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O de Bouteiller

Publications and source records attributed to O de Bouteiller.

At least 19 recordsLinked to original sources
PubMed2000

Dendritic cell biology and regulation of dendritic cell trafficking by chemokines.

DC (dendritic cells) represent an heterogeneous family of cells which function as sentinels of the immune system. They traffic from the blood to the tissues where, while immature, they capture antigens. Then, following inflammatory stimuli, they leave the tissues and move to the draining lymphoid organs where, converted into mature DC, they prime naive T cells. The key role of DC migration in their sentinel function led to the investigation of the chemokine responsiveness of DC populations during their development and maturation. These studies have shown that immature DC respond to many CC and CXC chemokines (MIP-1 alpha, MIP-1 beta, MIP-3 alpha, MIP-5, MCP-3, MCP-4, RANTES, TECK and SDF-1) which are inducible upon inflammatory stimuli. Importantly, each immature DC population displays a unique spectrum of chemokine responsiveness. For examples, Langerhans cells migrate selectively to MIP-3 alpha (via CCR6), blood CD11c+ DC to MCP chemokines (via CCR2), monocytes derived-DC respond to MIP-1 alpha/beta (via CCR1 and CCR5), while blood CD11c- DC precursors do not respond to any of these chemokines. All these chemokines are inducible upon inflammatory stimuli, in particular MIP-3 alpha, which is only detected within inflamed epithelium, a site of antigen entry known to be infiltrated by immature DC. In contrast to immature DC, mature DC lose their responsiveness to most of these inflammatory chemokines through receptor down-regulation or desensitization, but acquire responsiveness to ELC/MIP-3 beta and SLC/6Ckine as a consequence of CCR7 up-regulation. ELC/MIP-3 beta and SLC/6Ckine are specifically expressed in the T-cell-rich areas where mature DC home to become interdigitating DC. Altogether, these observations suggest that the inflammatory chemokines secreted at the site of pathogen invasion will determine the DC subset recruited and will influence the class of the immune response initiated. In contrast, MIP-3 beta/6Ckine have a determinant role in the accumulation of antigenloaded mature DC in T cell-rich areas of the draining lymph node, as illustrated by recent observations in mice deficient for CCR7 or SLC/6Ckine. A better understanding of the regulation of DC trafficking might offer new opportunities of therapeutic interventions to suppress, stimulate or deviate the immune response.

Animals↗
PubMed1998

Antigen receptor engagement turns off the V(D)J recombination machinery in human tonsil B cells.

The germinal center (GC) is an anatomic compartment found in peripheral lymphoid organs, wherein B cells undergo clonal expansion, somatic mutation, switch recombination, and reactivate immunoglobulin gene V(D)J recombination. As a result of somatic mutation, some GC B cells develop higher affinity antibodies, whereas others suffer mutations that decrease affinity, and still others may become self-reactive. It has been proposed that secondary V(D)J rearrangements in GCs might rescue B cells whose receptors are damaged by somatic mutations. Here we present evidence that mature human tonsil B cells coexpress conventional light chains and recombination associated genes, and that they extinguish recombination activating gene and terminal deoxynucleotidyl transferase expression when their receptors are cross-linked. Thus, the response of the recombinase to receptor engagement in peripheral B cells is the opposite of the response in developing B cells to the same stimulus. These observations suggest that receptor revision is a mechanism for receptor diversification that is turned off when antigen receptors are cross-linked by the cognate antigen.

Animals↗
PubMed1998

The normal counterpart of IgD myeloma cells in germinal center displays extensively mutated IgVH gene, Cmu-Cdelta switch, and lambda light chain expression.

Human myeloma are incurable hematologic cancers of immunoglobulin-secreting plasma cells in bone marrow. Although malignant plasma cells can be almost eradicated from the patient's bone marrow by chemotherapy, drug-resistant myeloma precursor cells persist in an apparently cryptic compartment. Controversy exists as to whether myeloma precursor cells are hematopoietic stem cells, pre-B cells, germinal center (GC) B cells, circulating memory cells, or plasma blasts. This situation reflects what has been a general problem in cancer research for years: how to compare a tumor with its normal counterpart. Although several studies have demonstrated somatically mutated immunoglobulin variable region genes in multiple myeloma, it is unclear if myeloma cells are derived from GCs or post-GC memory B cells. Immunoglobulin (Ig)D-secreting myeloma have two unique immunoglobulin features, including a biased lambda light chain expression and a Cmu-Cdelta isotype switch. Using surface markers, we have previously isolated a population of surface IgM-IgD+CD38+ GC B cells that carry the most impressive somatic mutation in their IgV genes. Here we show that this population of GC B cells displays the two molecular features of IgD-secreting myeloma cells: a biased lambda light chain expression and a C&mu-Cdelta isotype switch. The demonstration of these peculiar GC B cells to differentiate into IgD-secreting plasma cells but not memory B cells both in vivo and in vitro suggests that IgD-secreting plasma and myeloma cells are derived from GCs.

B-Lymphocytes↗
PubMed1998

Somatic hypermutation introduces insertions and deletions into immunoglobulin V genes.

During a germinal center reaction, random mutations are introduced into immunoglobulin V genes to increase the affinity of antibody molecules and to further diversify the B cell repertoire. Antigen-directed selection of B cell clones that generate high affinity surface Ig results in the affinity maturation of the antibody response. The mutations of Ig genes are typically basepair substitutions, although DNA insertions and deletions have been reported to occur at a low frequency. In this study, we describe five insertion and four deletion events in otherwise somatically mutated VH gene cDNA molecules. Two of these insertions and all four deletions were obtained through the sequencing of 395 cDNA clones (approximately 110,000 nucleotides) from CD38+IgD- germinal center, and CD38-IgD- memory B cell populations from a single human tonsil. No germline genes that could have encoded these six cDNA clones were found after an extensive characterization of the genomic VH4 repertoire of the tonsil donor. These six insertions or deletions and three additional insertion events isolated from other sources occurred as triplets or multiples thereof, leaving the transcripts in frame. Additionally, 8 of 9 of these events occurred in the CDR1 or CDR2, following a pattern consistent with selection, and making it unlikely that these events were artifacts of the experimental system. The lack of similar instances in unmutated IgD+CD38- follicular mantle cDNA clones statistically associates these events to the somatic hypermutation process (P = 0.014). Close scrutiny of the 9 insertion/deletion events reported here, and of 25 additional insertions or deletions collected from the literature, suggest that secondary structural elements in the DNA sequences capable of producing loop intermediates may be a prerequisite in most instances. Furthermore, these events most frequently involve sequence motifs resembling known intrinsic hotspots of somatic hypermutation. These insertion/deletion events are consistent with models of somatic hypermutation involving an unstable polymerase enzyme complex lacking proofreading capabilities, and suggest a downregulation or alteration of DNA repair at the V locus during the hypermutation process.

Amino Acid Sequence↗
PubMed1997

In vitro triggering of somatic mutation in human naive B cells.

During T cell-dependent immune response, germinal center B cells accumulate somatic mutations in their Ig V(D)J genes and give rise to affinity-selected B cells. We tested several culture conditions for triggering somatic mutation in human tonsillar naive slgD+CD23+ cells after cross-linking their membrane Igs. CD40 activation, in the presence of exogenous cytokines (IL-2, IL-4, and IL-10), induced proliferation and isotype switch without somatic mutation. In contrast, after coculture with anti-CD3-activated cloned T cells, somatic mutation accumulated in a fraction of naive B cells. Mutations included shared as well as independent events in clonally related sequences, allowing reconstitution of genealogic trees generated in vitro. Naive tonsillar B cells sorted for slgD expression can be induced to mutate their Ig V(H) gene upon coculture with activated T cells, thereby providing a model to study somatic hypermutation in vitro.

Amino Acid Sequence↗
PubMed1997

Germinal center founder cells display propensity for apoptosis before onset of somatic mutation.

B lymphocytes undergo affinity maturation of their antigen receptors within germinal centers. These anatomical structures develop in secondary lymphoid organs from the clonal expansion of a few antigen-specific founder B cells, whose isolation and characterization are reported here. Human germinal center founder cells express the naive B cell markers surface IgM and IgD as well as the germinal center B cell markers CD10 and CD38. They express low levels of Bcl-2, high levels of Fas, and undergo rapid apoptosis in culture. The smaller nonproliferating sIgM+IgD+CD38+ B cells displayed a lower level of somatic mutation in their immunoglobulin variable region genes compared with the large proliferating ones. Unmutated sIgM+IgD-CD38+ tonsillar B cells may thus represent germinal center founder cells in which the program for apoptotic cell death is triggered before the onset of somatic mutation, allowing the selection of the germline antibody repertoire at an early stage.

ADP-ribosyl Cyclase↗
PubMed1997

Follicular dendritic cells specifically express the long CR2/CD21 isoform.

This paper describes an antibody (mAb 7D6) that specifically recognizes human follicular dendritic cells (FDCs). By expression cloning, a cDNA clone encoding for the long human CR2/ CD21 isoform (CD21L) that contains an additional exon (10a) was isolated. We demonstrated that FDCs selectively express CD21L, while B cells selectively express the short CR2/CD21 lacking exon 10a (CD21S). By screening mouse Ltk- cells transfected with the CD21L cDNA, we further showed that the other two anti-human FDC mAbs DRC-1 and KiM4 also recognize CD21L. Thus, CD21L represents the first characterized human FDC-specific molecule, which may confer unique functions of FDCs in germinal center development.

Animals↗
PubMed1997

Molecular cloning of human RP105.

RP105 is a 105-kDa type I membrane protein of the leucine-rich repeat (LRR) family. Anti-RP105 sensitizes B cells to antigen-receptor-mediated apoptosis, but protects B cells from radiation-induced apoptosis and stimulates B cell proliferation. The sequence of the mouse RP105 has been reported. Here, we report the characterization of the human RP105. The 2.6-kb cDNA encodes a protein of 661 amino acids which displays 78% homology with mouse RP105. The 22 LRR and the 9 potential N-linked glycosylation sites within the extracellular region are conserved. While previous studies have shown that RP105 is expressed on surface IgM+IgD+2 B cells in mice, human RP105 was shown to be expressed on all subsets of mature B cells and dendritic cells. Human RP105 gene was mapped to the long arm of chromosome 5, where numerous cytokines and receptors have been localized.

Animals↗
PubMed1997

Mechanisms of selection and differentiation in germinal centers.

The choice between death and survival, a feature of early B cell development, is a choice also faced by mature B cells. If they survive this early developmental decision, mature B cells then face a second choice: either to undergo either terminal differentiation into plasma cells or to differentiate into memory B cells. Antigens, T cell signals (cytokines, CD40 ligand and Fas ligand) as well as the activation state of B cells determine their ultimate fate.

B-Lymphocytes↗
PubMed1996

Human germinal center B cells express the apoptosis-inducing genes Fas, c-myc, P53, and Bax but not the survival gene bcl-2.

During T cell-dependent antibody responses, B cells within germinal centers (GC) alter the affinity of their antigen receptor by introducing somatic mutations into variable region of immunoglobulin (IgV) genes. During this process, GC B cells are destined to die unless positively selected by antigens and CD40-ligand. To understand survival/death control of germinal center B cell, the expression of four apoptosis-inducing genes, Fas, c-myc, Bax, and P53, together with the survival gene bcl-2, has been analyzed herein among purified tonsillar naive, GC, and memory B cells. IgD+CD38- naive B cells were separated into CD23- (mature B cell [Bm]1) subset and CD23+ (Bm2), IgD-CD38+ GC B cells were separated into subsets of CD77+ centroblasts (Bm3) and CD77- centrocytes (Bm4), whereas IgD-CD38- cells represented the Bm5 memory B cell subset. Sequence analysis of IgV region genes indicated that somatic hypermutation was triggered in the Bm3 centroblast subset. Here we show that bcl-2 is only detectable with naive (Bm1 and 2) and memory B cell (Bm5) subsets, whereas all four apoptosis-inducing genes were most significantly expressed within GC B cells. Fas was equally expressed in Bm3 centroblasts and Bm4 centrocytes, whereas Bax was most significantly expressed in Bm4 centrocytes. c-myc, a positive regulator of cell cycle, was most significantly expressed in proliferating Bm3 centroblasts, whereas P53, a negative regulator of cell cycle, was most signficantly expressed in nonproliferating Bm4 centrocytes. The present results indicate that the survival/death of GC B cells are regulated by the up- and downregulation of multiple genes, among which the expression of c-myc and P53 in the absence of bcl-2 may prime the proliferating Bm3 centroblasts and nonproliferating Bm4 centrocytes to apoptosis.

Apoptosis↗
PubMed1996

Bcl-2+ tonsillar plasma cells are rescued from apoptosis by bone marrow fibroblasts.

Plasma cells represent the final stage of B lymphocyte differentiation. Most plasma cells in secondary lymphoid tissues live for a few days, whereas those in the lamina propria of mucosa and in bone marrow live for several weeks. To investigate the regulation of human plasma cell survival, plasma cells were isolated from tonsils according to high CD38 and low CD20 expression. Tonsillar plasma cells express CD9, CD19, CD24, CD37, CD40, CD74, and HLA-DR, but not CD10, HLA-DQ, CD28, CD56, and Fas/CD95. Although plasma cells express intracytoplasmic Bcl-2, they undergo swift apoptosis in vitro and do not respond to CD40 triggering. Bone marrow fibroblasts and rheumatoid synoviocytes, however, prevented plasma cells from undergoing apoptosis in a contact-dependent fashion. These data indicate that fibroblasts may form a microenvironment favorable for plasma cell survival under normal and pathological conditions.

Antigens, CD↗
PubMed1996

Sequential triggering of apoptosis, somatic mutation and isotype switch during germinal center development.

Using an approach similar to that used to study primary B-lymphocyte development within bone marrow and primary T-lymphocyte development within thymus, the peripheral B-cell maturation pathway within secondary lymphoid tissue (human tonsils) was analysed on the expression of discrete surface antigens. sIgD and CD38 permit the identification of four subpopulations of tonsillar B lymphocytes, including sIgD+ CD38-, sIgD+, CD38+, sIgD-CD38+ and sIgD-CD38- B cells. Further phenotypic, functional and Ig gene analysis (IgV gene sequences, expression of sterile transcripts and DNA switch circles) allowed us to conclude the following: (1) sIgM+ IgD+ CD38- B cells are naive B cells (Bm1 + 2), which carry unmutated antigen-receptors; (2) sIgM+ IgD+ CD38+ B cells are germinal center founder cells (Bm2'), which become prone to undergo apoptosis before the onset of somatic mutation; (3) sIgM-IgD+ CD38+ are germinal center B cells (Bm 3 delta), that have accumulated the highest number of somatic mutations ever reported in normal B cells; these cells may have undergone C mu-deletion by homologous recombination through sigma mu-sigma delta sequences: (4) sIgD-CD38+ CD77+ B cells are centroblasts (Bm3), in which somatic mutation machinery is activated; (5) sIgD-CD38+ CD77- B cells are centrocytes (Bm4), in which the isotype switching machinery is activated; (6) sIgD-CD38- cells (Bm5) represent somatically mutated resting memory B cells. In conclusion, human peripheral B-cell subpopulations corresponding to the differentiation stages before, during and after the triggering of apoptosis program, somatic mutation and isotype switch have been identified and isolated using a combination of surface markers.

Apoptosis↗
PubMed1996

Follicular dendritic cells and germinal centers.

Follicular dendritic cells (FDCs) are stromal cells unique to primary and secondary lymphoid follicles. Recirculating resting B cells migrate through the FDC networks, whereas antigen-activated B cells undergo clonal expansion within the FDC networks in a T cell-dependent fashion, thereby generating germinal centers. Here, B cells undergo somatic mutation, positive and negative selection, isotype switching and differentiation into high-affinity plasma cells and memory B cells. Since the discovery of FDCs by electron microscopy as long-term antigen-retaining cells 30 years ago isolation of FDCs and generation of FDC-like cells lines and of FDC-specific monoclonal antibodies have been achieved. FDCs express all three types of complement receptors as well as Ig-Fc receptors, through which antigen-antibody immune complexes are retained. However, the mechanism that prevents FDCs from internalizing the antigens and retaining them in native form for long periods of time remains obscure. Substantial evidence derived from cultures in vitro indicates that FDCs contribute directly to the survival and activation of peripheral B cells. The adhesion between FDCs and B cells is mediated by ICAM-1 (CD54)-LFA-1(CD11a) and VCAM-VLA-4. T cells may interact with FDCs in a CD40/CD40-ligand-dependent fashion. Whether FDCs originate from hematopoietic progenitors or from stromal elements is still a controversy. New evidence suggests the presence of two types of dendritic cells within human germinal centers: (i) the classic FDCs that express DRC-1, KiM4, and 7D6 antigens represent stromal cells; and (ii) the newly identified CD3-CD4-CD11c- germinal center dendritic cells (GCDC) represent hematopoietic cells that may be analogous to the antigen-transporting cells described in mice. Finally, FDCs appear to be involved in the growth of follicular lymphomas and in the pathogenesis of HIV infection.

Animals↗
PubMed1996

Within germinal centers, isotype switching of immunoglobulin genes occurs after the onset of somatic mutation.

Human tonsillar B cells were separated into naive IgD+CD38-CD23- (Bm1) and IgD+CD38-CD23 (Bm2), germinal center IgD-CD38+CD23- centroblasts (Bm3) and IgD-CD38+CD77- centrocytes (Bm4) and memory IgD-CD38- (Bm5) subsets. Previous IgVH sequence analysis concluded that the triggering of somatic mutations occurs during the transition from Bm2 subset into the Bm3 subset. To determine the initiation of isotype switching, sterile transcript expression was analyzed by amplification, cloning, and sequencing. A selective sterile I gamma, I alpha, and I epsilon expression was observed at centrocyte (Bm4) stage, suggesting that isotype switch is triggered within germinal centers, after somatic mutation is initiated with centroblasts (Bm3). Finally, the high level of 5'S gamma-S mu 3' DNA switching circles observed in germinal center B cells indicates that within human tonsils, germinal center is a major location for isotype switching.

B-Lymphocytes↗
PubMed1996

Normal human IgD+IgM- germinal center B cells can express up to 80 mutations in the variable region of their IgD transcripts.

Somatic hypermutation in immunoglobulin variable region genes occurs within germinal centers. Here, we describe a subset of germinal center dark zone centroblasts that express only sIgD and have accumulated up to 80 mutations per heavy chain variable region (IgVH delta gene). Over half of the hypermutated IgVH delta sequences were found to be clonally related. This level of mutation is not observed in either IgVH gamma transcripts from the same sample or IgVH delta transcripts from peripheral blood, suggesting that these cells neither undergo isotype switch nor mature into circulating memory B cells. Optimal growth of these cells in vitro depends on CD40 ligand, T cell cytokines, and a fibroblast stroma, a combination possibly mimicking the dark zone microenvironment. Our hypothesis is that these cells may be sequestered within germinal centers, where their somatic mutation machinery is triggered. The isolation of these hypermutated B cells may represent a critical step for studying both the biology and biochemistry of somatic hypermutation.

ADP-ribosyl Cyclase↗
PubMed1995

Human follicular dendritic cells enhance cytokine-dependent growth and differentiation of CD40-activated B cells.

Germinal centers constitute microanatomic subunits within secondary follicles where B cells undergo somatic mutations, isotype switch and affinity selection. This allows the generation of memory B cells and plasma cells, whose Igs bind to the eliciting Ag with a high affinity. T cells and follicular dendritic cells (FDCs) are thought to play key roles in the germinal center reaction. To study effects of FDCs on B cell growth and differentiation, we have isolated FDC-lymphocyte clusters from human tonsils by enzymatic digestion and centrifugation of the resulting cell suspension through BSA gradient. Irradiated FDC-lymphocyte clusters induced moderate proliferation of autologous B cells. IL-2 was the only cytokine able to enhance B cell proliferation cocultured with FDCs. When B cells were activated by soluble anti-CD40 Ab with or without IL-2, IL-3, IL-4, IL-10 or IL-13, addition of FDCs increased B cell proliferation. In the presence of FDCs, maximal B cell proliferation was observed in anti-CD40 stimulated cultures supplemented with either IL-4 + IL-10 or IL-2 + IL-10. Cultures performed in the presence of IL-2 and IL-10 resulted in high levels of Ig production in the presence of FDCs. In conclusion, the present study demonstrates that freshly isolated human FDCs can enhance the growth and differentiation of CD40-activated B cells.

B-Lymphocytes↗