Evaluation of systemic inflammation in patients being weaned from mechanical ventilation.

OBJECTIVES: The aim of this study was to evaluate systemic inflammatory factors and their relation to success or failure in a spontaneous ventilation test. METHODS: This cross-sectional study included a sample of 54 adult patients. Demographic data and clinical parameters were collected, and blood samples were collected in the first minute of the spontaneous ventilation test to evaluate interleukin (IL)-1ß, IL-6, IL-8, and IL-10, tumour necrosis factor alpha (TNFα) and C-reactive protein. RESULTS: Patients who experienced extubation failure presented a lower rapid shallow breathing index than those who passed, and these patients also showed a significant increase in C-reactive protein 48 hours after extubation. We observed, moreover, that each unit increase in inflammatory factors led to a higher risk of spontaneous ventilation test failure, with a risk of 2.27 (1.001 - 4.60, p=0.049) for TNFα, 2.23 (1.06 - 6.54, p=0.037) for IL-6, 2.66 (1.06 - 6.70, p=0.037) for IL-8 and 2.08 (1.01 - 4.31, p=0.04) for IL-10, and the rapid shallow breathing index was correlated with IL-1 (r=-0.51, p=0.04). CONCLUSIONS: C-reactive protein is increased in patients who fail the spontaneous ventilation test, and increased ILs are associated with a greater prevalence of failure in this process; the rapid shallow breathing index may not be effective in patients who present systemic inflammation.

Evaluation of systemic inflammation in patients being weaned from mechanical ventilation

OBJECTIVES: The aim of this study was to evaluate systemic inflammatory factors and their relation to success or failure in a spontaneous ventilation test. METHODS: This cross-sectional study included a sample of 54 adult patients. Demographic data and clinical parameters were collected, and blood samples were collected in the first minute of the spontaneous ventilation test to evaluate interleukin (IL)-1β, IL-6, IL-8, and IL-10, tumour necrosis factor alpha (TNFα) and C-reactive protein. RESULTS: Patients who experienced extubation failure presented a lower rapid shallow breathing index than those who passed, and these patients also showed a significant increase in C-reactive protein 48 hours after extubation. We observed, moreover, that each unit increase in inflammatory factors led to a higher risk of spontaneous ventilation test failure, with a risk of 2.27 (1.001 - 4.60, p=0.049) for TNFα, 2.23 (1.06 - 6.54, p=0.037) for IL-6, 2.66 (1.06 - 6.70, p=0.037) for IL-8 and 2.08 (1.01 - 4.31, p=0.04) for IL-10, and the rapid shallow breathing index was correlated with IL-1 (r=-0.51, p=0.04). CONCLUSIONS: C-reactive protein is increased in patients who fail the spontaneous ventilation test, and increased ILs are associated with a greater prevalence of failure in this process; the rapid shallow breathing index may not be effective in patients who present systemic inflammation.

Uncoupling protein-2 mRNA expression in mice subjected to intermittent hypoxia.

OBJECTIVE: To investigate the effect of intermittent hypoxia-a model of obstructive sleep apnea (OSA)-on pancreatic expression of uncoupling protein-2 (UCP2), as well as on glycemic and lipid profiles, in C57BL mice. METHODS: For 8 h/day over a 35-day period, male C57BL mice were exposed to intermittent hypoxia (hypoxia group) or to a sham procedure (normoxia group). The intermittent hypoxia condition involved exposing mice to an atmosphere of 92% N and 8% CO2 for 30 s, progressively reducing the fraction of inspired oxygen to 8 ± 1%, after which they were exposed to room air for 30 s and the cycle was repeated (480 cycles over the 8-h experimental period). Pancreases were dissected to isolate the islets. Real-time PCR was performed with TaqMan assays. RESULTS: Expression of UCP2 mRNA in pancreatic islets was 20% higher in the normoxia group than in the hypoxia group (p = 0.11). Fasting serum insulin was higher in the hypoxia group than in the normoxia group (p = 0.01). The homeostasis model assessment of insulin resistance indicated that, in comparison with the control mice, the mice exposed to intermittent hypoxia showed 15% lower insulin resistance (p = 0.09) and 21% higher pancreatic ß-cell function (p = 0.01). Immunohistochemical staining of the islets showed no significant differences between the two groups in terms of the area or intensity of α- and ß-cell staining for insulin and glucagon. CONCLUSIONS: To our knowledge, this is the first report of the effect of intermittent hypoxia on UCP2 expression. Our findings suggest that UCP2 regulates insulin production in OSA. Further study of the role that UCP2 plays in the glycemic control of OSA patients is warranted.

Uncoupling protein-2 mRNA expression in mice subjected to intermittent hypoxia / Expressão do mRNA da uncoupling protein-2 em camundongos submetidos à hipóxia intermitente

Objective: To investigate the effect of intermittent hypoxia-a model of obstructive sleep apnea (OSA)-on pancreatic expression of uncoupling protein-2 (UCP2), as well as on glycemic and lipid profiles, in C57BL mice. Methods: For 8 h/day over a 35-day period, male C57BL mice were exposed to intermittent hypoxia (hypoxia group) or to a sham procedure (normoxia group). The intermittent hypoxia condition involved exposing mice to an atmosphere of 92% N and 8% CO2 for 30 s, progressively reducing the fraction of inspired oxygen to 8 ± 1%, after which they were exposed to room air for 30 s and the cycle was repeated (480 cycles over the 8-h experimental period). Pancreases were dissected to isolate the islets. Real-time PCR was performed with TaqMan assays. Results: Expression of UCP2 mRNA in pancreatic islets was 20% higher in the normoxia group than in the hypoxia group (p = 0.11). Fasting serum insulin was higher in the hypoxia group than in the normoxia group (p = 0.01). The homeostasis model assessment of insulin resistance indicated that, in comparison with the control mice, the mice exposed to intermittent hypoxia showed 15% lower insulin resistance (p = 0.09) and 21% higher pancreatic β-cell function (p = 0.01). Immunohistochemical staining of the islets showed no significant differences between the two groups in terms of the area or intensity of α- and β-cell staining for insulin and glucagon. Conclusions: To our knowledge, this is the first report of the effect of intermittent hypoxia on UCP2 expression. Our findings suggest that UCP2 regulates insulin production in OSA. Further study of the role that UCP2 plays in the glycemic control of OSA patients is warranted. .

Objetivo: Investigar o efeito da hipóxia intermitente com um modelo de apneia obstrutiva do sono (AOS) sobre a expressão de uncoupling protein-2 (UCP2), assim como sobre perfis glicêmicos e lipídicos, em camundongos C57BL. Métodos: Camundongos C57BL machos foram expostos a hipóxia intermitente ou hipóxia simulada (grupo controle) 8 h/dia durante 35 dias. A condição de hipóxia intermitente envolveu a exposição dos camundongos a uma atmosfera de 92% de N e 8% de CO2 por 30 s, com redução progressiva de fração de O2 inspirado até 8 ± 1%, seguida por exposição a ar ambiente por 30 s e repetições do ciclo (480 ciclos no período experimental de 8 h). Os pâncreas foram dissecados para isolar as ilhotas. Foi realizada PCR em tempo real utilizando o método TaqMan. Resultados: A expressão do mRNA da UCP2 nas ilhotas pancreáticas foi 20% maior no grupo controle que no grupo hipóxia (p = 0,11). A insulina sérica de jejum foi maior no grupo hipóxia do que no grupo controle (p = 0,01). O modelo de avaliação da homeostase de resistência à insulina indicou que, em comparação com os camundongos controle, aqueles expostos à hipóxia intermitente apresentaram 15% menor resistência à insulina (p = 0,09) e 21% maior função das células beta (p = 0,01). A coloração das ilhotas pancreáticas por imuno-histoquímica não mostrou diferenças significativas entre os grupos em termos da área ou da intensidade das células alfa e beta, marcadas por insulina e glucagon. Conclusões: Segundo nosso conhecimento, esta é a primeira descrição do efeito da hipóxia intermitente sobre a expressão da UCP2. Nossos achados sugerem que UCP2 regula a produção de insulina na AOS. Futuras investigações sobre o papel da UCP2 no controle glicêmico em pacientes com AOS são justificadas. .

Antioxidants inhibit the inflammatory and apoptotic processes in an intermittent hypoxia model of sleep apnea.

BACKGROUND: Sleep apnea causes intermittent hypoxia (IH). We aimed to investigate the proteins related to oxidative stress, inflammation and apoptosis in liver tissue subjected to IH as a simulation of sleep apnea in conjunction with the administration of either melatonin (MEL, 200 µL/kg) or N-acetylcysteine (NAC, 10 mg/kg). METHODS: Seventy-two adult male Balb-C mice were divided: simulation of IH (SIH), SIH + MEL, SIH + NAC, IH, IH + MEL and IH + NAC. The animals were subjected to simulations of sleep apnea for 8 h a day for 35 days. The data were analyzed with ANOVA and Tukey tests with the significance set at p < 0.05. RESULTS: In IH, there was a significant increase in oxidative stress and expression of HIF-1a. In addition, we observed increase in the activation levels of NF-kB. This increase may be responsible for the increased expression of TNF-alpha and iNOS as well as the significant increase of VEGF signaling and expression of caspase-3 and caspase-6, which suggests an increase in apoptosis. In the groups treated with antioxidants, the analysis showed that the enzyme activity and protein levels were similar to those of the non-simulated group. CONCLUSIONS: Thus, we show that IH causes liver inflammation and apoptosis, which may be protected with either MEL or NAC.

N-acetylcysteine administration confers lung protection in different phases of lung ischaemia-reperfusion injury.

OBJECTIVES: To verify the effects of N-acetylcysteine (NAC) administered before and after ischaemia in an animal model of lung ischaemia-reperfusion (IR) injury. METHODS: Twenty-four Wistar rats were subjected to an experimental model of selective left pulmonary hilar clamping for 45 min followed by 2 h of reperfusion. The animals were divided into four groups: control group (SHAM), ischaemia-reperfusion, N-acetylcysteine-preischaemia (NAC-Pre) and NAC-postischaemia (NAC-Post). We recorded the haemodynamic parameters, blood gas analysis and histology. We measured the thiobarbituric acid reactive substances concentration; the expression of superoxide dismutase (SOD), inducible nitric oxide synthase (iNOS), nitrotyrosine, cleaved caspase 3, nuclear factor κB (NF-κB), NF-kappa-B inhibitor alpha (IκB-α), tumour necrosis factor alpha (TNF-α) and interleukin-1 beta (IL-1ß); myeloperoxidase activity (MPO). RESULTS: No significant differences were observed in the haemodynamic parameters, blood gas analysis and SOD activity among the groups. Lipid peroxidation was significantly higher in the IR and NAC-Pre groups (P < 0.01). The expression of nitrotyrosine, cleaved caspase 3, NF-κB, IκB-α, TNF-α and IL-1ß were significantly higher in the IR group when compared with the SHAM and NAC groups (P < 0.01). The NAC-Pre group showed a significantly higher expression of these proteins when compared with the SHAM and NAC-Post groups (P < 0.05). After reperfusion, the expression of iNOS increased almost uniformly in all groups when compared with the SHAM group (P < 0.01). The histological analysis showed fewer inflammatory cells in the NAC groups. CONCLUSIONS: The intravenous administration of NAC demonstrated protective properties against lung IR injury. The use of NAC immediately after reperfusion potentiates its protective effects.

Effect of the systemic administration of methylprednisolone on the lungs of brain-dead donor rats undergoing pulmonary transplantation.

OBJECTIVE: Most lung transplants are obtained from brain-dead donors. The physiopathology of brain death involves hemodynamics, the sympathetic nervous system, and inflammatory mechanisms. Administering methylprednisolone 60 min after inducing brain death in rats has been shown to modulate pulmonary inflammatory activity. Our objective was to evaluate the effects of methylprednisolone on transplanted rat lungs from donors treated 60 min after brain death. METHODS: Twelve Wistar rats were anesthetized, and brain death was induced. They were randomly divided into two groups (n=6), namely a control group, which was administered saline solution, and a methylprednisolone group, which received the drug 60 min after the induction of brain death. All of the animals were observed and ventilated for 2 h prior to being submitted to lung transplantation. We evaluated the hemodynamic and blood gas parameters, histological score, lung tissue levels of thiobarbituric acid-reactive substances, level of superoxide dismutase, level of tumor necrosis factor-alpha, and level of interleukin-1 beta. RESULTS: After transplantation, a significant reduction in the levels of tumor necrosis factor-alpha and IL-1ß was observed in the group that received methylprednisolone (p=0.0084 and p=0.0155, respectively). There were no significant differences in tumor necrosis factor-alpha and superoxide dismutase levels between the control and methylprednisolone groups (p=0.2644 and p=0.7461, respectively). There were no significant differences in the blood gas parameters, hemodynamics, and histological alterations between the groups. CONCLUSION: The administration of methylprednisolone after brain death in donor rats reduces inflammatory activity in transplanted lungs but has no influence on parameters related to oxidative stress.

Effect of the systemic administration of methylprednisolone on the lungs of brain-dead donor rats undergoing pulmonary transplantation

OBJECTIVE: Most lung transplants are obtained from brain-dead donors. The physiopathology of brain death involves hemodynamics, the sympathetic nervous system, and inflammatory mechanisms. Administering methylprednisolone 60 min after inducing brain death in rats has been shown to modulate pulmonary inflammatory activity. Our objective was to evaluate the effects of methylprednisolone on transplanted rat lungs from donors treated 60 min after brain death. METHODS: Twelve Wistar rats were anesthetized, and brain death was induced. They were randomly divided into two groups (n = 6), namely a control group, which was administered saline solution, and a methylprednisolone group, which received the drug 60 min after the induction of brain death. All of the animals were observed and ventilated for 2 h prior to being submitted to lung transplantation. We evaluated the hemodynamic and blood gas parameters, histological score, lung tissue levels of thiobarbituric acid-reactive substances, level of superoxide dismutase, level of tumor necrosis factor-alpha, and level of interleukin-1 beta. RESULTS: After transplantation, a significant reduction in the levels of tumor necrosis factor-alpha and IL-1β was observed in the group that received methylprednisolone (p = 0.0084 and p = 0.0155, respectively). There were no significant differences in tumor necrosis factor-alpha and superoxide dismutase levels between the control and methylprednisolone groups (p = 0.2644 and p = 0.7461, respectively). There were no significant differences in the blood gas parameters, hemodynamics, and histological alterations between the groups. CONCLUSION: The administration of methylprednisolone after brain death in donor rats reduces inflammatory activity in transplanted lungs but has no influence on parameters related to oxidative stress. .

Downregulation of uncoupling protein-1 mRNA expression and hypoadiponectinemia in a mouse model of sleep apnea.

PURPOSE: The knowledge on the effect of intermittent hypoxia on adipose tissue-mediated processes is incipient. The aim of the present study was to assess the effect of a sleep apnea model on a limited set of specific molecular, biochemical, histological, and behavioral parameters of adipose tissue function. METHODS: Mice were exposed to either intermittent hypoxia or sham hypoxia during 8 h a day for 37 days. Uncoupling protein-1 expression in brown adipose tissue was measured by real-time PCR and immunohistochemistry. Digital quantification of adipose cells and immunohistochemistry of uncoupling protein-1 were performed to determine cell dimensions, positive area, and staining intensity. Serum levels of leptin, adiponectin, and cortisol were measured by ELISA. RESULTS: In comparison with the control group, animals in the hypoxia group had significantly lower chow ingestion, weight gain, and smaller white and brown adipocytes on histological examination. Adiponectin levels were also lower in the hypoxia group. Uncoupling protein-1 mRNA was abolished in the mice exposed to hypoxia; accordingly, fewer cells positive for uncoupling protein-1 and lighter staining intensity were observed in brown adipocytes. CONCLUSIONS: An experimental model of sleep apnea produced changes in uncoupling protein-1 expression and adiponectin levels. These results confirm previous findings on the response of brown adipose tissue to intermittent hypoxia and indicate a yet-unknown interference of intermittent hypoxia on energy control, which may participate in the propensity to weight gain observed in patients with sleep apnea. Brown adipose tissue activity in this patient population needs to be further investigated.

Serum levels and polymorphisms of matrix metalloproteinases (MMPs) in carotid artery atherosclerosis: higher MMP-9 levels are associated with plaque vulnerability.

CONTEXT: Matrix metalloproteinases are involved in atherosclerosis and plaque vulnerability. OBJECTIVE: To investigate serum levels and genetic polymorphisms of matrix metalloproteinases (MMPs) -1, -3 and -9 in patients submitted to carotid endarterectomy. METHODS: Genetic polymorphisms were evaluated using polymerase chain reaction (PCR-RFLP); serum levels were measured using ELISA; histological sections were stained with Picrosirius Red to analyze the fibrous cap thickness, lipid core and collagen content and with hematoxylin--eosin to detect the presence of intraplaque hemorrhage. RESULTS: MMP-9 serum levels were significantly higher in patients with a thinner fibrous cap (p = 0.033) or acute or recent intraplaque hemorrhage (p = 0.008) on histology, as well as in patients with previous stroke (p = 0.009) or peripheral vascular disease (p = 0.049). No consistent associations were observed between different MMP genotypes and fibrous cap thickness, lipid core, collagen content or intraplaque hemorrhage. CONCLUSIONS: MMP-9 serum levels were consistently associated with markers of carotid atherosclerosis and lesion vulnerability, whereas specific MMP genotypes were not.

Neurobehavioral effects of L-carnitine and its ability to modulate genotoxicity and oxidative stress biomarkers in mice.

L-Carnitine, a natural vitamin-like compound supplied to the body by biosynthesis and dietary sources, has been shown to exert beneficial effects in disorders affecting cardiovascular, urinary, and nervous systems. However, the paucity of data on its effects does not guarantee the safe use of L-carnitine as a nutritional supplement, and further pre-clinical studies are required to assess toxicological aspects. The present study evaluated the effects of L-carnitine (10, 50 or, 100 mg/kg) in mice, in the open field test. Also, lipoperoxidation was assessed measuring thiobarbituric acid reactive substances (TBARS) and genotoxic/antigenotoxic activities were evaluated using the comet assay in several tissues. L-Carnitine 50 mg/kg impaired exploration, though with no effects on habituation to a novel environment. L-Carnitine increased TBARS in the brain and liver tissues, but it did not induce genotoxicity in any tissue. In ex vivo comet assay, a decrease in DNA damage in the blood and liver tissues was observed, while the opposite occurred in the brain tissue. In conclusion, L-carnitine may increase lipid peroxidation, though without inducing genotoxic effects, protect DNA against endogenous and induced oxidative damages in blood and liver; however, L-carnitine impaired exploratory behavior and increased the vulnerability of the brain tissue to oxidative stress, suggesting that the excessive consumption of L-carnitine may promote deleterious effects on the central nervous system.

Effects of methylprednisolone on inflammatory activity and oxidative stress in the lungs of brain-dead rats.

OBJECTIVE: To evaluate the effects that early and late systemic administration of methylprednisolone have on lungs in a rat model of brain death. METHODS: Twenty-four male Wistar rats were anesthetized and randomly divided into four groups (n = 6 per group): sham-operated (sham); brain death only (BD); brain death plus methylprednisolone (30 mg/kg i.v.) after 5 min (MP5); and brain death plus methylprednisolone (30 mg/kg i.v.) after 60 min (MP60). In the BD, MP5, and MP60 group rats, we induced brain death by inflating a balloon catheter in the extradural space. All of the animals were observed and ventilated for 120 min. We determined hemodynamic and arterial blood gas variables; wet/dry weight ratio; histological score; levels of thiobarbituric acid reactive substances (TBARS); superoxide dismutase (SOD) activity; and catalase activity. In BAL fluid, we determined differential white cell counts, total protein, and lactate dehydrogenase levels. Myeloperoxidase activity, lipid peroxidation, and TNF-α levels were assessed in lung tissue. RESULTS: No significant differences were found among the groups in terms of hemodynamics, arterial blood gases, wet/dry weight ratio, BAL fluid analysis, or histological score-nor in terms of SOD, myeloperoxidase, and catalase activity. The levels of TBARS were significantly higher in the MP5 and MP60 groups than in the sham and BD groups (p < 0.001). The levels of TNF-α were significantly lower in the MP5 and MP60 groups than in the BD group (p < 0.001). CONCLUSIONS: In this model of brain death, the early and late administration of methylprednisolone had similar effects on inflammatory activity and lipid peroxidation in lung tissue.

Efeitos da metilprednisolona na atividade inflamatória e estresse oxidativo nos pulmões de ratos com morte cerebral / Effects of methylprednisolone on inflammatory activity and oxidative stress in the lungs of brain-dead rats

OBJETIVO: Avaliar os efeitos da administração sistêmica precoce e tardia de metilprednisolona nos pulmões em um modelo de morte encefálica em ratos. MÉTODOS: Vinte e quatro ratos Wistar machos foram anestesiados e randomizados em quatro grupos (n = 6 por grupo): sham, somente morte encefálica (ME), metilprednisolona i.v. (30 mg/kg) administrada 5 min após a morte encefálica (MP5) e 60 min após a morte encefálica (MP60). Os grupos ME, MP5 e MP60 foram submetidos à morte encefálica por insuflação de um balão no espaço extradural. Todos os animais foram observados e ventilados durante 120 min. Foram determinadas variáveis hemodinâmicas e gasométricas, relação peso úmido/seco, escore histológico, thiobarbituric acid reactive substances (TBARS, substâncias reativas ao ácido tiobarbitúrico), atividade de superóxido dismutase (SOD) e de catalase, assim como contagem diferencial de células brancas, proteína total e nível de desidrogenase lática no LBA. A atividade da mieloperoxidase, peroxidação lipídica e níveis de TNF-α foram avaliados no tecido pulmonar. RESULTADOS: Não foram observadas diferenças significativas nas variáveis hemodinâmicas e gasométricas, relação peso úmido/seco, análises do LBA, escore histológico, SOD, mieloperoxidase e catalase entre os grupos. Os níveis de TBARS foram significativamente maiores nos grupos MP5 e MP60 do que nos grupos sham e ME (p < 0,001). Os níveis de TNF-α foram significativamente menores nos grupos MP5 e MP60 do que no grupo ME (p < 0,001). CONCLUSÕES: Neste modelo de morte cerebral, a administração precoce e tardia de metilprednisolona apresentou efeitos semelhantes sobre a inflamação e a peroxidação lipídica no tecido pulmonar.

OBJECTIVE: To evaluate the effects that early and late systemic administration of methylprednisolone have on lungs in a rat model of brain death. METHODS: Twenty-four male Wistar rats were anesthetized and randomly divided into four groups (n = 6 per group): sham-operated (sham); brain death only (BD); brain death plus methylprednisolone (30 mg/kg i.v.) after 5 min (MP5); and brain death plus methylprednisolone (30 mg/kg i.v.) after 60 min (MP60). In the BD, MP5, and MP60 group rats, we induced brain death by inflating a balloon catheter in the extradural space. All of the animals were observed and ventilated for 120 min. We determined hemodynamic and arterial blood gas variables; wet/dry weight ratio; histological score; levels of thiobarbituric acid reactive substances (TBARS); superoxide dismutase (SOD) activity; and catalase activity. In BAL fluid, we determined differential white cell counts, total protein, and lactate dehydrogenase levels. Myeloperoxidase activity, lipid peroxidation, and TNF-α levels were assessed in lung tissue. RESULTS: No significant differences were found among the groups in terms of hemodynamics, arterial blood gases, wet/dry weight ratio, BAL fluid analysis, or histological score-nor in terms of SOD, myeloperoxidase, and catalase activity. The levels of TBARS were significantly higher in the MP5 and MP60 groups than in the sham and BD groups (p < 0.001). The levels of TNF-α were significantly lower in the MP5 and MP60 groups than in the BD group (p < 0.001). CONCLUSIONS: In this model of brain death, the early and late administration of methylprednisolone had similar effects on inflammatory activity and lipid peroxidation in lung tissue.

Endobronchial perfluorocarbon administration decreases lung injury in an experimental model of ischemia and reperfusion.

OBJECTIVE: To verify the effects of liquid endobronchial perfluorocarbon (PFC) administered before reperfusion in an animal model of lung ischemia-reperfusion injury. METHODS: Eighteen Wistar rats were subjected to an experimental model of selective left pulmonary artery clamping for 45 min followed by reperfusion for 2 h. The animals were divided into three groups: the ischemia-reperfusion (IR) group, the sham group, and the PFC group. We recorded the hemodynamic parameters, blood gas analysis, and histology. A Western blot assay was used to measure the inducible nitric oxide synthase, caspase 3, and nuclear factor қB (subunit p65) activities. Lipid peroxidation was assessed by the thiobarbituric acid reactive substances assay and the activity of the antioxidant enzyme superoxide dismutase. RESULTS: No significant differences were observed in lipid peroxidation among the groups. The superoxide dismutase activity was increased (P < 0.05) in the PFC-treated group. The expressions of nuclear factor қB, inducible nitric oxide synthase, and caspase 3 were significantly lower in the PFC group than in the IR group (P < 0.05). The histologic analysis showed a reduction in lung injuries in the PFC group compared with the sham and IR groups. CONCLUSION: The use of endobronchial PFC reduces the inflammatory response, preserves the alveolar structure, and protects the lungs against the hazardous effects of ischemia-reperfusion injuries.

Quercetin decreases liver damage in mice with non-alcoholic steatohepatitis.

Non-alcoholic steatohepatitis (NASH) is a frequent condition in obese patients that may progress to end-stage liver disease. This study was designed to evaluate the modulation of this condition by use of quercetin (Q), a flavonoid largely found in vegetable foods, with known anti-inflammatory and antioxidant properties, in the experimental model of non-alcoholic steatohepatitis (NASH) using a diet deficient in methionine and choline (MCD). Male C57BL6 mice were divided into four groups (n = 16): (i) Control plus vehicle (control ration plus carboxymethylcellulose 1% used as vehicle, CO + V); (ii) Control ration plus Q 50 mg/kg (CO + Q); (iii) MCD diet plus vehicle (NASH + V); and (iv) MCD diet plus Q (NASH + Q). Diets were administered for 4 weeks. At the end of the experimental period, liver alterations, bioindicators of oxidative stress and DNA damage were assessed. NASH was diagnosed in 100% of the mice that were fed the MCD diet. In addition, a significant increase in DNA damage in liver tissue from NASH + V group was observed in comparison with CO + V. The group NASH + Q showed a significant decrease in hepatic damage enzymes, lipoperoxidation, DNA damage and a lower degree of macrovesicular steatosis, ballooning and inflammatory process. These findings suggest that Q may have protective effects by improving liver integrity in NASH.

Endobronchial perfluorocarbon reduces inflammatory activity before and after lung transplantation in an animal experimental model.

BACKGROUND: The aim of this study was to evaluate the use of liquid perfluorocarbon (PFC) as an adjuvant substance for lung preservation and assess its role in pulmonary protection after transplantation. METHODS: Seventy-two rat lungs were flushed with low-potassium dextran (LPD) solution and randomized into three main groups: control with LPD alone and experimental with 3 (PFC3) and 7 mL/kg (PFC7) of endobronchial PFC instilled just after harvest. Each group was divided into four subgroups according to preservation time (3, 6, 12, and 24 hours). Afterwards, we performed lung transplantation using rat lungs preserved for 12 hours with LPD alone or with 7 mL/kg of endobronchial PFC. RESULTS: There was a significant increase in oxidative stress in the control group at 6 h of cold ischemic time compared with the PFC3 and PFC7 groups. The apoptotic activity and NF-κB expression were significantly higher in the control group compared with the PFC groups at 3, 12, and 24 h of cold preservation. After transplantation, the NF-κB, iNOS, and nitrotyrosine expression as well as caspase 3 activity were significantly lower in the PFC groups. CONCLUSION: The use of endobronchial PFC as an adjuvant to the current preservation strategy improved graft viability.

When is injury potentially reversible in a lung ischemia-reperfusion model?

OBJECTIVE: To verify the impact of ischemic time on lung cell viability in an experimental model of lung ischemia-reperfusion (IR) injury and its repercussion on lung performance after reperfusion. METHODS: Twenty-four animals were subjected to selective clamping of the left pulmonary artery and divided into four groups (n = 6) according to ischemic time: 15 (IR15), 30 (IR30), 45 (IR45), and 60 min (IR60). All animals were observed for 120 min after reperfusion. The hemodynamics, arterial blood gases measurements, and histologic changes were analyzed. Immunofluorescence assays for caspase 3 and annexin V were performed. Lipid peroxidation was assessed by thiobarbituric acid-reactive substances, and caspase 3 activity was assessed by colorimetric extract. RESULTS: The partial pressure of arterial oxygen significantly decreased at the end of the observation period in the IR30, IR45, and IR60 groups (P < 0.05). The final mean arterial pressure significantly decreased in the IR60 group (P < 0.05). We observed a significant increase in caspase 3 activity and caspase 3-positive cells by immunofluorescence in the IR45 group compared with the other groups (P < 0.05). Additionally, there was an increase in necrotic cells assessed by annexin V in the IR60 group. The histologic score did not show differences among the different groups. CONCLUSIONS: The degree of cell damage had a negative impact on lung performance. Sixty minutes of lung ischemia and posterior reperfusion resulted in an increased number of necrotic cells, suggesting that these cells may not be able to reverse the effects of the IR injury because of the lack of viable cells.

Altered aquaporins in the brains of mice submitted to intermittent hypoxia model of sleep apnea.

Rostral fluid displacement has been proposed as a pathophysiologic mechanism of both central and obstructive sleep apnea. Aquaporins are membrane proteins that regulate water transport across the cell membrane and are involved in brain edema formation and resolution. The present study investigated the effect of intermittent hypoxia (IH), a model of sleep apnea, on brain aquaporins. Mice were exposed to intermittent hypoxia to a nadir of 7% oxygen fraction. Brain water content, Aquaporin-1 and Aquaporin-3 were measured in the cerebellum and hippocampus. Hematoxylin-eosin and immunohistochemistry stainings were performed to evaluate cell damage. Compared to the sham group, the hypoxia group presented higher brain water content, lower levels of Aquaporin-1 and similar levels of Aquaporin-3. Immunoreactivity to GFAP and S100B was stronger in the hypoxia group in areas of extensive gliosis, compatible with cytotoxic edema. These findings, although preliminary, indicate an effect of IH on aquaporins levels. Further investigation about the relevance of these data on the pathophysiology of OSA is warranted.

Simulating sleep apnea by exposure to intermittent hypoxia induces inflammation in the lung and liver.

Sleep apnea is a breathing disorder that results from momentary and cyclic collapse of the upper airway, leading to intermittent hypoxia (IH). IH can lead to the formation of free radicals that increase oxidative stress, and this mechanism may explain the association between central sleep apnea and nonalcoholic steatohepatitis. We assessed the level of inflammation in the lung and liver tissue from animals subjected to intermittent hypoxia and simulated sleep apnea. A total of 12 C57BL/6 mice were divided into two groups and then exposed to IH (n = 6) or a simulated IH (SIH) (n = 6) for 35 days. We observed an increase in oxidative damage and other changes to endogenous antioxidant enzymes in mice exposed to IH. Specifically, the expression of multiple transcription factors, including hypoxia inducible factor (HIF-1α), nuclear factor kappa B (NF-κB), and tumor necrosis factor (TNF-α), inducible NO synthase (iNOS), vascular endothelial growth factor (VEGF), and cleaved caspase 3 were shown to be increased in the IH group. Overall, we found that exposure to intermittent hypoxia for 35 days by simulating sleep apnea leads to oxidative stress, inflammation, and increased activity of caspase 3 in the liver and lung.

Blood collection for biochemical analysis in adult zebrafish.

The zebrafish has been used as an animal model for studies of several human diseases. It can serve as a powerful preclinical platform for studies of molecular events and therapeutic strategies as well as for evaluating the physiological mechanisms of some pathologies. There are relatively few publications related to adult zebrafish physiology of organs and systems, which may lead researchers to infer that the basic techniques needed to allow the exploration of zebrafish systems are lacking. Hematologic biochemical values of zebrafish were first reported in 2003 by Murtha and colleagues who employed a blood collection technique first described by Jagadeeswaran and colleagues in 1999. Briefly, blood was collected via a micropipette tip through a lateral incision, approximately 0.3 cm in length, in the region of the dorsal aorta. Because of the minute dimensions involved, this is a high-precision technique requiring a highly skilled practitioner. The same technique was used by the same group in another publication in that same year. In 2010, Eames and colleagues assessed whole blood glucose levels in zebrafish. They gained access to the blood by performing decapitations with scissors and then inserting a heparinized microcapillary collection tube into the pectoral articulation. They mention difficulties with hemolysis that were solved with an appropriate storage temperature based on the work Kilpatrick et al. When attempting to use Jagadeeswaran's technique in our laboratory, we found that it was difficult to make the incision in precisely the right place as not to allow a significant amount of blood to be lost before collection could be started. Recently, Gupta et al. described how to dissect adult zebrafish organs, Kinkle et al. described how to perform intraperitoneal injections, and Pugach et al. described how to perform retro-orbital injections. However, more work is needed to more fully explore basic techniques for research in zebrafish. The small size of zebrafish presents challenges for researchers using it as an experimental model. Furthermore, given this smallness of scale, it is important that simple techniques are developed to enable researchers to explore the advantages of the zebrafish model.