Background and objective: Asbestos can cause various pulmonary diseases including asbestosis, pleural plaques and pleural thickening. Animal and in vitro studies suggest that the toxicity of asbestos is due to the iron content of the fibres which not only generate oxidants directly, but also activate the inflammatory cells in the lung that contribute to oxidative stress. This study therefore sought to establish data in man to corroborate the animal and in vitro evidence. Exhaled breath condensate (EBC) collection is a novel, non-invasive technique to collect samples from the lung for investigating inflammatory biomarkers of lung diseases. This technique is harmless, rapid and easily repeatable which leads itself to the investigation of lung diseases such as asbestos-related diseases and pulmonary fibrosis (PF) that are otherwise difficult to study. The hypothesis tested was that oxidative and nitrosylative stress markers will be elevated in the EBC of patients with asbestos-related diseases and PF compared to normal control subjects. Methods: The study design was a cross-sectional and observational in vivo study whereby EBC was collected and fractional exhaled nitric oxide (FeNO) and carbon monoxide (eCO) were measured. EBC markers including pH, hydrogen peroxide (H2O2), total nitrogen oxides (NOx), 3-nitrotyrosine (3-NT), 8-isoprostane (8-iso), total protein and transforming growth factor-β1 (TGFβ1) were measured by microelectrode analysis, colorimetric and enzyme immunoassays. 3-NT and 8-iso were further examined by immunohistochemical techniques in samples of lung tissue. Results: Subjects with asbestosis had significantly raised levels of EBC H2O2, 8-iso, total protein and FeNO compared with healthy individuals. The same markers except H2O2, but with 3-NT and eCO were again significantly increased in those with other causes of PF, compared with control subjects. Heavy nitrotyrosine staining was found on the lung sections from patients with asbestosis and PF. Conclusions: This study confirmed that increased production of reactive oxygen and nitrogen species is associated with asbestos exposure and pulmonary fibrosis in vivo confirming animal and in vitro studies. Analysis of EBC may prove a useful non-invasive tool in exploring the basic pathophysiology of lung diseases in clinical research and may in the future be used to monitor progress in asbestosis and pulmonary fibrosis.

Healthcare costs are escalating in the U.S., with a projected 48 trillion dollars by 2021. More than ever medical researches are obligated to ensure that costly treatment modalities are safe and effective. Chronic obstructive pulmonary disease (COPD) is a costly and debilitating disease, ranked as the third leading cause of death in America. Currently, treatment for COPD consists of anti-inflammatory agents, bronchodilators, antibiotics and supplemental oxygen when hypoxemia or clinical manifestations ensue. Oxidative stress is central to the pathology of COPD. Supplemental oxygen has been substantiated as an instigator of oxidative stress; however, LFDO has not been evaluated as a mechanism of ongoing oxidative stress in individuals with COPD. Isofuran (IsoF), a biosynthetic relative of the validated oxidative stress biomarker 8-isoprostane, is preferentially synthesized during periods of increased tissue oxygen tension. This sort of specificity allows for refinement in the assessment of supplemental oxygen as a source of oxidative stress. To address this potential this study evaluated individuals diagnosed with COPD utilizing LFDO. The study also aimed to determine if IsoF possessed clinical application in predicting the standard pulmonary function test (PFT) parameters of FEV1, FVC, FEV1/FVC and FEF25-75. The exhaled breath condensate (EBC) of 52 individuals with COPD was evaluated for the presence of IsoF. An active control group not receiving LFDO (n=26) was compared to an active treatment group receiving LFDO for a minimum of 6 hrs/day (n=26). The groups showed no statistically significant demographic differences in age, gender, height, weight, ethnicity or smoking history or in the pulmonary function test parameters of FEV1, FVC, and FEV1/FVC, with the exception of the FEF25-75 (P=0.03). The active control group generated a mean EBC IsoF level of 35.81 ± 4.91 pg/ml (± SEM) compared to the active treatment group mean EBC IsoF level of 51.37 ± 8.27 pg/ml (P=0.057). Currently, no research has been conducted that defines baseline EBC IsoF levels in healthy or diseased lungs. No statistically significant differences in mean EBC IsoF levels were noted between the control and treatment groups; however, the results, in conjunction with the only two studies available utilizing EBC IsoF as an oxidative stress biomarker, may serve to provide benchmark information for future research regarding individuals with diseased lungs, specifically COPD.

O óxido nítrico (NO) tem papel relevante nas diversas funções fisiológicas do organismo e nos mais variados sistemas que o compõe. Sabe-se que esta molécula é um constituinte normal do ar exalado na respiração de humanos e de animais. Tem como funções modular respostas inflamatórias e imunes bem como atuar na regulação do tônus vascular da musculatura lisa das vias aéreas. Esta molécula vem sendo reconhecida como importante marcador de lesão pulmonar, o que despertou a intensa investigação do NO no ar exalado. Uma metodologia não invasiva, de baixo custo e de fácil aplicação está sendo extensivamente estudada para a captação do ar exalado através do seu resfriamento e condensação, denominada Condensado do Exalado Pulmonar (CEP). É grande o interesse no estudo de doenças pulmonares inflamatórias, no entanto, observa-se uma considerável carência de evidências acerca dos níveis de concentração de mediadores inflamatórios, como o NO em amostras de indivíduos hígidos utilizando o CEP. Diante disso, decidiu-se elaborar um protótipo de aparato para coleta do condensado e empregar esta técnica para determinar valores de concentração de NO em uma amostra de voluntários hígidos. A metodologia de coleta utilizada foi baseada nas recomendações da força tarefa da American Thoracic Society/European Respiratory Society (ATS/ERS) para procedimentos de coleta de condensado, com o intuito de evitar erros metodológicos, padronizar e trazer mais evidências para auxiliar a inserção do método na rotina clínica, além de disponibilizá-lo para novas pesquisas. Para isto foram selecionados 116 indivíduos adultos hígidos de ambos os sexos, com idade entre 20 e 70 anos. O protótipo de aparato utilizado na pesquisa foi de fabricação artesanal e desenvolvido na Divisão de Cirurgia Torácica e Cardiovascular da Faculdade de Medicina da Universidade de São Paulo- FMRP/USP, visto que os aparatos comerciais são todos importados, de custo elevado e que ainda não forneceram evidências suficientes de que algum destes seria o padrão ouro para a coleta. O aparato foi eficiente para coletar o CEP e o NO foi detectável em todas as amostras. A média de NO foi de 13±14,4µM, e não houve correlação com o gênero (p=0,847) e idade (p=0,811). Não foi observado correlação entre o NO e o volume de CEP coletado (p=0,944). O volume de CEP coletado é dependente do volume exalado total (p<0,001). A partir destes resultados pode-se verificar que o aparato é viável e eficiente. Não foram encontradas correlações entre a concentração de NO no CEP, idade e o gênero. O presente estudo poderá contribuir com futuras pesquisas de caráter comparativo e fornecer mais uma opção de aparato para coleta do condensado do exalado pulmonar, provendo baixo custo, reprodutibilidade e reutilização do aparato / Nitric oxide (NO) plays an important role in various physiological functions in the body and in various systems that compose it. It is known that this molecule is a normal constituent of exhaled breath in humans and animals. Its functions modulate inflammatory and immune responses as well as act in the regulation of vascular tone of smooth muscles of the airways. This molecule has been recognized as an important marker of lung injury, which sparked intense investigation of exhaled NO. A non-invasive method, low cost and easy application has been extensively studied to collect the exhaled air through its cooling and condensation, termed exhaled breath condensate. There is great interest in the study of inflammatory lung diseases, however, there is a considerable lack of evidence about the concentration levels of inflammatory mediators such NO in samples of healthy individuals using exhaled breath condensate. Given this, it was decided to develop a prototype apparatus for collecting exhaled breath condensate (EBC) and employ this technique to determine concentration values of nitric oxide in a sample of healthy volunteers. The sampling methodology based on the recommendations of the task force of the American Thoracic Society / European Respiratory Society (ATS / ERS) for procedures using for collection of condensate, in order to avoid methodological errors, standardize and bring more evidence to support the inclusion of the method in clinical routine, and make it available for further research. We selected 116 healthy adults of both gender, aged between 20 and 70. The prototype apparatus used in the research was designed, produced and developed in the Division of Thoracic and Cardiovascular Surgery, Faculty of Medicine, University of São Paulo-FMRP / USP. The commercial devices are all imported, expensive and not yet provided sufficient evidence that any of these would be the gold standard for collection. The apparatus was efficient to collect the EBC and NO was detectable in all samplesThe average NO was 14.4 ± 13 µM, and there was no correlation with gender (p = 0.847) and age (p = 0.811). There was no correlation between NO and the volume of collected EBC (p = 0.944). The volume of collected EBC is dependent on the total exhaled volume (p <0.001). From these results it can be seen that the apparatus is feasible and efficient. No correlation was found between the concentration of NO in the EBC, age and gender. The present study may contribute to future comparative studies of character and provide another option for apparatus for collecting condensate, providing low cost, reliability and reusability of the apparatus

Lung cancer (LC) is often diagnosed at advanced stage and as a result, survival rates are low. Recent studies describe exhaled breath and exhaled breath condensate (EBC) as a potential non-invasive method of sampling the airways for assessing inflammation of the respiratory system, and possibly for the early detection of LC. It was hypothesised that higher concentrations of markers and protein will be present in the EBC of LC patients compared to those of normal controls and healthy smokers, and may aid in assessing lung status. Methods: The gaseous phase of breath was investigated for volatile organic compound (VOC) patterns using an electronic nose (eNose) system, in addition to off-line measurements of carbon monoxide (CO) and nitric oxide (NO) levels. The aqueous phase, EBC, was collected during tidal breathing through a glass collection device cooled to 4??C by ice. Nitrite/nitrate (NOx) and pH levels were determined by a fluorescent modification of the Griess method, and silicon chip sensor pH meter, respectively. Protein levels in EBC were examined with a bicinchoninic acid (BCA) assay, silver staining and PAGE techniques, while the levels of tumour markers, CYFRA 21-1 and CEA, were quantified by enzyme-linked immunosorbent assays (ELISA). Results: The eNose machine was not able to produce characteristic VOC profiles from exhaled breath unique to each study group, while no significant difference was observed for mean NOx concentrations in the LC group when compared to other subjects (p=0.8824). Higher protein levels were found in the EBC of LC patient compared to normal controls (p=0.0204), with subsequent measurements of elevated CEA levels observed in the LC group when compared to non-smokers and smokers (p=0.023). Conclusion: This study showed that protein can be detected in the exhaled breath condensate of patients, with a significantly elevated amount in the samples from newly diagnosed LC patients. The mechanism for these differences remains to be determined but may be related to inflammatory changes within the airway, such as vascular protein leakage and release of mediators. Future work may aim to identify the upregulated proteins, and focus on proteomics and tissue microarrays to explore candidate proteins.

Lung cancer (LC) is often diagnosed at advanced stage and as a result, survival rates are low. Recent studies describe exhaled breath and exhaled breath condensate (EBC) as a potential non-invasive method of sampling the airways for assessing inflammation of the respiratory system, and possibly for the early detection of LC. It was hypothesised that higher concentrations of markers and protein will be present in the EBC of LC patients compared to those of normal controls and healthy smokers, and may aid in assessing lung status. Methods: The gaseous phase of breath was investigated for volatile organic compound (VOC) patterns using an electronic nose (eNose) system, in addition to off-line measurements of carbon monoxide (CO) and nitric oxide (NO) levels. The aqueous phase, EBC, was collected during tidal breathing through a glass collection device cooled to 4??C by ice. Nitrite/nitrate (NOx) and pH levels were determined by a fluorescent modification of the Griess method, and silicon chip sensor pH meter, respectively. Protein levels in EBC were examined with a bicinchoninic acid (BCA) assay, silver staining and PAGE techniques, while the levels of tumour markers, CYFRA 21-1 and CEA, were quantified by enzyme-linked immunosorbent assays (ELISA). Results: The eNose machine was not able to produce characteristic VOC profiles from exhaled breath unique to each study group, while no significant difference was observed for mean NOx concentrations in the LC group when compared to other subjects (p=0.8824). Higher protein levels were found in the EBC of LC patient compared to normal controls (p=0.0204), with subsequent measurements of elevated CEA levels observed in the LC group when compared to non-smokers and smokers (p=0.023). Conclusion: This study showed that protein can be detected in the exhaled breath condensate of patients, with a significantly elevated amount in the samples from newly diagnosed LC patients. The mechanism for these differences remains to be determined but may be related to inflammatory changes within the airway, such as vascular protein leakage and release of mediators. Future work may aim to identify the upregulated proteins, and focus on proteomics and tissue microarrays to explore candidate proteins.

O óxido nítrico (NO) tem papel relevante nas diversas funções fisiológicas do organismo e nos mais variados sistemas que o compõe. Sabe-se que esta molécula é um constituinte normal do ar exalado na respiração de humanos e de animais. Tem como funções modular respostas inflamatórias e imunes bem como atuar na regulação do tônus vascular da musculatura lisa das vias aéreas. Esta molécula vem sendo reconhecida como importante marcador de lesão pulmonar, o que despertou a intensa investigação do NO no ar exalado. Uma metodologia não invasiva, de baixo custo e de fácil aplicação está sendo extensivamente estudada para a captação do ar exalado através do seu resfriamento e condensação, denominada Condensado do Exalado Pulmonar (CEP). É grande o interesse no estudo de doenças pulmonares inflamatórias, no entanto, observa-se uma considerável carência de evidências acerca dos níveis de concentração de mediadores inflamatórios, como o NO em amostras de indivíduos hígidos utilizando o CEP. Diante disso, decidiu-se elaborar um protótipo de aparato para coleta do condensado e empregar esta técnica para determinar valores de concentração de NO em uma amostra de voluntários hígidos. A metodologia de coleta utilizada foi baseada nas recomendações da força tarefa da American Thoracic Society/European Respiratory Society (ATS/ERS) para procedimentos de coleta de condensado, com o intuito de evitar erros metodológicos, padronizar e trazer mais evidências para auxiliar a inserção do método na rotina clínica, além de disponibilizá-lo para novas pesquisas. Para isto foram selecionados 116 indivíduos adultos hígidos de ambos os sexos, com idade entre 20 e 70 anos. O protótipo de aparato utilizado na pesquisa foi de fabricação artesanal e desenvolvido na Divisão de Cirurgia Torácica e Cardiovascular da Faculdade de Medicina da Universidade de São Paulo- FMRP/USP, visto que os aparatos comerciais são todos importados, de custo elevado e que ainda não forneceram evidências suficientes de que algum destes seria o padrão ouro para a coleta. O aparato foi eficiente para coletar o CEP e o NO foi detectável em todas as amostras. A média de NO foi de 13±14,4µM, e não houve correlação com o gênero (p=0,847) e idade (p=0,811). Não foi observado correlação entre o NO e o volume de CEP coletado (p=0,944). O volume de CEP coletado é dependente do volume exalado total (p<0,001). A partir destes resultados pode-se verificar que o aparato é viável e eficiente. Não foram encontradas correlações entre a concentração de NO no CEP, idade e o gênero. O presente estudo poderá contribuir com futuras pesquisas de caráter comparativo e fornecer mais uma opção de aparato para coleta do condensado do exalado pulmonar, provendo baixo custo, reprodutibilidade e reutilização do aparato / Nitric oxide (NO) plays an important role in various physiological functions in the body and in various systems that compose it. It is known that this molecule is a normal constituent of exhaled breath in humans and animals. Its functions modulate inflammatory and immune responses as well as act in the regulation of vascular tone of smooth muscles of the airways. This molecule has been recognized as an important marker of lung injury, which sparked intense investigation of exhaled NO. A non-invasive method, low cost and easy application has been extensively studied to collect the exhaled air through its cooling and condensation, termed exhaled breath condensate. There is great interest in the study of inflammatory lung diseases, however, there is a considerable lack of evidence about the concentration levels of inflammatory mediators such NO in samples of healthy individuals using exhaled breath condensate. Given this, it was decided to develop a prototype apparatus for collecting exhaled breath condensate (EBC) and employ this technique to determine concentration values of nitric oxide in a sample of healthy volunteers. The sampling methodology based on the recommendations of the task force of the American Thoracic Society / European Respiratory Society (ATS / ERS) for procedures using for collection of condensate, in order to avoid methodological errors, standardize and bring more evidence to support the inclusion of the method in clinical routine, and make it available for further research. We selected 116 healthy adults of both gender, aged between 20 and 70. The prototype apparatus used in the research was designed, produced and developed in the Division of Thoracic and Cardiovascular Surgery, Faculty of Medicine, University of São Paulo-FMRP / USP. The commercial devices are all imported, expensive and not yet provided sufficient evidence that any of these would be the gold standard for collection. The apparatus was efficient to collect the EBC and NO was detectable in all samplesThe average NO was 14.4 ± 13 µM, and there was no correlation with gender (p = 0.847) and age (p = 0.811). There was no correlation between NO and the volume of collected EBC (p = 0.944). The volume of collected EBC is dependent on the total exhaled volume (p <0.001). From these results it can be seen that the apparatus is feasible and efficient. No correlation was found between the concentration of NO in the EBC, age and gender. The present study may contribute to future comparative studies of character and provide another option for apparatus for collecting condensate, providing low cost, reliability and reusability of the apparatus

Pacientes submetidos à cirurgia cardiotorácica com circulação extracorpórea (CEC) apresentam reações inflamatórias, desencadeadas por este procedimento e pela isquemiareperfusão, que acarretam disfunção pulmonar e lesão do endotélio vascular no pósoperatório. Estes processos relativos à cirurgia cardíaca podem afetar a produção e o consumo do óxido nítrico (NO) no pulmão e no endotélio vascular, principalmente na sua participação no processo inflamatório. No pulmão, o NO se difunde com facilidade sendo detectado na via aérea na fase gasosa ou no fluído pulmonar como nitrito e o nitrato. A presença do NO na via aérea pode em algumas situações refletir a sua produção pelo epitélio da via aérea e pelo endotélio microvascular pulmonar. No entanto, os processos que envolvem a produção e o consumo do NO na via aérea ainda não estão totalmente esclarecidos. O condensado do exalado pulmonar (CEP) é um fluido obtido através do resfriamento do ar exalado através de um método totalmente não invasivo e que pode ser utilizado para a investigação do NO nas vias aéreas distais. No desenvolvimento deste estudo foi utilizado um aparato de coleta artesanal e de baixo custo para obtenção do CEP. Assim, foi coletado o CEP de pacientes coronariopatas e valvopatas submetidos à cirurgia cardíaca com CEC nos períodos pré- operatório e pós-operatório (4 horas após a CEC e 12, 24, 48 e 72 horas após a extubação). As concentrações plasmáticas de nitrito/nitrato destes pacientes também foram avaliadas através de amostras de sangue colhidas imediatamente após cada coleta do CEP. O CEP e o plasma foram analisados pela técnica de quimioluminescência para obter as concentrações de nitrito/nitrato. O aparato utilizado obteve utilização reprodutível na rotina clínica da cirurgia cardíaca nesta pesquisa em pacientes sob ventilação espontânea e ventilação mecânica. Não foram encontradas diferenças significativas (p>0,05) nas concentrações de nitrito/nitrato no CEP e plasma entre os pacientes coronariopatas e valvopatas no período pré-operatório. Concentrações significativamente maiores de nitrito/nitrato (p=0,017) foram encontradas no CEP, mas não no plasma, de pacientes que utilizaram nitrato por via oral (dinitrato de isossorbida) no período pré-operatório. Em pacientes que não utilizaram medicação contendo doador de NO (nitroglicerina ou nitroprussiato de sódio) por via endovenosa no pósoperatório ocorreu uma tendência à elevação das concentrações de nitrito/nitrato no CEP que foi numericamente significativa no período de 48 horas do pós-operatório (p=0,008). Em pacientes que receberam ou não doador de NO no pós-operatório as concentrações de nitrito/nitrato foram significativamente maiores nos períodos de 48 horas (p=0,005) e 72 horas (p=0,037) do pós-operatório. A utilização de nitroglicerina (NTG) no período pósoperatório elevou significativamente as concentrações de nitrito/nitrato no CEP nos períodos de 12 horas (p=0,022), 48 horas (p=0,015) e 72 horas (0,048) e no plasma nos períodos de 12 horas (p=0,045). As concentrações de nitrito/nitrato plasmáticos foram significativamente reduzidas (p=0,045) no pós-operatório imediato 4 horas após a CEC em pacientes que receberam ou não doador de NO. A partir destes resultados pode se observar que não houve diferenças significativas nas concentrações de nitrito/nitrato no CEP dos pacientes coronariopatas e valvopatas no período pré-operatório, exceto elevação significativa encontrada em pacientes que utilizaram dinitrato de isossorbida por via oral. As concentrações de nitrito/nitrato no CEP tendem a elevar-se no período pós-operatório principalmente com a utilização de NTG. As concentrações de nitrito/nitrato no plasma foram significativamente reduzidas após 4 horas da CEC no pós-operatório imediato. No entanto foi observado elevação nas concentrações de nitrito/nitrato no plasma de pacientes que usaram NTG. Os achados deste estudo poderão contribuir em futuras pesquisas fornecendo dados comparativos a respeito da participação do NO na via aérea e no plasma nos processos que envolvem a cirurgia cardíaca nos períodos pré e pós-operatório. / Patients undergoing cardiothoracic surgery with cardiopulmonary bypass (CPB) have inflammatory reactions triggered by this procedure and by ischemia-reperfusion, which cause lung dysfunction and injury of vascular endothelium in the postoperative period. These processes related to cardiac surgery may affect the production and consumption of nitric oxide (NO) in the lung and vascular endothelium, especially in their participation in the inflammatory process. In the lung, NO diffuses easily and can be detected in the airway in the gas phase or in the lung fluid as nitrite and nitrate. The presence of NO in exhaled air may reflect the airway epithelium and lung microvascular endothelium in some situations. However, processes involving the production and consumption of NO in the airway are not totally understood. The exhaled breath condensate (EBC) is a fluid obtained by cooling exhaled air wich can be used for the investigation of NO in the distal airways. For this study we used an apparatus homemade and with low cost for collecting and obtain the EBC. The EBC was collected from patients with coronary artery disease and heart valve disease undergoing cardiac surgery with CPB in the preoperative and postoperative period (4 hours after CPB and 12, 24, 48 and 72 hours after extubation). Plasma concentrations of nitrite/nitrate in these patients were also evaluated using blood samples taken immediately after each collection of the EBC. The EBC and plasma were analyzed by chemiluminescence method to obtain the concentrations of nitrite/nitrate. The apparatus used was reproducible in clinical routine of cardiac surgery in patients under spontaneous ventilation or mechanical ventilation. There were no significant differences (p> 0.05) in concentrations of nitrite/nitrate in EBC and plasma in patients with coronary artery disease and heart valve disease in the preoperative period. Significantly higher concentrations of nitrite/nitrate (p=0.017) were found in EBC, but not in plasma of patients who used oral nitrate (isosorbide dinitrate) in the preoperative period. In patients who did not use medication containing NO donor (nitroglycerin or sodium nitroprusside) intravenously in the postoperative period there was a tendency to high concentrations of nitrite/nitrate in EBC that was numerically significant in the period of 48 hours postoperative (p=0.008). In patients who received or not NO donor in postoperative period the concentrations of nitrite/nitrate were significantly higher in periods of 48 hours (p=0.005) and 72 hours (p=0.037) after surgery. The use of nitroglycerin (GTN) in the postoperative period significantly elevated concentrations of nitrite/nitrate in the EBC in periods of 12 hours (p=0.022), 48 hours (p=0.015) and 72 hours (0.048) and plasma in period of 12 hours (p=0.045). The concentrations of nitrite/nitrate were significantly reduced (p=0.045) in the immediate postoperative period 4 hours after CPB in patients who received or not NO donor. From these results we could observe that there were no significant differences in concentrations of nitrite/nitrate in the EBC of patients with heart valve disease and coronary artery disease in the preoperative period, but we found a significant increase in these concentrations in patients who used isosorbide dinitrate orally. The concentrations of nitrite/nitrate in EBC presented a tendency to increase in postoperative period especially in patients who used GTN. The concentrations of nitrite/nitrate in plasma were significantly reduced after 4 hours of CPB in the immediate postoperative period. However, was observed that GTN tended to increase the concentration of nitrite/nitrate in plasma. These findings could help in future research providing comparative data about the role of NO in the airway and plasma in processes involving cardiac surgery in the pre and postoperative periods.

Pacientes submetidos à cirurgia cardiotorácica com circulação extracorpórea (CEC) apresentam reações inflamatórias, desencadeadas por este procedimento e pela isquemiareperfusão, que acarretam disfunção pulmonar e lesão do endotélio vascular no pósoperatório. Estes processos relativos à cirurgia cardíaca podem afetar a produção e o consumo do óxido nítrico (NO) no pulmão e no endotélio vascular, principalmente na sua participação no processo inflamatório. No pulmão, o NO se difunde com facilidade sendo detectado na via aérea na fase gasosa ou no fluído pulmonar como nitrito e o nitrato. A presença do NO na via aérea pode em algumas situações refletir a sua produção pelo epitélio da via aérea e pelo endotélio microvascular pulmonar. No entanto, os processos que envolvem a produção e o consumo do NO na via aérea ainda não estão totalmente esclarecidos. O condensado do exalado pulmonar (CEP) é um fluido obtido através do resfriamento do ar exalado através de um método totalmente não invasivo e que pode ser utilizado para a investigação do NO nas vias aéreas distais. No desenvolvimento deste estudo foi utilizado um aparato de coleta artesanal e de baixo custo para obtenção do CEP. Assim, foi coletado o CEP de pacientes coronariopatas e valvopatas submetidos à cirurgia cardíaca com CEC nos períodos pré- operatório e pós-operatório (4 horas após a CEC e 12, 24, 48 e 72 horas após a extubação). As concentrações plasmáticas de nitrito/nitrato destes pacientes também foram avaliadas através de amostras de sangue colhidas imediatamente após cada coleta do CEP. O CEP e o plasma foram analisados pela técnica de quimioluminescência para obter as concentrações de nitrito/nitrato. O aparato utilizado obteve utilização reprodutível na rotina clínica da cirurgia cardíaca nesta pesquisa em pacientes sob ventilação espontânea e ventilação mecânica. Não foram encontradas diferenças significativas (p>0,05) nas concentrações de nitrito/nitrato no CEP e plasma entre os pacientes coronariopatas e valvopatas no período pré-operatório. Concentrações significativamente maiores de nitrito/nitrato (p=0,017) foram encontradas no CEP, mas não no plasma, de pacientes que utilizaram nitrato por via oral (dinitrato de isossorbida) no período pré-operatório. Em pacientes que não utilizaram medicação contendo doador de NO (nitroglicerina ou nitroprussiato de sódio) por via endovenosa no pósoperatório ocorreu uma tendência à elevação das concentrações de nitrito/nitrato no CEP que foi numericamente significativa no período de 48 horas do pós-operatório (p=0,008). Em pacientes que receberam ou não doador de NO no pós-operatório as concentrações de nitrito/nitrato foram significativamente maiores nos períodos de 48 horas (p=0,005) e 72 horas (p=0,037) do pós-operatório. A utilização de nitroglicerina (NTG) no período pósoperatório elevou significativamente as concentrações de nitrito/nitrato no CEP nos períodos de 12 horas (p=0,022), 48 horas (p=0,015) e 72 horas (0,048) e no plasma nos períodos de 12 horas (p=0,045). As concentrações de nitrito/nitrato plasmáticos foram significativamente reduzidas (p=0,045) no pós-operatório imediato 4 horas após a CEC em pacientes que receberam ou não doador de NO. A partir destes resultados pode se observar que não houve diferenças significativas nas concentrações de nitrito/nitrato no CEP dos pacientes coronariopatas e valvopatas no período pré-operatório, exceto elevação significativa encontrada em pacientes que utilizaram dinitrato de isossorbida por via oral. As concentrações de nitrito/nitrato no CEP tendem a elevar-se no período pós-operatório principalmente com a utilização de NTG. As concentrações de nitrito/nitrato no plasma foram significativamente reduzidas após 4 horas da CEC no pós-operatório imediato. No entanto foi observado elevação nas concentrações de nitrito/nitrato no plasma de pacientes que usaram NTG. Os achados deste estudo poderão contribuir em futuras pesquisas fornecendo dados comparativos a respeito da participação do NO na via aérea e no plasma nos processos que envolvem a cirurgia cardíaca nos períodos pré e pós-operatório. / Patients undergoing cardiothoracic surgery with cardiopulmonary bypass (CPB) have inflammatory reactions triggered by this procedure and by ischemia-reperfusion, which cause lung dysfunction and injury of vascular endothelium in the postoperative period. These processes related to cardiac surgery may affect the production and consumption of nitric oxide (NO) in the lung and vascular endothelium, especially in their participation in the inflammatory process. In the lung, NO diffuses easily and can be detected in the airway in the gas phase or in the lung fluid as nitrite and nitrate. The presence of NO in exhaled air may reflect the airway epithelium and lung microvascular endothelium in some situations. However, processes involving the production and consumption of NO in the airway are not totally understood. The exhaled breath condensate (EBC) is a fluid obtained by cooling exhaled air wich can be used for the investigation of NO in the distal airways. For this study we used an apparatus homemade and with low cost for collecting and obtain the EBC. The EBC was collected from patients with coronary artery disease and heart valve disease undergoing cardiac surgery with CPB in the preoperative and postoperative period (4 hours after CPB and 12, 24, 48 and 72 hours after extubation). Plasma concentrations of nitrite/nitrate in these patients were also evaluated using blood samples taken immediately after each collection of the EBC. The EBC and plasma were analyzed by chemiluminescence method to obtain the concentrations of nitrite/nitrate. The apparatus used was reproducible in clinical routine of cardiac surgery in patients under spontaneous ventilation or mechanical ventilation. There were no significant differences (p> 0.05) in concentrations of nitrite/nitrate in EBC and plasma in patients with coronary artery disease and heart valve disease in the preoperative period. Significantly higher concentrations of nitrite/nitrate (p=0.017) were found in EBC, but not in plasma of patients who used oral nitrate (isosorbide dinitrate) in the preoperative period. In patients who did not use medication containing NO donor (nitroglycerin or sodium nitroprusside) intravenously in the postoperative period there was a tendency to high concentrations of nitrite/nitrate in EBC that was numerically significant in the period of 48 hours postoperative (p=0.008). In patients who received or not NO donor in postoperative period the concentrations of nitrite/nitrate were significantly higher in periods of 48 hours (p=0.005) and 72 hours (p=0.037) after surgery. The use of nitroglycerin (GTN) in the postoperative period significantly elevated concentrations of nitrite/nitrate in the EBC in periods of 12 hours (p=0.022), 48 hours (p=0.015) and 72 hours (0.048) and plasma in period of 12 hours (p=0.045). The concentrations of nitrite/nitrate were significantly reduced (p=0.045) in the immediate postoperative period 4 hours after CPB in patients who received or not NO donor. From these results we could observe that there were no significant differences in concentrations of nitrite/nitrate in the EBC of patients with heart valve disease and coronary artery disease in the preoperative period, but we found a significant increase in these concentrations in patients who used isosorbide dinitrate orally. The concentrations of nitrite/nitrate in EBC presented a tendency to increase in postoperative period especially in patients who used GTN. The concentrations of nitrite/nitrate in plasma were significantly reduced after 4 hours of CPB in the immediate postoperative period. However, was observed that GTN tended to increase the concentration of nitrite/nitrate in plasma. These findings could help in future research providing comparative data about the role of NO in the airway and plasma in processes involving cardiac surgery in the pre and postoperative periods.

acase@tulane.edu

Angesichts der nach wie vor hohen Mortalität und Morbidität des Bronchialkarzinoms ist die Entwicklung geeigneter Methoden zur früheren Diagnostik eine wichtige Notwendigkeit, um die geringe durchschnittliche 5-Jahres-Überlebensrate von 15% – 18% zu steigern. Unter diesem Gesichtspunkt wurde in der vorliegenden Arbeit das Atemkondensat von Patienten mit Bronchialkarzinom als nicht-invasiv und kostengünstig zu gewinnendes Medium auf das Vorliegen eines potentiellen Screeningmarkers – dem methylierten Tumorsuppressor-Gen p16 – untersucht. Dazu wurde ein Versuchsablauf entwickelt, bei dem trotz des geringen DNA-Gehaltes im Atemkondensat p16-Methylierungen nachgewiesen werden konnten. Die letztendlich etablierte Methode war eine methylierungsspezifische nested-PCR mit anschließendem Restriktionsverdau durch das Restriktionsenzym BstUI. Des Weiteren erfolgte die Untersuchung von in Paraffin eingebetteten Tumorpräparaten der Patienten. In der anschließenden statistischen Auswertung wurde der Einfluss von verschiedenen Faktoren wie COPD-Grad, Tumorlage, Tumorart, Nikotinabusus und stattgehabte Chemo- oder Strahlentherapie auf den Methylierungsstatus des p16-Gens analysiert.