Direct measurement of microvascular pressures in the isolated perfused dog lung.

Microvascular pressures in the pulmonary circulation were measured under the pleural surface of the isolated perfused dog lung by the servo-null technique. Strong glass micropipettes with short beveled tips were used, with a suction ring to stabilize the lung's surface. Of the total vascular resistance, 45 percent was in the alveolar wall capillaries themselves. Most of the remaining resistance was in the arterioles. There was negligible pressure drop in venules with diameters larger than 20 micrometers.

Beta-adrenergic agonists increase lung liquid clearance in anesthetized sheep.

We did experiments to determine whether beta-adrenergic agonists increase lung liquid clearance in anesthetized ventilated adult sheep and, if so, whether the increase is mediated by beta receptors and what mechanism is involved. We instilled 100 ml of autologous serum either alone or with a beta-adrenergic agonist (terbutaline, 10(-5) M, or epinephrine, 5.5 X 10(-6) M) into one lower lobe. After 4 h both terbutaline and epinephrine increased lung liquid clearance. The increase in lung liquid clearance was inhibited when propranolol (a beta blocker) or amiloride (a sodium channel blocker) was added to the terbutaline. Increased clearance was not explained by changes in pulmonary hemodynamics, pulmonary blood flow, or lung lymph flow. We conclude that beta-adrenergic agonists increase lung liquid clearance in anesthetized intact adult sheep. This increase is mediated through beta receptors and probably depends on increased active transport of sodium across the alveolar barrier.

Increased sheep lung vascular permeability caused by pseudomonas bacteremia.

In awake sheep, we compared the responses of lung lymph flow and lymph and plasma protein concentrations to steady state elevations of pulmonary vascular pressures made by inflating a left atrial balloon with those after an intravenous infusion of 10(5)-10(10)Pseudomonas aeruginosa. Lymph flow increased when pressure was increased, but lymph-plasma protein concentration ratios always fell and lymph protein flow (lymph flow x lymph protein concentration) increased only slightly. After Pseudomonas, sheep had transient chills, fever, leukopenia, hypoxemia, increased pulmonary artery pressure and lymph flow and decreased left atrial pressure and lymph protein concentration, 3-5 h after Pseudomonas, when vascular pressures and lymph protein concentrations had returned to near base line, lymph flow increased further to 3-10 times base line and remained at a steady level for many hours. During this steady state period, lymph-plasma protein concentration ratios were similar to base line and lymph protein flow was higher than in the increased pressure studies. Two sheep died of pulmonary edema 7 and 9 h after Pseudomonas, but in 16 studies, five other sheep appeared well during the period of highest lymph flow and all variables returned to base line in 24-72 h. Six serial indicator dilution lung water studies in five sheep changed insignificantly from base line after Pseudomonas. Postmortem lung water was high in the two sheep dead of pulmonary edema and one other, but six sheep killed 1-6 h after Pseudomonas had normal lung water. Because of the clear difference between the effects of increased pressure and Pseudomonas on lymphplasma protein concentration ratios and lymph protein flow, we conclude that Pseudomonas causes a prolonged increase in lung vessel permeability to protein. Because we saw lung lymph flow as high as 10 times base line without pulmonary edema, we conclude that lung lymphatics are a sensitive high-capacity mechanism for removing excess filtered fluid. An equivalent pore model of sheep lung vessels suggests that the changes we saw after Pseudomonas could result from small changes in the structure of exchanging vessel walls.

Relationship of pleural effusions to increased permeability pulmonary edema in anesthetized sheep.

We studied anesthetized sheep to determine the relationship between increased permeability pulmonary edema and the development and mechanism of pleural effusion formation. In 12 sheep with intact, closed thoraces, we studied the time course of pleural liquid formation after 0.12 ml/kg i.v. oleic acid. After 1 h, there were no pleural effusions, even though extravascular lung water increased 50% to 6.0 +/- 0.7 g/g dry lung. By 3 h pleural effusions had formed, they reached a maximum at 5 h (48.5 +/- 16.9 ml/thorax), and at 8 h there was no additional accumulation of pleural liquid (45.5 +/- 16.9 ml). Morphologic studies by light and electron microscopy demonstrated subpleural edema but no detectable injury to the visceral pleura, suggesting that the pleural liquid originated from the lung and not the pleura. In nine sheep, we quantified the rate of formation of pleural liquid by enclosing one lung in a plastic bag. By comparing in the same sheep the volume of pleural liquid collected from the enclosed lung to the volume found in the opposite intact chest, we estimated the rate of liquid absorption from the intact chest to be 0.32 ml/(kg.h); we had previously reported a liquid absorption rate of 0.28 ml/(kg.h) in normal sheep. These studies also supported the conclusion that the majority of the pleural liquid originated from the lung because we could account for all of the pleural liquid that was formed and cleared. The volume of pleural liquid collected from the enclosed lungs was equal to 21% of the excess lung liquid that formed after oleic acid-induced lung injury. Thus, the pleural space and parietal pleural lymphatic pathways are important pathways for the clearance of pulmonary edema liquid after experimentally induced increased permeability pulmonary edema.

Steady-state, hemoglobin-facilitated O2 transport in human erythrocytes.

We measured the rate of oxygen transport through thin (165 micro) films of packed erythrocytes (Hb concentration = 30 g/100 ml). Under optimal conditions steady-state O(2) diffusion was nearly three times that found when the hemoglobin was prevented from acting as a carrier molecule by carbon monoxide binding or high oxygen back pressure. After each experiment we measured hemolysis and found that it averaged less than 1%. Hemolysis could not account for the facilitation, thus proving that facilitated transport of O(2) by hemoglobin can occur in red blood cells. The rate of facilitated transport was identical for Hb solutions of equal concentration to the cells. We interpret this to mean that under the conditions of our experiments the red cell membrane offers no detectable diffusion resistance to O(2) and that the mobility of Hb in intact red cells is the same as in concentrated Hb solution.

The value of edema fluid protein measurement in patients with pulmonary edema.

Alveolar fluid and plasma proteins were analyzed in 24 patients with florid pulmonary edema, in 21 of whom pulmonary capillary wedge pressure (Pcw) was also measured. In all patients with Pcw less than 20 mm Hg, the edema fluid to plasma protein ratio exceeded 0.6; the mean edema fluid to plasma protein ratio in the four patients with cardiogenic edema (increased Pcw) was 0.46. In the 21 patients in whom full data were available, the net intravascular filtration force (Pcw - plasma colloid osmotic pressure) was less than -4 mm Hg, the value at which (according to others) pulmonary edema should occur, in only 10. When the interstitial colloid osmotic pressure, approximated by the osmotic pressure of edema fluid protein, was added, the net filtration force became positive in 17 of 21 patients. Comparison of the protein concentrations of edema fluid and plasma aids in the diagnostic separation of increased permeability from high hydrostatic pressure edema and adds to our understanding of the relative osmotic and hydrostatic forces that contribute to pulmonary edema when the alveolar-capillary membrane is damaged.

Pulmonary edema: physiologic approaches to management.

The integrity of the normal endothelial barrier is responsible for two of the three major safety factors preventing pulmonary edema. This is why edema due to increased pressure is usually not as severe as edema due to increased permeability. Management ought to follow sound physiologic principles. These principles are to lower microvascular hydrostatic pressure and to provide adequate supportive therapy. Positive end-expiratory pressure may improve arterial oxygen transport, but there is no evidence that it improves the balance of pulmonary fluid in edema. Raising the microvascular protein osmotic pressure may be beneficial in edema due to increased pressure but has no demonstrated rationale in edema due to increased permeability.

Effect of acute hemorrhagic shock on pulmonary microvascular fluid filtration and protein permeability in sheep.

In an attempt to further delineate the pathophysiology of the shock-lung syndrome, we studied the effect of hemorrhagic shock on the pulmonary microcirculation in the adult sheep with lung lymph flow and lymph protein transport as indices of fluid filtration and vascular permeability to protein. We noted that lymph flow remained at baseline levels during shock, despite significant decreases in pulmonary artery and left atrial pressures. This suggests that microvascular pressure is maintained by an increase in pulmonary venous resistance. We demonstrated that lymph protein transport does not increase during shock, which indicates that a change in pulmonary vascular permeability to protein does not occur.

Indirect estimate of perimicrovascular pressure rise in edema in isolated zone 1 lung.

To determine how liquid accumulation affects extra-alveolar perimicrovascular interstitial pressure, we measured filtration rate under zone 1 conditions (25 cmH2O alveolar pressure, 20 or 10 cmH2O vascular pressure) in isolated dog lung lobes in which all vessels were filled with autologous plasma. In the base-line condition, starting with normal extra-alveolar water content, filtration rate decreased by about one-half over 1 h as edema liquid slowly accumulated. We repeated each experiment after inducing edema (up to 100% lung weight gain). The absolute values and time course of filtration in the edema condition did not differ from base-line, i.e., the edema did not affect the time course of filtration. To compute the maximal initial and maximal change in extra-alveolar perimicrovascular pressure that occurred over each 1-h filtration study, we first assumed that the reflection coefficient is 0 in the Starling equation, then calculated perimicrovascular pressure and filtration coefficient from two equations with two unknowns. The mean filtration coefficient in 10 lobes is 0.063 g/(min X cmH2O X 100 g wet wt), and the initial perimicrovascular pressure is 3.9 cmH2O, rising by 4-7 cmH2O at 1 h. Finally we tested low protein perfusates and found the filtration rate was higher. We calculated an overall reflection coefficient = 0.44, a decrease in the initial perimicrovascular pressure to 1.9 cmH2O and a slightly lower increase after 1 h of edema formation, 2.2-6.6 cmH2O.

Filtration profile in isolated zone 1 and zone 3 dog lungs at constant high alveolar pressure.

We measured the rate of liquid filtration in isolated dog lung lobes inflated to a constant alveolar pressure of 25 cmH2O and with all open vessels filled with plasma. We measured lung weight gain at vascular pressures ranging from 5 to 40 cmH2O relative to pleural pressure. We confirmed that under zone 1 conditions the "arterial" and "venous" extra-alveolar segments have essentially the same filtration characteristics. Using the combined extra-alveolar vascular system, we determined when recruitment of filtration surface area occurred as we increased vascular pressure from 0 to 40 cmH2O. Based on an abrupt increase in filtration rate as vascular pressure approached the zone 1/3 boundary, we infer that a sudden recruitment of exchange surface area occurred at that point. Based on the slopes of the zone 1 and zone 3 filtration profiles, we conclude that extra-alveolar vascular segments contribute approximately 25% of total to filtration in the lung under zone 3 conditions, although the exact vessels filtering under zone 1 conditions have yet to be determined. Our analysis of the data supports the concept that there is a difference in the perimicrovascular pressure around alveolar and extra-alveolar vessels, which in part may account for the apparent high filtration fraction apportioned to extra-alveolar vessels.

Continuous measurement of protein osmotic pressure in blood and lymph of sheep.

We have continuously measured protein osmotic pressure of blood and lymph in sheep to compare two kinds of needle osmometers (rigid and flexible) with a membrane osmometer (Wescor). We also compared the averaged values of the continuous measurement with osmotic pressure calculated from total protein and albumin fraction, using the Yamada equation. The rigid-needle and membrane osmometers showed excellent correlation (y = 1.00x + 0.06; r greater than 0.99). The flexible-needle osmometer tended to overestimate osmotic pressure (avg 16%). We used the rigid-needle osmometer for continuous measurements of protein osmotic pressure of blood and lymph in anesthetized or unanesthetized sheep to observe changes in protein osmotic pressure of blood and lymph through the three different interventions. The relationship between the theoretical values (x) and the continuous measurements (y) of osmotic pressure was good (y = 0.99x + 0.16, r = 0.97), but after various interventions, the continuously measured protein osmotic pressure tended to exceed the calculated measurements. The continuous measurement should be monitored with spot samples measured in a stationary osmometer or by calculation of osmotic pressure from total protein concentration and albumin fraction.

Effect of particles on sheep lung hemodynamics parallels depletion and recovery of intravascular macrophages.

We previously showed in newborn lambs that the pulmonary hemodynamic responses to foreign particulate matter (liposomes; Monastral blue) developed in parallel with the maturation of the pulmonary intravascular macrophage system. We now report our use of the liposome-encapsulated heavy-metal-chelating agent dichloromethylene diphosphonate to deplete the intravascular macrophages of small lambs. Functionally and by quantitative histology, we depleted the vast majority of the intravascular macrophages (71% by Monastral blue particle retention, n = 22; 77% by histology; n = 2). Depletion success increased to > 90% as we optimized the liposome-depletion regime. Recovery of the lung hemodynamic response began within 3 days. By 2 wk, the functional responses had fully recovered (n = 8), and, according to quantitative histology, the macrophage population (n = 2) had recovered 65%. Macrophage depletion in lambs is relatively inexpensive and easy to achieve. It is a safe procedure and is followed by full recovery in approximately 2 wk, provided that an aseptic technique is used to prevent bacteremia.

Intrathoracic sources of caudal mediastinal lymph node efferent lymph in sheep.

We investigated the intrathoracic contributions to the caudal mediastinal lymph node (CMN) efferent lymph in 12 anesthetized sheep after removing possible systemic contributions from below the diaphragm. We interrupted various pathways that may send lymph to the CMN (chest wall, esophagus, lung). Because the experiment is destructive, we did the resections in various combinations and waited 1 h between steps. Base-line CMN efferent lymph flow averaged 0.90 +/- 0.52 g/15 min (mean +/- SD). Cutting the pulmonary ligaments bilaterally caused lymph flow to decrease by an average of 58%. In five sheep, cauterizing around the lung hila reduced lymph flow by 16% of base line, cauterizing along the esophagus reduced lymph flow by 18% of base line, and cauterizing along the chest wall increased lymph flow by 6% of base line. After complete isolation of the node, except for the bronchoesophageal artery, dorsal mediastinal vein, and CMN efferent duct, 14% of base-line flow remained. The lymph-to-plasma total protein concentration ratios did not change significantly with any procedure. Under the conditions of our experiments, approximately 74% of base-line intrathoracic CMN efferent flow comes from the lung.

Long-term clearance of liquid and protein from the lungs of unanesthetized sheep.

We measured the removal of 100 ml of autologous serum from the air spaces and lungs of unanesthetized, spontaneously breathing sheep at 4, 12, and 24 h. In the first 4 h, there was a rapid clearance of the liquid volume (8.3%/h), similar to our results in anesthetized ventilated sheep (Matthay et al., J. Appl. Physiol. 53: 96-104, 1982). However, liquid removal progressively slowed to 3.3 and 1.4%/h at 12 and 24 h, respectively. In contrast, protein clearance (as measured by 125I-albumin instilled with the serum) was monoexponential and slow (1%/h). The slowing of liquid clearance appears to be a function of the rising protein osmotic pressure of the residual protein in the air spaces (protein concentration doubled in 24 h). Because protein solutions are chemotactic for neutrophils, we quantified the movement of liquid from the extracellular space into the alveolar compartment with a plasma protein tracer (131I-albumin), so that our final calculation of alveolar liquid clearance would take into account bidirectional movement of liquid across the alveolar barrier. The corrected values for net liquid clearance are slightly faster (less than 10% of the instilled volume).

Intravascular macrophage depletion attenuates endotoxin lung injury in anesthetized sheep.

We recently showed that we can selectively and safely deplete most (average 85%) of the pulmonary intravascular macrophages in sheep by intravenously infusing liposomes containing dichloromethylene bisphosphonate. After a 1-h stable baseline, we made a 6-h comparison after a 30-min intravenous endotoxin infusion (1 microg/kg) between six anesthetized control lambs and six anesthetized lambs in which the intravascular macrophages had been depleted 24 h previously. Three of the control lambs had been macrophage depleted and allowed to recover their intravascular macrophage population for >/=2 wk. After depletion, both the early and late pulmonary arterial pressure rises were dramatically attenuated. Our main interest, however, was in the acute lung microvascular injury response. The early and late rises in lung lymph flow and the increase in lung lymph protein clearance (lymph flow x lymph-to-plasma protein concentration ratio) were >90% attenuated. We conclude the pulmonary intravascular macrophages are responsible for most of the endotoxin-induced pulmonary hypertension and increased lung microvascular leakiness in sheep, although the unavoidable injury of other intravascular macrophages by the depletion regime may also contribute something.

Liquid thickness vs. vertical pressure gradient in a model of the pleural space.

In recent studies using relatively noninvasive techniques, the vertical gradient in pleural liquid pressure was 0.2-0.5 cmH2O/cm ht, depending on body position, and pleural liquid pressure closely approximated lung recoil (J. Appl. Physiol. 59: 597-602, 1985). We built a model to discover why the vertical gradient in pleural pressure is less than hydrostatic (1 cmH2O/cm). A long rubber balloon of cylindrical shape was inflated in a plastic cylinder. The "pleural" space between the balloon and cylinder was filled with blue-dyed water. With the cylinder vertical, we measured pleural pressure by a transducer through side taps at 2-cm intervals up the cylinder. The pressure was measured with different amounts of water in the pleural space. With a clear separation between the balloon and the container, the vertical gradient in pleural liquid pressure was hydrostatic. As water was withdrawn from the pleural space, the balloon approached the wall of the container. Over an 8-cm-long midregion of the model where the balloon diameter matched the cylinder diameter, the vertical gradient was not hydrostatic and was virtually absent. In this region, the pleural liquid pressure was uniform and equal to the recoil of the balloon. In this section we could not see any pleural space. By scintillation imaging using 99mTc-diethylenetriamine pentaacetic acid in the water, we estimated the thickness of this flat "costal" pleural space to be approximately 20 microns. Radioactive tracer injected at the top of the pleural space appeared by 24 h at the bottom, which indicated a slow drainage of liquid by gravity.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of interstitial edema on lung lymph flow in goats in the absence of filtration.

We tested the effect of interstitial edema on lung lymph flow when no filtration occurred. In 16 anesthetized open-thorax ventilated supine goats, we set pulmonary arterial and left atrial pressures to nearly zero and measured lymph flow for 3 h from six lungs without edema and ten with edema. Lymph flow decreased exponentially in all experiments as soon as filtration ceased. In the normal lungs the mean half time of the lymph flow decrease was 12.7 +/- 4.8 (SD) min, which was significantly shorter (P less than 0.05) than the 29.1 +/- 14.8 min half time in the edematous lungs. When ventilation was stopped, lymph flow in the edematous lungs decreased as rapidly as in the normal lungs. The total quantity of lymph after filtration ceased was 2.7 +/- 0.8 ml in normal lungs and 9.5 +/- 6.3 ml in edematous lungs, even though extravascular lung water was doubled in the latter (8.4 +/- 2.4 vs. 3.3 +/- 0.4 g/g dry lung, P less than 0.01). Thus the maximum possible clearance of the interstitial edema liquid by the lymphatics was 6.3 +/- 4.8%. When we restarted pulmonary blood flow after 1-2 h in four additional goats, lymph flow recovered within 30 min to the baseline level. These findings support the hypothesis that lung lymph flow originates mainly from alveolar wall perimicrovascular interstitial liquid and that the contribution of the lung lymphatic system to the clearance of interstitial edema (bronchovascular cuffs, interlobular septa) is small.

Equivalent pore estimate for the alveolar-airway barrier in isolated dog lung.

In high-pressure pulmonary edema, lung interstitial and air space edema liquids have equal protein concentrations (Am. J. Physiol. 231: 1466, 1976). This suggests that the alveolar-airway barrier separating the air and interstitial spaces is relatively unrestrictive, even without apparent epithelial injury. To estimate the equivalent pore population of the alveolar-airway barrier we inflated each of 18 isolated dog lung lobes for 1 h with a solution of colored tracer of uniform radius. Tracer radii ranged from 1.3 to 405 nm. After freezing the lobes in liquid N2, we measured interstitial tracer concentrations in frozen perivascular cuffs or in samples thawed after dissection from frozen cuffs. Relative to the concentrations instilled, interstitial concentrations ranged from 0.34 for the smallest particles (1.3 and 3.5 nm radius) to zero for particles with radii of 405 nm. From the results we designed a pore model of the alveolar-airway barrier to reproduce the concentrations we measured. No single-pore model could be obtained, although a three-pore model fit the data well. The model results predict that pores with radii of 1, 40, and 400 nm would account for 68, 30, and 2% of total liquid flux, respectively. The majority of liquid flux (68%) would occur through passageways smaller than the smallest tracer we used (1.3 nm radius). We believe the alveolar-airway barrier consists not only of tight intercellular junctions that allow passage of only water and electrolytes but also of a smaller number of large leaks that allow passage of particles up to nearly 400 nm in radius.

Micropuncture pressure in small arteries or veins of perfused rabbit lungs.

Until now, direct micropuncture measurements of vascular pressure in lung have been limited to small vessels less than 100 microns on the pleural surface. On the other hand, direct pressure measurements using small catheters (less than 1-mm OD) in pulmonary vessels have been limited to those greater than 1.2 mm. We measured pressure in intermediate-sized microvessels (300-700 microns) using the micropuncture method in isolated perfused rabbit lungs. These microvessels are located 2 or 3 mm beneath the pleura. We exposed them by microsurgery and punctured the relatively thick-walled vessels with specially configured micropipettes. We exposed one pulmonary microvessel in each rabbit lung by microsurgery on the left middle lobe. In 15 rabbit lungs we measured pressure in a total of six small arteries (275- to 470-microns diam) and nine small veins (300- to 700-microns diam) under high zone 3 conditions, near the zone 2/3 boundary. We found approximately 35% of the total pulmonary vascular pressure drop in arteries greater than 275-microns diam and 7% in veins greater than 300-microns diam. In veins greater than 500-microns diam, there was no measurable pressure drop. After the measurements, we froze the lung and confirmed that there was no detectable interstitial or alveolar edema in the cross sections of the punctured site. Our data are compatible with those of other investigators who have used isolated perfused rabbit lungs under similar experimental conditions.

Clearance of lung edema into the pleural space of volume-loaded anesthetized sheep.

To determine whether lung edema leaks into the pleural space, we measured flow rates of visceral pleural liquid from exposed sheep lungs during volume loading and then compared the protein concentration of visceral pleural liquid and lung interstitial liquids (lymph and peribronchovascular cuff liquid). For 4 h, we volume loaded 24 anesthetized ventilated sheep with one side, both sides, or neither side of the chest open. During the experiment, we collected visceral pleural liquid from a bag surrounding the exposed lung and lung lymph; after the experiment, we collected peribronchovascular cuff liquid. We found that during volume loading visceral pleural liquid flow increased significantly by 2 h, and its protein concentration over the final hour was the same as that of lung interstitial liquids. The volume of visceral pleural liquid correlated with excess lung water and wedge pressure elevation. By our estimates, clearance of edema from the lung into the pleural space constituted 23-29% of all edema liquid collected, similar to measured lymph edema clearance. We conclude that edema liquid leaks directly from edematous sheep lungs into the pleural space and that this leakage provides an important additional route of edema clearance.