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E Baixeras

Publications and source records attributed to E Baixeras.

16 recordsLinked to original sources
PubMed1997

B cell receptor cross-linking prevents Fas-induced cell death by inactivating the IL-1 beta-converting enzyme protease and regulating Bcl-2/Bcl-x expression.

In the A20 cell line, we examined the mechanisms that modulate the Fas-mediated apoptotic pathway through the B cell receptor. As in other systems, Fas signaling activates cysteine proteases, leading to specific proteolysis of poly(ADP-ribose) polymerase (PARP) and protein kinase C (PKC) delta. We describe that PKC-epsilon and PKC-zeta proteins are two new IL-1 beta-converting enzyme (ICE) substrates; we found that ICE activation and its proteolytic effects are inhibited by surface IgG (sIgG) cross-linking. Apoptosis induced by Fas ligation is consequently abrogated after sIgG engagement, and sIgG signaling therefore interferes with the apoptotic signal upstream of ICE protease activation. Since the PKC inhibitor bisindolylmaleimide I completely abolishes the protective effect of the sIgG signal, a member of the PKC family is probably responsible for the prevention of ICE cascade activation. Direct activation of PKC by PMA partially mimics the protective effect of sIgG cross-linking against Fas-mediated death in A20 cells. Nevertheless, PMA inhibits neither ICE activation nor the subsequent proteolysis of ICE substrates, suggesting that the PKC responsible for ICE inactivation is a non-PMA-sensitive PKC. In this system, Fas ligation also triggers Bcl-2/Bcl-x down-regulation, an effect inhibited by sIgG cross-linking, the cysteine protease inhibitor acetyl-Tyr-Val-Ala-Asp-chloromethyl ketone, and PMA treatment. In A20 cells, Fas signaling may thus trigger both ICE activation and Bcl-x and Bcl-2 down-regulation. These results indicate that sIgG signaling gives rise to two pathways after PKC activation, one presumably promoted by non-PMA-sensitive PKC, which inactivates the ICE cascade, and another produced by PMA-sensitive PKC, which maintains normal Bcl-2/Bcl-x levels.

Amino Acid Chloromethyl Ketones↗
PubMed1996

CD2 ligation abrogates antigen-independent apoptosis in B cells.

We have described recently the prevention of apoptosis by CD2-soluble CD48 interaction on antigen B cell receptor occupancy. Here, we show that CD2 ligation is also able to interfere with B cell receptor-independent apoptosis pathways such as spontaneous death in spleen B cells or serum deprivation and hydrogen peroxide exposure in the BAL-17 cell line. In all cases, CD2 ligation induces a signal that prevents the downregulation of Bcl-2 expression. The specific CD2 signal pathway involved in this phenomenon is still unknown. As reported, CD2 did not appear to induce Ca2+ mobilization, phosphatidylinositol turnover, or PKC translocation in B cells. Nevertheless, we show that CD2 receptor ligation is coupled to the tyrosine phosphorylation pathway in B cells. These observations indicate that CD2 is functionally able to trigger at least an early signal that could play a role in apoptosis blockage B cells in addition to the adhesion one. The results suggest the participation of cellular membrane receptors other that CD40 in apoptosis rescue, not only in the antigen-dependent but also in the antigen-independent phases of B cell lymphopoiesis.

Animals↗
PubMed1996

Decrease in cAMP levels promoted by CD48-CD2 interaction correlates with inhibition of apoptosis in B cells.

The authors recently reported that CD2 ligation rescues B cells from antigen-induced apoptosis by upregulation of intracellular Bcl-2 levels. However, the characterization of the early signals involved in apoptosis rescue by CD2 ligation has not been well established. In this context, CD2 does not promote either phosphatidylinositol turnover or CA2+ mobilization in B cells. In this paper the authors show that CD2 interaction with its ligand CD48 also reduces the apoptosis induced by forskolin and the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine and, to a much lesser extent, the apoptosis induced by cholera toxin in murine B splenocytes. Using a cAMP detection system sensitive to the picomolar range, the authors demonstrate that CD2-CD48 interaction decreases the intracellular cAMP concentrations induced by forskolin but not by cholera toxin. In comparison with the CD2-CD48 interaction, CD40-CD40 ligand interaction completely inhibits the apoptosis induced by cAMP increases without affecting the intracellular cAMP levels promoted by forskolin or cholera toxin. These results indicate that CD2 can also control the apoptosis at the very early steps after receptor signalling, such as the adenylate cyclase activity. Given that heterotrimeric G-proteins can mediate the adenylate cyclase activity the authors suggest that CD2 signalling could act through these small proteins, which would explain the inability of CD2 signalling to rescue from the apoptosis induced by cholera toxin, a Gs-protein activator. Conversely, CD40 seems to control apoptosis further downstream of the cAMP-PKA pathway where the survival and apoptotic signals are confluent, which might therefore render it a more efficient system to block apoptosis.

Animals↗
PubMed1995

Pertussis toxin-sensitive GTP-binding proteins regulate activation-induced apoptotic cell death of human natural killer cells.

Apoptosis of natural killer (NK) cells can be induced by non-specific physical damage (UV irradiation, heat shock) or by simultaneous ligation of the CD16 and the interleukin-2 receptor (IL-2R) molecules, but not with either anti-CD16 or IL-2 alone. Whereas blockade of GTP-binding protein (G protein)-mediated signal transduction using ADP-ribosylating bacterial toxins or the GTPase-resistant GTP analog guanosine 5'-0-(3-thiotriphosphate (GTP gamma S) does not affect non-specific induction of NK cell apoptosis, such interventions do inhibit induction of apoptosis by anti-CD16/IL-2. The G proteins involved in the regulation of activation-induced NK apoptosis are sensitive to pertussis toxin (PTX) and to the non-specific GTP analog GTP gamma S but not to cholera toxin, Pseudomonas exotoxin A or diphtheria toxin. A pertussis toxin mutant that lacks ADP-ribosylating activity, but conserves the membrane translocating and T cell-mitogenic effects of the native molecule, fails to inhibit NK apoptosis. To exert their apoptosis-inhibitory effect, PTX and GTP gamma S must be employed before cells are activated. Later addition has no effect, suggesting the implication of G proteins in the transmission of apoptosis-inducing signals, but not in the effector stage of apoptosis. Pre-incubation with PTX or GTP gamma S does not affect the activation of NK cells by CD16 cross-linking, IL-2 stimulation- or both, as assessed by the induction of CD69 expression, protein tyrosine phosphorylation and calcium mobilization. Moreover, neither PTX nor GTP gamma S compromise the effector function of NK cells or the susceptibility of target cells to NK-mediated lysis. These data suggest apoptosis as a novel mechanism by which NK responses may be controlled in vivo, as well as an experimental and therapeutical strategy to counteract endogenous down-regulation of NK responses.

Apoptosis↗
PubMed1994

Pertussis toxin interferes with superantigen-induced deletion of peripheral T cells without affecting T cell activation in vivo. Inhibition of deletion and associated programmed cell death depends on ADP-ribosyltransferase activity.

Intravenous injection of a bacterial superantigen such as Staphylococcus aureus enterotoxin B (SEB) causes transient activation and expansion of SEB-reactive V beta 8+ T cells, as well as specific down-regulation of the immune response, through partial deletion of superantigen-reactive T cells. Here we demonstrate that co-administration of pertussis toxin (PTX) and SEB reduces the SEB-induced deletion of V beta 8+ T cells, although it does not affect T cell activation and proliferation. PTX abrogates the SEB-driven deletion of V beta 8+CD4+ (not V beta 8+CD8+) splenocytes that is observed early (12-24 h) after SEB injection. Moreover, it antagonizes the late (> or = 4 days) deletion of V beta 8+CD4+ and V beta 8+CD8+ peripheral T cells that follows transient expansion of such cells. This phenomenon is associated with significant reductions in apoptosis and endonucleolysis and is not caused by a compensatory increase in proliferation of SEB-reactive T cells, as we determined by using a combined fluorometric analysis of cell cycle and DNA alterations, which are associated with programmed cell death. These effects are also observed in thymectomized animals, thus excluding the possibility that PTX might act by enhancing the maturation and export of thymic T cells to the periphery. Moreover, the SEB-induced reduction of V beta 8+ splenocytes is antagonized by PTX in vitro. The capacity of PTX to reduce clonal deletion depends critically on its ADP-ribosyltransferase activity, inasmuch as a non-enzymatic PTX mutant fails to act in this biologic system. We conclude that PTX selectively antagonizes or impedes the delivery of negative signals to T cells, which are stimulated by superantigens, without interfering with the transmission of stimulatory signals.

Animals↗
PubMed1994

Differential in vivo effects of a superantigen and an antibody targeted to the same T cell receptor. Activation-induced cell death vs passive macrophage-dependent deletion.

Superantigens have multiple pleiotropic effects in vivo, causing the activation, proliferation, and deletion of specific T cells. In our study, we analyzed the effects of the bacterial superantigen Staphylococcus aureus enterotoxin B (SEB) on peripheral T cells in vivo. As an internal control we took advantage of a IgG2a mAb, F23.1 (anti-V beta 8), that recognizes products from the same V beta gene family as that recognized by SEB. Suprisingly, not only SEB, but also F23.1 primes peripheral T cells to undergo oligonucleosomal DNA fragmentation typical for programmed cell death (PCD). Nonetheless the deletion and induction of PCD imposed by both agents obey rather different principles. First, SEB, not F23.1-induced PCD, concerns T cells that have passed through the S phase of the cell cycle, as demonstrated by experiments in which the thymidine analogue 5-bromo-2'desoxyuridine was detected in mono- and oligonucleosomal fragments of T cells undergoing PCD. Second, deletion of V beta 8+ T cells induced by SEB, not F23.1, can be blocked in vivo by high doses of retinol and, during the early phase, by glucocorticoid receptor blockade with RU-38486. Inasmuch as retinol fails to antagonize the glucocorticoid-induced PCD, at least two pathways are involved in early SEB-driven deletion, one that depends on the presence of endogenous glucocorticoid, and another that can be inhibited by retinol. Third, depletion of phagocytes in vivo by means of liposome-encapsulated dichloromethylene diphosphonate does not impede the activation and deletion of V beta 8+ cells by SEB, although it partially prevents the elimination of T cells binding F23.1 in vivo. Thus, macrophages are not rate-limiting for the action of SEB. In a further series of experiments, we demonstrate that SEB causes the secretion of a variety of cytokines (IL-1, -2, -4, -10, granulocyte-macrophage-CSF, IFN-gamma, and TNF) that may cause lethal septic shock. In contrast, F23.1 that efficiently induces all these mediators in vitro, fails to do so in vivo. In synthesis, the elimination of T cells induced by two different agents specific for V beta 8 obeys different principles: activation-induced cell death in the case of SEB and passive macrophage-mediated elimination in the case of F23.1.

Animals↗
PubMed1992

Characterization of the lymphocyte activation gene 3-encoded protein. A new ligand for human leukocyte antigen class II antigens.

The lymphocyte activation gene 3 (LAG-3), expressed in human activated T and natural killer (NK) cells, is closely related to CD4 at the gene and protein levels. We report here the initial characterization of the LAG-3-encoded protein. We have generated two monoclonal antibodies after immunization of mice with a 30-amino acid peptide that corresponds to an exposed extra loop region present in the LAG-3 immunoglobulin-like first domain. The reactivity of these reagents is directed against LAG-3 since they recognize both membrane-expressed and soluble recombinant LAG-3 molecules produced in a baculovirus expression system. The two antibodies are likely to react with the same or closely related epitope (termed LAG-3.1) exposed on the LAG-3 first domain extra loop, as assessed in competition experiments on LAG-3-expressing activated lymphocytes. Cellular distribution analysis indicated that the LAG-3.1 epitope is expressed on activated T (both CD4+ and CD8+ subsets) and NK cells, and not on activated B cells or monocytes. In immunoprecipitation experiments performed on activated T and NK cell lysates, a 70-kD protein was detected after SDS-PAGE analysis. 45-kD protein species were also immunoprecipitated. Both the 70- and 45-kD proteins were shown to be N-glycosylated. In Western blot analysis, only the former molecule was recognized by the anti-LAG-3 antibodies, demonstrating that it is LAG-3 encoded. These anti-LAG-3 antibodies were used to investigate whether the LAG-3 protein interacts with the CD4 ligands. By using a high-level expression cellular system based on COS-7 cell transfection with recombinant CDM8 vectors and a quantitative cellular adhesion assay, we demonstrate that rosette formation between LAG-3-transfected COS-7 cells and human leukocyte antigen (HLA) class II-bearing B lymphocytes is specifically dependent on LAG-3/HLA class II interaction. In contrast to CD4, LAG-3 does not bind the human immunodeficiency virus gp120. This initial characterization will guide further studies on the functions of this molecule, which may play an important role in immune responses mediated by T and NK lymphocytes.

Amino Acid Sequence↗
PubMed1990

LAG-3, a novel lymphocyte activation gene closely related to CD4.

We have identified a novel human gene of the Ig superfamily, designated LAG-3. Expression of this gene is undetectable in resting PBL, while it is found (a 2-kb message) in activated T and NK cells. The LAG-3 gene includes eight exons; the corresponding cDNA encodes a 498-amino acid membrane protein with four extracellular IgSF domains. The first one belongs to the V-SET; it is particular since it includes an extra loop in the middle of the domain and an unusual intrachain disulphide bridge. The three other domains belong to the C2-SET. Strong internal homologies are found in the LAG-3 molecule between domains 1 and 3, as well as between domains 2 and 4. It is therefore likely that LAG-3 has evolved by duplication of a pre-existing gene encoding a two IgSF-domain structure. The compared analysis of LAG-3 and CD4, with respect to both their peptidic sequence as well as their exon/intron organization, indicated that the two molecules are closely related. This point is strengthened by the finding that both genes are located on the distal part of the short arm of chromosome 12.

Amino Acid Sequence↗
PubMed1990

Cloning and expression of a lymphocyte activation gene (LAG-1).

Using subtractive hybridization of a cDNA library we have identified a human gene, termed LAG-1 (for "Lymphocyte Activation Gene-1"). This cDNA codes for a 69 amino-acid polypeptide which belongs to a new class of recently described proteins secreted by activated lymphocytes and/or monocytes. The LAG-1 gene was cloned, sequenced and its chromosomal location assigned to chromosome 17 (q21 band). The promoter region of the LAG-1 gene was shown to include a GM-CSF-like decanucleotide sequence. Using a baculovirus vector expression system, we found that a 10 kDa recombinant LAG-1 protein is secreted by AcNPV infected SF9 cells, as determined in Western blot experiments by the reactivity of specific anti-peptidic heteroantibodies. Finally the natural LAG-1 protein was precipitated from the supernatant of internally labeled activated Nk cells and shown to migrate as a single entity of 14 kDa in SDS-PAGE analysis.

Amino Acid Sequence↗
PubMed1989

The T-cell receptor V delta genes predominantly used by human peripheral gamma/delta+ T lymphocytes are not rearranged in CD3- natural killer cells.

We have analyzed, in 19 CD3- natural killer cell clones, the genomic organization of the T-cell receptor delta locus with two distinct V delta probes, V delta 1 and V delta 2. These two V delta genes code for surface proteins expressed in more than 90% of peripheral blood T-cell receptor gamma/delta+ lymphocytes, as shown by double color immunofluorescence analysis with anti-TCR delta 1, anti-BB3, and anti-delta TCS1 monoclonal antibodies. The V delta 1 and V delta 2 genes were found to be in germline position in all these clones, which are distinct phenotypically and represent a variety of the corresponding peripheral natural killer cell populations. We also studied in these cloned cell lines the transcriptional activity of the T-cell receptor delta locus with a C delta probe: short transcripts (1.7 and 0.8 kb) were found exclusively. These experiments further suggest that CD3- natural killer peripheral cells are likely to constitute a unique lineage distinct from T lymphocytes.

Antibodies, Monoclonal↗
PubMed1987

Immunoferon (AM3) enhances the activities of early-type interferon inducers and natural killer cells.

The "in vivo" effect of Immunoferon (AM3) on the production of interferon (IFN) and natural killer (NK) activity in young and old mice was studied. Although AM3 is not an IFN inducer by itself, enhancements in the serum IFN levels were produced when drug was associated to Newcastle disease virus or bacterial lipopolysaccharides as IFN inducers. This effect appeared to be dependent on the time lapsed between the inducer agent and drug. In addition, a significant stimulating effect on NK cell activity was also produced by AM3 treatments. This effect could be a consequence of a marked IFN induction and/or a modifying effect in prostaglandin synthesis.

Animals↗
PubMed1984

Immunomodulatory activity of isoprinosine on experimental viral infections in avian models.

The immunomodulatory activity of Isoprinosine treatments have been experimentally verified on chicken infected by three different viruses: Newcastle disease, fowl plague and avian infectious bronchitis. In protection tests, positive variations in the mean day of death rather than in the mortality rate were found depending on the modality of treatment. A stimulatory influence on primary anti-Newcastle disease virus antibody response was observed. In the avian model the Isoprinosine antiviral effect appears as due mainly to the enhancement of interferon production and to a synergistic interferon-isoprinosine interaction.

Adjuvants, Immunologic↗