Ioannis Vouldoukis

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.

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‐γ (IFN‐γ) and tumour necrosis factor‐α (TNF‐α) 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 β‐hexosaminidase and the preformed pool of TNF‐α within minutes. Furthermore, direct cell‐parasite contact induced TNF‐α 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.

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.

Redox‐Dependent Apoptosis in Human Endothelial Cells after Adhesion of Plasmodium falciparum‐Infected Erythrocytes

Abstract: During Plasmodium falciparum infection leading to cerebral malaria, mechanisms such as cytokine generation and cytoadherence of parasitized red blood cells (PRBC) to post‐capillary venules are clearly involved. We demonstrated that PRBC adhesion to human lung endothelial cells (HLEC) upregulated TNF‐α superfamily genes and genes related to apoptosis and inflammation. Apoptosis was confirmed by standard techniques (annexin‐V binding, genomic DNA fragmentation, and caspases activation). This apoptotic process involved the cytoplasmic pathway from a death receptor (DR‐6, Fas, TNF‐R1) through caspase 8, and the mitochondrial pathway though Bad and caspase 9 activation. Oxidative stress has been implicated in apoptosis induction in various pathological models. Superoxide anion (O2•‐) is a key molecule in the oxidative stress pathway which can form peroxynitrites (ONOO‐) in association with nitric oxide (NO•). Even though the role of NO• in malaria physiopathology is still a matter of controversy, we demonstrated that PRBC‐induced apoptosis in endothelial cells is mediated through an oxidative stress pathway. The inhibition of NO• synthesis protected the endothelial cells suggesting a deleterious role for NO•. In addition, the superoxide dismutase mimetic, MnTBAP, also protected the HLEC against PRBC‐induced apoptosis, revealing the role of O2•‐ and ONOO‐.

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.

Induction of the CD23/nitric oxide pathway in endothelial cells downregulates ICAM-1 expression and decreases cytoadherence of Plasmodium falciparum -infected erythrocytes

Cytoadherence of parasitized red blood cells (PRBCs) to postcapillary venules and cytokine production are clearly involved in the pathogenesis of cerebral malaria. Nitric oxide and TNF‐α have been proposed as major effector molecules both in protective and physiopathological processes during malaria infections. Nitric oxide production has been shown to be induced by engagement of CD23 antigen. This study aimed to investigate the potential role of the CD23/nitric oxide pathway in the control of the cytoadherence of PRBCs on human endothelial cells. We demonstrate that normal human lung endothelial cells (HLECs) are able to express the low affinity receptor for IgE (Fc∈RII/CD23), following cell incubation with interleukin 4 or PRBCs. Ligation of the CD23 antigen by a specific anti‐CD23 monoclonal antibody at the cell surface of HLECs was found to induce iNOS mRNA and protein expression, NO release and P. falciparum killing. In addition, the specific CD23‐engagement on these cells also induced a significant decrease in ICAM‐1 expression, an adhesion molecule implicated in PRBCs cytoadherence. These data not only described for the first time the expression of a CD23 antigen at the cell surface of endothelial cells but also suggest a possible new regulatory mechanisms via the CD23/NO pathway during malaria infection.

Exploitation of parasite-derived antigen in therapeutic success against canine visceral leishmaniosis

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.

Vaccination and treatment trials against murine leishmaniasis with semi-purified Leishmania antigens

Antigens with molecular weight ranges of 94-67 kDa (LiF2), 30-20 kDa (LiF5), or below 20 kDa (LiF6), isolated from lysates of Leishmania infantum promastigotes by electroelution from polyacrylamide gels were injected into mice which were genetically either partially resistant (C57BL/6) or susceptible (BALB/c) to Leishmania infection. One month after the completion of the intravenous (C57BL/6) or subcutaneous (BALB/c) schedules, the mice were challenged with 1 x 10(3) L. major promastigotes. All mice immunized with LiF2, LiF5 and LiF6 were completely resistant. Furthermore, the C57BL/6 mice immunized with LiF2 resisted a second challenge with 1 x 10(4) L. major amastigotes. 5 months later, LiF2 antigen was used for immunotherapy of L. major leishmaniasis; parasites disappeared from the treated skin lesions, although ensuing systemic infection could not be averted.

Immunization of dogs with a Leishmania infantum-derived vaccine

A partially-purified extract of Leishmania infantum has been administered to healthy dogs. Post-immunization sera were found to neutralize the infectivity of L. infantum and to abate the development of L. major. Muramyl dipeptide and one of its derivates, murabutide, were the best adjuvants.

CD23 mediates antimycobacterial activity of human macrophages.

ABSTRACT Engagement of surface receptors contributes to the antimicrobial activity of human immune cells. We show here that infection of human monocyte-derived macrophages (MDM) with live Mycobacterium avium induced the expression of CD23 on their membrane. Subsequent cross-linking of surface CD23 by appropriate ligands induced a dose-dependent antibacterial activity of MDM and the elimination of most infected cells. The stimulation of inducible nitric oxide synthase-dependent generation of NO from MDM after CD23 activation played a major role during their anti-M. avium activity. CD23 activation also induced tumor necrosis factor alpha (TNF-α) production from MDM. Mycobacteria reduction was partially inhibited by the addition of neutralizing anti-TNF-α antibody to cell cultures without affecting NO levels, which suggested the role of this cytokine for optimal antimicrobial activity. Finally, interleukin-10, a Th2 cytokine known to downregulate CD23 pathway, is shown to decrease NO generation and mycobacteria elimination by macrophages. Therefore, (i) infection with M. avium promotes functional surface CD23 expression on human macrophages and (ii) subsequent signaling of this molecule contributes to the antimicrobial activity of these cells through an NO- and TNF-α-dependent pathway. This study reveals a new human immune response mechanism to counter mycobacterial infection involving CD23 and its related ligands.