Alexandre Rotta

Comparison of lung protective ventilation strategies in a rabbit model of acute lung injury.

Objective To determine the impact of different protective and nonprotective mechanical ventilation strategies on the degree of pulmonary inflammation, oxidative damage, and hemodynamic stability in a saline lavage model of acute lung injury. Design A prospective, randomized, controlled, in vivo animal laboratory study. Setting Animal research facility of a health sciences university. Subjects Forty-six New Zealand White rabbits. Interventions Mature rabbits were instrumented with a tracheostomy and vascular catheters. Lavage-injured rabbits were randomized to receive conventional ventilation with either a) low peak end-expiratory pressure (PEEP; tidal volume of 10 mL/kg, PEEP of 2 cm H2O); b) high PEEP (tidal volume of 10 mL/kg, PEEP of 10 cm H2O); c) low tidal volume with PEEP above Pflex (open lung strategy, tidal volume of 6 mL/kg, PEEP set 2 cm H2O > Pflex); or d) high-frequency oscillatory ventilation. Animals were ventilated for 4 hrs. Lung lavage fluid and tissue samples were obtained immediately after animals were killed. Lung lavage fluid was assayed for measurements of total protein, elastase activity, tumor necrosis factor-&agr;, and malondialdehyde. Lung tissue homogenates were assayed for measurements of myeloperoxidase activity and malondialdehyde. The need for inotropic support was recorded. Measurements and Main Results Animals that received a lung protective strategy (open lung or high-frequency oscillatory ventilation) exhibited more favorable oxygenation and lung mechanics compared with the low PEEP and high PEEP groups. Animals ventilated by a lung protective strategy also showed attenuation of inflammation (reduced tracheal fluid protein, tracheal fluid elastase, tracheal fluid tumor necrosis factor-&agr;, and pulmonary leukostasis). Animals treated with high-frequency oscillatory ventilation had attenuated oxidative injury to the lung and greater hemodynamic stability compared with the other experimental groups. Conclusions Both lung protective strategies were associated with improved oxygenation, attenuated inflammation, and decreased lung damage. However, in this small-animal model of acute lung injury, an open lung strategy with deliberate hypercapnia was associated with significant hemodynamic instability.

Partial liquid ventilation reduces pulmonary neutrophil accumulation in an experimental model of systemic endotoxemia and acute lung injury

OBJECTIVE To determine whether pulmonary neutrophil sequestration and lung injury are affected by partial liquid ventilation with perfluorocarbon in a model of acute lung injury (ALI). DESIGN A prospective, controlled, in vivo animal laboratory study. SETTING An animal research facility of a health sciences university. SUBJECTS Forty-one New Zealand White rabbits. INTERVENTIONS Mature New Zealand White rabbits were anesthetized and instrumented with a tracheostomy and vascular catheters. Animals were assigned to receive partial liquid ventilation (PLV, n = 15) with perflubron (18 mL/kg via endotracheal tube), conventional mechanical ventilation (CMV, n = 15) or high-frequency oscillatory ventilation (HFOV, n = 5). Animals were ventilated, using an FIO2 of 1.0, and ventilatory settings were required to achieve a normal PaCO2. Animals were then given 0.9 mg/kg of Escherichia coli endotoxin intravenously over 30 mins. Partial liquid ventilation, conventional mechanical ventilation, or high-frequency oscillatory ventilation was continued for an additional 4 hrs before the animals were killed. A group of animals not challenged with endotoxin underwent conventional ventilation for 4.5 hrs, serving as the control group (control, n = 6). Lungs were removed and samples were frozen at -70 degrees C. Representative samples were stained for histology. A visual count of neutrophils per high-power field (hpf) was performed in five randomly selected fields per sample in a blinded fashion by light microscopy. Lung samples were homogenized in triplicate in phosphate buffer, ultrasonified, freeze-thawed, and clarified by centrifugation. Supernatants were analyzed for myeloperoxidase (MPO) activity by spectrophotometry with o-dianisidine dihydrochloride and hydrogen peroxide at 460 nm. MEASUREMENTS AND MAIN RESULTS Histologic analysis of lung tissue obtained from control animals showed normal lung architecture. Specimens from the PLV and HFOV groups showed a marked decrease in alveolar proteinaceous fluid, pulmonary vascular congestion, edema, necrotic cell debris, and gross inflammatory infiltration when compared with the CMV group. Light microscopy of lung samples of animals supported with PLV and HFOV had significantly lower neutrophil counts when compared with CMV (PLV, 4 +/- 0.3 neutrophils/hpf; HFOV, 4 +/- 0.5 neutrophils/hpf; CMV, 10 +/- 0.9 neutrophils/hpf; p < .01). In addition, MPO activity from lung extracts of PLV and HFOV animals was significantly lower than that of CMV animals (PLV, 61 +/- 13.3 units of MPO activity/lung/kg; HFOV, 43.3 +/- 6.8 units of MPO activity/lung/kg; CMV, 140 +/- 28.5 units of MPO activity/lung/kg; p < .01). MPO activity from lungs of uninjured control animals was significantly lower than that of animals in the PLV, HFOV, and CMV groups (control, 2.2 +/- 2 units of MPO activity/lung/kg; p < .001). CONCLUSIONS Partial liquid ventilation decreases pulmonary neutrophil accumulation, as shown by decreased neutrophil counts and MPO activity, in an experimental animal model of ALI induced by systemic endotoxemia. The attenuation in pulmonary leukostasis in animals treated with PLV is equivalent to that obtained by a ventilation strategy that targets lung recruitment, such as HFOV.

PARTIAL LIQUID VENTILATION INFLUENCES PULMONARY HISTOPATHOLOGY IN AN ANIMAL MODEL OF ACUTE LUNG INJURY

PURPOSE The aim of this study was to assess the effect of partial liquid ventilation (PLV) and conventional mechanical ventilation (CMV) in the pattern of distribution of lung injury in a rabbit model of acute lung injury. MATERIALS AND METHODS Animals (1.5 to 3.5 kg) were assigned to receive CMV (tidal volume of 10 mL/kg and a PEEP of 5 cm H2O) or PLV with 18 mL/kg of intratracheal perflubron (tidal volume of 10 mL/kg and a PEEP of 5 cm H2O). Lung injury was elicited by intravenous administration of Escherichia coliendotoxin. Uninjured animals ventilated as the CMV group served as controls. After 4 hours of mechanical ventilation, the lungs were removed and tissue injury was assessed by light microscopy using a scoring system. RESULTS Animals in the CMV group had higher lung injury scores in comparison to the PLV group (10+/-4.5 vs. 5+/-3.3, respectively, P < .05). The injury scores were similar for nondependent lung regions (CMV: 8+/-4.3, PLV: 6+/-2.9) but significantly different for the dependent regions (CMV: 12+/-4.6, PLV: 5+/-3.8, P< .05). CONCLUSIONS PLV is associated with significant attenuation of lung injury, in comparison to CMV. This effect is predominantly due to attenuation of injury in the dependent region of the lung.

Gastric acid and particulate aspiration injury inhibits pulmonary bacterial clearance

ObjectiveTo establish a model of secondary bacterial pneumonia following gastric aspiration and to identify possible mechanisms involved in the suppressed antibacterial defenses following the initial pulmonary insult. DesignA controlled, in vivo laboratory study. SettingResearch laboratory of a health sciences university. SubjectsNinety-five Long-Evans rats. InterventionsAnimals were anesthetized for neck dissection and placement of a 14-gauge catheter in the trachea. Gastric aspirate (1.2 mL/kg of saline, pH 1.25, and 40 mg/mL sterile rat gastric particles) or an equal amount of normal saline (pH 5.3) was instilled intratracheally. One minute after this insult, animals received an intratracheal instillation of either 5.6 × 105 colony-forming units of Escherichia coli or an equal volume of normal saline. The animals remained in room air until kill at 4 hrs or 24 hrs after the intratracheal instillation. The lungs were homogenized for quantitative bacterial cultures. Bronchoalveolar lavage fluid was obtained for cell counts and measurements of albumin, tumor necrosis factor-&agr;, interleukin-1&bgr;, cytokine-induced neutrophil chemoattractant-1, macrophage inflammatory protein-2, monocyte chemoattractant protein-1, and interleukin 10. Measurements and Main ResultsAnimals that received gastric aspirate (followed by normal saline or E. coli) had increased injury as assessed by significant reductions in oxygenation and elevations in bronchoalveolar lavage albumin. At 24 hrs, animals that received gastric aspirate inoculation followed by E. coli had significantly higher pulmonary bacterial counts compared with animals that received E. coli alone. Gastric aspiration injury followed by bacterial inoculation also resulted in acute, but transient, increases in tumor necrosis factor-&agr;, interleukin-1&bgr;, cytokine-induced neutrophil chemoattractant-1, and macrophage inflammatory protein-2 and more sustained elevations of monocyte chemoattractant protein-1 and interleukin-10. ConclusionsLung injury increases and bacterial clearance decreases in this experimental model of E. coli pneumonia following gastric aspiration. Cytokine profiles suggest possible mechanisms for the impaired antibacterial host defense.

Fulminant pertussis: a multi-center study with new insights into the clinico-pathological mechanisms.

Pertussis carries a high risk of mortality in very young infants. The mechanism of refractory cardio‐respiratory failure is complex and not clearly delineated. We aimed to examine the clinico‐pathological features and suggest how they may be related to outcome, by multi‐center review of clinical records and post‐mortem findings of 10 patients with fulminant pertussis (FP). All cases were less than 8 weeks of age, and required ventilation for worsening respiratory symptoms and inotropic support for severe hemodynamic compromise. All died or underwent extra corporeal membrane oxygenation (ECMO) within 1 week. All had increased leukocyte counts (from 54 to 132 × 109/L) with prominent neutrophilia in 9/10. The post‐mortem demonstrated necrotizing bronchitis and bronchiolitis with extensive areas of necrosis of the alveolar epithelium. Hyaline membranes were present in those cases with viral co‐infection. Pulmonary blood vessels were filled with leukocytes without well‐organized thrombi. Immunodepletion of the thymus, spleen, and lymph nodes was a common feature. Other organisms were isolated as follows; 2/10 cases Para influenza type 3, 2/10 Moraxella catarrhalis, 1/10 each with respiratory syncytial virus (RSV), a coliform organism, methicillin‐resistant Staphylococcus aureus (MRSA), Haemophilus influenzae, Stenotrophomonas maltophilia, methicillin‐sensitive Staphylococcus aureus (MSSA), and candida tropicalis. We postulate that severe hypoxemia and intractable cardiac failure may be due to the effects of pertussis toxin, necrotizing bronchiolitis, extensive damage to the alveolar epithelium, tenacious airway secretions, and possibly leukostasis with activation of the immunological cascade, all contributing to increased pulmonary vascular resistance. Cellular apoptosis appeared to underlay much of these changes. The secondary immuno‐compromise may facilitate co‐infection. Pediatr Pulmonol. 2009; 44:970–980. ©2009 Wiley‐Liss, Inc.

Partial liquid ventilation with perflubron attenuates in vivo oxidative damage to proteins and lipids.

Objective: To determine the impact of partial liquid ventilation on the degree of pulmonary damage by reactive oxygen species in a model of acute lung injury caused by systemic endotoxemia. Design: A prospective, controlled, in vivo, animal laboratory study. Setting: Animal research facility of a health sciences university. Subjects: Forty New Zealand White rabbits. Interventions: Mature rabbits were anesthetized and instrumented with a tracheostomy and vascular catheters. Animals were assigned to receive either partial liquid ventilation (n = 16) with perflubron (18 mL/kg via endotracheal tube) or conventional mechanical ventilation (n = 16). Both groups were ventilated using similar strategies, with an FIO2 of 1.0 and tidal volume as required to obtain a normal PaCO2. Animals were then given 0.9 mg/kg Escherichia coli endotoxin intravenously over 30 mins. Eight uninjured instrumented and mechanically ventilated animals served as controls. Partial liquid ventilation or conventional ventilation was continued for 4 hrs before the animals were killed. Lung homogenates were analyzed for malondialdehyde (MDA) and 4‐hydroxy‐2(E)‐nonenal (4‐HNE) concentrations using a colorimetric assay. To assess protein oxidative damage, carbonyl groups in protein side chains were derivatized with 2,4‐dinitrophenylhydrazine followed by Western blotting with a dinitrophenylated‐specific primary antibody. Measurements and Main Results: MDA (713.42 ± 662 vs. 1601.4 ± 1156 nmol/g protein; p = .023) and MDA plus 4‐HNE (1480.24 ± 788 vs. 2675.2 ± 1628 nmol/g protein; p = .038) concentrations were lower in animals treated with partial liquid ventilation compared with conventionally ventilated animals, respectively. Animals treated with partial liquid ventilation exhibited attenuation of dinitrophenylated‐derivatized protein bands by Western blotting, indicating a reduction in protein oxidative damage. The presence of perfluorocarbon did not interfere with the MDA assay when assessed by independent analysis in vitro. Perflubron did not serve as a sink for peroxyl radicals produced in the aqueous phase during separate in vitro oxidation experiments. Conclusions: Partial liquid ventilation attenuates oxidative damage to lipids and proteins during experimental acute lung injury. This finding is not caused by binding of lipid peroxidation products to perflubron or by the peroxyl radical scavenging properties of perflubron.

Liquid ventilation attenuates pulmonary oxidative damage.

PURPOSE Liquid perfluorochemicals reduce the production of reaction oxygen species by alveolar macrophages. We sought to determine whether the use of liquid perfluorochemicals in vivo during liquid ventilation would attenuate oxidative damage to the lung. MATERIALS AND METHODS Healthy infant piglets (n = 16) were instrumented for mechanical ventilation and received intravenous oleic acid to create an acute lung injury. The animals were assigned to a nontreatment group receiving conventional mechanical ventilation or a treatment group receiving partial liquid ventilation with a liquid perfluorochemical. Following sacrifice, the bronchoalveolar lavage and lung parenchyma were analyzed for evidence of oxidative damage to lipids and proteins by determination of TBARS and carbonylated protein residues, respectively. RESULTS Mortality in the control group was 50% at the completion of the study compared with no deaths in the partial liquid ventilation group (P = .025). The alveolar-arterial oxygen difference was more favorable following injury in the partial liquid ventilation group. The liquid ventilation group demonstrated a 32% reduction in TBARS (P = .043) and a 14% reduction in carbonylated protein residues (P = .061). CONCLUSION These data suggest that partial liquid ventilation supports gas exchange and reduces mortality in association with a reduction in the production of reactive oxygen species and the concomitant attenuation of tissue damage during the early phase of acute lung injury.

Strict Glycemic Targets Need Not Be So Strict: A More Permissive Glycemic Range for Critically Ill Children

OBJECTIVE. The goal was to determine whether a more permissive glycemic target would be associated with a decreased incidence of hypoglycemia but not increased mortality rates in critically ill pediatric patients. METHODS. This retrospective study evaluated clinical and laboratory data for 177 patients who underwent 211 consecutive surgical procedures for repair or palliation of congenital heart defects at Driscoll Childrens Hospital. To establish the relationship between postoperative glycemia and subsequent morbidity and mortality rates, patients were stratified into 4 groups according to their median glucose levels, that is, euglycemia (60–125 mg/dL, 3.3–6.9 mmol/L), mild hyperglycemia (126–139 mg/dL, 6.9–7.7 mmol/L), moderate hyperglycemia (140–179 mg/dL, 7.7–9.9 mmol/L), or severe hyperglycemia (≥180 mg/dL, ≥9.9 mmol/L). Postoperative outcomes for those groups also were compared with outcomes for a more permissive glycemic target group (90–140 mg/dL, 5–7.7 mmol/L). RESULTS. The peak and mean blood glucose measurements and duration of hyperglycemia were not different for survivors and nonsurvivors in the first 24 hours after surgery. Nonsurvivors had higher peak glucose levels (389.3 ± 162 mg/dL vs 274.4 ± 106.3 mg/dL, 21.4 ± 8.9 mmol/L vs 15.1 ± 5.9 mmol/L) and longer duration of hyperglycemia (3.06 ± 1.67 days vs 2.11 ± 0.92 days) during the first 5 postoperative days, compared with survivors. Mortality rates were significantly higher for the moderate (38.8%) and severe (58.3%) hyperglycemia groups, compared with the euglycemia (6.02%) and permissive target (4.69%) groups. The incidence of hypoglycemia was significantly higher in the euglycemia group (31.8%), compared with the permissive target group (17.18%). CONCLUSIONS. Postoperative hyperglycemia is associated with increased morbidity and mortality rates in children after surgical repair of congenital heart defects. A more permissive glycemic target is associated with a lower incidence of hypoglycemia but not increased mortality rates in these patients.

O manejo da síndrome do desconforto respiratório agudo

OBJETIVO: Revisar as atuais estrategias de suporte e de tratamento da sindrome do desconforto respiratorio agudo (SDRA). FONTE DOS DADOS: Dados proprios de nosso laboratorio de pesquisa e bibliografia relacionada as areas de SDRA e lesao pulmonar aguda, pesquisados atraves do Medline. SINTESE DOS DADOS: Apesar de avancos no entendimento da sua patogenese, a SDRA ainda resulta em significativa morbidade e mortalidade. A ventilacao mecânica e a principal modalidade terapeutica na SDRA, sendo atualmente considerada nao mais apenas uma medida de suporte, mas sim uma terapia capaz de alterar o curso da patologia. Novas estrategias ventilatorias, como a ventilacao oscilatoria de alta frequencia (VOAF), tem-se mostrado promissoras. Neste texto, revisamos o conhecimento atual no manejo da SDRA, incluindo ventilacao mecânica convencional e nao convencional, uso de surfactante, oxido nitrico, moduladores do processo inflamatorio, oxigenacao extracorporea e posicao prona. CONCLUSOES: A ultima decada foi marcada por avancos significativos, como o conceito de ventilacao mecânica protetora na SDRA. O beneficio da aplicacao de estrategias alternativas, como a VOAF, assim como do uso do surfactante exogeno e moduladores de inflamacao continuam sendo alvo de estudo.

Perfluorooctyl bromide (perflubron) attenuates oxidative injury to biological and nonbiological systems.

Objective To examine whether perfluorooctyl bromide (perflubron) is capable of protecting biological and nonbiological systems against oxidative damage through a mechanism independent of its known anti-inflammatory property. Design A controlled, in vitro laboratory study. Setting Research laboratory of a health sciences university. Subjects Rat pulmonary artery endothelial cell cultures (biological system) and linoleic acid in sodium dodecyl sulfate micelles (nonbiological system). Interventions Rat pulmonary artery endothelial cells labeled with dichlorofluorescein diacetate and incubated with perflubron or culture media (control) were exposed to H2O2. H2O2-induced fluorescence of dichlorofluorescein diacetate was measured as an index of intracellular oxidative stress. In another experiment, linoleic acid in sodium dodecyl sulfate micelles was exposed to various concentrations of the azo initiator 2,2′-diazo-bis-(2-amidinopropane) dihydrochloride (2, 4, 20, and 50 mM) in the presence or absence of perflubron. Malondialdehyde measurements were obtained as a marker of oxidative damage to linoleic acid. Measurements and Main Results Cell monolayers incubated with perflubron exhibited 66.6% attenuation in intracellular fluorescence compared with controls (p < .05). Linoleic acid in sodium dodecyl sulfate micelles incubated with perflubron and exposed to 2, 4, 20, or 50 mM of 2,2′-diazo-bis-(2-amidinopropane) dihydrochloride showed less evidence of lipid peroxidation as indicated by lower malondialdehyde measurements at 240 mins (10.6%, 16%, 41%, and 14.2%, respectively) compared with controls. Conclusions Perflubron attenuates oxidative damage to both biological and nonbiological systems. This newly recognized property of perflubron is independent of its anti-inflammatory properties.