Thiol-mediated regulation of ICAM-1 expression in endotoxin-induced acute lung injury

The intracellular redox state regulates several aspects of cell function, suggesting that strategies directed toward altering the cellular redox state may modulate cell activation in inflammatory states. As the most abundant intracellular thiol, glutathione plays a critical role as an intracellular redox buffer. Using diethylmaleate (DEM) as a glutathione-depleting agent, we evaluated the effects of GSH depletion in a rodent model of polymorphonuclear neutrophil (PMN)-dependent acute lung injury. Rats received 500 microg of LPS by intratracheal challenge, inducing a 5.5-fold increase in lung permeability and sixfold increase in lung PMN content. Pretreatment with DEM prevented the LPS-induced increase in lung PMN influx and lung permeability. Northern analysis and immunohistochemical studies suggest that this effect may be mediated by preventing up-regulation of lung ICAM-1 mRNA and protein expression. This effect is specific to ICAM-1, because lung cytokine-induced neutrophil chemoattractant and TNF-alpha mRNA levels are unaffected. This finding is not unique to the lung, because a similar effect on PMN influx was recapitulated in a rodent model of chemical peritonitis. Further, in vitro studies demonstrated that pretreatment of HUVEC monolayers with DEM prevented both ICAM-1 up-regulation and PMN transendothelial migration. These data indicate the presence of a thiol-sensitive mechanism for modulating ICAM-1 gene expression and suggest a potential novel therapeutic strategy for diseases characterized by PMN-mediated tissue injury.     

Role of inducible nitric oxide synthase in endotoxin-induced acute lung injury

The role of nitric oxide (NO) in lung injury remains unclear. Both beneficial and detrimental roles have been proposed. In this study, we used mutant mice lacking the inducible nitric oxide synthase (iNOS) to assess the role of this isoform in sepsis-associated lung injury. Wild-type and iNOS knockout mice were injected with either saline or Escherichia coli endotoxin (LPS) 25 mg/kg and killed 6, 12, and 24 h later. Lung injury was evaluated by measuring lactate dehydrogenase activity in the bronchoalveolar lavage, pulmonary wet/dry ratio, and immunostaining for nitrotyrosine formation. In the wild-type mice, LPS injection elicited more than a 3-fold rise in lactate dehydrogenase activity, a significant rise in lung wet/dry ratio and extensive nitrotyrosine staining in large airway and alveolar epithelium, macrophages, and pulmonary vascular cells. This was accompanied by induction of iNOS protein and increased lung nitric oxide synthase activity. By comparison, LPS injection in iNOS knockout mice elicited no iNOS induction and no significant changes in lung NOS activity, lactate dehydrogenase activity, lung wet/dry ratio, or pulmonary nitrotyrosine staining. These results indicate that mice deficient in iNOS gene are more resistant to LPS-induced acute lung injury than are wild-type mice.     

Exogenous surfactant and positive end-expiratory pressure in the treatment of endotoxin-induced lung injury

OBJECTIVE: To evaluate the efficacy of treating endotoxin-induced lung injury with single dose exogenous surfactant and positive end-expiratory pressure (PEEP). DESIGN: Prospective trial. SETTING: Laboratory at a university medical center. SUBJECTS: Nineteen certified healthy pigs, weighing 15 to 20 kg. INTERVENTIONS: Pigs were anesthetized and surgically prepared for hemodynamic and lung function measurements. Animals were randomized into four groups: a) Control pigs (n = 4) received an intravenous infusion of saline without Escherichia colilipopolysaccharide (LPS); b) the LPS group (n = 5) received an intravenous infusion of saline containing LPS (100 microg/kg); c) the PEEP plus saline group (n = 5) received an intravenous infusion of saline containing LPS. Two hours after LPS infusion, saline was instilled into the lung as a control for surfactant instillation, and the animals were placed on 7.5 cm H2O of PEEP; d) the PEEP plus surfactant group (n = 5) received an intravenous infusion of saline containing LPS. Two hours following LPS infusion, surfactant (50 mg/kg) was instilled into the lung and the animals were placed on 7.5 cm H2O of PEEP. PEEP was applied first and surfactant or saline was instilled into the lung while maintaining positive pressure ventilation. All groups were studied for 6 hrs after the start of LPS injection. At necropsy, bronchoalveolar lavage was performed and the right middle lung lobe was fixed for histologic analysis. MEASUREMENTS AND MAIN RESULTS: Compared with LPS without treatment, PEEP plus surfactant significantly increased PaO2 (PEEP plus surfactant = 156.6 +/- 18.6 [SEM] torr [20.8 +/- 2.5 kPa]; LPS = 79.2 +/- 21.9 torr [10.5 +/- 2.9 kPa]; p<.05), and decreased venous admixture (PEEP plus surfactant = 12.5 +/- 2.0%; LPS = 46.9 +/- 14.2%; p< .05) 5 hrs after LPS infusion. These changes were not significant 6 hrs after LPS infusion. PEEP plus surfactant did not alter ventilatory efficiency index (VEI = 3800/[peak airway pressure - PEEP] x respiratory rate x PacO2), or static compliance as compared with LPS without treatment at any time point. Cytologic analysis of bronchoalveolar lavage fluid showed that surfactant treatment significantly increased the percentage of alveolar neutrophils as compared with LPS without treatment (PEEP plus surfactant = 39.1 +/- 5.5%; LPS = 17.4 +/- 6.6%; p< .05). Histologic analysis showed that LPS caused edema accumulation around the airways and pulmonary vessels, and a significant increase in the number of sequestered leukocytes (LPS group = 3.4 +/- 0.2 cells/6400 micro2; control group = 1.3 +/- 0.1 cells/6400 micro2; p < .05). PEEP plus saline and PEEP plus surfactant significantly increased the total number of sequestered leukocytes in the pulmonary parenchyma (PEEP plus surfactant = 8.2 +/- 0.7 cells/6400 micro2; PEEP plus saline = 3.9 +/- 0.2 cells/6400 micro2; p <.05) compared with the control and LPS groups. CONCLUSIONS: We conclude that PEEP plus surfactant treatment of endotoxin-induced lung injury transiently improves oxygenation, but is unable to maintain this salutary effect indefinitely. Thus, repeat bolus dosing of surfactant or bolus treatment followed by continuous aerosol delivery may be necessary for a continuous beneficial effect.     

Recombinant human soluble thrombomodulin reduces endotoxin-induced pulmonary vascular injury via protein C activation in rats

Adult respiratory distress syndrome (ARDS) is a serious complication of disseminated intravascular coagulation (DIC) or multiple organ failure. To determine whether recombinant soluble human thrombomodulin (rsTM) may be useful in treating ARDS due to sepsis, we investigated the effect of rsTM on lipopolysaccharide (LPS)-induced pulmonary vascular injury in rats. The intravenous administration of rsTM prevented the increase in pulmonary vascular permeability induced by LPS. Neither heparin plus antithrombin III (AT III) nor dansyl Glu Gly Arg chloromethyl ketone-treated factor Xa (DEGR-Xa), a selective inhibitor of thrombin generation, prevented LPS-induced vascular injury. The agents rsTM, heparin plus AT III, and DEGR-Xa all significantly inhibited the LPS-induced intravascular coagulation. Recombinant soluble TM pretreated with a monoclonal antibody (moAb) that inhibits protein C activation by rsTM did not prevent the LPS-induced vascular injury; in contrast, rsTM pretreated with a moAb that does not affect thrombin binding or protein C activation by rsTM prevented vascular injury. Administration of activated protein C (APC) also prevented vascular injury. LPS-induced pulmonary vascular injury was significantly reduced in rats with leukopenia induced by nitrogen mustard and by ONO-5046, a potent inhibitor of granulocyte elastase. Results suggest that rsTM prevents LPS-induced pulmonary vascular injury via protein C activation and that the APC-induced prevention of vascular injury is independent of its anticoagulant activity, but dependent on its ability to inhibit leukocyte activation.     

Ascorbic acid deficiency reduces the level of mRNA for cytochrome P-450 on the induction by polychlorinated biphenyls

Ascorbic acid (AsA) deficiency causes a decrease in hepatic concentration of cytochrome P-450 and a decrease in hepatic activity of drug-metabolizing enzymes in rats unable to synthesize AsA (ODS rats). To study the mechanism of the decrease in hepatic concentration of cytochrome P-450 isozymes by AsA deficiency, we chose the xenobiotics-inducible cytochrome P-450 and performed the experiments indicated below. AsA-deficient rats were fed polychlorinated biphenyls (PCB) which markedly induce both CYP1A subfamily and several isozymes in CYP2B subfamily. First, we assayed the activities of two drug-metabolizing enzymes so that one could be functionally distinguished from another. AsA deficiency significantly reduced the hepatic activity of aminopyrine-N-demethylase in ODS rats with and without dietary PCB, but had no effect on benzo(a)pyrene hydroxylase activity. Secondly, quantitative immunoblot analyses demonstrated that the levels of CYP2B1/2B2 and CYP1A1 in the AsA-deficiency rats fed PCB were approximately 60 and 80% lower than those found in rats fed AsA-supplemented diet. The degree of reduction in CYP2B1/2B2 was greater than CYP1A1. Thirdly, AsA deficiency caused a decrease in hepatic abundance of CYP2B1/2B2 mRNA, whereas it had no effect on the levels of CYP1A1 and 1A2 mRNA. These results indicated that dietary AsA selectively affects the levels of CYP2B1/2B2 mRNA among cytochrome P-450 induced by PCB and plays important roles for optimum induction of drug-inducible cytochrome P-450. We concluded that AsA deficiency decreases specific froms of drug-inducible cytochrome P-450, especially CYP2B1/2B2 and that the reduction of CYP2B1/2B2 mRNA level in AsA-deficient rats caused a decrease in cytochrome P-450 concentration and hepatic activity of drug-metabolizing enzymes.     

A possible involvement of oxidative lung injury in endotoxin-induced bronchial hyperresponsiveness to substance P in guinea pigs

Fragile-X syndrome and myotonic dystrophy are caused by triplet repeat expansions embedded in CpG islands in the transcribed non-coding regions of the FMR1 and the DMPK genes, respectively. Although initial reports emphasized differences in the mechanisms by which the expanded triplet repeats caused these diseases, results published in the past year highlight remarkable parallels in the likely molecular etiologies. At both loci, expansion is associated with altered chromatin, aberrant methylation, and suppressed expression of the adjacent FMR1 and DMAHP genes, implicating epigenetic mediation of these genetic diseases.     

Ascorbic acid stimulates chloride transport in the amphibian cornea

The cornea of the toad, Bufo marinus, actively transports chloride from the endothelial to the epithelial surface. This transport process has been related to the maintenance of the normal transparency of the cornea. Ion transport, as evidenced by the short-circuit current (SCC), is markedly stimulated by physiologic concentrations of ascorbic acid. Measurement of the unidirectional fluxes of 36Cl and 22Na shows that the increase in SCC is due primarily to a stimulation of the active transport of chloride.     

Donor treatment with the lazeroid U74389G reduces ischemia-reperfusion injury in a rat lung transplant model

BACKGROUND: Antioxidant treatment with lazeroids has proven beneficial for the amelioration of reperfusion injury in experimental lung transplantation. This study compares the effect of donor versus recipient treatment on immediate postoperative graft function. METHODS: A model of acute double-lung transplantation in rats was used to assess graft function. Transplanted controls after 2 (group I) and 16 hours of ischemia (group II) were compared to a recipient (group III; 16-hour ischemia) and a donor treatment group (group IV; 16-hour ischemia) using the lazeroid U74389G (6 mg/kg). Serial assessment of alveolar-arterial oxygen difference, dynamic lung compliance, airway and pulmonary vascular resistance was obtained during a 2-hour reperfusion period. Final analysis included survival, weight gain, and histologic examination. RESULTS: Graft function was significantly better after 2 hours of ischemia than in any of the three 16-hour ischemia groups (II, III, IV). After 16 hours of ischemia, donor treatment provided superior graft function with respect to dynamic lung compliance, airway resistance, and alveolar-arterial oxygen difference when compared with groups II and III. The pulmonary vascular resistance was significantly higher in group III when compared with groups II and IV. Graft weight increase reflecting edema was highest in groups III (104%) and II (98%). CONCLUSIONS: After prolonged ischemia only donor treatment with the lazeroid U74389G was able to significantly reduce ischemia-reperfusion-related graft dysfunction.     

Autonomic control of bronchial circulation in awake sheep during rest and behaviour

1. We tested the hypothesis that the pattern and the intensity of autonomic mechanisms causing vasoconstriction in the resting bronchial circulation of awake dogs also exists in awake sheep. It was also postulated that sighing behaviour and the associated bronchovascular dilatation induced by non-adrenergic, non-cholinergic (NANC) mechanisms observed in the dog exist in sheep. 2. Bronchial arterial blood flow to lower airways of both lungs of awake sheep was measured continuously using pulsed Doppler flow probes mounted on the bronchial artery at prior thoracotomy. 3. Cumulative and factorial analysis of responses to randomized combinations of autonomic alpha 1-, alpha 2-, beta 1- and beta 2-adrenoceptors and cholinoceptor autonomic blockade suggests that resting vasoconstrictor activity is less in sheep than in dogs. At normal aortic pressure, the autonomic activity of these receptor groups in the sheep lowers bronchial blood flow and conductance by 30%, whereas in the awake dog, the corresponding autonomic effect is 50%. 4. Tonic autonomic control of bronchial conductance can be partitioned in sheep to show significant and separate alpha- and beta-adrenoceptor vasoconstrictor activity at a ratio of 1.8:1, an effect normally offset by a weaker vasodilator alpha-/beta-adrenoceptor interaction. In contrast to the situation in awake dogs, cholinoceptors do not play a role in awake sheep. 5. Nitric oxide (NO) synthase inhibition in sheep using NG-nitro-L-arginine following blockade of alpha- and beta-adrenoceptors and cholinoceptors causes hypertension, but minor changes, if any, in pulmonary pressures or heart rate. Bronchial flow and conductance, however, fall from a higher resting conductance by approximately 50%, suggesting that, normally, resting bronchial flow conductance is dominated by strong tonic NO vasodilator effects that interact with weaker tonic autonomic vasoconstrictor effects. 6. Superimposed (respiratory) behaviours of sighing, sneezing and coughing, which involve negative swings in intrathoracic pressure and the movement of inspired air, evoke large active bronchovascular dilator effects. These appear to be largely NANC in origin and appear to be dependent, in part, on mechanisms associated with NO release. It is postulated that the C-fibre axon reflex using substance P, calcitonin gene-related peptide and neurokinin A may be involved. Vocalization and eructation do not evoke bronchovascular effects.     

Extreme hypoventilation reduces ventilator-induced lung injury during ventilation with low positive end-expiratory pressure in saline-lavaged rabbits

OBJECTIVE: To compare the degrees of ventilator-induced lung injury caused by two ventilation protocols. DESIGN: Randomized trial. SETTING: University animal laboratory. SUBJECTS: Sixteen New Zealand white rabbits. INTERVENTIONS: After five sequential saline lung lavages, eight pairs of anesthetized rabbits were allocated randomly to receive either of two ventilation protocols for 4 hrs during neuromuscular blockade. Both groups received 3 cm H2O of positive end-expiratory pressure and 100% oxygen. Control group animals received an estimated tidal volume of 12 mL/kg, an inspiratory time of 0.7 sec, and a ventilatory rate adjusted for a PaCO2 of 35 to 45 torr (4.7 to 6 kPa). Study group animals were ventilated through an intratracheal catheter, with inspiratory time of 1.5 secs, ventilatory rate of 20 breaths/min, and peak inspiratory pressure of 4 to 8 cm H2O, adjusted to maintain PaCO2 at 150 to 250 torr (20 to 33 kPa). MEASUREMENTS AND MAIN RESULTS: Arterial blood gases were measured every 30 mins. After 4 hrs, a final lung lavage was performed. Physiologic parameters, cell counts and protein concentration in the final lavage, and lung histology were compared between groups. The alveolar-arterial oxygen tension gradient was higher in the study group over the first 1.5 hrs, but the time profile showed significantly (p = .001) greater improvement in the study group. After 4 hrs, the mean alveolar-arterial oxygen tension gradient was lower in the study group (94 torr [12.5 kPa] vs. 201 torr [26.8 kPa]). The increase in neutrophil count from the initial to the final lung lavage was lower in the study group (0.27 x 10(7) cells/L vs. 2.01 x 10(7) cells/L, p = .037), as was the absolute value of the neutrophil count in the final lavage (1.33 x 10(7) cells/L vs. 3.02 x 10(7) cells/L, p = .04). The median hyaline membrane score was lower in the study group (0.5 vs. 3.0) but the difference was not statistically significant. CONCLUSION: These findings suggest that a very low tidal volume reduces ventilator-induced lung injury in saline-lavaged rabbits during ventilation at low lung volume.     

Pulmonary vasoconstriction in oleic acid induced lung injury. A morphometric study

Distribution and severity of active vasoconstriction of muscular pulmonary arteries were morphometrically assessed in anaesthetized, paralysed and mechanically ventilated pigs with respiratory distress, induced by oleic acid. Vasoconstriction was deduced from the medial thickness which was measured and expressed as a percentage of external diameter. Six pigs received oleic acid (0.12 +/- 0.07 ml/kg), dissolved 1:1 in 96% alcohol, in multiple injections of 0.1 ml. Six pigs were used as controls. After the oleic acid injections a stable hypoxaemia (PaO2 = 57 +/- 8 mmHg, at an inspiratory oxygen fraction of 0.6) and pulmonary hypertension (mean Ppa = 36 +/- 2 mmHg) were obtained for several hours. Electron microscopy revealed swelling of endothelial cells with signs of degeneration. Medial thickness was far greater in the oleic acid group than in the control group; overall mean values were 8.1 +/- 3.2 and 3.8 +/- 1.7% respectively (P < 0.001). Arteries with prominent vasoconstriction were lying in clusters. This pattern was the same in dependent and non-dependent regions. We concluded that in oleic acid induced respiratory distress active vasoconstriction of muscular pulmonary arteries is an important factor in the development of pulmonary hypertension. Besides vasoconstriction, endothelial swelling and intravascular clotting may contribute to the development of pulmonary hypertension.     

Leukocyte adhesion molecules in the liver and plasma cytokine levels in endotoxin-induced rat liver injury

BACKGROUND: The interactions between polymorphonuclear neutrophils (PMNs) and sinusoidal endothelial cells (SECs) have been known to be involved in the pathogenesis of acute liver injury. It has been also reported that tumor necrosis factor-alpha (TNF-alpha) up-regulates ICAM-1 expression on SECs and that interleukin-8 (IL-8) provokes rapid activation of CD11/CD18 on PMNs. These findings expand into the relationship between the expression of leukocyte adhesion molecules (ICAM-1, CD11a/CD18 and CD11b/CD18) in liver tissues and plasma TNF and IL-8 levels after lipopolysaccharide (LPS)-induced liver injury in rats. METHODS: Male Wistar rats weighing 200-250 g were treated with 2 mg LPS/kg intravenously in a 0.2- to 0.25-ml volume. Liver and blood samples were obtained at 1, 3, 8, and 12 h after LPS exposure. Plasma TNF and IL-8 levels were measured using bioassay and specific enzyme-linked immunosorbent assay, respectively. Liver samples were fixed and studied by immunohistochemistry using specific monoclonal antibodies against ICAM-1, CD11a, and CD11b. RESULTS: The TNF level showed a peak at 1 h (23.3 +/- 11.4 IU/ml), and the IL-8 level showed a peak at 3 h (343.1 +/- 110.5 ng/ml) after LPS exposure. An increase in the number of PMNs in the liver was observed as early as 1 h and continued until 12 h after LPS exposure. PMNs adhered to degenerated SECs and hepatocytes. ICAM-1 on SECs was diffusely and strongly expressed at 8 h, and PMNs adhered to SECs expressed both CD11a and CD11b. ICAM-1 was also observed on hepatocytes. CONCLUSION: These data suggest that PMN-SEC and PMN-hepatocyte interactions via leukocyte adhesion molecules, related to inflammatory cytokines such as TNF and IL-8, exist and play an important role in the pathogenesis of acute liver injury.     

Release of lactate by the lung in acute lung injury

The pathogenesis of hyperlactatemia during sepsis is poorly understood. We have previously described an increase in lactate concentration across the lung in the dog during early endotoxemia. Accordingly, we sought to determine if the lung releases lactate in humans and what relation this has with lung injury. METHODS: We measured lactate concentrations across the lung and lung injury scores (LIS) in two groups of patients. Group 1 consisted of nine patients with acute lung injury (LIS > or = 2.0) and elevated lactate concentrations (> 2.0 mmol/L). Group 2 contained 12 patients with no acute lung injury (LIS scores < or = 1.5), with or without increased lactate concentrations. Simultaneous measurements of plasma lactate and blood gases were obtained from indwelling arterial and pulmonary artery catheters. Measurements of cardiac output were also obtained. Lactate measurements were done using a lactate analyzer (YSI; Yellow Springs, Ohio). RESULTS: For each patient with acute lung injury and hyperlactatemia, an arterial-venous lactate gradient existed demonstrating release of lactate by the lung. This gradient persisted after correction for changes in hemoconcentration across the lung. The lactate gradient across the lung was 0.4 +/- 0.2 mmol/L for group 1 vs 0.05 +/- 0.1 mmol/L for group 2 (p = 0.001). This corresponded to a mean pulmonary lactate flux of 231.3 +/- 211.3 vs 5.0 +/- 37.2 mmol/h (p = 0.001). The lactate flux and the arterial-venous lactate difference correlated with LIS both for the entire sample and for the subgroup with hyperlactatemia (r = 0.69, p < 0.01). Pulmonary lactate flux was not related to arterial lactate levels (r = 0.25). CONCLUSION: In patients with acute lung injury and hyperlactatemia, the lung is a major source of lactate and lactate flux correlates with LIS. This lactate flux could explain some of the hyperlactatemia seen in sepsis.     

Ventilatory responses in awake guinea pigs exposed to acid aerosols

This study reports experiments designed to evaluate the dose and temporal effects of an atmospheric pollutant, sulfuric acid (H2SO4) aerosol, on the dynamic components of the respiratory cycle. Ventilation was measured in a whole-body barometric plethysmograph in unanesthetized, unrestrained animals following a 4-h exposure to H2SO4 aerosol at 14.1, 20.1, or 43.3 mg/m3. Lung injury was assessed by histopathology and bronchoalveolar lavage (BAL). Aerosol exposure with H2SO4 caused marked alterations in both the magnitude and composition of the ventilatory response, which were both dose and time dependent. At the highest concentration tested, there was a significant increase in tidal volume (deltaVt) and a decrease in breathing frequency (f) immediately after exposure. Analysis of BAL fluid at this time showed increased inflammatory cells and protein in the acid exposed animals, and histology showed hyaline membranes and acute inflammatory cells in the proximal acinar region. By 24 h postexposure, f significantly increased whereas deltaVt decreased. This pattern of breathing was interspersed with short periods of apnea. The onset of rapid, shallow breathing was associated with histological evidence of diffuse pulmonary edema. By contrast, the immediate postexposure period at the lowest concentration of H2SO4 aerosol was characterized by a significant increase in f and little or no effect on deltaVt. These effects diminished with time, and at 24 h postexposure ventilatory parameters were indistinguishable from baseline values. An apparent crossover between the effects associated with the high and low exposure concentrations was seen at the intermediate exposure concentration; however, closer inspection of these findings on an animal-by-animal basis revealed two populations of animals with respiratory characteristics of either the high-exposure or low-exposure groups. The data suggest that the guinea pig exhibits complex interactions between dose and time to response that are consistent with the activation of neural reflexes. The indirect plethysmographic method provides a simple means to assess these responses in a model system that avoids the use of anesthetics, surgery, and restraint.     

Effects of pretreatment with SDZ MRL 953, a novel immunostimulatory lipid A analog, on endotoxin-induced acute lung injury in guinea pigs

SDZ MRL 953 (SDZ), a novel immunostimulatory lipid A analog, has been reported to have immunopharmacological activities similar to those of lipopolysaccharide (LPS) but to have little of the toxicity of LPS. We investigated the effects of pretreatment with SDZ on Escherichia coli endotoxin-induced acute lung injury in guinea pigs. Four experimental groups consisted of saline control (n = 16), SDZ (-12 h) plus LPS (2 mg/kg of SDZ per kg of body weight injected intravenously 12 h before intravenous injection of 2 mg of LPS per kg; n = 15), SDZ (-10 min) plus LPS (SDZ injected 10 min before LPS injection; n = 10), and LPS alone (n = 16). The animals were sacrificed, and lung tissue was sampled 4 h after LPS or saline infusion. Lung injury was assessed by measuring the wet weight-to-dry weight ratio and the level of 125I-labeled albumin accumulation in bronchoalveolar lavage fluid relative to that in plasma. In the SDZ (-12 h) plus LPS group, these two parameters of acute lung injury were decreased compared with those in the LPS alone group. However, they were not decreased in the SDZ (-10 min) plus LPS group. We conclude that SDZ attenuates endotoxin-induced acute lung injury when it is administered 12 h before LPS injection. The attenuating effects of SDZ are speculated to be due to down regulation of the response to endotoxin rather than to receptor blocking.     

Aprotinin reduces injury of the spinal cord in transient ischemia

OBJECTIVE: The protective effect of aprotinin, which is a protease inhibitor, was assessed in a rabbit spinal cord ischemia model. DESIGN: Randomized, controlled, prospective study. SETTING: University research laboratory. SUBJECTS: New Zealand white rabbits (36) of both sexes. METHODS: In 24 animals, ischemia was induced with midline laparotomy and clamping the aorta just distal to left renal artery and proximal to aortic bifurcation for 20 min. Aprotinin was given 30000 KIU as a short intravenous injection after anesthesia, and was followed by 10000 KIU/h by continuous infusion in group 1 (n = 12). Similar volume of saline solution was used in control group of animals (group 2, n = 12). Group 3 of animals (sham group, n = 12) were anesthetized and subjected to laparotomy without aortic occlusion. Physiological parameters and somatosensory evoked-potentials (SEP) were monitored in animals before ischemia, during ischemia and in the first 60 min of reperfusion. Their neurological outcome was clinically evaluated up to 48 h postischemia. Their motor function was scored, and the intergroup differences were compared. The animals were sacrificed after two days of postischemia. Their spinal cord, abdominal aorta, and its branches were processed for histopathological examination. RESULTS: In group 3, SEP amplitudes did not change during the procedures, and all animals recovered without neurologic deficits. At the end of ischemic period, the average amplitude was reduced to 53+/-7% of the baseline in all ischemic animals. This was followed by a gradual return to 89+/-8 and 81+/-13% of the initial amplitude after 60 min of reperfusion in group 1 and group 2 correspondingly (P > 0.05). The average motor function score was significantly higher in group 1 than group 2 at 24 and 48 h after the ischemic insult (P < 0.05). Histological observations were clearly correlated with the neurological findings. CONCLUSION: The results suggest that aprotinin reduces spinal cord injury and preserves neurologic function in transient spinal cord ischemia in rabbits.     

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.     

Effect of lung volume on lung resistance and elastance in awake subjects measured during sinusoidal forcing

BACKGROUND: Although lung volume may be changed by certain procedures during anesthesia and mechanical ventilation, dependence of the dynamic mechanical properties of the lungs on lung volume are not clear. Based on studies in dogs, the authors hypothesized that changes in lung mechanics caused by anesthesia in healthy humans could be accounted for by immediate changes in lung volume and that lung resistance will not be decreased by positive end-expiratory airway pressure if tidal volume and respiratory frequency are in the normal ranges. METHODS: Lung resistance and dynamic lung elastance were measured in six healthy, relaxed, seated subjects during sinusoidal volume oscillations at the mouth (5 mL/kg; 0.4 Hz) delivered at mean airway pressure from -9 to +25 cmH2O. Changes in lung volume from functional residual capacity were measured with inductance plethysmographic belts. RESULTS: Decreases in mean mean airway pressure that caused decreases in lung volume from functional residual capacity comparable to those typically observed during anesthesia were associated with significant increases in both dynamic lung elastance and lung resistance. Increases in mean mean airway pressure that caused increases in lung volume from functional residual capacity did not increase lung resistance and increased dynamic lung elastance only above about 15 cmH2O. CONCLUSIONS: Increases in dynamic lung elastance and lung resistance with anesthesia can be explained by the accompanying, acute decreases in lung volume, although other factors may be involved. Increasing lung volume by increasing mean airway pressure with positive end-expiratory pressure will decrease lung resistance only if the original lung volume is low compared to awake, seated functional residual capacity.     

Effects of a synthetic lung surfactant on pharyngeal patency in awake human subjects

BACKGROUND: This study presents the use of physiological principles and assessment techniques in addressing four objectives that can enhance a swimmer's likelihood of successfully swimming the English Channel. The four objective were: (1) to prescribe training intensities and determine ideal swimming pace; (2) to determine the amount of insulation needed, relative to heat produced, to diminish the likelihood of the swimmer suffering from hypothermia; (3) to calculate the caloric expenditure for the swim and the necessary glucose replacement required to prevent glycogen depletion; and (4) to determine the rate of acclimatization to cold water (15.56 C/60 F). METHODS: The subject participated in several pool swimming data collection sessions including a tethered swim incremental protocol to determine peak oxygen consumption and onset of lactate accumulation and several steady state swims to determine ideal swimming pace at 4.0 mM/L of lactate. Additionally, these swims provided information on oxygen consumption, which in combination with ultrasound assessment of subcutaneous fat was used to assess heat production and insulation capabilities. Finally, the subject participated in 18 cold water immersions to document acclimatization rate. RESULTS: The data demonstrated the high fitness level of this subject and indicated that at a stroke rate of 63 stokes/min, HR was 130 heats/min and lactate was 4 mM/L. At this swimming pace the swimmer would need to consume 470 kcal of glucose/hr. In addition, the energy produced at this swim pace was 13.25 kcal/min while the energy lost at the present subcutaneous fat quantity was 13.40 kcal/min, requiring a fat weight gain of 6,363.03 g (13.88 lbs) to resist heat loss. CONCLUSIONS: Finally, the data from the cold water immersions suggested that acclimatization occurred following two weeks of immersions. There results were provided to the swimmer and utilized in making decisions in preparation for the swim.     

Ascorbic acid and reducing agents regulate the fates and functions of S-nitrosothiols

The antibody-direct epifluorescent filter (Ab-DEFT) technique was evaluated as a rapid alternative to the most probable number (MPN) method for enumeration of artificially inoculated Listeria monocytogenes in ready-to-eat packaged salads and other fresh vegetables. Ab-DEFT was performed by homogenization of food in mesh-lined Stomacher bags, followed by prefiltration of homogenate through a 5 microns pore nylon filter, and passage of filtrate through a 0.4 micron pore black polycarbonate filter to collect and concentrate Listeria cells. After cells were stained with a fluorochrome-labeled polyclonal antibody to Listeria, the filter surface was examined by epifluorescence microscopy, and fluorescent cells were counted. A 3-tube MPN procedure was performed by successive enrichments of homogenized foods in Listeria enrichment and Fraser broths, followed by selective plating. Ab-DEFT provided quantitative determinations of Listeria cells that correlated with plate counts and MPN estimates in a linear response over a range of cell concentrations from 10 to 10(7) colony forming units (CFU)/mL. Microbial backgrounds as high as 10(8) CFU/mL did not affect performance of Ab-DEFT. In contrast to the MPN method, which required 5 days to perform, quantitation by Ab-DEFT could be completed in less than 1 h. Despite cross-reactivities demonstrated by the polyclonal fluorescent antibody, the potential of Ab-DEFT as a rapid alternative to MPN for microbial cell enumeration was evident.