 1817-1822 ◽  
Author(s):  
Gaëtan Jego ◽  
Régis Bataille ◽  
Catherine Pellat-Deceunynck
Keyword(s):  
B Cells ◽  
Growth Factor ◽  
Interleukin 6 ◽  
Plasma Cells ◽  
Fold Increase ◽  
Human Memory ◽  
B Cell Differentiation ◽  
Activated B Cells

Interleukin-6 (IL-6), although often regarded as a B-cell differentiation factor, was recently described as the essential survival factor for human plasmablasts in vivo in reactive plasmacytosis. The present study reinvestigated the roles of IL-6 and IL-2 in the generation of plasma cells from human memory B cells in vitro. The cells involved in this differentiation process were identified as preplasmablasts (CD20±CD38±CD138−), plasmablasts (CD20−CD38++CD138−), and early plasma cells (CD20−CD38+++CD138+++). IL-2 or IL-10 induced a strong generation of plasmablasts and early plasma cells (PCs). Compared to IL-2 or IL-10, IL-6 alone was inefficient at PC generation. However, when combined with IL-2 or IL-10, IL-6 enhanced generation of early PCs. Moreover, anti–IL-6 monoclonal antibody markedly reduced IL-2–induced generation of early plasma cells, but not of plasmablasts. These roles of IL-2 and IL-6 were consistent with the difference in the expression of their respective receptors (R). CD25 (IL-2Rα) was increased 72 ± 10-fold on activated B cells, but decreased and then disappeared on plasmablasts. Conversely, CD126 (IL-6Rα) was barely expressed on activated B cells, but increased 18 ± 2-fold on preplasmablasts. Finally, IL-6 enhanced the proliferation (2-fold increase) of IL-2–generated plasmablasts. In conclusion, the data indicate that IL-6 is a growth factor for nonmalignant human plasmablasts.

scholarly journals Repression of c-myc Is Necessary but Not Sufficient for Terminal Differentiation of B Lymphocytes In Vitro

2000 ◽  
Vol 20 (23) ◽  
pp. 8684-8695 ◽  
Author(s):  
Kuo-I Lin ◽  
Yi Lin ◽  
Kathryn Calame
Keyword(s):  
Cell Differentiation ◽  
Target Genes ◽  
Ectopic Expression ◽  
Constitutive Expression ◽  
Dominant Negative ◽  
B Cell Differentiation ◽  
Dominant Negative Form ◽  
Proliferation And Differentiation ◽  
Negative Form

ABSTRACT The importance of c-myc as a target of the Blimp-1 repressor has been studied in BCL-1 cells, in which Blimp-1 is sufficient to trigger terminal B-cell differentiation. Our data show that Blimp-1-dependent repression of c-myc is required for BCL-1 differentiation, since constitutive expression of c-Myc blocked differentiation. Furthermore, ectopic expression of cyclin E mimicked the effects of c-Myc on both proliferation and differentiation, indicating that the ability of c-Myc to drive proliferation is responsible for blocking BCL-1 differentiation. However, inhibition of c-Myc by a dominant negative form was not sufficient to drive BCL-1 differentiation. Thus, during Blimp-1-dependent plasma cell differentiation, repression of c-myc is necessary but not sufficient, demonstrating the existence of additional Blimp-1 target genes.

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