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Abstract |
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Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
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The alveolar epithelium is not injured by the apical application of moderate doses of Pseudomonas aeruginosa strains that produce protease. To determine the effect of Pseudomonas proteases on the basolateral surface of the alveolar epithelium, a series of experiments were done, in which P. aeruginosa strains that produce and do not produce proteases were administered intravenously. Subsequently, an innocuous dose of bacteria was instilled into the lungs of the rabbits. Although all the intravenous Pseudomonas strains increased the extravascular lung water to a similar degree, only the intravenous administration of the protease-producing P. aeruginosa strain increased the vulnerability of the alveolar epithelium to injury by the subsequent airspace bacteria. Bacteremia secondary to P. aeruginosa strains producing proteases could increase the chances of developing acute lung injury.
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Introduction |
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Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
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The instillation of live Pseudomonas aeruginosa into the airspaces of sheep, rabbits, and rats caused an increase in the permeability of the lung epithelium to protein, produced infected pleural effusions, and injured both the alveolar epithelial type I and type II cells (1). The extent of the alveolar epithelial injury caused by these P. aeruginosa strains was dependent on the size of the bacterial inoculum and on the production of specific toxic bacterial exoproducts by the strains, specifically exoenzyme S and phospholipase C (2, 4).
The production of elastase by these P. aeruginosa strains did not affect the extent of alveolar epithelial injury (2, 4). In vitro studies have confirmed that Pseudomonas elastase does not diminish epithelial cell viability (5). However, Pseudomonas elastase was found to increase epithelial permeability by damaging tight junction-associated proteins in these in vitro studies (5).
To further investigate the role of P. aeruginosa proteases in causing lung injury, two wild-type strains of P. aeruginosa (one strain that produces elastase and alkaline protease [PA01] and one strain that naturally does not produce elastase [PA103]), were compared for their ability to cause lung injury. We had previously shown that the intravenous administration of P. aeruginosa did not lead to increased alveolar epithelial permeability (6). The present experiments were done to compare the effects of the intravenous administration of the two strains of P. aeruginosa and the effects of the production of proteases by the bacteria on the resistance of the alveolar epithelium to a second injury.
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Materials and Methods |
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Abstract
Introduction
Materials & Methods
Results
Discussion
References
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Surgical Preparation and Ventilation
New Zealand White rabbits (range, 3-5 kg) were anesthetized with 3% halothane and anesthesia was maintained with 1% halothane. A 20-gauge intravenous catheter was inserted into an ear vein for intravenous access. A right carotid arterial line was inserted to monitor systemic blood pressure and for blood sampling. Pancuronium (0.3 mg/kg) was given for neuromuscular blockade. An endotracheal tube was inserted through a tracheostomy. The rabbits were placed supine and ventilated with a constant-volume pump (Harvard, Millis, MA) with an inspired O2 fraction of 1.0, peak pressures of 13-17 cm H2O, and positive end-expiratory pressure of 3 cm H2O. The respiratory rate was adjusted to maintain an arterial PCO2 between 35 and 45 mm Hg. All animal experiments were done in compliance with the Animal Care Committee of the University of California at San Francisco.
Preparation of Instilled Solution
A 5% rabbit albumin solution was prepared using Ringer's lactate and adjusted with NaCl to be isoosmolar with the circulating plasma of the rabbit. The solution was then filtered through a 0.2-µm pore size filter (Nalge Company, Rochester, NY) and 2 mg of anhydrous Evan's blue and 3 µCi of 125I-labeled human serum albumin (Merck-Frosst, Quebec, Canada) were added to the rabbit albumin solution, as we have done in prior studies (2, 4, 7). Evan's blue was used to ascertain that the instilled solution was confined to the instilled lower lobe. The bacteria used in the instillate never varied; PA01 Fisher immunotype 7 P. aeruginosa (107 colony-forming units [cfu]/ml) was added to the protein solution. A sample of the instillate was saved for radioactivity counts, total protein measurement, and water-to-dry weight ratio measurement so that the dry weight of the protein solution could be subtracted from the final lung water calculation.
Culture Conditions
All bacteria were taken from frozen explants, streaked onto blood agar plates, and grown overnight in broth. The wild-type PA01 (Fisher immunotype 7) and the PA0R1 mutant (generously donated by Dr. Barbara Iglewski, University of Rochester, Rochester, NY) were grown at 37°C for 24 h in trypticase soy broth with 10 mM nitrilotriacetic acid added to the dialyzed medium. PA103 bacteria (Fisher immunotype 2) were cultured at 32°C for 24 h in trypticase soy broth and with nitrilotriacetic acid, which was added to the dialyzed medium to enhance exoenzyme S production. Cultures were centrifuged at 8,500 × g for 5 min and the bacterial pellet was washed three times with phosphate-buffered saline prior to the addition of the bacteria to the instillate. The number of viable bacteria added to the instillate was determined by serial dilution of the bacterial suspension prior to addition to the instillate.
Bacterial Strains
Table 1 lists the P. aeruginosa strains used in this study, with the relevant phenotypic characteristics. Strains PA01 and PA103 have been described (8, 9). Strain PA01 has been shown to produce elastase, exotoxin A, and exoenzyme S, whereas strain PA103 naturally does not produce elastase, but hyperproduces exotoxin A and exoenzyme S. To explore the importance of bacterial proteases in causing injury to the lung endothelium and the alveolar epithelium, we also used PA0R1, an isogenic mutant of PA01, which does not produce either elastase or alkaline protease. The mutation is in the lasR gene of PA01, and was constructed using a deletion and insertion strategy similar to that described by Ostroff and coworkers (10). lasR has been shown to be required for transcription of lasB, the elastase structural gene (11). This isogenic strain, PA0R1, has been shown to produce all other bacterial exoproducts in normal quantities (4).
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General Experimental Protocol
For all experimental studies, we used the following general protocol. After surgery, a 1-h baseline with stable heart rate and blood pressure was established prior to the instillation of the solution containing protein and/or bacteria. Three microcuries of 131I-labeled human albumin was injected intravenously at least 30 min prior to the instillation. The rabbits were placed in the right lateral decubitus position to facilitate liquid deposition into their right lungs. Using a 12-ml syringe and pediatric feeding tube (5F), the instillate (3 ml/kg) was delivered to the right lung (primarily lower lobe) over 20 min, as we have done previously (2, 4, 7).
After 8 h, the abdomen was opened and the rabbits were exsanguinated. Urine, pericardial fluid, right and left pleural fluids, and liver were sampled for bacterial cultures and radioactivity counts. The lungs were removed through a sternotomy and the remaining alveolar liquid was obtained by passing a Silastic tube (0.74-mm internal diameter [i.d.]) into a wedged position in the right lower lobe. After centrifugation, the total protein concentration and radioactivity of the alveolar liquid sample were measured. We have previously reported that the concentration of total protein in the liquid sampled by a catheter wedged into the distal airways was the same as in a directly obtained alveolar micropuncture sample (12). Extravascular lung water in each lung was measured.
Specific Protocols
Instillation of the protein solution alone (n = 6). After the baseline period, a 5% rabbit albumin solution (3 ml/kg) was instilled into the right lower lobe of the anesthetized rabbits. After monitoring for 8 h, the rabbits were exsanguinated and processed as described in the general experimental protocol.
Airspace instillation of PA01 P. aeruginosa (n = 4). After the baseline period, PA01 P. aeruginosa (107 cfu/ml instillate) was mixed with a 5% rabbit albumin solution in Ringer's lactate and instilled (3 ml/kg) into the right lower lobe. We have previously reported that the instillation of a protein solution containing PA01 P. aeruginosa (107 cfu/ml instillate) into the airspaces does not cause any injury to the alveolar epithelial barrier (2). The rabbits were monitored and processed as described in the general experimental protocol.
Effect of intravenous bolus of PA01 P. aeruginosa (n = 6). After the baseline period, a bolus of PA01 P. aeruginosa (109 cfu) was administered intravenously 60 min before the 5% rabbit albumin solution was instilled into the right lower lobe. The rabbits were monitored and lungs processed as described in the general experimental protocol.
Effect of intravenous and airspace PA01 P. aeruginosa (n = 4). After the baseline period, a bolus of PA01 P. aeruginosa (109 cfu) was administered intravenously 60 min prior to the airspace instillation of PA01 P. aeruginosa (107 cfu/ml instillate). These rabbits were monitored and their lungs were processed as described in the general experimental protocol.
Effect of P. aeruginosa virulence exoproducts (n = 17). The intravenous PA01 was replaced by strains of P. aeruginosa that did not produce elastase, PA0R1, the isogenic mutant strain, or by PA103, the strain that naturally does not produce elastase (Table ). Sixty minutes after the intravenous administration of one of these P. aeruginosa strains, the PA01 P. aeruginosa (107 cfu/ml instillate) was instilled into the right lower lung lobe of the experimental rabbit. Control studies were done in which the PA0R1 or PA103 (109 cfu) was administered intravenously 60 min prior to the instillation of a protein solution not containing bacteria.
Tracer Binding Measurement
To determine if the experimental conditions caused dissociation of the radiolabeled iodine from the protein tracer, we incubated 109 bacteria of each strain in 5% rabbit albumin in Ringer's lactate solution at 37°C for 8 h, and measured specific activity. Trichloroacetic acid (TCA) was added to all tubes and the tubes were centrifuged to obtain the supernatant for measurement of free 125I radioactivity. The results were expressed as the percentage of the unbound 125I radioactivity relative to the total amount of 125I-labeled albumin radioactivity instilled.
Measurements of Hemodynamics and Protein Concentration
Systemic arterial and airway pressures were measured at 30-min intervals. Total protein concentration of plasma, and instilled and final samples from the airspaces were measured using an automated analyzer (AAII; Technicon, Tarrytown, NJ).
Measurement of Alveolar Epithelial and Endothelial Barrier Protein Permeability
Two different methods were used to measure the protein permeability across the alveolar epithelial barrier (2, 4, 7). The first method required the measurement of the residual 125I-labeled albumin (the alveolar protein tracer) in the lung as well as the accumulation of 125I-labeled albumin in the plasma. The second method measured 131I-labeled albumin (the vascular protein tracer) in the airspace compartment of the lung.
The total 125I-labeled albumin instilled into the lung
was determined by measuring total radioactivity (counts · min
1 · g1) in the instillate and multiplying this value by
the total volume instilled. To determine the residual 125I-labeled albumin in the lung after 8 h, the average of the
specific activity in two 0.5-g samples of lung homogenate
was multiplied by the total volume of the homogenates.
The lung homogenate radioactivity was added to the
counts recovered in the aspirated alveolar fluid to determine the total 125I-labeled albumin remaining in the lung
after 8 h. The plasma 125I-labeled albumin in the circulation was measured from a sample of plasma obtained at 8 h.
The total number of counts in the plasma was calculated
by multiplying the counts per milliliter by the total plasma
volume [body weight × 0.07(1
Hematocrit)].
The second method involved measurement of the movement of the vascular protein tracer, 131I-labeled albumin, into the final alveolar liquid. Plasma 131I-labeled albumin counts were averaged over the course of the experiment, and the 131I-labeled albumin counts in the airspaces were expressed as a ratio to the plasma counts. This ratio provides a good index of equilibration of the vascular protein tracer into the alveolar compartment, as has been shown (2, 4, 7).
Also, the accumulation of the vascular protein tracer in the extravascular spaces of the lungs was used as an additional index of lung endothelial permeability. The movement of plasma into the extravascular spaces of the lungs was determined by measuring the total extravascular counts of 131I-labeled albumin in the lung divided by the average counts in the plasma over 8 h (13).
Changes in the protein concentration of the instilled fluid were determined over 8 h. When the protein concentration in the final fluid sampled from the distal airspaces is significantly less than the protein concentration measured in the samples from control animals, either the alveolar epithelium can no longer actively transport sodium owing to injury, or there is a continued influx of fluid into the airspaces overwhelming the capacity of the alveolar epithelium for active transport (2, 4, 7).
Lung Fluid Balance
Extravascular lung water was determined by calculating the water-to-dry weight ratio, as previously described (7). The volume of the excess lung water (ELW) of the instilled experimental lungs was calculated as the difference between the water-to-dry weight ratios of the experimental and contralateral lung multiplied by the dry weight of the instilled lung. The "dry weight" of the instilled experimental lungs was calculated by subtracting the dry weight of the instilled protein remaining in the lung at the end of the experiment. To determine the mass of protein remaining in the instilled lungs, the dry weight of the instillate was multiplied by the fraction of 125I-labeled albumin remaining in these lungs. These values were then subtracted from the weights of the experimental lungs.
The equation used for these calculations is ELW (ml) = [We/(De P) Wc/Dc](De P), where W and D are
extravascular lung water and blood-free dry weight of the
experimental lung (e) and control lung (c), and P is the
blood-free dry weight of the initial alveolar fluid multiplied by the fraction of 125I-labeled albumin remaining in
the lung. This equation does not account for the possibility
that some of the circulating plasma may enter the instilled
experimental lung. To estimate the quantity of plasma that
entered the instilled lung, we measured the transfer of the
vascular protein tracer, 131I-labeled albumin, into the extravascular spaces of the instilled lung.
In addition, changes in the water-to-dry weight ratio of the contralateral (noninstilled lung) secondary to the intravenous administration of P. aeruginosa were used as another index of lung endothelial injury. In preliminary experiments, we found that both lungs gained the same amount of water per gram of tissue over 8 h secondary to the intravenous administration of P. aeruginosa.
Statistics
All data are presented as mean ± SE. Data were analyzed by a one-way analysis of variance; Fisher's test was used for comparisons. Culture data were analyzed using chi-square and contingency tables. A value of P < 0.05 was accepted as statistically significant (14).
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Results |
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Abstract
Introduction
Materials & Methods
Results
Discussion
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Alveolar Epithelial Injury
The only rabbits that had evidence of increased alveolar epithelial permeability were those that had received both the intravenous and airspace PA01 bacteria. Eight percent of the alveolar protein tracer (125I-labeled albumin) left the lungs of the rabbits treated with both intravenous and airspace PA01 bacteria (Table 2).
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These rabbits also had a significant increase in the alveolar-to-plasma 131I-labeled albumin ratio recovered in their airspaces (Figure 1). In contrast, there was not a significant increase in the ratio when the rabbits received the airspace protein solution or the airspace PA01 or when the rabbits received intravenous PA01 P. aeruginosa. Nor was there an increase in this ratio when the rabbits received the intravenous PA103, the strain that naturally does not produce elastase, and then received the airspace PA01 (Figure 2). Finally, the intravenous administration of PA0R1, the isogenic mutated strain that does not produce proteases, followed by the airspace instillation of PA01 did not lead to an increase in the alveolar-to-plasma 131I-labeled albumin ratio (Figure 2).
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Additional evidence for epithelial injury in the rabbits given both intravenous and airspace PA01 bacteria was the significantly lower protein concentration of their final airspace fluid (P < 0.05) when compared to the protein concentration in the final airspace fluid from the rabbits who had received a single intravenous or airspace injection (Table 3).
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Quantitative cultures of all the instilled solutions verified that 107 cfu/ml PA01 P. aeruginosa had been instilled in the rabbits designated to receive bacterial airspace instillates. Terminally, there was a five-fold increase in the number of PA01 cultured from the instilled lungs of the rabbits that had received both the intravenous PA01 and the airspace PA01 (Figure 3). In contrast, there was a four-fold decrease in the number of PA01 cultured from the lungs of the rabbits that had received only airspace PA01. The rabbits that had received intravenous PA01 only had, on average, 5 × 105 cfu of PA01 cultured from their lungs.
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Lung Endothelial Injury and Systemic Hemodynamic Effects
Rabbits that receive any strain of intravenous P. aeruginosa developed lung edema as indicated by a significant increase (P < 0.05) in extravascular lung water when compared with values measured in rabbits that had received airspace protein solutions or airspace PA01 alone (Figure 4). All rabbits that received intravenous P. aeruginosa developed systemic arterial hypotension and metabolic acidosis after 8 h (final values for rabbits that received intravenous and airspace PA01: mean arterial pressure 61 ± 6 mm Hg; pH: 7.31 ± 0.04, respectively). Similar results were obtained when the intravenous PA01 was replaced by PA0R1 or PA103, the pseudomonal strains that do not produce proteases.
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In contrast, rabbits that had received airspace protein solutions or that had received airspace PA01 alone did not develop metabolic acidosis. These rabbits had final mean systemic arterial pressures of 87 ± 7 mm Hg and the final arterial pH values were pH 7.42 ± 0.04 in the rabbits that had received airspace protein solutions and pH 7.46 ± 0.03 in the rabbits that had received airspace PA01. The initial and final arterial PO2 were 461 ± 34 and 452 ± 29 mm Hg, respectively, in the rabbits that received airspace protein solutions; the arterial PO2 values were not significantly different in any of the experimental groups. Because ventilation was controlled, arterial PCO2 was kept constant.
Tracer Binding
The percentages of unbound 125I after 8 h of incubation at 37°C were 1.11% for PA01, 0.28% for PA103, and 0.25% for PA0R1. Trichloracetic acid protein precipitation was also done on the final alveolar fluid obtained from rabbits that received intra-alveolar P. aeruginosa. These aspirates never had greater than 0.6% of unbound iodine present. The percentage of free 125I from samples of the tracer taken from the stock solution was 0.20% and when a sample of 125I-labeled albumin was left for 8 h in phosphate-buffered saline (PBS) the free 125I was found to be 0.24%.
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Discussion |
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Abstract
Introduction
Materials & Methods
Results
Discussion
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When intravenous bacteria were administered to produce acute lung injury, alveolar epithelial injury has not been demonstrated. Large doses of intravenous Escherichia coli endotoxin (1) or repeated intravenous boluses of P. aeruginosa (6) administered to sheep, rats, and rabbits did not injure the alveolar epithelial barrier despite the development of severe systemic arterial hypotension and metabolic acidosis. In contrast, sepsis-induced acute lung injury in humans often involves flooding of the airspaces with protein-rich edema fluid, indicating an increase in the alveolar epithelial permeability to protein (15, 16). We hypothesized that bacterial products affect the basolateral surface of alveolar epithelium so that subsequent apical challenges cause epithelial injury.
To test this hypothesis, PA01 was given intravenously in combination with a noninjurious dose of PA01 into the airspaces of anesthetized rabbits and alveolar epithelial injury was produced. The injury was manifested by a significant increase in the alveolar epithelial permeability to protein, a significant decrease in ability of the alveolar epithelium to transport fluid out of the airspaces, and the production of infected pleural effusions (data not shown). In contrast, the single instillation of the bacteria PA01 into the airspaces or the single administration of PA01 intravenously did not produce alveolar epithelial permeability changes or injury.
To determine the mechanism of this injury, other experiments were performed in which bacterial strains that could (PA01) or could not (PA0R1 or PA103) produce proteases were administered intravenously prior to the instillation of PA01. An increase in alveolar epithelial injury was produced only when the bacteria administered intravenously produced proteases. These results were the same for both the administration of the wild-type strain (PA103) that naturally does not produce proteases or for the administration of the isogenic, mutant strain PA0R1, that had been engineered not to produce proteases. The results were not due to differences in bacterial clearance; the numbers of bacteria recovered at the end of the experiments from the instilled lungs were similar in all of the experimental groups in which intravenous and airspace bacteria were administered.
Note that the strain of P. aeruginosa that does not produce elastase, PA013, has been found to cause severe epithelial injury when administered into the airspaces of rabbits or rats (3, 4). However, when PA103 was administered intravenously prior to the airspace PA01 instillation, alveolar epithelial injury was not produced. Therefore, although injurious when instilled into the airspaces of the lung, the intravenous administration of PA103 does not reduce the resistance of the alveolar epithelium to injury.
Previous in vivo and in vitro studies have reported that the mucosal side of the alveolar epithelium is resistant to injury from P. aeruginosa elastase. For example, the nebulization of 20 mg of P. aeruginosa elastase to the lungs of anesthetized sheep did not increase extravascular lung water or alter lung endothelial or epithelial permeability to protein. Gas exchange remained normal despite this treatment (17). Similarly, the airspace instillation of PA0R1, the mutant that does not produce elastase, caused the same degree of alveolar epithelial injury as seen after the instillation of wild-type elastase-producing PA01 in rabbits (2).
In contrast, there is in vitro experimental evidence suggesting that P. aeruginosa elastase decreases the permeability of the apical and basolateral surfaces of cultured alveolar epithelial cells as well as the permeability of other cultured epithelial cells (5). The discrepancy between the results in this investigation and the results found in the in vivo experiments may be explained by a very small amount of protease production by P. aeruginosa in vivo. This could occur as there is clearly in vivo regulation of the secretion of virulence products of live bacteria. There are data that bacterial virulence products are produced only after contact with host cells (18). In fact, P. aeruginosa, as well as other pathogenic gram-negative bacteria, have a type III secretory system that allows these bacteria to inject virulence products directly into the host cells (19). Therefore, P. aeruginosa may only produce proteases once it is in close proximity to host cells.
There have been in vivo experiments using the intravenous administration of Psuedomonas elastase, rather than the intravenous injection of live P. aeruginosa that produce proteases. In guinea pigs, the intravenous administration of P. aeruginosa elastase caused the development of hypotension due to decreased systemic vascular resistance (20) that was associated with the activation of the kallikrein- kinin system (21). Other in vitro experiments using P. aeruginosa elastase demonstrated that the P. aeruginosa protease cleaved Fe-transferrin to form iron complexes that catalyzed the formation of hydroxyl radicals from neutrophil products. The investigators concluded that the hydroxyl formation might serve as the mechanism for tissue injury at sites of bacterial infection (22).
The present experiments and these reports suggest that the exposure of the lung endothelium or the basolateral surface of the alveolar epithelium to P. aeruginosa protease facilitates the development of alveolar epithelial injury when the apical side of the alveolar epithelium is exposed to small doses of bacteria. The experiments in this report and in our previous investigations (2, 4) further suggest that each barrier of the lung can be injured by different P. aeruginosa exoproducts.
In conclusion, injury to the alveolar epithelial barrier occurred only after a combined administration of intravenous PA01 and a small, innocuous airspace innoculum of PA01. The Pseudomonas protease produced by the intravenous bacteria was critical for the development of this alveolar epithelial injury. Specific exoproducts of P. aeruginosa are injurious to the mucosal surface of the alveolar epithelium and other exoproducts from this bacteria injure the basolateral surface of the alveolar epithelium. The implication of these studies is that bacterial strains may be virulent at one cell surface, and less pathogenic at other cellular locations. Bacteremia may lead to acute lung injury when the intravenous bacteria produce proteases or exoproducts that increase the susceptibility of the alveolar epithelium to subsequent injury.
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Footnotes |
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Address correspondence to: Jean-Francois Pittet, M.D., Department of Anesthesia, Room 3C-38, San Francisco General Hospital, 1001 Potrero Avenue, San Francisco, CA 94110. E-mail: jean_pittet{at}quickmail.ucsf.edu
(Received in original form September 24, 1996 and in revised form June 3, 1997).
Acknowledgments: The authors wish to thank Dr. Michael A. Matthay (UCSF) for his comments regarding this work, and Richard Shanks for his technical assistance in carrying out these studies. This work was primarily supported by National Heart, Lung, and Blood Institute Grants HL 49810 and HL 55980.
Abbreviations cfu, colony-forming unit; ELW, excess lung water; i.d., internal diameter; TCA, trichloroacetic acid.
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References |
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Abstract
Introduction
Materials & Methods
Results
Discussion
References
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1. Wiener-Kronish, J. P., K. Albertine, and M. A. Matthay. 1991. Differential responses of the endothelial and epithelial barriers of the lung in sheep to E. coli endotoxin. J. Clin. Invest. 88: 864-875 .
2.
Wiener-Kronish, J. P.,
T. Sakuma,
I. Kudoh,
J. F. Pittet,
D. Frank,
L. Dobbs,
M. Vasil, and
M. A. Matthay.
1993.
Alveolar epithelial injury and
pleural empyema in acute P. aeruginosa pneumonia in anesthetized rabbits.
J. Appl. Physiol.
75:
1661-1669
3.
McElroy, M. C.,
J. F. Pittet,
L. Allen,
J. P. Wiener-Kronish, and
L. G. Dobbs.
1995.
Alveolar fluid content of a specific type I cell protein is an index of epithelial injury in a rat model of acute pneumonia.
Am. J. Physiol.
268:
L181-L186
4.
Kudoh, I.,
J. P. Wiener-Kronish,
S. Hashimoto,
J. F. Pittet, and
D. Frank.
1994.
Exoproduct secretions of P. aeruginosa strains influence severity of
alveolar epithelial injury.
Am. J. Physiol
267:
L551-L556
5. Azghani, A. O.. 1996. Pseudomonas aeruginosa and epithelial permeability: Role of virulence factors elastase and exotoxin A. Am. J. Respir. Cell Mol. Biol. 15: 132-140 [Abstract].
6. Pittet, J. F., J. P. Wiener-Kronish, M. C. McElroy, H. G. Folkesson, and M. A. Matthay. 1994. Stimulation of alveolar epithelial liquid clearance by endogenous release of catecholamines in septic shock. J. Clin. Invest 94: 663-671 .
7.
Smedira, N.,
L. Gates,
R. Hastings,
C. Jayr,
T. Sakuma,
J. F. Pittet, and
M. A. Matthay.
1991.
Alveolar and lung liquid clearance in anesthetized rabbits.
J. Appl. Physiol.
70:
1827-1835
8.
Holloway, B. W.,
V. Krishnapillae, and
A. F. Morgan.
1979.
Chromosomal
genetics of Pseudomonas.
Microbiol. Rev.
43:
73-102
9. Liu, P. V.. 1974. Extracellular toxins of Pseudomonas aeruginosa. J. Infect. Dis. 130: S94-S99 .
10.
Ostroff, R. M.,
B. Wretlind, and
M. L. Vasil.
1989.
Mutations in the hemolytic-phospholipase C operon result in decreased virulence of Pseudomonas aeruginosa PA01 grown under phosphate-limiting conditions.
Infect. Immun.
57:
1369-1373
11.
Gambello, M. G., and
B. H. Iglewski.
1991.
Cloning and characterization of
the Pseudomonas las R gene, a transcriptional activator of elastase expression.
J. Bacteriol.
173:
3000-3009
12.
Berthiaume, Y.,
V. C. Broaddus,
M. A. Gropper,
T. Tanita, and
M. A. Matthay.
1988.
Alveolar liquid and protein clearance from normal dog lungs.
J. Appl. Physiol.
65:
585-593
13. Pittet, J. F., J. P. Wiener-Kronish, V. S. Serikov, and M. A. Matthay. 1995. Resistance of the alveolar epithelium to injury from septic shock in sheep. Am. J. Respir. Crit. Care Med 151: 1093-1100 [Abstract].
14. Zar, J. H. 1974. Biostatistical Analysis. Prentice Hall, Englewood Cliffs, NJ.
15. Matthay, M. A., and J. P. Wiener-Kronish. 1990. Intact epithelial barrier function is critical for the resolution of alveolar edema in humans. Am. Rev. Respir. Dis. 142: 1250-1257 [Medline].
16.
Clark, J. G.,
J. A. Milberg,
K. P. Steinberg, and
L. D. Hudson.
1995.
Type III procollagen peptide in the adult respiratory distress syndrome: association of increased peptide levels in bronchoalveolar lavage fluid with increased risk for death.
Ann. Intern. Med.
122:
17-23
17.
Peterson, P. T.,
M. L. Collins,
L. D. Gray, and
A. O. Azghani.
1992.
Aerosolized Pseudomonas elastase and lung fluid balance in anesthetized
sheep.
J. Appl. Physiol.
72:
1927-1933
18. Pettersson, J., R. Nordfelth, E. Dubinina, T. Bergman, M. Gustafsson, K. E. Magnusson, and H. Wolf-Watz. 1996. Modulation of virulence factor expression by pathogen target cell contact. Science 273: 1231-1233 [Abstract].
19. Yahr, T. L., J. Goranson, and D. W. Frank. 1996. Exoenzyme of Pseudomonas aeruginosa is secreted by a type III pathway. Mol. Microbiol. 22: 991-1003 [Medline].
20. Khan, M. M. H., T. Yamamoto, H. Araki, Y. Ijiri, Y. Shibuya, M. Okamoto, and T. Kambara. 1993. Pseudomonal elastase injection causes low vascular resistant shock in guinea pigs. Biochim. Biophys. Acta 1182: 83-93 [Medline].
21. Khan, M. M. H., T. Yamamoto, H. Araki, Y. Shibuya, and T. Kambara. 1993. Role of Hageman factor/kallikrein-kinin system in pseudomonal elastase-induced shock model. Biochim. Biophys. Acta 1157: 119-126 [Medline].
22. Miller, R. A., and B. E. Britigan. 1995. Protease-cleaved iron-transferrin augments oxidant-mediated endothelial cell injury via hydroxyl radical formation. J. Clin. Invest. 95: 2491-2500 .
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