Astma yra lėtinė kvėpavimo takų uždegimo liga. Mokslininkai neabejoja, jog 2-o tipo T limfocitai pagalbininkai bei eozinofilinis kvėpavimo takų uždegimas yra astmos patogenezės pagrindas. Tačiau šis mechanizmas ne visuomet gali paaiškinti astmos metu esančio kvėpavimo takų uždegimo bei klinikinių simptomų įvairumą, eigos ypatumus ir net skirtingą atsaką į gydymą. Eksperimentiniai tyrimai su gyvūnais parodė 17-o tipo T limfocitų pagalbininkų (Th17) svarbą alerginės astmos vystymuisi. Todėl šio mokslinio tyrimo tikslas buvo įvertinti Th17 limfocitų vaidmenį sergant alergine astma. Tyrimo metu sergantiesiems alergine astma nustatytas didesnis Th17 limfocitų kiekis periferiniame kraujyje bei didesnė interleukino (IL)-17 koncentracija serume ir indukuotuose skrepliuose, lyginant su sveikais asmenimis. Be to, bronchų provokacija su Dermatophagoides pteronyssinus alergenu sukėlė Th17 limfocitų ir IL-17 kiekio padidėjimą praėjus 7 ir 24 val.po jos, ypatingai ryškų sergantiems alergine astma su ankstyva ir vėlyva bronchų obstrukcija. Atlikto tyrimo rezultatai parodė, kad Dermatophagoides pteronyssinus alergenas sukelia vietinį ir sisteminį Th17 limfocitų imuninį atsaką kuris yra susiję su vėlyvos fazės kvėpavimo takų uždegimu. / Allergic asthma is a chronic inflammatory disease of the airways associated with the response of predominant type 2 helper T lymphocytes to an inhaled allergen. However, differences in inflammation and clinical symptoms of this disease not always can be explained by this mechanism. Recent animal model studies have shown the importance of type 17 helper T lymphocytes (Th17) in the development of allergic asthma. The role of these cells in causing allergen-induced airway inflammation as well as systemic inflammatory response in human is still not well defined. Therefore, we investigated the peripheral blood Th17 lymphocyte response to inhaled Dermatophagoides pteronyssinus (D. pteronyssinu) allergen in patients with allergic asthma. The present study has shown that patients with allergic asthma have a higher percentage of peripheral blood Th17 lymphocytes and elevated serum as well as induced sputum interleukin-17 levels compared with healthy subjects. Moreover, all studied allergic asthma patients, especially with early- and late-phase asthmatic reaction, showed an enhanced airway and systemic Th17 lymphocyte response 7 h and 24 h after bronchial challenge. We documented an enhanced local and systemic Th17 lymphocyte response to inhaled D. pteronyssinus in association with late-phase allergen-induced airway inflammation in patients with allergic asthma.

A asma alérgica é uma doença inflamatória crônica das vias aéreas que envolve ativação de fibroblastos pulmonares. Esta ativação é induzida por TGF-<font face=\"Symbol\">b e este processo é regulado por moléculas anti-fibróticas. Nosso objetivo foi elucidar mecanismos envolvidos na fibrose das vias aéreas em um modelo de asma. Na primeira parte, investigamos o eixo síntese/resposta da PGE2. A PGE2 e seu análogo forskolina inibiram síntese de colágeno I e proliferação de fibroblastos. Estas células apresentaram perda tempo-dependente na capacidade de sintetizar PGE2 sob estímulo com IL-1<font face=\"Symbol\">b, e menor expressão de COX-2 e mPGEs-1. Na segunda parte, estudamos a relação TGF-<font face=\"Symbol\">b1/BMP-7 na fibrose das vias aéreas. Há predomínio da molécula pró-fibrótica TGF-<font face=\"Symbol\">b1 sobre a molécula anti-fibrótica BMP-7 nos pulmões de animais asmáticos. Em fibroblastos, a BMP-7 inibe a síntese de colágeno tipo I induzida pelo TGF-<font face=\"Symbol\">b1 e as vias de SMAD-2, SMAD-3 e p38. O tratamento dos animais com BMP-7 causou diminuição significativa da fibrose. Os resultados implicam estes mecanismos na fibrose das vias aéreas na asma. / Allergic asthma is a chronic inflammatory disease of the airways that involves activation of lung fibroblasts. This activation is induced by TGF-<font face=\"Symbol\">b and this process is regulated by anti-fibrotic molecules. Our goal was to elucidate mechanisms involved in airway fibrosis in an animal model of asthma. In the first part, we investigated the PGE2 synthesis/response axis. PGE2 and its analog forskolin inhibited collagen I synthesis and proliferation of fibroblasts. These cells showed a time-dependent loss in the ability to synthesize PGE2 under IL-1<font face=\"Symbol\">b stimulation, and downregulated COX-2 and mPGEs-1. In the second part, we studied the ratio TGF-<font face=\"Symbol\">b1/BMP-7 in airway fibrosis. There is predominance of the pro-fibrotic TGF-<font face=\"Symbol\">b1 over the anti-fibrotic BMP-7 in the lungs of asthmatic animals. In fibroblasts, BMP-7 inhibits TGF-<font face=\"Symbol\">b1-induced type I collagen synthesis and the SMAD-2, SMAD-3 and p38 pathways. Treatment of the animals with BMP-7 caused significant decrease in fibrosis. The results implicate these mechanisms in airway fibrosis in asthma.

Submitted by Renata Lopes (renatasil82@gmail.com) on 2017-01-11T12:38:06Z No. of bitstreams: 1 marcelamota.pdf: 3222712 bytes, checksum: b3a35bc0417b3d343e69a2bd71784fe2 (MD5) / Approved for entry into archive by Diamantino Mayra (mayra.diamantino@ufjf.edu.br) on 2017-01-31T11:19:18Z (GMT) No. of bitstreams: 1 marcelamota.pdf: 3222712 bytes, checksum: b3a35bc0417b3d343e69a2bd71784fe2 (MD5) / Made available in DSpace on 2017-01-31T11:19:18Z (GMT). No. of bitstreams: 1 marcelamota.pdf: 3222712 bytes, checksum: b3a35bc0417b3d343e69a2bd71784fe2 (MD5) Previous issue date: 2016-06-30 / CAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / A asma alérgica é uma doença inflamatória crônica das vias aéreas, caracterizada por sintomas como tosse, broncoespasmo, e hiperreatividade das vias aéreas associada à inflamação eosinofílica. A doença atinge cerca de 300 milhões de pessoas no mundo, exercendo um forte impacto financeiro e social, e sua prevalência em países desenvolvidos vem aumentando consideravelmente. Tal fato está associado à “Hipótese da Higiene”, a qual preconiza que uma menor exposição dos indivíduos a micro-organismos durante a infância pode predispô-los ao desenvolvimento de doenças alérgicas. Considerando o efeito modulador exercido pelo BCG, e visto que a modulação da resposta imune se dá principalmente nos primeiros anos de vida do indivíduo, infere-se que a relação entre a idade do indivíduo e o contato com certos antígenos bacterianos, virais e fúngicos pode reduzir a incidência de doenças respiratórias na população. O presente estudo investigou o efeito modulador exercido pelo BCG na resposta imune inata em camundongos neonatos sensibilizados com OVA. Observou-se uma redução de características ligadas à asma alérgica, tais como produção de citocinas de perfil Th2 e inflamação eosinofílica. Tal fato foi atribuído à modulação da resposta imune, induzida por um aumento de citocinas reguladoras, além da ativação de células dendríticas CD103+CD8α+, presentes na mucosa pulmonar. Estas células são responsáveis pela apresentação do alérgeno e do BCG, através da ativação de receptores tipo Toll 2, 4 e 9, ativando-os e culminando na produção de citocinas. Além disso, atuaram aumentando o percentual de expressão da molécula co-estimuladora PD-L1 após o tratamento, agindo de maneira antagônica na expressão da molécula PD-L2, a qual apresentou uma tendência de redução após o tratamento. Já foi observado que tais moléculas possuem efeitos opostos, sendo que a primeira está relacionada à diminuição da inflação alérgica pulmonar, enquanto a segunda está associada à hiperreatividade das vias aéreas. Os resultados indicam que o BCG é um potente modulador da resposta imune alérgica, e que sua regulação é mais eficaz no período neonatal. / Allergic asthma is a chronic inflammatory disease of the airways, characterized by symptoms such as cough, bronchoconstriction and airway hyperreactivity, associated with eosinophilic inflammation. This disease affects about 300 million people worldwide, and have a huge financial and social impact, and its prevalence has increased considerably in developed countries. This fact is associated to "Hygiene Hypothesis", which proposes that a reduced exposure of individuals to microorganisms during childhood may predispose them to the development of allergic diseases. Considering the modulatory effect exerted by BCG, and since the modulation of immune response is major in the first years of age, implied that relationship between the age of individual and the contact with certain bacterial, viral and fungal antigens, can reduce the incidence of respiratory diseases in population. The present study investigated the modulating effect exerted by BCG in the innate immune response in neonates mice sensitized with OVA. There was a reduction of allergic asthma features, such as the production of Th2 cytokine profile and eosinophilic inflammation. This was attributed to modulation of immune response induced by an increase in regulatory cytokines, as well as activation of dendritic cells CD103+CD8α+ belonging to the lung mucosa. These cells are responsible for allergens and BCG presentation through activation of Toll-like receptors 2, 4 and 9, triggering activation and cytokines production. Furthermore, they acted by increasing the percentage of costimulatory molecule PD-L1 expression after treatment, acting antagonistically in PD-L2 molecule expression, which showed a tendency to decrease after treatment. These molecules have opposite effects, which the first is associated to decrease in allergic lung inflammation, while the second is associated to airway hyperreactivity. The results indicate that BCG is a potent modulator of allergic immune response, and this regulation is more effective in the neonatal period.

Background: It is unclear how far the superior efficacy of omalizumab, established in randomized controlled clinical trials of patients with severe allergic asthma (SAA), translates into routine practice and when compared to matched controls. Methods: New-onset omalizumab-treated (OT) patients with SAA (n = 53) were compared to a matched control group of usual-care (UC) patients (n = 53). Treatment and procedures were naturalistic. Subsequent to a baseline assessment, patients were followed up over at least 6 months with at least two follow-up assessments. Primary clinical outcomes were the number of asthma attacks, persistence of asthma symptoms and degree of control [asthma control test (ACT), Global Initiative for Asthma]. Secondary outcome criteria were quality of life (Euro-Qol 5D) and number of medications. For each outcome we compared within-group effects from baseline to 6-month follow-up as well as between-group effects. Results: OT patients showed significant improvements in number [effect size (ES) = 0.03] and frequency (ES = 0.04) of asthma attacks as well as asthma control (ES = 0.09), whereas controls revealed no significant improvements in these measures. Further improvements in the OT group were found for ‘perceived control always’ (ACT, p = 0.006), no impairment (ACT, p = 0.02), reduction of sickness days (p = 0.002) and number of medications needed (p = 0.001). Conclusions: Substantial beneficial effects of omalizumab, similar to those observed in controlled trials and after marketing studies, were confirmed, particularly with regard to the reduction of asthma attacks, persistence of symptoms, asthma control and reduction of concomitant asthma medications. This study provides a tougher test and generalizable evidence for the effectiveness of omalizumab in routine care.

No description available.

No / Background  Interleukin (IL)-33 is a member of the IL-1 family and has been implicated in Th2-driven allergic diseases such as atopic dermatitis (AD) and asthma. The principal Th2 cytokine IL-4, found highly expressed in acute allergic eczema, is known to downregulate human β-defensin 2 (hBD2) expression in human keratinocytes and this is associated with superinfection in patients with AD. Objectives  To investigate the effect of IL-33 on the expression of hBD2 in human keratinocytes. Methods  hBD2 production by stimulated keratinocytes was measured by enzyme-linked immunosorbent assay. Results  Our results showed that serum is a very potent inducer of hBD2 and 2·5% human serum was much more potent in inducing hBD2 than 20 ng mL−1 of tumour necrosis factor-α. Interestingly, serum from patients with AD showed an impaired ability to induce hBD2 in normal keratinocytes. IL-33 significantly downregulated serum-induced hBD2. The downregulatory capacity of IL-33 was found to be 1·5- to 2-fold weaker compared with IL-4. Conclusions  Our data suggest that IL-33 can significantly contribute to the decreased expression of hBD2 in acute eczematous reaction clinically characterized by spongiosis and oozing – thus indicative for contact of the epidermis with serum components.

Allergic diseases are chronic inflammatory conditions, characterised by eosinophil inflammation systemically and in target organs, where cytotoxic granule proteins are responsible for tissue injury. Allergic rhinitis is known to be a risk factor for the development of asthma, yet not all with rhinitis develop asthma. The overall aim was to investigate the involvement of eosinophils in allergic rhinitis and allergic asthma in vivo and in experimental settings, with a focus on differences between rhinitis and asthma. Birch pollen allergy was used as a model and patients were studied during pollen season and after nasal and bronchial allergen challenge. During pollen season and at baseline, allergic rhinitis and allergic asthma had the same degree of systemic eosinophil inflammation. Despite this, impairment in lung function during season and increased bronchial responsiveness at baseline were more common in the asthmatics. Systemic inflammation was more pronounced after seasonal exposure than after experimental challenge. Allergic rhinitis and allergic asthma had the same degree of eosinophil airway inflammation after bronchial challenge, but only the asthmatics had increased bronchial responsiveness measured as PD20 for birch allergen. Allergen primed eosinophils were investigated in vitro for C3b-induced degranulation after seasonal and experimental challenge. The released amount of eosinophil granule proteins was within the same range for all three allergen challenge models with just minor differences in propensity for degranulation between rhinitics and asthmatics. Signalling through PI3K for degranulation was studied with the specific inhibitor Wortmannin. PI3K signalling for eosinophil degranulation was clearly involved in allergic rhinitis and allergic asthma irrespective of the model for allergen exposure. Asthmatics demonstrated less inhibition of degranulation through PI3K during pollen season, indicating that other pathways contribute to eosinophil degranulation in allergic asthmatics. Conclusion: Allergic rhinitis and allergic asthma present with the same degree of systemic and local eosinophil inflammation. The eosinophils are primed for degranulation equally and follow the same pathway through PI3K for degranulation. Our data indicates that eosinophil inflammation per se is not sufficient for the development of asthma.

Made available in DSpace on 2015-05-14T13:00:11Z (GMT). No. of bitstreams: 1 arquivototal.pdf: 2644910 bytes, checksum: 94bc3ca3dd5252435423f525d32a2ea9 (MD5) Previous issue date: 2009-04-01 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / Asthma is a chronic inflammatory disease characterized by cell migration and bronchial hyperreactivity (HRB). One strategy to treat allergic diseases is the development of new drugs with good efficacy and few side effects. Flavonoids are plant compounds known for its antioxidant, antiallergic and anti-inflammatory activities. Based on this premise, the study aimed to select flavonoids by the immunopharmacological screening and to evaluate prophylactic and therapeutic treatments in the experimental asthma model. For this purpose the flavonoids tested were myricetin, isoramnetin and glycosylated kampherol known as 3-O-[β-D-glycopiranosil-(1→6)-α-L-rhamnopyranosyl]-7-O-α-L-rhamnopyranosyl-kamferol (GRRK). The experimental model was BALB/c mice sensitized and challenged with ovalbumin (OVA) and the treatments were before (prophylactic) or after (therapeutic) to establishment of the experimental asthma model. The three flavonoids inhibited the animal death during the anaphylactic chock reaction induced by OVA and the inflammatory cell migration to the lung; however only the GRRK was used in the following experiments because of its best yield in the purification process. Both prophylactic and therapeutic treatments with GRRK (30 mg/kg) decreased significantly the total number of inflammatory cells (P< 0.05), eosinophils cells (P< 0.05), IL-5 (P< 0.05) and IL-13 (P< 0.05) production from bronchoalveolar lavage (BAL), IgE OVA-specific title (P< 0.01) in the sera, the bronchial hyperreactivity (HRB) (P< 0.05) induced by methacholine and mucus production (P< 0.001) by the globbet cells from the lung as compared with non-treated GRRK animals. The results obtained in the different treatments with GRRK were statistically comparable with that observed in the OVA-sensitized group treated with dexametasone. Moreover, treatment with GRRK was able to reduce the number of CD4+ T cells (p <0.001), B cells (p <0.001), expression of major complex of compatibility II class (MHC II class) and CD40 molecules in the antigen presenting cells (CD11b+ CD11c+) from BAL. Although the GRRK induced a suppressive effect in the Th2 immune response it was not able to exacerbate the Th1 immune response by IFN- production. The production of this cytokine was not modified when compared with non treated animals. The GRRK treatment also was not able to modify the regulatory immune response because there was no significant change in the production of TGF- and the number of TCD4+Foxp3+, a marker of LT regulatory (TReg). These results demonstrated that GRRK treatment before (prophylactic) or after (therapeutic) the establishment of allergic asthma, restored the morpho-functional changes in the airways by Th2-dependent immunossupression, although independent of Th1 and TReg. / A asma é uma doença inflamatória crônica caracterizada por migração celular para os pulmões e hiperreatividade brônquica (HRB). Uma estratégia para o tratamento de doenças alérgicas é o desenvolvimento de novos medicamentos com boa eficácia e poucos efeitos colaterais. Flavonóides são compostos derivados de plantas conhecidos por suas atividades antioxidante, antialérgica e antiinflamatória. Partindo desta premissa, o trabalho teve como objetivo selecionar flavonóides a partir de análises imunofarmacológicas e avaliar os tratamentos profiláticos ou terapêuticos no modelo de asma alérgica experimental. Para tal, os flavonóides testados foram: myricetina, isoramnetina e kanferol glicosilado conhecido por 3-O-[β-D-glycopiranosil-(1→6)-α-L-ramnopiranosil]-7-O-α-L-ramnopiranosil-kanferol (GRRK). O modelo experimental foi realizado em camundongos BALB/c sensibilizados e desafiados com ovalbumina (OVA) e os tratamentos foram antes (profilático) ou após (terapêutico) estabelecer o modelo de asma alérgica experimental. Os três flavonóides inibiram a morte dos animais durante o choque anafilático induzido por OVA e a migração de células da inflamação para os pulmões, entretanto, apenas o GRRK foi usado nos demais experimentos, devido ao seu melhor rendimento no processo de purificação. Os tratamentos profilático e terapêutico com GRRK (30 mg/kg) diminuíram, de forma significativa, o número total de células inflamatórias (P< 0,05), de eosinófilos (P< 0,05) a produção de IL-5 (P< 0,05) e IL-13 (P< 0,05) no lavado broncoalveolar (LBA), o título de IgE-OVA-específica (P< 0,01) no soro, a hiperreatividade brônquica (HRB) (P< 0,05) induzida com metacolina e a produção de muco (P< 0,001) pelas células caliciformes de pulmão quando comparados com os animais não tratados com GRRK e sensibilizado com OVA. Os resultados obtidos nos diferentes tratamentos com GRRK foram comparáveis estatisticamente aos efeitos observados no grupo de animais tratados com a droga padrão dexametasona. Além disso, o tratamento com GRRK foi capaz de diminuir: o número de linfócitos T CD4+ (P< 0,001) e de linfócitos B (P< 0,001), a expressão de moléculas de classe II do complexo principal de histocompatibilidade (MHC II) e da molécula CD40 em células apresentadoras de antígeno (CD11b+CD11c+) do LBA. Embora o tratamento com GRRK tenha induzido um efeito supressor na resposta imune de perfil Th2, não foi capaz de exacerbar a resposta imune Th1 com a produção de IFN- A produção desta citocina se manteve inalterada quando comparada ao grupo de animais não tratados. O tratamento também não foi capaz de alterar a reposta imune reguladora, pois não houve mudança significativa na produção de TGF- e no número de células TCD4+Foxp3+, marcador de LT regulador (LTReg). Esses resultados demonstram que o tratamento com GRRK antes (profilático) ou depois (terapêutico) de estabelecer a asma alérgica experimental, restabeleceu as alterações morfofuncionais das vias aéreas, por um mecanismo modulador das células Th2, embora independente de Th1 e de Treg

Une cellule dendritique (CD) qui exprime le facteur de transcription GILZ durant l’apprêtement de l’antigène et sa présentation aux cellules effectrices, génère des lymphocytes T régulateurs (LTregs) CD25high Foxp3+ sécréteurs d’IL-10. La production de GILZ est dépendante de l’action des glucocorticoïdes, de l’IL-10 et du TGF-.Nous avons mis en évidence chez l’homme qu’une corticothérapie orale de 48h induit l’expression de GILZ dans les cellules présentatrices de l’antigène circulantes (CPAs) de sujets allergiques. Les CPAs isolées après la corticothérapie génèrent des LTregs CD25high Foxp3+ IL-10+ spécifiques de l’allergène.Nous également constaté in vitro que les mastocytes participent à l’activation des CDs au cours des réactions allergiques en régulant l’expression de GILZ. Les médiateurs d’origine mastocytaire, dont l’histamine, diminuent l’expression de GILZ dans les CDs et altèrent ainsi leur capacité à activer des LTregs. Nous avons identifié le mécanisme par lequel l’histamine diminue l’expression de GILZ dans les CDs humaines. L’histamine inhibe l’activité transcriptionnelle de Foxo3, un facteur de transcription régulant l’expression de GILZ. Enfin, nous avons démontré que des souris transgéniques qui surexpriment GILZ constitutivement dans les CDs sont protégées contre le développement de l’asthme allergique. L’ensemble de ces résultats permet d’envisager de nouvelles stratégies d’immunomodulation dans l’allergie, centrée sur la régulation de l’expression de GILZ dans les CDs. / We previously showed in vitro that DCs with a high level of GILZ activate regulatory T cells (Tregs) whereas DCs with low level of GILZ trigger effector T lymphocytes. Glucocorticoids (GCs), IL-10 and TGF- are potent inducers of GILZ expression. The aim of this thesis is to extend the above findings to induction of tolerance to allergens. Modulation of GILZ expression by DCs should induce allergen-specific Tregs able to inhibit the activation and proliferation of allergen specific T cell clones. In order to validate this concept we demonstrated that:- allergen-specific tolerance can be achieved in allergic patients treated with oral GC through the induction of GILZ expression in their antigen-presenting cells, and the role of allergen-specific Tregs in this effect,- mast cells play a role in the activation of DCs by inhibiting their expression of GILZ and thus their ability to stimulate Tregs against harmless environmental allergens,- GILZ-expressing DCs protect against allergic asthma in a model of transgenic mice over-expressing GILZ in their DCs.The present study supports the concept of an immune regulation of allergic responses through the modulation of GILZ expression by DCs and opens new perspectives in the development of innovative immunotherapies in the treatment of allergic diseases.

Allergic asthma is an inflammatory disease of the airways and is characterized by eosinophilic inflammation and increased airway reactivity. In the studies presented in this thesis, lung mechanics and measurements of airway reactivity were assessed in anaesthetized tracheostomized mice by using an animal ventilator (flexiVent®). A forced oscillation technique makes it possible to measure of both airway and tissue mechanics with a potential to distinguish between central and peripheral airways. The results of the experiments on lung mechanics imply that it is important to understand how altered lung mechanics can affect the airway physiology in order to assess the relevance of different animal models of asthma. We have investigated the effects of changing different components of the lung mechanical measurements, such as administering bronchoconstrictive agents via inhalation or intravenously and implementing deep inhalation in animals with airway inflammation. We have also investigated the relation between airway inflammation and oxidative stress. We found that the formation and time-course of F2-isoprostanes, a marker of oxidative stress, and tissue damage were associated with the degree of inflammation and with the degree of heterogeneous airway airflow. Finally we wished to investigate the hypothesis that nitric oxide (NO) may interact with glucocorticoid (GC) treatment because we see a potential for finding new strategies to increase the therapeutic effect in poor responders or patients resistant to GC treatment. NO plays a central role in physiological regulation of the airway function, and is involved in asthma. We found that the concomitant administration of NO and GC attenuated the airway reactivity more than either treatment alone. In conclusion, with the information presented in this thesis, we hope to contribute to the development of better experimental tools and to improved understanding of murine models of asthma for investigating and understanding the underlying pathophysiology of asthma.