INTRODUÇÃO: Recentes estudos avaliam o uso da solução salina hipertônica na concentração de 3% no tratamento de pacientes com traumatismos cranioencefálicos, entretanto, poucos trabalhos têm analisado a sua eficácia no tratamento do choque hemorrágico. O objetivo deste trabalho é avaliar os efeitos do tratamento do choque hemorrágico com a solução salina hipertônica a 3%, analisando principalmente seus possíveis efeitos benéficos na atenuação da resposta inflamatória decorrrente do choque. Para tal, esta solução será comparada a outras duas, amplamente estudadas: a solução salina hipertônica a 7,5% e a solução de Ringer lactato. MÉTODOS: Foram utilizados, neste estudo, 26 ratos Wistar. Os animais foram anestesiados com pentobarbital sódico por via intraperitoneal (50 mg/Kg) e, então, submetidos a um protocolo de choque hemorrágico controlado. Neste protocolo, os animais foram sangrados até que fosse atingida uma pressão arterial média (PAM) de 35 mmHg, em um período de 10 minutos, sendo este nível de PAM mantido por 50 minutos. Ao término deste período de choque, os animais foram randomizados em três grupos para reposição volêmica: reposição com solução de Ringer lactato (grupo RL, n=7), na dose de 33 mL/Kg; reposição com solução salina hipertônica a 3% (grupo SH3%, n=7), na dose de 10 mL/Kg; reposição com solução salina hipertônica a 7,5% (grupo SH7,5%, n=7), na dose de 4 mL/Kg. Após a infusão das soluções, metade do volume de sangue retirado foi reinfundido em todos os animais. Sangue arterial foi coletado para análise de gasometria, lactato, concentração plasmática de sódio e osmolaridade plasmática. Para avaliação da resposta inflamatória, os animais foram sacrificados quatro horas após o início do experimento, sendo obtidas amostras de sangue para determinação das concentrações plasmáticas de interleucina (IL) -6 e fator de necrose tumoral (TNF) -alfa. Amostras de tecido pulmonar e intestinal foram obtidas para avaliação histopatológica de lesão orgânica, sendo as lâminas analisadas por dois patologistas sem conhecimento prévio dos grupos, determinando-se, assim, um escore de lesão baseado em um sistema de pontuação. RESULTADOS: Todos os animais submetidos à reposição volêmica apresentaram valores adequados de PAM ao término do tratamento. Nos animais tratados com as duas concentrações de solução salina hipertônica, a concentração plasmática de sódio e o valor da osmolaridade plasmática foram significativamente maiores, quando comparados aos grupos CT e RL. A concentração plasmática de TNF-alfa foi significativamente maior nos animais tratados com a solução de Ringer lactato, não havendo, para tanto, diferenças estatísticas entre os grupos CT, SH3% e SH7,5%. Em relação a IL-6, não se observou diferenças estatisticamente significantes entre os grupos CT, SH3% e SH7,5%, sendo a concentração plasmática deste mediador inflamatório significativamente elevada no grupo RL, quando comparado ao grupo CT. O escore de lesão pulmonar foi significativamente maior no grupo RL, em comparação aos grupos SH3% e SH7,5% (5,7 ± 0,7, 2,7 ± 0,5, 2,1 ± 0,4, respectivamente). Os animais dos grupos SH3% e SH7,5% apresentaram atenuação da lesão intestinal pós-choque em comparação aos animais do grupo RL (2,3 ± 0,4, 2,0 ± 0,6, 5,9 ± 0,6, respectivamente). CONCLUSÕES: O tratamento do choque hemorrágico com as duas concentrações de solução salina hipertônica resultou em atenuação da resposta inflamatória pós-choque. A solução salina hipertônica a 3% apresentou efeitos metabólicos e imunológicos semelhantes à solução salina hipertônica a 7,5%, sendo ambas superiores em relação aos parâmetros estudados à solução de Ringer lactato. / BACKGROUND: Recent studies have been conducted examining the efficacy of 3% hypertonic saline solution (HSS) in traumatic brain injury; however, few studies have analyzed the effects of 3%HSS during hemorrhagic shock. The aim of this study was to test the potential immunomodulatory benefits of 3%HSS resuscitation over standard fluid resuscitation. METHODS: Wistar rats were bled to a mean arterial pressure (MAP) of 35 mmHg and then randomized in 3 groups: LR (lactated Ringer, 33mL/Kg, n=7), 3%HSS (10mL/Kg, n=7) and 7.5%HSS (4mL/Kg, n=7). Half of the shed blood was infused after fluid resuscitation. Animals who did not undergo shock served as controls (CT,n=5). Four hours after HS, blood was collected for evaluation of tumor necrosis factor (TNF)-alpha and interleukin (IL)-6 by enzyme immunoassay. Lung and intestinal samples were obtained for histopathological analysis. RESULTS: Animals in HSS groups had significantly higher MAP than LR one hour after treatment. Osmolarity and sodium levels were markedly elevated in HSS groups. TNF-alpha and IL-6 levels were similar between CT and HSS groups, but significantly higher in LR (p<0.05). Lung injury score was significantly higher in LR when compared to 7.5%HSS and 3%HSS (5.7 ± 0.7, 2.1 ± 0.4 and 2.7 ± 0.5, respectively). Intestinal injury was attenuated in the 7.5%HSS and 3%HSS groups when compared to LR (2.0 ± 0.6, 2.3 ± 0.4 and 5.9 ± 0.6, respectively). CONCLUSIONS: Small volume resuscitation strategy modulates the inflammatory response and decrease the end-organ damage after HS. 3%HSS provides immunomodulatory and metabolic effects similar to those observed with conventional concentration of HSS.

Tao Dacheng. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (leaves 70-75). / Abstracts in English and Chinese. / Chapter Chapter 1 --- Introduction --- p.1 / Chapter 1.1 --- Content-based Image Retrieval --- p.1 / Chapter 1.2 --- SVM based RF in CBIR --- p.3 / Chapter 1.3 --- DA based RF in CBIR --- p.4 / Chapter 1.4 --- Existing CBIR Engines --- p.5 / Chapter 1.5 --- Practical Applications of CBIR --- p.10 / Chapter 1.6 --- Organization of this thesis --- p.11 / Chapter Chapter 2 --- Statistical Learning Theory and Support Vector Machine --- p.12 / Chapter 2.1 --- The Recognition Problem --- p.12 / Chapter 2.2 --- Regularization --- p.14 / Chapter 2.3 --- The VC Dimension --- p.14 / Chapter 2.4 --- Structure Risk Minimization --- p.15 / Chapter 2.5 --- Support Vector Machine --- p.15 / Chapter 2.6 --- Kernel Space --- p.17 / Chapter Chapter 3 --- Discriminant Analysis --- p.18 / Chapter 3.1 --- PCA --- p.18 / Chapter 3.2 --- KPCA --- p.18 / Chapter 3.3 --- LDA --- p.20 / Chapter 3.4 --- BDA --- p.20 / Chapter 3.5 --- KBDA --- p.21 / Chapter Chapter 4 --- Random Sampling Based SVM --- p.24 / Chapter 4.1 --- Asymmetric Bagging SVM --- p.25 / Chapter 4.2 --- Random Subspace Method SVM --- p.26 / Chapter 4.3 --- Asymmetric Bagging RSM SVM --- p.26 / Chapter 4.4 --- Aggregation Model --- p.30 / Chapter 4.5 --- Dissimilarity Measure --- p.31 / Chapter 4.6 --- Computational Complexity Analysis --- p.31 / Chapter 4.7 --- QueryGo Image Retrieval System --- p.32 / Chapter 4.8 --- Toy Experiments --- p.35 / Chapter 4.9 --- Statistical Experimental Results --- p.36 / Chapter Chapter 5 --- SSS Problems in KBDA RF --- p.42 / Chapter 5.1 --- DKBDA --- p.43 / Chapter 5.1.1 --- DLDA --- p.43 / Chapter 5.1.2 --- DKBDA --- p.43 / Chapter 5.2 --- NKBDA --- p.48 / Chapter 5.2.1 --- NLDA --- p.48 / Chapter 5.2.2 --- NKBDA --- p.48 / Chapter 5.3 --- FKBDA --- p.49 / Chapter 5.3.1 --- FLDA --- p.49 / Chapter 5.3.2 --- FKBDA --- p.49 / Chapter 5.4 --- Experimental Results --- p.50 / Chapter Chapter 6 --- NDA based RF for CBIR --- p.52 / Chapter 6.1 --- NDA --- p.52 / Chapter 6.2 --- SSS Problem in NDA --- p.53 / Chapter 6.2.1 --- Regularization method --- p.53 / Chapter 6.2.2 --- Null-space method --- p.54 / Chapter 6.2.3 --- Full-space method --- p.54 / Chapter 6.3 --- Experimental results --- p.55 / Chapter 6.3.1 --- K nearest neighbor evaluation for NDA --- p.55 / Chapter 6.3.2 --- SSS problem --- p.56 / Chapter 6.3.3 --- Evaluation experiments --- p.57 / Chapter Chapter 7 --- Medical Image Classification --- p.59 / Chapter 7.1 --- Introduction --- p.59 / Chapter 7.2 --- Region-based Co-occurrence Matrix Texture Feature --- p.60 / Chapter 7.3 --- Multi-level Feature Selection --- p.62 / Chapter 7.4 --- Experimental Results --- p.63 / Chapter 7.4.1 --- Data Set --- p.64 / Chapter 7.4.2 --- Classification Using Traditional Features --- p.65 / Chapter 7.4.3 --- Classification Using the New Features --- p.66 / Chapter Chapter 8 --- Conclusion --- p.68 / Bibliography --- p.70

Wang Ying. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (leaves 124-135). / Abstracts in English and Chinese. / Acknowledgements / Abstract / 摘要 / Table of contents / List of figures / List of tables / List of abbreviations / CHAPTER / Chapter 1. --- Introduction / Chapter 1.1 --- Background of SARS and epidemiology / Chapter 1.2 --- SARS symptoms and infected regions / Chapter 1.3 --- SARS virus / Chapter 1.4 --- Treatment for SARS at present / Chapter 1.5 --- Vaccine development is a more effective way to fight against SARS / Chapter 1.6 --- Vaccine candidates / Chapter 1.6.1 --- Truncated S protein as a vaccine candidate / Chapter 1.6.2 --- Full-length N protein as a vaccine candidate / Chapter 1.7 --- E.coli expression system / Chapter 1.8 --- Baculovirus expression system / Chapter 1.8.1 --- Characteristics of baculovirus / Chapter 1.8.2 --- Infection cycle of baculovirus / Chapter 1.8.3 --- Control of viral gene expression in virus-infected cells / Chapter 1.8.4 --- Merits of baculovirus expression system / Chapter 1.9 --- Aim of study / Chapter 2. --- "Bacterial expression and purification of rS1-1000(E), rS401-1000(E) and rN(E)" / Chapter 2.1 --- Introduction / Chapter 2.2 --- Materials / Chapter 2.2.1 --- Reagents for bacterial culture / Chapter 2.2.2 --- Reagents for agarose gel electrophoresis / Chapter 2.2.3 --- 2'-deoxyribonucleoside 5'-triphosphate (dNTP) mix for polymerase chain reaction (PCR) / Chapter 2.2.4 --- Sonication buffer / Chapter 2.2.5 --- Reagents for immobilized metal affinity chromatography (IMAC) purification / Chapter 2.2.6 --- Reagents for gel filtration chromatography / Chapter 2.2.7 --- Reagents for sodium dodecylsulphate polyacrylamide gel electrophoresis (SDS-PAGE) / Chapter 2.2.8 --- Reagents for Western blotting / Chapter 2.3 --- Methods / Chapter 2.3.1 --- General techniques in molecular cloning / Chapter 2.3.2 --- "PCR amplification of the S1-400,S401-1000" / Chapter 2.3.3 --- Construction of clone pET-S 1-400 and PET-s401-1000 / Chapter 2.3.4 --- Construction of clone pAC-N / Chapter 2.3.5 --- Expression / Chapter 2.3.6 --- Inclusion bodies preparation / Chapter 2.3.7 --- Inclusion bodies solubilization using urea / Chapter 2.3.8 --- Protein refolding by rapid dilution and dialysis / Chapter 2.3.9 --- Purification of recombinant protein by nickel ion chelating Sepharose fast flow column (IMAC) / Chapter 2.3.10 --- Gel filtration chromatography for further purification / Chapter 2.3.11 --- Bradford assay for the protein concentration analysis / Chapter 2.3.12 --- Protein analysis / Chapter 2.4 --- Results / Chapter 2.4.1 --- SDS-PAGE analysis of the expressed proteins / Chapter 2.4.2 --- Western blot analysis of the bacterial cell lysate / Chapter 2.4.3 --- Protein purification by IMAC / Chapter 2.4.4 --- Purification of rS401-1000(E) by gel filtration / Chapter 2.4.5 --- Determination of production yield of recombinant fusion proteins / Chapter 2.5 --- Discussion / Chapter 2.5.1 --- Expression vector selected for rS1-400(E) and rS401-1000(E) expression / Chapter 2.5.2 --- Protein expression in E.coli / Chapter 2.5.3 --- Purification process / Chapter 3. --- Baculovirus expression and purification of rS401-1000(ACN) and rN(BMN) protein / Chapter 3.1 --- Introduction / Chapter 3.2 --- Materials / Chapter 3.2.1 --- Reagents for insect cell culture and virus work / Chapter 3.3 --- Methods / Chapter 3.3.1 --- "PCR amplification of N and cloning of S401-1000, N genes into the transfer vector pVL1393" / Chapter 3.3.2 --- Cloning of S401-1000 into transfer vector pFastBac HT B / Chapter 3.3.3 --- Virus works / Chapter 3.3.4 --- Identification of recombinant BmNPV or AcMNPV / Chapter 3.3.5 --- Manipulation of silkworm / Chapter 3.3.6 --- Mouse immunization for polyclonal antibody against rN(E) protein / Chapter 3.4 --- Results / Chapter 3.4.1 --- Expression of rN(BMN) in baculovirus / Chapter 3.4.2 --- Expression of rS401-1000(BMN) and rS401-1000(ACN) in baculovirus / Chapter 3.5 --- Discussion / Chapter 3.5.1 --- The expression level of rN(BMN) in both in vitro and invivo / Chapter 3.5.2 --- The rS401-1000(ACN) protein expression level in vitro / Chapter 3.5.3 --- Failure in generating rS401-1000(BMN) / Chapter 3.5.4 --- Purification process of rN(BMN) by IMAC / Chapter 4. --- "Characterization of recombinant rS1-400(E), rN(E), rN(BMN), rS401_1000(E) and rS401-1000(ACN)" / Chapter 4.1 --- Introduction / Chapter 4.2 --- Materials / Chapter 4.2.1 --- Reagents for enzyme-linked immunosorbent assay (ELISA) / Chapter 4.2.2 --- Reagents for purification of human IgG / Chapter 4.2.3 --- Source and identity of Immune sera / Chapter 4.3 --- Methods / Chapter 4.3.1 --- ELISA / Chapter 4.3.2 --- Purification process of human IgG / Chapter 4.4 --- Results / Chapter 4.4.1 --- Validation of Immune sera using SARS viral lysate / Chapter 4.4.2 --- Immunoreactivities of rS1-400(E) and rN(E) against pooled patients sera and normal human serum / Chapter 4.4.3 --- Immunoreactivity comparison of rN(E) and rN(BMN) / Chapter 4.4.4 --- Comparison of the immunoreactivities of rS401-1000(E) and rS401-1000(ACN) / Chapter 4.4.5 --- Immunoreactivity of SARS related proteins against Anti-SARS Antibody (Equine) / Chapter 4.5 --- Discussion / Chapter 4.5.1 --- Comparison of the immunoreactivities of SARS related proteins expressed in the present study / References

I established the phage antibody library platform for the identification of specific antibodies. In the first part of my study, I tried to identify antibody against SARS-CoV. Two fragments on the spike protein, which is responsible for inducing viral entry, was chosen as target for the selection of antibody. An antibody was identified which can selectively recognize the SARS-CoV infected cells, but not non-infected cells. Although this antibody was found to retain no neutralizing ability, this specific antibody may have potential to develop for diagnostic purpose. / I utilized the phage system-based cloning method as an attractive approach to screen and identify virus-specific antibodies that can be encoded by the human genome. Once a useful phage clone is identified, unlimited amounts of human monoclonal virus-specific antibodies can be manufactured, and potentially applied clinically for prophylactic and therapeutic uses. The study focuses on two of these new infections, both of which cause severe respiratory disease: SARS and avian influenza. / Identification of specific antibodies, either for diagnostic or therapeutic use, was successfully demonstrated in the two infectious disease models. The phage antibody platform offers a fast and cost-effective method to identify phage antibodies, which can easily be converted to human viral specific monoclonal antibodies for clinical use. / In the 21st century, a number of novel infectious diseases emerged suddenly and spread rapidly, endangering the lives and well-being of people around the world. Severe acute respiratory syndrome (SARS) is a life threatening form of atypical pneumonia that ravaged Hong Kong, Taiwan, China, Canada and many cities in 2003. In the same year, novel avian influenza viruses infected human beings on two continents. Both of these diseases originated in animals and crossed over into the human population. These emerging diseases pose significant public health threats while providing a chilling reminder that another influenza pandemic could occur at any time. Thus, the development of effective therapeutics to control the disease is of paramount importance. Although several vaccines against SARS and avian influenza are available nowadays, the poor clinical performance and frequent mutation of viral strains may limit the practical use and value of the vaccines. Moreover, there are no promising antiviral drugs available for the treatment. Therefore, I aimed to develop an immunotherapy as an alternative treatment option against these diseases. / In the second part of my study, the extracellular domain of matrix protein of avian influenza virus was chosen as target for the selection of antibody. I successfully identified an antibody which can neutralize the avian influenza virus infection. This promising result indicated this antibody has potential to develop for therapeutic use and these antibodies can be easily manufactured in unlimited amounts for clinical application. / Leung, Ka Man. / Adviser: Kwok Pui Fung. / Source: Dissertation Abstracts International, Volume: 71-01, Section: B, page: 0212. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2009. / Includes bibliographical references (leaves 112-123). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. Ann Arbor, MI : ProQuest Information and Learning Company, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese.

A disfunção de vias aéreas em pacientes com Síndrome do Desconforto Respiratória Agudo (SDRA) é caracterizada por limitação do fluxo expiratório e hiperinsuflação dinâmica. As alterações morfológicas possivelmente associadas com tais alterações funcionais têm sido investigadas em modelos experimentais de lesão pulmonar aguda, que mostram necrose e descamação epitelial em vias aéreas distais. Entretanto, até o momento, essa avaliação não foi realizada em humanos. O objetivo deste estudo foi investigar as alterações estruturais e inflamatórias nas pequenas vias aéreas de pacientes com SDRA. Com este propósito, estudamos, retrospectivamente, o tecido pulmonar de 31 pacientes com SDRA (A: PaO2/FIO2<=200, 45±14anos, 16 homens) e 11 controles (C: 52±16anos, 7 homens) submetidos à autópsia. Por meio de análise de imagem, quantificamos a extensão das alterações epiteliais, a inflamação bronquiolar, a espessura da parede da via aérea e o conteúdo de proteínas da matriz extracelular (MEC) nas pequenas vias aéreas. As vias aéreas dos pacientes com SDRA apresentaram menor extensão de epitélio normal (A: 32,9±27.2%, C: 76,7±32.7%, p<0,001), maior extensão de descamação epitelial (A: 52,6±35.2%, C: 21,8±32.1%, p<0,01), maior índice de inflamação [A: 1(3), C: 0(1), p= 0,03], maior espessura da parede da via aérea (A: 138,7 ± 54,3 ?m, C: 86,4 ± 33,3 ?m, p< 0,01) e maior conteúdo de colágeno I, fibronectina, versicam e MMP-9 comparado aos controles (p<=0,03). Nos pacientes com SDRA, a extensão de epitélio normal apresentou correlação positiva com a PaO2/FiO2 (r=0,58; p=0,02) e correlação negativa com a pressão de platô utilizada (r=-0,52; p=0,04). A extensão de epitélio descamado apresentou correlação negativa com a PaO2/FiO2 (r=-0,52; p=0,04). Nossos dados mostram que as pequenas vias aéreas dos pacientes com SDRA apresentam alterações estruturais caracterizadas por descamação epitelial, inflamação e espessamento da parede com deposição de MEC. Estas alterações podem contribuir para as alterações funcionais observadas em pacientes com SDRA. / Airway dysfunction in patients with acute respiratory distress syndrome (ARDS) is evidenced by expiratory flow limitation and dynamic hyperinflation. The morphological alterations potentially associated with these functional changes have been investigated in experimental models of Acute Lung Injury, which show epithelial necrosis and denudation in distal airways. To date, however, no study has focused on the morphological airway changes in lungs from human subjects with ARDS. Objective: To evaluate structural and inflammatory changes in distal airways in ARDS patients. Methods and Results: We retrospectively studied autopsy lung tissue from 31 ARDS patients (A: PaO2/FIO2<=200, 45±14years, 16 males) and 11 controls (C: 52±16years, 7 males). Using image analysis, we quantified the extension of epithelial changes, bronchiolar inflammation, airway wall thickness, and extracellular matrix (ECM) protein content in distal airways. ARDS airways showed a shorter extension of normal epithelium (A:32.9±27.2%, C:76.7±32.7%, p<0.001), a larger extension of epithelium denudation (A:52.6±35.2%, C:21.8±32.1%, p<0.01), increased airway inflammation (p=0.03), higher airway wall thickness (A:138.7±54.3?m, C:86.4±33.3?m, p<0.01), and higher airway content of collagen I, fibronectin, versican and MMP-9 compared to controls (p<=0.03). The extension of normal epithelium showed a positive correlation with PaO2/FiO2 (r=0.58; p=0.02) and a negative correlation with plateau pressure (r=-0.52; p=0.04). The extension of denuded epithelium showed a negative correlation with PaO2/FiO2 (r=-0.52; p=0.04). Conclusion: Structural changes in small airways of patients with ARDS were characterized by epithelial denudation, inflammation and airway wall thickening with ECM remodeling. These changes are likely to contribute to functional airway changes in patients with ARDS.

Alguns estudos sugerem que as pequenas vias aéreas têm um papel importante na fisiopatologia da lesão pulmonar aguda/ síndrome do desconforto respiratório agudo (LPA/SDRA). O epitélio respiratório que reveste as vias aéreas é capaz de liberar mediadores inflamatórios e está relacionado ainda com a produção de surfactante nas vias aéreas. Até o presente momento, existem poucos estudos que avaliaram se estas funções do epitélio que reveste as pequenas vias aéreas encontram-se alteradas na SDRA. No presente estudo, nós mensuramos a expressão da proteína de surfactante (PS) A e PS-B, a expressão de citocinas inflamatórias interleucina (IL)-6 e IL-8, e um índice de apoptose do epitélio que reveste as pequenas vias aéreas de pacientes com SDRA que foram submetidos a autópsia e comparamos estes resultados com os de indivíduos controle. Foram incluídos no estudo pulmões de autópsia de 31 pacientes com SDRA (PaO2/FiO2200, 45±14 anos, 16 homens) e 11 controles (52±16 anos, 7 homens). A expressão de IL-6, IL-8, PS-A e PS-B no epitélio das pequenas vias aéreas (diâmetro2.0mm) foi verificada através de reações de imunohistoquímica e análise de imagem. O índice de apoptose epitelial das vias aéreas foi avaliado através do método de TUNEL e da expressão de FAS/FASL. Avaliou-se ainda a densidade de células inflamatórias positivas para IL-6 e IL-8 na parede das pequenas vias aéreas. As vias aéreas dos pacientes com SDRA apresentaram maior expressão epitelial de IL-8 (p=0,006) e maior densidade de células inflamatórias expressando IL-6 (p=0,004) e IL-8 (p<0,001) quando comparadas com o grupo controle. Não houve diferenças na expressão epitelial de PS-A e PS-B ou no índice de apoptose epitelial entre os grupos SDRA e controle. Nossos resultados mostram que as pequenas vias aéreas participam da inflamação pulmonar de pacientes com SDRA, caracterizada pelo aumento na expressão de interleucinas próinflamatórias tanto em células inflamatórias da parede da via aérea quanto no epitélio. Nossos resultados sugerem ainda que a apoptose não é um mecanismo importante de morte de células epiteliais das vias aéreas de pacientes com SDRA / Recent studies suggest a role for distal airway injury in the pathophysiology of human ALI/ARDS. The epithelium lining the airways modulates airway function secreting a large number of molecules such as surfactant components and inflammatory mediators. So far, there is little information on how these secretory functions of the small airways are altered in ARDS. In the present study we assessed the airway expression of surfactant protein (SP) A and SP B, the expression of inflammatory cytokines IL-6 and IL-8, and an index of airway epithelial apoptosis of patients with ARDS submitted to autopsy and compared the results with those of control subjects. We studied autopsy lungs of 31 ARDS patients (PaO2/FiO2200, 45±14 years, 16 males) and 11 controls (52±16 years, 7 males). Using immunohistochemistry and image analysis, we quantified the expression of IL-6, IL-8 and SP-A and SP-B in the epithelium of small airways (diameter2.0mm). Airway epithelial apoptosis index was obtained with the TUNEL assay and FAS/FASL expression. We also quantified the density of inflammatory cells expressing IL-6 and IL-8 within the small airway walls. ARDS airways showed an increase in the epithelial expression of IL-8 (p=0.006) and an increased density of inflammatory cells expressing IL-6 (p=0.004) and IL-8 (p<0.001) when compared to controls. There were no differences in SP-A and SP-B epithelium expression or in epithelial apoptosis index between ARDS and controls. Our results show that the distal airways are involved in ARDS lung inflammation with higher expression of pro-inflammatory interleukins in both airway epithelial and inflammatory cells. Our results also suggest that apoptosis is not a major mechanism of airway epithelial cell death in ARDS

INTRODUÇÃO: Existem muitas controvérsias, nos estudos epidemiológicos existentes, a respeito da incidência e desfechos da síndrome de lesão pulmonar aguda. A incidência e as características clínicas da síndrome dependem principalmente da definição utilizada e da metodologia empregada no estudo, bem como da disponibilização e utilização dos leitos nas unidades de terapia intensiva da região estudada. Pela ausência de dados epidemiológicos existentes de lesão pulmonar aguda na Grande Vitória, no Espírito Santo, realizamos este estudo para analisar a incidência, características, sobrevida aos 28 dias e mortalidade hospitalar. MÉTODOS: Os pacientes internados nas 14 unidades de terapia intensiva da Grande Vitória, durante o período de 15 meses, submetidos à ventilação mecânica e que preencheram os critérios de lesão pulmonar aguda da Conferência de Consenso Européia-Americana de 1994 foram selecionados prospectivamente para o estudo. Os pacientes também foram classificados de acordo com a nova definição de Berlim. Avaliamos as características clínicas e funcionais no primeiro dia de internação, durante a primeira semana, no 14º dia e no 28º dia de evolução. Foram calculadas a incidência da síndrome acumulada/ano, a sobrevida aos 28 dias e a mortalidade hospitalar. RESULTADOS: Foram avaliados 7.133 pacientes admitidos nas unidades de terapia intensiva, dos quais 130 (1,8%) foram selecionados. A mediana de tempo para o diagnóstico de lesão pulmonar aguda foi de 2 dias (IQ: 0-3 dias), sendo 25,4% dos diagnósticos realizados no momento da internação na unidade de terapia intensiva. Os fatores de risco foram principalmente pneumonia (35,3%), sepse não pulmonar (31,5%) e trauma (16,9%). A média de idade dos pacientes foi de 44,2 ± 15,9 anos, sendo 61,5% do sexo masculino. A média do APACHE II foi de 20,7 ± 7,9 e a média da PaO2/FiO2, de 206,7 ± 61,6. O tempo médio em ventilação mecânica foi de 21 ± 15 dias e o tempo médio de permanência na unidade de terapia intensiva foi de 26,4 ± 18,7 dias. De acordo com a nova definição de Berlim, os pacientes com a síndrome de desconforto respiratório agudo foram classificados em: leve, com 49 casos (37,7%); moderada, com 68(52,3%); e grave, com 13(10%). A incidência acumulada de LPA foi de 10,1 casos/100.000 habitantes/ano, sendo 3,8 casos/100.000 habitantes/ano para LPA sem SDRA e 6,3 casos/100.000 habitantes/ano para SDRA, representando 1,7% das admissões no ano. A relação PaO2/FiO2 nos dias 6 e 7 de evolução após o diagnóstico da síndrome foi um fator preditor independente para a mortalidade aos 28 dias, que foi de 38,5% (95% IC, 30,1-46,8). A mortalidade intrahospitalar foi de 49,2% (95% IC, 40,6-57,8), não diferindo entre os pacientes com LPA sem SDRA e SDRA. CONCLUSÕES: A incidência de LPA nos pacientes submetidos à ventilação mecânica invasiva na região da Grande Vitória, Espírito Santo, foi baixa, sendo a maioria dos casos diagnosticada 2 dias após a admissão nas unidades de terapia intensiva. A mortalidade aos 28 dias e a hospitalar dos pacientes com LPA sem SDRA e com SDRA não foram estatisticamente diferentes neste estudo. As mudanças nas práticas assistenciais nas unidades de terapia intensiva poderão contribuir para a redução da incidência da SDRA intrahospitalar / INTRODUCTION: There are many controversies in the existing epidemiological studies regarding the incidence and outcomes in acute lung injury. The incidence and clinical features of the syndrome mainly depend on the definition adopted and on the methodology employed in the study, as well as on the availability and use of beds in intensive therapy units in the regions studied. Due to the absence of existing epidemiological data concerning acute lung injury in Vitória, Espírito Santo, we conducted this study to analyze the incidence, clinical characteristics, survival rate at 28 days, and mortality rate. METHODS: The patients hospitalized in the 14 units of intensive therapy in the region of Grande Vitória for the period of 15 months submitted to mechanical ventilation, who fulfilled the criteria of acute lung injury as defined by the Conference of European-American Consensus of 1994, were prospectively selected for the study. These patients were also classified according to the new Berlin definition. We evaluated the clinical and functional characteristics on the first day of hospitalization, during the first week, on day 14 and on day 28 of clinical evolution. We calculated the cumulative incidence/year for the syndrome, the survival rate at 28 days, and hospital mortality. RESULTS: A total of 7,133 patients admitted to the intensive care units was evaluated, of whom 130 (1.8%) were selected. The median time to diagnosis of acute lung injury was 2 days (IQR: 0-3 days), 25.4% of diagnoses being made at admission to the intensive care unit. The risk factors were mainly pneumonia (35.3%), nonpulmonary sepsis (31.5%) and trauma (16.9%). The patients\' mean age was 44.2 ± 15.9 years, 61.5% being male. The APACHE II prognostic score averaged 20.7 ± 7.9, mean arterial oxygenation variable PaO2/FiO2 206 ± 61.6 and time on mechanical ventilation with a mean of 21 ± 15 days. The average length of stay in intensive care unit was 26.4 ± 18.7 days. Based on the new Berlin definition, patients with acute respiratory distress syndrome were classified as mild: 49 (37.7%); moderate: 68 (52.3%); and severe: 13 (10%). The cumulative incidence was 10.1 cases per 100,000 inhabitants /year for ALI, of which 3.8 cases per 100,000 inhabitants / year were for non-ARDS ALI and 6.3 cases per 100,000 inhabitants / year were for ARDS, representing 1.7% of admissions in the year. The variable arterial oxygenation on days 6 and 7 of evolution after the diagnosis of the syndrome was an independent factor for mortality at 28 days, which was 38.5% (95% CI, 30.1 to 46.8). In-hospital mortality was 49.2% (95% CI, 40.6 to 57.8), and did not differ between patients with ALI non-ARDS and acute respiratory distress syndrome Summary (ARDS). CONCLUSIONS: The incidence of acute lung injury in patients undergoing invasive mechanical ventilation in the region of Grande Vitória, Espírito Santo was low, most of them being diagnosed 2 days after admission to intensive care units. Mortality at 28 days and hospital mortality of patients with ALI non-ARDS were not statistically different in this study. Changes in care practices in intensive therapy units can contribute to reduce the incidence of in-hospital ARDS

INTRODUÇÃO: A insuficiência respiratória acomete grande parte dos pacientes oncológicos levando a altos índices de mortalidade. A ventilação não invasiva (VNI) pode auxiliar seu manejo, mas seus efeitos ainda são pouco conhecidos sobre os mecanismos de defesa pulmonar. OBJETIVOS: Observar o efeito da VNI com máscara facial usando-se geradores de fluxo com pressão positiva contínua (CPAP) e ventilador microprocessado no modo pressão de suporte + pressão positiva ao final da expiração (PSV + PEEP), a fim de verificar impacto nas propriedades viscoelásticas do muco respiratório e o conforto proporcionado ao paciente. MÉTODOS: A VNI foi instalada após diagnóstico de insuficiência respiratória em dezenove pacientes, admitidos nas unidades de tratamento intensivo do Hospital A. C. Camargo, sendo nove submetidos ao CPAP e dez com PSV + PEEP. Foram colhidos antes e após uma hora de VNI: os dados clínicos, secreção nasal, gasometria, e o grau de conforto através de uma escala visual. As propriedades físicas do muco (transportabilidade in vitro, adesividade e wettabilidade ou hidrofobicidade) foram avaliadas respectivamente no palato de rã, máquina da tosse e ângulo de contato. RESULTADOS: Os grupos eram homogêneos entre si em relação à idade, sexo, tipo e estadiamento do tumor e SAPS II. Em relação às propriedades físicas do muco, houve um aumento da transportabilidade in vitro do muco nasal com o sistema PSV + PEEP (p = 0,04) e um aumento na wettabilidade no grupo CPAP (p = 0,06). Os dois sistemas foram eficazes em melhorar significativamente os sinais vitais, a PaO2/FiO2, o padrão e o conforto respiratório e em evitar a intubação traqueal nas primeiras 24 horas (p < 0,05). Entretanto, independentemente do tipo de sistema de VNI usado, foram encontrados altos índices de intubação endotraqueal e mortalidade no seguimento destes pacientes. CONCLUSÃO: As propriedades físicas do muco (transportabilidade in vitro e wettabilidade) se alteraram após uma hora de uso da VNI e parecem ser dependentes da temperatura e umidificação dentro da máscara. A VNI mostrou-se útil em reverter a insuficiência respiratória em pacientes selecionados, ou pelo menos em trazer conforto para pacientes hipoxêmicos que a princípio recusam a intubação endotraqueal / INTRODUCTION: Respiratory failure is a common situation among cancer patients leading to high rates of mortality. Noninvasive ventilation (NIV) can help its management, but its effects are still unknown regarding the pulmonary defense mechanisms. OBJECTIVES: Observe the effect of NIV with facial mask using a flow generator with continuous positive pressure (CPAP) and standard intensive care unit ventilator using pressure support ventilation + positive end expiratory pressure (PSV + PEEP), to verify impact on the physical properties of respiratory mucus and the comfort provided to the patient. METHODS: NIV was started after diagnosis of respiratory failure in nineteen patients, admitted in the intensive care unit of the A. C. Camargo Hospital. Nine patients were submitted to CPAP and ten to PSV + PEEP. Nasal mucus, blood gases, and the degree of comfort through a visual scale were accessed before and after one hour. The physical properties of nasal mucus (transportabilility in vitro, adhesivity and wettability or hydrofobicity) were evaluated respectively by frog palate, cough machine and contact angle. RESULTS: Groups had similar characteristics about age, sex, tumor and SAPS II score. Regarding the physical properties of the mucus, there was an increase in mucus transportability (by the frog palate model) with the system PSV + PEEP (p = 0.04) and an increase in the contact angle in the CPAP groupo (p = 0.06). The two systems were effective in improving the vital signs, the PaO2/FiO2, the respiratory pattern and comfort and avoiding endotracheal intubation in the first 24 hours (p < 0.05). However, regardless of the type of NIV system used, high rates of endotracheal intubation and mortality were found. CONCLUSION: The physical properties of the mucus (transportability in vitro and wettability) changed after an hour of use of the NIV as a result of temperature and humidification into the mask. NIV was useful in reversing the respiratory failure in selected patients, or at least in bringing comfort for those who refuse endotracheal intubation

<p>The human coronavirus NL63 was discovered in 2004 by a team of researchers in Amsterdam. Since its discovery it has been shown to have worldwide spread and affects mainly children, aged 0-5 years old, the immunocompromised and the elderly. Infection with HCoV-NL63 commonly results in mild upper respiratory tract infections and presents as the common cold, with symptoms including fever, cough, sore throat and rhinorrhoea. Lower respiratory tract findings are less common but may develop into more serious complications including bronchiolitis, pneumonia and croup. The primary function of the HCoV-NL63 nucleocapsid (N) protein is the formation of theprotective ribonucleocapsid core. For this particle to assemble, the N-protein undergoes N-N dimerization and then interacts with viral RNA. Besides the primary structural role of the Nprotein, it is also understood to be involved in viral RNA transcription, translation and replication, including several other physiological functions. The N-protein is also highly antigenic and elicits a strong immune response in infected patients. For this reason the N-protein may serve as a target for the development of diagnostic assays. We have used bioinformatic analysis to analyze the HCoV-NL63 N-protein and compared it to coronavirus N-homologues. This bioinformatic analysis provided the data to generate recombinant clones for expression in a bacterial system. We constructed recombinant clones of the N-protein of SARS-CoV and HCoV-NL63 and synthesized truncated clones corresponding to the N- and C-terminal of the HCoV-NL63 N-protein. These heterologously expressed proteins will serve the basis for several post-expression studies including characterizing the immunogenic epitope of the N-protein as well identifying any antibody crossreactivity between coronavirus species.</p>

<p>Human Coronaviruses (HCoVs) are found within the family Coronaviridae (genus, Coronavirus) and are enveloped, single-stranded, positive-sense RNA viruses. Infections of humans by&nbsp / coronaviruses are not normally associated with severe diseases. However, the identification of the coronavirus responsible for the outbreak of severe acute respiratory syndrome (SARS-CoV)&nbsp / showed that highly pathogenic coronaviruses can enter the human population. The SARS-CoV epidemic resulted in 8 422 cases with 916 deaths globally (case fatality rate: 10.9%). In 2004 a&nbsp / group 1 Coronavirus, designated Human Coronavirus NL63 (HCoV-NL63), was isolated from a 7 month old Dutch child suffering from bronchiolitis. In addition, HCoV-NL63 causes disease in&nbsp / children (detected in approximately 10% of respiratory tract infections), the elderly and the immunocompromised. This study was designed to express the full length nucleocapsid (N) proteins of&nbsp / HCoV-NL63 and SARS-CoV for antibody production in an animal model. The NL63-N/pFN2A and SARSN/ pFN2A plasmid constructs were used for this study. The presence of the insert on the Flexi &reg / vector was confirmed by restriction endonuclease digest and sequence verification. The sequenced chromatographs obtained from Inqaba Biotec were consistent with sequences from&nbsp / the NCBI Gen_Bank. Proteins were expressed in a KRX Escherichia coli bacterial system and analysed using 15% SDS-PAGE and Western Blotting. Thereafter, GST-tagged proteins were purified&nbsp / ith an affinity column purification system. Purified fusion proteins were subsequently cleaved with Pro-TEV Plus protease, separated on 15% SDS-PAGE gel and stained with Coomassie&nbsp / Brilliant Blue R250. The viral fusion proteins were subsequently used to immunize Balbc mice in order to produce polyclonal antibodies. A direct ELISA was used to analyze and validate the&nbsp / production of polyclonal antibodies by the individual mice. This is a preliminary study for development of diagnostic tools for the detection of HCoV-NL63 from patient samples collected in the&nbsp / Western Cape.</p>