<p>Uvod: Interleukin 33 (IL-33) ima značajnu ulogu u inflamatornim i autoimunskim oboljenjima, ali se sve vi&scaron;e proučava njegov značaj u imunopatogenezi različitih alergijskih oboljenja, uključujući i alergijsku astmu (AA). Cilj: Ispitivanje vrednosti IL-33 u serumu pacijenata sa AA pre i posle &scaron;estomesečne inhalatorne kortikosteroidne terapije (ICS Th) i povezanosti dobijenih vrednosti IL-33 sa određenim kliničkim i laboratorijskim karakteristikama ovih pacijenata. Metode: Vrednost IL-33 u serumu određena je kod 61 pacijenta sa AA pre započinjanja i posle sprovedne &scaron;estomesečne ICS Th i kod 30 zdrave dece. U obradi podataka primenjene su standardne metode deskriptivne i analitičke statistike. Rezultati: Kod pacijenata sa nelečenom AA, serumske vrednosti IL-33 su signifikantno veće u odnosu na pacijente kod kojih je sprovedena &scaron;estomesečna ICS Th (p&lt;0,05), kao i u odnosu na zdravu decu (p&lt;0,01). Pacijenti sa AA koji su tokom 6 meseci lečeni sa ICS Th i zdrava deca imaju slične vrednosti IL-33 u serumu (p&gt;0,05). Kod pacijenata sa AA pre započinjanja i 6 meseci posle primene ICS Th ne postoji signifikantna korelacija između vrednosti IL-33 u serumu i eozinofilnih granulocita periferne krvi (p&gt;0,05), eozinofilnih granulocita u nazalnom sekretu (p&gt;0,05) i ukupnog IgE u serumu (p&gt;0,05). Kod pacijenata sa nelečenom AA postoji signifikantna negativna korelacija između vrednosti serumskog nivoa IL-33 i sledećih parametara plućne fukcije: FEV1 (p&lt;0,05), FEV1/FVC (p&lt;0,05), PEF(p&lt;0,05) i MEF 25/75 (p&lt;0,05). Posle &scaron;estomesečne ICS Th pobolj&scaron;ava se plućna funkcija, odnosno dolazi do porasta brzine protoka vazduha u disajnim putevima kao i promena u plućnim volumenima u zavisnosti od stepena opstrukcije u odnosu na vrednosti pre uključenja antiinflamatorne terapije (FEV1, FVC, FEV1/FVC, PEF, MEF 25/75, za sve vrednosti p&lt;0,01). Dok je signifikantna negativna korelacija dokazana između IL-33 i vrednosti FEV1 (p&lt;0,01), FVC (p&lt;0,01) i PEF (p&lt;0,05). Zaključak: Serumski nivo IL-33 je značajno povi&scaron;en kod dece sa nelečenom, odnosno nekontrolisanom AA. &Scaron;estomesečna primena ICS dovodi do značajne redukcije IL-33 u serumu čije su vrednosti u negativnoj korelaciji sa vrednostima FEV1, FVC i PEF, odnosno pozitivnoj korelaciji sa težinom i kontrolom AA. Rezultati na&scaron;e studije ističu da IL-33 ima značajnu ulogu u imunopatogenezi AA. Određivanje serumske vrednosti IL-33 može biti koristan indikator težine AA.</p> / <p>Introduction: Interleukin 33 (IL-33) plays a significant role in inflammatory and autoimmune diseases, but its significance in the immunopathogenesis of various allergic diseases including allergic asthma (AA) has gained increasing attention in research over recent years. Objective: Testing serum levels of IL-33 in patients with AA before and after a six-month inhaled corticosteroid therapy (ICS Th) and correlation of IL-33 values with specific clinical and laboratory characteristics of these patients. Methods: Serum levels of IL-33 were determined in 61 patients with AA prior to the initiation of ICS Th and following the six-month ICS Th as well as in 30 healthy children. Data processing was performed applying standard methods of descriptive and analytical statistics. Results: In patients with untreated AA, serum levels of IL-33 were significantly higher as compared to the patients who have received a six month ICS Th (p &lt;0.05) as well as to healthy children (p &lt;0.01). Patients with AA, who were treated with ICS Th for six months, and healthy children have similar serum IL-33 (p&gt; 0.05). In patients with AA, significant correlation between serum IL-33 levels and eosinophilic peripheral blood granulocytes (p&gt;0.05), eosinophilic granulocytes in nasal secretion (p&gt;0.05) and the total IgE in serum has not been observed for the period prior to initiation and 6 months after the administration of ICS Th. In patients with untreated AA, there is significant negative correlation between serum IL-33 and the following pulmonary functions test results: FEV1 (p&lt;0.05), FEV1/FVC (p&lt;0.05), PEF (p&lt;0.05) and MEF 25/75 (p&lt;0.05). After a six-month ICS Th, significant improvement of pulmonary functions was evident, that is, increase in airflow speed and lung volume change as compared to the values determined before the initiation of the anti-inflammatory therapy (FEV1, FVC, FEV1/FVC, PEF, MEF 25/75, for all values p&lt;0.01). Significant negative correlation between IL-33 and the values of FEV1 (p&lt;0.01), FVC (p&lt;0.01)and PEF (p&lt;0.05) has been established. Conclusion: Serum level of IL-33 is significantly elevated in children with untreated, i.e., uncontrolled AA. A six-month ICS Th asthma treatment results in significant reduction of serum levels of IL-33. This level is negatively correlated with FEV1, FVC and PEF values while positively correlated with the severity of the disease and control of AA. The results of our study point out that IL-33 plays an important role in the immunopathogenesis of AA. Quantification of serum IL-33 levels can be a useful indicator of the severity of AA.</p>

A dermatite de contato alérgica (DCA) ao íon níquel (Ni2+) é uma dermatose inflamatória frequente nos países industrializados. Envolve a ativação de células T específicas ao Ni2+, seguida da proliferação e indução de um perfil misto de citocinas, tanto pró-inflamatórias quanto reguladoras, sugerindo que vários subtipos de células T (helper - Th e citotóxica - Tc) estão envolvidos na resposta imune. Este estudo teve como objetivo a análise das citocinas TNF-alfa, INF-y, IL-2, IL-4, IL-10, IL-13, IL-17 e IL-23 pela técnica de imuno-histoquímica, para tentar identificar a prevalência de um ou mais subtipos de células T (Th/Tc), nos eczemas crônico e agudo de pacientes com DCA ao Ni2+. Avaliamos 20 pacientes (17 mulheres e 3 homens, com idade mediana de 46 anos) apresentando eczema crônico, pelo contato cotidiano do paciente com o Ni2+. Foram coletadas duas biópsias de pele em cada um dos 20 pacientes, a primeira no local do eczema crônico ao Ni2+, antes da aplicação do teste de contato (TC); e a segunda no local do eczema agudo, provocado pelo TC com o sulfato de níquel, 48 horas após sua fixação, nas leituras positivo forte (++) ou positivo muito forte (+++). Foram 160 amostras de eczema agudo e 160 de eczema crônico, perfazendo um total de 320 amostras. Apenas três amostras foram excluídas devido a algum tipo de falha técnica, como, por exemplo, o descolamento dos cortes de pele da lâmina. Para a análise dos dados utilizou-se o software estatístico STATA versão 13. As amostras coradas revelaram resultados positivos para as oito citocinas estudadas, e estas apresentaram valores heterogêneos. Esta heterogeneidade foi medida pelo coeficiente de variação, indicando a variabilidade do conjunto dos dados obtidos. O TNF-alfa, IFN-y, IL-4, IL-13 e IL-17 tiveram prevalência maior no eczema crônico do que no eczema agudo, a IL-2 e IL-23 apresentaram maior prevalência no eczema agudo, em comparação com o eczema crônico e a IL-10 apresentou prevalência similar tanto no eczema agudo quanto no crônico, porém, estas prevalências foram muito baixas, em ambos os eczemas. O TNF-alfa foi a citocina que mais prevaleceu no eczema crônico e a IL-2 foi a mais prevalente no eczema agudo. Porém, estas prevalências foram estatisticamente significantes apenas para a IL-4 e IL-13. Verificamos, nos eczemas crônico e agudo, a presença de um perfil misto de citocinas dos subtipos de células T (Th/Tc), sugerindo que as respostas imunes são expressas ao mesmo tempo. Entretanto, são necessários mais estudos para uma compreensão mais ampla sobre o perfil das citocinas na DCA ao Ni2+, o que poderia levar a novas abordagens terapêuticas / Allergic contact dermatitis (ACD) to nickel (Ni+2) is a inflammatory dermatosis, common in industrialized countries. It involves the activation of nickel-specific T cells, followed by the proliferation and induction of a mixed profile of both proinflammatory and regulatory cytokines, suggesting that several T cell subtypes (helper - Th and cytotoxic - Tc) are involved in the immune response. This study aimed to analyze the cytokines TNF-alfa, INF-y, IL-2, IL-4, IL-10, IL-13, IL-17 and IL-23 using the immunohistochemistry technique in order to try to identify the prevalence of one or more T cell subtypes (Th/Tc) in the chronic and acute eczema of patients with ACD to Ni+2. We evaluated 20 patients (17 women and 3 men, median age of 46 years) with chronic eczema, by the patient\'s daily contact with Ni+2. Two skin biopsies were collected in each of the 20 patients, the first at the site of the chronic eczema to Ni+2, prior to the application of the contact test (CT); and the second at the site of acute eczema caused by CT with nickel sulphate, 48 hours after its fixation in the strong positive (++) or very strong positive (+++) readings. There were 160 samples of acute eczema and 160 of chronic eczema, a total of 320 samples. Only three samples were excluded due to some kind of technical failure, such as detachment of the skin cuts from the microscope slide. Statistical software STATA version 13 was used to analyze the data. The stained samples showed positive results for the eight cytokines studied, and these presented heterogeneous values. This heterogeneity was measured by the coefficient of variation, indicating the variability of the data set obtained. TNF-alfa, IFN-y, IL-4, IL-13 and IL-17 had a higher prevalence in chronic eczema than in acute eczema, IL-2 and IL-23 were more prevalent in acute eczema compared to chronic eczema and IL-10 presented similar prevalence in both acute and chronic eczema, however, a very low prevalence in both eczema. TNF-alfa was the most prevalent cytokine in chronic eczema and IL-2 was the most prevalent in acute eczema. However, these prevalences were statistically significant only for IL-4 and IL-13. In chronic and acute eczema, we observed the presence of a mixed cytokine profile of the T cell subtypes (Th/Tc), suggesting that immune responses are expressed at the same time. However, further studies are needed for a broader understanding of the cytokine profile in ACD to Ni+2, which could lead to new therapeutic approaches

A asma é uma doença inflamatória crônica, predominantemente das vias aéreas, mas também envolve o sistema vascular e parênquima pulmonar, na qual as células inflamatórias, a musculatura lisa e o epitélio brônquico têm um papel fundamental na fase inicial, progressão e perpetuação da doença. O treinamento físico aeróbio tem sido indicado como uma relevante forma de auxílio no tratamento de pacientes asmáticos por melhorar a qualidade de vida e reduzir sintomas e o uso de medicamentos. No entanto, não existe um consenso a respeito sobre a intensidade de treinamento ideal para esses pacientes assim como também existem pouquíssimos estudos a respeito dos possíveis mecanismos da atividade física aeróbia para esses pacientes. Por esses motivos, os objetivos do presente estudo foram avaliar os efeitos de duas intensidades de atividade física aeróbia (leve e moderada) sobre um modelo de inflamação pulmonar alérgica crônica em camundongos. Os animais foram sensibilizados com ovalbumina por 51 dias. A atividade física aeróbia teve início no dia 21 e perdurou por 30 dias. Os resultados demonstraram que ambas as intensidades de atividade aeróbia inibiram o desenvolvimento da inflamação predominantemente eosinofílica no lavado broncoalveolar, nos compartimentos peribrônquico, perivascular e no parênquima pulmonar, a expressão de interleucina 4 e 5 pelas células inflamatórias nestes três compartimentos pulmonares. Ambas intensidades de atividade física aeróbia também inibiram significativamente a deposição de fibras colágenas e elásticas (nas vias aéreas e vasos pulmonares) e também o espessamento da musculatura lisa brônquica e vascular assim como da camada epitelial brônquica. Por outro lado, ambas intensidades de atividade física aeróbia não inibiram a síntese de anticorpos anafiláticos IgE e IgG1 e a hiperresponsividade brônquica. Portanto, concluímos que a atividade física aeróbia de intensidade leve e moderada são capazes de inibir o desenvolvimento da inflamação e do remodelamento pulmonar, mas não a hiperresponsividade num modelo experimental de inflamação pulmonar alérgica crônica em camundongos. / Asthma is a chronic inflammatory disease, predominantly involving the airways, but also involving the pulmonary vessels and parenchyma, in which the inflammatory cells, bronchial smooth muscle and epithelium have a central role in the initial phase, progression and perpetuation of the disease. The low and moderate intensity of aerobic physical training have been indicated as a relevant mean to help in the treatment of asthmatic patients, improving life quality, decreasing symptoms and the use of medicines. However, there is not a consensus about the best intensity of training to these patients and also there are few studies about the possible mechanisms of aerobic physical training for asthmatic patients. Therefore, the aims of this study was to evaluate the effects of two intensities of aerobic physical training (low and moderate) on an experimental model of chronic allergic lung inflamattion in mice. The animals were sensitizes with ovalbuim during 51 days. The aerobic physical training started to 21st day and endures for 30 days. The results showed that as low as moderate intensities of aerobic physical training inhibited the eosinophilic inflammation in the bronchoalveolar lavage, peribronchial, perivascular and in the pulmonary parenchyma, as well as the expression of interleukin 4 and 5 by inflammatory cells in these three pulmonary compartments. Both intensities of aerobic physical training also inhibited the collagen and elastic fibers deposition (in the airways and in the pulmonary vessels) and also the thickness of smooth muscle in the airways and vessels, as well as of the airway epithelial layer. On other hand, both intensities of aerobic physical training did not inhibit the synthesis of anaphylactic antibodies IgE and IgG1 and the hyperresponsiveness. Therefore, we conclude that aerobic physical training, to both intensities, were capable of inhibit the development of pulmonary inflammation and remodeling, but not of hyperresponsiveness in an experimental model of allergic lung inflammation in mice.

O objetivo deste trabalho foi esclarecer qual das frações protéicas da soja é mais alergênica, avaliando-se sua imunogenicidade e alergenicidade por imunoensaios e teste de anafilaxia cutânea passiva. Além desse propósito, este estudo visou a produção de anticorpos monoclonais para utilizá-lo como ferramenta na padronização de um ensaio imunoenzimático com a finalidade de detectar proteínas da soja, em produtos comercializados. A purificação das frações 2S,7S e 11S da soja foi realizada com base em estudos prévios, padronizando-se a metodologia dentro das condições experimentais do laboratório. A constatação da pureza foi realizada por eletroforese em gel de poliacrilamida (gel de empilhamento a 5% e de separação em gradiente de 7 a 15%), onde se verificou as bandas protéicas características de cada fração. Evidenciou-se as subunidades &#945; (63,17 KDa) &#945;\' (58,06 KDa) e &#946; (42,09 KDa), correspondentes à fração 7S da soja e as subunidades ácida (38,8 KDa) e básica (21,04 KDa), correspondentes à fração 11S da soja. A fração 2S da soja mostrou uma banda protéica de 20 KDa, nesse ensaio. Os resultados da cinética demonstraram que as frações 7S e 11S da soja são muito imunogênicas, visto que induziram uma grande produção de anticorpos das classes IgM e IgG, em camundongos BALB/c. Por outro lado, a fração 2S da soja induziu uma pequena produção de anticorpos IgG, principalmente no 30&#186; dia após a imunização. No teste de anafilaxia cutânea passiva, quando se avaliou a capacidade das frações protéicas da soja de induzirem a produção de anticorpos IgE, em camundongos BALB/c, verificou-se que a fração 7S da soja é alergênica, nesse modelo experimental. Entretanto, as frações 2S e 11S não estimularam o sistema imunológico desses animais a produzir essa classe de anticorpo. Quando se investigou a presença de anticorpos IgE dirigidos contra as frações protéicas 2S, 7S e 11S da soja, no soro de pacientes alérgicos, obteve-se resultado negativo com todas as três proteínas estudadas. Entretanto, na análise da presença de IgG, tanto o soro dos pacientes alérgicos como o dos controles mostraram-se reativos às frações 2S e 7S da soja, sugerindo a presença do anticorpo IgG4 anafilático no soro teste. Os quatro anticorpos monoclonais reativos às frações 7S e 11S da soja, obtidos neste estudo, mostraram-se reativos até as diluições 1/8000 (2A8 e 1H4) e 1/10000 (1F9) para a fração 7S e 1/12000 (3F2) para a fração 11S da soja. A identificação dos determinantes antigênicos reconhecidos pelos anticorpos monoclonais foi realizado por Imunotransferência. Observou-se que os anticorpos monoclonais anti-fração protéica 7S da soja (1H4, 2A8 e 1F9) reconheceram as subunidades &#945;, &#945; e &#946; (1F9), &#945;\' e &#946; (1H4) e &#946; (2A8), além de outras proteínas. Por outro lado, o anticorpo monoclonal anti-fração protéica 11S da soja (3F2) reagiu apenas com a subunidade básica dessa proteína. A padronização do ensaio imunoenzimático com o objetivo de detectar proteínas de soja, em produtos comercializados, mostrou que os anticorpos monoclonais dirigidos contra as frações protéicas 7S e 11S reconhecem, apenas, as proteínas da soja. Em conclusão, esses resultados sugerem que as frações protéicas 7S e 11S da soja são imunogênicas em camundongos, enquanto que na espécie humana, essa resposta foi observada com as frações 2S e 7S da soja. No teste de anafilaxia cutânea passiva, a fração protéica 7S da soja mostrou a capacidade de induzir anticorpos da classe IgE em camundongos BALB/c, sugerindo que essa fração é alergênica nesse ensaio. Por outro lado, os anticorpos monoclonais reativos às frações 7S e 11S detectaram as proteínas da soja nos produtos comercializados testados. / The goal of this study was to elucidate which of the soy protein fractions is more allergenic by using enzyme immunoassays and passive cutaneous anaphylactic activity test. Furthermore, monoclonal antibodies were obtained in order to standardize immunoassays to detect soy protein fractions in commercial soy-derived products. Purification of 2S, 7S and 11S fractions from soy was based on previous studies which were adapted to our experimental conditions. Purity of the isolated fractions was determined by polyacrilamide gel electrophoresis (7-15%). The subunits &#945; (63,17 KDa) &#945;\' (58,06 KDa) and &#946; (42,09 KDa), corresponding to 7S fraction as well as the acid (38,8 KDa) and basic (21,04 Kda) subunits of 11S fraction were visualized. The 2S fraction showed a 20 KDa band only. Data showed that the 7S and 11S soy fractions are immunogenic as they elicited IgM and IgG antibodies in BALB/c mice. In contrast, the 2S fraction was not immunogenic in this specie. The passive cutaneous anaphylactic activity test showed that 7S fraction is allergenic as it elicited IgE production in BALC mice. However, this test was negative for 2S and 7S fractions. No IgE reactive against the 2S, 7S e 11S was found in sera of patients (n=5) presenting previous food allergy symptoms. However, either these patients or controls (n=5) showed IgG reactive against 2S and 7S fractions in their sera. The four monoclonal antibodies obtained in this study were reactive up to 1/8000 (2A8 and 1H4) and 1/10000 (1F9) against 7S fraction, and up to 1/12000 (3F2) for 11S fraction. Immunoblot analysis showed that the monoclonal antibodies anti-7S fraction (1H4, 2A8 e 1F9) recognized the &#945; &#945;\' and &#946; (1F9), &#945;\' and &#946; (1H4) and &#945; (2A8) subunits. The monoclonal antibody anti-11S fraction (3F2) reacted with the basic subunit of this fraction only. By using ELISA and the monoclonal antibodies anti-7S and anti-11S it was possible to detect especifically these soy protein fractions in commercial soy-derived products. In conclusion, data suggest that 7S and 11S soy protein fractions are immunogenic in mice while 2S and 7S fraction are immunogenic in humans. The IgE response to 7S fraction in BALB/c mice showed by the passive cutaneous anaphylactic activity test indicates that this soy protein fraction is allergenic in this experimental model. Finally, the monoclonal antibodies anti-7S and anti-11S obtained in this study did not react with other vegetable or animal proteins being suitable to be used in ELISA for detection of these proteins in commercial soy-derived products.

Diss. Linköping : Univ., 2004.

Influenza is a preventable infectious disease, against which vaccination is the primary means of protection. Health care workers (HCW) are among the most vulnerable to the illness and are likely to be sources of infection transmission while caring for patients. Circumstantial evidence suggests higher rates of vaccination coverage by HCW will coincide with a lower incidence of influenza transmission, yet a gap remains in the literature regarding governing health agencies' (i.e., licensing boards, medical and nursing associations) influence on the influenza vaccination practices of their constituents. Moreover, discrepancies exist between governing health agencies' and the National Vaccine Advisory Committee's recommendations on mandatory influenza vaccination for HCW. The main purpose of this quantitative cross-sectional study was to explore the relationship between influenza vaccination uptake by HCW and guidance from governing health agencies to vaccinate. The health belief model and social cognitive theory were used to identify the most influential determinant for HCW to vaccinate against influenza. The sample consisted of 388 HCW who provided direct patient care at the same hospital. Data were analyzed using Fisher's exact test. Study findings suggest that a workplace mandate for influenza vaccination has an influence on HCW uptake of the vaccine and that governing agencies' lack of uniformity on the matter has minimal impact on their constituents' beliefs and behavior. It is recommended that a universal policy be adopted for health agencies' implementation of an influenza vaccine mandate, which could lead to positive social change by supporting preventive self-care practices, minimizing spread of the disease to workers and patients, and maintaining workplace productivity.

Nonalcoholic fatty liver disease (NAFLD) has become one of the most common hepatic diseases worldwide, making the diagnosis and management of NAFLD an emerging public health issue. Theories associated with NAFLD surmise that inflammation may be the root cause, along with the complex interplay of other chronic conditions such as obesity, metabolic syndrome, diabetes, dyslipidemia, and cardiovascular disease (CVD). It is unknown if other inflammatory conditions such as rheumatoid arthritis (RA), along with the use of methotrexate (MTX), might confer increased risk for NAFLD. Longitudinal data collected from a retrospective cohort of 17,481 adult RA patients in the United States were used to determine the incidence and factors associated with the development of NAFLD using a noninvasive tool (Fibrosis-4 score). Results of the Kaplan Meier analysis showed that 31% of this cohort developed NAFLD, in about 7 years from baseline, with most having mild to moderate disease and only 1.4% with advanced disease. RA patients also had a prevalence of chronic conditions associated with NAFLD, as seen in the general population. In the Cox proportional hazard multivariate analysis, age (middle and elderly), hypertension, CVD, dyslipidemia, metabolic syndrome, exercise, use of MTX, and non-MTX antirheumatic drugs were independent predictors for the development of NAFLD. This research could improve early diagnosis of NAFLD using a novel noninvasive tool. Increase awareness of the prevalence and causes of NALFD inform clinical practice and management of the disease and influence policy about this chronic condition in patients with RA.

Rationale: The purpose of this study is to investigate the stored stability of dilutions of neuromuscular blocking agents (NMBAs), namely cisatracurium, rocuronium, and vecuronium, for skin prick/intradermal testing. Methods: Concentrations of NMBAs were monitored by liquid chromatography-mass spectrometry (LC-MS/MS) for a period of 14 days. Dilutions of NMBAs were prepared in saline by factors of 10x, 100x, 1,000x, 10,000x, and 100,000x as sensitivity of the assay allowed. Diluted drug products were stored in a laboratory refrigerator until sampling. On sampling days, aliquots of each dilution were removed and compared to a freshly prepared set of reference dilutions. Results: The results are measured as beyond use date (BUD) defined as recovery of drug versus the reference (90-110%). Based on the LC-MS/MS data, the BUD for cisatracurium diluted to 10x and 100x is 96 hours. Higher dilutions (1,000x to100,000x) should be used immediately following preparation (within less than 24 hours). Vecuronium at 10x and 100x, also has a BUD of 96 hours, and the 1,000x dilution is stable for 24 hours. The 10,000x dilution should be used immediately. Rocuronium at 10x to 1,000x has a BUD of 48 hours, yet higher dilutions (10,000x and 100,000x) should be used immediately. Conclusions: With increasing dilution factors, the stability of these drugs in saline decreases, increasing deviation between samples and references. The most stable dilutions for each of the drugs tested were 10x and 100x. Stability of these drugs is likely compromised by hydrolysis of the ester bonds in the drug molecules.

<p>Food allergies are immune system-driven diseases that lead to reproducible adverse reactions which can be fatal. These severe systemic reactions are primary mediated by immunoglobulin E (IgE) that is derived from B cells which have been activated by T helper type 2 (Th2) cells. While much work has advanced the clinical and pharmacological management of patients with allergic diseases, much remains to be elucidated about how individuals initially acquire allergy. This Thesis details a mechanism linking initial gastrointestinal exposure to peanut (PN) allergen, to the generation of Th2 cells: PN allergen activates epithelial cell secretion of interleukin (IL)-33 and eosinophil degranulation of eosinophil peroxidase, which causes CD103+ dendritic cell (DC) activation and migration to mesenteric lymph nodes where DC OX40L engages naïve T cells to secrete IL-4 in an autocrine/paracrine manner to promote and consolidate Th2 cell differentiation. These events are followed by B cell activation and PN-specific IgE production, which sensitizes mast cells to be hypersensitive to PN re-exposure by causing immediate allergic reactions including anaphylaxis. This is later followed by eosinophilic inflammation that is partially mediated by innate lymphoid cells. As food allergy also serves as a unique model to better understand mechanisms of adaptive immunity, especially Th2 immunobiology, both basic science and clinical implications are discussed in this Thesis. Major themes include Th2 and disease heterogeneity, identification of ‘the original source of IL-4’, an unprecedented <em>in vivo </em>requirement for eosinophils in priming adaptive immune responses, and the need to weigh basic science findings against the human disease <em>in natura </em>litmus test. Looking forward, many questions remain to be answered in the field of food allergy research, but the findings of this Thesis may be one step towards the prevention, management or cure of a disease with growing public concern, potentially fatal consequences, and an unmet need in understanding its pathogenesis.</p> / Doctor of Philosophy (Medical Science)

INTRODUÇÃO: A importância da resposta mecânica do parênquima pulmonar na fisiopatologia da asma tem sido recentemente reconhecida. O óxido nítrico é um mediador que controla o tônus muscular liso das vias aéreas, porém este efeito no parênquima pulmonar periférico ainda não foi previamente investigado. Nossa hipótese é que a inibição crônica das óxido nítrico sintases por meio do tratamento com L-NAME (falso substrato para todas as óxido nítrico sintases) pode modular a mecânica do parênquima pulmonar, o recrutamento eosinofílico e o remodelamento da matriz extracelular em modelo de inflamação alérgica crônica pulmonar em cobaias. MÉTODOS: Os animais foram expostos a sete inalações com soro fisiológico ou com ovoalbumina em doses crescentes (1~5mg/ml - 4 semanas) e tratadas ou não com L-NAME (60 mg/kg/ por dia /por animal) na água de beber. Setenta e duas horas após a sétima inalação os animais foram anestesiados, exsanguinados e a mecânica oscilatória do parênquima pulmonar foi medida na condição pré e após desafio (0.1%). Utilizando a técnica de morfometria foram avaliadas a densidade de eosinófilos, o número de células nNOS e iNOS positivas, a densidade de actina, das fibras colágenas e das fibras elásticas bem como a proporção de volume de 8-iso-PGF2 no septo alveolar. RESULTADOS: Os animais que foram expostos à ovoalbumina apresentaram um aumento da resistência e da elastância tecidual (resposta basal e após desafio antigênico), na densidade de eosinófilos, no número de células nNOS e iNOS positivas, na densidade de fibras colágenas e de fibras elásticas bem como na expressão de 8-isoPGF2 no septo alveolar comparativamente aos grupos controles (p<0,05). O tratamento com L-NAME em animais expostos à ovoalbumina atenuou todas as respostas de mecânica do tecido pulmonar periférico (p<0, 01), reduziu o número de células nNOS e iNOS positivas (p<0.01), o conteúdo de fibras elásticas (p<0,001) e de 8-iso-PGF2 no septo alveolar (p<0,001). No entanto, este tratamento não afetou o número total de eosinófilos e o conteúdo de fibras colágenas. Este trabalho sugere que o óxido nítrico contribui para a constrição do parênquima pulmonar e para a deposição de fibras elásticas neste modelo. Estes efeitos foram associados à ativação de iNOS e nNOS em células do parênquima distal e aumento na via do estresse oxidativo / The importance of lung tissue mechanical responses in asthma pathophysiology has been recently recognized. Although nitric oxide (NO) is a mediator that controls smooth muscle tonus control in the airways, its effects on lung tissue responsiveness has not been previously investigated. We hypothesized that chronic nitric oxide synthase inhibition by L-NAME (false substrate for all nitric oxide synthases) treatment may modulate lung tissue mechanics, eosinophilic recruitment and extracellular matrix remodeling in a model of chronic pulmonary allergic inflammation. Guinea pigs were submitted to seven normal saline or ovalbumin exposures with increasing doses (1~5mg/mL-4weeks) and treated or not with L-NAME in drinking water. Seventy-two hours after the seventh inhalation the animals were anesthetized, exsanguinated, and oscillatory mechanics of lung tissue strips was performed in baseline condition and after ovalbumin challenge (0.1%). Using morphometry, we assessed the density of eosinophils, the number of iNOS and nNOS-positive cells, the density of actin, the collagen and elastic fibers content and the volume proportion of 8-iso-PGF2 in the alveolar septa. Ovalbumin-exposed animals presented an increase in baseline and maximal tissue resistance and elastance responses, eosinophil density, in the number of iNOS and nNOS positive cells, in the amount of collagen and elastic fibers and in the volume proportion of 8-iso-PGF2 in the alveolar septa compared to controls (p<0.05). L-NAME treatment in ovalbumin-exposed animals attenuated all lung tissue mechanical responses (p<0.01), reduced the number of iNOS and nNOS positive cells (p<0.01), elastic fiber content (p<0.001) and 8-isoPGF2 in the alveolar septa (p<0.001). However, this treatment did not affect the total number of eosinophils and collagen deposition. These data suggest that NO contributes to distal lung parenchyma constriction and to elastic fibers deposition in this model. These effects were associated to iNOS and nNOS activation in pulmonary parenchyma and with an increase in oxidative stress pathway activation