Comparison of Celsior and Perfadex lung preservation solutions in rat lungs subjected to 6 and 12 hours of ischemia using an ex-vivo lung perfusion system

OBJECTIVE: This study evaluated the performance of lungs that were preserved with different solutions (Celsior, Perfadex or saline) in an ex vivo rat lung perfusion system. METHODS: Sixty Wistar rats were anesthetized, anticoagulated and randomized into three groups (n = 20). The rats were subjected to antegrade perfusion via the pulmonary artery with Perfadex, Celsior, or saline, followed by 6 or 12 hours of ischemia (4 degrees C, n = 10 in each group). Respiratory mechanics, gas exchange and hemodynamics were measured at 10-minute intervals during the reperfusion of heart-lung blocks in an ex vivo system (IL2-Isolated Perfused Rat or Guinea Pig Lung System, Harvard Apparatus, Holliston, Massachusetts, USA; Hugo Sachs Elektronik, Germany) for 60 minutes. The lungs were prepared for histopathology and evaluated for edema following reperfusion. Group comparisons were performed using ANOVA and the Kruskal-Wallis test with a 5% level of significance. RESULTS: Gas exchange was not significantly different between lungs perfused with either Perfadex or Celsior at the same ischemic times, but it was very low in lungs that were preserved with saline. Airway resistance was greater in the lungs that were preserved for 12 hours. Celsior lungs that were preserved for 6 and 12 hours exhibited lower airway resistance (p = 0.01) compared to Perfadex lungs. Pulmonary artery pressure was not different between the groups, and no significant differences in histopathology and apoptosis were observed between the groups. CONCLUSIONS: Lungs that were preserved with Celsior or Perfadex exhibited similar gas exchange and histopathological findings. Airway resistance was slightly lower in the Celsior-preserved lungs compared with the Perfadex-preserved lungs.

Calorie restriction increases cerebral mitochondrial respiratory capacity in a NO center dot-mediated mechanism: Impact on neuronal survival

Calorie restriction (CR) enhances animal life span and prevents age-related diseases, including neurological decline. Recent evidence suggests that a mechanism involved in CR-induced life-span extension is NO-stimulated mitochondrial biogenesis. We examine here the effects of CR on brain mitochondrial content. CR increased eNOS and nNOS and the content of mitochondria] proteins (cytochrome c oxidase, citrate synthase, and mitofusin) in the brain. Furthermore, we established an in vitro system to study the neurological effects of CR using serum extracted from animals on this diet. In cultured neurons, CR serum enhanced nNOS expression and increased levels of nitrite (a NO product). CR serum also enhanced the levels of cytochrome c oxidase and increased citrate synthase activity and respiratory rates in neurons. CR serum effects were inhibited by L-NAME and mimicked by the NO donor SNAP. Furthermore, both CR sera and SNAP were capable of improving neuronal survival. Overall, our results indicate that CR increases mitochondrial biogenesis in a NO-mediated manner, resulting in enhanced reserve respiratory capacity and improved survival in neurons. (C) 2012 Elsevier Inc. All rights reserved.

Economic Cost of Air Pollution in Cubatao - SP Based on Health Expenses Related to Diseases of the Respiratory and Circulatory Systems

The aim of this research was to evaluate economic costs of respiratory and circulatory diseases in the municipality of Cubatao, in the state of Sao Paulo, Brazil. Data on hospital admissions and on missed working days due to hospitalization (for age group 14 to 70 years old) from the database of Sistema Unico de Sa de (SUS - Brazilian National Health System) were used. Results: Based on these data, it was calculated that R$ 22.1 million were spent in the period 2000 to 2009 due to diseases of the respiratory and circulatory systems. Part of these expenses can be directly related to the emission of atmospheric pollutants in the city. In order to estimate the costs related to air pollution, data on Cubatao were compared to data from two other municipalities that are also located at the coast side (Guaruja and Peru be), but which have little industrial activity in comparison to Cubatao. It was verified that, in both, average per capita costs were lower when compared to Cubatao, but that this difference has been decreasing in recent years.

The neonatal immune system: immunomodulation of infections in early life

The innate and adaptive immune responses in neonates are usually functionally impaired when compared with their adult counterparts. The qualitative and quantitative differences in the neonatal immune response put them at risk for the development of bacterial and viral infections, resulting in increased mortality. Newborns often exhibit decreased production of Th1-polarizing cytokines and are biased toward Th2-type responses. Studies aimed at understanding the plasticity of the immune response in the neonatal and early infant periods or that seek to improve neonatal innate immune function with adjuvants or special formulations are crucial for preventing the infectious disease burden in this susceptible group. Considerable studies focused on identifying potential immunomodulatory therapies have been performed in murine models. This article highlights the strategies used in the emerging field of immunomodulation in bacterial and viral pathogens, focusing on preclinical studies carried out in animal models with particular emphasis on neonatal-specific immune deficits.

Behavior of respiratory metabolism and pyridine nucleotide levels in Oxyrrhis marina during starvation

Máster en Oceanografía

Respiratory electron transport activity in plankton: determinants and detection

[ES]La presente tesis, se centra en el estudio del Sistema de Transporte de Electrones (ETS) en organismos del plancton marino, los factores que lo influencian la interpretación de estas mediciones y su detección mediante espectrofotometría y espectrofluorometría, en muestras oceánicas naturales y en cultivos de organismos marinos. Se pudo establecer, la biomasa, la respiración (R) y la respiración potencial (ɸ), en tres transectos en los océanos Índico y Atlántico Norte Sur. A su vez, se determino el estado fisiológico, en tres tamaños del zooplancton, midiendo la relación R/ɸ. Se exploró los efectos de la inanición sobre la R y la variación con respecto a la ɸ en el zooplancton

The Influence of Baseline Metabolic Rate and Exercise Training on Cardio-Respiratory Dynamics

Speeding the VO2 kinetics results in a reduction of the O2 deficit. Two factors might determine VO2 kinetics: oxygen delivery to muscle (Tschakovsky and Hughson 1999) and a muscle 'metabolic inertia' (Grassi et al. 1996). Therefore, in study 1 we investigated VO2 kinetics and cardiovascular system adaptations during step exercise transitions in different regions of the moderate domain. In study 2 we investigated muscle oxygenation and cardio-pulmonary adaptations during step exercise tests before, after and over a period of training. Study 1 methods: Seven subjects (26 ± 8 yr; 176 ± 5 cm; 69 ± 6 kg) performed 4 types of step transition from rest (0-50W; 0-100W) or elevate baseline (25-75W; 25-125W). GET and VO2max were assessed before testing. O2 uptake and were measured during testing. Study 2 methods: 10 subjects (25 ± 4 yr; 175 ± 9 cm; 71 ± 12 kg) performed a step transition test (0 to 100 W) before, after and during 4 weeks of endurance training (ET). VO2max and GET were assessed before and after of ET (40 minutes, 3 times a week, 60% O2max). VO2 uptake, Q and deoxyheamoglobin were measured during testing. Study 1 results: VO2 τ and the functional gain were slower in the upper regions of the moderate domain. Q increased more abruptly during rest to work condition. Q τ was faster than VO2 τ for each exercise step. Study 2 results: VO2 τ became faster after ET (25%) and particularly after 1 training session (4%). Q kinetics changed after 4 training sessions nevertheless it was always faster than VO2 τ. An attenuation in ∆[HHb] /∆VO2 was detectible. Conclusion: these investigations suggest that muscle fibres recruitment exerts a influence on the VO2 response within the moderate domain either during different forms of step transition or following ET.

Risk assessment in the first fifteen minutes: a prospective cohort study of a simple physiological scoring system in the emergency department

Introduction The survival of patients admitted to an emergency department is determined by the severity of acute illness and the quality of care provided. The high number and the wide spectrum of severity of illness of admitted patients make an immediate assessment of all patients unrealistic. The aim of this study is to evaluate a scoring system based on readily available physiological parameters immediately after admission to an emergency department (ED) for the purpose of identification of at-risk patients. Methods This prospective observational cohort study includes 4,388 consecutive adult patients admitted via the ED of a 960-bed tertiary referral hospital over a period of six months. Occurrence of each of seven potential vital sign abnormalities (threat to airway, abnormal respiratory rate, oxygen saturation, systolic blood pressure, heart rate, low Glasgow Coma Scale and seizures) was collected and added up to generate the vital sign score (VSS). VSSinitial was defined as the VSS in the first 15 minutes after admission, VSSmax as the maximum VSS throughout the stay in ED. Occurrence of single vital sign abnormalities in the first 15 minutes and VSSinitial and VSSmax were evaluated as potential predictors of hospital mortality. Results Logistic regression analysis identified all evaluated single vital sign abnormalities except seizures and abnormal respiratory rate to be independent predictors of hospital mortality. Increasing VSSinitial and VSSmax were significantly correlated to hospital mortality (odds ratio (OR) 2.80, 95% confidence interval (CI) 2.50 to 3.14, P < 0.0001 for VSSinitial; OR 2.36, 95% CI 2.15 to 2.60, P < 0.0001 for VSSmax). The predictive power of VSS was highest if collected in the first 15 minutes after ED admission (log rank Chi-square 468.1, P < 0.0001 for VSSinitial;,log rank Chi square 361.5, P < 0.0001 for VSSmax). Conclusions Vital sign abnormalities and VSS collected in the first minutes after ED admission can identify patients at risk of an unfavourable outcome.

The relationship between extravascular lung water and oxygenation in three patients with influenza A (H1N1)-induced respiratory failure

This case series reports the correlation between extravascular lung water (EVLW) and the partial arterial oxygen pressure/fractional inspiratory oxygen (PaO(2)/FiO(2)) ratio in three patients with severe influenza A (H1N1)-induced respiratory failure. All patients suffered from grave hypoxia (PaO(2), 26-42 mmHg) and were mechanically ventilated using biphasic airway pressure (PEEP, 12-15 mmHg; FiO(2), 0.8-1) in combination with prone positioning at 12 hourly intervals. All patients were monitored using the PICCO system for 8-11 days. During mechanical ventilation, a total of 62 simultaneous determinations of the PaO(2)/FiO(2) ratio and EVLW were performed. A significant correlation between EVLW and the PaO(2)/FiO(2) ratio (Spearman-rho correlation coefficient, -0.852; p < 0.001) was observed. In all patients, a decrease in EVLW was accompanied by an improvement in oxygenation. Serum lactate dehydrogenase levels were elevated in all patients and significantly correlated with EVLW during the intensive care unit stay (Spearman-rho correlation coefficient, 0.786; p < 0.001). In conclusion, EVLW seems increased in patients with severe H1N1-induced respiratory failure and appears to be closely correlated with impairments of oxygenatory function.

Lung on Chip: In vitro HGF effects on injured alveolar A549 epithelial cells in microfluidic system

Microfluidic systems have become competitive tools in the invitro modelling of diseases and promising alternatives to animal studies. They allow obtaining more invivo like conditions for cellular assays. Research in idiopathic pulmonary fibrosis could benefit from this novel methodological approach to understand the pathophysiology of the disease & develop efficient therapies. The use of hepatocyte growth factor (HGF) for alveolar reepithelisation is a promising approach. In this study, we show a new microfluidic system to analyse the effects of HGF on injured alveolar epithelial cells. Microfluidic systems in polydimethylsiloxane were fabricated by soft lithography. The alveolar A549 epithelial cells (10,000 cells) were seeded and studied in these microfluidic systems with media perfusion (1μl/30min). Injury tests were made on the cells by the perfusion with media containing H2O2 or bleomycin. The degree of injury was then assessed by a metabolic and an apoptotic assays. Wound assays were also performed with a central laminar flow of trypsin. Monitoring of wound closure with HGF vs control media was assessed. The alveolar A549 epithelial cells grew and proliferated in the microfluidic system. In the wound closure assay, the degree of wound closure after 5 hours was (53.3±1.3%) with HGF compared to (9.8±2.4%) without HGF (P <0.001). We present a novel microfluidic model that allows culture, injury and wounding of A549 epithelial cells and represents the first step towards the development of an invitro reconstitution of the alveolar-capillary interface. We were also able to confirm that HGF increased alveolar epithelial repair in this system.

Lung on a chip: Co-culture of alveolar epithelial cells and bone marrow derived stromal stem cells in a microfluidic system

Background: Microfluidics system are novel tools to study cell-cell interactions in vitro. This project focuses on the development of a new microfluidic device to co-culture alveolar epithelial cells and mesenchymal stem cells to study cellular interactions involved in healing the injured alveolar epithelium. Methods: Microfluidic systems in polydimethylsiloxane were fabricated by soft lithography. The alveolar A549 epithelial cells were seeded and injury tests were made on the cells by perfusion with media containing H2O2 or bleomycin during 6 or 18hrs. Rat Bone marrow derived stromal cells (BMSC) were then introduced into the system and cell-cell interaction was studied over 24 hrs. Results: A successful co-culture of A549 alveolar epithelial cells and BMS was achieved in the microfluidic system. The seeded alveolar epithelial cells and BMSC adhered to the bottom surface of the microfluidic device and proliferated under constant perfusion. Epithelial injury to mimic mechanisms seen in idiopathic pulmonary fibrosis was induced in the microchannels by perfusing with H2O2 or bleomycin. Migration of BMSC towards the injured epithelium was observed as well as cell-cell interaction between the two cell types was also seen. Conclusion: We demonstrate a novel microfluidic device aimed at showing interactions between different cell types on the basis of a changing microenvironment. Also we were able to confirm interaction between injured alvolar epithelium and BMSC, and showed that BMSC try to heal the injured epitelium.

A realistic validation study of a new nitrogen multiple-breath washout system

Background For reliable assessment of ventilation inhomogeneity, multiple-breath washout (MBW) systems should be realistically validated. We describe a new lung model for in vitro validation under physiological conditions and the assessment of a new nitrogen (N2)MBW system. Methods The N2MBW setup indirectly measures the N2 fraction (FN2) from main-stream carbon dioxide (CO2) and side-stream oxygen (O2) signals: FN2 = 1−FO2−FCO2−FArgon. For in vitro N2MBW, a double chamber plastic lung model was filled with water, heated to 37°C, and ventilated at various lung volumes, respiratory rates, and FCO2. In vivo N2MBW was undertaken in triplets on two occasions in 30 healthy adults. Primary N2MBW outcome was functional residual capacity (FRC). We assessed in vitro error (√[difference]2) between measured and model FRC (100–4174 mL), and error between tests of in vivo FRC, lung clearance index (LCI), and normalized phase III slope indices (Sacin and Scond). Results The model generated 145 FRCs under BTPS conditions and various breathing patterns. Mean (SD) error was 2.3 (1.7)%. In 500 to 4174 mL FRCs, 121 (98%) of FRCs were within 5%. In 100 to 400 mL FRCs, the error was better than 7%. In vivo FRC error between tests was 10.1 (8.2)%. LCI was the most reproducible ventilation inhomogeneity index. Conclusion The lung model generates lung volumes under the conditions encountered during clinical MBW testing and enables realistic validation of MBW systems. The new N2MBW system reliably measures lung volumes and delivers reproducible LCI values.

An optimized in vitro model of the respiratory tract wall to study particle cell interactions

As a part of the respiratory tissue barrier, lung epithelial cells play an important role against the penetration of the body by inhaled particulate foreign materials. In most cell culture models, which are designed to study particle-cell interactions, the cells are immersed in medium. This does not reflect the physiological condition of lung epithelial cells which are exposed to air, separated from it only by a very thin liquid lining layer with a surfactant film at the air-liquid interface. In this study, A549 epithelial cells were grown on microporous membranes in a two chamber system. After the formation of a confluent monolayer the cells were exposed to air. The morphology of the cells and the expression of tight junction proteins were studied with confocal laser scanning and transmission electron microscopy. Air-exposed cells maintained monolayer structure for 2 days, expressed tight junctions and developed transepithelial electrical resistance. Surfactant was produced and released at the apical side of the air-exposed epithelial cells. In order to study particle-cell interactions fluorescent 1 microm polystyrene particles were sprayed over the epithelial surface. After 4 h, 8.8% of particles were found inside the epithelium. This fraction increased to 38% after 24 h. During all observations, particles were always found in the cells but never between them. In this study, we present an in vitro model of the respiratory tract wall consisting of air-exposed lung epithelial cells covered by a liquid lining layer with a surfactant film to study particle-cell interactions.

Alteration of the pulmonary surfactant system in full-term infants with hereditary ABCA3 deficiency

RATIONALE: ABCA3 mutations are known to cause fatal surfactant deficiency. OBJECTIVE: We studied ABCA3 protein expression in full-term newborns with unexplained respiratory distress syndrome (URDS) as well as the relevance of ABCA3 mutations for surfactant homeostasis. METHODS: Lung tissue of infants with URDS was analyzed for the expression of ABCA3 in type II pneumocytes. Coding exons of the ABCA3 gene were sequenced. Surfactant protein expression was studied by immunohistochemistry, immunoelectron microscopy, and Western blotting. RESULTS: ABCA3 protein expression was found to be greatly reduced or absent in 10 of 14 infants with URDS. Direct sequencing revealed distinct ABCA3 mutations clustering within vulnerable domains of the ABCA3 protein. A strong expression of precursors of surfactant protein B (pro-SP-B) but only low levels and aggregates of mature surfactant protein B (SP-B) within electron-dense bodies in type II pneumocytes were found. Within the matrix of electron-dense bodies, we detected precursors of SP-C (pro-SP-C) and cathepsin D. SP-A was localized in small intracellular vesicles, but not in electron-dense bodies. SP-A and pro-SP-B were shown to accumulate in the intraalveolar space, whereas mature SP-B and SP-C were reduced or absent, respectively. CONCLUSION: Our data provide evidence that ABCA3 mutations are associated not only with a deficiency of ABCA3 but also with an abnormal processing and routing of SP-B and SP-C, leading to severe alterations of surfactant homeostasis and respiratory distress syndrome. To identify infants with hereditary ABCA3 deficiency, we suggest a combined diagnostic approach including immunohistochemical, ultrastructural, and mutation analysis.

Effects of tributyltin on the immune system of Japanese flounder (Paralichthys olivaceus)

Effects of tributyltin (TBT) which has been used for antifouling paint of ship's hulls and fishing nets on the immune system in Japanese flounder (Paralichthys olivaceus) were investigated. After short-term exposure to a high level of TBT, leucocytes in the head kidney from 1-year-old flounder were examined for the proportion of neutrophils in total leucocytes. Also examined were their respiratory burst activities using flow cytometry, the reduction of nitroblue tetrazolium (NBT) and lysozyme activities. Furthermore, long-term exposures to a relatively low level of TBT using young flounder were also carried out. The proportion of neutrophils in total leucocytes prepared from head kidney in each fish exposed to TBT at 20 microg/L for 5 days and the reduction of NBT by leucocytes prepared from the same experimental conditions increase compared to the control group. The contents were 42.0+/-6.8 and 52.5+/-6.3%, respectively. Significant differences of the NBT reduction were observed between 0 and 20 microg/L TBT exposure groups. On the other hand, the respiratory burst activity of cells in the exposure group clearly showed a tendency to decrease compared to the control group. Furthermore, high level of TBT also inhibited lysozyme activity which plays an important role for the bacteriocidal procedures. However, similar results were not obtained in the exposure group with a relatively low level of TBT. To determine the immunotoxic effects of TBT, infection experiments using pathogens which are naturally occurring should be further investigated.