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Papers by Eric Muraille

European Journal of Immunology, 2010

Experimental animal and human studies have demonstrated that probiotic strains have beneficial ef... more Experimental animal and human studies have demonstrated that probiotic strains have beneficial effects on allergy. Here we report that the probiotic Escherichia coli Nissle 1917 strain (EcN) is able to activate DC, as shown by important cytokine synthesis together with up-regulation of membrane expression of CD40, CD80 and CD86. This EcN-induced DC activation was strictly dependent on the TLR4 signaling pathway and was also associated with stimulation of NF-κB and MAPK. We next investigated the prophylactic potential of i.n. co-administration of EcN with a recombinant form of Der p 1 (ProDer p 1) in a murine model of mite allergy. I.n. vaccinations with EcN plus ProDer p 1 prevented the subsequent allergic response following Der p 1 sensitization and airway challenge with aerosolized mite extracts through the induction of an allergen-specific IgG2a response, the prevention of specific IgE production and a strong reduction of IL-5 secretion by allergen-restimulated splenocytes. EcN alone or in combination with ProDer p 1 inhibited the development of airway eosinophilia and neutrophilia. This in vivo protective effect of EcN was, in part, mediated by TLR4 signaling. Our results suggest that EcN represents an efficient adjuvant to prevent allergic responses.

Biochemical Journal, 1999

The termination of activation signals is a critical step in the control of the immune response ; ... more The termination of activation signals is a critical step in the control of the immune response ; perturbation of inhibitory feedback pathways results in profound immune defects culminating in autoimmunity and overwhelming inflammation. FcγRIIB receptor is a well described inhibitory receptor. The ligation of B-cell receptor (BCR) and FcγRIIB leads to the inhibition of B-cell activation. Numerous studies have demonstrated that the SH2-domain-containing inositol 5-phosphatase SHIP (referred hereto as SHIP-1) is essential in this process. The cDNA encoding a second SH2-domain-containing inositol 5phosphatase, SHIP-2, has been cloned Biochem. Biophys. Res. Commun. 239, 697-700]. Here we report the distribution of SHIP-2 in

Neuroscience, 2001

AbstractöThe germinative ventricular zone of embryonic brain contains neural lineage progenitor c... more AbstractöThe germinative ventricular zone of embryonic brain contains neural lineage progenitor cells that give rise to neurons, astrocytes and oligodendrocytes. The ability to generate neurons persists at adulthood in restricted brain areas. During development, many growth factors exert their e¡ects by interacting with tyrosine kinase receptors and activate the phosphatidylinositol 3-kinase and the Ras/MAP kinase pathways. By its ability to modulate these pathways, the recently identi¢ed Src homology 2 domain-containing inositol polyphosphate 5-phosphatase 2, SHIP2, has the potential to regulate neuronal development. Using in situ hybridization technique with multiple synthetic oligonucleotides, we demonstrated that SHIP2 mRNA was highly expressed in the ventricular zone at early embryonic stages and subventricular zones at latter stages of brain and spinal cord and in the sympathetic chain. No signi¢cant expression was seen in di¡erentiated ¢elds. This restricted expression was maintained from embryonic day 11.5 to birth. In the periphery, large expression was detected in muscle and kidney and moderate expression in thyroid, pituitary gland, digestive system and bone. In the adult brain, SHIP2 was mainly restricted in structures containing neural stem cells such as the anterior subventricular zone, the rostral migratory stream and the olfactory tubercle. SHIP2 was also detected in the choroid plexuses and the granular layer of the cerebellum. The speci¢city of SHIP2 expression in neural stem cells was further demonstrated by (i) the dramatic increase in SHIP2 mRNA signal in neural cell adhesion molecule (N-CAM)-de¢cient mice, which present an accumulation of progenitor cells in the anterior subventricular zone and the rostral migratory stream, (ii) the abundant expression of 160-kDa SHIP2 by western blotting in proliferating neurospheres in culture and its downregulation in non-proliferating di¡erentiated neurospheres.

Immunology Letters, 2000

FcgRIIB are single-chain low-affinity receptors for IgG that bear an immunoreceptor tyrosine-base... more FcgRIIB are single-chain low-affinity receptors for IgG that bear an immunoreceptor tyrosine-based inhibition motif (ITIM) in their intracytoplasmic domain and that negatively regulate immunoreceptor tyrosine-based activation motif (ITAM)-dependent cell activation. In B cells, coaggregation of the B cell receptor (BCR) and FcgRIIB leads to an inhibition of B cell activation. Inhibitory properties of FcgRIIB have been related to the recruitment of SHIP, an SH2 domain-containing inositol 5-phosphatase (referred to as SHIP1), via ITIM phosphorylated FcgRIIB. Here, we demonstrate that the second SH2 domain-containing inositol 5-phosphatase SHIP2 could also bind to the FcgRIIB ITIM. As a model, a FcgRIIB deficient B cell line (IIA1.6), transfected with a cDNA encoding either w.t. FcgRIIB1% or FcgRIIB1% whose ITIM tyrosine was mutated has been used. SHIP2 tyrosine phosphorylation and association to the adaptator protein Shc were only found in transfectants expressing w.t. FcgRIIB1%. SHIP2 was also found to bind to a phosphopeptide corresponding to the ITIM sequence of FcgRIIB. There was no binding to the nonphosphorylated peptide. Finally, both SHIP2 and SHIP1 were coprecipitated with FcgRIIB1% upon coaggregation with BCR in IIA1.6 transfectants.

Infection and Immunity, 2002

The role of interleukin-12 (IL-12) has been clearly established in the resistance of C57BL/6 (B6)... more The role of interleukin-12 (IL-12) has been clearly established in the resistance of C57BL/6 (B6) mice to Leishmania major infection, but its involvement in the control of Leishmania mexicana infection remains to be determined. Here, we show the following. (i) L. mexicana, in contrast to L. major, induces the development of nonhealing lesions in B6 mice. (ii) Cells expressing IL-12p40, gamma interferon (IFN-␥), NOS2, and CD40L are numerous in the footpad lesion and/or the draining popliteal lymph node of animals infected for up to 14 weeks with L. mexicana. (iii) B6 mice, either IL-12p40 deficient or treated with IL-12p40-neutralizing antibodies, display a dramatic enhancement of primary and secondary lesions leading to death 10 weeks after inoculation with L. mexicana. (iv) Splenocytes harvested 4 and 8 weeks after infection of IL-12p40 ؊/؊ B6 mice with L. mexicana are unable to produce IFN-␥, but secrete IL-4, IL-10, and IL-18. Thus, the early control of L. mexicana infection by B6 mice is independent of IL-12, whereas IL-12 and Th1 responses play a key role in controlling the late stages of L. mexicana infection. However, they fail to resolve lesions, in contrast to L. major infection, emphasizing the different outcomes induced by these two Leishmania species in B6 mice.

PLOS Pathogens, 2012

Brucella are facultative intracellular bacteria that chronically infect humans and animals causin... more Brucella are facultative intracellular bacteria that chronically infect humans and animals causing brucellosis. Brucella are able to invade and replicate in a broad range of cell lines in vitro, however the cells supporting bacterial growth in vivo are largely unknown. In order to identify these, we used a Brucella melitensis strain stably expressing mCherry fluorescent protein to determine the phenotype of infected cells in spleen and liver, two major sites of B. melitensis growth in mice. In both tissues, the majority of primary infected cells expressed the F4/80 myeloid marker. The peak of infection correlated with granuloma development. These structures were mainly composed of CD11b + F4/80 + MHC-II + cells expressing iNOS/NOS2 enzyme. A fraction of these cells also expressed CD11c marker and appeared similar to inflammatory dendritic cells (DCs). Analysis of genetically deficient mice revealed that differentiation of iNOS + inflammatory DC, granuloma formation and control of bacterial growth were deeply affected by the absence of MyD88, IL-12p35 and IFN-c molecules. During chronic phase of infection in susceptible mice, we identified a particular subset of DC expressing both CD11c and CD205, serving as a reservoir for the bacteria. Taken together, our results describe the cellular nature of immune effectors involved during Brucella infection and reveal a previously unappreciated role for DC subsets, both as effectors and reservoir cells, in the pathogenesis of brucellosis.

Allergy, 2009

Background: Selected lactic acid bacteria were reported to prevent atopic dermatitis and experim... more Background: Selected lactic acid bacteria were reported to prevent atopic dermatitis and experimental asthma but the mechanisms of their immunomodulatory effects are not fully elucidated. In this study, the signaling pathways triggered by Lactobacillus plantarum NCIMB8826 were investigated and the potential use of this strain producing a variant of the mite allergen Der p 1 as live vaccine vehicle was evaluated.Methods: Mouse bone marrow-derived dendritic cells were stimulated with wild-type or a L. plantarum teichoic acid mutant to evaluate the secretion of cytokines. A recombinant L. plantarum expressing Der p 1 was engineered, its in vitro immunomodulatory properties were characterized and its prophylactic potential was evaluated in a Der p 1-sensitization murine model.Results: Mouse dendritic cells stimulated by L. plantarum triggered the release of interleukin-10 (IL-10), IL-12 p40, IL-12 p70 and tumor necrosis factor-alpha (TNF-α). IL-12 p40 secretion was dependent on nuclear factor-κB (NF-κB), mitogen-activated protein (MAP) kinases, Toll-like receptor 2 (TLR2), TLR9 and on the bacterial teichoic acid composition. Recombinant L. plantarum producing Der p 1 exhibited similar immunostimulatory properties as wild-type. Prophylactic intranasal pretreatment of mice with this recombinant strain prevented the development of the typical Th2-biased allergic response by a drastic reduction of specific IgE and the induction of protective allergen-specific IgG2a antibodies. Moreover, both wild-type or recombinant L. plantarum reduced airway eosinophilia following aerosolized allergen exposure and IL-5 secretion upon allergen restimulation.Conclusion: By combining both Th1-type immunostimulatory properties and an efficient allergen delivery capacity, recombinant L. plantarum producing Der p 1 represents a promising vaccine against house dust mite allergy.

Cellular Immunology, 1995

Immunology, 1996

The aim of this study was to investigate the mechanisms by which B7-related costimulatory molecul... more The aim of this study was to investigate the mechanisms by which B7-related costimulatory molecules (CD80, CD86) increase T-cell responsiveness to extracellular ligands. As a model study, the in vitro response of purified splenic CD4+ T cells to a bacterial superantigen, SEB, was characterized. Previous analysis of this experimental model led us to conclude that expression of B7-related molecules is strictly required in order to activate CD4+ T cells in the presence of bacterial superantigens. In the present report, we demonstrate that antigen-presenting cell-derived costimulatory signals regulate the kinetics of interleukin-2 (IL-2) production by SEB-activated splenic CD4+ T cells. Indeed, experiments performed with purified subpopulations of antigenpresenting cells and using B7-transfected cell lines indicated that increased levels of CD80 and/or CD86 cell surface expression is associated with a faster kinetics of IL-2 production in response to SEB. Accordingly, blocking of CD80 or CD86-derived signals by specific monoclonal antibodies led to a slower kinetics of IL-2 production in response to SEB. Thus these data demonstrate that similar strength of signal through the T-cell receptor can lead to immune responses displaying distinct kinetics depending on the level of costimulatory ligands on APC.

European Journal of Immunology, 2008

DiC14-amidine cationic liposomes were recently shown to promote Th1 responses when mixed with all... more DiC14-amidine cationic liposomes were recently shown to promote Th1 responses when mixed with allergen. To further define the mode of action of diC14-amidine as potential vaccine adjuvant, we characterized its effects on mouse and human myeloid dendritic cells (DC). First, we observed that, as compared with two other cationic liposomes, only diC14-amidine liposomes induced the production of IL-12p40 and TNF-α by mouse bone marrow-derived DC. DiC14-amidine liposomes also activated human DC, as shown by synthesis of IL-12p40 and TNF-α, accumulation of IL-6, IFN-β and CXCL10 mRNA, and up-regulation of membrane expression of CD80 and CD86. DC stimulation by diC14-amidine liposomes was associated with activation of NF-κB, ERK1/2, JNK and p38 MAP kinases. Finally, we demonstrated in mouse and human cells that diC14-amidine liposomes use Toll-like receptor 4 to elicit both MyD88-dependent and Toll/IL-1R-containing adaptor inducing interferon IFN-β (TRIF)-dependent responses.Supporting Information for this article is available at www.wiley-vch.de/contents/jc_2040/2008/37998_s.pdf

Scandinavian Journal of Immunology, 2007

The identification of subsets of CD4+ helper cells producing distinct pattern of cytokines has pr... more The identification of subsets of CD4+ helper cells producing distinct pattern of cytokines has provided a valuable framework for understanding how different effector populations of immune cells can be recruited in vivo during infection. In the view of most investigators, Th1 and Th2 cells produce factors that serve as their own autocrine factors and cytokines exerting suppressive activities on each other's development and activity. This concept intuitively explains the natural tendency of immune responses to become progressively polarized. However, several experimental observations appear difficult to rationalize with a simple, ‘symmetrical’ Th1/Th2 paradigm including those that Th1 cells do not produce their own growth factor; that both Th1 and Th2 cells can promote inflammatory responses; that interleukin-10 (IL-10) inhibits inflammatory responses in a Th1/Th2-independent fashion; that IL-10 promotes the development of Th1-type effector cells; and that IL-12 can amplify pre-established Th2 responses. The purpose of the present analysis is to provide a revised model for better understanding how cytokines regulate immune responses in vivo.

PLOS Pathogens, 2009

Leishmania major parasites reside and multiply in late endosomal compartments of host phagocytic ... more Leishmania major parasites reside and multiply in late endosomal compartments of host phagocytic cells. Immune control of Leishmania growth absolutely requires expression of inducible Nitric Oxide Synthase (iNOS/NOS2) and subsequent production of NO. Here, we show that CD11b + CD11c + Ly-6C + MHC-II + cells are the main iNOS-producing cells in the footpad lesion and in the draining lymph node of Leishmania major-infected C57BL/6 mice. These cells are phenotypically similar to iNOS-producing inflammatory DC (iNOS-DC) observed in the mouse models of Listeria monocytogenes and Brucella melitensis infection. The use of DsRed-expressing parasites demonstrated that these iNOS-producing cells are the major infected population in the lesions and the draining lymph nodes. Analysis of various genetically deficient mouse strains revealed the requirement of CCR2 expression for the recruitment of iNOS-DC in the draining lymph nodes, whereas their activation is strongly dependent on CD40, IL-12, IFN-c and MyD88 molecules with a partial contribution of TNF-a and TLR9. In contrast, STAT-6 deficiency enhanced iNOS-DC recruitment and activation in susceptible BALB/c mice, demonstrating a key role for IL-4 and IL-13 as negative regulators. Taken together, our results suggest that iNOS-DC represent a major class of Th1-regulated effector cell population and constitute the most frequent infected cell type during chronic Leishmania major infection phase of C57BL/6 resistant mice.

European Journal of Immunology, 1995

Exogenous glucocorticoid hormones are widely used as therapeutical agents, whereas endogenous glu... more Exogenous glucocorticoid hormones are widely used as therapeutical agents, whereas endogenous glucocorticoids may act as physiological immunosuppressants involved in the control of immune and inflammatory responses. The optimal activation of T lymphocytes requires two distinct signals: the major histocompatibility complex-restricted presentation of the antigen and an additional co-stimulatory signal provided by the antigen-presenting cells. There is ample evidence that, among the cells able to present the antigen, the dendritic cells (DC) have the unique property to activate antigen-specific, naive T cells in vitro and in vivo, and are therefore required for the induction of primary immune responses. In this work, we tested whether glucocorticoids affected the capacity of DC to sensitize naive T cells. Our data show that, in vitro, the steroid hormone analog dexamethasone (Dex) affects the viability of DC, selectively downregulates the expression of co-stimulatory molecules on viable DC, and strongly reduces their immunostimulatory properties. In vivo, a single injection of Dex results in impaired antigen presenting function, a finding which correlates with reduced numbers of splenic DC. These results show that glucocorticoids regulate DC maturation and immune function in vitro and in vivo and suggest that this mechanism may play a role in preventing overstimulation of the immune system.

Infection and Immunity, 2003

Cells of the dendritic cell (DC) lineage, by their unique ability to stimulate naive T cells, may... more Cells of the dendritic cell (DC) lineage, by their unique ability to stimulate naive T cells, may be of crucial importance in the development of protective immune responses to Leishmania parasites. The aim of this study was to compare the impact of L. major infection on DCs in BALB/c (susceptible, developing Th2 responses), C57BL/6 (resistant, developing Th1 responses), and tumor necrosis factor (TNF) ؊/؊ C57BL/6 mice (susceptible, developing delayed and reduced Th1 responses). We analyzed by immunohistochemistry the phenotype of infected cells in vivo. Granulocytes (GR1 ؉ ) and macrophages (CD11b ؉ ) appear as the mainly infected cells in primary lesions. In contrast, cells expressing CD11c, a DC specific marker, are the most frequently infected cells in draining lymph nodes of all mice tested. These infected CD11c ؉ cells harbored a particular morphology and cell surface phenotype in infected C57BL/6 and BALB/c mice. CD11c ؉ infected cells from C57BL/6 and TNF ؊/؊ C57BL/6 mice displayed a weak parasitic load and a dendritic morphology and frequently expressed CD11b or F4/80 myeloid differentiation markers. In contrast, some CD11c ؉ infected cells from BALB/c mice were multinucleated giant cells. Giant cells presented a dramatic accumulation of parasites and differentiation markers were not detectable at their surface. In all mice, lymph node CD11c ؉ infected cells expressed a low major histocompatibility complex II level and no detectable CD86 expression. Our results suggest that infected CD11c ؉ DC-like cells might constitute a reservoir of parasites in lymph nodes.

European Journal of Immunology, 1995

Excessive T cell activation induced by bacterial superantigens plays an important role in the pat... more Excessive T cell activation induced by bacterial superantigens plays an important role in the pathology associated with Gram-positive bacteremia. To gain insight into the early phases of T cell activation by bacterial enterotoxins in vivo, we investigated the ability of antibodies to well-defined co-stimulatory molecules to inhibit T cell activation and the subsequent toxic shock syndrome induced in BALB/c mice following the injection of staphylococcal enterotoxin B (SEB). We demonstrate here that a single dose of anti-B7.2 antibodies, but not anti-B7.1 antibodies, significantly inhibits T cell activation, as judged by lower systemic IL-2 release, blastogenesis and IL-2 receptor expression, and reduces the lethal effect of SEB in D-galactosamine-sensitized mice. These results demonstrate that co-stimulation through the B7.2 molecule plays an important role in the activation of T cells in response to SEB in vivo and suggest alternative therapies for septic shock caused by bacterial enterotoxins based on blocking antibodies to co-stimulatory molecules.

Immunology Letters, 2008

Dendritic cells (DCs) have been shown to express the tryptophan catabolizing enzyme indoleamine 2... more Dendritic cells (DCs) have been shown to express the tryptophan catabolizing enzyme indoleamine 2,3dioxygenase (IDO-1), a protein presently thought to exert dual and possibly contrasting effects on the immune response. Depletion of tryptophan and release of tryptophan catabolites have been shown to exert a tolerogenic influence on T cell responses, while the IDO enzymatic activity has been recently suggested to promote DC maturation. In this report, we have explored the putative role of IDO-1 in regulating DC biology by analyzing DC development and function from IDO-1 deficient mice. In keeping with previous observations, lack of IDO-1 expression was found to affect in vitro DC generation from bone mouse precursors cultured in the presence of GM-CSF. However, change in growth factor (Flt3L) and/or culture conditions (low-adherence vessels) abolished the difference observed between wt (wild type) and IDO-1-deficient, in vitro generated DCs. Moreover, IDO-1-deficient mice displayed a normal DC compartment in vivo, suggesting that IDO-1 does not play a significant role in DC development and function in vivo. Collectively, these observations suggest that despite a possible role for IDO-1 expression in regulating DC differentiation in vitro under commonly used culture conditions, IDO-1 is largely dispensable for DC development and function in vivo.

Cellular Immunology, 1999

Based on the observation that pathogen-derived lectins play an important role in cell adhesion an... more Based on the observation that pathogen-derived lectins play an important role in cell adhesion and invasion, we examined the possible role of host carbohydrate-bearing molecules in inducing the secretion of IL-12, a crucial proinflammatory cytokine. The ability of 12 plant lectins to recognize and stimulate naive murine mononuclear cells in vitro has been characterized in this study. Mitogenic lectins (comprising Con A, PHA, PSA, and LCA) were found to induce the secretion of multiple cytokines in vitro, including IL-2, interferon (IFN)-␥, and IL-12. Of interest, WGA, a nonmitogenic lectin unable to promote IL-2 secretion, was found to induce IL-12 and IFN-␥ production in a T and B cell-independent fashion. The functional properties of WGA were inhibited by N-acetylneuraminic acid and N,N-diacetylchitobiose. WGA therefore represents a potentially useful tool for the study of membrane glycoproteins involved in the early proinflammatory response characteristic of innate immunity.

Dendritic cells (DC) are described as "nature's adjuvant," since they have the capacity to sensit... more Dendritic cells (DC) are described as "nature's adjuvant," since they have the capacity to sensitize T cells in vivo upon first encounter with the antigen. The potent accessory properties of DC appear to develop sequentially. In particular, the ability to process antigens and to sensitize naive T cells develops in sequence, a process termed "maturation" that is well described in vitro. Here, we obtain evidence for maturation in vivo in response to the bacterial product lipopolysaccharide (LPS). Before LPS treatment, many DC are found at the margin between the red and white pulp. These cells lack the M342 and DEC-205 markers, but process soluble proteins effectively. 6 h after LPS, DC with the M342 and DEC-205 markers are found in increased numbers in the T cell areas. These cells have a reduced capacity to process proteins, but show increases in the B7 costimulator and T cell stimulatory capacity. 48 h after LPS, the number of DC in the spleen is reduced markedly. We interpret these findings to mean that LPS can cause DC in the marginal zone to mature and to migrate into and then out of the T cell areas.

European Journal of Immunology, 2010

Experimental animal and human studies have demonstrated that probiotic strains have beneficial ef... more Experimental animal and human studies have demonstrated that probiotic strains have beneficial effects on allergy. Here we report that the probiotic Escherichia coli Nissle 1917 strain (EcN) is able to activate DC, as shown by important cytokine synthesis together with up-regulation of membrane expression of CD40, CD80 and CD86. This EcN-induced DC activation was strictly dependent on the TLR4 signaling pathway and was also associated with stimulation of NF-κB and MAPK. We next investigated the prophylactic potential of i.n. co-administration of EcN with a recombinant form of Der p 1 (ProDer p 1) in a murine model of mite allergy. I.n. vaccinations with EcN plus ProDer p 1 prevented the subsequent allergic response following Der p 1 sensitization and airway challenge with aerosolized mite extracts through the induction of an allergen-specific IgG2a response, the prevention of specific IgE production and a strong reduction of IL-5 secretion by allergen-restimulated splenocytes. EcN alone or in combination with ProDer p 1 inhibited the development of airway eosinophilia and neutrophilia. This in vivo protective effect of EcN was, in part, mediated by TLR4 signaling. Our results suggest that EcN represents an efficient adjuvant to prevent allergic responses.

Biochemical Journal, 1999

The termination of activation signals is a critical step in the control of the immune response ; ... more The termination of activation signals is a critical step in the control of the immune response ; perturbation of inhibitory feedback pathways results in profound immune defects culminating in autoimmunity and overwhelming inflammation. FcγRIIB receptor is a well described inhibitory receptor. The ligation of B-cell receptor (BCR) and FcγRIIB leads to the inhibition of B-cell activation. Numerous studies have demonstrated that the SH2-domain-containing inositol 5-phosphatase SHIP (referred hereto as SHIP-1) is essential in this process. The cDNA encoding a second SH2-domain-containing inositol 5phosphatase, SHIP-2, has been cloned Biochem. Biophys. Res. Commun. 239, 697-700]. Here we report the distribution of SHIP-2 in

Neuroscience, 2001

AbstractöThe germinative ventricular zone of embryonic brain contains neural lineage progenitor c... more AbstractöThe germinative ventricular zone of embryonic brain contains neural lineage progenitor cells that give rise to neurons, astrocytes and oligodendrocytes. The ability to generate neurons persists at adulthood in restricted brain areas. During development, many growth factors exert their e¡ects by interacting with tyrosine kinase receptors and activate the phosphatidylinositol 3-kinase and the Ras/MAP kinase pathways. By its ability to modulate these pathways, the recently identi¢ed Src homology 2 domain-containing inositol polyphosphate 5-phosphatase 2, SHIP2, has the potential to regulate neuronal development. Using in situ hybridization technique with multiple synthetic oligonucleotides, we demonstrated that SHIP2 mRNA was highly expressed in the ventricular zone at early embryonic stages and subventricular zones at latter stages of brain and spinal cord and in the sympathetic chain. No signi¢cant expression was seen in di¡erentiated ¢elds. This restricted expression was maintained from embryonic day 11.5 to birth. In the periphery, large expression was detected in muscle and kidney and moderate expression in thyroid, pituitary gland, digestive system and bone. In the adult brain, SHIP2 was mainly restricted in structures containing neural stem cells such as the anterior subventricular zone, the rostral migratory stream and the olfactory tubercle. SHIP2 was also detected in the choroid plexuses and the granular layer of the cerebellum. The speci¢city of SHIP2 expression in neural stem cells was further demonstrated by (i) the dramatic increase in SHIP2 mRNA signal in neural cell adhesion molecule (N-CAM)-de¢cient mice, which present an accumulation of progenitor cells in the anterior subventricular zone and the rostral migratory stream, (ii) the abundant expression of 160-kDa SHIP2 by western blotting in proliferating neurospheres in culture and its downregulation in non-proliferating di¡erentiated neurospheres.

Immunology Letters, 2000

FcgRIIB are single-chain low-affinity receptors for IgG that bear an immunoreceptor tyrosine-base... more FcgRIIB are single-chain low-affinity receptors for IgG that bear an immunoreceptor tyrosine-based inhibition motif (ITIM) in their intracytoplasmic domain and that negatively regulate immunoreceptor tyrosine-based activation motif (ITAM)-dependent cell activation. In B cells, coaggregation of the B cell receptor (BCR) and FcgRIIB leads to an inhibition of B cell activation. Inhibitory properties of FcgRIIB have been related to the recruitment of SHIP, an SH2 domain-containing inositol 5-phosphatase (referred to as SHIP1), via ITIM phosphorylated FcgRIIB. Here, we demonstrate that the second SH2 domain-containing inositol 5-phosphatase SHIP2 could also bind to the FcgRIIB ITIM. As a model, a FcgRIIB deficient B cell line (IIA1.6), transfected with a cDNA encoding either w.t. FcgRIIB1% or FcgRIIB1% whose ITIM tyrosine was mutated has been used. SHIP2 tyrosine phosphorylation and association to the adaptator protein Shc were only found in transfectants expressing w.t. FcgRIIB1%. SHIP2 was also found to bind to a phosphopeptide corresponding to the ITIM sequence of FcgRIIB. There was no binding to the nonphosphorylated peptide. Finally, both SHIP2 and SHIP1 were coprecipitated with FcgRIIB1% upon coaggregation with BCR in IIA1.6 transfectants.

Infection and Immunity, 2002

The role of interleukin-12 (IL-12) has been clearly established in the resistance of C57BL/6 (B6)... more The role of interleukin-12 (IL-12) has been clearly established in the resistance of C57BL/6 (B6) mice to Leishmania major infection, but its involvement in the control of Leishmania mexicana infection remains to be determined. Here, we show the following. (i) L. mexicana, in contrast to L. major, induces the development of nonhealing lesions in B6 mice. (ii) Cells expressing IL-12p40, gamma interferon (IFN-␥), NOS2, and CD40L are numerous in the footpad lesion and/or the draining popliteal lymph node of animals infected for up to 14 weeks with L. mexicana. (iii) B6 mice, either IL-12p40 deficient or treated with IL-12p40-neutralizing antibodies, display a dramatic enhancement of primary and secondary lesions leading to death 10 weeks after inoculation with L. mexicana. (iv) Splenocytes harvested 4 and 8 weeks after infection of IL-12p40 ؊/؊ B6 mice with L. mexicana are unable to produce IFN-␥, but secrete IL-4, IL-10, and IL-18. Thus, the early control of L. mexicana infection by B6 mice is independent of IL-12, whereas IL-12 and Th1 responses play a key role in controlling the late stages of L. mexicana infection. However, they fail to resolve lesions, in contrast to L. major infection, emphasizing the different outcomes induced by these two Leishmania species in B6 mice.

PLOS Pathogens, 2012

Brucella are facultative intracellular bacteria that chronically infect humans and animals causin... more Brucella are facultative intracellular bacteria that chronically infect humans and animals causing brucellosis. Brucella are able to invade and replicate in a broad range of cell lines in vitro, however the cells supporting bacterial growth in vivo are largely unknown. In order to identify these, we used a Brucella melitensis strain stably expressing mCherry fluorescent protein to determine the phenotype of infected cells in spleen and liver, two major sites of B. melitensis growth in mice. In both tissues, the majority of primary infected cells expressed the F4/80 myeloid marker. The peak of infection correlated with granuloma development. These structures were mainly composed of CD11b + F4/80 + MHC-II + cells expressing iNOS/NOS2 enzyme. A fraction of these cells also expressed CD11c marker and appeared similar to inflammatory dendritic cells (DCs). Analysis of genetically deficient mice revealed that differentiation of iNOS + inflammatory DC, granuloma formation and control of bacterial growth were deeply affected by the absence of MyD88, IL-12p35 and IFN-c molecules. During chronic phase of infection in susceptible mice, we identified a particular subset of DC expressing both CD11c and CD205, serving as a reservoir for the bacteria. Taken together, our results describe the cellular nature of immune effectors involved during Brucella infection and reveal a previously unappreciated role for DC subsets, both as effectors and reservoir cells, in the pathogenesis of brucellosis.

Allergy, 2009

Background: Selected lactic acid bacteria were reported to prevent atopic dermatitis and experim... more Background: Selected lactic acid bacteria were reported to prevent atopic dermatitis and experimental asthma but the mechanisms of their immunomodulatory effects are not fully elucidated. In this study, the signaling pathways triggered by Lactobacillus plantarum NCIMB8826 were investigated and the potential use of this strain producing a variant of the mite allergen Der p 1 as live vaccine vehicle was evaluated.Methods: Mouse bone marrow-derived dendritic cells were stimulated with wild-type or a L. plantarum teichoic acid mutant to evaluate the secretion of cytokines. A recombinant L. plantarum expressing Der p 1 was engineered, its in vitro immunomodulatory properties were characterized and its prophylactic potential was evaluated in a Der p 1-sensitization murine model.Results: Mouse dendritic cells stimulated by L. plantarum triggered the release of interleukin-10 (IL-10), IL-12 p40, IL-12 p70 and tumor necrosis factor-alpha (TNF-α). IL-12 p40 secretion was dependent on nuclear factor-κB (NF-κB), mitogen-activated protein (MAP) kinases, Toll-like receptor 2 (TLR2), TLR9 and on the bacterial teichoic acid composition. Recombinant L. plantarum producing Der p 1 exhibited similar immunostimulatory properties as wild-type. Prophylactic intranasal pretreatment of mice with this recombinant strain prevented the development of the typical Th2-biased allergic response by a drastic reduction of specific IgE and the induction of protective allergen-specific IgG2a antibodies. Moreover, both wild-type or recombinant L. plantarum reduced airway eosinophilia following aerosolized allergen exposure and IL-5 secretion upon allergen restimulation.Conclusion: By combining both Th1-type immunostimulatory properties and an efficient allergen delivery capacity, recombinant L. plantarum producing Der p 1 represents a promising vaccine against house dust mite allergy.

Cellular Immunology, 1995

Immunology, 1996

The aim of this study was to investigate the mechanisms by which B7-related costimulatory molecul... more The aim of this study was to investigate the mechanisms by which B7-related costimulatory molecules (CD80, CD86) increase T-cell responsiveness to extracellular ligands. As a model study, the in vitro response of purified splenic CD4+ T cells to a bacterial superantigen, SEB, was characterized. Previous analysis of this experimental model led us to conclude that expression of B7-related molecules is strictly required in order to activate CD4+ T cells in the presence of bacterial superantigens. In the present report, we demonstrate that antigen-presenting cell-derived costimulatory signals regulate the kinetics of interleukin-2 (IL-2) production by SEB-activated splenic CD4+ T cells. Indeed, experiments performed with purified subpopulations of antigenpresenting cells and using B7-transfected cell lines indicated that increased levels of CD80 and/or CD86 cell surface expression is associated with a faster kinetics of IL-2 production in response to SEB. Accordingly, blocking of CD80 or CD86-derived signals by specific monoclonal antibodies led to a slower kinetics of IL-2 production in response to SEB. Thus these data demonstrate that similar strength of signal through the T-cell receptor can lead to immune responses displaying distinct kinetics depending on the level of costimulatory ligands on APC.

European Journal of Immunology, 2008

DiC14-amidine cationic liposomes were recently shown to promote Th1 responses when mixed with all... more DiC14-amidine cationic liposomes were recently shown to promote Th1 responses when mixed with allergen. To further define the mode of action of diC14-amidine as potential vaccine adjuvant, we characterized its effects on mouse and human myeloid dendritic cells (DC). First, we observed that, as compared with two other cationic liposomes, only diC14-amidine liposomes induced the production of IL-12p40 and TNF-α by mouse bone marrow-derived DC. DiC14-amidine liposomes also activated human DC, as shown by synthesis of IL-12p40 and TNF-α, accumulation of IL-6, IFN-β and CXCL10 mRNA, and up-regulation of membrane expression of CD80 and CD86. DC stimulation by diC14-amidine liposomes was associated with activation of NF-κB, ERK1/2, JNK and p38 MAP kinases. Finally, we demonstrated in mouse and human cells that diC14-amidine liposomes use Toll-like receptor 4 to elicit both MyD88-dependent and Toll/IL-1R-containing adaptor inducing interferon IFN-β (TRIF)-dependent responses.Supporting Information for this article is available at www.wiley-vch.de/contents/jc_2040/2008/37998_s.pdf

Scandinavian Journal of Immunology, 2007

The identification of subsets of CD4+ helper cells producing distinct pattern of cytokines has pr... more The identification of subsets of CD4+ helper cells producing distinct pattern of cytokines has provided a valuable framework for understanding how different effector populations of immune cells can be recruited in vivo during infection. In the view of most investigators, Th1 and Th2 cells produce factors that serve as their own autocrine factors and cytokines exerting suppressive activities on each other's development and activity. This concept intuitively explains the natural tendency of immune responses to become progressively polarized. However, several experimental observations appear difficult to rationalize with a simple, ‘symmetrical’ Th1/Th2 paradigm including those that Th1 cells do not produce their own growth factor; that both Th1 and Th2 cells can promote inflammatory responses; that interleukin-10 (IL-10) inhibits inflammatory responses in a Th1/Th2-independent fashion; that IL-10 promotes the development of Th1-type effector cells; and that IL-12 can amplify pre-established Th2 responses. The purpose of the present analysis is to provide a revised model for better understanding how cytokines regulate immune responses in vivo.

PLOS Pathogens, 2009

Leishmania major parasites reside and multiply in late endosomal compartments of host phagocytic ... more Leishmania major parasites reside and multiply in late endosomal compartments of host phagocytic cells. Immune control of Leishmania growth absolutely requires expression of inducible Nitric Oxide Synthase (iNOS/NOS2) and subsequent production of NO. Here, we show that CD11b + CD11c + Ly-6C + MHC-II + cells are the main iNOS-producing cells in the footpad lesion and in the draining lymph node of Leishmania major-infected C57BL/6 mice. These cells are phenotypically similar to iNOS-producing inflammatory DC (iNOS-DC) observed in the mouse models of Listeria monocytogenes and Brucella melitensis infection. The use of DsRed-expressing parasites demonstrated that these iNOS-producing cells are the major infected population in the lesions and the draining lymph nodes. Analysis of various genetically deficient mouse strains revealed the requirement of CCR2 expression for the recruitment of iNOS-DC in the draining lymph nodes, whereas their activation is strongly dependent on CD40, IL-12, IFN-c and MyD88 molecules with a partial contribution of TNF-a and TLR9. In contrast, STAT-6 deficiency enhanced iNOS-DC recruitment and activation in susceptible BALB/c mice, demonstrating a key role for IL-4 and IL-13 as negative regulators. Taken together, our results suggest that iNOS-DC represent a major class of Th1-regulated effector cell population and constitute the most frequent infected cell type during chronic Leishmania major infection phase of C57BL/6 resistant mice.

European Journal of Immunology, 1995

Exogenous glucocorticoid hormones are widely used as therapeutical agents, whereas endogenous glu... more Exogenous glucocorticoid hormones are widely used as therapeutical agents, whereas endogenous glucocorticoids may act as physiological immunosuppressants involved in the control of immune and inflammatory responses. The optimal activation of T lymphocytes requires two distinct signals: the major histocompatibility complex-restricted presentation of the antigen and an additional co-stimulatory signal provided by the antigen-presenting cells. There is ample evidence that, among the cells able to present the antigen, the dendritic cells (DC) have the unique property to activate antigen-specific, naive T cells in vitro and in vivo, and are therefore required for the induction of primary immune responses. In this work, we tested whether glucocorticoids affected the capacity of DC to sensitize naive T cells. Our data show that, in vitro, the steroid hormone analog dexamethasone (Dex) affects the viability of DC, selectively downregulates the expression of co-stimulatory molecules on viable DC, and strongly reduces their immunostimulatory properties. In vivo, a single injection of Dex results in impaired antigen presenting function, a finding which correlates with reduced numbers of splenic DC. These results show that glucocorticoids regulate DC maturation and immune function in vitro and in vivo and suggest that this mechanism may play a role in preventing overstimulation of the immune system.

Infection and Immunity, 2003

Cells of the dendritic cell (DC) lineage, by their unique ability to stimulate naive T cells, may... more Cells of the dendritic cell (DC) lineage, by their unique ability to stimulate naive T cells, may be of crucial importance in the development of protective immune responses to Leishmania parasites. The aim of this study was to compare the impact of L. major infection on DCs in BALB/c (susceptible, developing Th2 responses), C57BL/6 (resistant, developing Th1 responses), and tumor necrosis factor (TNF) ؊/؊ C57BL/6 mice (susceptible, developing delayed and reduced Th1 responses). We analyzed by immunohistochemistry the phenotype of infected cells in vivo. Granulocytes (GR1 ؉ ) and macrophages (CD11b ؉ ) appear as the mainly infected cells in primary lesions. In contrast, cells expressing CD11c, a DC specific marker, are the most frequently infected cells in draining lymph nodes of all mice tested. These infected CD11c ؉ cells harbored a particular morphology and cell surface phenotype in infected C57BL/6 and BALB/c mice. CD11c ؉ infected cells from C57BL/6 and TNF ؊/؊ C57BL/6 mice displayed a weak parasitic load and a dendritic morphology and frequently expressed CD11b or F4/80 myeloid differentiation markers. In contrast, some CD11c ؉ infected cells from BALB/c mice were multinucleated giant cells. Giant cells presented a dramatic accumulation of parasites and differentiation markers were not detectable at their surface. In all mice, lymph node CD11c ؉ infected cells expressed a low major histocompatibility complex II level and no detectable CD86 expression. Our results suggest that infected CD11c ؉ DC-like cells might constitute a reservoir of parasites in lymph nodes.

European Journal of Immunology, 1995

Excessive T cell activation induced by bacterial superantigens plays an important role in the pat... more Excessive T cell activation induced by bacterial superantigens plays an important role in the pathology associated with Gram-positive bacteremia. To gain insight into the early phases of T cell activation by bacterial enterotoxins in vivo, we investigated the ability of antibodies to well-defined co-stimulatory molecules to inhibit T cell activation and the subsequent toxic shock syndrome induced in BALB/c mice following the injection of staphylococcal enterotoxin B (SEB). We demonstrate here that a single dose of anti-B7.2 antibodies, but not anti-B7.1 antibodies, significantly inhibits T cell activation, as judged by lower systemic IL-2 release, blastogenesis and IL-2 receptor expression, and reduces the lethal effect of SEB in D-galactosamine-sensitized mice. These results demonstrate that co-stimulation through the B7.2 molecule plays an important role in the activation of T cells in response to SEB in vivo and suggest alternative therapies for septic shock caused by bacterial enterotoxins based on blocking antibodies to co-stimulatory molecules.

Immunology Letters, 2008

Dendritic cells (DCs) have been shown to express the tryptophan catabolizing enzyme indoleamine 2... more Dendritic cells (DCs) have been shown to express the tryptophan catabolizing enzyme indoleamine 2,3dioxygenase (IDO-1), a protein presently thought to exert dual and possibly contrasting effects on the immune response. Depletion of tryptophan and release of tryptophan catabolites have been shown to exert a tolerogenic influence on T cell responses, while the IDO enzymatic activity has been recently suggested to promote DC maturation. In this report, we have explored the putative role of IDO-1 in regulating DC biology by analyzing DC development and function from IDO-1 deficient mice. In keeping with previous observations, lack of IDO-1 expression was found to affect in vitro DC generation from bone mouse precursors cultured in the presence of GM-CSF. However, change in growth factor (Flt3L) and/or culture conditions (low-adherence vessels) abolished the difference observed between wt (wild type) and IDO-1-deficient, in vitro generated DCs. Moreover, IDO-1-deficient mice displayed a normal DC compartment in vivo, suggesting that IDO-1 does not play a significant role in DC development and function in vivo. Collectively, these observations suggest that despite a possible role for IDO-1 expression in regulating DC differentiation in vitro under commonly used culture conditions, IDO-1 is largely dispensable for DC development and function in vivo.

Cellular Immunology, 1999

Based on the observation that pathogen-derived lectins play an important role in cell adhesion an... more Based on the observation that pathogen-derived lectins play an important role in cell adhesion and invasion, we examined the possible role of host carbohydrate-bearing molecules in inducing the secretion of IL-12, a crucial proinflammatory cytokine. The ability of 12 plant lectins to recognize and stimulate naive murine mononuclear cells in vitro has been characterized in this study. Mitogenic lectins (comprising Con A, PHA, PSA, and LCA) were found to induce the secretion of multiple cytokines in vitro, including IL-2, interferon (IFN)-␥, and IL-12. Of interest, WGA, a nonmitogenic lectin unable to promote IL-2 secretion, was found to induce IL-12 and IFN-␥ production in a T and B cell-independent fashion. The functional properties of WGA were inhibited by N-acetylneuraminic acid and N,N-diacetylchitobiose. WGA therefore represents a potentially useful tool for the study of membrane glycoproteins involved in the early proinflammatory response characteristic of innate immunity.

Dendritic cells (DC) are described as "nature's adjuvant," since they have the capacity to sensit... more Dendritic cells (DC) are described as "nature's adjuvant," since they have the capacity to sensitize T cells in vivo upon first encounter with the antigen. The potent accessory properties of DC appear to develop sequentially. In particular, the ability to process antigens and to sensitize naive T cells develops in sequence, a process termed "maturation" that is well described in vitro. Here, we obtain evidence for maturation in vivo in response to the bacterial product lipopolysaccharide (LPS). Before LPS treatment, many DC are found at the margin between the red and white pulp. These cells lack the M342 and DEC-205 markers, but process soluble proteins effectively. 6 h after LPS, DC with the M342 and DEC-205 markers are found in increased numbers in the T cell areas. These cells have a reduced capacity to process proteins, but show increases in the B7 costimulator and T cell stimulatory capacity. 48 h after LPS, the number of DC in the spleen is reduced markedly. We interpret these findings to mean that LPS can cause DC in the marginal zone to mature and to migrate into and then out of the T cell areas.