Biomedical subjects

B Dugas

Publications and source records attributed to B Dugas.

At least 37 records · Page 2Linked to original sources

PubMed1997

Characterization of a constitutive type III nitric oxide synthase in human U937 monocytic cells: stimulation by soluble CD23.

The soluble cleavage fragment of the low-affinity immunoglobulin E (IgE) receptor/CD23 (sCD23 25000 MW) and antibodies directed against their receptors on monocytes, CD11b and CD11c, stimulate the production of nitric oxide (NO) by these cells and we have suggested that the enzyme involved could be related to the endothelial constitutive type III nitric oxide synthase (ecNOS). In the present work, we have analysed the characteristic properties of this NOS isoform in the model of the human promonocytic cells U937 By reverse-transcription polymerase chain reaction (RT-PCR), the presence of an mRNA coding for type III NOS was found in U937 cells and the corresponding protein was detected by immunofluorescence in permeabilized cells with a specific anti-ecNOS monoclonal antibody (mAb). Membrane extracts displayed a NOS activity dependent on the presence of calcium and calmodulin in the reaction medium and that was abrogated in the presence of EGTA. Recombinant soluble CD23 (25000 MW) was found to trigger an NO-dependent cGMP accumulation in these cells, which was abrogated by calcium chelators and inhibitors of the calcium/calmodulin complex. Moreover, sCD23 elicited a transient augmentation of intracytoplasmic free calcium concentration [Ca2+]i that was dependent on the presence of calcium in the external buffer and was prevented in the presence of EGTA, indicating that it was due to a calcium influx. In conclusion, human promonocytic cells such as U937 exhibit a functional type III NOS that can be stimulated by calcium-raising agents, such as sCD23.

Calcium↗

PubMed1996

Role of nitric oxide in the anti-tumoral effect of retinoic acid and 1,25-dihydroxyvitamin D3 on human promonocytic leukemic cells.

All trans retinoic acid and vitamin D3 derivatives are well known for their antileukemic activity, while the precise mechanism of this effect remains to be clarified. Using human leukemic U937 and THP-1 promonocytic cell lines, we analyzed the effect of all-trans retinoic acid (RA) and/or 1,25-dihydroxyvitamin D3 (VD) on the generation of nitric oxide (NO), a potent antitumoral mediator. U937 cell differentiation with VD or with both RA and VD (RA/VD) correlated with gene transcription and functional expression of inducible nitric oxide synthase (iNOS). Nitrites and L-citrulline were also detected in U937 cell supernatants as soon as 24 hours following cell incubation with VD or RA/VD, but not in cells treated with RA alone. Inhibition of iNOS activity by NG-monomethyl-L-arginine (LNMMA) significantly decreased in vitro U937 cell differentiation with VD and RA/VD as shown by the expression of cell differentiation markers (CD14 and CD68) and by the capacity of these cells to undergo a luminol-dependent chemiluminescence in response to opsonized zymosan. Similar results were obtained using the THP-1 cell line strengthening the role of NO in the VD- and RA/VD-induced growth arrest and terminal differentiation of promonocytic leukemia cells.

Antineoplastic Agents↗

PubMed1996

Triggering of CD23b antigen by anti-CD23 monoclonal antibodies induces interleukin-10 production by human macrophages.

The aim of this study was to evaluate the capacity of human macrophages to produce interleukin (IL)-10 upon stimulation of membrane CD23. An anti-CD23 monoclonal antibody (mAb) was found to elicit the expression of the specific mRNA for IL-10 in CD23-bearing macrophages, and to induce a time-dependent production of this cytokine with a maximal effect reached after 12 h. Inasmuch as we previously reported that CD23 ligation evoked the generation of nitric oxide and of cAMP, the effect of the Rp diastereoisomer of adenosine 3', 5'-cyclic phosphorothioate (Rp-cAMP, an inhibitor of the cAMP pathway) and of NG-monomethyl-L-arginine (L-NMMA, an inhibitor of the nitric oxide pathway) were evaluated on CD23-induced IL-10 production. In the presence of Rp-cAMP, the CD23-induced production of IL-10 and of the pro-inflammatory cytokine tumor necrosis factor-alpha (TNF-alpha) was totally abrogated, whereas, in the presence of L-NMMA, IL-10 production was enhanced and TNF-alpha production was suppressed. In addition, neutralization of IL-10 with an anti-IL-10 mAb increased both the magnitude and duration of CD23-driven TNF-alpha production. Such an inducing effect was observed with different anti-CD23 mAb (clone 135, MHM6 and 25), indicating that the triggering of the CD23 molecule at the surface of human macrophages induced the generation of IL-10 through a cAMP-dependent mechanism. Concomitantly this generation of IL-10 was down-regulated by nitric oxide, which was also produced after triggering of the CD23 antigen. Taken together these data indicated that human macrophages produced IL-10 after triggering of the CD23 molecule and that this production could regulate the inflammatory state of these cells.

Base Sequence↗

PubMed1996

Role of leukotriene B4 in the interleukin-4-induced human mononuclear phagocyte activation.

Interleukin-4 (IL-4) induced a time- and dose-dependent production of leukotriene B4 (LTB4) by human resting monocytes indicating that IL-4 induced the activation of the 5-lipoxygenase pathway in resting human monocytes. Maximal effect was observed in the presence of 10 ng/ml IL-4, and in kinetics experiments LTB4 production plateaued 40 min after the onset of stimulation. When stimulated for 48 hr with IL-4, resting human monocytes expressed and released the low-affinity receptor for IgE (CD23) and were partially inhibited in the presence of a highly non-redox 5-lipoxygenase inhibitor (BW B70C), suggesting that the production of LTB4 partially contributed to the IL-4-induced CD23 expression and release. This hypothesis was strengthened by the fact that exogenous LTB4 (10 nM) was found to increase the effect of a suboptimal dose of IL-4 (1 ng/ml). In addition to these phenotypical changes, IL-4 primed the phorbol-12-myristate-13-acetate (PMA)-induced luminol-dependent chemiluminescence response (LDCL) by normal human monocytes, this priming effect being abrogated in the presence of BW B70C. Taken together, these data indicated that IL-4 induced the production of LTB4 by activation of the 5-lipoxygenase pathway in human monocytes, and that the activation of this pathway could upregulate the expression and release of CD23 and the respiratory burst of these cells.

Cell Culture Techniques↗

PubMed1996

Canine visceral leishmaniasis: successful chemotherapy induces macrophage antileishmanial activity via the L-arginine nitric oxide pathway.

Following successful chemotherapy in canine visceral leishmaniasis, monocyte-derived macrophages can induce antileishmanial activity via a gamma interferon-dependent mechanism in the presence of autologous lymphocytes. The killing of leishmania correlated with the induction of the NO synthase pathway, because it correlated with the generation of nitrogen derivative production and was abrogated in the presence of NG-monomethyl-L-arginine, a competitive inhibitor of the NO synthase pathway. The level of L-citrulline in serum, which was produced after activation of the NO synthase pathway, was markedly enhanced in dogs receiving successful chemotherapy. Taken together, these data indicate that following successful chemotherapy of visceral leishmaniasis, leishmania parasites are killed by macrophages activated by gamma interferon-producing lymphocytes via an NO-dependent mechanism.

Animals↗

PubMed1996

Role of IgE immune complexes in the regulation of HIV-1 replication and increased cell death of infected U1 monocytes: involvement of CD23/Fc epsilon RII-mediated nitric oxide and cyclic AMP pathways.

BACKGROUND: IgE/anti-IgE immune complexes (IgE-IC) induce the release of multiple mediators from monocytes/macrophages and the monocytic cell line U937 following the ligation of the low-affinity Fc epsilon receptors (Fc epsilon RII/CD23). These effects are mediated through an accumulation of cAMP and the generation of L-arginine-dependent nitric oxide (NO). Since high IgE levels predict more rapid progression to acquired immunodeficiency syndrome, we attempted to define the effects of IgE-IC on human immunodeficiency virus (HIV) production in monocytes. MATERIALS AND METHODS: Two variants of HIV-1 chronically infected monocytic U1 cells were stimulated with IgE-IC and virus replication was quantified. NO and cAMP involvement was tested through the use of agonistic and antagonistic chemicals of these two pathways. RESULTS: IgE-IC induced p24 production by U1 cells with low-level constitutive expression of HIV-1 mRNAs and extracellular HIV capsid protein p24 levels (U1low), upon their pretreatment with interleukin 4 (IL-4) or IL-13. This effect was due to the crosslinking of CD23, as it was reversed by blocking the IgE binding site on CD23. The IgE-IC effect could also be mimicked by crosslinking of CD23 by a specific monoclonal antibody. p24 induction by IgE-IC was then shown to be due to CD23-mediated stimulation of cAMP, NO, and tumor necrosis factor alpha (TNF alpha) generation. In another variant of U1 cells with > 1 log higher constitutive production of p24 levels (U1high), IgE-IC addition dramatically decreased all cell functions tested and accelerated cell death. This phenomenon was reversed by blocking the nitric oxide generation. CONCLUSIONS: These data point out a regulatory role of IgE-IC on HIV-1 production in monocytic cells, through CD23-mediated stimulation of cAMP and NO pathways. IgE-IC can also stimulate increased cell death in high HIV producing cells through the NO pathway.

Antigen-Antibody Complex↗

PubMed1995

The killing of Leishmania major by human macrophages is mediated by nitric oxide induced after ligation of the Fc epsilon RII/CD23 surface antigen.

Serum IgE concentrations and the expression of the low-affinity receptor for IgE (Fc epsilon RII/CD23) are increased in cutaneous leishmaniasis or after immune challenge with Leishmania antigens. In vitro, the ligation of CD23 by IgE-anti-IgE immune complexes (IgE-IC) or by anti-CD23 monoclonal antibody (mAb) induces nitric oxide (NO) synthase and the generation of various cytokines by human monocytes/macrophages. The present study shows that IgE-IC, via CD23 binding, induce intracellular killing of Leishmania major in human monocyte-derived macrophages through the induction of the L-arginine:NO pathway. This was demonstrated by increased generation of nitrite (NO2-), the stable oxidation product of NO, and by the ability of NG-monomethyl-L-arginine to block both NO generation and parasite killing. A similar NO-dependent effect was observed with interferon gamma-treated cells. Tumor necrosis factor alpha is involved in this process, since both the induction of NO synthase and the killing of parasites caused by anti-CD23 mAb were inhibited by an anti-tumor necrosis factor alpha mAb. Treatment of noninfected CD23+ macrophages with IgE-IC provided protection against subsequent in vitro infection of these cells by Leishmania major promastigotes. Thus, IgE-IC promote killing of L. major by inducing NO synthase in human macrophages.

Amino Acid Oxidoreductases↗

PubMed1995

Interleukin-10 inhibits IgE-mediated nitric oxide synthase induction and cytokine synthesis in normal human keratinocytes.

Human keratinocytes (HK) generate nitric oxide (NO) and proinflammatory mediators following activation with either IgE/anti-IgE immune complexes or a combination of lipopolysaccharide (LPS) and interferon-gamma (IFN-gamma). Recently, interleukin-10 (IL-10) has been shown to down-regulate various inflammatory responses and to be secreted by lymphocytes and dendritic cells during skin inflammatory reactions. We show here that IL-10 down-regulates the production of tumor necrosis factor (TNF)-alpha and IL-6 by activated HK. Also, induction of inducible nitric oxide synthase (iNOS) expression in HK by IgE/anti-IgE or LPS/IFN-gamma is significantly reduced by the addition of IL-10. This effect is dose dependent and correlates with reduction of iNOS mRNA production and enzyme level. Therefore, IL-10 down-regulates NO-mediated HK inflammatory responses and may thus participate in the regulation of the skin immune network.

Antibodies, Anti-Idiotypic↗

PubMed1995

Ligation of CD23 activates soluble guanylate cyclase in human monocytes via an L-arginine-dependent mechanism.

Transduction through Fc epsilon R2/CD23 was analyzed in normal human monocytes using immunoglobulin E (IgE)-anti-IgE immune complexes (IgE ICs) and monoclonal antibodies (mAbs) to CD23. Anti-CD23 mAb and IgE IC triggered a time-dependent increase in cGMP and cAMP in interleukin-4-preincubated (CD23+) but not in unstimulated (CD23-) monocytes. Maximal cGMP and cAMP accumulations were observed 10 and 20 min, respectively, after the onset of CD23 ligation. The increase in cGMP was inhibited with N omega-monomethyl-L-arginine (L-NMMA), which also partially affected cAMP accumulation. Addition of an anti-CD23 mAb Fab fragment inhibited the IgE IC- and the anti-CD23 mAb-induced cGMP and cAMP accumulation, confirming the engagement of CD23. In addition, IgE IC and anti-CD23 mAb induced, at least in some donors, a production of nitrite that was inhibited in the presence of L-NMMA. Taken together, these findings suggest a possible involvement of the nitric oxide synthase pathway in IgE IC-mediated activation of CD23+ monocytes.

Antibodies, Monoclonal↗

PubMed1995

Evidence for a role of Fc epsilon RII/CD23 in the IL-4-induced nitric oxide production by normal human mononuclear phagocytes.

IL-4 stimulates NO production by human monocytes. After 6 days of culture with IL-4, human monocytes released detectable amounts of nitrite and L-citrulline that were inhibited in the presence of nitro-L-arginine (1 mM). Incubation with an anti-CD23 mAb Fab fragment that suppressed the biological effect of CD23 led to a strong reduction (50 to 70%) of the IL-4-induced nitrite and L-citrulline production. Ligation of membrane-associated CD23 or stimulation with recombinant soluble CD23 elicited monocytes to release nitrite and L-citrulline that was suppressed by nitro-L-arginine. Preactivation of human monocytes with IFN-gamma led to subsequent increased IL-4- and CD23-driven nitrite and L-citrulline productions that were also suppressed in the presence of either nitro-L-arginine or the anti-CD23 mAb Fab fragment. The CD23 molecule under its membrane or soluble form thus regulates NO generation by human monocytes. In addition, the IL-4-induced NO production could be mediated, at least in part, by CD23.

Antibodies, Monoclonal↗

PubMed1995

Role of cyclic nucleotides and nitric oxide in blood mononuclear cell IgE production stimulated by IL-4.

The involvement of cyclic nucleotides and of phosphodiesterase activities in IL-4-induced IgE production and release of the soluble form of the low affinity receptor for IgE (sCD23) by normal human peripheral blood mononuclear cells (PBMC) was evaluated. PBMC were stimulated by a suboptimal dose of IL-4 (10 ng/ml) cAMP inducers, adrenaline (ADR) and cholera toxin (CTx), which were found to potentiate IL-4-induced IgE production and sCD23 release after 12 days of culture. In the presence of an optimal dose of IL-4 (30 ng/ml), both ADR and CTx inhibited the production of both IgE and sCD23. In the presence of a chemical cGMP inducer, Sin-1, the production of IgE induced by 10 ng/ml IL-4 appeared to be potentiated whereas in the same experimental situation the sCD23 production was decreased. Sin-1 was found to inhibit the production of both IgE and sCD23 as effectively as cAMP inducers when an optimal dose of IL-4 was used. Since Sin-1 is a nitric oxide (NO) generating compound, we evaluated the possible involvement of the L-arginine metabolic pathway using a competitive inhibitor of L-arginine, NG-monomethyl-L-arginine (LNMMA). In the presence of 1 mM LNMMA both IgE and sCD23 production induced by either a sub-optimal or an optimal dose were partially inhibited (from 50 to 80% inhibition depending on the donor). The generation of cAMP and cGMP in the cells is controlled by cyclic nucleotide phosphodiesterases (CN-PDE), so we evaluated the effect of a CN-PDE inhibitor, isobutyl-methyl xanthine (IBMX), on the IL-4-induced IgE and sCD23 production.(ABSTRACT TRUNCATED AT 250 WORDS)

1-Methyl-3-isobutylxanthine↗

PubMed1995

Nitric oxide production by human monocytes: evidence for a role of CD23.

Nitric oxide (NO) appears to be an important and pleiotropic bioregulator of immune responses. The existence of the NO synthase (NOS) pathway in human monocytes/macrophages remains a subject of controversy, despite an increasing number of reports suggesting that human monocytes produce NO in vitro in response to various stimuli. Here, Bernard Dugas and colleagues consider the arguments supporting these conclusions, with particular emphasis on the results obtained by ligation of the low-affinity IgE receptor (Fcepsilon RIIb/CD23b).

Animals↗

PubMed1995

CD23 regulates monocyte activation through a novel interaction with the adhesion molecules CD11b-CD18 and CD11c-CD18.

CD23 is expressed on a variety of haemopoietic cells and displays pleiotropic activities in vitro. We report that in addition to CD21 and IgE, CD23 interacts specifically with the CD11b and CD11c, the alpha chains of the beta 2 integrin adhesion molecule complexes CD11b-CD18 and CD11c-CD18, on monocytes. Full-length recombinant CD23 incorporated into fluorescent liposomes was shown to bind to COS cells transfected with cDNA encoding either CD11b-CD18 or CD11c-CD18 but not with CD11a-CD18. The interaction was specifically inhibited by anti-CD11b or anti-CD11c, respectively, and by anti-CD23 MAbs. The functional significance of this ligand pairing was demonstrated by triggering CD11b and CD11c on monocytes with either recombinant CD23 or anti-CD11b and anti-CD11c MAbs to cause a marked increase in nitrite-oxidative products and pro-inflammatory cytokines (IL-1 beta, IL-6, and TNF alpha). These CD23-mediated activities were decreased by Fab fragments of MAbs to CD11b, CD11c, and CD23. These results demonstrate that CD11b and CD11c are receptors for CD23 and that this novel ligand pairing regulates important activities of monocytes.

CD11 Antigens↗

PubMed1995

Regulation of IgE production from human mononuclear cells by beta 2-adrenoceptor agonists.

The present study examined the effect of beta 2-adrenoceptor agonists on the interleukin-4 (IL-4)-driven IgE production and on the possible mechanisms of action of these compounds. We present evidence that salbutamol and fenoterol potentiated the IL-4-induced IgE production by human peripheral blood mononuclear cells (PBMC). No significant effect of incubation in the presence of beta 2-adrenoceptor agonists on IgG, IgA and IgM production was observed. Salbutamol and fenoterol inhibited interferon-(IFN)-gamma production by PHA-activated human PBMC suggesting that the blockade of the production of this cytokine could possibly explain the enhancement of IgE production. Salbutamol and fenoterol potentiated the IL-4-induced production of sCD23 whereas no effect on CD23 expression was observed. The potentiating effect of salbutamol on IgE production was blocked by two antagonists of beta 2-adrenoceptor, namely butoxamine and D,L-propranolol, suggesting a beta-adrenoceptor-mediated event. These results demonstrate that beta 2-adrenoceptor stimulation results in an increase in IgE production by human B lymphocytes.

Adrenergic beta-Agonists↗

PubMed1995

Impairment of circulating lactoferrin in HIV-1 infection.

Levels of plasma lactoferrin are decreased in HIV-1-infected patients in relation to the progression of the disease. Plasma lactoferrin concentrations were determined using a specific and sensitive enzyme immunoassay. 97 plasma were studied (22 asymptomatic, 45 symptomatic patients compared to 30 healthy controls) and the results showed a highly significant decrease (p < 0.001) of the level of lactoferrin in HIV-1-infected patients (respectively 2.79 +/- 1.2 and 0.68 +/- 0.22 micrograms/ml) compared to controls (4.37 +/- 0.83 micrograms/ml). Since it is well established that plasma lactoferrin level could be influenced by the number of neutrophils, the experiments were reproduced in neutropenic patients who represent 10% of recruitment (6 among 45 symptomatic patients). The plasma from neutropenic symptomatic patients (neutrophils < or = 1,300/mm3) showed their mean lactoferrin level at 0.36 micrograms/ml still far above the normal values. In view of the different reported biological effects of lactoferrin that are of great importance in the non-specific defences, the real biological place of the lack of such a molecule could be one important component of the multifactorial nature of HIV-1 infection.

AIDS-Related Complex↗

PubMed1994

Growth arrest and terminal differentiation of leukemic myelomonocytic cells induced through ligation of surface CD23 antigen.

Acute myelogenous leukemia (AML) cells express CD23 surface antigen after in vitro treatment with various cytokines, including interleukin-4 (IL-4) and interferon gamma. Subsequent ligation of CD23 by specific monoclonal antibody (MoAb) induces substantial morphologic and functional modifications in these cells. In the present study, we investigated the role of CD23 in the proliferation and the maturation of leukemic cells from AML patients or the U937 cell line. CD23+ cell treatment with CD23 MoAb inhibited the proliferation of leukemic cells. This correlated with their terminal differentiation after 7 to 9 days incubation because they (1) definitively lost their growth capacity; (2) adhered to culture flasks and became monocyte/macrophage-like; and (3) expressed mature monocyte markers including nonspecific esterases. Intracellular mechanism of this antitumoral effect was then analyzed in U937 cells. Induction of high-density surface CD23 expression by IL-4 or granulocyte-macrophage colony-stimulating factor coincided with a transient decrease of U937 cell proliferation. CD23 ligation during this low-proliferative phase induced a rapid activation of L-arginine-dependent pathway and the intracellular accumulation of cyclic guanosine monophosphate and cyclic adenosine monophosphate (cAMP). Induction of these early messengers was followed by the activation of nuclear factor-kB transcription factor and the modulation of proto-oncogene expression by U937 cells. Whereas U937 cell treatment with IL-4 decreased c-fos/c-jun expression, CD23 MoAb reinduced c-fos/c-jun and promoted the expression of cell maturation-associated proto-oncogenes junB and c-fms, during the first 24 hours. Both IL-4 and CD23 MoAb downregulated the expression of c-myb. CD23 ligation also induced the production of TNF alpha by U937 cells. Inhibitors of cAMP and nitric oxide reversed CD23-mediated modification in U937 cells. These data evidence the ability of CD23 surface antigen to mediate terminal differentiation of early leukemic myelomonocytic cells.

Antibodies, Monoclonal↗

PubMed1994

Involvement of Fc epsilon RII/CD23 and L-arginine dependent pathway in IgE-mediated activation of human eosinophils.

Eosinophils display various receptors for immunoglobulin E (IgE) including the high affinity receptor for IgE (Fc epsilon RI), CD23 (Fc epsilon RII), and Mac-2/epsilon BP. We attempted here to clarify the role of these receptors in IgE-mediated activation of eosinophils from normal human bone marrow cultures. Pretreatment of eosinophils with IL-4 is required for IgE/anti-IgE-mediated stimulation of TNF-alpha and peroxydes production. TNF-alpha release from eosinophils was also induced following ligation of CD23 and to a lesser extent with anti-Mac-2, while Fc epsilon RI-ligation had no effect. IgE/anti-IgE effect dramatically decreased when eosinophils were pretreated with Fab fragments of CD23-mAb. In addition, this effect could also be reversed by inhibiting CD23-dependent nitric oxide pathway by NG-monomethyl-L-arginine. Nitric oxide chemical donor, SIN-1, induced TNF-alpha release from eosinophils. CD23 and nitric oxide pathway are thus involved in IgE-mediated stimulation of normodense human eosinophils.

Antibodies, Monoclonal↗

PubMed1994

Interleukin-4 stimulates cGMP production by IFN-gamma-activated human monocytes. Involvement of the nitric oxide synthase pathway.

Resting human blood monocytes from some donors were found to produce a small amount of 3'-5' guanine cyclic monophosphate (cGMP) in response to interleukin 4 (IL-4). A much higher response was observed when monocytes were preincubated with interferon (IFN-gamma), which alone was ineffective. Preincubation of monocytes with IL-4 led, in contrast, to their subsequent incapacity to generate cGMP in response to IL-4. The accumulation of cGMP induced by IL-4 in IFN-gamma preincubated monocytes was dose-dependent and peaked about 15 min after its addition. It was inhibited in the presence of NG-mono-methyl-L-arginine (L-NMMA), an inhibitor of the nitric oxide synthase pathway. This suppressive effect of L-NMMA was reverted by an excess of L- but not of D-arginine. Accumulation of cGMP was significantly reduced by addition of soluble guanylyl cyclase inhibitors, such as LY83583 [correction of LY83853] and methylene blue, but was not impaired in the presence of EGTA, suggesting that the pathway involved is calcium independent. In addition, IL-4 induced an increased secretion of nitrite by monocytes, that was potentiated by IFN-gamma and inhibited by L-NMMA. Taken together, these results suggest that the sequential exposure of monocytes to IFN-gamma and IL-4 elicits the release of NO from L-arginine, which in turn is capable to stimulate soluble guanylyl cyclase.

Amino Acid Oxidoreductases↗