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Based on studies reporting specific antibody titers, it is recommended to vaccinate preterm infants against Bordetella pertussis according to their chronological age. However, as specific T-cell responses also are involved in the protection against B. pertussis, we have determined whether highly preterm infants (<31 weeks) are able to mount these immune responses during vaccination. Forty-eight premature infants were vaccinated at 2, 3, and 4 months of their chronological age with an acellular (Pa; n = 24) or a whole-cell (Pw; n = 24) tetravalent diphtheria-tetanus-pertussis-polio vaccine, and blood samples were collected at 2, 3, and 6 months of age. Most of the Pa- and Pw-vaccinated infants developed at 3 or 6 months of age a gamma interferon (IFN-gamma) response to the B. pertussis antigens, accompanied by an interleukin-5 (IL-5) and IL-13 secretion for the Pa-vaccinated infants. No association was found between a very low infant birth weight, the occurrence of severe infections, and corticosteroid treatment or the administration of gammaglobulins with a low level of antigen-induced IFN-gamma secretion. We conclude that like full-term infants, most preterm infants are able to mount a specific cellular immune response to the administration of the first doses of an acellular or a whole-cell pertussis vaccine., Journal Article, Research Support, Non-U.S. Gov't, info:eu-repo/semantics/published

12e réunion commune de la Société de Néphrologie et de la Société Francophone de Dialyse (Bruxelles, 28/9 au 1/10/2010), info:eu-repo/semantics/published

Antibody deficiencies constitute the largest group of symptomatic primary immunodeficiency diseases. In several patients, mutations in CD19 have been found to underlie disease, demonstrating the critical role for the protein encoded by this gene in antibody responses; CD19 functions in a complex with CD21, CD81, and CD225 to signal with the B cell receptor upon antigen recognition. We report here a patient with severe nephropathy and profound hypogammaglobulinemia. The immunodeficiency was characterized by decreased memory B cell numbers, impaired specific antibody responses, and an absence of CD19 expression on B cells. The patient had normal CD19 alleles but carried a homozygous CD81 mutation resulting in a complete lack of CD81 expression on blood leukocytes. Retroviral transduction and glycosylation experiments on EBV-transformed B cells from the patient revealed that CD19 membrane expression critically depended on CD81. Similar to CD19-deficient patients, CD81-deficient patients had B cells that showed impaired activation upon stimulation via the B cell antigen receptor but no overt T cell subset or function defects. In this study, we present what we believe to be the first antibody deficiency syndrome caused by a mutation in the CD81 gene and consequent disruption of the CD19 complex on B cells. These findings may contribute to unraveling the genetic basis of antibody deficiency syndromes and the nonredundant functions of CD81 in humans., Case Reports, Journal Article, Research Support, Non-U.S. Gov't, SCOPUS: ar.j, info:eu-repo/semantics/published

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Peptide microarrays are useful tools for characterizing the humoral response against methylated antigens. They are usually prepared by printing unmodified and methylated peptides on substrates such as functionalized microscope glass slides. The preferential capture of antibodies by methylated peptides suggests the specific recognition of methylated epitopes. However, unmodified peptide epitopes can be masked due to their interaction with the substrate. The accessibility of unmodified peptides and thus the specificity of the recognition of methylated peptide epitopes can be probed using the in situ methylation procedure described here. Alternately, the in situ methylation of peptide microarrays allows probing the presence of antibodies directed toward methylated epitopes starting from easy-to-make and cost-effective unmodified peptide libraries. In situ methylation was performed using formaldehyde in the presence of sodium cyanoborohydride and nickel chloride. This chemical procedure converts lysine residues into mono- or dimethyl lysines., info:eu-repo/semantics/published

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Malgré la disponibilité d’un traitement curatif et un vaccin largement utilisé, l’OMS estime qu’approximativement un tiers de la population mondiale est infectée par Mycobacterium tuberculosis, l’agent étiologique de la tuberculose, et qu’environ 2 millions de personnes en meurent chaque année. La compréhension de l’épidémiologie de la tuberculose et les actions de contrôle de la maladie ont été, récemment, compliquées par l’émergence de bacilles tuberculeux résistants aux antibiotiques et par la synergie fournie par la co-infection avec le VIH. Une tendance alarmante pour la santé publique est l’émergence de souches résistantes à plusieurs antibiotiques (multi-résistantes, MDR), définies comme des isolats résistants au moins à l’isoniazide (INH) et la rifampicine (RIF), les deux agents anti-tuberculeux les plus puissants.La sélection de mutants résistants se produit chez le patient lorsque les taux d’antibiotiques présents dans le corps sont sub-thérapeutiques ou lorsque la thérapie est inappropriée. Un des facteurs favorisant est l’exceptionnelle durée de la chémothérapie. Le besoin de maintenir des taux élevés d’antibiotiques pendant des mois, combiné avec la toxicité inhérente des agents, résultent en une observance incomplète du traitement par le patient et le risque d’acquérir des résistances. La résistance aux antibiotiques chez M. tuberculosis résulte d’altérations dans des gènes chromosomiques spécifiques. Les causes génétiques de la résistance ont été définies pour certains antibiotiques bien que plusieurs inconnues persistent. Le présent travail a consisté en l’étude du problème de la résistance aux antibiotiques anti-tuberculeux par différentes approches :l’analyse génétique des mécanismes de résistance, l’évaluation de l’activité thérapeutique de nouvelles molécules et la caractérisation du profil de résistance de souches cliniques. L’acide p-aminosalicylique (PAS) est un antibiotique bactériostatique de deuxième ligne dont le mécanism, Doctorat en Sciences biomédicales et pharmaceutiques, info:eu-repo/semantics/nonPublished

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Two different types of pertussis vaccines are currently available to protect children against whooping cough, the first-generation whole-cell (Pw) vaccines and the more recent acellular (Pa) vaccines. Both types provide good protection, yet induce different types of immune responses in 6-month-old infants, with a strong Th1 response induced by Pw vaccines compared to a mixed Th1/Th2 response and a delay in non-specific IFN-gamma secretions after the administration of Pa vaccines. We show here that at 13 months of age, most Pw- or Pa-vaccinated children display Bordetella pertussis-specific T-cell responses, in addition to significant antibody levels, although a higher Th2/Th1 cytokine ratio remained in Pa recipients compared to Pw recipients. In contrast, the proportion of children with tetanus toxin-specific T-cell responses was lower in Pa than in Pw vaccine recipients, although most children had protective anti-tetanus toxin IgG levels. In addition, the global Th2 bias observed in 6-month-old infants vaccinated with a Pa vaccine was normalized at 13 months., Journal Article, Research Support, Non-U.S. Gov't, info:eu-repo/semantics/published

Antigen-specific gamma interferon (IFN-gamma) has been demonstrated to participate in protection against Bordetella pertussis infection. Circulating mononuclear cells from B. pertussis-infected and from pertussis-vaccinated infants secrete high amounts of IFN-gamma after in vitro stimulation by B. pertussis antigens, but with a large variation in the secreted IFN-gamma levels between individuals. We show here that the inhibition of the specific IFN-gamma response can be at least partially attributed to IL-10 secretion by monocytes. This IL-10 secretion was not associated with polymorphisms at positions -1082, -819, and -592 of the IL-10 gene promoter, suggesting that other genetic or environmental factors affect IL-10 expression and secretion., Journal Article, Research Support, Non-U.S. Gov't, info:eu-repo/semantics/published

A number of different interferon-gamma ELISpot protocols are in use in laboratories studying antigen-specific immune responses. It is therefore unclear how results from different assays compare, and what factors most significantly influence assay outcome. One such difference is that some laboratories use a short in vitro stimulation period of cells before they are transferred to the ELISpot plate; this is commonly done in the case of frozen cells, in order to enhance assay sensitivity. Other differences that may be significant include antibody coating of plates, the use of media with or without serum, the serum source and the number of cells added to the wells. The aim of this paper was to identify which components of the different ELISpot protocols influenced assay sensitivity and inter-laboratory variation. Four laboratories provided protocols for quantifying numbers of interferon-gamma spot forming cells in human peripheral blood mononuclear cells stimulated with Mycobacterium tuberculosis derived antigens. The differences in the protocols were compared directly. We found that several sources of variation in assay protocols can be eliminated, for example by avoiding serum supplementation and using AIM-V serum free medium. In addition, the number of cells added to ELISpot wells should also be standardised. Importantly, delays in peripheral blood mononuclear cell processing before stimulation had a marked effect on the number of detectable spot forming cells; processing delay thus should be minimised as well as standardised. Finally, a pre-stimulation culture period improved the sensitivity of the assay, however this effect may be both antigen and donor dependent. In conclusion, small differences in ELISpot protocols in routine use can affect the results obtained and care should be given to conditions selected for use in a given study. A pre-stimulation step may improve the sensitivity of the assay, particularly when cells have been previously frozen., Journal Article, Research Support, Non-U.S. Gov't, info:eu-repo/semantics/published

BACKGROUND: Positive skin prick tests (SPT) for food allergens and specific IgE (sIgE) in serum indicate sensitization but do not enable distinction between sensitized but tolerant and clinically allergic patients. OBJECTIVE: Herein, we evaluate the clinical relevance of basophil activation tests (BATs) for peanut or egg allergy diagnosis. METHODS: Thirty-two peanut-allergic, 14 peanut-sensitized (sIgE(+) and/or SPT(+) to peanuts) but tolerant children and 29 controls with no history of an adverse reaction to peanuts were included. Similarly, 31 egg-allergic, 14 egg-sensitized children (sIgE(+) and/or SPT(+) to egg white) and 22 controls were studied. Flow cytometric analysis of CD63 expression or CD203c upregulation on basophils and the production of leukotrienes (LT) were performed in response to an in vitro crude peanut extract or ovalbumin (OVA) challenge. RESULTS: After in vitro peanut challenge, the basophils from peanut-allergic children showed significantly higher levels of activation than those from controls (P<0.001). After OVA challenge, a similar distinction (P<0.001) was observed between egg-allergics and controls. Interestingly, the majority of egg- or peanut-sensitized children failed to activate basophils, respectively, in response to OVA and peanut challenge. The sensitivity of the CD63, CD203c and LT assay was 86.7%, 89.5% and 76.0% with a specificity of 94.1%, 97.1% and 94.6% for peanut allergy diagnosis. The corresponding performances of BATs applied to egg allergy diagnosis were 88.9%, 62.5% and 77.8% for the sensitivity and 100%, 96.4% and 96.4% for the specificity. CONCLUSION: Neither conventional tests nor BATs are sensitive and specific enough to predict food allergy accurately. However, BATs may helpfully complete conventional tests, especially SPT, allowing improved discrimination between allergic and non-allergic individuals., Journal Article, Research Support, Non-U.S. Gov't, SCOPUS: ar.j, FLWIN, info:eu-repo/semantics/published

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The role of tumor-associated macrophages (TAMs) is controversial. Although most studies on different cancer types associate them with a poorer prognosis, interestingly in colon cancer, most articles indicate that TAMs prevent tumor development; patients with high TAMs have better prognosis and survival rate. M1-polarized macrophages produce high level of tumor necrosis factor-alpha, interleukin-1 beta or reactive oxygen species, which can effectively kill susceptible tumor cells. In contrast, M2-polarized macrophages can secrete different factors that promote tumor cell growth and survival or favor angiogenesis and tissue invasion. Considering the beneficial role of TAMs in colon cancer, we speculated that they may not display the M2 polarization commonly observed in tumor microenvironment, but rather develop M1 properties. Therefore, we used an in vitro model to analyze the effects of supernatants from M1-polarized macrophages on DLD-1 colon cancer cells. Our data indicate that the conditioned medium from LPS-activated macrophages (CM-LAM) contains a high level of granulocyte-macrophage colony-stimulating factor, interleukins-1 beta, -6, -8 and tumor necrosis factor-alpha, and that it exerts a marked growth inhibitory activity on DLD-1 cells. Prolonged exposure to CM-LAM results in cell death by apoptosis. Such exposure to CM-LAM leads to the modulation of gal-3 expression: we observed a marked downregulation of gal-3 mRNA and protein expression following CM-LAM treatment. We also describe that the knockdown of gal-3 sensitizes DLD-1 cells to CM-LAM. These data suggest an involvement of gal-3 in the response of colon cancer cells to proinflammatory stimuli, such as the conditioned medium from activated macrophages.

Le système immunitaire nous protège contre les infections et les cancers. Cependant, des réponses immunitaires excessives ou incontrôlées peuvent causer des pathologies potentiellement mortelles comme le choc endotoxinique, des maladies auto-immunes, ou le rejet d’allogreffe. Une compréhension claire des mécanismes de régulation des réponses immunes permettrait d’envisager de nouvelles thérapeutiques plus ciblées. Dans ce travail réalisé chez la souris, la modulation de la réponse inflammatoire à une toxine bactérienne, le lipopolysaccharide (LPS), a été utilisée comme archétype de la régulation immunitaire. Ceci nous a permis de démontrer que la régulation de la prolifération homéostatique de lymphocytes T CD4+CD25- par des lymphocytes T régulateurs naturels tempère la réponse inflammatoire au LPS. Cet effet est notamment dépendant d’une diminution de la sécrétion d’IFN-γ par les lymphocytes activés par la prolifération induite par la lymphopénie. Nous avons aussi observé, que la désensibilisation du système immunitaire inné vis-à-vis du LPS, suite à des injections répétées d’endotoxine, induit le développement de cellules myéloïdes suppressives (myeloïd-derived suppressor cells MDSC) capables de réguler des réponses lymphocytaires T in vitro et in vivo. Nous avons pu mettre en évidence que l’effet suppresseur des MDSC est dépendant de leur expression de l’enzyme hème oxygénase-1 et de leur production d’IL-10. Le dialogue constant entre les cellules de l’immunité innée et de l’immunité acquise assure donc à la fois l’activation et la régulation du système immunitaire. Dans la majorité des cas, ceci permet aux réponses immunitaires d’être efficaces sans être excessives. La mise en évidence de ces processus identifie les lymphocytes T régulateurs et les cellules myéloïdes suppressives comme des éléments clefs de la régulation d’un processus inflammatoire. Les traitements immunosuppresseurs, les thérapies cellulaires et les greffes de moelle ont des effets variables e, Doctorat en Sciences médicales, info:eu-repo/semantics/nonPublished

Globalement, un tiers de la population mondiale est infectée par Mycobacterium tuberculosis, l'agent infectieux de la tuberculose (TB). Fort heureusement, seuls 5 à 10 % des individus infectés développent un jour une TB active. Les individus non malades restent cependant infectés à vie, on parle d'infection latente. Chaque année, 8-10 millions nouveaux cas de tuberculose active sont recensés et M. tuberculosis est responsable de 1,5 à 2 millions de décès. Depuis plus d'une décennie, M. tuberculosis s'est étroitement associé à l'infection par le virus de l'immunodéficience humaine. Cette alliance néfaste représente une importante menace pour les pays en voie de développement, car ces 2 pathogènes déciment les forces vives de ces populations. Il faut malheureusement rajouter à ce triste tableau une fréquence grandissante de souches multi-résistantes, voire extensivement multi-résistantes. Face à ces souches, les avancées thérapeutiques du siècle dernier sont pratiquement réduites à néant. Considérant ces données, il est désormais crucial d'améliorer nos outils de dépistage de l'infection latente, de diagnostic de la maladie active, de prévention (vaccins) et de traitement. Pour atteindre ces objectifs, une des pistes est la caractérisation détaillée des réponses immunitaires. En comparant les réponses immunitaires des sujets infectés de manière latente à celles liées à la maladie active, nous pourrons peut-être comprendre certains mécanismes de protection. L'étude des réponses immunitaires induites par la « Heparin-Binding-Hemagglutinin » (HBHA) s'est faite dans cet objectif. La HBHA est une adhésine exprimée par le complexe M. tuberculosis. Elle est impliquée dans la dissémination extrapulmonaire du bacille et constitue donc un facteur de virulence. Par ailleurs, une vaccination de souris par seulement 3 doses de 5 µg de HBHA suffit à protéger de l'infection avec une efficacité comparable à celle du vaccin BCG. Chez l'homme, les sujets sains mais infectés développ, Doctorat en Sciences biomédicales et pharmaceutiques, info:eu-repo/semantics/nonPublished

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10ème réunion commune de la Société de Néphrologie et de la Société Francophone de Dialyse (Marrakech, 26-29 novembre 2008), info:eu-repo/semantics/published

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10ème réunion commune de la Société de Néphrologie et de la Société Francophone de Dialyse (Marrakech, 26-29 novembre 2008), info:eu-repo/semantics/published

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