Pulmonary microvascular response to LTB4: effects of perfusate composition

We examined the effects of leukotriene B4 (LTB4) on pulmonary hemodynamics and vascular permeability using isolated perfused guinea pig lungs and cultured monolayers of pulmonary arterial endothelial cells. In lungs perfused with Ringer solution, containing 0.5 g/100 ml albumin (R-alb), LTB4 (4 micrograms) transiently increased pulmonary arterial pressure (Ppa) and capillary pressure (Pcap). Pulmonary edema developed within 70 min after LTB4 injection despite a normal Pcap. The LTB4 metabolite, 20-COOH-LTB4 (4 micrograms), did not induce hemodynamic and lung weight changes. In lungs perfused with autologous blood hematocrit = 12 +/- 1%; protein concentration = 1.5 +/- 0.2 g/100 ml), the increases in Ppa and Pcap were greater, and both pressures remained elevated. The lung weight did not increase in blood-perfused lungs. In lungs perfused with R-alb (1.5 g/100 ml albumin) to match the blood perfusate protein concentration, LTB4 induced similar hemodynamic changes as R-alb (0.5 g/100 ml) perfusate, but the additional albumin prevented the pulmonary edema. LTB4 (10(-11)-10(-6) M) with or without the addition of neutrophils to the monolayer did not increase endothelial 125I-albumin permeability. Therefore LTB4 induces pulmonary edema when the perfusate contains a low albumin concentration, but increasing the albumin concentration or adding blood cells prevents the edema. The edema is not due to increased endothelial permeability to protein and is independent of hemodynamic alterations. Protection at higher protein-concentration may be the result of LTB4 binding to albumin.     

Increased lung vascular permeability after arachidonic acid and hydrostatic challenge

Arachidonic acid (AA) metabolites are known to be potent vasoactive substances in the pulmonary circulation, whereas their influence on lung vascular permeability is still uncertain. We investigated the effect of AA bolus injection on the capillary filtration coefficient (Kf,C) of isolated rabbit lungs, recirculatingly perfused with Krebs-Henseleit albumin (1%) buffer. Kf,C was measured using repetitive sudden venous pressure elevations (7.5 Torr) and time zero extrapolation of the slope of the weight gain curve. It ranged from 1.3 to 2.4 cm3 X s-1 X Torr-1 X g-1 X 10(-4) in control lungs. Pulmonary arterial injection of AA (100 microM; in presence of 20 microM indomethacin to suppress pulmonary arterial pressure rise) during an acute hydrostatic challenge, but not at zero venous pressure, caused a greater than 10-fold increase in Kf,C. Vascular compliance was not altered. Additional experiments, performed under zero-flow conditions to avoid any ambiguity in microvascular pressure, corroborated the severalfold increase in vascular permeability, detectable within 3 min after AA application during acute hydrostatic challenge.     

Alveolar epithelial barrier functions in ventilated perfused rabbit lungs

We employed ultrasonic nebulization for homogeneous alveolar tracer deposition into ventilated perfused rabbit lungs. (22)Na and (125)I-albumin transit kinetics were monitored on-line with gamma detectors placed around the lung and the perfusate reservoir. [(3)H]mannitol was measured by repetitive counting of perfusion fluid samples. Volume of the alveolar epithelial lining fluid was estimated with bronchoalveolar lavage with sodium-free isosmolar mannitol solutions. Sodium clearance rate was -2.2 +/- 0.3%/min. This rate was significantly reduced by preadministration of ouabain/amiloride and enhanced by pretreatment with aerosolized terbutaline. The (125)I-albumin clearance rate was -0.40 +/- 0.05%/min. The appearance of [(3)H]mannitol in the perfusate was not influenced by ouabain/amiloride or terbutaline but was markedly enhanced by pretreatment with aerosolized protamine. An epithelial lining fluid volume of 1.22 +/- 0.21 ml was calculated in control lungs. Fluid absorption rate was 1.23 microl x g lung weight(-1) x min(-1), which was blunted after pretreatment with ouabain/amiloride. We conclude that alveolar tracer loading by aerosolization is a feasible technique to assess alveolar epithelial barrier properties in aerated lungs. Data on active and passive sodium flux, paracellular solute transit, and net fluid absorption correspond well to those in previous studies in fluid-filled lungs; however, albumin clearance rates were markedly higher in the currently investigated aerated lungs.     

IL-2 induces pulmonary edema and vasoconstriction independent of circulating lymphocytes

We investigated the effect of IL-2 in the isolated guinea pig lung perfused with phosphate-buffered Ringer's solution (containing 0.5 g/100 ml albumin and 5.5 mM dextrose) to determine the mechanism of IL-2-induced pulmonary edema. IL-2 (0 to 10,000 U/ml) was added to the perfusate following a 10 min baseline steady-state period. Pulmonary arterial pressure (Ppa), pulmonary capillary pressure (Ppc), and change in lung weight (as a measure of developing pulmonary edema) were recorded at 0, 10, 30, 40, and 60 min. The capillary filtration coefficient (Kf.c), an index of vascular permeability to water, was measured at 30 and 60 min. Infusion of IL-2 increased Ppc (from 3.9 +/- 0.1 cm H2O at baseline to 8.8 +/- 1.1 cm H2O at 60 min for IL-2 at 2000 U/ml, p less than 0.01; and from 3.8 +/- 0.1 cm H2O at baseline to 8.9 +/- 0.6 cm H2O at 60 min for IL-2 at 10,000 U/ml, p less than 0.01. The lung weight also increased (32% at IL-2 concentration of 2000 U/ml, and 26% at IL-2 concentration of 10,000 U/ml) The capillary filtration coefficient did not change with IL-2 infusion. The IL-2 response was prevented using the pulmonary vasodilator, papaverine. The infusion of IL-2 was associated with the generation of thromboxane A2(TxA2) in the effluent perfusate. Inhibition of TxA2 synthetase using Dazoxiben prevented the pulmonary vasoconstriction and edema response to IL-2. In addition, IL-2 had no effect on the transendothelial clearance of 125I-albumin. The results indicate that IL-2 causes pulmonary edema secondary to an increase in Ppc. The response is mediated by IL-2 stimulation of TxA2 generation from the lung.     

Physiological and biochemical markers of alveolar epithelial barrier dysfunction in perfused human lungs

To study air space fluid clearance (AFC) under conditions that resemble the clinical setting of pulmonary edema in patients, we developed a new perfused human lung preparation. We measured AFC in 20 human lungs rejected for transplantation and determined the contribution of AFC to lung fluid balance. AFC was then compared with air space and perfusate levels of a biological marker of epithelial injury. The majority of human lungs rejected for transplant had intact basal (75%) and beta(2)-adrenergic agonist-stimulated (70%) AFC. For lungs with both basal and stimulated AFC, the basal AFC rate was 19 +/- 10%/h, and the beta(2)-adrenergic-stimulated AFC rate was 43 +/- 13%/h. Higher rates of AFC were associated with less lung weight gain (Pearson coefficient -0.90, P < 0.0001). Air space and perfusate levels of the type I pneumocyte marker receptor for advanced glycation end products (RAGE) were threefold and sixfold higher, respectively, in lungs without basal AFC compared with lungs with AFC (P < 0.05). These data show that preserved AFC is a critical determinant of favorable lung fluid balance in the perfused human lung, raising the possibility that beta(2)-agonist therapy to increase edema fluid clearance may be of value for patients with acute lung injury and pulmonary edema. Also, although additional studies are needed, a biological marker of alveolar epithelial injury may be useful clinically in predicting preserved AFC.     

Lung endothelial and epithelial permeability after platelet-activating factor

We investigated whether platelet-activating factor (PAF) increased epithelial or endothelial permeability in isolated-perfused rabbit lungs. PAF was either injected into the pulmonary artery or instilled into the airway of lungs perfused with Tyrode's solution containing 1% bovine serum albumin. The effect of adding neutrophils or platelets to the perfusate was also tested. Perfusion was maintained 20-40 min after adding PAF and then a fluid filtration coefficient (Kf) was determined to assess vascular permeability. At the end of each experiment, one lung was lavaged, and the lavagate protein concentration (BALP) was determined. Wet weight-to-dry weight ratios (W/D) were determined on the other lung. PAF added to the vascular space increased peak pulmonary arterial pressure (Ppa) from 13.5 +/- 3.1 (mean +/- SE) to 24.2 +/- 3.3 cmH2O (P less than 0.05). The effect was amplified by platelets [Ppa to 70.8 +/- 8.0 cmH2O (P less than 0.05)] but not by neutrophils [Ppa to 22.0 +/- 1.4 cmH2O (P less than 0.05)]. Minimal changes in Ppa were observed after instilling PAF into the airway. The Kf, W/D, and BALP of untreated lungs were not increased by injecting PAF into the vasculature or into the air space. The effect of PAF on Kf, W/D, and BALP was unaltered by adding platelets or neutrophils to the perfusate. PAF increases intravascular pressure (at a constant rate of perfusion) but does not increase epithelial or endothelial permeability in isolated-perfused rabbit lungs.     

Evans blue dye as a marker of albumin clearance in cultured endothelial monolayer and isolated lung.

Determination of protein transfer across the endothelial barrier or the entire alveolar capillary membrane is critical for investigation of mechanisms leading to pulmonary edema. The purpose of this study was to evaluate Evans blue dye for determination of protein clearance across cultured bovine pulmonary artery endothelial cell monolayers and as a quantitative marker for albumin leakage to the air spaces in isolated perfused rat lungs. Evans blue dye bound tightly to albumin (EBA) as determined by lack of transfer through dialysis membranes and specific elution with albumin from a molecular exclusion column. EBA was equivalent to 125I-labeled albumin for calculation of albumin clearance rates (Calb) across intact and challenged monolayers [Calb (+ vehicle) = 0.12 microliters/min; Calb (+10 nM alpha-thrombin) = 0.47 microliters/min; Calb (+5 mg/ml trypsin) = 1.29 microliters/min]. Transfer of EBA was linear with time in both the endothelial cell monolayer model and the perfused lung. EBA was a sensitive marker for early edema in the perfused lung (before detectable weight gain) as well as for severe edema in the oxidant-injured lung (marked EBA accumulation in lavage fluid) and was a more specific marker for protein transfer than lavage fluid protein. EBA transfer is a convenient, reproducible, and accurate means to assess alterations in vascular permeability.     

Importance of vasoconstriction in lipid mediator-induced pulmonary edema

Lipid mediators of inflammation cause pulmonary edema, yet it is unclear to what degree hemodynamic alterations or increased vascular permeability contribute to lung edema formation. The isolated rat lung preparation was used to examine the effect of leukotriene C4 (LTC4) and platelet-activating factor (PAF) on pulmonary arterial pressure (Ppa), lung microvascular pressure (Pmv), lung wet-to-dry weight ratio, and the 125I-albumin escape index. We first defined the response of the isolated rat lung perfused with protein-free salt solution to hydrodynamic stress by raising the lung outflow pressure. Sustained elevation of the lung outflow pressure less than 5.5 cmH2O (4.01 mmHg) caused a negligible increase in Ppa and wet-to-dry lung weight ratio. Elevation of outflow pressures greater than 7.5 cmH2O (5.4 mmHg) increased the vascular albumin escape index more than the lung wet-to-dry weight ratio. Dibutyryl adenosine 3',5'-cyclic monophosphate (db-cAMP) inhibited the increase in albumin escape index because of increased lung outflow pressure, suggesting perhaps a pressure-independent microvascular membrane effect of db-cAMP. Both LTC4 (2-micrograms bolus) and PAF (2-2,000 ng/ml perfusate) increased the albumin escape index in association with increases in Ppa and Pmv. Because the increased albumin escape index after LTC4 or PAF injection was largely accounted for by the increased vascular pressures and because db-cAMP and papaverine inhibited the rise in vascular pressures and in the albumin escape index, we conclude that vasoconstriction is an important contributor to LTC4- and PAF-induced edema formation in rat lungs.     

Time course of changes in lung permeability and edema in the rat exposed to 100% oxygen

Rats were exposed to 100% oxygen for up to 60 h to determine early changes in lung permeability leading to the development of pulmonary edema. The time course of development of increased solute flux was assessed by the clearance of 99mTc-labeled diethylenetriamine pentaacetate (99mTc-DTPA) from the lung and the accumulation of 125I-labeled albumin (125I-albumin) in the lung. These end points were related to the development of pulmonary edema by the measurement of the wet-to-dry weight ratio of the lung and the weight of fluid in the pleural cavity. No significant changes occurred until 48 h of hyperoxia, when sharp increases in both indexes of lung permeability and wet-to-dry weight ratio occurred. By 60 h of exposure, pleural effusions had developed. The volume of this effusion was significantly correlated to both 99mTc-DTPA clearance and 125I-albumin flux.     

Canine pulmonary filtration coefficient calculated from optical, radioisotope, and weight measurements.

Three independent methods were used to estimate filtration coefficient (Kf) in isolated dog lungs perfused with low-hematocrit (Hct) blood. Pulmonary vascular pressure was increased by 12-23 cmH2O to induce fluid filtration. Average Kf (ml.min-1 x cmH2O-1 x 100 g dry wt-1) for six lungs was 0.26 +/- 0.05 (SE) with use of equations describing conservation of optically measured protein labeled with indocyanine green. Good agreement was found when a simplified version of the multiequation theory was applied to the data (0.24 +/- 0.05). Both optical estimates were lower than those predicted by constant slope (0.55 +/- 0.07) or extrapolation (1.20 +/- 0.15) techniques, which are based on changes in total lung weight. Subsequent studies in five dog lungs investigated whether the higher Kf from weight analyses could be caused by prolonged pulmonary vascular filling. We found that 51Cr-labeled red blood cells (RBCs), monitored over the lung, continued to accumulate for 30 min after vascular pressure elevations of 9-16 cmH2O.Kf was determined by subtracting computed vascular filling from total weight change (0.28 +/- 0.06) and by perfusate Hct changes determined from radiolabeled RBCs (0.23 +/- 0.04). These values were similar to those obtained from analysis of optical data with the complete model (0.30 +/- 0.06), the simplified version (0.26 +/- 0.05), and from optically determined perfusate Hct (0.16 +/- 0.03). However, constant slope (0.47 +/- 0.04) and extrapolation (0.57 +/- 0.07) computations of Kf were higher than estimates from the other methods. Our studies indicate that prolonged blood volume changes may accompany vascular pressure elevations and produce overestimates of Kf with standard weight measurement techniques. However, Kf computed from optical measurements is independent of pulmonary blood volume changes.     

Sites of leakage in three models of acute lung injury

Fluid leaking from arterial and venous extra-alveolar vessels (EAV's) may account for up to 60% of the total transvascular fluid flux when edema occurs in the setting of normal vascular permeability. We determined if the permeability and relative contribution of EAV's was altered after inducing acute lung injury in rabbits by administering oleic acid (0.1 ml/kg) into the pulmonary artery, HCl (5 ml/kg of 0.1 N) into the trachea, or air emboli (0.03 ml.kg-1.min-1) into the right atrium for 90 min. Subsequently, the lungs were excised and continuously weighed while they were maintained in a warmed, humidified chamber with alveolar and pulmonary vascular pressures controlled and the lungs either ventilated or distended with 5% CO2 in air. The vascular system was filled with autologous blood and saline (1:1) to which papaverine (0.1 mg/ml) was added to inhibit vasospasm. Vascular pressures were referenced to the lung base. After a transient hydrostatic stress to maximize recruitment, vascular pressures were set at 5 cmH2O, and lungs were allowed to become isogravimetric (30-60 min). A fluid filtration coefficient (Kf) was determined by the use of a modification of the method of Drake and colleagues [Am. J. Physiol. 234 (Heart Circ. Physiol. 3): H266-H274, 1978]. EAV's were isolated by zoning techniques. In control preparations arterial and venous EAV's accounted for 26% (n = 9) and 38% (n = 11) of the total leakage, respectively. In all three models Kf increased two- to fourfold when the lungs were in zone 3 (alveolar vessels and arterial and venous EAV's contributing to the leakage).(ABSTRACT TRUNCATED AT 250 WORDS)     

Protective effect of alpha-human atrial natriuretic polypeptide (alpha-hANP) on chemical-induced pulmonary edema

It has been established that alpha-hANP, the newly discovered peptide extracted from human cardiac atria, has potent natriuretic and hypotensive actions. Our present investigation is the first to demonstrate that alpha-hANP is capable of protecting against pulmonary edema caused by various chemicals, using isolated perfused guinea pig lung system. Lungs were perfused via pulmonary artery with Krebs-Ringer bicarbonate buffer at 5.0 ml/min, and wet weight of lungs and perfusion pressure of pulmonary artery (Pa) were monitored. Bolus injection of Triton-X or CHAPS into cannulated pulmonary artery produced edema as indicated by a massive increase in wet weight and a slight increase in Pa. Constant infusion of alpha-hANP through pulmonary artery at 200 ng/ml was effective in causing decrease in wet weight of lung. Perfusion of lung with paraquat or PGF2 alpha, and repeated bolus injection of arachidonic acid or PGE2 caused elevation in both wet weight of lung and Pa. The treatment with alpha-hANP similar to that described above also protected against edema caused by paraquat or arachidonic acid. Bolus administration of epinephrine induced a slight increase in wet weight and Pa, and alpha-hANP was effective in decreasing the elevated lung wet weight and Pa of lungs. Infusion or bolus administration of alpha-hANP into control lungs increased cGMP level in outflow perfusate as well as in lung tissue significantly. In lungs with edema which were induced by Triton-X or paraquat, there was a slight increase in cGMP level in Triton-X treated and no increase in paraquat treated lung tissues. In either cases, was there any increase in cGMP level in perfusate. The specific binding study of [125I]alpha-hANP revealed that the lack of increase in cGMP was not due to a loss of receptor in Triton-X or paraquat treated lungs. Thus our study demonstrated that alpha-hANP had a direct anti-edematic action(s) in lung which was not secondary to the systemic natriuretic and/or hypotensive action(s).     

Spontaneous injury in isolated sheep lungs: role of perfusate leukocytes and platelets

Perfusion of isolated sheep lungs with blood causes spontaneous edema and hypertension preceded by decreases in perfusate concentrations of leukocytes (WBC) and platelets (PLT). To determine whether these decreases were caused by pulmonary sequestration, we continuously measured blood flow and collected pulmonary arterial and left atrial blood for cell concentration measurements in six lungs early in perfusion. Significant sequestration occurred in the lung, but not in the extracorporeal circuit. To determine the contribution of these cells to spontaneous injury in this model, lungs perfused in situ with a constant flow (100 ml.kg-1.min-1) of homologous leukopenic (WBC = 540 mm-3, n = 8) or thrombocytopenic blood (PLT = 10,000 mm-3, n = 6) were compared with control lungs perfused with untreated homologous blood (WBC = 5,320, PLT = 422,000, n = 8). Perfusion of control lungs caused a rapid fall in WBC and PLT followed by transient increases in pulmonary arterial pressure, lung lymph flow, and perfusate concentrations of 6-ketoprostaglandin F1 alpha and thromboxane B2. The negative value of reservoir weight (delta W) was measured as an index of fluid entry into the lung extravascular space during perfusion. delta W increased rapidly for 60 min and then more gradually to 242 g at 180 min. This was accompanied by a rise in the lymph-to-plasma oncotic pressure ratio (pi L/pi P). Relative to control, leukopenic perfusion decreased the ratio of wet weight to dry weight, the intra- plus extravascular blood weight, and the incidence of bloody lymph. Thrombocytopenic perfusion increased lung lymph flow and the rate of delta W, decreased pi L/pi P and perfusate thromboxane B2, and delayed the peak pulmonary arterial pressure. These results suggest that perfusate leukocytes sequestered in the lung and contributed to hemorrhage but were not necessary for hypertension and edema. Platelets were an important source of thromboxane but protected against edema by an unknown mechanism.     

Promethazine or DPPD pretreatment attenuates oleic acid-induced injury in isolated canine lungs

Oleic acid causes pulmonary edema by increasing capillary endothelial permeability, although the mechanism of this action is uncertain. We tested the hypothesis that the damage is an oxidant injury initiated by oleic acid, using isolated blood-perfused canine lung lobes. The lobes were dilated with papaverine and perfused in zone III with a constant airway pressure of 3 cmH2O. Changes in isogravimetric capillary pressure (Pc,i) and capillary filtration coefficient (Kf,C) were used as indices of alterations in microvascular permeability in lungs treated with silicone fluid (n = 3), oleic acid (n = 11), oleic acid after pretreatment with the antioxidants promethazine HCl (n = 11) or N,N'-diphenyl-p-phenylenediamine (DPPD; n = 4), or oleic acid following pretreatment with methylprednisolone (n = 4). Kf,C averaged 0.21 +/- 0.02 ml X min-1 X cmH2O-1 X 100 g-1 in control and increased to 0.55 +/- 0.05 and 0.47 +/- 0.05 when measured 20 and 180 min after the administration of oleic acid. When oleic acid was infused into lungs pretreated with promethazine, Kf,C increased to only 0.38 +/- 0.05 ml X min-1 X cmH2O-1 X 100 g-1 after 20 min and had returned to control levels by 180 min. Pretreatment with DPPD, but not methylprednisolone, similarly attenuated the increase in Kf,C following oleic acid. Silicone fluid had no effect on Kf,C. That oleic acid increases vascular permeability was also evidenced by a fall (P less than 0.05) in Pc,i from control when measured at 180 min in every group.(ABSTRACT TRUNCATED AT 250 WORDS)     

Venous occlusion pressure and vascular permeability in the dog lung after air embolization

Pulmonary edema has frequently been associated with air embolization of the lung. In the present study the hemodynamic effects of air emboli (AE) were studied in the isolated mechanically ventilated canine right lower lung lobe (RLL), pump perfused at a constant blood flow. Air was infused via the pulmonary artery (n = 7) at 0.6 ml/min until pulmonary arterial pressure (Pa) rose 250%. While Pa rose from 12.4 +/- 0.6 to 44.6 +/- 2.0 (SE) cmH2O (P less than 0.05), venous occlusion pressure remained constant (7.0 +/- 0.5 to 6.8 +/- 0.6 cmH2O; P greater than 0.05). Lobar vascular resistance (RT) increased from 2.8 +/- 0.3 to 12.1 +/- 0.2 Torr.ml-1.min.10(-2) (P less than 0.05), whereas the venous occlusion technique used to determine the segmental distribution of vascular resistance indicated the increase in RT was confined to vessels upstream to the veins. Control lobes (n = 7) administered saline at a similar rate showed no significant hemodynamic changes. As an index of microvascular injury the pulmonary filtration coefficient (Kf) was obtained by sequential elevations of lobar vascular pressures. The Kf was 0.11 +/- 0.01 and 0.07 +/- 0.01 ml.min-1.Torr-1.100 g RLL-1 in AE and control lobes, respectively (P less than 0.05). Despite a higher Kf in AE lobes, total lobe weight gains did not differ and airway fluid was not seen in the AE group. Although air embolization caused an increase in upstream resistance and vascular permeability, venous occlusion pressure did not increase, and marked edema did not occur.     

Edema from cyclooxygenase products of endogenous arachidonic acid in isolated lung

We infused A23187, a calcium ionophore, into the pulmonary circulation of dextran-salt-perfused isolated rabbit lungs to release endogenous arachidonic acid. This led to elevations in pulmonary arterial pressure and to pulmonary edema as measured by extravascular wet-to-dry weight ratios. The increase in pressure and edema was prevented by indomethacin, a cyclooxygenase enzyme inhibitor, and by 1-benzylimidazole, a selective inhibitor of thromboxane (Tx) A2 synthesis. Transvascular flux of 125I-albumin from vascular to extravascular spaces of the lung was not elevated by A23187 but was elevated by infusion of oleic acid, an agent known to produce permeability pulmonary edema. We confirmed that A23187 leads to elevations in cyclooxygenase products and that indomethacin and 1-benzylimidazole inhibit synthesis of all cyclooxygenase products and TxA2, respectively, by measuring perfusate levels of prostaglandin (PG) I2 as 6-ketoprostaglandin F1 alpha, PGE2, and PGF2 alpha and TxA2 as TxB2. We conclude that release of endogenous pulmonary arachidonic acid can lead to pulmonary edema from conversion of such arachidonic acid to cyclooxygenase products, most notably TxA2. This edema was most likely from a net hydrostatic accumulation of extravascular lung water with an unchanged permeability of the vascular space, since an index of permeability-surface area product (i.e., transvascular albumin flux) was not increased.     

Mechanisms of increased pulmonary microvascular permeability induced by FMLP in isolated rabbit lungs.

We observed that the chemotactic peptide N-formyl-L-methionyl-L-leucyl-L- phenylalanine (FMLP) induced pulmonary edema when polymorphonuclear leukocytes (PMNs) were added to isolated constant-flow buffer-perfused rabbit lungs. This study was designed to test the hypothesis that PMNs activated by FMLP induced lung injury by the modulation of reactive oxygen species (ROS), cyclooxygenase products, or cysteinyl leukotrienes (LTs). Addition of FMLP alone did not increase microvascular permeability (Kf). When PMNs were added to the isolated lung, FMLP caused an 80% increase in Kf. Wet-to-dry weight ratio was also significantly increased with PMNs + FMLP compared with FMLP only. There was a significant positive correlation between total myeloperoxidase activity in lung tissue and Kf values after FMLP (30 min). Pretreatment with two dissimilar cyclooxygenase inhibitors, meclofenamate or ibuprofen, had no effect on the PMN + FMLP-induced increase in Kf. However, the ROS inhibitor catalase and the nonantioxidant LT synthesis blocker MK 886 inhibited the PMN + FMLP increase in Kf. Perfusate levels of LTs (LTC4, -D4, and -E4) were significantly increased from baseline values 30 min after FMLP. Both MK 886 and catalase suppressed the elevation of LTs after PMN + FMLP. These results indicate that FMLP increased a pulmonary microvascular permeability in isolated buffer-perfused rabbit lungs that is PMN dependent and mediated by LT produced possibly by a result of ROS production.     

Effects of arachidonic acid and prostaglandin F2 alpha in isolated rat lungs

Isolated rat lungs were ventilated and perfused by saline-Ficoll perfusate at a constant flow. The baseline perfusion pressure (PAP) correlated with the concentration of 6-keto-PGF1 alpha the stable metabolite of PGI2 (r = 0.83) and with the 6-keto-PGF1 alpha/TXB2 ratio (r = 0.82). A bolus of 10 micrograms exogenous arachidonic acid (AA) injected into the arterial cannula of the isolated lungs caused significant decrease in pulmonary vascular resistance (PVR) which was followed by a progressive increase of PVR and edema formation. Changes in perfusion pressure induced by AA injection also correlated with concentrations of the stable metabolites (6-keto-PGF1 alpha: r = -0.77, TxB2: -0.76), and their ratio: (6-keto-PGF1 alpha/TXB2: r = -0.73). Injection of 10 and 100 micrograms of PGF2 alpha into the pulmonary artery stimulated the dose-dependent production of TXB2 and 6-keto-PGF1 alpha. No significant correlations were found between the perfusion pressure (PAP) which was increased by the PGF2 alpha and the concentrations of the former stable metabolites. The results show that AA has a biphasic effect on the isolated lung vasculature even in low dose. The most potent vasoactive metabolites of cyclooxygenase, prostacyclin and thromboxane A2 influence substantially not only the basal but also the increased tone of the pulmonary vessels.     

Actions of lipoxygenase metabolites in isolated rat lungs

Leukotrienes constrict smooth muscle and could be important for the regulation of the pulmonary circulation. We examined the production and action of lipoxygenase metabolites in isolated lungs, where we controlled the perfusing fluid used. Arachidonate injected into isolated rat lungs perfused with cell- and protein-free physiological salt solution caused a transient pressor response. Following indomethacin, arachidonate caused a delayed slow pressure rise followed by edema. The lung effluent contracted the guinea pig ileum. High-pressure liquid chromatography (HPLC) analysis of the perfusate demonstrated the presence of leukotrienes (LTC4 and LTD4). Diethylcarbamazine, a leukotriene synthesis inhibitor, prevented the slow pressure rise and edema seen after indomethacin plus arachidonate. In lungs perfused with cell- and protein-free physiological salt solution, LTC4, but not LTD4, caused a transient pressure rise followed by a sustained pressure rise. The sustained rise was abolished by a leukotriene-receptor blocker (FPL 55712) but not by indomethacin. In blood-perfused lungs, LTC4 caused only the transient pressure rise that was not blocked by FPL 55712. In lungs perfused with physiological salt solution containing albumin, LTC4 had no effect. We concluded that 1) perfused nonsensitized rat lungs produced LTC4 and LTD4; 2) LTC4 may be a major pulmonary vasoconstrictor; and 3) albumin binding limits the pressor effect of LTC4.     

Cyclooxygenase inhibition prevents PMA-induced increases in lung vascular permeability

The effect of cyclooxygenase inhibition in phorbol myristate acetate (PMA)-induced acute lung injury was studied in isolated constant-flow blood-perfused rabbit lungs. PMA caused a 51% increase in pulmonary arterial pressure (localized in the arterial and middle segments as measured by vascular occlusion pressures), a 71% increase in microvascular permeability (measured by the microvascular fluid filtration coefficient, Kf), and a nearly threefold increase in perfusate thromboxane (Tx) B2 levels. Cyclooxygenase inhibition with three chemically dissimilar inhibitors, indomethacin (10(-7) and 10(-6) M), meclofenamate (10(-6) M), and ibuprofen (10(-5) M), prevented the Kf increase without affecting the pulmonary arterial pressure increase or resistance distribution changes after PMA administration. The specific role of TxA2 was investigated by pretreatment with OKY-046, a specific Tx synthase inhibitor, or infusion of SQ 29548, a TxA2 receptor antagonist; both compounds failed to protect against either the PMA-induced permeability or the vascular resistance increase. These results indicate that cyclooxygenase-mediated products of arachidonic acid other than TxA2 mediate the PMA-induced permeability increase but not the hypertension.