Increase In Pulmonary Microvascular Permeability Research Articles

Role of β-arrestin-1 in penehyclidine hydrochloride-induced inhibition of LPS-caused increase in pulmonary microvascular permeability in human pulmonary microvascular endothelial cells

Objective To evaluate the role of β-arrestin-1 in penehyclidine hydrochloride(PHC)-induced inhibition of lipopolysaccharide(LPS)-caused increase in pulmonary microvascular permeability in human pulmonary microvascular endothelial cells(PMVECs). Methods Human PMVECs were seeded in 6-well plates(2 ml/well)or in culture flasks(4 ml/flask)at the density of 1×105 cells/ml and divided into 5 groups(n=15 each)using a random number table: empty plasmid transfection group(group C), LPS plus empty plasmid transfection group(LPS group), PHC plus LPS plus empty plasmid transfection group(P+ LPS group), LPS plus β-arrestin-1 short hairpin RNA(shRNA)transfection group(LPS+ shRNA group)and PHC plus LPS plus β-arrestin-1 shRNA transfection group(P+ LPS+ shRNA group). In LPS and LPS+ shRNA groups, the cells were transfected with empty plasmid 1.5 μg or with plasmid containing 15 nmol/L β-arrestin-1 shRNA, LPS with the final concentration of 0.1 μg/ml was added at 24 h of incubation, and the cells were then incubated for 1 h. In P+ LPS and P+ LPS+ shRNA groups, the cells were transfected with empty plasmid 1.5 μg or with plasmid containing 15 nmol/L β-arrestin-1 shRNA, PHC with the final concentration of 2 μg/ml was added at 24 h of incubation, LPS with the final concentration of 0.1 μg/ml was added at 1 h of incubation, and the cells were then incubated for 1 h. The cell permeability was measured using Transwell chambers.The expression of heat shock protein(HSP27)was detected by immunofluorescence.The expression of β-arrestin-1, p38 mitogen-activated protein kinase(p38MAPK)and phosphorylated p38MAPK(p-p38MAPK)was detected by Western blot.The ratio of p-p38MAPK/p38MAPK was calculated. Results Compared with group C, the cell permeability was significantly increased, the expression of HSP27 was up-regulated, p-p38MAPK/p38MAPK ratio was increased, and the expression of β-arrestin-1 was down-regulated in LPS, LPS+ shRNA and P+ LPS+ shRNA groups(P 0.05). Compared with group LPS, the cell permeability was significantly decreased, the expression of HSP27 was down-regulated, p-p38MAPK/p38MAPK ratio was decreased, and the expression of β-arrestin-1 was up-regulated in group P+ LPS, and p-p38MAPK/p38MAPK ratio was significantly increased(P 0.05). Compared with group P+ LPS, the cell permeability was significantly increased, the expression of HSP27 was up-regulated, p-p38MAPK/p38MAPK ratio was increased, and the expression of β-arrestin-1 was down-regulated in group P+ LPS+ shRNA(P<0.05). Conclusion The mechanism by which PHC inhibits LPS-induced increase in pulmonary microvascular permeability is totally related to β-arrestin-1 in human PMVECs. Key words: Arrestins; Cholinergic antagonists; Endotoxins; Lung; Capillary permeability

The mechanism underlying alpinetin-mediated alleviation of pancreatitis-associated lung injury through upregulating aquaporin-1.

Characterized by its acute onset, critical condition, poor prognosis, and high mortality rate, severe acute pancreatitis (SAP) can cause multiple organ failure at its early stage, particularly acute lung injury (ALI). The pathogenesis of ALI is diffuse alveolar damage, including an increase in pulmonary microvascular permeability, a decrease in compliance, and invasion of many inflammatory cells. Corticosteroids are the main treatment method for ALI; however, the associated high toxicity and side effects induce pain in patients. Recent studies show that the effective components in many traditional Chinese medicines can effectively inhibit inflammation with few side effects, which can decrease the complications caused by steroid consumption. Based on these observations, the main objective of the current study is to investigate the effect of alpinetin, which is a flavonoid extracted from Alpinia katsumadai Hayata, on treating lung injury induced by SAP and to explore the mechanism underlying the alpinetin-mediated decrease in the extent of ALI. In this study, we have shown through in vitro experiments that a therapeutic dose of alpinetin can promote human pulmonary microvascular endothelial cell proliferation. We have also shown via in vitro and in vivo experiments that alpinetin upregulates aquaporin-1 and, thereby, inhibits tumor necrosis factor-α expression as well as reduces the degree of lung injury. Overall, our study shows that alpinetin alleviates SAP-induced ALI. The likely molecular mechanism includes upregulated aquaporin expression, which inhibits tumor necrosis factor-α and, thus, alleviates SAP-induced ALI.

Cardiopulmonary bypass increases pulmonary microvascular permeability through the Src kinase pathway: Involvement of caveolin-1 and vascular endothelial cadherin.

Changes in pulmonary microvascular permeability following cardiopulmonary bypass (CPB) and the underlying mechanisms have not yet been established. Therefore, the aim of the present study was to elucidate the alterations in pulmonary microvascular permeability following CPB and the underlying mechanism. The pulmonary microvascular permeability was measured using Evans Blue dye (EBD) exclusion, and the neutrophil infiltration and proinflammatory cytokine secretion was investigated. In addition, the activation of Src kinase and the phosphorylation of caveolin-1 and vascular endothelial cadherin (VE-cadherin) was examined. The results revealed that CPB increased pulmonary microvascular leakage, neutrophil count and proinflammatory cytokines in the bronchoalveolar lavage fluid, and activated Src kinase. The administration of PP2, an inhibitor of Src kinase, decreased the activation of Src kinase and attenuated the increase in pulmonary microvascular permeability observed following CPB. Two important proteins associated with vascular permeability, caveolin-1 and VE-cadherin, were significantly activated at 24 h in the lung tissues following CPB, which correlated with the alterations in pulmonary microvascular permeability and Src kinase. PP2 administration inhibited their activation, suggesting that they are downstream factors of Src kinase activation. The data indicated that the Src kinase pathway increased pulmonary microvascular permeability following CPB, and the activation of caveolin-1 and VE-cadherin may be involved. Inhibition of this pathway may provide a potential therapy for acute lung injury following cardiac surgery.

Documentation of pulmonary capillary permeability in the adult respiratory distress syndrome accompanying human sepsis.

Current evidence suggests that pulmonary edema accompanying human sepsis may result either from changes in the serum oncotic and hydrostatic pressures or an increase in the permeability of the pulmonary microvasculature. In this study, we compared the "clearance" of injected 131I-labeled human serum albumin from blood to bronchoalveolar secretions in intubated patients with pulmonary edema secondary to sepsis or myocardial infarction. A significantly increased mean +/- SE clearance of the radionuclide was seen in patients with sepsis (0.34 +/- 0.03 ml per hour) compared to those with myocardial infarction (0.043 +/- 0.008 ml per hour) (P less than 0.001), although both groups had similar degrees of edema on chest radiographs. Because the patients with sepsis had no severe decrease in serum oncotic pressure (18.4 +/- 5.0 mm Hg) or evidence of left heart failure, as determined by the pulmonary wedge pressure (11.0 +/- 6.8 mm Hg), we concluded that the genesis of the pulmonary edema in sepsis was due to an increase in pulmonary microvascular permeability, as measured by the increased clearance of 131I-labeled human serum albumin.

SC5b-9-Induced Pulmonary Microvascular Endothelial Hyperpermeability Participates in Ventilator-Induced Lung Injury

Mechanical ventilation with large tidal volumes can increase lung alveolar permeability and initiate inflammatory responses, termed ventilator-induced lung injury (VILI). VILI is characterized by an influx of inflammatory cells, increased pulmonary permeability, and endothelial and epithelial cell death. But the underlying molecular mechanisms that regulate VILI remain unclear. The purpose of this study was to investigate the mechanisms that regulate pulmonary endothelial barrier in an animal model of VILI. These data suggest that SC5b-9, as the production of the complement activation, causes increase in rat pulmonary microvascular permeability by inducing activation of RhoA and subsequent phosphorylation of myosin light chain and contraction of endothelial cells, resulting in gap formation. In general, the complement-mediated increase in pulmonary microvascular permeability may participate in VILI.

Administration of a Peroxynitrite Decomposition Catalyst Into the Bronchial Artery Attenuates Pulmonary Dysfunction After Smoke Inhalation and Burn Injury in Sheep

Reactive nitrogen species such as peroxynitrite play a significant role in burn and smoke inhalation injury. The bronchial circulation increases more than 10-fold in response to this combination injury. We hypothesized that direct delivery of low-dose WW-85, a peroxynitrite decomposition catalyst, into the bronchial artery would attenuate burn- and smoke inhalation-induced acute lung injury. In adult female sheep (n = 17), the bronchial artery was cannulated in preparation surgery. After a 5- to 7-day recovery period, sheep were subjected to a burn (40% total body surface area, third degree) and inhalation injury (48 breaths of cotton smoke, <40°C). The animals were divided into three groups following the injury: (i) WW-85 group: 1 h after injury, WW-85 (0.002 mg/kg per hour) was continuously infused into the bronchial artery, n = 5; (ii) control group: 1 h after injury, an equivalent amount of saline was injected into the bronchial artery, n = 6; (iii) sham group: no injury, no treatment, same operation and anesthesia, n = 6. All animals were mechanically ventilated and fluid resuscitated equally. In the control group, the injury induced a severe deterioration of pulmonary oxygenation and shunting and an increase in pulmonary microvascular permeability toward sham. The injury was further associated with an increase in reactive nitrogen species in lung tissues of the control group. All these alterations were significantly attenuated in the WW-85 group. We demonstrated that a low dosage of WW-85 directly administered into the bronchial artery attenuated pulmonary dysfunction to the same extent as higher systemically administered doses in previous experiments. Our data strongly suggest that local airway production of peroxynitrite contributes to pulmonary dysfunction following smoke inhalation and burn injury.

Endothelial connexin43 mediates acid-induced increases in pulmonary microvascular permeability

Acid aspiration, a common cause of acute lung injury, leads to alveolar edema. Increase in lung vascular permeability underlies this pathology. To define mechanisms, isolated rat lungs were perfused with autologous blood. Hydrochloric acid and rhodamine-dextran 70 kDa (RDx70) were coinstilled into an alveolus by micropuncture. RDx70 fluorescence was used to establish the spatial distribution of acid. Subsequently, FITC-dextran 20 kDa (FDx20) was infused into microvessels for 60 min followed by a 10-min HEPES-buffered saline wash. During the infusion, FITC fluorescence changes were recorded to quantify the ratio of peak to postwash fluorescence. The ratio, termed normalized fluorescence, was low for acid compared with buffer instillation both in microvessels abutting acid-treated alveoli and those located more than 700 μm away. In contrast, the normalized fluorescence was similar to buffer controls when a higher molecular weight tracer (FITC-dextran 70 kDa) was infused instead of FDx20, suggesting that normalized FDx20 fluorescence faithfully represented microvascular permeability. Inhibiting endothelial connexin43 (Cx43) gap junction communication with Gap27 blunted the acid-induced reduction in normalized fluorescence, although scrambled Gap27 did not have any effect. The blunting was evident not only in microvessels away from the site of injury, but also in those abutting directly injured alveoli. Thus the new fluorescence-based method reveals that acid increases microvascular permeability both at acid-instilled and away sites. Inhibiting endothelial Cx43 blocked the permeability increase even at the direct injury sites. These data indicate for the first time that Cx43-dependent mechanisms mediate acid-induced increases in microvascular permeability. Cx43 may be a therapeutic target in acid injury.

Ulinastatin Improves Pulmonary Function in Severe Burn-Induced Acute Lung Injury by Attenuating Inflammatory Response

Acute systemic inflammatory response to severe skin burn injury mediates burn-induced acute lung injury. Ulinastatin is potentially an effective intervention, because it attenuates the systemic inflammatory response induced by endotoxin and improves myocardial function during ischemic shock and reperfusion. Rats received full-thickness burn wounds to 30% total body surface area followed by delayed resuscitation. The treatment group received 50,000 U/kg of ulinastatin and the burn group was given vehicle only. A sham group was not burned but otherwise was treated identically. After killing, blood and lung samples were harvested for histology and measurement of inflammatory mediators. Administration of ulinastatin significantly decreased the mRNA and protein levels of tumor necrosis factor-alpha, interleukin-1β, -6, and -8 both locally and systemically in burn-injured rats. The secretion of neutrophil elastase and myeloperoxidase in the lung and the expression of intercellular adhesion molecule-1 on the surface of lung epithelium were inhibited by ulinastatin. Ulinastatin also reduced the increase in pulmonary microvascular permeability. Consistent with these findings, ulinastatin ameliorated the lung edema and pulmonary oxygenation in burn-injured rats. These results indicate that the inhibitory effects of ulinastatin on inflammatory mediator production, neutrophil activation, and microvascular permeability are associated with the recovery of pulmonary functions in severe burn-induced acute lung injury and suggest that ulinastatin may serve as a potential therapeutic administration in critical burn care.

Bone Marrow Progenitor Cells Induce Endothelial Adherens Junction Integrity by Sphingosine-1-Phosphate–Mediated Rac1 and Cdc42 Signaling

Little is known about the contribution of bone marrow-derived progenitor cells (BMPCs) in the regulation endothelial barrier function as defined by microvascular permeability alterations at the level of adherens junctions (AJs). We investigated the role of BMPCs in annealing AJs and thereby in preventing lung edema formation induced by endotoxin (LPS). We observed that BMPCs enhanced basal endothelial barrier function and prevented the increase in pulmonary microvascular permeability and edema formation in mice after LPS challenge. Coculture of BMPCs with endothelial cells induced Rac1 and Cdc42 activation and AJ assembly in endothelial cells. However, transplantation of BMPCs isolated from sphingosine kinase-1-null mice (SPHK1(-/-)), having impaired S1P production, failed to activate Rac1 and Cdc42 or protect the endothelial barrier. These results demonstrate that BMPCs have the ability to reanneal endothelial AJs by paracrine S1P release in the inflammatory milieu and the consequent activation of Rac-1 and Cdc42 in endothelial cells.

Drotrecogin Alfa (activated) Prevents Smoke-Induced Increases in Pulmonary Microvascular Permeability and Proinflammatory Cytokine IL-1β in Rats

In an established rat model of smoke inhalation injury, we conducted a dose-response study to examine the protective effects of Xigris [drotrecogin alfa (activated) (DrotAA)], a recombinant form of human activated protein C (APC). DrotAA is a serine protease (approximately 55 kD molecular weight) with the same amino acid sequence and the glycosylation site as human plasma-derived APC. A total of 120 F344/NH rats (half each gender, approximately 175 g body weight) were randomly divided into five groups and exposed nose-only to air or diesel fuel smoke for 20 min. These rats were then i.v. administered with DrotAA in 0, 5, 10, and 20 mg/kg body weight, respectively, immediately following smoke exposure. Treatment with DrotAA significantly attenuated smoke inhalation injury in a dose-dependent manner at 2 hours after insult, as indicated by preserving microvascular permeability and proinflammatory cytokine IL-1beta (but not TNF-alpha and neuropeptide substance P) in bronchoalveolar lavage fluid (BALF). Moreover, the rats treated with 20 mg/ kg of DrotAA had an improvement of the expiration phase of pulmonary dynamic compliance. At all dosages, however, DrotAA also significantly increased all phases of pulmonary resistance compared with either the controls or to smoke inhalation alone. Generally, these data suggest that DrotAA may exert an anti-inflammatory effect by inhibiting cytokine-mediated inflammatory amplification. However, additional studies following a clinical course are needed to confirm the maximum efficiency and possible side effects of this recombined human activated protein C.

Recombinant plasma gelsolin infusion attenuates burn-induced pulmonary microvascular dysfunction.

Reduced plasma concentrations of the extracellular actin-binding proteins gelsolin and Gc-globulin correlate with pulmonary failure and death in humans after injury. The purpose of this study was to investigate the role of plasma gelsolin in the pathophysiology of inflammation-induced lung injury. We postulated that plasma gelsolin levels decrease at an early time point after burn injury and that the intravenous infusion of gelsolin prevents burn-induced pulmonary microvascular dysfunction. Adult Sprague-Dawley rats were randomized to undergo a 40% body surface area thermal injury (Burn) or manipulation without burn (Sham). Plasma gelsolin and Gc-globulin concentrations were determined at various times during the first 6 days of injury by Western blotting. Other animals were randomized to receive either recombinant human gelsolin (0.078, 0.78, or 7.8 mg) or albumin (7.8 mg) before and 8 h after Burn or Sham. Twenty-four hours later, pulmonary microvascular permeability was assessed by measuring the capillary filtration by use of an isolated, perfused lung model. We found that plasma gelsolin levels of burn-injured rats decreased to 10% of normal levels within 12 h and remained below normal levels for up to 6 days postinjury. Gc-globulin values also fall, but to a lesser extent and only transiently. Treatment of burned animals with intravenous infusions of recombinant human gelsolin prevented the increase in pulmonary microvascular permeability that accompanies this injury. Our findings are consistent with the hypothesis that plasma gelsolin depletion contributes to the pathophysiology of pulmonary microvascular dysfunction during inflammation.

Abrogation of thrombin-induced increase in pulmonary microvascular permeability in PAR-1 knockout mice.

We investigated the function of proteinase-activated receptor-1 (PAR-1) in the regulation of pulmonary microvascular permeability in response to thrombin challenge using PAR-1 knockout mice (-/-). Lungs were isolated and perfused with albumin (5 g/100 ml)-Krebs solution at constant flow (2 ml/min). Lung wet weight and pulmonary artery pressure (P(pa)) were continuously monitored. We determined the capillary filtration coefficient (K(fc)) and (125)I-labeled albumin (BSA) permeability-surface area product (PS) to assess changes in pulmonary microvessel permeability to liquid and albumin, respectively. Normal and PAR-1-null lung preparations received in the perfusate: 1) thrombin or 2) selective PAR-1 agonist peptide (TFLLRNPNDK-NH(2)). In control PAR-1 (+/+) mouse lungs, (125)I-albumin PS and K(fc) were significantly increased over baseline (by approximately 7- and 1.5-fold, respectively) within 20 min of alpha-thrombin (100 nM) challenge. PAR-1 agonist peptide (5 microM) gave similar results, whereas control peptide (5 microM; FTLLRNPNDK-NH(2)) was ineffective. At relatively high concentrations, thrombin (500 nM) or PAR-1 agonist peptide (10 microM) also induced increases in P(pa) and lung wet weight. All effects of thrombin (100 or 500 nM) or PAR-1 agonist peptide (5 or 10 microM) were prevented in PAR-1-null lung preparations. Baseline measures of microvessel permeability and P(pa) in the PAR-1-null preparations were indistinguishable from those in normal lungs. Moreover, PAR-1-null preparations gave normal vasoconstrictor response to thromboxane analog, U-46619 (100 nM). The results indicate that the PAR-1 receptor is critical in mediating the permeability-increasing and vasoconstrictor effects of thrombin in pulmonary microvessels.

Effect of femoral fracture and intramedullary fixation on lung capillary leak.

Pulmonary injury is an important complication in the trauma patient with long-bone fractures. The purpose of this study was to determine the effect of femoral fracture or fracture and intramedullary fixation on lung capillary leak. The contribution of leukocytes to lung injury in this model was also determined. The pulmonary capillary filtration coefficient was determined in lungs of rats after femur fracture or fracture and reamed or unreamed intramedullary fixation. Pulmonary arterial vascular resistance and lung neutrophil content were also determined. Fracture alone did not cause lung injury, whereas fracture and intramedullary fixation elicited lung capillary leak. Fracture alone and intramedullary fixation increased pulmonary vascular resistance, whereas unreamed intramedullary fixation caused lung leukosequestration. Femoral fracture alone does not cause an increase in pulmonary microvascular permeability. Femoral fracture and intramedullary fixation causes lung capillary leak, which is not increased by reaming the femoral canal.

Improvement of Rat Lung Structure and Function after Preservation with Celsior

Ischemia/reperfusion-induced increase in pulmonary microvascular permeability was shown to be reduced after preservation with Celsior. We investigated reimplantation-induced lung injury in isolated, reperfused rat lungs after preservation via the pulmonary artery with Celsior, Celsior + prostacyclin, and reduced-potassium (40 mmol) Euro-Collins solution (40 ml/kg/body wt each) followed by 2 h of cold ischemia. Arterial and veneous oxygen tensions were recorded during 50 min ofin vitroreperfusion after which the lungs (10 right lungs per experimental group) were fixed by vascular perfusion. The tissue was further processed for microscopy, and histological changes were quantified stereologically. Lung preservation with Celsior resulted in a significantly higher volume of air-filled alveolar space with a large proportion of widely distended alveoli compared with the other groups. In the Euro-Collins group the fraction of atelectatic alveoli exceeded that observed in Celsior-preserved lungs. In accordance, the difference between arterial and venous oxygen tensions was significant among Euro-Collins- and Celsior-protected lungs, with improved oxgenation values in the Celsior group. In contrast, addition of prostacyclin to Celsior treatment resulted in rather variable structural as well as functional data. There were no differences in the volumes of intraalveolar edema among the groups tested. However, the volume of alveolar tissue was increased in the Euro-Collins group. In conclusion, compared with Euro-Collins and Celsior + prostacyclin solutions, preservation with Celsior resulted in improved structural characteristics which in combination with improved oxygenation parameters supports the prospective advantage of Celsior in clinical organ preservation.

Aspiration of activated charcoal elicits an increase in lung microvascular permeability.

Gastric decontamination with orally administered activated charcoal is the recommended treatment for many poisonings. However, ingestion of central nervous system depressants resulting in loss of protective airway reflexes may result in pulmonary aspiration of activated charcoal. Although activated charcoal has been reported to be an inert substance, evidence suggests that pulmonary aspiration of charcoal is associated with lung edema formation and pulmonary compromise. This study tested the hypothesis that intratracheal instillation of activated charcoal disrupts the integrity of the lung microvascular barrier. The capillary filtration coefficient (Kf,c), a sensitive measure of lung microvascular permeability, was determined isogravimetrically prior to and after intratracheal instillation of activated charcoal 0.04 g/kg (12% weight/vol solution, pH 7.4) or an equal volume of sterile water in isolated, perfused rat lungs. Arterial blood gas analysis was determined prior to and after tracheal instillation of activated charcoal or sterile water in a separate group of animals. Intratracheal instillation of activated charcoal resulted in a significant increase in pulmonary microvascular permeability compared to lungs treated with sterile water or control lungs (delta Kf,c = +0.21 +/- 0.076; -0.014 +/- 0.04; and -0.041 +/- 0.02 mL/min/cm H2O/100 g lung tissue, respectively, p < 0.05 ANOVA). There was no significant difference in baseline blood gases in the 3 experimental groups. There was a significant decrease in arterial Po2, bicarbonate, and pH in animals administered activated charcoal compared to time-matched controls and animals administered sterile water. Intratracheal instillation of activated charcoal is associated with a significant increase in lung microvascular permeability and arterial blood gas derangements. The effects of activated charcoal on pulmonary microvascular barrier integrity may contribute to the lung edema formation and pulmonary compromise observed following clinical aspiration of activated charcoal.

Rolipram and Isoproterenol Reverse Platelet Activating Factor-Induced Increases in Pulmonary Microvascular Permeability and Vascular Resistance

Platelet activating factor (PAF) is an important mediator of pulmonary microvascular endothelial cell (PMVEC) injury in sepsis. Membrane receptors for PAF have been identified on PMVECs and mediate its actions at least in part by protein kinase C activation. Since rolipram, a family IV cyclic AMP phosphodiesterase inhibitor, and isoproterenol, an adenylate cyclase activator, both reverse ischemia-reperfusion-induced lung permeability, we studied the effects of these agents on PAF-induced pulmonary microvascular permeability. The isolated rat lung model was used in which lungs were ventilated and buffer perfused at constant flow while suspended from a force transducer to monitor lung weight along with arterial ( P a) and venous ( P v) pressures. Control lungs ( n = 6) were infused with PAF (40 nmole/kg) via an arterial port and the capillary permeability coefficient ( K f,c) was determined at 0, 15, and 60 min. The remaining lungs were randomized for infusion with either rolipram ( n = 4, 20 μmole/kg) or isoproterenol ( n = 4, 5 μmole/kg) via an arterial port 30 min after injury with PAF. In the rolipram- and isoproterenol-treated groups, the K f,c was determined 15 and 60 min postinfusion with these agents. The control group showed significant elevation in the K f,c and total pulmonary resistance ( R t). At 15 and 60 min, rolipram and isoproterenol reversed PAF injury as shown by the significant improvement in the K f,c and R t. These findings support the concept that increased cyclic AMP is an important mediator in the reversal of PAF-increased PMVEC permeability and pulmonary resistance.

Dose-dependent mediation of leukotriene D 4-induced airway microvascular leakage and bronchoconstriction in the guinea pig

The i.v. administration of leukotriene (LT)D 4 to anesthetized guinea pigs produced dose-dependent increases in pulmonary microvascular permeability, as measured by extravasation of Evans blue dye into the trachea, main bronchi, and small airways, with an ED 50 of approximately 0.05 μg/kg. When LTD 4 was administered at 0.3 μg/kg, the resulting plasma extravasation into all three airway sections was markedly reduced by pretreatment with a cyclooxygenase inhibitor, meclofenamic acid (2.5 mg/kg, i.v.), a thromboxane (TX) receptor antagonist, SQ 29,548 (0.1 or 1 mg/kg, i.v.), or a peptidoleukotriene receptor antagonist, pranlukast (SB 205312) (0.1 or 1 mg/kg, i.v.), but not by the H 1 histamine receptor antagonist, pyrilamine. When LTD 4 was administered at 1.0 μg/kg, meclofenamate (2.5 or 5 mg/kg, i.v.) or SQ 29,548 slightly attenuated plasma extravasation only in the small airway, whereas pranlukast was effective in all three airway segments. Administration of the 5-lipoxygenase inhibitor, zileuton (10 mg/kg, i.v.), or the PAF antagonist, L-659,989 (5 mg/kg, i.v.), did not affect the microvascular leakage response to 1.0 μg/kg LTD 4. In addition, i.v.-administered LTD 4 (0.3 or 1.0 μg/kg) or the prostaglandin (PG)/TXA 2 receptor agaonist, U-46619 (3.0 μg/kg), produced significant bronchoconstriction as measured by increases in pulmonary insufflation pressure. The bronchoconstrictor responses to LTD 4 were markedly attenuated by the same inhibitors, namely meclofenamic acid, SQ 29,548, and pranlukast, that reduced the 0.3 μg/kg LTD 4-induced plasma extravasation throughout the airways and the 1.0 μg/kg LTD 4-induced extravasation into the small airways. U-46619-induced bronchoconstriction was blocked only by SQ 29,548. Based on the profile of antagonists employed in the present studies, we conclude that microvascular leakage and bronchoconstriction elicited by i.v. LTD 4 are mediated through a direct and an indirect pathway which includes the formation of cyclooxygenase product(s) which act at PG/TXA 2 receptors. The pattern of inhibition of the LTD 4-induced bronchoconstriction most closely mimicked the inhibitor profile of microvascular leakage in the small airways, suggesting a greater relative contribution of this site to activity resulting from cyclooxygenase product formation or action. The ability of pranlukast to abolish both responses to i.v. LTD 4 suggests that the phospholipase(s) activated to produce the substrate for cyclooxygenase metabolism are pranlukast sensitive.

Chemokine expression during hepatic ischemia/reperfusion-induced lung injury in the rat. The role of epithelial neutrophil activating protein.

The liver is highly susceptible to a number of pathological insults, including ischemia/reperfusion injury. One of the striking consequences of liver injury is the associated pulmonary dysfunction that may be related to the release of hepatic-derived cytokines. We have previously employed an animal model of hepatic ischemia/reperfusion injury, and demonstrated that this injury causes the production and release of hepatic-derived TNF, which mediates a neutrophil-dependent pulmonary microvascular injury. In this study, we have extended these previous observations to assess whether an interrelationship between TNF and the neutrophil chemoattractant/activating factor, epithelial neutrophil activating protein-78 (ENA-78), exists that may be accountable for the pathology of lung injury found in this model. In the context of hepatic ischemia/reperfusion injury, we demonstrated the following alterations in lung pathophysiology: (a) an increase in pulmonary microvascular permeability, lung neutrophil sequestration, and production of pulmonary-derived ENA-78; (b) passive immunization with neutralizing TNF antiserum resulted in a significant suppression of pulmonary-derived ENA-78; and (c) passive immunization with neutralizing ENA-78 antiserum resulted in a significant attenuation of pulmonary neutrophil sequestration and microvascular permeability similar to our previous studies with anti-TNF. These findings support the notion that pulmonary ENA-78 produced in response to hepatic-derived TNF is an important mediator of lung injury.

Association of Pulmonary Inflammation and Increased Microvascular Permeability During the Development of Bronchopulmonary Dysplasia: A Sequential Analysis of Inflammatory Mediators in Respiratory Fluids of High-Risk Preterm Neonates

Bronchopulmonary dysplasia (BPD) of preterm neonates is associated with an increased recruitment of inflammatory cells into the airways. To evaluate further the role of inflammation in the pathogenesis of BPD, tracheobronchial aspirate fluid of neonates with birth weight < 1200 g (n = 59) was sequentially analyzed in a prospective study. Tracheobronchial aspirate fluid was assessed for chemotactic activity, neutrophil cell count, concentrations of elastase-alpha 1-proteinase inhibitor and activity of free elastase, concentrations of chemoattractants (complement component C5-derived anaphylatoxin, leukotriene B4, interleukin-8), and albumin concentrations as well as alpha 1-proteinase inhibitor activity. The secretory component for immunoglobulin A was used as reference protein. Only specimens without evidence of microbiological colonization were studied. In neonates with prolonged respiratory disease (BPD-risk neonates, n = 24, fraction of inspired oxygen > or = 0.3 and/or peak inspiratory pressure > or = 16 cm H2O at day 10 postnatal age, birth weight 892 +/- 36 g, gestational age 27.2 +/- 0.3 weeks) chemotactic activity, cell count, concentrations of the chemoattractants complement component C5-derived anaphylatoxin, leukotriene B4, interleukin-8, as well as levels of elastase-alpha 1-proteinase inhibitor were significantly higher at day 10 and/or day 15 of postnatal age compared with neonates without chronic pulmonary disease (total n = 35; day 10, n = 11; day 15, n = 8). There was no difference in free elastolytic activity. Concentrations of albumin as well as alpha 1-proteinase inhibitor activity were higher in BPD-risk patients on day 15, indicating an increased pulmonary leak. We conclude that preterm neonates at risk for the development of BPD show an enhanced inflammatory reaction in the lungs and an associated increase in pulmonary microvascular permeability. We speculate that inflammation may play an important role in the pathogenesis of BPD.

Roles of neutrophil elastase and superoxide anion in leukotriene B4-induced lung injury in rabbit

The effects of a competitive neutrophil elastase (NE) inhibitor, ONO-5046, and a recombinant human superoxide dismutase on leukotriene B4 (LTB4)-induced polymorphonuclear leukocyte (PMN)-mediated increase in microvascular permeability in isolated non-blood-perfused rabbit lungs were studied. Pulmonary microvascular permeability and lung edema were evaluated by use of the fluid filtration coefficient (Kf) and the wet-to-dry lung weight ratio (W/D), respectively. Pulmonary capillary pressure was estimated by the double occlusion technique. NE activity in the perfusate was determined using a spectrophotometric method. The PMNs (2-3 x 10(8) cells) were added into the perfusate in all groups of lungs. Injection of LTB4 (5 micrograms) increased Kf and W/D without affecting pulmonary arterial or capillary pressure. The LTB4-induced lung injury was closely associated with the increase in NE activity in the perfusate. Infusion of ONO-5046 (1 or 10 mg.kg-1 x h-1) inhibited the LTB4-induced increases in Kf, W/D, and perfusate NE activity in a dose-dependent fashion. Infusion of recombinant human superoxide dismutase (80,000 U.kg-1 x h-1) attenuated the LTB4-induced increases in Kf and W/D, although it did not influence the elevation of perfusate NE activity induced by LTB4. These results suggest that both NE and superoxide anion play important roles in the LTB4-induced PMN-mediated increase in pulmonary microvascular permeability.