Influence of PDE5 inhibitor on MRI measurement of clitoral volume response in women with FSAD: a feasibility study of a potential technique for evaluating drug response.

The purpose of this study was to determine if magnetic resonance imaging (MRI) could quantify a difference in clitoral response following administration of a vasoactive medication, in 12 women with female sexual arousal disorder (FSAD). Subjects were entered into a double-blind, randomized two-way crossover study of sildenafil 50 mg vs placebo administered 1 h prior to genital MRI. Each subject underwent two MR studies, performed while subjects viewed alternating segments of nonerotic and erotic video. MR images were analyzed for change in clitoral volume during each session. The mean change in clitoral volume for the entire group was higher in the sildenafil MRI session (1282 mm(3)) compared with placebo (849 mm(3)) but did not reach statistical significance (P=0.064). Comparison using analysis of variance between the two sessions for each individual subject revealed a significant increase in clitoral volume following sildenafil compared with placebo in 6 of 12 subjects, no significant change in either imaging session in three subjects and in three subjects, there was a robust clitoral response in both MR sessions. In conclusion, MR measurements of clitoral volume can provide an objective measure of engorgement change following a vasoactive medication in women with FSAD.

Traceless, self-cleaving solid- and solution-phase parallel synthesis of 3,4,7-trisubstituted 3,4-dihydroquinoxalin-2-ones.

This article describes a new methodology for the parallel synthesis of 3,4-dihydroquinoxalin-2-ones containing three points of diversity. The synthesis begins with commercially available resin-bound alpha-amino acids as the source of the first diversity element and employs a combination of solid- and solution-phase chemistry to introduce the other two. The key step is an intramolecular cyclization and simultaneous traceless cleavage from the solid support to give a disubstituted 3,4-dihydroquinoxalin-2-one. The third substituent is introduced in solution by N-alkylation of the aniline nitrogen using a scavenger resin to dispose of excess reagent. All the reactions in the sequence take place at room temperature without the need to use strong acids or to maintain an inert atmosphere, thereby preserving the chiral integrity of the starting alpha-amino acid and facilitating the generation of libraries in a high-throughput parallel format.

Modulation by pentobarbital of neutrophil responses to inhaled E. coli endotoxin in sheep: role of lung epithelium.

Neutrophils (PMNs) are implicated in the pathogenesis of acute respiratory distress syndrome (ARDS). The role of the epithelium in the modulation of PMN migration within the lungs was examined. Epithelial integrity and PMN concentrations in the lung air spaces and lymph were measured in sheep anaesthetized with either halothane (1-2.5%) or intravenous pentobarbital (12+/-4 mg x kg(-1) x h(-1)). Ventilation with an aerosol containing 25 mg Escherichia Coli endotoxin (lipopolysaccharide; LPS) effected neutrophil recruitment to the air spaces. Lymphatic clearance of aerosolized 99mTc-DTPA provided an index of epithelial integrity. Three hours after the deposition of LPS, the lung lining fluid of sheep anaesthetized with halothane (n=7) had 4.9+/-3.2x10(6) PMN.mL(-1), but the lung lymph had almost no PMNs (3+/-8%). Sheep anaesthetized with pentobarbital (n=6) had fewer PMNs in the air spaces (2.4+/-1.2X10(6) mL(-1)) and more PMNs in the lung lymph (30+/-20%). Control sheep (n=5) that received no LPS had almost no PMNs in the airspaces or lung lymph, regardless of the anaesthesia. Three additional sheep that remained awake after receiving LPS also had no PMNs in the lung lymph. The PMN fraction in the lung lymph correlated well with the extra-alveolar epithelial permeability measured by lymphatic clearance of aerosolized diethylenetriamine penta-acetic acid (r=0.81, p<0.001). Aerosolized lipopolysaccharide recruits neutrophils into the lungs of sheep, but they appear to remain in the airspaces unless extra-alveolar permeability is increased by agents such as pentobarbital.

Virulence factors from Pseudomonas aeruginosa increase lung epithelial permeability.

Pseudomonas aeruginosa infection frequently complicates lung injury and can be fatal in immunocompromised or debilitated individuals. Previous studies from our laboratory indicate that elastase from P. aeruginosa increases epithelial permeability by disrupting tight junctions between epithelial cells. Because the inflammatory reaction of the host is a prominent feature of bacterial infection, we reasoned that additional virulence factors from this organism could extend and augment the initial pulmonary injury by prompting accumulation of neutrophils. To test this hypothesis, we compared responses of guinea pigs to aerosols of elastase (PE) and lipopolysaccharide (LPS) from P. aeruginosa. After each treatment, we measured epithelial permeability and accumulation of neutrophils, interleukin 8 (IL-8), and beta-glucuronidase in epithelial lining fluid (ELF). We found that PE increased epithelial permeability, as measured by both the clearance of aerosolized radiolabeled albumin from the air spaces and the concentration of plasma albumin in epithelial lining fluid, but it was less effective than LPS at recruiting neutrophils into the lungs. In contrast, LPS had no significant effect on epithelium, but it increased the concentration of neutrophils, IL-8, and beta-glucuronidase in ELF. Increased epithelial permeability induced by PE does not cause lung inflammation, but it may facilitate the LPS-induced influx of neutrophils.

Neutrophil influx and migration in rabbit airways in response to staphylococcal enterotoxin-A.

Single-cycle lavages performed on the lungs of rabbits 7 hours after they received intravenous staphylococcal enterotoxin-A (SEA) showed that most of the neutrophils were found in the second of 5 fractions of fluid removed from the lungs. To determine whether this finding reflected an accumulation of neutrophils in the airways versus the alveolar regions of the lungs, we counted the neutrophils in histologic sections (20 microns) and estimated the concentration of neutrophils in the epithelial lining fluid of the airways versus the alveoli. Comparisons of the morphometric measurements and the lavage data showed a strong correlation between the 2 methods (r = .90, P < .001), but the lavage technique underestimated the morphometric value by a factor of 7. The histologic examination confirmed that the concentration of neutrophils in the airways was 3-4 times higher than in the alveoli. There were no differences between the alveoli and the airways with respect to the concentration of endogenous proteins and the chemoattractant, interleukin (IL)-8. Lavages performed in additional rabbits 4 and 18 hours after injection of SEA showed that neutrophils most likely entered the alveolar region in response to SEA and migrated into the airways. The single-cycle technique provides a reliable measure of the relative concentration of neutrophils in the airways versus the alveoli.

Clearance of proteins from the air spaces following cardiogenic edema in sheep.

Studies on the clearance of instilled fluid and protein into the lungs of sheep show that fluid initially clears rapidly from the lungs resulting in an increase in an increase in air space protein concentrations. This is followed by a slower monoexponential clearance of proteins during the next few days. To determine whether the clearance of edema fluid follows the same time course as instilled fluid, alveolar edema was induced in 11 sheep by inflating a balloon in the left atrium for 2 h to increase left atrial pressure 35-40 cm H2O. Protein concentrations in the epithelial lining fluid (ELF) were monitored by performing single-cycle lavages immediately after deflation of the left atrial balloon, and again 3, 21, 48, and 96 h later. During the first 3 h of recovery, 44% of the fluid cleared and ELF protein concentrations rose from 7 +/- 2 to 19 +/- 6 mg/mL in the 7 sheep that recovered well. The ELF protein concentration in these sheep remained elevated for 48 h and then cleared at rates similar to that seen following instillation of proteins. We conclude that insights about mechanisms of clearance of edema fluid obtained from studies of clearance of instilled fluid are valid in cardiogenic edema, but the time course of the clearance differs between the two models.

Peptide inhibitor of interleukin-8 (IL-8) reduces staphylococcal enterotoxin-A (SEA) induced neutrophil trafficking to the lung.

Staphylococcal enterotoxin A (SEA) is a superantigen, produced by some strains of Staphylococcus aureus (S. aureus), which can cause a variety of clinical manifestations ranging from food poisoning to shock. SEA can also stimulate human alveolar macrophages to produce interleukin-8 (IL-8), a member of the alpha-chemokine subfamily that activates and is chemotactic for neutrophils. In these studies we showed that in rabbits, intravenous SEA significantly decreased the circulating white blood cell population from a baseline value of 6409 +/- 2027 x 10(3) cells/ml to 1943 +/- 862 x 10(3) cells/ml in 7 h. There was a concommitent increase in IL-8 in the circulating plasma (baseline: 60 +/- 34 pg/ml, 7 h post SEA: 109 +/- 64 pg/ml). The increase in circulating IL-8 was accompanied by a much greater increase in the IL-8 concentration of the epithelial lining fluid (ELF) where the IL-8 increased from 0.05 +/- 0.08 ng/ml (control) to 13.8 +/- 9.3 ng/ml (SEA treated). The increase in IL-8 concentration in the alveolar spaces was paralleled by an increase in both the percentage of neutrophils (1.4 +/- 0.9% (control) to 26.0 +/- 10.8% (SEA treated)) and total number of neutrophils (0.04 +/- 0.02 x 10(6)/ml (control) to 4.8 +/- 3.3 10(6)/ml (SEA treated) in the airspaces, and the numbers of neutrophils in the ELF correlated with the IL-8 concentration r = 0.62, p = 0.006). When antileukinate, a hexapeptide which inhibits the binding of IL-8 to neutrophils, was administered to animals receiving SEA, the IL-8 concentration in the ELF (14.8 +/- 10.7 ng/ml) was not significantly different from the concentration of IL-8 in those animals receiving SEA alone). However, both the percentage of neutrophils (9.5 +/- 3.2%), and the total number of neutrophils (1.3 +/- 1.0 x 10(6)/ml) in the ELF following SEA and antileukinate administration was significantly lower than in animals which only received SEA (p < 0.05). The findings suggest that SEA released into the circulation during a Staphylococcal infection can cause an inflammatory reaction in the lung. Since this reaction is at least partially mediated by IL-8, antileukinate may have pharmacologic potential in reducing the inflammatory reaction.

Differential effects of salmeterol on lung endothelial and epithelial leakage in sheep.

Salmeterol has been shown to prevent the influx of proteins into the air spaces of lungs of guinea pigs given intravenous histamine. To determine whether the salmeterol acts to stabilize the epithelial or endothelial barrier, we ventilated anesthetized sheep with aerosolized salmeterol before infusing histamine intravenously at a rate of 4 micrograms.kg-1.min-1 for 3 h. Changes in endothelial permeability were assessed by measuring the flow of lymph and proteins from the lungs. The influx of proteins into the air spaces was detected by performing single-cycle lavages to measure the concentration of circulating 125I-albumin in the epithelial lining fluid. Intravenous histamine increased the lymph flow to 13.2 +/- 6.8 ml/h compared with the control value of 5.6 +/- 2.8 ml/h (P < 0.05). Histamine also increased the concentration of 125I-albumin in the epithelial lining fluid from 1.8 +/- 0.9 to 8.5 +/- 2.5% of the plasma concentration (P < 0.01) and the postmortem lung water volume from 3.5 +/- 0.5 to 5.0 +/- 0.8 mg/g dry lung wt (P < 0.05). Pretreatment with 2.5 mg of aerosolized salmeterol prevented the influx of proteins into the air spaces and the increase in the postmortem lung water volume but it also increased the lung lymph flow even further to 20.0 +/- 5.6 ml/h (P < 0.05), increased the lymph-to-plasma protein ratio from 0.77 to 0.91, and tripled the increase in alveolar-arterial oxygen gradient caused by histamine alone. Pretreatment with 2.5 mg of intravenous salmeterol had essentially the same effect as salmeterol administered by aerosol. We conclude that salmeterol decreases lung epithelial permeability but increases lung endothelial permeability due to intravenous histamine in sheep.

Single-cycle bronchoalveolar lavage to determine solute concentrations in epithelial lining fluid.

An accurate and reproducible measure of solute concentration of lung epithelial lining fluid (ELF) by bronchoalveolar lavage would be valuable in lung research and in patient care. Measurements of the albumin/total protein ratio in a previously proposed rewash lavage procedure showed that albumin enters the lavage fluid. Therefore, the rewash lavage may measure ELF volume accurately, but it overestimates ELF protein concentration (PELF). To avoid problems of solute exchange, we examined five sequential fractions of lavage fluid obtained from sheep after a single 60-ml lavage containing a 99mTcO4- tracer. Assays of albumin, 99mTcO4-, total protein, and endogenous urea concentrations allowed calculation of PELF from each fraction. PELF was 8 +/- 4 mg/ml when calculated from dilution of either endogenous urea or 99mTcO4- in fractions collected after the first 15 to 20 ml. Paired lavages provided a reproducible measure of PELF (SD, 1.2 mg/ml) that was unaffected by any solute exchange that occurred during the 40-s procedure. Accuracy was verified by comparisons of lung lymph and ELF protein concentrations during high pressure lung edema in anesthetized sheep. The single-cycle lavage procedure is an accurate and reproducible procedure for measuring PELF.

Albumin fraction and measurement of total protein concentration.

The standard curve of a typical colorimetric assay for total protein is often nonlinear and dependent on the albumin fraction of the protein standard. We developed a simple mathematical transformation to make the standard curve linear and a computational method to correct for differences in albumin concentrations among the samples. This method uses data from total protein assays on two sets of standards (albumin and gamma globulin) and provides accurate measures of total protein over the full range of albumin fractions. Comparison of this two-standard method with the a method that uses only albumin as a standard shows that this method prevents physiologically significant overestimations in total protein concentration and calculated protein osmotic pressure differences in the lungs.

Clearance of 99mTc-labeled albumin from lungs in anesthetized guinea pigs.

Gamma imaging was used to measure the rate of clearance of aerosolized 99mTc-human serum albumin (HSA) from the lungs of control guinea pigs and guinea pigs that received increased lung inflation or lung injury. Anesthetized guinea pigs were ventilated for 6 min with an aerosol of HSA and the radioactivity in the chest was monitored for 2 h with a gamma camera to determine whether the clearance rate would be a reliable assessment of lung epithelial permeability. Increased lung volumes were effected by application of 5 or 7 cm H2O positive end-expired pressure (5-PEEP and 7-PEEP, respectively). Lung injury was induced either by intravenous oleic acid (OA, 27-73 microliters/kg) or inhalation of nitrogen dioxide (NO2, 80-100 ppm) for 2 h. Postmortem extravascular lung water volume (EVLW) provided an assessment of the degree of lung injury. Tracer clearance rates in animals receiving 5 or 7 cm H2O PEEP were not significantly different from controls (K = 0.15 +/- 0.05 and 0.24 +/- 0.10 vs 0.12 +/- 0.03%/min, respectively, p > .05). Animals exposed to NO2 had faster tracer clearance rates (K = 0.33 +/- 0.21%/min, p < .05) and higher EVLW (5.8 +/- 3.0 vs 3.7 +/- 0.2 mL/g dry lung, p < .05) than controls. Clearance rates of HSA from the lungs of NO2-exposed guinea pigs correlated well with injury as assessed by EVLW (r = .93, p < .01). Clearance rates of HSA and EVLW in animals receiving oleic acid were significantly higher than controls and the group receiving 5 cm H2O PEEP (K = 0.58 +/- 0.41%/min, EVLW = 8.1 +/- 0.8 mL/g dry lung tissue, p < .05), but there was no correlation between these parameters in this injury model. It is concluded that imaging of the disappearance of radiolabeled HSA in the guinea pig can be a useful index of lung epithelial permeability, but this technique is limited to certain models of lung injury.

Aerosolized Pseudomonas elastase and lung fluid balance in anesthetized sheep.

The role of the lung epithelium in lung fluid balance was studied by ventilating anesthetized sheep with an aerosol of 20 mg of elastase from Pseudomonas aeruginosa (Ps. elastase) to increase lung epithelial permeability without affecting lung endothelial permeability or lung vascular pressures. Ps. elastase had no effect on the lung vascular pressures, the alveolar-arterial PO2 gradient (A-aPO2), the flow or protein concentration of the lung lymph, or the postmortem water volume of the lungs. The morphological alveolar flooding score in these sheep was 2.5 times the control level, but this was only marginally significant. Elevation of the left atrial pressure by 20 cmH2O alone increased the postmortem lung water volume but had no effect on A-aPO2, the alveolar flooding score, or the lung epithelial permeability assessed by the clearance of 99mTc-labeled human serum albumin. Addition of aerosolized Ps. elastase to these sheep had no effect on the total lung water volume, but it caused a redistribution of water into the air spaces, as evidenced by significant increases in the alveolar flooding score and A-aPO2 (P less than 0.01). Elevation of the left atrial pressure by 40 cmH2O without elastase caused the same response as elevation of the left atrial pressure by 20 cmH2O with elastase, except the higher pressure caused a greater increase in the total lung water volume. We conclude that alteration of the integrity of the lung epithelium with aerosolized Ps. elastase causes a redistribution of lung water into the alveoli without affecting the total lung water volume.(ABSTRACT TRUNCATED AT 250 WORDS)

Permeability: theory vs. practice in lung research.

An increase in permeability of the pulmonary endothelium or epithelium to proteins and other macromolecules is an important part of many types of lung injury. Assessment of lung alveolar-capillary membrane integrity has been the focus of many investigations, but no single method is best for all conditions. Methods using in vitro preparations, such as cell monolayers, provide the most accurate estimates of biophysical parameters defined from membrane theory, but these models may not accurately represent the in vivo condition. Methods using in vivo preparations, such as indicator dilution techniques and the sheep lung-lymph preparation, are more accurate representations of the clinical condition, but considerable assumptions must be made to estimate the parameters of permeability. The most productive studies of lung permeability make use of at least two methods carefully selected according to their strengths and weaknesses. The challenge is to select the right tools for the job and to interpret the results in light of the weaknesses of the methods used. This will lead to a better understanding of the role of altered permeability in the pathophysiology of lung injury.

Rewash bronchoalveolar lavage.

A significant limitation of standard bronchoalveolar lavage (BAL) technique is the inability to measure or calculate epithelial lining fluid (ELF) volume and, therefore, in vivo concentrations of substances in the ELF. We evaluated a new rewash BAL procedure with the radiolabeled tracer technetium pertechnetate (99mTcO4-) that theoretically should be immune to even exaggerated fluid shifts during BAL. To test this theory, we measured ELF volume in control sheep using isosmotic (280 mosm/L) hypoosmotic (140 mosm/L) and hyperosmotic (570 mosm/L) BAL solutions to induce exaggerated fluid shifts during the lavage procedure. The mean ELF volume of the lavaged lung segment was not significantly different for the three solutions (isosmotic, 1.7 +/- 0.8 ml; hypoosmotic, 1.1 +/- 1.2 ml; hyperosmotic, 2.1 +/- 1.6 ml). The slope of the 99mTcO4- disappearance curve, however, was significantly steeper for the hyperosmotic solution (-0.40 +/- 0.04%/min) compared with the other solutions (isosmotic, -0.14 +/- .01%/min; hypoosmotic, -0.12 +/- 0.07%/min). Calculation of ELF volume using sodium as an endogenous tracer gave consistently smaller values with each of the mannitol solutions (isosmotic, 0.21 +/- 0.30 ml; hypoosmotic, 0.02 +/- 0.03 ml; hyperosmotic, 0.18 +/- 0.18 ml). The failure of sodium to provide accurate estimates of the ELF volume may be due to complicated sodium movement in the lung and errors in our assumption of the initial concentration of sodium in the ELF fluid. We conclude that the rewash BAL technique with 99mTcO4- gives values of ELF volume that are not significantly affected by even exaggeration of the fluid flux that invariably accompanies BAL.

Concentration of aerosolized 99mTc-albumin in the pulmonary lymph of anesthetized sheep.

We examined the lymphatic concentration of 99mTc-albumin deposited in the air spaces of anesthetized sheep to determine whether changes in the concentration reflected changes in lung epithelial function. Five control sheep were ventilated with an aerosol of 99mTc-albumin for 6 min, and the lung lymphatic concentration of the tracer was monitored for the next 2 h. During the last 45 min the lymphatic concentration stabilized at a value that was 0.03 +/- 0.01% of the estimated value in the air spaces. Pulmonary vascular hypertension, induced in seven sheep by increasing the left atrial pressure 20 cmH2O for 4 h, increased the lung lymph flow from a base-line value of 3 +/- 2 to 21 +/- 14 ml/h. This caused the concentration of the 99mTc-albumin in the lymph to double to 0.07 +/- 0.03% of the air space concentration (P less than 0.01). Lung injury induced by infusing 0.08-0.10 ml/kg oleic acid intravenously in seven other sheep increased the lymphatic concentration of the 99mTc-albumin 10-fold to 0.31 +/- 0.09% of the air space concentration (P less than 0.01). The increased tracer concentration in the sheep with pulmonary vascular hypertension could be the result of the increased lymph flow causing a diversion of tracer into the lymphatics. However, a mathematical model showed that the 10-fold increase in the lymphatic concentration in the sheep with lung injury was primarily the result of an increase in both permeability and surface area of the epithelium that participated in the transfer of the 99mTc-albumin from the air spaces into the lung tissue drained by the lymphatics.(ABSTRACT TRUNCATED AT 250 WORDS)

A modified bronchoalveolar lavage procedure that allows measurement of lung epithelial lining fluid volume.

We replaced the standard serial bronchoalveolar lavage technique with a new "rewash" lavage procedure to allow estimation of the volume and protein concentration of the epithelial lining fluid (ELF) in anesthetized sheep. A bronchoscope 6.0 mm in diameter wedged in an airway was used to lavage a segment of lung with four cycles of instillation and aspiration of the lavage solution containing a radioactive tracer (technetium pertechnetate, 99mTcO4-). Errors caused by the fall in concentration of the tracer during the lavage were minimized by extrapolating the tracer concentration back to time zero when the lavage solution had mixed with the ELF, but had not had time to be affected by loss of the tracer or influx of fluid from the interstitium. In control sheep, the ELF of these lavaged segments had a mean volume of 1.6 +/- 1.0 ml and a mean protein concentration that was 26 +/- 19% of the protein concentration measured in the plasma. Increasing the left atrial pressure 19 +/- 5 cm H2O to cause "cardiac lung edema" had no significant effect on the ELF volume, but it increased the mean protein concentration to 57 +/- 30% of the plasma value (p less than 0.01). Lung injury caused by intravenous oleic acid caused lung edema, increased the mean ELF volume to 6.8 +/- 2.2 ml, and increased the mean ELF protein concentration to 86 +/- 26% of the plasma value (p less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of Pseudomonas aeruginosa elastase on alveolar epithelial permeability in guinea pigs.

Elastase-deficient mutants of Pseudomonas aeruginosa are less virulent than the wild type and are easily cleared from the lungs of guinea pigs. The effect of P. aeruginosa elastase on lung epithelium, however, is not yet understood. We addressed the hypothesis that breach of the epithelial barrier by elastase from P. aeruginosa allows invading organisms and toxic substances to penetrate the interstitium. We measured the clearance of aerosolized technetium-labeled albumin (molecular weight, 69,000) from the lungs of anesthetized guinea pigs with the aid of a gamma camera and a dedicated computer. Aerosols of the elastase (0.1 to 5 micrograms) increased the rate of clearance of labeled albumin from the lungs in proportion to the elastase dose. Electron microscopic studies using horseradish peroxidase as a tracer revealed that elastase interrupts intercellular tight junctions of the epithelial lining, thereby increasing the permeability to macromolecules. The amounts of elastase used in this report did not cause interstitial or alveolar edema, as determined by both postmortem extravascular lung water volume measurement and morphological examination. The data indicate that the elastase is a potentially important virulence factor in acute lung infection.

Comparison of three tracers for detecting lung epithelial injury in anesthetized sheep.

We compared the ability of three aerosolized tracers to discriminate among control, lung inflation with a positive end expired pressure of 10 cmH2O, lung vascular hypertension and edema without lung injury, and lung edema with lung injury due to intravenous oleic acid. The tracers were 99mTc-diethylenetriaminepentaacetate (99mTc-DTPA, mol wt 492), 99mTc-human serum albumin (99mTc-ALB, mol wt 69,000), and 99mTc-aggregated albumin (99mTc-AGG ALB, mol wt 383,000). 99mTc-DTPA clearance measurements were not able to discriminate lung injury from lung inflation. The 99mTc-AGG ALB clearance rate was unchanged by lung inflation and increased slightly with lung injury. The 99mTc-ALB clearance rate (0.06 +/- 0.02%/min) was unchanged by lung inflation (0.09 +/- 0.02%/min, P greater than 0.05) or 4 h of hypertension without injury (0.09 +/- 0.04%/min, P greater than 0.05). Deposition of 99mTc-ALB within 15 min of the administration of the oleic acid increased the clearance rate to 0.19 +/- 0.06%/min, which correlated well with the postmortem lung water volume (r = 0.92, P less than 0.01). This did not occur when there was a 60-min delay in the deposition of 99mTc-ALB. We conclude that 99mTc-ALB is the best indicator for studying the effects of lung epithelial injury on protein and fluid transport into and out of the air spaces of the lungs in a minimally invasive manner.

Local abnormalities of coagulation and fibrinolysis and alveolar fibrin deposition in sheep with oleic acid-induced lung injury.

Extravascular, primarily intra-alveolar, fibrin deposition is a histologic hallmark of acute lung injury in humans and experimental animals, but the mechanisms leading to this finding are poorly understood. To determine whether local abnormalities in the fibrinolytic-procoagulant balance contribute to alveolar fibrin deposition in acute lung injury, we studied bronchoalveolar lavage (BAL) fluids of anesthetized sheep that received intravenous oleic acid. Prominent alveolar fibrin deposition was observed within 2 h after oleic acid-induced lung injury. Procoagulant and fibrinolytic activities were determined in BAL samples of anesthetized, mechanically ventilated sheep before and 2 h after intravenous oleic acid or saline. BAL procoagulant activity was found to be due mainly to tissue factor associated with Factor VII. In baseline BAL samples, we found relatively low levels of procoagulant activity and relatively high levels of fibrinolytic activity. After induction of oleic acid-induced lung injury, the procoagulant activity of BAL was markedly increased, whereas fibrinolytic activity was either depressed or undetectable. Antiplasmin activity was detectable in BAL of sheep after oleic acid-induced lung injury, which contributed at least in part to the depressed fibrinolytic activity observed. These perturbations occurred with the appearance of extensive alveolar fibrin deposition. In control sheep, BAL fibrinolytic activity was decreased, and antiplasmin activity increased modestly after 2 h of mechanical ventilation, but procoagulant activity was unchanged and alveolar fibrin was not observed. Procoagulant activity in lung lymph and plasma after lung injury did not differ from baseline values, and fibrinolytic activity was undetectable in lymph or plasma samples. These data indicate that increased procoagulant activity and concurrent disruption of the balance of coagulation and fibrinolysis establish local conditions that promote acute fibrin deposition in the alveoli of mechanically ventilated, oleic acid-injured sheep.