Biomedical subjects

I Vouldoukis

Publications and source records attributed to I Vouldoukis.

At least 19 recordsLinked to original sources

PubMed2001

Oestrus ovis (Diptera: Oestridae): effects of larval excretory/secretory products on nitric oxide production by murine RAW 264.7 macrophages.

Larvae of Oestrus ovis (Insecta: Diptera: Oestridae) are common parasites of nasal and sinus cavities of sheep and goats. Previous studies revealed that crude extracts of larvae modify NO synthesis by ovine monocyte derived macrophages. The aim of this study was to investigate the larval excretory/secretory products effects on nitric oxide production by murine tumour macrophages RAW 264.7. Stimulation of RAW macrophages by excretory/secretory products of the three instars larvae (25 microg/ml) significantly increased nitrite concentrations in culture supernatants compared to negative and positive Escherichia coli lipopolysaccharide control. This effect was time and dose dependent. Nitrite production in culture supernatants was due to induction of isoform NOS-2 because both NG monomethyl L-arginine (100 microM) and dexamethasone (20 microM) inhibited, by 60 and 50%, respectively, nitrite accumulation in culture supernatants. First steps of purification, by ion exchange chromatography, indicated that one protein of 29 kDa was able to induce NO synthesis by macrophages. Further studies are needed for a better characterization of these molecule and to investigate their immunogenicity for a vaccine approach.

Animals↗

PubMed2000

Ascaris lumbricoides infection is associated with protection from cerebral malaria.

Following reports of increased IgE in severe malaria and hypothesizing that helminth coinfections could modify its outcome, we conducted a retrospective case-control study to establish whether helminths affect the evolution of Plasmodium falciparum malaria. Some 182 severe cases, 315 mild controls and 40 controls with circulating schizonts were examined for intestinal helminths. Comparing cerebral malaria with mild controls, Ascaris lumbricoides was associated with a protective adjusted odds ratio (OR) of 0.58 (0. 32-1.03) P = 0.06, for coinfection with Ascaris and Necator americanus, OR = 0.39 (0.17-0.88) P = 0.02. Protection followed a dose-effect trend (P = 0.008). When comparing cerebral malaria cases and controls with circulating schizonts the OR was 0.25 (0.009-0.67) P = 0.006. We hypothesized that Ascaris infected patients may have had decreased cyto-adherence, possibly through endothelial cell receptor downregulation and/or decreased splenic clearance leading to the absence of selection of virulent P. falciparum strains. IgE-anti-IgE immune complexes resulting from helminth preinfection may have an important role in influencing clinical presentation of severe malaria, and in establishing malaria tolerance, through the CD23/NO pathway.

Acute Kidney Injury↗

PubMed2000

Fc-receptor-mediated intracellular delivery of Cu/Zn-superoxide dismutase (SOD1) protects against redox-induced apoptosis through a nitric oxide dependent mechanism.

BACKGROUND: Using specific antibodies against bovine Cu/Zn-superoxide dismutase (EC 1.15.1.1, SOD1) we demonstrated that anti-SOD antibodies (IgG1) are able to promote the intracellular translocation of the antioxidant enzyme. The transduction signalling mediated by IgG1 immune complexes are known to promote a concomitant production of superoxide and nitric oxide leading to the production of peroxynitrites and cell death by apoptosis. The Fc-mediated intracellular delivery of SOD1 thus limited the endogenous production of superoxide. It was thus of interest to confirm that in the absence of superoxide anion, the production of nitric oxide protected cells against apoptosis. Study in greater detail clearly stated that under superoxide anion-free conditions, nitric oxide promoted the cell antioxidant armature and thus protected cells against redox-induced apoptosis. MATERIALS AND METHODS: The murine macrophage cell-lines J774 A1 were preactivated or not with interferon-gamma and were then stimulated by IgG1 immune complexes (IC), free SOD1 or SOD1 IC and superoxide anion, nitric oxide, peroxynitrite, and tumor necrosis factor-alpha (TNF-alpha) production was evaluated. The redox consequences of these activation processes were also evaluated on mitochondrial respiration and apoptosis as well as on the controlled expression of the cellular antioxidant armature. RESULTS: We demonstrated that SOD1 IC induced a Fcgamma receptor (FcgammaR)-dependent intracellular delivery of the antioxidant enzyme in IFN-gamma activated murine macrophages (the J774 AI cell line). The concomitant stimulation of the FcyR and the translocation of the SOD1 in the cytoplasm of IFN-gamma-activated macrophages not only reduced the production of superoxide anion but also induced the expression of the inducible form of nitric oxide synthase (iNOS) and the related NO production. This inducing effect in the absence of superoxide anion production reduced mitochondrial damages and cell death by apoptosis and promoted the intracellular antioxidant armature. CONCLUSIONS: To define the pharmacologic mechanism of action of bovine SOD1, we attempted to identify the second messengers that are induced by SOD1 IC. In this work, we propose that Fc-mediated intracellular delivery of the SOD1 that reduced the production of superoxide anion and of peroxynitrite, promoted a NO-induced protective effect in inducing the antioxidant armature of the cells. Taken together, these data suggested that specific immune responses against antigenic SOD1 could promote the pharmacological properties of the antioxidant enzyme likely via a NO-dependent mechanism.

Animals↗

PubMed1999

The human immune response during cutaneous leishmaniasis: NO problem.

During some helminth infections, increased expression of the low-affinity receptor for IgE (CD23/FcepsilonRII) by macrophages and/or increased levels of plasma IgE have been seen, but their role in host protection or disease progression remains unclear. Recently, crosslinking of CD23 was shown to promote intracellular killing of Leishmania parasites in human macrophages, a phenomenon involving the production of tumor necrosis factor alpha and nitric oxide (NO). Based upon various in vitro and in vivo studies of human cutaneous leishmaniasis, Djavad Mossalayi, Michel Arock, Dominique Mazier, Philipe Vincendeau and Ioannis Vouldoukis here propose a model for an immune response that involves CD23-IgE-mediated NO release during protection, as well as during active cutaneous leishmaniasis.

Humans↗

PubMed1998

Nitric oxide production in human macrophagic cells phagocytizing opsonized zymosan: direct characterization by measurement of the luminol dependent chemiluminescence.

When differentiated into mature macrophages by the combination of all-trans retinoic acid and 1,25-dihydroxyvitamin D3, the human promonocytic cell lines U937 and THP-1 expressed inducible nitric oxide synthase (iNOS) transcripts. During their differentiation, the cells acquired the capacity to produce not only superoxide anion (O2.-) but also nitric oxide (.NO) in response to IgG (or IgE)-opsonized zymosan. The inhibitors of the iNOS pathway, aminoguanidine and NG-monomethyl-L-arginine (L-NMMA), suppressed the production of .NO and enhanced the steady-state concentration of O2.- determined. Conversely, superoxide dismutase (SOD) scavenged the O2.- released and increased the .NO-derived nitrite concentration detected. These data suggested a possible interaction between O2.- and .NO. In differentiated U937 (or THP-1) cells, IgG or IgE-opsonized zymosan induced a strong time-dependent luminol-dependent chemiluminescence (LDCL), which was abrogated by SOD and partially inhibited by aminoguanidine or L-NMMA. Since the iNOS inhibitors did not directly scavenge O2.-, LDCL determination in the presence or absence of SOD and/or iNOS inhibitors demonstrated a concomitant production of O2.- and .NO. These radicals induced the formation of a .NO-derived product(s), probably peroxynitrite (ONOO-), which was required to elicit maximal LDCL. Finally, LDCL measurement provided a convenient tool to characterize iNOS triggering and demonstrated an interaction between NADPH oxidase and iNOS products in human macrophagic cells phagocytizing opsonized-zymosan. These findings show that in activated macrophages, iNOS activity can be involved in LDCL and support the debated hypothesis of iNOS participation to the microbicidal activity of human macrophages.

Calcitriol↗

PubMed1997

Interleukin-10 and interleukin-4 inhibit intracellular killing of Leishmania infantum and Leishmania major by human macrophages by decreasing nitric oxide generation.

The host response to Leishmania infection is regulated by a specific pattern of local cytokine production. We investigated the effect of interleukin (IL)-10 and IL-4 on the leishmanicidal activity of human macrophages (M phi). As with L. major, intracellular killing of L. infantum by human M phi was obtained following ligation of surface CD23 or cell treatment with interferon-gamma (IFN-gamma). This leishmanicidal activity required nitric oxide (NO) generation by activated M phi, and it was partially mimicked by cell treatment with chemical NO donors. Addition of recombinant human IL-10 or IL-4 to CD23 mAb or IFN-gamma decreased L. infantum and L. major killing by infected M phi. IL-10 was more potent than IL-4 in inhibiting the leishmanicidal activity of human M phi. Inhibition of Leishmania killing by IL-4 and IL-10 correlated with decreased NO generation from M phi, and was reversed when exogenous NO was added to cell cultures. Therefore, IL-10 and IL-4 down-regulate leishmanicidal activity of human M phi, in part by inhibiting NO generation by these cells.

Animals↗

PubMed1997

Nitric oxide pathway is induced by Fc epsilon RI and up-regulated by stem cell factor in mouse mast cells.

Murine stem cell factor (SCF) induces the differentiation of mucosal mast cells (MMC) into connective tissue mast cells (CTMC) and potentiates mediator release induced by aggregation of high-affinity IgE receptors (Fc epsilon RI). In the present work, we investigated the effect of Fc epsilon RI aggregation on nitric oxide (NO) pathway induction in the different subsets of mast cells, as well as the contribution of SCF in this induction. Inducible NO synthase (iNOs) expression was not evidenced in non-stimulated MMC obtained by culture of hematopoietic progenitors in the presence of interleukin-3, whereas IgE-antigen-stimulated MMC expressed iNOs mRNA and protein and synthesized nitrites. Long-term treatment of MMC with SCF, allowing them to differentiate into CTMC, induced iNOs expression in non-stimulated cells and up-regulated iNOs expression and generation of NO derivatives induced by IgE-antigen stimulation. Thus, NO derivatives generated by mast cells could participate in inflammatory reactions during allergic stimulation.

Animals↗

PubMed1997

Evidence for direct interaction between mast cells and Leishmania parasites.

When stimulated through IgE-(or IgG-) immune complexes with parasite antigens, mast cells can release several cytokines, including IL-4, IL-6, IL-10, IL-12, Interferon-gamma (IFN-gamma) and tumour necrosis factor-alpha (TNF-alpha) that may influence the host response to Leishmania major in modulating lesion size and persistence during experimental infection in the mouse. Moreover, recent data demonstrated that mast cells are able to be antibody-independently activated by direct contact with bacteria, making them important elements in innate immunity. Given these data, we asked whether cell-parasite contact could directly induce mast cell mediator release and whether mast cells could be infected by L. major or L. infantum parasites. In this study, we showed that a pure homogeneous population of mouse bone marrow derived mast cells (BMMC) in contact with living L. major or L. infantum promastigotes, but not with attenuated parasites or soluble parasite antigens, released preformed mediators such as beta-hexosaminidase and the preformed pool of TNF-alpha within minutes. Furthermore, direct cell-parasite contact induced TNF-alpha synthesis by mast cells within hours. Moreover, we demonstrated by in vitro co-culture experiments that metacyclic L. major or L. infantum promastigotes are directly infective for a significant proportion of BMMC and are transformed into intracellular amastigotes. Taken together, these data suggest that mast cell can participate in the first line of defence, i.e. innate immunity, during local cutaneous infection with Leishmania parasites.

Animals↗

PubMed1996

Broad spectrum antibiotic activity of the skin-PYY.

Neuropeptide Y (NPY) and polypeptide YY (PYY) are two ubiquitous neuropeptides, found in brain and intestines, respectively, where they exert important regulatory functions. In this study, a new member of the YY family recently isolated from amphibian skin, skin-PYY (SPYY), is reported to inhibit irreversibly the proliferation of a broad spectrum of pathogenic microorganisms. NPY and PYY are shown to be endowed with the same activity. Their potency is similar to that of other antibacterial peptides which have been shown to exert their function by disintegrating the bacterial membrane. These findings and the fact that the C-terminal alpha-helical domain SPYY14-36, which is highly conserved among family members, was responsible for killing microorganisms and for permeation of phospholipid vesicles, suggested that the antibiotic activity may emerge via a membrane permeation mechanism. These findings also raise the question whether NPY and PYY exert in vivo a similar function in mammals.

Amino Acid Sequence↗

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

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

Antigenic components of partially purified antigens of Leishmania donovani infantum recognized by sera from dogs with asymptomatic or active visceral leishmaniasis.

The antigenic components of a semipurified fraction of Leishmania donovani infantum were determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot analysis using 14 serum samples from dogs with symptomatic visceral leishmaniasis and 11 serum samples from apparently healthy dogs collected in an area endemic for canine visceral leishmaniasis (CVL). It was found that these antigens were composed of many polypeptides, among which seven components recognized by symptomatic CVL sera, had molecular weights of approximately 18, 28, 30, 33, 63, 70, and 72 kilodaltons (kD); two components of 63 and 70 kD were recognized by three of 11 healthly dog sera. These findings suggest that specific antigens induce humoral immune response in dogs with asymptomatic or active visceral leishmaniasis. Infected dogs are not readily identifiable by their symptoms. The potential interest of the immunoblot test for CVL diagnostic purposes is discussed.

Animals↗

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

A monoclonal antibody against blood forms of Trypanosoma cruzi lyses the parasite in vitro and inhibits host cell invasion.

Monoclonal antibodies (mAbs) of the IgM isotypes were produced from mice immunized with blood forms of Trypansoma cruzi Y strain. Characterization of the epitope recognized by one of the mAbs, 164C11, as well as the effects of this mAb on complement-mediated lysis and host cell invasion are reported. Immunocytochemical analysis showed that the mAb was reactive with various strains of T. cruzi (Y, WSL, and Colombiana) as well as other trypanosomatids. The mAb 164C11 demonstrated a high complement-mediated lytic activity against bloodstream trypomastigotes, being more effective than chronic mouse serum. A protein with an apparent molecular weight of 72 kDa was detected by this mAb on all developmental stages of T. cruzi. Studies using periodate and endoglycosidase treatments suggested that the epitope is not a carbohydrate and seems to be located on the parasite membrane. In addition, preliminary results are presented, suggesting that the 72-kDa protein is involved in adhesion/or internalization of bloodstream trypomastigotes.

Animals↗

PubMed1995

In vitro study of the anti-leishmanial activity of biodegradable nanoparticles.

Leishmania are obligate intracellular parasites, responsible for leishmaniasis. Leishmaniasis are transmitted via insect vector to vertebrate hosts including humans. The infection was reproduced in vitro with promastigotes which can infect murine resident peritoneal cells. Amphotericin B was incorporated into poly(D, L-lactide-co-glycolide) nanoparticles, biodegradable drug carriers, to allow specific targeting inside the cell. The interaction of the drug with infected cells was determined by exposing macrophage cultures to drug carriers. The toxic effects of polymeric drug carriers were defined prior to exposing cells to drug-loaded nanoparticles. For contact times up to 4h, cells tolerated polymer concentrations of 0.01%. The viability of parasites after treatment was determined. Infected macrophages were incubated at 26 degrees C (which allows the transformation of amastigote to promastigote) along with loaded and unloaded nanoparticles, as well as the free drug alone, and a count of the parasites in the medium was recorded. Anti-leishmanial activity was observed with drug-free nanoparticles. This activity may arise through the release of hydrogen peroxide following the activation of macrophages. The incorporation of amphotericin B did not enhance this effect. Interestingly, trehalose, a cryoprotector of the freeze-dried nanoparticles, altered parasite growth and activated macrophages.

Amphotericin B↗

PubMed1994

Exploitation of parasite-derived antigen in therapeutic success against canine visceral leishmaniosis. Veterinary Group of Lupino.

In an attempt to obtain therapeutic success against canine visceral leishmaniosis, the potential of LiF2 antigen (Leishmania infantum-derived Fraction 2, 94-67 kDa), given alone or in combination with the chemotherapeutic agent N-methylglucamine antimonate, was compared with conventional chemotherapy with that drug. Absence of any parasite in direct microscopic examination of bone-marrow aspirates in treated dogs was considered a parasitological cure, i.e. therapeutic success. Results showed that the disappearance of clinical symptoms did not always indicate parasitological healing in dogs. The parasitological healing rates with chemotherapy and immunotherapy alone were 37.5% and 25% respectively, in contrast to the 100% cure rate observed with chemotherapy combined with immunotherapy. The development of a protective response in dogs, as measured by the in vitro leishmanicidal activity of monocyte-derived macrophages in the presence of autologous lymphocytes, was found to correlate well with the success of therapy. The overall findings of this study give an important insight into the immunotherapeutic strategy by which therapeutic success can be achieved in canine visceral leishmaniosis.

Animals↗

PubMed1994

CD23 and IgE expression during the human immune response to cutaneous leishmaniasis: possible role in monocyte activation.

Leishmania brasiliensis causes cutaneous leishmaniasis (CL) in humans. During this infection, a variety of inflammatory mediators are produced by T cells and monocytes/macrophages. In the present study, we analysed serum IgE levels and their correlation with in situ expression of the low affinity receptor for IgE (Fc epsilon RII/CD23) in patients infected with L. brasiliensis before and following therapy. These analyses were compared to in situ expression of tumour necrosis factor-alpha (TNF alpha), interleukin 3 (IL3), interferon-gamma (IFN gamma) and IL4. Disease-free individuals from the same endemic area sensitized with L. brasiliensis antigens were also included in this work. Our data indicate that during infection, serum levels of IgE and TNF alpha increased and correlated with elevated in situ expression of CD23, IL4 and TNF alpha mRNA. This expression disappeared following successful treatment, but persisted in patients resistant to anti-leishmania therapy. Patients resistant to therapy differed from other cases by a dramatic decrease in their in vivo expression of IFN gamma protein. Analysis of CD23 function in purified human monocytes indicated that this antigen mediates IgE/anti-IgE-dependent TNF alpha production. These data suggest a possible in vivo role of CD23 in acute immune responses in human CL.

Animals↗

PubMed1993

IL-4 release by human leukemic and activated normal basophils.

Recently, authors have addressed the ability of human basophils to produce IL-4. We report here the detection of significant serum IL-4 levels in a case of acute transformation of chronic myelogenous leukemia with a predominant basophilic cell population. Leukemic basophils were isolated from patients' PBMC and assayed for their IL-4-mRNA expression and their ability to secrete this cytokine in vitro. Leukemic basophilic cells (> 90% toluidine blue positive) but not other PBMC expressed IL-4-mRNA, contained IL-4 protein, and secreted this cytokine. These cells had a spontaneous IL-4 secretion ability, without a need for an exogenous activator. Meanwhile, IL-4 release was significantly increased following leukemic cell activation through Fc epsilon RI-ligation or by Ca2+ ionophore. IL-4 and its mRNA were also detected in leukemic basophils from three other chronic myelogenous leukemia patients with moderate basophilia (13, 14, and 23% basophils in PBMC). To confirm these data in normal human cells, we have developed a method to obtain large numbers of purified basophils from human bone marrow cell cultures. In contrast to leukemic basophils, normal cells required in vitro activation through Fc epsilon RI ligation or by Ca2+ ionophore to express and secrete IL-4. Leukemic and normal basophils secreted histamine following in vitro activation, but were negative for tryptase. These data thus demonstrate the in vivo and in vitro ability of human basophils to produce IL-4.

Basophils↗