Search PubMed⌕ Search

Biomedical subjects

F Rousset

Publications and source records attributed to F Rousset.

At least 37 records · Page 2Linked to original sources
PubMed1996

Human IgG monoclonal antibodies that modulate the binding of specific IgE to birch pollen Bet v 1.

Birch pollen allergy is a very frequent pathology in Europe and North America. More than 95% of the tree pollen allergic patients display IgE reactivity against Bet v 1, the major birch pollen allergen. Starting with PBL from a patient desensitized by immunotherapy, we have generated five B cell lines (BAB1 to BAB5) that secrete human IgG mAbs of high affinity for Bet v 1. Although competition studies indicated that these human IgG mAb recognized different epitopes, broad cross-reactivity was found with Bet v 1 homologous allergens present in tree pollens and plant-derived foods. When tested for interference with allergic patients' IgE, BAB1 inhibited (by 80-100%) the binding of IgE to nitrocellulose-blotted Bet v 1, while BAB2 enhanced it. The biologic significance of the ability of BAB1 to interfere with patients' IgE binding is indicated by the finding that BAB1 completely inhibited Bet v 1-induced histamine release from allergic patients' basophils. Allergen-specific human IgG mAbs such as BAB1, which presents high blocking activity in both immunochemical and cellular IgE competition experiments, might have therapeutical application.

Adjuvants, Immunologic↗
PubMed1996

Inability to produce IL-6 is a functional feature of human germinal center B lymphocytes.

In response to Ag encounter, B lymphocytes undergo a complex maturation process yielding phenotypically distinct subpopulations that are located in highly organized compartments of secondary lymphoid organs. This study describes the patterns of cytokine secretion of naive, memory, and germinal center (GC) human tonsillar B lymphocytes, activated either through CD40 or B cell receptor or with Staphylococcus aureus Cowan I particles. The three B cell subpopulations produced comparable levels of IL-10 and TNF-alpha, regardless of the stimulation pathway. Interestingly, activated GC B lymphocytes fail to express IL-6, as determined both at mRNA and at protein levels, whereas both naive and memory B cells can be induced to secrete IL-6. Likewise, naive B lymphocytes undergoing dual ligation of CD40 and B cell receptor fail to express IL-6, since they acquire a GC-like phenotype. IL-6 receptors are up-regulated on both ex vivo-purified GC B lymphocytes and in vitro generated GC-like B cells, following CD40 activation. Consistent with this, addition of exogenous IL-6 sustains growth of CD40-stimulated GC B lymphocytes. Taken together, these results demonstrate that loss of IL-6 secretion is a functional characteristic of human GC B lymphocytes. The swap from an autocrine to a paracrine IL-6 response may permit a better control of B cell growth and differentiation during the germinal center reaction.

Antilymphocyte Serum↗
PubMed1996

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

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.

Antibodies, Anti-Idiotypic↗
PubMed1996

Four IgG anti-islet human monoclonal antibodies isolated from a type 1 diabetes patient recognize distinct epitopes of glutamic acid decarboxylase 65 and are somatically mutated.

The selective destruction by an autoimmune process of the beta cells in the pancreas is the hallmark of the type 1 insulin-dependent diabetes mellitus. What triggers islet cell-specific autoreactive T and B cells, however, remains unclear. Identification of the targets of the anti-islet cell autoantibodies frequently found in insulin-dependent diabetes mellitus patients and analysis of their sequences should provide some insights into the nature of this disease. We have combined EBV transformation with CD40 activation of peripheral B cells from one patient with insulin-dependent diabetes mellitus to isolate four B cell clones that secrete islet cell-specific autoantibodies, These four human monoclonal autoantibodies are of the IgG1 isotype, and they each recognize a different epitope of the glutamic acid decarboxylase enzyme. Analysis of their variable gene sequences shows that, while clonally unrelated, three of the four human monoclonal autoantibodies use a member of the VH4 family, and two have rearranged the same delta light chain variable gene. The IgG1 isotype of the four autoantibodies as well as the presence of somatic mutations in both heavy and light chain genes provide concrete evidence for their derivation by a T cell-dependent immune response.

Adult↗
PubMed1996

CD40 and B cell antigen receptor dual triggering of resting B lymphocytes turns on a partial germinal center phenotype.

Phenotypic alterations occur when resting human B lymphocytes become germinal center (GC) cells. These include the induction of surface CD38, CD95 (FAS/APO-1), and carboxy-peptidase-M (CPM), a recently described GC marker. However, the factors that govern the in vivo induction of these surface molecules on B cells remain unknown. Here, we purified resting (CD38-) human B lymphocytes from tonsils in an attempt to establish culture conditions resulting in the induction of these three GC markers. We show that interferon (IFN) alpha or IFN-gamma, as well as antibodies against the B cell antigen receptor (BCR), could induce CD38 on resting B lymphocytes, a phenomenon further enhanced by CD40 stimulation. Concomitantly, CD95 was upregulated by CD40 ligation and, to a lesser extent, by IFN-gamma. By contrast, CPM expression could be upregulated only through BCR triggering. This CPM induction was specifically enhanced by CD19 or CD40 ligation. CD40 + BCR stimulation of resting B cells with CD40 ligand-transfected fibroblastic cells in the presence of cross-linked anti-BCR monoclonal antibodies resulted in the coexpression of CD38, CD95, and CPM. As GC cells, these cells also expressed CD71, CD80 (B7.1), and CD86 (B7.2), but not CD24. However, CD10+ or CD44- B cells could not be detected in these culture conditions, suggesting that yet other signals are required for the induction of these GC markers. Consistent with a GC phenotype, CD40 + BCR-stimulated cells exhibited reduced viability when cultured for 20 h in the absence of stimulus. These results first demonstrate that cotriggering of resting B cells through BCR and CD40 induces both phenotypic and functional GC features. They also show that IFN and CD19 triggering of resting B cells specifically modulate the expression of GC markers.

ADP-ribosyl Cyclase↗
PubMed1996

Equilibrium values of measures of population subdivision for stepwise mutation processes.

Expected values of WRIGHT's F-statistics are functions of probabilities of identity in state. These values may be quite different under an infinite allele model and under stepwise mutation processes such as those occurring at microsatellite loci. However, a relationship between the probability of identity in state in stepwise mutation models and the distribution of coalescence times can be deduced from the relationship between probabilities of identity by descent and the distribution of coalescence times. The values of FIS and FST can be computed using this property. Examination of the conditional probability of identity in state given some coalescence time and of the distribution of coalescence times are also useful for explaining the properties of FIS an FST at high mutation rate loci, as shown here in an island model of population structure.

Alleles↗
PubMed1996

Testing differentiation in diploid populations.

We examine the power of different exact tests of differentiation for diploid populations. Since there is not necessarily random mating within populations, the appropriate hypothesis to construct exact tests is that of independent sampling of genotypes. There are two categories of tests, FST-estimator tests and goodness of fit tests. In this latter category, we distinguish "allelic statistics", which account for the nature of alleles within genotypes, from "genotypic statistics" that do not. We show that the power of FST-estimator tests and of allelic goodness of fit tests are similar when sampling is balanced, and higher than the power of genotypic goodness of fit tests. When sampling is unbalanced, the most powerful tests are shown to belong to the allelic goodness of fit group.

Alleles↗
PubMed1996

Molecular identification of a Wolbachia endosymbiont in a Tetranychus urticae strain (Acari: Tetranychidae).

Wolbachia, a maternally inherited bacterium, is involved in alterations of arthropod sexuality. Reproductive incompatibilities are often observed in miles, but the existence of this microorganism in their cytoplasm has not yet been demonstrated. We identified the presence of Wolbachia in a strain of the spider mite Tetranychus urticae based on the amplification and sequencing of part of the 16S rDNA and ftsZ genes. In order to establish the phylogenetic relationships between Wolbachia found in T. urticae and in other arthropods, we aligned the resulting sequences with already published ones. For both 16S and ftsZ genes the Wolbachia carried by T. urticae clustered together with Wolbachia found in other arthropods.

Animals↗
PubMed1995

Evolution of single and double Wolbachia symbioses during speciation in the Drosophila simulans complex.

Maternally inherited bacteria of the genus Wolbachia are responsible for the early death of embryos in crosses between uninfected females and infected males in several insect species. This phenomenon, known as cytoplasmic incompatibility, also occurs between strains infected by different symbionts in some species, including Drosophila simulans. Wolbachia was found in two species closely related to D. simulans, Drosophila mauritiana, and Drosophila sechellia, and shown to cause incompatibility in the latter species but not in D. mauritiana. Comparison of bacterial and mtDNA history clarifies the origins of bacterial and incompatibility polymorphisms in D. simulans. Infection in D. mauritiana is probably the result of introgression of an infected D. simulans cytoplasm. Some D. simulans and D. sechellia cytoplasmic lineages harbor two bacteria as a consequence of a double infection which probably occurred in a common ancestor. The descendant symbionts in each species are associated with similar incompatibility relationships, which suggests that little variation of incompatibility types has occurred within maternal lineages beyond that related to the density of symbionts in their hosts.

Animals↗
PubMed1995

Endogenous IL-6 and IL-10 contribute to the differentiation of CD40-activated human B lymphocytes.

This study was initiated to explore the contribution of endogenous cytokines to CD40-induced B cell proliferation and differentiation. First, both CD40 and Ag receptor (AgR) cross-linking induced purified tonsillar human B lymphocytes to secrete the same pattern of cytokines, including IL-1 beta, IL-6, IL-10, granulocyte-macrophage-CSF, and TNF-alpha, whereas IL-1 alpha, IL-2, IL-3, IL-4, IL-5, IL-7, granulocyte (G)-CSF, or IFN-gamma were not detected. Second, cotriggering of CD40 and AgR resulted in additive secretion of both IL-6 and IL-10. Addition of IL-4 to CD40-activated B cells increased IL-6 levels but decreased IL-10 levels. In contrast, exogenous IL-10 diminished IL-6 levels. Neutralization of IL-6 and IL-10 using blocking Abs did not alter CD40-induced B cell growth. In contrast, IL-6 neutralization markedly inhibited the IL-4-induced IgE secretion (57 +/- 10%) as well as the IgG and IgM production resulting from AgR and CD40 cotriggering (49 +/- 16.5 and 29.5 +/- 4.5%, respectively). Blocking IL-10 inhibited the IgA secretion (25 +/- 2.7%) obtained after CD40 activation and the production of IgG, IgA, and IgM (24.1 +/- 5.6, 25 +/- 8, and 42 +/- 6.5%, respectively) by B lymphocytes undergoing dual ligation of CD40 Ag and AgR. Simultaneous neutralization of both endogenous IL-6 and IL-10 resulted in an increased inhibition of Ig secretion for B cells cotriggered by CD40 Ag and AgR (65-75%). Thus, endogenously produced IL-6 and IL-10 are involved in the differentiation of CD40-activated B cell.

Antigens, CD↗
PubMed1995

Anti-CD40 plus interleukin-4-activated human naive B cell lines express unmutated immunoglobulin genes with intraclonal heavy chain isotype variability.

Combination of anti-CD40 antibody and interleukin-4 (IL-4) induces B cell clonal expansion reminiscent of the T-dependent proliferation following antigenic challenge in vivo. We have analyzed the usage of CH genes and the presence or absence of somatic mutations within the progeny of a single human naive B cell activated with anti-CD40 + IL-4. To address this issue, single-cell cultures of naive (sIgD+) tonsillar B lymphocytes expressing the VH1-restricted G8 idiotype were set up. After culture and RNA extraction, VH1+ Ig mRNA were reverse-transcribed, amplified by polymerase chain reaction and sequenced. A single sIgD+ B cell could generate clones expressing mu, gamma 1, gamma 3, or epsilon, illustrating that the progeny of a single cell can express different isotypes in response to the same stimulus in vitro. The rate of somatic mutations affecting the immunoglobulin variable heavy chain gene was indistinguishable from the background of errors introduced by Taq polymerase.

Antibodies, Anti-Idiotypic↗
PubMed1995

Testing heterozygote excess and deficiency.

Currently used tests of Hardy-Weinberg proportions do not take into account the nature of the alternative hypothesis, which is generally a heterozygote deficiency. Different exact tests, appropriate for small sample size and large number of alleles, are proposed in this perspective, and their properties are evaluated by power comparisons. Some tests are found to be close to optimal for the detection of inbreeding or heterozygote excess, one of which is a score test closely related to Robertson and Hill's estimator of the inbreeding coefficient. This test is also easily applied to multiple samples. Such tests are not always the most appropriate if alternative hypotheses differ from those considered here.

Alleles↗
PubMed1995

Long-term cultured CD40-activated B lymphocytes differentiate into plasma cells in response to IL-10 but not IL-4.

We compared the effects of IL-10 and IL-4 on the functions of B lymphocytes triggered through their CD40. During the initial phase, IL-10 was as potent as IL-4 in inducing the expansion of viable B cells. Then, cellular expansion slowed down and after approximately 3 weeks the number of B cells started to decline. While the combination of IL-10 and IL-4 was synergistic during the first 2 weeks of culture, B cell recovery declined after 3 weeks, indicating that IL-10 prevails over IL-4. Those effects were not restricted to a specific B cell subset as both sIgD+ B cells and sIgD- B cells behaved in a similar way, though the latter population responded with a slightly accelerated kinetic. Inverted microscope examination and scanning electron microscopy showed that in response to IL-10, CD40-activated B cell cultures were heterogeneous with loose aggregates of cells as well as free floating large ovoid cells. In contrast, in the presence of IL-4, CD40-activated B cell cultures were essentially composed of tight cell clumps. IL-10 progressively induced all B cells to differentiate into non-replicating cells with intracytoplasmic Ig that secreted Ig at a high rate. Cytologic analysis indicated that IL-10 cultured cells display a basophilic cytoplasm with an arcoplasm and a low nucleus/cytoplasm ratio. Transmission electron microscopy demonstrated that when IL-10 was added to the culture, B cells displayed structures for excretion with extended endoplasmic reticulum and dilated cisternae containing paracrystalline structures, typical of plasmablasts cells. Taken together, these results indicate that IL-10 acts as a plasma cell differentiation factor for CD40-activated B cells.

Antibody-Producing Cells↗
PubMed1994

Human monoclonal autoantibodies specific for the bullous pemphigoid antigen 1 (BPAg 1).

Bullous pemphigoid (BP) is an acquired blistering skin disease associated with the production of IgG autoantibodies to the 230-kDa BP Ag (BPAg1). To better characterize the epitopes of BPAg1, we generated immortalized B cell lines secreting human mAbs (HumAbs) to BPAg1 from two BP patients whose sera reacted with native BPAg1 but not with a recombinant BP55 carboxyl-terminal peptide. Ab-producing B cell lines were established by EBV infection of CD40-activated PBMCs. Three independent clonal lines were obtained that secreted IgG HumAbs, including one IgG1 kappa (BP3) and two distinct IgG4 kappa (BP1 and BP2). These three HumAbs immunoprecipitated BPAg1. Blocking immunofluorescence experiments and phylogenetic studies showed that these Abs recognize different epitopes of BPAg1. This analysis with HumAbs further extends the serologic demonstration of the wide variety of epitopes recognized by BPAg1 autoantibodies which contrasts with the limited number of epitopes recognized by thyroid peroxidase monoclonal autoantibodies.

Antibodies, Monoclonal↗