Die Infektion mit Schistosoma mansoni resultiert in einer Th2-Immunantwort mit Eosinophilie und erhöhtem IgE-Titer, wobei der wasserlösliche Extrakt der S. mansoni Eier (SEA) ausreicht um diese Reaktion auszulösen. In der vorliegenden Arbeit konnte demonstriert werden, dass sich IL-4-produzierende CD4+ T-Lymphozyten in Zellkulturen mit SEA-konditionierten Dendritischen Zellen (DCs) trotz gleichzeitig vorkommenden IFN-gamma entwickeln und SEA die Expression von Faktoren in DCs, die üblicherweise mit einer Th1-Antwort einhergehen, auf Transkriptions- und Proteinebene selektiv hemmt. Um den Faktor aus S. mansoni Eiern zu isolieren, der zur Expression von IL-4 in CD4+ Zellen und zur Inhibition von IL-12 in DCs führt, wurde eine Gelfiltrationschromatographie der exkretorisch/sekretorischen Ei-antigene (ES) durchgeführt und die Fraktionen in vitro getestet. Darin wurde gezeigt, dass Fraktionen mit einer Proteinbande von 30 kD die Expression von IL-4 in CD4+ Zellen induzieren. Dieses ES-Protein wurde durch N-terminale Sequenzierung als hepatotoxische Ribonuclease Omega-1 identifiziert, welches ebenfalls die Expression von IL-12 in DCs inhibiert und die Produktion von IL-4 in CD4+ Zellen bei einer 10-fach geringeren Proteinkonzentration als mit dem Kontrollansatz SEA induziert. Zudem sollte untersucht werden, inwieweit Toll-like Rezeptoren in der Generierung einer Th2 Antwort gegen schistosomale Antigene involviert sind. Dazu wurden TLR2-, TLR3-, TLR4- und MyD88-defiziente Mäuse mit S. mansoni infiziert und immunologische und pathologische Daten in der akuten und chronischen Phase der Infektion analysiert. Demnach sind TLR2, TLR3, TLR4 und MyD88-abhängige Signaltransduktionswege nicht für eine-Th2 Antwort notwendig, jedoch ist letzteres Molekül in der Ausprägung der typischen Leberfibrose involviert. / Infection with Schistosoma mansoni results in the induction of a Th2 immune response, eosinophilia and increased levels of IgE. The water-soluble extract of S. mansoni eggs (SEA) is sufficient to promote TH2 polarization in a dendritic cell-dependent manner. In this thesis, it was demonstrated that IL-4+ CD4+ cells emerge in cultures with SEA-conditioned dendritic cells (DCs) in the presence of IFN-gamma and that SEA inhibits selectively the expression of IL-12 and co-stimulatory markers in DCs on the transcriptional and protein level. To identify the putative protein in S. mansoni eggs mediating a Th2 induction, a gel filtration chromatography of the excretory/secretory egg antigens (ES) was conducted and the fractions tested in vitro. Fractions containing a single band of 30 kD were sufficient to promote IL-4 induction in naïve CD4+ cells. Using N-terminal sequencing this ES-protein was identified as the hepatotoxic S. mansoni ribonuclease omega-1 which displayed both biological functions observed with SEA: inhibition of IL-12 in LPS-stimulated DCs and induction of IL-4+ CD4 cells at a 10 fold lower protein concentration than SEA. In order to understand, if the innate immune receptors TLR2, TLR3, TLR4 or the TLR adaptor molecule MyD88 are involved in the generation of the Th2 response against schistosomal antigens, the respective knock out mice were infected and immunological and pathological parameters were analyzed during acute and chronic phase of infection. This study showed that during S. mansoni infection TLR2, TLR3, TLR4 and TLR activation through the MyD88-dependent pathway are neither required for the induction (priming and polarization) nor for the down-regulation of Th2 responses, however, the fibrotic response against S. mansoni eggs was significantly reduced in MyD88-deficient mice suggesting a detrimental role of this pathway in liver pathology.

CD4+ T-Helferzellen (Th) entwickeln sich zu Th1 und Th2 Zellen, die nach ihrer Funktion und Zytokinexpression eingeteilt werden. Die differentielle Induktion von Th Zellen, die Th1 oder Th2 Zytokine exprimieren, ist der Schlüssel zur Regulation von Immunantworten bei Infektionskrankheiten, Allergien und Autoimmunerkrankungen. Daher können stabil exprimierte Oberflächenmoleküle, die spezifisch für die funktionell unterschiedlichen Th Zellen sind, von besonderer Bedeutung für die Analyse und selektive funktionelle Modulation von Th Subtypen sein und erlauben es neue therapeutische Strategien für die Behandlung von allergischen und Autoimmunerkrankungen zu etablieren. TIM-3 wurde kürzlich identifiziert als ein Molekül, welches selektiv auf der Oberfläche von Th1 Zellen exprimiert wird und welches möglicherweise eine Rolle bei der Induktion von Autoimmunerkrankungen spielt. Um monoklonale Antikörper gegen humanes TIM-3 zu produzieren, wurde die humane TIM-3 cDNA von in vitro generierten dendritischen Zellen kloniert. Der extrazelluläre Teil des Gens wurde in den prokaryotischen Expressionsvektor pQE100S insertiert und in E.coli BL21(DE3) exprimiert. Die gesamte codierende Sequenz wurde in den eukaryotischen Expressionsvektor pIRES2EGFP subkloniert und auf der Oberfläche von Säugetierzellen exprimiert. Stabile Transfektanten der CHO-K1 und HEK293 Zelllinie wurde etabliert. Die Balb/c Mäuse wurden mit löslichem und unlöslichem rekombinanten humanem TIM-3 sowie mit stabilen Transfektanten für humanes TIM-3 immunisiert. Milzzellen dieser Tiere wurden mit der Myelomzelllinie P3 X 63 Ag8.653 fusioniert. Die entwickelten Hybridome wurden im ELISA und mittels FACS auf Spezifität gegen humanes TIM-3 hin untersucht. Ein Klon der generierten Hybridome war positiv im ELISA zeigte jedoch kein Signal gegen TIM-3 auf der Oberfläche von Zellen. / CD4+ T helper (Th) cells develop into effector Th1 and Th2 cells, which are frequently categorized according to their function and cytokine expression. The differential induction of Th cells expressing Th1 or Th2 cytokines is key to the regulation of immune responses by infectious diseases, allergies and autoimmnune diseases. Thus, stably expressed surface molecules, significant for functionally different types of Th cells could be of utmost importance for the analysis and selective functional modulation of Th subsets and provide new therapeutic strategies for the trestment of allergic or autoimmune diseases. TIM-3 was recently identified as a molecule that is selectively expressed on the surface of Th1 cells and that may have a role in the induction of autoimmune disease. To produce monoclonal antibody of human TIM-3, the human TIM-3 cDNA was cloned from in vitro generated dendritic cells. The extracellular domain of human TIM-3 was inserted into prokaryotic expression vector pQE100S and expressed in E.coli BL21(DE3). The whole coding region was subcloned into eukaryotic expression vector pIRES2EGFP and expressed on the surface of mammalian cells. The stable transfectants of CHO-K1 and HEK-293 cell line was established. The BALB/c mice were immunized with soluble and insoluble recombinant human TIM-3 and also with stable transfectants. Splenocytes were fused with P3 X 63 Ag8.653 myeloma cells. The generated hybridomas were tested in ELISA and FACS for specificity against human TIM-3. A generated clone was positive in ELISA but did not respond to the TIM-3 molecule on the cell surface.

0 objetivo deste trabalho foi definir se a carga parasitaria na fase aguda da doenga de Chagas experimental afeta a parasitemia, a patologia e a resposta imune na fase cr6nica. Para obtengelo de diferentes cargas parasitoirias na fase aguda, camundongos A/J foram infectados corn 103 OU 105 formas tripomastigotas de T. cruzi e analisados urn ano depois. Os animais cr6nicos infectados corn 105 formas tripomastigotas apresentaram maior nivel de parasitemia residual, maior intensidade de inflamagclo no coragtio e no moscuio esquel6tico e maior grau de ativa95o do sistema imune do que os animais infectados corn 103 formas. Em reiagclo aos parametros imuno16gicos analisados, observou-se nos animais infectados corn 105 formas: i) expansio das populag6es B220-CD5- e CD8\'; ii) freq0@ncia maior de blastos nas populag6es linfocit@rias B220\', CD8\' e CD4\'; iii) mudanga acentuada nas c61ulas CD4+ para o fen6tipo CD4+CD45RBI-ow, indicando urn aumento das c61ulas efetoras elou de mem6ria; iv) freqGC=ncias elevadas de blastos CD4+CD45RB Hig\' e CD4+CD45RB Low; vi) nomero superior de c61ulas secretoras de lg principaimente IgG2a; v) niveis superiores de anticorpos IgG2a e IgGl especificos e vii) maior produgclo de IFN-Y e de IL-4. Estes resultados indicam que a carga parasitaria na fase aguda da infecggto influencia a ativagclo do sistema imune e o desenvolvimento da patologia na fase cr6nica da doenga de Chagas. / The objective of this project is to evaluate if the parasite load in the acute phase experimental Chagas\' disease affects the parasitemias, the pathology and the immune response in the chronic phase. To obtain low- and high-parasite loads in the acute phase of the disease, AlJ mice were infected with 103 or 105 T. cruzi trypomastigotes of the Y strain, and treated on day 6 with Benzonidazol. One year later, chronic mice were screened for subpatent parasitemias, tissue pathology and immune response. Mice infected with the high parasite inoculum showed higher levels of chronic parasitemias, heart and striated muscle inflammation and activation of the immune system when compared to mice infected with the low¬dose inoculum. Concerning the activation of the immune system, the main findings in high-dose infected mice were: i) increased numbers of splenocytes, with preferential expansion of CD8+ and B220-CDS- cells, many of them bearing a macrophage phenotype; ii) higher frequencies of B (B220+), CD4+ and CD8+ large lymphocytes; iii) a shift of CD4+ cells towards a CD4SRBLow phenotype; iv) increased frequencies of both CD4SRBLow and CD4SRBHigh large CD4+ cells; v) augmented numbers of total Ig-secreting cells, with predominance of IgG2a¬producing cells, and; vi) increased production of IFN-y and IL-4. In addition, these mice presented lower IgM and higher IgG2a and IgG1 parasite-specific serum antibody levels. Our results indicate that the parasite load at the acute phase of T. cruzi infection influences the activation of the immune system and development of Chagas pathology at the late chronic phase of the disease.

Les canaux calciques incluent notamment les canaux dépendants des stocks (ORAI) et dépendants du voltage (Cav) qui sont considérés de première importance pour l'entrée du calcium dans les cellules non-excitables et excitables, respectivement. Les canaux calciques voltage-dépendants tels que Cav1 sont essentiels pour le fonctionnement des cellules excitables, notamment la transmission neuronale, la contraction musculaire ou la sécrétion hormonale. Cependant, de nombreuses études montrent désormais que les canaux Cav1 sont aussi exprimés dans des cellules non excitables, et sont importants pour les fonctions effectrices des lymphocytes T. Les canaux Cav1 sont constitués de la sous-unité a1 formant le pore ionique et des sous-unités auxiliaires ß et a2δ. Ces sous-unités sont importantes pour l'activité électrique du canal, mais aussi pour sa régulation, sa stabilité et son expression à la membrane plasmique dans les cellules excitables. Les travaux de notre groupe ont clairement identifié les sous-unités a1 de Cav1.2 et Cav1.3 comme essentielles pour la fonction des lymphocytes Th2, une sous-population de cellules T responsable des maladies allergiques. L'inhibition pharmacologique et génétique de ces canaux réduit de manière significative l'expression des cytokines dans les lymphocytes Th2 chez la souris, mais pas dans les Th1. Le but de mon travail a été de comprendre si les sous-unités auxiliaires des canaux Cav, et plus particulièrement la sous-unité ß, sont nécessaires au fonctionnement des canaux Cav1 dans les lymphocytes Th2 qui ne sont pas des cellules excitables. Nous avons utilisé des oligonucléotides antisens ciblant toutes les sous-unités ß afin de réduire l'expression de ß1 et ß3, les deux sous-unités exprimées dans des lymphocytes Th2. La transfection de Th2 murines et humaines avec ces oligonucléotides diminue l'influx de calcium dépendant du TCR et l'expression des cytokines. En outre, l'effet des oligonucléotides antisens semble résulter de la perte d'expression de la sous-unité a1 selon un mécanisme similaire à celui décrit dans les neurones. De plus, L'utilisation de shRNA spécifiques de ß1 et ß3 dans les Th2 de souris montre un rôle essentiel la sous-unité ß1 dans la réponse fonctionnelle des lymphocytes Th2. Enfin, les antisens Cavß diminuent l'inflammation dans un modèle d'asthme allergique chez la souris, de même qu'un inhibiteur pharmacologique des sous-unités a2δ. Ce travail a donc permis d'identifier les sous-unités auxiliaires des canaux Cav comme de nouvelles cibles thérapeutiques potentielles dans le cadre des maladies allergiques telles que l'asthme. / Calcium channels include store-operated (ORAI) and voltage-gated (Cav) channels that are considered to be important for calcium entry in non-excitable and excitable cells, respectively. Voltage-gated calcium channels such as Cav1 are essential for excitable cell function, including neuronal transmission, muscle contraction or hormone secretion. However, numerous studies show that Cav1 channels are expressed in non-excitable cells as well, and are important for T cell effector functions. Cav1 channels are composed of the a1 subunit forming the ion pore and auxiliary subunits ß and a2δ. These subunits are important for the electric activity of the channel but also for its regulation, its stability and its expression at the plasma membrane in excitable cells. Our group clearly identified the a1 subunit of Cav1.2 and Cav1.3 channels as essential for the function of Th2 lymphocytes, a T cell subset responsible for allergic diseases. Pharmacological and genetic inhibition of these channels significantly reduces the expression of cytokines in mouse Th2 cells, but not in Th1 cells. The goal of my work was to understand whether the auxiliary subunits of Cav channels, particularly the ß subunit, are necessary for the function of Cav1 channels in Th2 lymphocytes that are not excitable cells. We used antisense oligonucleotides targeting all ß subunits to reduce the expression of ß1 and ß3, the two subunits expressed in Th2 lymphocytes. Transfection of murine and human Th2 with these oligonucleotides decreases TCR-dependent calcium influx and cytokine expression. In addition, the effect of the Cavß antisense oligonucleotides seems to result from the loss of expression of the a1 subunit, as similarly described in neurons. In addition, the use of shRNA specific to ß1 and ß3 in mouse Th2 shows a critical role the ß1 subunit in the functional response of Th2 lymphocytes. Finally, the Cavß antisense oligonucleotides reduce the airway inflammation in an allergic asthma model in mice, as well as a pharmacological inhibitor of a2δ subunits. This work has identified auxiliary subunits of Cav channels as new potential therapeutic targets in allergic diseases such as asthma.

A forma indeterminada é a fase inicial da hanseníase, que se caracteriza histologicamente pelo infiltrado inflamatório leve, não granulomatoso, de linfócitos e histiócitos ao redor de vasos, anexos e nervos. No local de entrada do M. leprae, as células apresentadoras de antígeno do tipo células dendríticas são as primeiras a encontrar o bacilo. Este, no interior de células dendríticas, desencadeia a produção local de citocinas e quimiocinas, que resultam em proliferação de linfócitos T helper 1 ou T helper 2, assim definindo uma resposta imune celular ou humoral, respectivamente. As lesões tuberculoides mostram predominância das citocinas de padrão Th1 como IL-2, TNF-alfa, IFN-y, IL-12 e IL-18, enquanto que nas lesões virchowianas predominam citocinas de padrão Th2, como IL-4, IL-5, IL-10 e TGF-beta. Na pele, as principais células dendríticas são células dendríticas mieloides, células de Langerhans e alguns dendrócitos dérmicos. São identificadas respectivamente pela expressão imuno-histoquímica de S100, CD1a e Fator XIIIa. Células de Langerhans e dendrócitos dérmicos Fator XIIIa positivos estão aumentados em quantidade nas lesões tuberculoides quando comparadas com lesões virchowianas. Os objetivos do presente estudo foram: 1) caracterizar a inflamação \"in situ\" na hanseníase indeterminada através da quantificação das marcações imuno-histoquímicas de: CD57, CD4, CD8, CD1a, S100, FXIIIa, CD68, Foxp3, CD123, IL-1, IL-2r, IL-4, IL-6, IL-8, IL-10, IL-12, IL-18, IFN-, TNF-alfa, TGF-beta, iNOS, granzima B, receptor Toll-like 2/4, e antígeno BCG, 2) comparar o perfil fenotípico e citocínico das lesões na hanseníase indeterminada entre grupos de reação de Mitsuda positiva e negativa, a fim de investigar se existem padrões que possam prever para qual forma a doença evoluiria, e 3) revisar a histopatologia da forma indeterminada através da análise semiquantitativa das alterações vistas à coloração de hematoxilina/eosina. Foram selecionadas 15 lesões de pacientes com hanseníase indeterminada. Foram usados grupos controles de expressão de Fator XIIIa e CD1a em 10 casos de pele normal. A histopatologia mostrou discretas alterações epidérmicas, como alteração vacuolar e exocitose de linfócitos (33% dos casos cada), apoptose de queratinócitos (26%), atrofia e acantose (06% dos casos cada); infiltrado inflamatório linfomononuclear neural (100%), perivascular superficial (100%), perivascular profundo (93%), peri-écrino (40%) e peri-folículo pilossebáceo (20%), além de melanófagos em 93% dos casos. Esses achados mostraram que nem sempre todos os ambientes estão acometidos por inflamação na histopatologia. Em 66% dos pacientes foi encontrado antígeno bacilar (por Fite-Faraco ou técnica imuno-histoquímica anti-BCG), portanto a forma indeterminada não deve ser considerada sistematicamente como paucibacilar. Não houve diferença significativa de padrões de marcadores entre os grupos Mitsuda positivo e negativo. No microambiente inflamatório das lesões houve expressões significativas de TLR 2/4, Fator XIIIa, CD4, CD8, IL-2r, IL-4, IL-10, iNOS e TGF-beta. A expressão importante de IL-4, IL-10 e TGF- beta nas lesões de hanseníase indeterminada significaram tendência de resposta imune para o polo Th2, um ambiente de tolerância à permanência do bacilo. A baixa expressão de IFN-y colaborou para a inexpresiva resposta Th1. Não houve diferença significativa na expressão de CD1a entre as lesões e pele normal. Fator XIIIa foi expresso em mais que 50 células/mm2 em todos os casos, com quantidades significativamente maiores que outras células dendríticas nas lesões (S100, CD68, CD123) e que a pele normal. Estes achados demonstraram a importância dos dendrócitos dérmicos Fator XIIIa positivos na apresentação de antígeno na fase inicial da hanseníase / The indeterminate form is the initial stage of leprosy, which is characterized histologically by mild inflammatory infiltrate, non granulomatous, with lymphocytes and histiocytes around vessels, nerves and adnexals. When M. leprae enter the host, antigen-presenting cells of dendritic type are the first cells to find the bacillus. Once inside dendritic cells, the bacillus elicits local production of cytokines and chemokines, which result in proliferation of T lymphocytes helper 1 or T helper 2, thereby defining a cellular or humoral immune response, respectively. The tuberculoid lesions show predominance of Th1 cytokines such as IL-2, TNF-alfa, IFN-y, IL-12 and IL-18, whereas in the lepromatous lesions predominate cytokines of Th2 pattern such as IL-4, IL-5 IL-10 and TGF-beta. In the skin, main dendritic cells are myeloid dendritic cells, Langerhans cells, and some dermal dendrocytes. They are identified respectively by immunohistochemical expression of S100, CD1a and Factor XIIIa. Langerhans cells and dermal dendrocytes Factor XIIIa positive are increased in number in tuberculoid lesions compared with lepromatous lesions. The objectives of this study were: 1) to characterize \"in situ\" inflammation in indeterminate leprosy through the quantification of immunohistochemical markers: CD57, CD4, CD8, CD1a, S100, FXIIIa, CD68, Foxp3, CD123, IL-1, IL-2r, IL-4, IL-6, IL-8, IL-10, IL-12, IL-18, IFN-y, TNF-alfa, TGF-beta, iNOS, granzyme B, Toll-like receptor 2/4, and BCG antigen, 2) compare the phenotypic and cytokinic profile of indeterminate leprosy lesions between positive and negative Mitsuda reaction groups in order to investigate if there are patterns that can predict which way the disease may evolve, and 3 ) review the histopathology of the indetermate form by semi-quantitative analysis of changes seen in hematoxylin / eosin. Fifteen lesions of patients with indeterminate leprosy were selected. There was used control groups of Factor XIIIa and CD1a expression in 10 cases of normal skin. Histopathology showed discrete epidermal changes, such as vacuolar changes and lymphocyte exocytosis (in 33% of cases each), keratinocyte apoptosis (26%), atrophy and acanthosis (in 06% of cases each); neural lymphocytic inflammatory infiltrate (100%), superficial perivascular (100%), deep perivascular (93%), peri-eccrine (40%), peri-pilosebaceous follicle (20%), and melanophages in 93% of cases. These findings showed that not always all environments are affected by inflammation in histopathology. In 66% of patients it was found bacterial antigen (by Fite-Faraco or immunohistochemical technique anti-BCG), so the indeterminate form should not be systematically considered as paucibacillary. There was no significant difference in phenotypic and cytokinic patterns between the positive and negative Mitsuda groups. In the microenvironment of inflammatory lesions there was significant expression of TLR 2/4, Factor XIIIa, CD4, CD8, IL-2r, IL-4, IL-10, TGF-beta and iNOS. The important expression of IL-4, IL-10 and TGF-beta in indeterminate leprosy meant tendency to Th2 immune response pole, an environment of tolerance to permanence of bacillus. Low IFN-? expression contributed to the negligible Th1 response. There was no significant difference in the expression of CD1a between the lesions and normal skin. Factor XIIIa was expressed as greater than 50 cells / mm2 in all cases, with significantly larger quantities than other dendritic cells in lesions (S100, CD68, CD123) and than normal skin. These findings demonstrate the importance of Factor XIIIa positive dermal dendrocytes in antigen presentation at the initial stage of leprosy

Avaliou-se a capacidade dos cimentos endodônticos: Sealapex®, Activ-GP® e AH-Plus® de alterarem o perfil das citocinas nas respostas Th1, Th2 e Th17, após a implantação destes em subcutâneo de camundongos. A quantificação das citocinas IL-2, IL-6, TNF-α, IFN-γ, IL-4, IL-10 e IL-17 foi realizada in vivo, no tecido reacional circundante aos implantes, os quais foram confeccionados a partir de sondas nasogástricas estéreis e apirogênicas de cloreto de polivinila preenchidas com os cimentos, tendo um grupo controle com sondas vazias. Utilizou-se camundongos isogênicos da linhagem C57BL/6 machos de 6/8 semanas de vida sendo que cada um recebeu 2 implantes na região dorsal (lado direito e esquerdo). Após os períodos experimentais de 7, 21 e 63 dias, com os camundongos anestesiados, os implantes foram removidos juntamente com o tecido circundante, e os animais sacrificados em seguida por deslocamento cervical. As amostras de cada grupo foram divididas sendo: duas, contendo implante/tecido, processadas histotecnicamente e as demais com apenas tecido (sem implante) foram homogeneizadas e centrifugadas com solução formada por tampão RIPA e inibidor de protease. O sobrenadante, fruto deste processo, foi coletado e a dosagem das citocinas realizada através do kit CBA mouse-Th1/Th2/Th17 Cytokine Kit (BD Cytometric Bead Array, San Jose, CA, USA) em análise por citometria de fluxo. Os parâmetros avaliados foram a concentração da citocina em função do cimento testado em cada período experimental. Submeteu-se os resultados à análise estatística por meio do teste t com a correção de Welch\'s. Para todos os testes o nível de significância foi de 5%. Com relação à IL-2 houve diferenças estatísticas significantes entre os grupos Activ-GP® e AH-Plus® (p=0,0391). No período de 21 dias foram detectadas diferenças entre o grupo controle e AH-Plus® (p=0,0402) e entre o grupo Sealapex® e AH-Plus® (p=0,0244). O AH-Plus® induziou um maior aumento na IL-6, aos 7 dias em relação ao Activ-GP® (p=0,0286) e aos 21 dias entre o grupo controle (p=0.0402) e Activ-GP® (p=0.0244). Os níveis de TNF-α foram estatisticamente superiores após 7 dias quando comparou-se o grupo AH-Plus® com os demais. Observou-se que no grupo controle aos 7 e 21 dias ocorreram diferenças estatísticas em relação ao Sealapex® e AH-Plus® respectivamente quando avaliadas as concentrações de IFN-γ. Houve também diferenças estatísticas entre o grupo controle e Sealapex® (p=0,0158) no período de 7 dias para a citocina IL-4. Os valores de IL-10 foram estatisticamente superiores para o grupo controle em relação ao Activ-GP® no período de 21 dias (p=0,0471). Com relação a IL-17 no período de 21 dias, observou-se os maiores valores para o grupo controle, seguido pelo Sealapex®, Activ-GP® e AH-Plus®. Foram detectadas diferenças entre os grupos controle e AH-Plus® (p=0,0121), controle e Activ-GP® (p=0,0262) e entre Sealapex® e Activ GP® (p=0,0314). Baseado nesses resultados podê-se concluir que: os cimentos endodônticos são capazes de modular as respostas Th1, Th2 e Th17 através da inibição ou estimulação da liberação das citocinas testadas. O cimento AH-Plus® promoveu as maiores diferenças no perfil de resposta Th1. / It was evaluated the capacity of the following endodontic sealers: Sealapex, Activ-GP and AH-Plus to modify the cytokine profile in Th1, Th2 and Th17 responses, after their implantation in the subcutaneous tissue of mice. Quantification of IL-2, IL-6, TNF-α, IFN-γ, IL-4, IL-10 and IL-17 was performed in vivo, in the reactional tissue surrounding the implants, which were made from sterile nasogastric probes and apyrogenic of polyvinyl chloride filled with sealer, and a control group of empty probes. It was used isogenic mice of C57BL/6 lineage, 6/8 weeks old males, each of which received two implants in the dorsal region (left and right). After the experimental time of 7, 21 and 63 days, the mice were anesthetized and the implants were removed along with the surrounding tissue, the animals were then sacrificed by cervical dislocation. Samples from each group were divided as follows: two containing implant / tissue processed histologically and with only the remaining tissue (without implant) were mixed and centrifuged with a solution formed by RIPA buffer and protease inhibitors. The supernatant result of this process was collected and cytokine dosage accomplished by mouse-Th1/Th2/Th17 Cytokine CBA Kit Kit (BD cytometric Bead Array, San Jose, CA, USA) for flow cytometry analysis. The evaluated parameters were the cytokine concentration in function of sealer tested in each trial. The results were submitted to statistical analysis using the t test with Welch\'s correction. For all tests the significance level was 5%. With respect to IL-2 there were significant statistical differences between groups-Activ GP and AH-Plus (p=0.0391). In the period of 21 days differences were found between the control group and AH-Plus (p=0.0402) and between the group Sealapex and AH-Plus (p=0.0244). The AH-Plus induced a greater increase in IL-6, at 7 days compared to Activ-GP (p=0.0286) and at 21 days between the control group (p=0.0402) and Activ-GP (p=0.0244). The levels of TNF-α were significantly higher after 7 days when the AH-Plus group was compared with others. It was observed that in the control group at 7 and 21 days there were statistical differences in relation to Sealapex and AH-Plus respectively when evaluated concentrations of IFN-γ. There were also significant differences between the control group and Sealapex (p=0.0158) within 7 days for the cytokine IL-4. The amounts of IL-10 were statistically higher in the control group compared to the Activ GP in a period of 21 days (p=0.0471). With respect to IL-17 in a period of 21 days, it was observed the highest values for the control group, followed by Sealapex, Activ-GP and AH-Plus. Differences were found between the control groups and AH-Plus (p=0.0121), control and Activ-GP (p=0.0262) and between Sealapex and Activ-GP (p=0.0314). Based on the presented results theendodontic sealers are able to promote changes in the response cytokine profile Th1, Th2 and Th17; Sealer AH-Plus produced the greatest changes, in the Th1 response profile.

Avaliou-se a capacidade dos cimentos endodônticos: Sealapex®, Activ-GP® e AH-Plus® de alterarem o perfil das citocinas nas respostas Th1, Th2 e Th17, após a implantação destes em subcutâneo de camundongos. A quantificação das citocinas IL-2, IL-6, TNF-α, IFN-γ, IL-4, IL-10 e IL-17 foi realizada in vivo, no tecido reacional circundante aos implantes, os quais foram confeccionados a partir de sondas nasogástricas estéreis e apirogênicas de cloreto de polivinila preenchidas com os cimentos, tendo um grupo controle com sondas vazias. Utilizou-se camundongos isogênicos da linhagem C57BL/6 machos de 6/8 semanas de vida sendo que cada um recebeu 2 implantes na região dorsal (lado direito e esquerdo). Após os períodos experimentais de 7, 21 e 63 dias, com os camundongos anestesiados, os implantes foram removidos juntamente com o tecido circundante, e os animais sacrificados em seguida por deslocamento cervical. As amostras de cada grupo foram divididas sendo: duas, contendo implante/tecido, processadas histotecnicamente e as demais com apenas tecido (sem implante) foram homogeneizadas e centrifugadas com solução formada por tampão RIPA e inibidor de protease. O sobrenadante, fruto deste processo, foi coletado e a dosagem das citocinas realizada através do kit CBA mouse-Th1/Th2/Th17 Cytokine Kit (BD Cytometric Bead Array, San Jose, CA, USA) em análise por citometria de fluxo. Os parâmetros avaliados foram a concentração da citocina em função do cimento testado em cada período experimental. Submeteu-se os resultados à análise estatística por meio do teste t com a correção de Welch\'s. Para todos os testes o nível de significância foi de 5%. Com relação à IL-2 houve diferenças estatísticas significantes entre os grupos Activ-GP® e AH-Plus® (p=0,0391). No período de 21 dias foram detectadas diferenças entre o grupo controle e AH-Plus® (p=0,0402) e entre o grupo Sealapex® e AH-Plus® (p=0,0244). O AH-Plus® induziou um maior aumento na IL-6, aos 7 dias em relação ao Activ-GP® (p=0,0286) e aos 21 dias entre o grupo controle (p=0.0402) e Activ-GP® (p=0.0244). Os níveis de TNF-α foram estatisticamente superiores após 7 dias quando comparou-se o grupo AH-Plus® com os demais. Observou-se que no grupo controle aos 7 e 21 dias ocorreram diferenças estatísticas em relação ao Sealapex® e AH-Plus® respectivamente quando avaliadas as concentrações de IFN-γ. Houve também diferenças estatísticas entre o grupo controle e Sealapex® (p=0,0158) no período de 7 dias para a citocina IL-4. Os valores de IL-10 foram estatisticamente superiores para o grupo controle em relação ao Activ-GP® no período de 21 dias (p=0,0471). Com relação a IL-17 no período de 21 dias, observou-se os maiores valores para o grupo controle, seguido pelo Sealapex®, Activ-GP® e AH-Plus®. Foram detectadas diferenças entre os grupos controle e AH-Plus® (p=0,0121), controle e Activ-GP® (p=0,0262) e entre Sealapex® e Activ GP® (p=0,0314). Baseado nesses resultados podê-se concluir que: os cimentos endodônticos são capazes de modular as respostas Th1, Th2 e Th17 através da inibição ou estimulação da liberação das citocinas testadas. O cimento AH-Plus® promoveu as maiores diferenças no perfil de resposta Th1. / It was evaluated the capacity of the following endodontic sealers: Sealapex, Activ-GP and AH-Plus to modify the cytokine profile in Th1, Th2 and Th17 responses, after their implantation in the subcutaneous tissue of mice. Quantification of IL-2, IL-6, TNF-α, IFN-γ, IL-4, IL-10 and IL-17 was performed in vivo, in the reactional tissue surrounding the implants, which were made from sterile nasogastric probes and apyrogenic of polyvinyl chloride filled with sealer, and a control group of empty probes. It was used isogenic mice of C57BL/6 lineage, 6/8 weeks old males, each of which received two implants in the dorsal region (left and right). After the experimental time of 7, 21 and 63 days, the mice were anesthetized and the implants were removed along with the surrounding tissue, the animals were then sacrificed by cervical dislocation. Samples from each group were divided as follows: two containing implant / tissue processed histologically and with only the remaining tissue (without implant) were mixed and centrifuged with a solution formed by RIPA buffer and protease inhibitors. The supernatant result of this process was collected and cytokine dosage accomplished by mouse-Th1/Th2/Th17 Cytokine CBA Kit Kit (BD cytometric Bead Array, San Jose, CA, USA) for flow cytometry analysis. The evaluated parameters were the cytokine concentration in function of sealer tested in each trial. The results were submitted to statistical analysis using the t test with Welch\'s correction. For all tests the significance level was 5%. With respect to IL-2 there were significant statistical differences between groups-Activ GP and AH-Plus (p=0.0391). In the period of 21 days differences were found between the control group and AH-Plus (p=0.0402) and between the group Sealapex and AH-Plus (p=0.0244). The AH-Plus induced a greater increase in IL-6, at 7 days compared to Activ-GP (p=0.0286) and at 21 days between the control group (p=0.0402) and Activ-GP (p=0.0244). The levels of TNF-α were significantly higher after 7 days when the AH-Plus group was compared with others. It was observed that in the control group at 7 and 21 days there were statistical differences in relation to Sealapex and AH-Plus respectively when evaluated concentrations of IFN-γ. There were also significant differences between the control group and Sealapex (p=0.0158) within 7 days for the cytokine IL-4. The amounts of IL-10 were statistically higher in the control group compared to the Activ GP in a period of 21 days (p=0.0471). With respect to IL-17 in a period of 21 days, it was observed the highest values for the control group, followed by Sealapex, Activ-GP and AH-Plus. Differences were found between the control groups and AH-Plus (p=0.0121), control and Activ-GP (p=0.0262) and between Sealapex and Activ-GP (p=0.0314). Based on the presented results theendodontic sealers are able to promote changes in the response cytokine profile Th1, Th2 and Th17; Sealer AH-Plus produced the greatest changes, in the Th1 response profile.

A forma indeterminada é a fase inicial da hanseníase, que se caracteriza histologicamente pelo infiltrado inflamatório leve, não granulomatoso, de linfócitos e histiócitos ao redor de vasos, anexos e nervos. No local de entrada do M. leprae, as células apresentadoras de antígeno do tipo células dendríticas são as primeiras a encontrar o bacilo. Este, no interior de células dendríticas, desencadeia a produção local de citocinas e quimiocinas, que resultam em proliferação de linfócitos T helper 1 ou T helper 2, assim definindo uma resposta imune celular ou humoral, respectivamente. As lesões tuberculoides mostram predominância das citocinas de padrão Th1 como IL-2, TNF-alfa, IFN-y, IL-12 e IL-18, enquanto que nas lesões virchowianas predominam citocinas de padrão Th2, como IL-4, IL-5, IL-10 e TGF-beta. Na pele, as principais células dendríticas são células dendríticas mieloides, células de Langerhans e alguns dendrócitos dérmicos. São identificadas respectivamente pela expressão imuno-histoquímica de S100, CD1a e Fator XIIIa. Células de Langerhans e dendrócitos dérmicos Fator XIIIa positivos estão aumentados em quantidade nas lesões tuberculoides quando comparadas com lesões virchowianas. Os objetivos do presente estudo foram: 1) caracterizar a inflamação \"in situ\" na hanseníase indeterminada através da quantificação das marcações imuno-histoquímicas de: CD57, CD4, CD8, CD1a, S100, FXIIIa, CD68, Foxp3, CD123, IL-1, IL-2r, IL-4, IL-6, IL-8, IL-10, IL-12, IL-18, IFN-, TNF-alfa, TGF-beta, iNOS, granzima B, receptor Toll-like 2/4, e antígeno BCG, 2) comparar o perfil fenotípico e citocínico das lesões na hanseníase indeterminada entre grupos de reação de Mitsuda positiva e negativa, a fim de investigar se existem padrões que possam prever para qual forma a doença evoluiria, e 3) revisar a histopatologia da forma indeterminada através da análise semiquantitativa das alterações vistas à coloração de hematoxilina/eosina. Foram selecionadas 15 lesões de pacientes com hanseníase indeterminada. Foram usados grupos controles de expressão de Fator XIIIa e CD1a em 10 casos de pele normal. A histopatologia mostrou discretas alterações epidérmicas, como alteração vacuolar e exocitose de linfócitos (33% dos casos cada), apoptose de queratinócitos (26%), atrofia e acantose (06% dos casos cada); infiltrado inflamatório linfomononuclear neural (100%), perivascular superficial (100%), perivascular profundo (93%), peri-écrino (40%) e peri-folículo pilossebáceo (20%), além de melanófagos em 93% dos casos. Esses achados mostraram que nem sempre todos os ambientes estão acometidos por inflamação na histopatologia. Em 66% dos pacientes foi encontrado antígeno bacilar (por Fite-Faraco ou técnica imuno-histoquímica anti-BCG), portanto a forma indeterminada não deve ser considerada sistematicamente como paucibacilar. Não houve diferença significativa de padrões de marcadores entre os grupos Mitsuda positivo e negativo. No microambiente inflamatório das lesões houve expressões significativas de TLR 2/4, Fator XIIIa, CD4, CD8, IL-2r, IL-4, IL-10, iNOS e TGF-beta. A expressão importante de IL-4, IL-10 e TGF- beta nas lesões de hanseníase indeterminada significaram tendência de resposta imune para o polo Th2, um ambiente de tolerância à permanência do bacilo. A baixa expressão de IFN-y colaborou para a inexpresiva resposta Th1. Não houve diferença significativa na expressão de CD1a entre as lesões e pele normal. Fator XIIIa foi expresso em mais que 50 células/mm2 em todos os casos, com quantidades significativamente maiores que outras células dendríticas nas lesões (S100, CD68, CD123) e que a pele normal. Estes achados demonstraram a importância dos dendrócitos dérmicos Fator XIIIa positivos na apresentação de antígeno na fase inicial da hanseníase / The indeterminate form is the initial stage of leprosy, which is characterized histologically by mild inflammatory infiltrate, non granulomatous, with lymphocytes and histiocytes around vessels, nerves and adnexals. When M. leprae enter the host, antigen-presenting cells of dendritic type are the first cells to find the bacillus. Once inside dendritic cells, the bacillus elicits local production of cytokines and chemokines, which result in proliferation of T lymphocytes helper 1 or T helper 2, thereby defining a cellular or humoral immune response, respectively. The tuberculoid lesions show predominance of Th1 cytokines such as IL-2, TNF-alfa, IFN-y, IL-12 and IL-18, whereas in the lepromatous lesions predominate cytokines of Th2 pattern such as IL-4, IL-5 IL-10 and TGF-beta. In the skin, main dendritic cells are myeloid dendritic cells, Langerhans cells, and some dermal dendrocytes. They are identified respectively by immunohistochemical expression of S100, CD1a and Factor XIIIa. Langerhans cells and dermal dendrocytes Factor XIIIa positive are increased in number in tuberculoid lesions compared with lepromatous lesions. The objectives of this study were: 1) to characterize \"in situ\" inflammation in indeterminate leprosy through the quantification of immunohistochemical markers: CD57, CD4, CD8, CD1a, S100, FXIIIa, CD68, Foxp3, CD123, IL-1, IL-2r, IL-4, IL-6, IL-8, IL-10, IL-12, IL-18, IFN-y, TNF-alfa, TGF-beta, iNOS, granzyme B, Toll-like receptor 2/4, and BCG antigen, 2) compare the phenotypic and cytokinic profile of indeterminate leprosy lesions between positive and negative Mitsuda reaction groups in order to investigate if there are patterns that can predict which way the disease may evolve, and 3 ) review the histopathology of the indetermate form by semi-quantitative analysis of changes seen in hematoxylin / eosin. Fifteen lesions of patients with indeterminate leprosy were selected. There was used control groups of Factor XIIIa and CD1a expression in 10 cases of normal skin. Histopathology showed discrete epidermal changes, such as vacuolar changes and lymphocyte exocytosis (in 33% of cases each), keratinocyte apoptosis (26%), atrophy and acanthosis (in 06% of cases each); neural lymphocytic inflammatory infiltrate (100%), superficial perivascular (100%), deep perivascular (93%), peri-eccrine (40%), peri-pilosebaceous follicle (20%), and melanophages in 93% of cases. These findings showed that not always all environments are affected by inflammation in histopathology. In 66% of patients it was found bacterial antigen (by Fite-Faraco or immunohistochemical technique anti-BCG), so the indeterminate form should not be systematically considered as paucibacillary. There was no significant difference in phenotypic and cytokinic patterns between the positive and negative Mitsuda groups. In the microenvironment of inflammatory lesions there was significant expression of TLR 2/4, Factor XIIIa, CD4, CD8, IL-2r, IL-4, IL-10, TGF-beta and iNOS. The important expression of IL-4, IL-10 and TGF-beta in indeterminate leprosy meant tendency to Th2 immune response pole, an environment of tolerance to permanence of bacillus. Low IFN-? expression contributed to the negligible Th1 response. There was no significant difference in the expression of CD1a between the lesions and normal skin. Factor XIIIa was expressed as greater than 50 cells / mm2 in all cases, with significantly larger quantities than other dendritic cells in lesions (S100, CD68, CD123) and than normal skin. These findings demonstrate the importance of Factor XIIIa positive dermal dendrocytes in antigen presentation at the initial stage of leprosy

As relações bidirecionais entre o Sistema Nervoso e o Sistema Imune são relevantes para a manutenção da homeostase do organismo. Estudos realizados em nosso laboratório mostraram que 14 dias de coabitação com um conspecífico doente (injetado com células do tumor de Ehrlich-TAE) produziu mudanças comportamentais, endócrinas e imunológicas. Este estudo analisa os efeitos da convivência com um animal portador de tumor de Ehrlich em camundongos OVA sensibilizados e desafiados sobre a resposta alérgica pulmonar. Pares de camundongos machos foram separados em três grupos: naïve, controle e experimental. Os animais do grupo naïve não foram manipulados sendo utilizados para a avaliação de parâmetros basais. Um animal de cada par dos grupos experimental e controle foi imunizado com OVA. No dia D(0), os animais imunizados receberam uma dose reforço de OVA. No dia D(0) os camundongos do grupo experimental que não foram manipulados foram inoculados com 5x106 células de tumor de Ehrlich; seus companheiros de gaiola moradia foram designados CAD (companheiro do animal doente). Os camundongos não perturbados de cada par do grupo controle foram tratados (i.p.) em D(0) com 0,9% de NaCl, sendo designados CAS (companheiro do animal saudável). O desafio intranasal com OVA foi realizado nos camundongos CAS e CAD nos dias D(12) e D(13); colheram-se o sangue e os tecidos no dia D(14). Em comparação com o grupo CAS, os camundongos do grupo CAD apresentaram 14 dias após a coabitação: (1) aumento do número de eosinófilos e neutrófilos no LBA, (2) diminuição na contagem de células da medula óssea, (3) aumento do níveis de IL-4 e IL-5 e diminuição de IL-10 e INF-ϒ no sobrenadante do LBA, (4) aumento dos níveis de IgG1-OVA, diminuição dos níveis de IgG2a-OVA e nenhuma alteração na IgE-OVA no sangue periférico, (5) aumento na expressão de ICAM-1, VCAM-1 e L-selectina em granulócitos do LBA, (6) diminuição da reatividade da traquéia à metacolina in vitro, (7) aumento da desgranulação de mastócitos, (8) nenhuma alteração nos níveis plasmáticos de corticosterona, (9) aumento dos níveis de adrenalina e noradrenalina plasmáticas, (10) diminuição no tempo de permanência e entradas nos braços abertos do labirinto em cruz elevado, (11) diminuição da expressão de IL-6 no PVN e (12) diminuição da expressão de C-fos no PFC. Estes resultados mostram que a convivência forçada com um animal portador de um tumor ascitico de Ehrlich exacerba a inflamação alérgica pulmonar de camundongos. Eles foram discutidos como decorrentes da estimulação do Sistema Nervoso Autônomo Simpático (SNS) pelo estresse psicológico gerado pela coabitação com o parceiro doente, via liberação de adrenalina e noradrenalina e consequente mudança no perfil de citocinas Th1/Th2 para uma resposta do tipo Th2. Esta alteração seria, provavelmente, um dos mecanismos responsáveis pelo aumento do recrutamento celular para as vias aéreas dos camundongos do grupo CAD. / The bidirectional relationship between the nervous system and the imune system is relevant for homeostatic organism maintenance. Studies from our laboratory showed that 14 days of cohabitation with a sick conspecific (injected with Ehrlich tumor cells-TAE) produced behavioral, endocrinological and immunological changes. This study analyzes the effects of cohabitation with an Ehrlich tumor-bearing animal on ovalbumin (OVA)-induced lung inflammatory response in mice. Pairs of male mice were separate into three groups: naïve, control and experimental. Animals of the naïve group were kept undisturbed being used for assessment of basal parameters. One animal of each experimental and control pair of mice was immunized with OVA. On D(0), these OVA-immunized animals received an OVA booster. At this day (D(0)) the experimental mice that were kept undisturbed were inoculated with 5x106 Ehrlich tumor cells; their immunized cage-mates were then referred as to CSP(companion of sick partner). The undisturbed mice of each control pair were i.p. treated on D(0) with 0.9% NaCl; their sensitized cage-mate were subsequently referred as CHP (companion of health partner). The intranasal OVA challenge was performed on CSP and CHP mice on D(12) and D(13); blood and tissue collection were performed on D (14). Fourteen days after cohabitation, in comparison to the CHP mice, the CSP mice displayed the following: (1) an increased number of eosinophils and neutrophils in the BAL, (2) a decreased bone marrow cell count, (3) increased levels of IL-4 and IL-5 and decreased levels of IL-10 and INF-ϒ in the BAL supernatant, (4) increased levels of IgG1-OVA, decreased levels of IgG2a-OVA and no changes in OVA-specific IgE in the peripheral blood, (5) increased expression of ICAM-1, VCAM-1 and L-selectin in the BAL granulocytes, (6) decreased tracheal reactivity to metacholine measured in vitro , (7) increased mast cell degranulation, (8) no changes in plasma corticosterone levels (9) increased levels of plasmatic adrenaline and noradrenaline, (10) decreased time and % of entries on open arms of elevated plus maze, (11) decreased expression of IL-6 on PVN and (12) decreased expression of C-fos on PFC. These results suggest that cohabitation with an Ehrlich tumor bearing mice exacerbates allergic lung inflammatory response in mice. Most probably, the changes observed in CSP mice are a consequence of the psychological stress induced by forced cohabitation with the sick partner. Strong involvement of the sympathetic nervous system through adrenaline and noradrenaline release and a shift of the Th1/Th2 cytokine profile toward a Th2 response were considered to be the mechanisms underlying the cell recruitment to the animal´s airways.

L’asthme est une maladie multifactorielle hétérogène qui engendre une inflammation pulmonaire associée à une variété de manifestations cliniques, dont des difficultés respiratoires graves. Globalement, l’asthme touche environ une personne sur 6 et présente actuellement un sérieux problème de santé publique. Bien que de nombreux traitements soient disponibles pour soulager les symptômes de la maladie, aucun traitement curatif n’est actuellement disponible. La compréhension des mécanismes qui régissent l’état inflammatoire au cours de la maladie est primordiale à la découverte de nouvelles cibles thérapeutiques efficaces. Les cellules dendritiques captent les allergènes dans les poumons et migrent vers les ganglions drainants pour les présenter aux cellules T et engendrer la réponse inflammatoire pathogénique chez les asthmatiques. Nous avons contribué à l’avancement des connaissances mécanistiques de l’asthme en identifiant chez la souris la sous-population de cellules dendritiques responsable de l’initiation et du maintien de la réponse inflammatoire locale et systémique associée à l’asthme. En effet, nous avons démontré que le SIRPα, récepteur extracellulaire impliqué dans la régulation de la réponse immune, est sélectivement exprimé à la surface des cellules dendritiques immunogéniques. L’interruption de la liaison entre le SIRPα et son ligand, le CD47, interfère avec la migration des cellules dendritiques SIRPα+ et renverse la réponse inflammatoire allergique. Ce mécanisme constitue une avenue thérapeutique prometteuse. D’ailleurs, les molécules de fusion CD47-Fc et SIRPα-Fc se sont avérées efficaces pour inhiber l’asthme allergique dans le modèle murin. Nous avons également démontré l’implication des cellules dendritiques SIRPα dans un modèle d’inflammation pulmonaire sévère. L’administration répétée de ces cellules, localement par la voie intra-trachéale et systémiquement par la voie intra-veineuse, mène au développement d’une réponse inflammatoire mixte, de type Th2-Th17, similaire à celle observée chez les patients atteints d’asthme sévère. La présence de cellules T exprimant à la fois l’IL-17, l’IL-4, l’IL-13 et le GATA3 a été mise en évidence pour la première fois in vitro et in vivo dans les poumons et les ganglions médiastinaux grâce à ce modèle. Nos expériences suggèrent que ces cellules Th2-Th17 exploitent la plasticité des cellules T et sont générées à partir de la conversion de cellules Th17 qui acquièrent un phénotype Th2, et non l’inverse. Ces résultats approfondissent la compréhension des mécanismes impliqués dans l’initiation et le maintien de l’asthme allergique et non allergique, en plus d’ouvrir la voie à l’élaboration d’un traitement spécifique pour les patients asthmatiques, particulièrement ceux pour qui aucun traitement efficace n’est actuellement disponible. / Asthma is a heterogeneous multifactorial disease resulting in airway inflammation associated with a variety of clinical manifestations, which include severe breathing difficulties. Asthma affects approximately one out of six people and is currently a serious public health problem. As of now, many treatments are available to relieve the symptoms of the disease, but no definitive cure is available. Understanding the mechanisms that regulate the inflammatory condition during the disease is essential to the discovery of effective new therapeutic targets. Dendritic cells capture allergens in the lungs, migrate to the draining lymph nodes where they activate cognate T cells, which cause the pathogenic inflammatory response. My work help defined and deepened the mechanistic understanding of asthma by identifying the subpopulation of dendritic cells responsible for the initiation and maintenance of local and systemic inflammatory response. We demonstrated that SIRPα is selectively expressed on the surface of immunogenic dendritic cells. Indeed, the interruption of the ligation between SIRPα and its ligand, CD47, interferes with the migration of SIRPα+ dendritic cells and reverses the allergic inflammatory response. This mechanism is a promising new therapeutic avenue. Moreover, we showed that the soluble fusion molecules CD47-Fc and SIRPα-Fc are potent inhibitors of the allergic asthma in a mouse model. In addition, we demonstrated the involvement of SIRPα+ dendritic cells in a model of severe airway inflammation induced upon local and systemic repeated administration of those cells. Either treatment led to the development of a mixed Th2-Th17 inflammation, a phenotype recently described in patients with severe asthma. This model allowed us to show the presence of T cells expressing at once IL-17, IL-4, IL-13 and GATA3 in vitro and in vivo in the lungs and in the mediastinal lymph nodes. Our results suggest that these Th2-Th17 cells are generated from the conversion of Th17 cells acquiring a Th2 phenotype, and not the other way around, a hallmark of Th17 cells plasticity. These results deepen the understanding of the mechanisms involved in the initiation and maintenance of allergic and non-allergic asthma. Besides, we open a way to the development of a specific treatment for asthmatic patients, particularly those for whom no effective treatment is currently available.