The innate immune protein surfactant protein (SP-) D is a carbohydrate binding protein that was originally isolated from mucosal lung tissues. Recently, studies show that SP-D binds to antibodies, including immunoglobulin M (IgM), which interacts with late apoptotic cells. Here we focus on the interaction between SP-D and IgM as they pertain to late apoptotic cell clearance. We hypothesized that the three-way interaction between IgM, SP-D and late apoptotic cells is functionally applicable to clearing late apoptotic cells from the lungs, thereby reducing lung inflammation. We show that SP-D binds to IgM and that IgM binds to the late apoptotic subclass of dying cells. We demonstrate that IgM and SP-D can both bind to late apoptotic cells in mutually distinct regions while also displaying some regional overlap. We show evidence that during LPS-induced lung inflammation both IgM and SP-D levels are elevated and this corresponds to an augmentation of apoptotic cell clearance. We illustrate that the protein interaction of IgM and SP-D is functionally relevant to apoptotic cell clearance in the lungs by showing that late apoptotic cells coated in IgM and/or SP-D are cleared more efficiently than control cells, by alveolar macrophages in vivo. Our ex vivo studies further show that these cells internalize apoptotic cells by engulfing very small particles released from the dying cells. We then showed that IgM preferentially directs the engulfment of small particles (~1 μm) by macrophages, in an apparent size-specific antibody-dependent particle clearance function. Our data reveals a novel relationship amongst IgM, SP-D, apoptotic cells, and alveolar macrophages that contributes to our understanding of apoptotic cell clearance, which may be used in the future to generate strategies addressing apoptotic cell accumulation or clearance deficiency in disease.

The innate immune protein surfactant protein (SP-) D is a carbohydrate binding protein that was originally isolated from mucosal lung tissues. Recently, studies show that SP-D binds to antibodies, including immunoglobulin M (IgM), which interacts with late apoptotic cells. Here we focus on the interaction between SP-D and IgM as they pertain to late apoptotic cell clearance. We hypothesized that the three-way interaction between IgM, SP-D and late apoptotic cells is functionally applicable to clearing late apoptotic cells from the lungs, thereby reducing lung inflammation. We show that SP-D binds to IgM and that IgM binds to the late apoptotic subclass of dying cells. We demonstrate that IgM and SP-D can both bind to late apoptotic cells in mutually distinct regions while also displaying some regional overlap. We show evidence that during LPS-induced lung inflammation both IgM and SP-D levels are elevated and this corresponds to an augmentation of apoptotic cell clearance. We illustrate that the protein interaction of IgM and SP-D is functionally relevant to apoptotic cell clearance in the lungs by showing that late apoptotic cells coated in IgM and/or SP-D are cleared more efficiently than control cells, by alveolar macrophages in vivo. Our ex vivo studies further show that these cells internalize apoptotic cells by engulfing very small particles released from the dying cells. We then showed that IgM preferentially directs the engulfment of small particles (~1 μm) by macrophages, in an apparent size-specific antibody-dependent particle clearance function. Our data reveals a novel relationship amongst IgM, SP-D, apoptotic cells, and alveolar macrophages that contributes to our understanding of apoptotic cell clearance, which may be used in the future to generate strategies addressing apoptotic cell accumulation or clearance deficiency in disease.

[Truncated abstract] Fibrosis is a feature of many pulmonary conditions, including idiopathic pulmonary fibrosis (IPF), which is characterised by the accumulation of fibroblasts/myofibroblasts and excessive deposition of collagen. IPF is a disease of unknown aetiology that is unresponsive to current therapy and is typically fatal. The inflammatory cytokine interleukin (IL)-6 is elevated in patients with IPF and recent studies have shown that IL-6-induced signalling is altered in lung fibroblasts from patients with IPF. IL-6 belongs to the gp130 cytokine family, which is a group of ten structurally related cytokines, that all require the membrane bound glycoprotein gp130 to activate intracellular signalling pathways. Gp130 activates intracellular signalling through the Shp2-ERK1/2 and STAT1/3 pathways to mediate cellular activities. This thesis tests the hypothesis that gp130-mediated signalling is dysregulated in the development and progression of pulmonary fibrosis. To address this hypothesis, I assessed the role of gp130-mediated signalling in a mouse model of bleomycin-induced lung fibrosis. This thesis utilised two novel gp130 mutant mice strains with directed and enhanced gp130-mediated Shp2-ERK1/2 (gp130¿STAT/¿STAT) or STAT1/3 (gp130757F/757F) signalling. I observed complete protection from fibrosis in gp130¿STAT/¿STAT mice up to 60 days after bleomycin treatment and profound fibrosis in gp130757F/757F mice compared to wt controls. The enhanced fibrosis observed in gp130757F/757F mice was diminished by monoallelic deletion of STAT3 (gp130757F/757F;STAT3+/-), identifying gp130-STAT3 signalling as a novel promoter of lung fibrosis. ... In addition, IL-6/11 activation of gp130-mediated signalling modulated transforming growth factor (TGF)-ß-induced effects on adult fibroblast proliferation and myofibroblast differentiation. Interaction between IL-6/11 and TGF-ß1 on fibroblast proliferation was dependent on both the gp130-ERK1/2 and gp130-STAT1/3 pathways. Loss of either pathway abrogated the effects of IL-6 and IL-11 on TGF-ß1- 4 induced fibroblast proliferation. However, it was clear that gp130-STAT3 signalling inhibited TGF-ß1-induced myofibroblast differentiation of primary lung fibroblasts. The inhibition of myofibroblast differentiation was associated with gp130-STAT3 dependent inhibition of TGF-ß1-induced Smad3 phosphorylation. These results indicate that IL-6 and IL-11 promote myofibroblastic differentiation of lung fibroblasts, while gp130-STAT3 signalling inhibits TGF-ß1-induced Smad3 phosphorylation and myofibroblastic differentiation of lung fibroblasts While the pathogenesis of IPF is unknown, it is believed that excessive collagen deposition, aberrant fibroblast behaviour and an inflammatory response are critical to the progression of this disease. It has been shown here that IL-6 family cytokines mediate the development and progression of bleomycin-induced lung fibrosis by increasing collagen synthesis, fibroblast proliferation, myofibroblast differentiation and inflammation through gp130-STAT3 signalling. This thesis has demonstrated that differential activation of cytoplasmic signalling pathways by a membrane bound receptor can have a profound effect on pulmonary responses to injury. Furthermore, this thesis is the first study to identify the gp130-STAT3 pathway as a therapeutic target in the treatment of IPF.

The idiopathic inflammatory myopathies (IIMs) are a heterogeneous group of chronic muscle diseases with frequent extramuscular organ involvement that contributes to serious prognosis. The presence of autoantibodies and composition of muscle infiltrates both support autoimmune nature of the disease and pathogenic role of B lymphocytes. Besides the traditional diagnostic subgroups, autoantibody characterised phenotype subsets have been identified with presumed similar pathogenic mechanisms. The best known is the antisynthetase syndrome which is characterised by presence of myositis, antisynthetase autoantibodies (with anti-Jo-1 being the most frequent), interstitial lung disease and other extramuscular manifestations. BAFF (B cell-Activating Factor of the TNF Family) is a key factor in B cell homeostasis modulation. In high levels, it allows survival of autoreactive B cell clones and thus participates in the pathogenesis of autoimmune diseases. Its expression is induced by type I interferons (IFN-1). The aim of the PhD thesis was to explore the role of BAFF in pathogenesis of IIMs by analysis of its serum levels, the receptors for BAFF in muscle tissue, their associations to IFN-1 and expression of BAFF gene mRNA transcription variants in peripheral blood cells. Further aspect was to study a possible...

A Fibrose Cística (FC) é uma doença letal, de caráter autossômico recessivo, que acomete populações de diferentes etnias. A doença caracteriza-se pelo comprometimento sistêmico das glândulas exócrinas e, na maioria dos pacientes, a doença pulmonar acaba tornando-se a patologia predominante. A infecção por P. aeruginosa é a principal causa de mortalidade dos pacientes com FC. O Sistema de Secreção Tipo III da bactéria é expresso na fase aguda da doença e é responsável por injetar proteínas citotóxicas no interior da célula eucariótica. Há um grande interesse em se investigar a resposta de anticorpos anti P. aeruginosa em pacientes com FC a fim de diagnosticar a colonização e ou infecção pulmonar antes da cultura, permitindo a antibioticoterapia preventiva, a fim de se evitar a infecção pulmonar crônica. Nesta tese, investigamos a resposta de anticorpos (IgG+IgM+IgA) contra as proteínas do SSTT de P. aeruginosa, através do Western-Blot. Participaram do estudo 51 pacientes com FC, de 1.1 a 16.8 anos acompanhados no Departamento de Pneumologia do Instituto Fernandes Figueira - FioCruz, durante um período aproximado de 2 anos. De cada paciente foram coletadas de 1 a 4 amostras de sangue, com intervalo médio de 6 meses entre as coletas. O grupo controle negativo consistiu de 28 indivíduos não fibrocísticos, de 2 a 17 anos, atendidos no Hospital Universitário Pedro Ernesto - HUPE UERJ. As proteínas do SSTT foram extraídas das cepas PAO1 e PAOΔExsA (regulador da expressão do SSTT) de P. aeruginosa. Controles positivos e negativos foram utilizados em todas as reações. Para a identificação das proteínas do SSTT na reação utilizou-se antisoro de camundongos imunizados com a proteína recombinante PcrV. Doze (75%) dos 16 pacientes fibrocísticos considerados não infectados por P. aeruginosa tiveram a primeira sorologia positiva para PopB e 15 (93,75%) para ExoS/ExoT, indicando a colonização ou infecção por P. aeruginosa. Aproximadamente 25% e 35,7% dos soros do grupo controle mostraram reatividade fraca com PopB ou ExoS/ExoT, respectivamente. O tempo decorrido entre a primeira sorologia positiva e o primeiro isolamento de P. aeruginosa nestes pacientes variou de 18 a 30 meses. Concluindo, é possível fazer o diagnóstico sorológico da infecção pulmonar por P. aeruginosa antes do isolamento da bactéria pela cultura. / Cystic Fibrosis (CF) is a lethal disease of autosomal recessive character, which affects people of different ethnicities. The disease is characterized by the involvement of systemic exocrine glands and in most patients, the lung disease becomes the predominant pathology. The infection with P. aeruginosa is the leading cause of mortality in patients with CF. The Type III Secretion System (TTSS) of bacteria is expressed during acute disease and injects cytotoxic proteins inside the host cell. There is a great interesting in investigate the antibody response to P. aeruginosa in CF patients in order to diagnose a pulmonary infection or colonization before the culture. Then, preventive antibiotic treatment can be initiated before the installation of chronic lung infection. We investigated the antibody response (IgG + IgA + IgM) against TTSS proteins of P. aeruginosa by Western-blot. The study included 51 patients with CF, from 1.1 to 16.8 years attending the Pediatric Pulmonology Unit of Fernandes Figueira Institute (IFF) FIOCRUZ, Rio de Janeiro, for a period of approximately 2 years. Most patients had or 4 blood samples collected for antibody analyses. Samples were obtained with a mean interval of 6 months. The negative control group consisted of 28 non-CF individuals, from 2 to 17 years, attended at Pedro Ernesto University Hospital - HUPE - UERJ. The TTSS proteins were extracted from strains PAO1 and PAOΔExsA (regulator of TTSS proteins expression) of P. aeruginosa. Positive and negative controls were used in all reactions. For the identification of TTSS proteins in the reaction we used antisera from mice immunized with the recombinant protein PcrV. Twelve (75%) of 16 CF patients considered not infected by P. aeruginosa had their first serology positive for "PopB" and 15 (93.75%) for "ExoS/ExoT. These results indicated that these patients were colonized or infected by P. aeruginosa. About 25% e 35,7% of negative control sera showed a weak reactivity with PopB or ExoS, respectively. The time between the first positive serology and the first isolation of P. aeruginosa in these patients ranged from 18 to 30 months. In conclusion, it is possible to make a serological diagnosis of pulmonary infection by P. aeruginosa before the isolation of the bacterium by culture.

A Fibrose Cística (FC) é uma doença letal, de caráter autossômico recessivo, que acomete populações de diferentes etnias. A doença caracteriza-se pelo comprometimento sistêmico das glândulas exócrinas e, na maioria dos pacientes, a doença pulmonar acaba tornando-se a patologia predominante. A infecção por P. aeruginosa é a principal causa de mortalidade dos pacientes com FC. O Sistema de Secreção Tipo III da bactéria é expresso na fase aguda da doença e é responsável por injetar proteínas citotóxicas no interior da célula eucariótica. Há um grande interesse em se investigar a resposta de anticorpos anti P. aeruginosa em pacientes com FC a fim de diagnosticar a colonização e ou infecção pulmonar antes da cultura, permitindo a antibioticoterapia preventiva, a fim de se evitar a infecção pulmonar crônica. Nesta tese, investigamos a resposta de anticorpos (IgG+IgM+IgA) contra as proteínas do SSTT de P. aeruginosa, através do Western-Blot. Participaram do estudo 51 pacientes com FC, de 1.1 a 16.8 anos acompanhados no Departamento de Pneumologia do Instituto Fernandes Figueira - FioCruz, durante um período aproximado de 2 anos. De cada paciente foram coletadas de 1 a 4 amostras de sangue, com intervalo médio de 6 meses entre as coletas. O grupo controle negativo consistiu de 28 indivíduos não fibrocísticos, de 2 a 17 anos, atendidos no Hospital Universitário Pedro Ernesto - HUPE UERJ. As proteínas do SSTT foram extraídas das cepas PAO1 e PAOΔExsA (regulador da expressão do SSTT) de P. aeruginosa. Controles positivos e negativos foram utilizados em todas as reações. Para a identificação das proteínas do SSTT na reação utilizou-se antisoro de camundongos imunizados com a proteína recombinante PcrV. Doze (75%) dos 16 pacientes fibrocísticos considerados não infectados por P. aeruginosa tiveram a primeira sorologia positiva para PopB e 15 (93,75%) para ExoS/ExoT, indicando a colonização ou infecção por P. aeruginosa. Aproximadamente 25% e 35,7% dos soros do grupo controle mostraram reatividade fraca com PopB ou ExoS/ExoT, respectivamente. O tempo decorrido entre a primeira sorologia positiva e o primeiro isolamento de P. aeruginosa nestes pacientes variou de 18 a 30 meses. Concluindo, é possível fazer o diagnóstico sorológico da infecção pulmonar por P. aeruginosa antes do isolamento da bactéria pela cultura. / Cystic Fibrosis (CF) is a lethal disease of autosomal recessive character, which affects people of different ethnicities. The disease is characterized by the involvement of systemic exocrine glands and in most patients, the lung disease becomes the predominant pathology. The infection with P. aeruginosa is the leading cause of mortality in patients with CF. The Type III Secretion System (TTSS) of bacteria is expressed during acute disease and injects cytotoxic proteins inside the host cell. There is a great interesting in investigate the antibody response to P. aeruginosa in CF patients in order to diagnose a pulmonary infection or colonization before the culture. Then, preventive antibiotic treatment can be initiated before the installation of chronic lung infection. We investigated the antibody response (IgG + IgA + IgM) against TTSS proteins of P. aeruginosa by Western-blot. The study included 51 patients with CF, from 1.1 to 16.8 years attending the Pediatric Pulmonology Unit of Fernandes Figueira Institute (IFF) FIOCRUZ, Rio de Janeiro, for a period of approximately 2 years. Most patients had or 4 blood samples collected for antibody analyses. Samples were obtained with a mean interval of 6 months. The negative control group consisted of 28 non-CF individuals, from 2 to 17 years, attended at Pedro Ernesto University Hospital - HUPE - UERJ. The TTSS proteins were extracted from strains PAO1 and PAOΔExsA (regulator of TTSS proteins expression) of P. aeruginosa. Positive and negative controls were used in all reactions. For the identification of TTSS proteins in the reaction we used antisera from mice immunized with the recombinant protein PcrV. Twelve (75%) of 16 CF patients considered not infected by P. aeruginosa had their first serology positive for "PopB" and 15 (93.75%) for "ExoS/ExoT. These results indicated that these patients were colonized or infected by P. aeruginosa. About 25% e 35,7% of negative control sera showed a weak reactivity with PopB or ExoS, respectively. The time between the first positive serology and the first isolation of P. aeruginosa in these patients ranged from 18 to 30 months. In conclusion, it is possible to make a serological diagnosis of pulmonary infection by P. aeruginosa before the isolation of the bacterium by culture.

A tolerância pode ser considerada um dos pilares da imunologia. Sabe-se que a tolerância a um antígeno pode gerar tolerância a outro antígeno não relacionado, fenômeno conhecido como tolerância cruzada. Neste trabalho caracterizamos a tolerância cruzada utilizando a OVA como tolerógeno e extrato de Blomia tropicalis (Bt) ou Hemocianina de Keyhole limpet (KLH) como alérgenos. Verificamos que é possível reproduzir o fenômeno da tolerância cruzada neste modelo de inflamação alérgica induzida tanto pelo KLH quanto pela Bt, com diminuição do infiltrado inflamatório no pulmão, eosinófilos, IgE total e produção de muco. Ainda, a estratégia de utilizar a tolerância cruzada terapeuticamente, i.é., após a sensibilização com KLH, a indução de tolerância cruzada não foi capaz de prevenir a resposta alérgica. Em conjunto, nossos dados mostram que a tolerância à OVA modifica as respostas alérgicas tanto à Bt quanto ao KLH no modelo de inflamação pulmonar experimental de forma profilática, mas que a tolerância cruzada não é eficiente em animais já sensibilizados. / Tolerance is among the Immunology pillars. Experimental data indicate that tolerance towards an antigen can promote tolerance to an unrelated antigen, a phenomenon known as cross-tolerance. Here we sought to characterize cross tolerance using OVA as a tolerogen and Blomia tropicalis (Bt) extract or Keyhole limpet Hemocianina (KLH) as allergens. We found that cross tolerance can be reproduced in the model of allergic lung disease induced by KLH or Bt, with less inflammatory infiltrate in the lung, eosinophils, total IgE and mucus production. Using cross tolerance therapeutically, i.e., after KLH sensitization, was not effective, since the allergic lung response was not modulated. Altogether, our data shows that OVA tolerance modulate allergic lung disease induced either by Bt or KLH when used as prophylactic model, however cross tolerance is ineffective in sensitized animals.

Comentando-se algumas evidências da literatura e da análise de dados secundários de morbidade e mortalidade, estabelece-se a hipótese de que as doenças chiadoras recorrentes da infância possam constituir-se em fator de risco para o desenvolvimento de infecções pulmonares. Um estudo caso-controle é desenvolvido para testar esta hipótese reunindo 51 casos de pneumonia pareados por sexo e idade a 51 controles sadios e 51 controles doentes não respiratórios. A amostra é colhida entre pacientes do Hospital Universitário da USP sendo condição de entrada para os casos um diagnóstico de pneumonia adquirida na comunidade e livre de tratamento anterior. Os controles são selecionados dentro da mesma clientela entre pacientes com outro diagnóstico e crianças sadias usuárias dos mesmos serviços, identificadas entre acompanhantes de pacientes. Casos e controles são submetidos a idêntica investigação quanto a presença de doença (pneumonia) e de exposição ao fator de risco investigado (doença chiadora recorrente) através de anamnese e exame físico padronizados, realizados independentemente por dois observadores distintos. Ambos os observadores são pediatras designados pelo Departamento de Pediatria do Hospital para esta tarefa e recebem orientação e supervisão para uma observação padronizada. Os dados assim recolhidos são processados em análises estatísticas uni e multivariadas para explorar diferenças entre casos e controles. A amostra estudada resulta constituída por crianças de idade média de 2 anos (com variação entre um mês e sete anos), entre as quais 47 por cento são meninos. O diagnóstico de pneumonia é validado através da aplicação de análise discriminante multivariada das informações relativas a sinais clínicos, encontrando-se uma compatibilidade entre a conclusão clínica e estes sinais da ordem de pelo menos 75 por cento . O diagnóstico de exposição a doença chiadora é validado pela presença de história compatível segundo premissas pré-estabelecidas nos métodos do estudo (diagnóstico de asma e pelo menos um episódio de dispnéia nos últimos 12 meses ou história de chiado recorrente que melhora com medicação e pelo menos dois episódios nos últimos 12 meses) em 40 dos 41 expostos identificados. O questionário de identificação da exposição tem a repitibilidade medida através de sua reaplicação pelo mesmo observador a uma amostra de 20 por cento do total de crianças examinadas. Encontra-se um nível geral de concordância entre a primeira e segunda aplicação do questionário de 76,7 por cento e um índice Kappa de 0,65. A associação entre pneumonia e doença chiadora recorrente é analisada através de regressão logística com controle para todas as variáveis que em análise univariada mostram frequência estatisticamente significante entre casos e controles. Encontra-se que o risco de pneumonia entre crianças expostas a doença chiadora é 7 vezes maior do que entre crianças não expostas, controladas a renda familiar e a situação de aglomeração no quarto de dormir, também identificadas como fatores de risco para pneumonia (\"odds ratio\" de 5,6 e 2,4 para rendas baixa e média comparadas com renda alta e \"odds ratio\" de 1,5 para cada pessoa a mais no quarto de dormir). Calcula-se que para a comunidade hospitalar estudada a doença chiadora recorrente represente um risco atribuível para pneumonia entre 33 por cento e 51 por cento , conforme a aplicação de diferentes técnicas de cálculo. Conclui-se que as doenças chiadoras recorrentes da infância constituem-se em importante fator de risco para pneumonia e que seu controle, através da inclusão de assistência sistemática a pacientes com este diagnóstico nos programas de Saúde Pública para o controle de doenças respiratórias da infância, pode ter importante impacto sobre a incidência de pneumonias. / Taking into account some evidences from the literature and from analysis of available data, a hyphothesis that childhood wheezing diseases are related to pneunomia is established. A case-control study is designed to test this hypothesis taking 51 cases of pneumonia matched by sex and age to 51 healthy and 51 non-respiratory controls. The sample is drawn from patients of the \"Hospital Universitário da USP\". Entry condition for cases is to bear a community acquired pneumonia free of previous treatment and for controls is to be custommer of the same health services. Cases and controls are equally investigated with regards to the presence of pneumonia and history of wheezing diseases, investigation being conducted by two independent observers. Both are paediatricians selected by the Hospital Paediatric Department and are dully trained and supervised as to assure a standardized observation. Data are processed in oneway and multivariate statistical analyses to explore diferences between cases and controls. The sample studied comprises children aged 2 years in average (range between one month and seven years) and male subjects account for 47 per cent of the total. The diagnosis of pneumonia is validated through multivariate discriminant analysis which shows that clinical opinion is compatible to clinical signs in at least 75 per cent of the cases. The clinical conclusion of presence of wheezing disease is found compatible to previously defined criteria in 40 out of 41 patients. Repeatability of such information is assessed by re-aplication of the questionnaire for wheezing disease investigation to a sample of 20 per cent of the total number of children, which is carried out by the same observer of the first interview. An overall agreement of 76.7 per cent and a Kappa of 0.65 is found. Association between pneumonia and wheezing disease is analysed through logistic regression controlling the effects of all variables which have shown statistically significant differences between cases and controls. It is found that children who bear a wheezing disease have a risk to pneumonia which is 7 times greater than those who do not, allowing for socioeconomic status and bedroom crowding, both also risk fators: odds ratio of 5.6 and 2.4 for low and medium family income as opposed to high income and odds ratio of 1.5 for each increase of one person in bedroom.Derived from different techniques, an attributable risk for wheezing disease ranging from 33 per cent to 51 per cent is calculated. It is concluded that wheezing diseases of childhood are an important risk factor to pneumonia and that its control, by means of regular medical assistance of patients being included among the items of current public health programmes, should result in an important effect over the frequency of pneumonia.

O desenvolvimento tecnológico proporciona uma melhoria na qualidade de vida devido à facilidade, rapidez e flexibilidade no acesso à informação. Na área biomédica, a tecnologia é reconhecidamente uma importante aliada, permitindo o rápido desenvolvimento de métodos e técnicas que auxiliam o profissional na atenção à saúde. Recentes avanços na análise computadorizada de imagens médicas contribuem para o diagnóstico precoce de uma série de doenças. Nesse trabalho é apresentada uma metodologia para o desenvolvimento de um sistema computacional para caracterização de padrões em imagens pulmonares, baseado em técnicas de redes neurais artificiais. No estudo, buscou-se verificar a utilização de redes neurais auto-organizáveis como ferramenta de extração de atributos e redução de dimensionalidade de imagens radiográficas de tórax, objetivando a caracterização de lesões intersticiais de pulmão. Para a redução de dimensionalidade e extração de atributos, implementou-se um algoritmo baseado nos mapas auto-organizáveis (SOM), com algumas variações, obtendo-se uma redução dos cerca de 3 milhões de pixels que compõe uma imagem, para 240 elementos. Para a classificação dos padrões, utilizou-se uma rede Perceptron multi-camadas (MLP), validada com a metodologia leave-one-out. Com uma base contendo 79 exemplos de padrão linear, 37 exemplos de padrão nodular, 30 exemplos de padrão misto, e 72 exemplos de padrão normal, o classificador obteve a média de 89,5% de acerto, sendo 100% de classificação correta para o padrão linear, 67,5% para o padrão nodular, 63,3% para o padrão misto, e 100% para o padrão normal. Os resultados obtidos comprovam a validade da metodologia. / The technological development provides an improvement in the quality of life due to easiness, speed and flexibility in the access to the information. In the biomedical area, the technology is admitted as an important allied, allowing the fast development of methods and techniques that assist the professional in the health care. Recent advances in the computerized analysis of medical images contribute for the precocious diagnosis of a series of diseases. In this work a methodology for the development of a computational system for characterization of patterns in pulmonary images, based in techniques of artificial neural networks is presented. In the study, has searched for the verification the use of self-organizing neural networks as a feature extraction and dimensionality reduction tool of chest radiographs, willing to characterize interstitial lung disease. For the dimensionality reduction and feature extraction, an algorithm based on Self-Organizing Maps (SOM) was implemented, with some variations, getting a reduction of about 3 million pixels that it composes an image, for 240 elements. For the pattern classification, a Multilayer Perceptron (MLP) was used, validated with the leave-one-out methodology. With a database containing 79 samples of linear pattern, 37 samples of nodular pattern, 30 samples of mixed pattern, and 72 samples of normal pattern, the classifier provided an average result of 89.5% of right classification, with 100% of right classification for linear pattern, 67.5% for nodular pattern, 63.3% for mixed pattern, and 100% for normal pattern. The results prove the validity of the methodology.

Orientador: Lourenço Sbragia Neto / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas / Made available in DSpace on 2018-08-12T11:31:54Z (GMT). No. of bitstreams: 1 Nassr_AzizeCristinaCapelli_D.pdf: 6642584 bytes, checksum: 60fb43046c34664559a19ebb364b282b (MD5) Previous issue date: 2008 / Resumo: A Hérnia Diafragmática Congênita (HDC) é um defeito da formação do músculo diafragma que incide em aproximadamente 1:2500 nascidos vivos e apresenta altos índices de mortalidade fetal e neonatal decorrentes da hipoplasia e da hipertensão pulmonares. Este defeito pode ser induzido experimentalmente em ratas grávidas administrando o herbicida nitrofen que causa HDC em 24% dos fetos. A análise microscópica do pulmão da HDC demonstra a presença de hipolasia pulmonar além de alveolização e vascularização alterada. Um dos fatores de crescimento envolvidos no desenvolvimento vascular é o VEGF (vascular endothelial growth factor) e seus receptores, no entanto ainda não se conhece como a expressão desta glicoproteína e de seus receptores varia ao longo do desenvolvimento pulmonar fetal nesta doença. Utilizando o modelo experimental de HDC induzido pelo nitrofen (2,4-dicloro-4'nitrodifenil éter) investigamos o grau de hipoplasia pulmonar e por meio de análise imunoistoquímica, comparamos a expressão dos receptores para o VEGF em três fases do desenvolvimento pulmonar, pseudoglandular, canalicular e sacular de fetos de ratos normais e com HDC. Dividimos o experimento em ratas da raça Sprague-Dawley em três grupos: controle externo (CE), exposto ao óleo de oliva (OO) e expostas ao nitrofen com e sem HDC. Estudamos quatro grupos de 20 fetos cada em cinco dias gestacionais (DG) diferentes 17,5, 18,5, 19,5, 20,5 e 21,5. As variáveis morfológicas estudadas foram: peso corporal (PC), peso pulmonar total (PPT), peso do pulmão esquerdo (PPE), relação PPT/PC, volume pulmonar total (VPT) e volume do pulmão esquerdo (VPE). As variáveis histométricas estudadas foram: parênquima pulmonar (Par), espaço aéreo (EA), densidade do parênquima (DAP) e volume do parênquima do pulmão esquerdo. A avaliação imumohistoquímica foi realizada por meio da contagem de pontos de receptor de VEGFR-1 e 2. Obtivemos 37 % (100/270) de HDC nas ratas expostas ao nitrofen, todas variáveis morfológicas e histométricas indicam diminuição dos resultados no grupo nitrofen com e sem HDC em relação aos demais, mas que se acentuam mais ainda no grupo HDC. Essas alterações são mais evidentes a partir dos DG 18,5 e 19,5. A imunomarcação para os receptores VEGFR-1 aumentou nos grupos nitrofen e foram progressivamente maiores no grupo nitrofen com HDC (p<0,005) que os fetos dos grupo CE e OO a partir do dia gestacional 17,5, fase pseudoglandular com pico máximo no dia gestacional 19,5. O mesmo ocorreu com os receptores de VEGFR-2 a partir do dia gestacional 17,5, fase pseudoglandular até o dia 21,5 fase sacular do desenvolvimento pulmonar. Concluímos que o modelo é valido e que os fetos expostos ao nitrofen com e sem HDC apresentam hipoplasia pulmonar primária sendo mais acentuada nos fetos portadores de HDC. O mesmo resultado ocorre com imunomarcação para os receptores de VEGFR-1 e 2 que foram maiores na HDC. / Abstract: The Congenital Diaphragmatic Hernia (CDH) is a defect in the embryogenesis of the diaphragm with an incidence of 1:2500 liveborns and high fetal and neonatal mortality due to pulmonary hypoplasia and hypertension. This defect can be experimentally induced in fetuses of pregnant rats by the administration of Nitrofen, an herbicide that causes CDH in 24% of the fetuses. The histology of lungs in CDH shows pulmonary hipoplasia and not only the alveolarization but also the vascularization are affected. These changes lead to a high neonatal mortality because of the thickening of the middle layer of the arterioles causing pulmonary hypertension. One of the factors involved in the growth of the arterioles is VEGF (vascular endothelial growth factor) and its receptors; however, it is not known how the expression of this glycoprotein and its receptors change during lung development in this disease. In Brazil, the experimental model has never been tested. So, we tested the model and verified the degree of pulmonary hipoplasia and, using imunohystochemistry, we compared the expression of the receptor of VEGF in three different stages of lung development, pseudoglandular, canalicular and saccular, of normal rat fetuses and fetuses with CDH. Female Sprague-Dawley rats were divided in three groups: external control (EC), exposed to olive oil (OO) and exposed to nitrofen (N). We studied four groups - EC, OO, N with CDH and N without CDH - with 20 fetuses in each five different gestational days (GD) 17,5, 18,5, 19,5, 20,5, 21,5. The morphologic variables studied were: body weight (BW), total lung weight (TLW), left lung weight (LLW), relationship TLW/BW, total lung volume (TLV) and left lung volume (LLV). The hystometric variables studied were: lung parenchyma (LP), air space (AS), left lung parenchyma density (PD) and left lung parenchyma volume (PV). The immunohystochemistry variables were: points positive and negative for the receptor for VEGF 1 and 2. We had 37% (100/270) of CDH frequency in the fetuses exposed to nitrofen. All the morphological and hystometrical variables show a reduction in the nitrofen group with and without CDH, which were more pronounced in the group of fetuses with CDH. These changes are more evident from the GD 18,5 and 19,5 on. The receptors VEGFR-1 e 2 are increased in the nitrofen groups with and without CDH, but this increase is higher in the fetuses with CDH. We conclude that the model is valid and that the fetuses exposed to nitrofen with and without CDH show primary pulmonary hypoplasia that is more pronounced in CDH, the same is also observed in the receptors of VEGFR-1 and 2. / Doutorado / Pesquisa Experimental / Doutor em Cirurgia