House dust mites are a major source of allergens in house dust and, thus, the main trigger of perennial allergic respiratory diseases [1,2,3,4,5]. Scientific research on the life cycle, diet, and reproductive behavior of dust mites and on the biology of mite allergens has uncovered mechanisms leading to the development of respiratory allergies and suggests measures that can minimize exposure to dust mite allergens. Here, we discuss the evidence linking house dust mite exposure and respiratory allergies and present the efficacy of avoidance measures and their scientific evidence.

Submitted by PPG Pediatria e Sa?de da Crian?a (pediatria-pg@pucrs.br) on 2018-02-16T19:15:31Z No. of bitstreams: 1 Tese final 2018 - artigo publicado.pdf: 1647298 bytes, checksum: 03a39af4be2d469330c9618fb70f9cdd (MD5) / Approved for entry into archive by Caroline Xavier (caroline.xavier@pucrs.br) on 2018-02-23T16:28:05Z (GMT) No. of bitstreams: 1 Tese final 2018 - artigo publicado.pdf: 1647298 bytes, checksum: 03a39af4be2d469330c9618fb70f9cdd (MD5) / Made available in DSpace on 2018-02-23T16:35:09Z (GMT). No. of bitstreams: 1 Tese final 2018 - artigo publicado.pdf: 1647298 bytes, checksum: 03a39af4be2d469330c9618fb70f9cdd (MD5) Previous issue date: 2017-07-12 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior - CAPES / Background: Asthma is a chronic disease of the airways, characterized by bronchial inflammation and hyperresponsiveness, which affects approximately 300 million people around the world. The increase in the prevalence of asthma in recent years has been associated with an increase in vitamin D deficiency. About 1 billion people in the world have insufficient levels of vitamin D due to many factors such as reduced outdoor activities, sunscreens use and a diet low in vitamin D. In addition, many studies suggest that vitamin D deficiency has a direct effect on lung function, leading to changes in the structure of the airways and in the inflammatory process. Objectives: To evaluate the effect of vitamin D deficiency at different life stages in a murine model of allergic airways disease house dust mite (HDM) induced on inflammation and lung function. Methods: Female BALB / c mice were placed in a diet replete or deficient in vitamin D at three-week old. At 8 weeks, females were mated with males on a diet replete in vitamin D. At birth, pups were cross-fostered to assess the effects of vitamin D deficiency at different stages of life, in utero (Vit D -/+), postnatal (Vit D +/-) and whole-life (Vit D - / -) compared to the control group whole-life vitamin D replete (Vit D + / +). At 8 weeks of age, mice of both sexes were challenged for 10 consecutive days intranasally with either HDM extract or saline solution after mild anesthesia. The animals were anesthetized for lung function test and then submitted to euthanasia for bronchoalveolar lavage (BAL) and lung tissue removal 24 hours after the last intranasal challenge. The total BAL cell count and collagen quantification of lung tissue homogenized were evaluated. Results: Vitamin D deficiency did not affect HDM-induced inflammation, which was characterized by BAL eosinophilia. Vitamin D deficiency at any life stage (in utero, postnatal and all life) caused impairment of lung function, increased tissue damping and tissue elastance, being particularly observed in females. On the other hand, the asthma HDM-induced decreased airway distensibility, but only in females and vitamin D do not altered this response. Conclusion: Our results suggest that vitamin D and HDM have different mechanisms that influence in the development of allergic lung disease and furthermore the effects appear to be sex-specific. / Introdu??o: A asma ? uma doen?a cr?nica das vias a?reas, caracterizada por inflama??o e hiperresponsividade br?nquica, que atinge aproximadamente 300 milh?es de pessoas ao redor do mundo. O aumento da preval?ncia da asma nos ?ltimos anos tem sido associado ao aumento da defici?ncia de vitamina D. Cerca de 1 bilh?o de pessoas no mundo apresenta n?veis insuficientes de vitamina D em fun??o de diversos fatores, tais como a redu??o de atividades ao ar livre, uso de protetor solar e dieta pobre em vitamina D. Al?m disso, estudos sugerem que a defici?ncia de vitamina D possui um efeito direto na fun??o pulmonar, causando altera??es na estrutura das vias a?reas e no processo inflamat?rio. Objetivo: Avaliar o efeito da defici?ncia de vitamina D em diferentes est?gios da vida de camundongos com asma induzida por ?caro domiciliar (house dust mite; HDM) sobre a inflama??o e fun??o pulmonar. M?todos: Camundongos BALB/c f?meas receberam dieta rica ou deficiente em vitamina D a partir da terceira semana de vida. Com 8 semanas de vida, as f?meas foram acasaladas com machos em dieta rica em vitamina D. Ao nascimento foi realizado o cross-fostering (ado??o cruzada) com a prole para que fosse poss?vel avaliar o efeito da defici?ncia de vitamina D em diferentes est?gios da vida, in utero (Vit D -/+), p?s-natal (Vit D +/-) e durante toda a vida (Vit D -/-), em compara??o ao grupo controle, que recebeu dieta rica em vitamina D durante toda a vida (Vit D +/+). Com 8 semanas de vida, camundongos de ambos os sexos foram desafiados por 10 dias consecutivos por via intranasal, com extrato de HDM ou apenas solu??o salina. Os animais foram anestesiados para a realiza??o do teste de fun??o pulmonar e ent?o submetidos a eutan?sia para a realiza??o do lavado broncoalveolar (LBA) e retirada do tecido pulmonar, 24 horas ap?s o ?ltimo desafio intranasal. Foi avaliada a contagem total de c?lulas do LBA e quantifica??o de col?geno no homogeneizado de tecido pulmonar. Resultados: A defici?ncia de vitamina D n?o afetou a inflama??o induzida por HDM, que foi caracterizada por eosinofilia no LBA. A defici?ncia de vitamina D em qualquer fase da vida dos camundongos (in utero, p?s-natal e durante toda a vida) causou uma piora na fun??o pulmonar, aumentando o tissue damping e tissue elastance, sendo observado particularmente em f?meas. Por outro lado, a asma induzida por HDM diminuiu a distensibilidade das vias a?reas apenas em f?meas e a vitamina D n?o alterou essa resposta. Conclus?o: Nossos resultados sugerem que a vitamina D e HDM possuem diferentes mecanismos que influenciam no desenvolvimento da doen?a pulmonar al?rgica e, al?m disso, os efeitos parecem ser dependentes do sexo.

<p>The overall aims of our studies were to identify risk factors for respiratory symptoms and asthma in indoor environment but even to look at some personal risk factors such as body mass index and gastroesophageal reflux. The study population is based on participants of the European Community Respiratory Health Survey I and II. </p><p> In the first study, water damage and visible moulds were reported in 7.4% and 17% of the homes respectively. The combination of water damage and visible moulds was independently associated with attacks of breathlessness when resting and after activity and also to long term cough. In the second study, the prevalence of nocturnal GER increased with higher BMI and the same pattern could be seen for habitual snoring. Reported onset of asthma, wheeze and night-time symptoms increased in prevalence along with the BMI gradient. In the multivariable analysis, obesity and nocturnal GER were independent risk factors for onset of asthma, wheeze and night-time symptoms. Habitual snoring was an independent risk factor for onset of wheeze and night-time symptoms, but not for onset of asthma. In the third study, a total of 18% of the subjects reported indoor dampness in the last 12 months and 27% of the subjects reported indoor dampness since the previous survey. Respiratory symptoms and asthma were significantly more prevalent in individuals exposed to indoor dampness and indoor dampness was a risk factor for respiratory symptoms and asthma after adjusting for possible confounders. Indoor dampness was an independent risk factor for onset of respiratory symptoms but not for asthma onset. Remission of respiratory symptoms was less likely to occur if subjects reported indoor dampness. In the fourth study, the lowest prevalence of atopy and the lowest levels of all indoor allergens, bacteria and moulds were found in Iceland. A positive association was found, between cat allergen exposure and asthma symptoms and between bronchial hyperresponsiveness and the amount of viable mould in indoor air.</p>

Les effets anti-allergiques des bactéries lactiques sont suggérés par plusieurs études épidémiologiques, des essais cliniques et des modèles expérimentaux d’allergie. Cependant, les propriétés immunomodulatrices des bactéries lactiques sont sous-exploitées par les stratégies vaccinales développées pour combattre l’allergie et les mécanismes empruntés par ces bactéries pour moduler l’allergie restent peu caractérisés. Dès lors, nous avons caractérisé les propriétés immunomodulatrices qu’exerce Lactobacillus plantarum NCIMB8826, une bactérie lactique modèle, sur la cellule dendritique étant donné le rôle déterminant de cette cellule sur la réponse allergique. Nous montrons que L. plantarum induit une forte sécrétion d’IL-12 p40, d’IL-12 p70, de TNF-a mais une faible production d’IL-10. Cette faculté à induire la sécrétion de cytokines polarisantes dépend de TLR2, de TLR9, de MyD88, de NF-kB, des MAPKs (en particulier JNK, p38 et ERK 1/2), de la composition de l’acide lipotéichoïque de L. plantarum et de CD14. Nous montrons aussi que l’ADN génomique de L. plantarum est un agoniste de TLR9 et que CD14 et CD36 facilitent la liaison de la cellule dendritique avec L. plantarum. Ensuite, nous avons évalué le potentiel vaccinal d’une coadministration L. plantarum + Der p 1 dans un modèle murin d’allergie à Der p 1. Cette formulation vaccinale prévient la production d’IgE Der p 1-spécifique et atténue l’éosinophilie pulmonaire tout en stimulant une forte production d’anticorps IgG2a Der p 1-spécifiques et d’IFN-g par les cellules spléniques. Ces effets bénéfiques nous ont conduit à élaborer une bactérie lactique recombinante dérivée de L. plantarum produisant Der p 1 pour la vaccination contre l’allergie aux acariens. La forme antigénique que nous avons réussi à faire produire par L. plantarum correspond à une protéine de fusion entre la Maltose Binding Protein de E. coli et ProDer p 1 (le zymogène de Der p 1), la présence de ce partenaire de fusion étant indispensable à la production de ProDer p 1. En prophylaxie, la vaccination par cette bactérie recombinante prévient la production d’anticorps IgE-Der p 1-spécifiques et stimule la production d’anticorps IgG2a spécifiques, reproduisant les effets de la coadministration L. plantarum + Der p 1. Elle réduit de manière drastique la production d’IL-5 des cellules spléniques et des cellules ganglionnaires médiastinales et prévient l’éosinophilie pulmonaire mais n’a pas d’effet sur l’hyperréactivité bronchique. Der p 1 étant un des allergènes d’acarien les plus immunodominants, cet ensemble de données montre donc que cette bactérie recombinante constitue un vaccin prophylactique prometteur pour la prévention de l’allergie aux acariens. Des résultats préliminaires obtenus à partir de cellules dendritiques humaines et lymphocytes T autologues montrent la forte capacité de cette bactérie recombinante à induire le développement d’une réponse Th1 fortement polarisée (production d’IFN-g en l’absence de production d’IL-4 et d’IL-5), ce qui suggère que l’utilisation de cette bactérie recombinante pourrait être envisagée pour le traitement de l’allergie chez l’homme.

The major allergen Der p 1 of the house dust mite Dermatophagoides pteronyssinus is a papain-like cysteine protease (CA1) associated to the development of allergic diseases such as asthma, rhinitis or atopic dermatitis. This allergen is expressed as an inactive precursor, called proDer p 1, formed by a 25 kDa catalytic domain downstream to an 10 kDa N-terminal propeptide, which blocks the active site cleft. The propeptide of Der p 1 exhibits a specific fold, which makes it unique in the CA1 propeptide family as it is characterised by the presence of four alpha helices and the absence of ERFNIN motif. In this study, we investigated the activation steps involved in the maturation of recombinant proDer p 1 expressed in Pichia pastoris under acidic conditions and we studied the influence of acidic pH on the structure of both propeptide and catalytic domain. Therefore, we characterized the interaction between the propeptide and mature Der p 1 at different pH values in terms of activity inhibition, structural stability and proteolytic susceptibility. According to our results, the auto-activation of proDer p 1 is a multistep mechanism, characterized by at least two intermediates (ATFE- and SNGG-) corresponding to the loss of the first and second propeptide alpha helices, respectively. The propeptide strongly inhibits unglycosylated and glycosylated recombinant Der p 1 (KD= 7 nM) at neutral pH. This inhibition is pH dependent, decreasing from pH 7 to pH 4 and can be related to structural changes of the propeptide initiated by the protonation of the aspartate residue of Lys17-Asp51-Tyr19 structural triad presents within the propeptide N-terminal domain. This protonation triggers conformational changes of the first propeptide alpha helix leading to an increase of the propeptide flexibility, an increase of its proteolytic sensitivity and the formation of a molten globule state. In addition, we compare mature protease, zymogen and propeptide pH unfolding and stability and highlights that the presence of the propeptide does not influence the catalytic domain pH unfolding and stability as the propeptide displays a weaker pH stability than the protease domain. These results confirmed that the propeptide unfolding is the key event of the activation process. Finally, we unravel the intermolecular contribution of mature Der p 1 in the activation process and highlights that activation of the precursor can be achieved, under acidic conditions, by intermolecular process but initial auto-activation most probably occurs through an intramolecular process or by the proteolysis by the catalytic domain of another zymogen in which the propeptide is unfolded. According to our results, we proposed that activation of the zymogen at pH 4 reflects a compromise between activity preservation and propeptide unfolding and that the location of the activation sites on the propeptide structure is a compromise between sequence recognition specificity and proteolytic susceptibility of the corresponding area.

The overall aims of our studies were to identify risk factors for respiratory symptoms and asthma in indoor environment but even to look at some personal risk factors such as body mass index and gastroesophageal reflux. The study population is based on participants of the European Community Respiratory Health Survey I and II. In the first study, water damage and visible moulds were reported in 7.4% and 17% of the homes respectively. The combination of water damage and visible moulds was independently associated with attacks of breathlessness when resting and after activity and also to long term cough. In the second study, the prevalence of nocturnal GER increased with higher BMI and the same pattern could be seen for habitual snoring. Reported onset of asthma, wheeze and night-time symptoms increased in prevalence along with the BMI gradient. In the multivariable analysis, obesity and nocturnal GER were independent risk factors for onset of asthma, wheeze and night-time symptoms. Habitual snoring was an independent risk factor for onset of wheeze and night-time symptoms, but not for onset of asthma. In the third study, a total of 18% of the subjects reported indoor dampness in the last 12 months and 27% of the subjects reported indoor dampness since the previous survey. Respiratory symptoms and asthma were significantly more prevalent in individuals exposed to indoor dampness and indoor dampness was a risk factor for respiratory symptoms and asthma after adjusting for possible confounders. Indoor dampness was an independent risk factor for onset of respiratory symptoms but not for asthma onset. Remission of respiratory symptoms was less likely to occur if subjects reported indoor dampness. In the fourth study, the lowest prevalence of atopy and the lowest levels of all indoor allergens, bacteria and moulds were found in Iceland. A positive association was found, between cat allergen exposure and asthma symptoms and between bronchial hyperresponsiveness and the amount of viable mould in indoor air.

Submitted by Lara Oliveira (lara@ufersa.edu.br) on 2017-05-19T20:57:26Z No. of bitstreams: 1 Ismênia_DISSERT.pdf: 2286785 bytes, checksum: fe512f9cb09ab8ccaeca35cce30794cc (MD5) / Made available in DSpace on 2017-05-19T20:57:26Z (GMT). No. of bitstreams: 1 Ismênia_DISSERT.pdf: 2286785 bytes, checksum: fe512f9cb09ab8ccaeca35cce30794cc (MD5) Previous issue date: 2017-02-17 / Leprosy is a chronic infectious disease caused by Mycobacterium leprae and persists as a serious public health problem in Brazil. The fact that this microorganism is inculcable makes it difficult to diagnose and elucidate details of its transmissive chain. Aiming to clarify the role of house dust in the disease transmission route, this case-control study carried out in the Barrocas, Mossoró, RN, Brazil, the largest cluster in the state, compared clinical, epidemiological, slit skin smear and serological results of 22 newly diagnosed patients of the cluster, with molecular results of detection of the specific RLEP and 16S rRNA genomic regions of this bacillus in the nasal swab samples, saliva and house dust of these individuals and of the controls (44 household contacts and 44 endemic contacts) between november 2015 to november 2016. There was greater detection in women and school-age youth, with degree of disability, with predominance of paucibacillary forms. There was no statistical difference between the number of BCG vaccine scars from patients and endemic contacts, and 31.8% of the patients had no vaccine scar. The two rapid serological tests evaluated, ML-flow and OrangeLife® presented similar results, with a higher positivity among paucibacillary patients through the latter (54.5%). Both were superior to slit skin smear (9.1%). Regarding molecular research, the positivity in nasal swab and saliva of multibacillary patients with primer RLEP was 16.7% and 33.3%, respectively. There was no detection of bacterial DNA in domestic dust or between paucibacillary. Regarding the DNA analysis in saliva of endemic multibacillary contacts, there was 27.2% positivity. Associating molecular and serological results, it was possible to identify 4.5% of subclinicity between home contacts and 15% of asymptomatic carrier status. Regarding the risk variables, residing in dwellings with up to two windows offered 3.79 times more chance of progressing to this outcome, having a history of cases of leprosy in the family increased 2.89 times the risk of presenting seropositivity by the OrangeLife® serological test In this community and being over 60 years of age gave 3.6 times more chances of having positive serological reaction for leprosy with the test evaluated, which denotes prolonged exposure to the bacillus. Although not statistically significant, it was noted that the pattern of low schooling, low income, environmental insalubrity, population agglomeration and inadequate living habits made up a sociodemographic and epidemiological profile that attests to the context of social vulnerability of the affected population, requiring integrated actions of rehabilitation that go beyond the physical aspect of their dwellings, since they need to contemplate the human and social development of these people in the broad sense / A hanseníase é uma doença infecciosa crônica causada pelo Mycobacterium leprae e persiste como grave problema de saúde pública no Brasil. O fato deste micro-organismo ser incultivável dificulta o diagnóstico e elucidação de detalhes da sua cadeia transmissiva. Objetivando analisar a dinâmica de transmissão ambiental da hanseníase, este estudo caso-controle realizado no Bairro Barrocas, Mossoró/RN, maior cluster do estado, confrontou resultados de avaliação clínica, epidemiológica, baciloscópica e sorológica de 22 doentes recém-diagnosticados nas Barrocas, com resultados moleculares de detecção das regiões genômicas específicas RLEP e 16S rRNA deste bacilo nas amostras de swab nasal, saliva e poeira domiciliar destes indivíduos e dos seus controles (44 contactantes domiciliares e 44 contactantes endêmicos), entre novembro de 2015 a novembro de 2016. Os indivíduos foram diagnosticados através de busca ativa nos domicílios, nas escolas e na sede da unidade básica de saúde local. Houve maior detecção da doença em mulheres, nos menores de 15 anos, com grau 0 de incapacidade e predominância de formas paucibacilares. Não houve diferença estatística entre o número de cicatrizes vacinais de BCG de doentes e de contatos endêmicos, e 31,8% dos hansenianos não apresentaram cicatriz vacinal. Os dois testes rápidos sorológicos avaliados, ML-flow (IgM ND-O-BSA) e OrangeLife® (IgM e IgG anti NDO-LID 1) apresentaram resultados semelhantes, com maior positividade entre os paucibacilares através deste último (54,5%). Ambos foram superiores à baciloscopia (9,1%). Quanto à pesquisa molecular, a positividade em swab nasal e saliva de doentes multibacilares com primer RLEP foi de 16,7% e 33,3%, respectivamente. Não houve detecção de DNA bacteriano na poeira doméstica nem entre os paucibacilares. Já em relação à pesquisa de DNA em saliva de contatos endêmicos de multibacilares, houve 27,2% de positividade. Associando-se resultados moleculares e sorológicos, foi possível identificar 4,5% de subclinicidade e 15% de status portador assintomático entre contatos domiciliares. Em relação às variáveis de risco de soropositivar com o teste OrangeLife®, este estudo revelou que residir em moradias com até duas janelas ofereceu 3,79 vezes mais chance de evoluir para este desfecho, ter histórico de casos de hanseníase na família aumentou 2,89 vezes o risco e ter mais de 60 anos de idade conferiu 3,6 vezes mais chances, o que denota exposição prolongada destes indivíduos ao bacilo. Embora sem significância estatística, fez-se notório que o padrão de baixa escolaridade, baixa renda, insalubridade ambiental, aglomeração populacional e hábitos de vida inadequados compuseram um perfil sociodemográfico e epidemiológico que atestou o contexto de vulnerabilidade social da população afetada, exigindo ações integradas de reabilitação que vão além do aspecto físico de suas moradias, pois necessitam contemplar o desenvolvimento humano e social dessas pessoas no seu sentido amplo / 2017-05-18

Canine atopic dermatitis (CAD) is a common dermatosis, defined as a genetic-based disease, which predisposes to cutaneous inflammation and pruritus, mediated by class IgE immunoglobulins directed against specific antigens in most cases. Clinical diagnosis may be later complemented by skin allergic and/or serological tests (ELISA). The aim of these tests is to identify possible allergens in order to enable the clinicians to select candidate antigens for allergen specific immunotherapy. This work aimed to identify the sensitization profile of 58 dogs with atopic dermatitis diagnosis. All animals were submitted to intradermic test (IDT) and screened for the presence of antibodies against different allergens using a serologic test. House dust mites are described as the most frequent allergens in all continents. However, the positivity to C. dactylon is not commonly described and may be characteristic for the region. With this work it was possible to identify the main allergens involved in the immunologic response of atopic dogs residing in Rio Grande do Sul, pointing to the importance to include C. dactylon in screening tests for allergy. / A dermatite atópica canina (DAC) é uma dermatose comum, definida como uma doença de cunho genético que predispõe à inflamação e ao prurido cutâneo, mediada por imunoglobulinas da classe IgE dirigidas contra antígenos específicos na maior parte dos casos. O diagnóstico da DAC é clínico e pode ser posteriormente complementado por testes alérgicos cutâneos e/ou sorológicos. O objetivo desses testes é identificar possíveis alérgenos e, com isso, possibilitar ao clínico a seleção de antígenos candidatos para a imunoterapia alérgeno-específica. No presente trabalho buscou-se identificar o perfil de sensibilização de 58 cães diagnosticados com dermatite atópica. Todos os animais foram submetidos ao teste intradérmico (TID) e à detecção de anticorpos específicos para diferentes alérgenos através de teste sorológico (ELISA). Os ácaros domiciliares são descritos como os alérgenos mais frequentes em todos os continentes. Entretanto, a positividade ao C. dactylon não é usualmente descrita e pode ser característica da região. Com esse trabalho foi possível identificar os principais alérgenos envolvidos na resposta imunológica de cães atópicos residentes no Rio Grande do Sul, ressaltando-se a importância da inclusão do extrato de C. dactylon em testes alérgicos.

<p>Allergic asthma is a chronic and complex disease of the airways characterized by dysregulated immune-inflammatory responses to aeroallergens and reversible airflow obstruction. The prevalence and economic burden of allergic asthma have increased substantially over the last five decades. Despite remarkable progress in our understanding of the immunobiology and pathophysiology of asthma, the ontogeny of the disease remains elusive. As a result, there is a lack of effective preventative strategies. Here, we used a murine model of allergic asthma to house dust mite (HDM), the most pervasive indoor aeroallergen worldwide to address issues pertaining to the development of allergic asthma. First, we provided a comprehensive computational view of the impact of dose and length of HDM exposure on both local and systemic allergic outcomes (Chapter 2). Parameters, such as thresholds of responsiveness, and non-linear relationships between allergen exposure, allergic sensitization and airway inflammation were identified. We, then, investigated molecular signatures implicated in the onset of allergic responses (Chapter 3). HDM exposure was associated with production of the epithelial-associated cytokines TSLP, IL-25 and IL-33. However, only IL-33 signaling was necessary for intact Th2 immunity to HDM, likely because of its superior ability to induce the critical co-stimulatory molecule OX40L on dendritic cells and expand innate lymphoid cells. Lastly, as individuals are most likely exposed to allergens concomitantly to other environmental immunogenic agents, we studied the impact of an initial immune perturbation on allergic responses to sub-threshold amounts of HDM (Chapter 4). We showed that transient expression of GM-CSF in the airway substantially lowers the threshold of allergen required to generate robust, HDM-specific Th2 immunity, likely through increasing IL-33 production from alveolar type II cells. These studies favor a paradigm whereby distinct molecular pathways can elicit type 2 immunity, intimating the need to classify asthma into distinct clinical subsets.</p> / Doctor of Philosophy (PhD)

Les maladies allergiques sont de plus en plus fréquentent. Elles atteignent souvent l’enfant jeune chez qui l’allergie respiratoire et l’allergie alimentaire sont les principales pathologies. L’unique traitement curatif est l’immunothérapie spécifique d’antigène (ITA), largement développée dans l’allergie respiratoire et encore à ses débuts dans l’allergie alimentaire. Pour adapter au mieux la prise en charge du patient, le diagnostic précis de l’allergie est indispensable et il n’existe actuellement pas d’examen biologique totalement fiable. Seul, la présence d’IgE spécifiques permet de diagnostiquer une sensibilisation à un allergène mais pas une allergie cliniquement symptomatique. Dans une première partie, nous avons étudié l’intérêt d’un test fonctionnel, l’ELISpot (Enzyme-linked immunosorbent spot), dans le diagnostic de l’allergie aux acariens chez l’enfant asthmatique. Le nombre de lymphocytes T circulants spécifiques d’acariens sécréteur d’interleukine (IL)-4 ou d’IL-13 était associé à la présence d’une allergie symptomatique, indépendamment des IgE spécifiques. Il était plus élevé dans le cas d’une rhinite allergique sévère et plus faible dans le cas d’une rhinite allergique légère. De plus, il variait au cours de l’année en fonction des saisons avec un pic en automne et un pic en début de printemps. Dans une deuxième partie, nous avons étudié l’intérêt de l’ELISpot dans le diagnostic de l’allergie au lait de vache chez l’enfant, confirmée par un test de provocation orale en double aveugle. Nous avons décrit que le nombre de lymphocytes T spécifiques de la caséine et sécréteurs d’IL-4 et d’IL-13 était associé à l’allergie au lait de vache avec une sensibilité de 100%. Par ailleurs, le nombre de lymphocytes T spécifiques de la caséine était également associé à la dose maximale de lait tolérée par l’enfant.Enfin, dans une troisième partie, nous avons étudié la réponse lymphocytaire T au cours d’une ITA sub-linguale (SLIT) d’une part et sous-cutanée (SCIT) d’autre part, chez des enfants asthmatiques allergiques aux acariens suivis pendant une année. Nous avons décrit une diminution des lymphocytes Th2 (sécréteurs d’IL-4 et IL-13) spécifiques d’acariens après 12 mois de SLIT associée à une augmentation des cellules sécrétrices d’IL-10 (Tr1) spécifiques d’acariens après 6 mois de SLIT. De plus, les lymphocytes T régulateurs (CD4+CD25hiCD127loFoxp3+) étaient augmentés après 12 mois de SCIT. Nous n’avons pas retrouvé de production accrue d’interféron γ (IFNγ) par les lymphocytes T spécifiques d’acariens au cours de la désensibilisation.Au total, ce travail nous a permis de décrire qu’un test fonctionnel, l’ELISpot, permet de réaliser un diagnostic fiable de l’allergie aux acariens et de l’allergie au lait de vache chez l’enfant. Par ailleurs, l’ITA induit une diminution des cellules Th2 et une augmentation des cellules Tr1 par voie sub-linguale ainsi qu’une augmentation des Treg Foxp3+ par voie sous-cutanée sans immunodéviation Th2/Th1, chez l’enfant allergique aux acariens. / Allergic diseases are steadily increasing steadily and especially in children. Allergen specific immunotherapy (desensitization) is the only curative treatment for which accurate diagnosis of allergy is essential. Currently, the presence of specific IgE diagnoses a sensitization to an allergen but not a clinically symptomatic allergy. In a first part, we studied the value of a functional test, the ELISpot (Enzyme-linked immunosorbent spot) in the diagnosis of allergy to house dust mites (HDM). The number of circulating HDM-specific IL-4 and IL-13 secreting T cells was associated with the presence of symptoms, regardless of specific IgE and was higher in severe rhinitis than in mild rhinitis. In addition, it varied according to the season with a peak in autumn and a peak in early spring (wet periods with greater allergen exposure). In a second part, we studied the value of ELISpot for the diagnosis of cow's milk allergy in children, confirmed by double blind placebo control food challenge. We found that the number of casein-specific IL-4 and IL -13 secreting T-cells was associated with allergy to cow's milk. It was also inversely correlated to the cow’s milk tolerated cumulative dose. Receiver-operating characteristic (ROC) curve of combined IL-4 and IL-13 analysis was generated. AUC was 0,98 (95% CI 0.90-1.06). For a cut-off of 10 IL-4- and 12 IL-13 secreting T-cells, sensitivity and negative predictive value were 100%.Finally, in the third part, we monitored antigen specific T-cell response in HDM allergic children treated with sublingual ITA (SLIT) on the one hand and subcutaneous ITA (SCIT) on the other hand, during one year. We found a decrease in HDM specific Th2 cells after 12 months of SLIT associated with an increase in HDM specific IL-10 secreting T-cells after 6 months of SLIT. In addition, regulatory T cells (CD4 + CD25hiCD127loFoxp3+) were increased after 12 months of SCIT. In conclusion, this work has allowed us to describe a functional test, the ELISpot, as a reliable tool for the diagnosis of mite allergy and cow's milk allergy in children. In addition, in HDM allergic children, a decrease of Th2 cells and an increase of IL-10 secreting T-cells was found in children treated with SLIT to HDM as well as an increase in Foxp3+ Treg in children treated with SCIT.