scholarly journals Antigen-driven B cell differentiation in vivo.

1993 ◽  
Vol 178 (1) ◽  
pp. 295-307 ◽  
Author(s):  
M G McHeyzer-Williams ◽  
M J McLean ◽  
P A Lalor ◽  
G J Nossal
Keyword(s):  
B Cells ◽  
Somatic Mutation ◽  
Germinal Center ◽  
Somatic Hypermutation ◽  
Hypervariable Region ◽  
Germinal Centers ◽  
B Cell Differentiation ◽  
The One ◽  
Selection Of

The secretion of specific antibodies and the development of somatically mutated memory B cells in germinal centers are consequences of T cell-dependent challenge with the hapten (4-hydroxy-3-nitrophenyl)acetyl (NP). Using six-parameter flow cytometry and single cell molecular analysis we can directly monitor the extent of somatic hypermutation in individual responsive (isotype switched) antigen-specific B cells. The current study provides a direct quantitative assessment of recruitment into the antibody-secreting compartment on the one hand, and the germinal center pathway to memory on the other. Cellular expansion in both compartments is exponential and independent during the first week after challenge. The first evidence of somatic mutation, towards the end of the first week, was restricted to the germinal center pathway. Furthermore, germinal center cells express a significantly shorter third hypervariable region (CDR3), even when unmutated, than their antibody-secreting counterparts, suggesting a secondary selection event may occur at the bifurcation of these two pathways in vivo. By the end of the second week, the majority of mutated clones express a shorter CDR3 and affinity-increasing mutations as evidence of further selection after somatic mutation. These data provide evidence for substantial proliferation within germinal centers before the initiation of somatic mutation and the subsequent selection of a significant frequency of mutated clonotypes into the memory compartment.

scholarly journals Germinal centers in human lymph nodes contain reactivated memory B cells

2007 ◽  
Vol 204 (11) ◽  
pp. 2655-2665 ◽  
Author(s):  
Richard J. Bende ◽  
Febe van Maldegem ◽  
Martijn Triesscheijn ◽  
Thera A.M. Wormhoudt ◽  
Richard Guijt ◽  
...  
Keyword(s):  
B Cells ◽  
Lymph Nodes ◽  
Germinal Center ◽  
Lymphoid Tissue ◽  
Somatic Hypermutation ◽  
Germinal Centers ◽  
Memory B Cells ◽  
Cell Clones ◽  
Clonal Origin ◽  
Successive Recall

To reveal migration trails of antigen-responsive B cells in lymphoid tissue, we analyzed immunoglobulin (Ig)M-VH and IgG-VH transcripts of germinal center (GC) samples microdissected from three reactive human lymph nodes. Single B cell clones were found in multiple GCs, one clone even in as many as 19 GCs. In several GCs, IgM and IgG variants of the same clonal origin were identified. The offspring of individual hypermutated IgG memory clones were traced in multiple GCs, indicating repeated engagement of memory B cells in GC reactions. These findings imply that recurring somatic hypermutation progressively drives the Ig repertoire of memory B cells to higher affinities and infer that transforming genetic hits in non-Ig genes during lymphomagenesis do not have to arise during a single GC passage, but can be collected during successive recall responses.

Dysregulation of MMSET Is Tumorigenic In-Vivo.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 74-74 ◽  
Author(s):  
Marta Chesi ◽  
Kruti Naik ◽  
Davide F. Robbiani ◽  
Maurizio Affer ◽  
Helen D. Nickerson ◽  
...  
Keyword(s):  
T Cells ◽  
B Cells ◽  
Transgenic Mice ◽  
Cell Lines ◽  
Germinal Center ◽  
Somatic Hypermutation ◽  
Critical Role ◽  
Lymphoid Cells ◽  
Proximal Promoter

Abstract Approximately 15% of multiple myeloma (MM) is characterized by a t(4;14) translocation that causes the simultaneous dysregulation of MMSET on der(4) and fibroblast growth factor receptor 3 gene (FGFR3) on der(14). We reported several lines of evidence indicating a role for FGFR3 in myeloma tumorigenesis. First, activated FGFR3 is an oncogene capable of transforming fibroblasts. Second, FGFR3 activating mutations are acquired by MM cells during tumor progression. Third, targeted inhibition of FGFR3 leads to terminal differentiation and apoptosis in two t(4;14) MM cell lines. However, expression of FGFR3, but never of MMSET, is lost in about 25% of t(4;14) MM. Therefore, the overexpression of MMSET in all MM tumors with a t(4;14) translocation, and its homology to MLL, the oncogene on 11q23 translocated in acute leukemia suggest a critical role for MMSET in MM. To determine whether MMSET is an oncogene in vivo, we have generated transgenic mice in which MMSET expression in driven in lymphocytes by the lck proximal promoter juxtaposed to the Emu enhancer. Using the same expression vector we and others have obtained specific, high levels of transgene expression in B and T cells from spleen, bone marrow and thymus. Four transgenic lines were generated and although we detected MMSET expression in T cells in each of them, unexpectedly no expression in B cells was seen. This is consistent with our inability to ectopically express MMSET in B cell lines. Nevertheless B lymphoid tumors expressing MMSET developed at 23 month of age in each line (18/51 mice). Only 1/19 wild type matching control mice developed a splenomegaly. By Southern blot, monoclonal rearrangements of IgH, IgL and TCR β were detected within the same tumor population. In conclusion, this is the first report that MMSET is an oncogene capable of transforming lymphoid cells in an animal model. We are currently crossing these mice with FGFR3 transgenic mice to assess cooperation between these two oncogenes in tumorigenesis. Obviously a more restricted expression of MMSET in germinal center cells is required to investigate the role of MMSET in MM. Therefore, as we have done for c-myc, we are generating new transgenic mice in which MMSET expression will be activated sporadically in germinal center B cells by somatic hypermutation.

scholarly journals In situ studies of the primary immune response to (4-hydroxy-3-nitrophenyl)acetyl. III. The kinetics of V region mutation and selection in germinal center B cells.

1993 ◽  
Vol 178 (4) ◽  
pp. 1293-1307 ◽  
Author(s):  
J Jacob ◽  
J Przylepa ◽  
C Miller ◽  
G Kelsoe
Keyword(s):  
B Cells ◽  
Germinal Center ◽  
Somatic Hypermutation ◽  
Point Mutations ◽  
Variable Region ◽  
Germinal Centers ◽  
Primary Immune Response ◽  
Kinetics Of ◽  
Mutation And Selection ◽  
Germinal Center B Cells

In the murine spleen, germinal centers are the anatomic sites for antigen-driven hypermutation and selection of immunoglobulin (Ig) genes. To detail the kinetics of Ig mutation and selection, 178 VDJ sequences from 16 antigen-induced germinal centers were analyzed. Although germinal centers appeared by day 4, mutation was not observed in germinal center B cells until day 8 postimmunization; thereafter, point mutations favoring asymmetrical transversions accumulated until day 14. During this period, strong phenotypic selection on the mutant B lymphocytes was inferred from progressively biased distributions of mutations within the Ig variable region, the loss of crippling mutations, decreased relative clonal diversity, and increasingly restricted use of canonical gene segments. The period of most intense selection on germinal center B cell populations preceded significant levels of mutation and may represent a physiologically determined restriction on B cells permitted to enter the memory pathway. Noncanonical Ig genes recovered from germinal centers were mostly unmutated although they probably came from antigen-reactive cells. Together, these observations demonstrate that the germinal center microenvironment is rich and temporally complex but may not be constitutive for somatic hypermutation.

scholarly journals Negative selection of human germinal center B cells by prolonged BCR cross-linking.

1996 ◽  
Vol 183 (5) ◽  
pp. 2075-2085 ◽  
Author(s):  
L Galibert ◽  
N Burdin ◽  
C Barthélémy ◽  
G Meffre ◽  
I Durand ◽  
...  
Keyword(s):  
Cell Differentiation ◽  
B Cells ◽  
B Cell ◽  
B Lymphocytes ◽  
Germinal Center ◽  
B Lymphocyte ◽  
Calcium Flux ◽  
Cross Linking ◽  
B Cell Differentiation ◽  
Selection Of

The antigen receptors on T and B lymphocytes can transduce both agonist and antagonist signals leading either to activation/survival or anergy/death. The outcome of B lymphocyte antigen receptor (BCR) triggering depends upon multiple parameters which include (a) antigen concentration and valency, (b) duration of BCR occupancy, (c) receptor affinity, and (d) B cell differentiation stages. Herein, using anti-immunoglobulin kappa and lambda light chain antibodies, we analyzed the response of human naive, germinal center (GC) or memory B cells to BCR cross-linking regardless of heavy chain Ig isotype or intrinsic BCR specificity. We show that after CD40-activation, anti-BCR (kappa + gamma) can elicit an intracellular calcium flux on both GC and non-GC cells. However, prolonged BCR cross-linking induces death of CD40-activated GC B cells but enhances proliferation of naive or memory cells. Anti-kappa antibody only kills kappa + GC B cells without affecting surrounding gamma + GC B cells, thus demonstrating that BCR-mediated killing of GC B lymphocytes is a direct effect that does not involve a paracrine mechanism. BCR-mediated killing of CD40-activated GC B cells could be partially antagonized by the addition of IL-4. Moreover, in the presence of IL-4, prestimulation through CD40 could prevent subsequent anti-Ig-mediated cell death, suggesting a specific role of this combination in selection of GC B cells. This report provides evidence that in human, susceptibility to BCR killing is regulated along peripheral B cell differentiation pathway.

scholarly journals TLR4 signaling augments B lymphocyte migration and overcomes the restriction that limits access to germinal center dark zones

2009 ◽  
Vol 206 (12) ◽  
pp. 2641-2657 ◽  
Author(s):  
Il-Young Hwang ◽  
Chung Park ◽  
Kathleen Harrison ◽  
John H. Kehrl
Keyword(s):  
B Cells ◽  
Chronic Inflammation ◽  
Germinal Center ◽  
B Lymphocyte ◽  
Plasma Cells ◽  
Germinal Centers ◽  
Lymphocyte Migration ◽  
Tlr4 Signaling ◽  
Cell Compartments

B lymphocyte–intrinsic Toll-like receptor (TLR) signals amplify humoral immunity and can exacerbate autoimmune diseases. We identify a new mechanism by which TLR signals may contribute to autoimmunity and chronic inflammation. We show that TLR4 signaling enhances B lymphocyte trafficking into lymph nodes (LNs), induces B lymphocyte clustering and interactions within LN follicles, leads to sustained in vivo B cell proliferation, overcomes the restriction that limits the access of nonantigen-activated B cells to germinal center dark zones, and enhances the generation of memory and plasma cells. Intravital microscopy and in vivo tracking studies of B cells transferred to recipient mice revealed that TLR4-activated, but not nonstimulated, B cells accumulated within the dark zones of preexisting germinal centers even when transferred with antigen-specific B cells. The TLR4-activated cells persist much better than nonstimulated cells, expanding both within the memory and plasma cell compartments. TLR-mediated activation of B cells may help to feed and stabilize the spontaneous and ectopic germinal centers that are so commonly found in autoimmune individuals and that accompany chronic inflammation.

scholarly journals Altered Germinal-Center Metabolism in B Cells in Autoimmunity

Metabolites ◽  
2022 ◽  
Vol 12 (1) ◽  
pp. 40
Author(s):  
Ashton K. Shiraz ◽  
Eric J. Panther ◽  
Christopher M. Reilly
Keyword(s):  
B Cells ◽  
B Cell ◽  
Lupus Erythematosus ◽  
Germinal Center ◽  
Plasma Cells ◽  
Somatic Hypermutation ◽  
Cell Metabolism ◽  
Affinity Maturation ◽  
B Cell Differentiation ◽  
Contributing Factors

B lymphocytes play an important role in the pathophysiology of many autoimmune disorders by producing autoantibodies, secreting cytokines, and presenting antigens. B cells undergo extreme physiological changes as they develop and differentiate. Aberrant function in tolerogenic checkpoints and the metabolic state of B cells might be the contributing factors to the dysfunctionality of autoimmune B cells. Understanding B-cell metabolism in autoimmunity is important as it can give rise to new treatments. Recent investigations have revealed that alterations in metabolism occur in the activation of B cells. Several reports have suggested that germinal center (GC) B cells of individuals with systemic lupus erythematosus (SLE) have altered metabolic function. GCs are unique microenvironments in which the delicate and complex process of B-cell affinity maturation occurs through somatic hypermutation (SHM) and class switching recombination (CSR) and where Bcl6 tightly regulates B-cell differentiation into memory B-cells or plasma cells. GC B cells rely heavily on glucose, fatty acids, and oxidative phosphorylation (OXPHOS) for their energy requirements. However, the complicated association between GC B cells and their metabolism is still not clearly understood. Here, we review several studies of B-cell metabolism, highlighting the significant transformations that occur in GC progression, and suggest possible approaches that may be investigated to more precisely target aberrant B-cell metabolism in SLE.

DNA Breaks in Hypermutating Immunoglobulin Genes: Evidence for a Break-and-Repair Pathway of Somatic Hypermutation

Genetics ◽  
2001 ◽  
Vol 158 (1) ◽  
pp. 369-378 ◽  
Author(s):  
Qingzhong Kong ◽  
Nancy Maizels
Keyword(s):  
B Cells ◽  
Germinal Center ◽  
Somatic Hypermutation ◽  
Sequence Motif ◽  
Immunoglobulin Gene ◽  
Sequence Motifs ◽  
Dna Breaks ◽  
Dna Strands ◽  
Germinal Center B Cells

Abstract To test the hypothesis that immunoglobulin gene hypermutation in vivo employs a pathway in which DNA breaks are introduced and subsequently repaired to produce mutations, we have used a PCR-based assay to detect and identify single-strand DNA breaks in λ1 genes of actively hypermutating primary murine germinal center B cells. We find that there is a two- to threefold excess of breaks in λ1 genes of hypermutating B cells, relative to nonhypermutating B cells, and that 1.3% of germinal center B cells contain breaks in the λ1 gene that are associated with hypermutation. Breaks were found in both top and bottom DNA strands and were localized to the region of λ1 that actively hypermutates, but duplex breaks accounted for only a subset of breaks identified. Almost half of the breaks in hypermutating B cells occurred at hotspots, sites at which two or more independent breaks were identified. Breaksite hotspots were associated with characteristic sequence motifs: a pyrimidine-rich motif, either RCTYT or CCYC; and RGYW, a sequence motif associated with hypermutation hotspots. The sequence motifs identified at breaksite hotspots should inform the design of substrates for characterization of activities that participate in the hypermutation pathway.

scholarly journals Histone deacetylase 3 controls a transcriptional network required for B cell maturation

2019 ◽  
Vol 47 (20) ◽  
pp. 10612-10627 ◽  
Author(s):  
Kristy R Stengel ◽  
Srividya Bhaskara ◽  
Jing Wang ◽  
Qi Liu ◽  
Jacob D Ellis ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Histone Deacetylase ◽  
Germinal Center ◽  
Hdac Inhibitors ◽  
Germinal Centers ◽  
Cell Of Origin ◽  
Histone Deacetylase 3 ◽  
Germinal Center B Cells

Abstract Histone deacetylase 3 (Hdac3) is a target of the FDA approved HDAC inhibitors, which are used for the treatment of lymphoid malignancies. Here, we used Cd19-Cre to conditionally delete Hdac3 to define its role in germinal center B cells, which represent the cell of origin for many B cell malignancies. Cd19-Cre-Hdac3−/− mice showed impaired germinal center formation along with a defect in plasmablast production. Analysis of Hdac3−/− germinal centers revealed a reduction in dark zone centroblasts and accumulation of light zone centrocytes. RNA-seq revealed a significant correlation between genes up-regulated upon Hdac3 loss and those up-regulated in Foxo1-deleted germinal center B cells, even though Foxo1 typically activates transcription. Therefore, to determine whether gene expression changes observed in Hdac3−/− germinal centers were a result of direct effects of Hdac3 deacetylase activity, we used an HDAC3 selective inhibitor and examined nascent transcription in germinal center-derived cell lines. Transcriptional changes upon HDAC3 inhibition were enriched for light zone gene signatures as observed in germinal centers. Further comparison of PRO-seq data with ChIP-seq/exo data for BCL6, SMRT, FOXO1 and H3K27ac identified direct targets of HDAC3 function including CD86, CD83 and CXCR5 that are likely responsible for driving the light zone phenotype observed in vivo.

scholarly journals bcl-2 Transgene Expression Inhibits Apoptosis in the Germinal Center and Reveals Differences in the Selection of Memory B Cells and Bone Marrow Antibody-Forming Cells

2000 ◽  
Vol 191 (3) ◽  
pp. 475-484 ◽  
Author(s):  
Kenneth G.C. Smith ◽  
Amanda Light ◽  
Lorraine A. O'Reilly ◽  
Soon-Meng Ang ◽  
Andreas Strasser ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
Germinal Center ◽  
Low Frequency ◽  
Germinal Centers ◽  
Affinity Maturation ◽  
Memory B Cells ◽  
High Affinity ◽  
Excessive Number ◽  
Selection Of

Immunization with T cell–dependent antigens generates long-lived memory B cells and antibody-forming cells (AFCs). Both populations originate in germinal centers and, predominantly, produce antibodies with high affinity for antigen. The means by which germinal center B cells are recruited into these populations remains unclear. We have examined affinity maturation of antigen-specific B cells in mice expressing the cell death inhibitor bcl-2 as a transgene. Such mice had reduced apoptosis in germinal centers and an excessive number of memory B cells with a low frequency of V gene somatic mutation, including those mutations encoding amino acid exchanges known to enhance affinity. Despite the frequency of AFCs being increased in bcl-2–transgenic mice, the fraction secreting high-affinity antibody in the bone marrow at day 42 remained unchanged compared with controls. The inability of BCL-2 to alter selection of bone marrow AFCs is consistent with these cells being selected within the germinal center on the basis of their affinity being above some threshold rather than their survival being due to a selective competition for an antigen-based signal. Continuous competition for antigen does, however, explain formation of the memory compartment.

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