bluuebunny/crossref_metadata_2025_split huggingface dataset:
Staphylococcal enterotoxin B inhibits the production of interleukin‐4 in a human mast‐cell line HMC‐1
Authors: Ackermann L. , Pelkonen J. , Harvima I. T. 
Published: March 2003

Staphylococcal enterotoxins belong to the recently characterized group of immunocytotropic bacterial superantigens that are potent mitogens for human T cells. Superantigens are presented, but without intracellular processing, to T cells by monocyte/macrophages, Langerhans’ cells and keratinocytes via the class II major histocompatibility complex (MHC) molecules. Superantigens have been demonstrated to act as potent inducers of several proinflammatory cytokines in the antigen‐presenting cells such as interleukin‐1 (IL‐1) and tumour necrosis factor‐α (TNF‐α). As mast cells participate in the pathogenesis of several inflammatory skin disorders such as atopic dermatitis (AD), which is often aggravated by staphylococcal infections, we studied the effect of staphylococcal enterotoxin B (SEB) superantigen on the histamine release and IL‐4 expression in a human mast‐cell line (HMC‐1). Incubation of SEB (50 μg/ml) with HMC‐1 cells for 45 min, could not induce any histamine release. The HMC‐1 cells were also stimulated with various concentrations of SEB (0, 1, 10, 20, 50 μg/ml) for 1, 2, 3 and 4 days. Clear dose‐dependent inhibition of IL‐4 protein production and release was observed on day 4 without any observed effect on cell viability. Compared with unstimulated HMC‐1 cells, after 50 μg/ml SEB stimulation, the IL‐4 mRNA levels decreased steadily in the 2, 6, 18 and 24 hr samples in repeated experiments as measured with the reverse transcription–polymerase chain reaction (RT–PCR) method. In comparison, a biphasic decrease in TNF‐α expression was found. Our results show that in human leukaemic mast cells, superantigen stimulation downregulates the production of IL‐4.IL, interleukin TNF‐α, tumour necrosis factor‐alpha AD, atopic dermatitis SEB, Staphylococcal enterotoxin B Th, T helper IFN‐γ, interferon‐γ ICAM‐1, intercellular adhesion molecule‐1 RT–PCR, reverse transcription–PCR.
Both adhesion to immobilized vitronectin and FcɛRI cross‐linking cause enhanced focal adhesion kinase phosphorylation in murine mast cells
Authors: Bhattacharyya S. P. , Mekori Y. A. , Hoh D. , Paolini R. , Metcalfe D. D. , Bianchine P. J. 
Published: March 2003

SummaryMurine mast cells adhere spontaneously to plate‐bound vitronectin (VNPB) via αv‐containing integrins, and this adhesive interaction results in an augmented interleukin‐3 (IL‐3)‐dependent mast‐cell proliferation. In this report we demonstrate that the activation of murine mast cells through αv‐integrin, as well as through the high affinity immunoglobulin E (IgE) receptor (FcɛRI), results in enhanced tyrosine phosphorylation of focal adhesion kinase (FAK), a cytoplasmic protein tyrosine kinase involved in mitogenic and oncogenic signal transduction. While mast cell adhesion to VNPB resulted in enhanced FAK phosphorylation, treatment with soluble vitronectin (VNSOL) failed to do so. Spontaneous mast cell adhesion to entactin (EN) did not induce tyrosine phosphorylation of FAK, demonstrating that not all adhesive interactions lead to the same sequence of biochemical events. Because FAK has intrinsic tyrosine kinase activity, we examined whether activating mast cells via αv‐integrins, or via FcɛRI‐cross‐linking stimulated the in vitro kinase activity of FAK. Both pathways were found independently to activate FAK in mast cells and together appeared additive. Protein kinase C depletion in mast cells and calcium depletion in the medium caused decreased tyrosine phosphorylation of FAK, indicating that optimal tyrosine phosphorylation of FAK is regulated by both pathways. These data are consistent with the conclusion that the tyrosine phosphorylation of FAK represents at least one example of a point of convergence in the intracellular tyrosine phosphorylation cascades induced by αv integrin‐and FcɛRI‐mediated signal transduction pathways in mast cells.
Induction of interactions between CD44 and hyaluronic acid by a short exposure of human T cells to diverse pro‐inflammatory mediators
Authors: Ariel A. , Lider, O. , Brill A. , Cahalon L. , Savion N. , Varon D. , Hershkoviz R. 
Published: March 2003

SummaryMigration of T cells into extravascular sites of inflammation is mediated by cell–cell and cell–matrix adhesion receptors, including the hyaluronan‐binding glycoprotein, CD44. The biochemical nature of CD44 variants and the ligand specificity, function and the regulation of activation of CD44 expressed on various cell types have been extensively studied. However, little is still known about the short‐term influence of cytokines and chemokines on the activation of CD44 on human T cells. Therefore, we studied the role of inflammatory mediators in regulating the adhesion of T cells from human peripheral blood to immobilized hyaluronan under static or shear stress conditions. We found that the CD44‐dependent adhesion, under static and shear stress (i.e. relative gradual resistance to flow of 150 and 1500 s−1) conditions, of T cells to hyaluronan requires a T‐cell activation of 2–3 hr and is regulated by the cross‐linking of CD3, cytokines (e.g. interleukin‐2 and tumour necrosis factor‐α), and chemokines (e.g. MIP‐1β, interleukin‐8, and RANTES). This T‐cell adhesion was manifested by polarization, spreading and co‐localization of cell surface CD44 with a rearranged actin cytoskeleton in hyaluronan‐bound T cells. Thus, cytokines and chemokines present in the vicinities of blood vessel walls or present intravascularly in tissues where immune reactions take place, can rapidly activate the CD44 molecules expressed on T cells.
Signals sustaining human immunoglobulin V gene hypermutation in isolated germinal centre B cells
Authors: Dahlenborg K. , Pound J. D. , Gordon J. , Borrebaeck C. A. K. , Carlsson R. 
Published: March 2003

SummaryAffinity maturation of antibody responses depends on somatic hypermutation of the immunoglobulin V genes. Hypermutation is initiated specifically in proliferating B cells in lymphoid germinal centres but the signals driving this process remain unknown. This study identifies signals that promote V gene mutation in human germinal centre (GC) B cells in vitro. Single GC B cells were cultured by limiting dilution to allow detection of mutations arising during proliferation in vitro. Cells were first cultured in the presence of CD32L cell transfectants and CD40 antibody (the ‘CD40 system’) supplemented with combinations of cytokines capable of supporting similar levels of CD40‐dependent GC B‐cell growth [interleukin (IL)‐10 + IL‐1β + IL‐2 and IL‐10 + IL‐7 + IL‐4]. Components of the ‘EL4 system’ were then added to drive differentiation, providing sufficient immunoglobulin mRNA for analysis. Analysis of VH3 genes from cultured cells by reverse transcription–polymerase chain reaction (RT–PCR)‐based single‐strand conformation polymorphism indicated that the combination IL‐10 + IL‐1β + IL‐2 promoted active V gene mutation whereas IL‐10 + IL‐7 + IL‐4 was ineffective. This was confirmed by sequencing which also revealed that the de novo generated mutations were located in framework and complementarity‐determining regions and shared characteristics with those arising in vivo. Somatic mutation in the target GC B‐cell population may therefore be actively cytokine driven and not simply a consequence of continued proliferation. The experimental approach we describe should facilitate further studies of the mechanisms underlying V gene hypermutation.
CD5‐positive and CD5‐negative human B cells converge to an indistinguishable population on signalling through B‐cell receptors and CD40
Authors: Gagro A. , Mccloskey N. , Challa A. , Holder M. , Grafton G. , Pound J. D. , Gordon J. 
Published: March 2003

SummaryWhether CD5 on B cells marks a subset functionally distinct from the conventional CD5 negative (CD5neg) adult population or is more an indicator of activation, remains contentious. Here we have investigated whether CD5 positive (CD5pos) and CD5neg B cells can be distinguished in terms of their response to surrogate signals aimed to model, in vitro, T‐cell dependent (TD) and T‐independent (TI) encounters with antigen in vivo: the predominantly CD5pos B‐cell population found in cord blood, CD5 B cells positively selected from tonsils and their CD5neg counterparts, were compared. Neonatal B cells displayed a near‐identical phenotype to that of adult CD5pos B cells, being characterized by uniform immunoglobulin M (IgM), immunoglobulin D (IgD), CD23 and CD44 coexpression. When cultured with anti‐IgM maintained at high density on CD32‐tranfected mouse L cells to model TI responses or on CD40 ligand (CD40L)‐bearing L cells (with or without captured anti‐IgM) to model TD encounters, DNA synthesis was stimulated to a similar extent in all three populations. Focusing on CD5 and CD23, we found that – although the signals delivered promoted distinct profiles of expression – under each condition of activation, the phenotypes that emerged for adult CD5pos and CD5neg B cells were remarkably similar. Neonatal B cells displayed a greater diminution in CD5 expression than adult CD5pos B cells following CD40 signals but otherwise the two populations again behaved similarly. The inclusion of interleukin‐4 (IL‐4) to cultures where cells were costimulated via surface (s)IgM and CD40 resulted in a complete loss of CD5 expression and a corresponding hyperexpression of CD23, irrespective of the population studied. The near‐identical response of CD5pos and CD5neg B cells to surrogate TD or TI signals in vitro and their convergence to indistinguishable phenotypes is wholly supportive of CD5 being a fluctuating marker of activation rather than it delineating functionally distinct subsets.
A novel subset of murine B cells that expresses unmasked forms of CD22 is enriched in the bone marrow: implications for B‐cell homing to the bone marrow
Authors: Floyd H. , Nitschke L. , Crocker P. R. 
Published: March 2003

SummaryCD22 is a B‐cell‐restricted transmembrane protein, which acts as a negative regulator of B‐cell signalling. CD22 also has lectin‐like adhesive properties. When expressed on transfected fibroblasts, it is capable of mediating adhesion to other cells via recognition of cell‐surface glycoconjugates terminating in α2,6‐linked sialic acids. In previous studies in the mouse, CD22 was implicated as a bone marrow homing receptor for recirculating immunoglobulin D+ (IgD+) B cells through recognition of sialylated ligands on marrow sinusoidal endothelium. As the adhesive function of CD22 can be masked when α2,6‐linked sialic acids are co‐expressed at the cell surface, the aim of the present study was to investigate whether recirculating B cells have unmasked forms of CD22 that could be involved in bone marrow homing. Using α2,6‐sialyllactose coupled to biotinylated polyacrylamide as a probe for detection of unmasked CD22, we showed that ≈ 2–5% of IgD+ murine B cells in the spleen and mesenteric lymph nodes were able to bind this synthetic ligand. In the bone marrow, however, the fraction of IgD+ B cells with unmasked CD22 was increased by two‐ to fivefold. B cells from CD22‐deficient mice were not stained with the polyacrylamide probe, confirming that staining of B cells in wild‐type mice was caused by CD22 and not by other potential sialic acid‐binding lectins. In conclusion, we have identified a new subset of mature B cells in the mouse with unmasked CD22. This subset of recirculating B cells may bind to CD22 ligands on bone marrow sinusoidal endothelium, leading to their selective homing and subsequent enrichment in this tissue.
Graft‐versus‐host‐disease‐associated donor cell engraftment in an F1 hybrid model is dependent upon the Fas pathway
Authors: Iwasaki T. , Hamano T. , Saheki K. , Kuroiwa T. , Kataoka Y. , Takemoto Y. , Ogata A. , Fujimoto J. , Kakishita E. 
Published: March 2003

SummaryThe graft‐versus‐host disease (GVHD) generated in BDF1 mice by the injection of spleen cells from the C57BL/6 parental strain induces a direct cell‐mediated attack on host lymphohaematopoietic populations, resulting in the reconstitution of the host with donor cells. We examined Fas–Fas ligand (FasL) interactions in donor and host haematopoietic cells over a prolonged period of parental‐induced GVHD. Fas expression on bone marrow cells of both donor and host origin increased at 2 weeks. Host cell incubation with anti‐Fas antibody induced apoptosis, and the number of haematopoietic progenitor cells decreased. Fas‐induced apoptosis by the repopulating donor cells, however, did not increase until 12 weeks, when more than 90% of the cells were donor cells. The expression of various cytokines, such as interferon‐γ (IFN‐γ) and tumour necrosis factor‐α (TNF‐α), and FasL gene expression in the bone marrow increased concomitantly. To examine directly whether FasL has a major role in the development of donor cell engraftment, FasL‐deficient (gld) mice were used as donors. Injection of B6/gld spleen cells induced significantly less host lymphohaematopoietic depletion, resulting in a failure of donor cell engraftment. Furthermore, injection of IFN‐γ gene knockout (gko) B6 spleen cells failed to augment Fas and FasL expression in recipient mice, resulting in a failure of donor cell engraftment. This suggests that the induction of apoptosis by Fas–FasL interactions in host cells may contribute to a reconstitution of the host with donor cells and that donor‐derived IFN‐γ plays a significant role for Fas–FasL interactions in host cells during parental‐induced GVHD.
Bovine tuberculosis: immune responses in the peripheral blood and at the site of active disease
Authors: Rhodes S. G. , Buddle B. M. , Hewinson R. G. , Vordermeier H. M. 
Published: March 2003

SummaryThis report describes a comparison of immune responses in the peripheral blood and at the site of active disease in cattle 20 weeks after experimental infection with Mycobacterium bovis. Lymphocyte proliferation, and the production of interferon‐γ (IFN‐γ) and interleukin (IL)‐2 were measured in response to tuberculin and a number of mycobacterial antigens, including ESAT‐6, MPB64, MPB70, MPB83, hsp 16.1, hsp 65, hsp 70 and the 38 000 MW lipoprotein antigen. The level of transforming growth factor‐β (TGF‐β) was measured following stimulation of cells with tuberculin. Our results suggest little difference in the responses of peripheral blood and lymph node cells to most of the antigens used. However, tuberculin purified protein derivative (PPD) and ESAT‐6 elicited stronger responses in the peripheral blood compared with lymph node cells. Investigation of the responding T‐cell subpopulations in the peripheral blood showed that both CD4+ and, to a lesser extent, γδ T‐cell receptor‐positive (TCR+) T cells contributed to these responses. This is the first report to compare peripheral and local immune responses in bovine tuberculosis. Unlike cases of human tuberculosis where immune activity at the site of disease and anergy in the peripheral blood have been reported, our results suggest that for bovine tuberculosis immune responses occurring in the peripheral blood reflect those at the site of disease.
Neutralizing antibodies to granulocyte–macrophage colony‐stimulating factor, interleukin‐1α and interferon‐α but not other cytokines in human immunoglobulin preparations
Authors: Wadhwa M. , Meager A. , Dilger P. , Bird C. , Dolman C. , Das R. G. , Thorpe R. 
Published: March 2003

SummaryHuman immunoglobulin preparations are used therapeutically for various disorders. Such therapy is generally safe but adverse effects occasionally occur in recipients. It has been suggested that antibodies to cytokines present in clinical immunoglobulin products may contribute to undesirable effects in recipients. Therefore, we investigated intravenous and intramuscular immunoglobulin products for the presence of cytokine‐specific neutralizing antibodies. Using validated bioassays, we detected neutralizing activity against human granulocyte–macrophage colony‐stimulating factor (GM‐CSF), interferon‐α2a (IFN‐α2a) and interleukin‐1α (IL‐1α) in immunoglobulin products. We found no neutralization of granulocyte colony‐stimulating factor, macrophage colony‐stimulating factor, stem cell factor, IL‐1β, IL‐2, IL‐3, IL‐4, IL‐6, IL‐9, IL‐10, IL‐12, tumour necrosis factor‐α, oncostatin M (OSM) and IFN‐γ. Most batches which neutralized IFN‐α2a activity also neutralized other IFN‐α subtypes, IFN‐ω and IFN‐β. Most products (94%) neutralized the biological activity of GM‐CSF. No correlation between batches and their ability to neutralize bioactivities of GM‐CSF, IFN‐α2a and IL‐1α was found. This neutralizing activity could be traced to plasma pools used for manufacture of immunoglobulins. The neutralization was mediated by specific cytokine antibodies contained within immunoglobulin products as it was present in specific immunoglobulin G (IgG) fractions eluted from cytokine affinity chromatography columns. Specific binding of such IgG fractions to cytokines in immunoblots and in enzyme‐linked immunosorbent assays (ELISAs) was observed. This contrasts with the broad non‐specific recognition of cytokine proteins observed using unfractionated immunoglobulins in ELISAs. This is the first comprehensive study showing the presence of neutralizing antibodies against GM‐CSF, IL‐1α, or IFN‐α2a in immunoglobulin products.
Characterization and role in experimental systemic lupus erythematosus of T‐cell lines specific to peptides based on complementarity‐determining region‐1 and complementarity‐determining region‐3 of a pathogenic anti‐DNA monoclonal antibody
Authors: Brosh N. , Eilat E. , Zinger H. , Mozes E. 
Published: March 2003

SummaryPeptides based on the complementarity‐determining region 1 (CDR1) and CDR3 of an anti‐DNA monoclonal antibody (mAb) carrying the 16/6 idiotype (Id) were shown to induce experimental systemic lupus erythematosus (SLE) in susceptible mouse strains. In the present study, T‐cell lines specific to the pCDR1 and pCDR3 peptides were established in BALB/c and in SJL mice, respectively. The T‐cell lines were characterized and analysed for their pathogenicity upon administration to syngeneic mouse strains. Both T‐cell lines expressed the αβ T‐cell receptor (TCR) and the CD4+ CD8– phenotype. Additionally, both cell lines secreted interleukin (IL)‐4 and IL‐10 upon stimulation with their specific peptide, thus belonged to the T helper 2 (Th2) subset. Upon immunization, the pCDR3‐specific T‐cell line induced experimental SLE in SJL mice. The animals produced high levels of autoimmune anti‐DNA and antinuclear protein antibodies, as well as anti‐16/6 Id antibodies (Abs). Furthermore, the mice developed clinical manifestations, including leukopenia, proteinuria and accumulation of immune complex deposits in their kidneys. The pCDR1‐specific T‐cell line failed to induce SLE when injected into BALB/c mice. It is thus suggested that pCDR3 is an immunodominant epitope in experimental SLE and that pCDR3‐specific T cells initiate autoimmunity, leading to SLE, probably via epitope spreading.
Effective formation of major histocompatibility complex class II–peptide complexes from endogenous antigen by thyroid epithelial cells
Authors: Maile R. , Elsegood K. A. , Harding T. C. , Uney J. B. , Stewart C. E. H. , Banting G. , Dayan C. M. 
Published: March 2003

SummaryIn autoimmune thyroid disease, thyroid epithelial cells (TEC) express major histocompatibility complex (MHC) class II molecules, potentially enabling them to present thyroid self‐antigens to CD4‐positive T cells. However, despite this, TEC may fail to present endogenous antigen as a result of limited processing or MHC class II loading capacity, or inadequate MHC class II levels. We addressed these issues using the cloned rat TEC line, Fischer rat thyroid cell line (FRTL5), which was transfected using an adenoviral expression vector that expressed ovalbumin (OVA) as an integral membrane protein. OVA‐expressing FRTL5 cells very efficiently activated a panel of OVA‐specific, class II‐restricted T‐cell hybridomas. This response was dependent on induction of MHC class II molecules by interferon‐γ (IFN‐γ) and was blocked by anti‐MHC class II antibodies. Poor responses were seen to exogenously added OVA or OVA peptides. These results provide the most direct evidence to date that TEC can form MHC class II–peptide complexes derived from self‐antigen in sufficient quantities to activate T cells.
Structural and functional analysis of the human CD45 gene (PTPRC) upstream region: evidence for a functional promoter within the first intron of the gene
Authors: Timón M. , Beverley P. C. L. 
Published: March 2003

SummaryExpression of the leucocyte common antigen (CD45) in mammals is restricted to the nucleated lineages of haematopoietic cells. It appears in early progenitors in the bone marrow and is expressed at the surface of these cells throughout their differentiation. However, at least in T cells, the pattern of expression switches between different isoforms during the successive stages of differentiation in the thymus and after activation in the periphery. In order to understand the mechanisms controlling the transcription of the human CD45 gene, 2·7 kbp of the 5′‐flanking region were sequenced and analysed for their ability to direct expression of a reporter gene. The only region with promoter activity was localized within the first intron of the gene. This promoter shows no tissue specificity but could be enhanced by a heterologous enhancer. Mobility shift assays showed complex but specific protein binding. The sequence in this region lacks similarity with known promoters or initiators but is highly conserved in evolution. No transcription initiation could be detected within or downstream of this region, suggesting that this might be a new type of RNA polymerase II promoter able to drive transcription from an upstream sequence. An additional exon was also found upstream of exon 1. The two exons 1 (1a and 1b) are mutually exclusive and both are spliced to exon 2. This makes the structure of the 5′ region of the human CD45 gene identical to its mouse homologue.
Polymorphism of the human α1 immunoglobulin gene 3′ enhancer hs1,2 and its relation to gene expression
Authors: Denizot Yves , Pinaud Eric , Aupetit Corinne , Le Morvan Caroline , Magnoux Emmanuelle , Aldigier Jean Claude , Cogné Michel 
Published: March 2003

SummaryWe studied the hs1,2 transcriptional enhancer identified downstream of the human α1 gene of the immunoglobulin H (IgH) locus, for which two different allelic configurations (a and b) were previously reported by Southern blotting. By using a polymerase chain reaction (PCR) method we amplified minisatellites within the hs1,2 core enhancer, with variable numbers of tandem repeats (VNTR) defining three ‘PCR alleles’α1A, α1B and α1C (including one, two and three repeats, respectively). Five different α1 h1,2 genotypes were encountered in a population of 513 donors, representing 13·8, 34·5, 49·7, 1·3 and 0·6% for the AA, BB, AB, AC and BC genotypes, respectively. Luciferase assays showed that increasing the number of minisatellites increased the transcriptional strength of the α1 hs1,2 enhancer. Simultaneous determination of Southern blot alleles and VNTR alleles only showed a partial linkage between both types of polymorphism, altogether defining at least six different allelic forms of the 3′α1 region. In conclusion, the present study further demonstrates the genetic instability of the 3′α region, for which multiple alleles have been generated through inversions and internal deletions and/or duplications. This study also strengthens the hypothesis that the polymorphism at the IgH 3′ regulatory region of the α1 gene could play a role in the outcome of diseases involving immunoglobulin secretion.