Surfactant Replacement Improves Lung Recoil in Rabbit Lungs after Acid Aspiration 1- 3

WAYNE J. E. LAMM and RICHARD K. ALBERT

Introduction Several observations have suggested that surfactant replacement might be an efficacious treatment for the adult respiratory distress syndrome (ARDS). Ashbaugh and colleagues (1) reported that surface activity was decreased in the first report describing ARDS, and they suggested that surfactant deficiency might be intrinsic to the pathophysiology of the syndrome. Subsequently, both qualitative and quantitative surfactant abnormalities have been found in fluid that has been lavaged or suctioned from the lungs of patients with ARDS (2-4). We (5) and others (6-8) have found that surfactant inactivation or depletion augments edema formation, probably by decreasing the interstitial hydrostatic pressure in one or more perivascular interstitial compartments (2, 9-12). Surfactant abnormalities might occur in ARDS because of one or more of the following problems: (1) direct epithelial damage (13), (2) pulmonary thrombosisassociated oligemia caused by metabolic dysfunction (as pulmonary arterial occlusion has been found to alter surfactant) (14), (3) surfactant destruction by lecithinases generated by bacteria (15, 16), (4) alveolar edema, hemoglobin, lipids, fibrinogen, fibrin monomers, and/or other surfactant inhibitor proteins gaining entry to the alveolar space as a result of the ARDS-associated increase in epithelial permeability (17- 23), (5) sequestration of surfactant because of hyaline membranes (24), and/or (6) displacement of surfactant into the airway (25). Surfactant replacement has a variety of beneficial effects in animal models of the infant respiratory distress syndrome (IRDS) (summarized in 26). In infants, a single intratracheal instillation of surfactant improves lung function for as long as 72 h (27), reduces the incidence of pneumothorax, has a preventive effect in infants at risk for the syndrome, and may reduce mortality in those more severely affected (26, 28). Surfactant replacement also improves the respiratory

SUMMARY We tested the hypothesis that surfactant replacement would be beneficial In the acldaspiration model of acute lung Injury. HCI (0.1 N, 2 mllkg) was Injected Into the trachea of excised 8). Control lungs (n 4) had no Intervention. All were perfused with lYrode's rabbit lungs (n solution mixed 1:1 with autologous whole blood at 40 mllmin/kg for 30 min, and then degassed. A modified natural surfactant (Survanta@, Ross Laboratories) was then Injected Into the trachea of four lungs Injured with HCI (100 mg/kg at 25 mg/ml). lWo quasi-static pressure-volume curves were determined. The mean alveolar pressures at 50,60,70,80, and 90% of TLC were greater In the HCI group than In the control group (p < 0.05). However, no difference was observed between the control lungs and those that received HCI + Survanta. In 13 anesthetized, paralyzed, and ventilated rabbits, deflation pressure-volume curvas were determined from TLC to FRC (measured by helium dilution). Then, 0.1 N HCI (3 ml/kg) was Injected into the trachea and, in seven, Survanta was Instilled 5 min later. The mean alveolar pressures at 60, 70, 80, and 90% TLC were higher at 15 and 60 min In the HCI group compared with their pre-HCI time point (p < 0.05). In the HCI + Survanta group, no differences were seen at 15 min, and only slight Increased were seen at 60 min. No effect of surfactant replacement on arterial blood gases was observed. HCI aspiration Increased recoil In both excised and in "i"o lungs, and surfactant replacement with Survanta returned recoil to normal. AM REV RESPIR DIS 1990; 142:1279-1283

=

=

distress induced by bilateral cervical vagotomy (29). These findings have stimulated studies of surfactant replacement in the oxygen toxicity model of subacute lung injury (30), and in theN-nitroso-Nmethylurethane model of subacute epithelial cell injury (31). The purpose of the present investigation was to determine if surfactant replacement improved the function of lungs acutely injured by acid aspiration. Methods New Zealand white rabbits weighing 2 to 3.5 kg were anesthetized intravenously with sodium pentobarbital (30 to 50 mg/kg). A tracheotomy was performed, and ventilation was started using a tidal volume of 25 to 35 ml and a rate of 20 breaths/min. Studies were done in excised lungs and in lungs in vivo. Excised Lung Studies Lung preparation. The thorax was opened, an intracardiac injection of heparin (1,000 units) was given, and the animals were killed by exsanguination. Ventilation was temporarily interrupted while cannulas were tied into the pulmonary artery and left atrium, and the lungs and heart were excised en bloc and suspended from a force transducer (FT03OC; Grass Instruments, Quincy, MA) in a humidified chamber warmed to 32° C. Ventilation was restarted with a positive end-expiratory

pressure of 2 cm H 20. The cannulas were connected to a perfusion pump, the vascular pressures were referenced to the most dependent part of the lung, and the vasculature was flushed at 40 mllmin/kg with 100 to 200 ml ofTRIS-buffered Tyrode's solution that had been warmed to 37° C. After the flush, perfusion was started with a 1:1 mixture of autologous whole blood and the buffered Tyrode's solution. Lung weight, end-inspiratory airway (Paw), and mean pulmonary arterial (Ppa) and venous (Ppv) pressures were amplified (model 1-183; Bell and Howell, San Dimes, CA) and recorded continuously. Ppv was set at approximately 5 cm H 2 0. The pH of the perfusate was kept at 7.4 ± 0.1 throughout the experiment. Lung weight and vascular pressures were allowed to stabilize for 10 to 15 min.

(Received in originaljorm September 12, 1989 and in revised jorm July 2, 1990) 1 From the Medical Service, Veterans Administration Medical Center, and the Department of Medicine, University of Washington, Seattle, Washington. 2 Supported by General Medical Research of the Veterans Administration and by Ross Laboratories. 3 Correspondence and requests for reprints should be addressed to Richard K. Albert, M.D., University of Washington Medical Center, Pulmonary and Critical Care Medicine, Mail Stop RM-12, Seattle, WA 98195.

1279

LAMM AND ALBERT

1280

Experimental Groups 1. Control (n = 4). Perfusion and ventila-

tion were maintained for 30 min with no intervention. 2. HCl (n = 4). With the lungs in the upright position ventilation was interrupted, and 0.1 N HCl (2 ml/kg) was injected into the trachea as a bolus, followed by 32 ml of air. Ventilation was restarted, and the perfusate pH was readjusted to 7.4 by adding sodium hydroxide. After 30 min of perfusion, all lungs (control and HCl) were disconnected from the ventilator and from the perfusion pump and placed in a Plexiglasill box for vacuum degassing (2 cycles). 3. HCl + Surfactant (n = 4). An additional four lungs were treated exactly as those given HCl with one addition: after degassing, a modified natural surfactant consisting of an organic solvent extract of minced cow lung supplemented with dipalmitoylphosphatidylcholine, palmitic acid, and tripalmitin (Survantaill; Ross Laboratories, Columbus, OH) was added. The Survanta was dispersed in aqueous saline (25 mg of phospholipid/mI), autoclaved for sterilization, and frozen for storage. At the time of use, Survanta was warmed to room temperature and instilled into the airways at a dose of 4 ml/kg.

Measurement of Lung Recoil After degassing (and surfactant instillation in four of the eight lungs that received HCl), lung recoil was assessed by two quasi-static pressure-volume (P-V) curves. Curves were obtained by inflating and deflating the lungs in lO-ml steps (except near TLC and Rv) every 10 s with a 15D-ml syringe. TLC was measured from the second deflation cycle and was considered to be that lung volume present at an alveolar pressure (PA) of 25 cm H 1 0. Lung volume was corrected for gas compression and, in the surfactant-treated group, for the volume of surfactant added. To compare the results obtained in different animals, lung volume was expressed as a percent of TLC. In Vivo Lung Studies Thirteen New Zealand white rabbits were anesthetized, intubated, and ventilated as described above, except that the gas used for ventilation was 1000/0 oxygen. They were placed supine on a heating pad set at 37° C, and cannulas were inserted into the left carotid artery to measure systemic blood pressure and to obtain arterial blood for blood gas (ABO) analysis, and into the carotid vein to administer medications. The rabbits were then paralyzed intravenously with Pavulon (0.1 mg/kg). Supplemental doses of pentobarbital and Pavulon were given as needed. Ventilation was interrupted, and control P-V curves were obtained by inflating the lungs rapidly (10 to 15 s) to PA = 40 cm H 1 0 (a pressure assumed to produce total lung expansion in this closed-chest situation) using 10% helium in oxygen. Paw was measured as the lung volume was decreased in lO-ml increments

from TLC to FRC. FRC was measured by helium dilution. The rabbits were then tilted to ahead-up position, and 3 ml/kgofO.1 N HCl, followed by 52 ml of air, were injected as a bolus into the trachea (time = 0 min). The rabbits were returned supine, and ventilation was resumed. Within the first 3 min all rabbits were given 45 mEq of sodium bicarbonate intravenously. At t = 5 min, Survanta (4 ml/kg at a concentration of 25 mg/mI) was instilled into the trachea of seven of the 13 rabbits in the same manner as the acid was given. All rabbits had P-V curves repeated at t = 15 and 60 min using 100% oxygen. ABO measurements were obtained prior to each P-V curve, and also at t = 30 min. On the final deflation curve (done at t = 60 min) the trachea was clamped at FRC, and the lungs were degassed. The chest was opened, and the lung and heart were removed en bloc. The left lung was tied off and used for wet/dry (W/D) weight ratio determination.

Distribution of Hel and Surfactant To determine if the method of instilling HCl and surfactant resulted in a uniform dispersion to all lobes, two additional rabbits were studied. HCl and surfactant were given to both in the identical fashion as that described above except that 0.14 ml of 99mTc DTPA (lC-99, 140 j.1Ci) was mixed with the HCl , and 0.1 ml of 670a (Ga-67, 100 j.1Ci) was mixed with the surfactant prior to instillation. One minute after surfactant was instilled, the trachea was clamped and the animal was given an overdose of pentobarbital. The chest was opened, and the vasculature was flushed clear of blood with 60 ml of saline. Each lobe was removed separately, divided into central and peripheral regions, counted in a gamma counter, dried, and weighed. Statistical Analysis Lung volume was plotted against PA. In excised lungs the PAS producing lung volumes of 50, 60, 70, 80, and 90% TLC in control lungs, HCl-injured lungs, and HCl-injured lungs given surfactant replacement were compared using analysis of variance followed by Fisher's least squares test. For the in vivo studies the PAS producing 50 to 90% TLC for control, 15, and 60 min were compared first by a two-way analysis of variance with repeated measures, then by Student's paired t test

at specific volumes. Body weights, lung volumes, WID ratios, and the ABO measurements at each time point were compared using Student's unpaired t test. Results

Excised Lung Studies No significant differences were observed between the body weights or the initial mean Ppa values or end-inspiratory Paw values in the three experimental groups (table 1). The changes in Ppa, Paw, and lung weight (Wt) gains during the 30 min of perfusion under control conditions and after HCI are shown in table 1. No changes were observed in the control group. The Ppa and Paw values markedly increased in the lungs receiving HCl. Despite subtracting the volume of acid injected into the trachea, the HCI-injured lungs had nearly a 2000/0 increase in lung weight (p < 0.001). In three lungs, HCI was mixed with Evans blue prior to the intratracheal injection. In these, blue staining of both central and peripheral regions of all lung lobes was observed. At 30 min the HCIinjured lungs that subsequently received Survanta had a degree of edema that was similar to, or greater than, those that received HCI without surfactant replacement (table 1). The mean deflation P-V curves from the second cycle for each of the three groups are shown in figure 1. No significant difference was observed between the absolute volumes observed at TLC or at a transpulmonary pressure of zero cm H 20 in the three groups (table 1). The P-V curve obtained from the HCI-injured lungs that did not receive surfactant replacement was shifted to the right (p < 0.05 for volumes of 50, 60, 70 and 80% TLC). The mean P-V curve of the acidinjured lungs that did receive surfactant replacement was shifted to a much lesser degree, and it was not found to be significantly different from that of the control group (p = NS at all lung volumes).

TABLE 1 EXCISED LUNG STUDIES·

--

Group Control HCI HCI + surfactant

Body Weight

Pulmonary Arterial Pressuret

Airway Pressuret:l:

Number

(kg)

(em H.O)

4 4 4

3.1 ± 0.2 3.2 ± 0.2 2.9 ± 0.1

3.2 ± 1.3 10.2 ± 5.4 12.3 ± 4.1

• Values are mean ± SEM. Change in pressure during course of experiment. Measured at end-inhalation. § p < 0.05 versus control value.

t

*

TLC

RV

(em H.O)

Weight Change (gl30 min)

(m/)

(m/)

0.4 ± 0.2 13.6 ± 0.8§ 13.1 ± 1.5§

0.0 ± 0.5 14.7 ± 3.1§ 21.4 ± 4.6§

102 ± 10 102 ± 8 101 ± 9

18 ± 5 14 ± 2 32 ± 9

1281

SURFACTANT REPLACEMENT IMPROVES LUNG RECOIL AFTER ACID ASPIRATION

100

90

\,\,,- /*

80 """

-r-----h"

70

0

",",~" ,,-..........

...I

I-

~

, , _ Acid-Aspiration

"""

-/....~

60

; ;

Fig. 1. Excised lung deflation P-V curves for the three groups. Only the HCI aspiration group was significantly shifted from the control group. Mean ± SEM, n = 4 for each group.

Acid-Aspiration + Surfactant Replacement

/ * I

LJ..

50

:' 1*

mean *SEM

*

/' I

40

P < 0.05 vs Control ANOVA

30 0

10

20

ALVEOLAR PRESSURE (cm HP)

In Vivo Lung Studies The body weights and the initial TLC and FRC values were similar in the animals that either did or did not receive surfactant replacement (table 2). The initial deflation P-V curves obtained under control conditions prior to HCI-aspiration were the same for the two groups (figure 2). The P-V curves of the lungs that received HCI without surfactant replacement were shifted to the right at 15 min (p < 0.05), and were shifted further by 60 min (P < 0.05). No P-V shift was observed at 15 min in the rabbits receiving surfactant replacement 5 min after H CI aspiration, and the shift at 60 min was slight (figure 2). HCI aspiration decreased the arterial pH and Pa0 2 while increasing the Pac02 and the calculated AaP02 (figure 3). Gas exchange was not improved in the rabbits given surfactant replacement. The WID ratios were similarly elevated in the HCI and the HCI + surfactant replacement groups even though approximately 12 m1 of additional fluid were instilled directly into the air space of the treated lungs as surfactant replacement (table 2). Addition of 12 ml of fluid to the wet weight of normal rabbit lungs not

subjected to this experimental protocol, and dividing the sum by the normal dry weight, we observe would increase the WID to approximately 11.

Distribution of Hel and Surfactant The distribution of HCI and surfactant to each of the lobes of both rabbits is shown in figure 4. Activity was normalized as a fraction of the total activity and dry weight of each lobe. The dispersion between lobes was rather uniform as was the distribution between central and peripheral regions (data not shown). All lobes received at least some HCI and some surfactant. Discussion

The important findings of this study are that with marked edema caused by HCI aspiration, surfactant replacement with Survanta (1) reduced the recoil of both excised and in vivo lungs, and (2) did not improve gas exchange in vivo. The increase in weight that occurred during the 30 min of perfusion after HCI aspiration could have resulted from increases in pulmonary vascular volume as well as from interstitial and/or alveolar edema. However, the substantial increase

TABLE 2 IN VIVO LUNG STUDIES·

Group HCI HCI + surfactant • Values are mean '" SEM.

Number 6

7

Body Weighl (kg)

TLC (m/)

Rv (m/)

WetlDry Weight

2.7 ± 0.2 3.0 ± 0.4

122 ± 6 122 ± 7

33 ± 6 31 ± 6

10.7 ± 0.7 10.6 ± 0.6

in Ppa seen after HCl aspiration suggests that pulmonary arterial constriction occurred since flow and Ppv were held constant. Without pulmonary venoconstriction (or a decrease in pulmonary venous compliance) it is likely that very little of the weight gain observed resulted from vascular distension and that the large majority represented edema. This suggestion is supported by our finding foam in the airways as well as by the markedly increased WID ratios. Because the lungs were ventilated at a constant volume, the increases in Paw that occurred after HCI aspiration could represent changes in airway resistance and/or ventilation at a higher lung volume. Because we observed a step change in Paw immediately after giving the HCI, we assumed that the majority of the increase resulted from the HCI filling some of the air spaces, causing the remaining alveoli to be ventilated at a higher volume. However, in addition to the step change in Paw, a progressive increase in Paw also occurred over time. This could have resulted from a subsequent, more gradually occurring, increase in airway resistance or from progressive alveolar edema. The P-V curves were obtained under quasi-static conditions so as to eliminate any potential effects of changes in airway resistance. The excised lungs receiving HCI were degassed prior to surfactant administration. Accordingly, the volume of surfactant administered was included in the value for TLC reported in table 1 for the animals in the group receiving HCI + surfactant. In all groups TLC was assumed to be present at a transpulmonary pressure of 25 cm H 20. In lungs receiving HCI + surfactant it took only approxiD;l.ately 90 ml of air to achieve this pressure, and the TLC was assumed to be 90 ml plus the volume of surfactant added to the air space. If all of the surfactant were absorbed out of the air spaces, this correction would have underestimated the true TLC by a maximum of 14 ml (the volume of surfactant given was 4 mllkg and the heaviest rabbit weighed 3.5 kg) or approximately 14070 (as the TLC values were approximately 100 ml). Such an underestimate would have caused an artifactual rightward shift of the P-V curve of those lungs receiving HCL + surfactant. This would bias the results against a beneficial effect of surfactant replacement. The TLC values from the in vivo lung experiments are summarized in table 2. For these it was not possible to measure TLC directly. TLC was considered to be

1282

LAMM AND ALBERT

100

air space. This underestimate would again bias the data against a beneficial surfactant effect as it would move the P-V curve to the right. The observation that the TLC values in the in vivo lungs were approximately 20070 greater than those measured in the excised lungs supports the possibility that the data collected from the excised lungs were biased against a beneficial effect of surfactant. For both the excised and in vivo studies we know of no mechanism by which the TLC could have been overestimated, thus resulting in an artifactual bias towards a beneficial surfactant effect. As opposed to the results of several other studies, we did not find that surfactant replacement improved the AaPo2 • Horbar and colleagues (27) found that the AaP02 did not begin to improve until approximately 36 h after surfactant replacement. Because our studies lasted only 1 h, they may have been too brief to see an improvement. However, several other groups have found that surfactant replacement acutely improves the AaP0 2 in infants with ARDS (32-35) and animals that were premature (36, 37) or had acute lung injury (31). Accordingly, we considered several other explanations for why the AaP02 remained unchanged. The improvement in the P-V curves that we observed indicates that lower trans-

90

80

0

~ 70 ;,!! 0

60

mean t.SEM • p < 0.05 vs Control Paired Student-t

50

40 0

10

5

15

20

25

o

5

10

15

25

20

ALVEOLAR PRESSURE (em H20) Fig. 2. In vivo lung deflation P-V curves for HCI-injured lungs that did not (A) or did (B) receive surfactant replacement. Curves were repeated 15 and 60 min after acid injury; n = 6 and 7 for A and B, respectively.

present at an alveolar pressure of 40 cm H 2 0. We elected to apply a greater alveolar pressure because the lungs were within a closed chest, and we wanted to as-

7.5

sure that we had reached the plateau of the P-V curves in the lungs in each of the three experimental conditions. As above, the volume of surfactant added was included when positioning the P-V curves on the volume axis. Again, these estimates of TLC would have been lower than the true value to the extent that surfactant was rapidly absorbed from the

PapH

0.8

I-

60

0.6

::I:

Cl

50

iii

:s:w

pac~ (torr)

40

III

0

30

...J Fig. 4. Distribution of HCI and surfactant by right upper (RU), right middle (RM), right lower (Rl), caudal (C), left upper (LU), and left lower (ll) lobes. Activity was normalized as a fraction of the total activity and dry weight of each lobe. Closed bars = HCI (TC99); hatched bars = surfactant (GA67).

600 500 400 paC:! 300 (torr) 200 100 I

500

~ CJ

-

en 'E

~

I-

«

'E

(,)

«

0.2

0.0

:::J

0

:>

::J

0

(ij

'0

-

0.8

Ul

:::J

0.6

0

g 0.4

:iE

A-a D0 400 2 (Iorr) 300

0::

mean",SEM

200 100

--

E

> t-

C W N

------

600

>

0:: C

0.4

0

20

40

60

TIME (min)

0

Z

0.2

0.0 RU

Fig. 3. Changes in arterial blood gases with time after HCI aspiration with and without surfactant replacement.

RM

RL

LOBE

C

LU

LL

SURFACTANT REPLACEMENT IMPROVES WNG RECOIL AFTER ACID ASPIRATION

pulmonary pressures would have been needed both to open atelectatic alveoli during inhalation and to keep these alveoli open during exhalation. If the shunt that occurs in ARDS results from perfusion of atelectatic alveoli, this effect should translate into improved gas exchange. However, the P-V curves that we reported were obtained after inflation to TLC. A disparity between the effects on the P-V curve and gas exchange could have resulted if the peak transpulmonary pressures developed during tidal breathing in vivo were insufficient to open the atelectatic alveoli. We elected to administer Survanta 5 min after HCI aspiration in an attempt to mimic a sequence of events that would be most optimal for observing a beneficial effect yet remain clinically realistic (i.e., administering surfactant as soon after the acute event as could occur after witnessing gastric aspiration in a hospital setting). If surfactant replacement must be given earlier than 5 min after the event resulting in ARDS to be efficacious, it is unlikely that it will be a clinically useful intervention for this condition. We cannot exclude the possibility that a greater effect might have been observed if we had delayed surfactant replacement. However, this would go against the observations in the pediatric literature suggesting that surfactant replacement is most beneficial when given before the first breath. Jacobs and colleagues (38) administered 1\veen~-20 or surfactant replacement to surfactant-deficient lambs and found that while both agents improved peak inspiratory pressures and dynamic compliances over those present in control animals, only surfactant replacement reduced the minimal surface tension of the alveolar wash and improved gas exchange. Accordingly, they suggested that the properties of surfactant considered to be essential for improving lung compliance were quite different from those resulting in improved gas exchange. The pathophysiology of IRDS suggests that surfactant replacement for a short period of time (one or two doses) may overcome the problem. However, the ongoing inflammation associated with ARDS implies that multiple doses over a more protracted interval might be needed. Accordingly, sensitization to the surfactant-associated proteins is a concern. Hull and Whitsett (39) have preliminary data showing that no antibodies to surfactant-associated proteins B or C have

been observed at 7,28, or 180 days in infants given surfactant replacement. References 1. Ashbaugh 00, Bigelow DB, Petty TL, Levine BE. Acute respiratory distress in adults. Lancet 1967; 2:319-23. 2. Petty TL, Reiss OK, Paul OW, Silvers OW, Elkins ND. Characteristics of pulmonary surfactant in adult respiratory distress syndrome associated with trauma and shock. Am Rev Respir Dis 1977; 115:531-6. 3. von Wichert P, Kohl FV. Decreased dipalmitoyl lecithin content found in lung specimens from patients with so-called shock-lung. Intensive Care Med 1977; 3:27-30. 4. Hallman M. Spragg R, Harrell HA, Moser KM, Gluck L. Evidence of lung surfactant abnormality in respiratory failure. J Clin Invest 1982; 70:673-83. 5. Albert RK, Lakshminarayan S, Hildebrandt J, Kirk W, Butler J. Increased surface tension favors pulmonary edema formation in anesthetized dogs' lungs. J Clin Invest 1979; 63:1015-8. 6. Bredenberg CE, Paskanik AM, Nieman GF. High surface tension pulmonary edema. J Surg Res 1983; 34:515-23. 7. Nieman OF, Bredenberg CEo High surface tension pulmonary edema induced by detergent aerosol. J Appl Physiol 1985; 58:129-36. 8. Jefferies AL, Kawano T, Mori S, Burger R. Effect of increased surface tension and assisted ventilation on 99mTc_IJfPA clearance. J Appl Physiol 1988; 64:562-8. 9. Beck KC, Lai-Fook S-J. Alveolar liquid pressure in excised edematous dog lungs with increased static recoil. J Appl Physiol 1983; 55:1277-83. 10. Bruderman I, Somers K, Hamilton WI(, Tholey WH, Butler J. Effect of surface tension on circulation in the excised lungs of dogs. J Appl Physiol 1964; 19:707-12. 11. Lloyd TC, Wright GW. Pulmonary vascular resistance and vascular transmural gradient. J Appl Physiol 1960; 15:241-5. 12. Sun RY, Nieman GF, Hakim TS, Chang HK. Effects of lung volume and alveolar surface tension on pulmonary vascular resistance. J Appl Physiol 1987; 62:1622-6. 13. De la Monte S, Hutchins GM, Moore OW. Respiratory epithelial cell necrosis in the earliest lesion of hyaline membrane disease of the newborn. Am J Pathol 1986; 123:155-60. 14. Shepard JW, Hauer D, Miyai K, Moser KM. Lamellar body depletion in dogs undergoing pulmonary artery occlusion. J Clin Invest 1980; 66:36-42. 15. Pattie RE, Burgess F. The lung lining film in some pathological conditions. J Pathol Bacteriol 1961; 82:315-31. 16. Sutnick AI, Soloff LA. Atelectasis with pneumonia: a pathophysiologic study. Ann Intern Med 1964; 60:39-46. 17. Johnson JWC, Permutt S, Sipple JH, Salem ES. Effect of intra-alveolar fluid on pulmonary surface tension properties. J Appl Physiol 1964; 19:769-77. 18. Holm BA, Notter RH, Findelstein IN. Surface property changes from interactions of albumin with natural lung surfactant and extracted lung lipids. Chem Phys Lipids 1985; 38:287-98. 19. Holm BA, Notter RH. Effects of hemoglobin and cell membrane lipids of pulmonary surfactant activity. J Appl Physiol 1987; 63:1434-42. 20. Fuchimukai T, Fujiwara T, Takahashi A, Enhoming O. Artificial pulmonary surfactant inhibited by proteins. J Appl Physiol 1981; 62:429-37.

1283 21. Se~ger W, Stohr G, Wolf HRD, Neuhof H. AlteratIOn of. su.rfactan~ function due to protein leakage: speCial mteractlOn with fibrin monomer J Appl Physiol 1985; 58:326-38. . 22. Ikegami M, Jobe A, Jacobs H, Lam R. A protein from the airways of premature lambs that inhibits surfactant function. J Appl Physiol 1984; 57:1134-42. 23. Ikegami M, Jobe A, Berry D. A protein that inhibits surfactant in respiratory distress syndrome. Bioi Neonate. 1986; 50:121-32. 24. Balis JU, Shelley SA, McCue MJ, Rappaport E. Mechanisms of damage to the lung surfactant system. Ultrastructure and quantitation of normal and in vitro inactivated lung surfactant. Exp Mol Pathol 1971; 14:243-62. 25. Said SI, Avery ME, Davis RK, Banerjee CM, EI-Oohary M. Pulmonary surface activity in induced pulmonary edema. J Clin Invest 1965; 44:458-64. 26. Jobe A, Ikegami M. Surfactant for the treatment of respiratory distress syndrome. Am Rev Respir Dis 1987; 136:1256-75. 27. Horbar JD, Stoll RF, Sutherland JM, et a/. A multicenter, placebo-controlled trial of surfactant therapy for respiratory distress syndrome. N Engl J Med 1989; 320:959-65. 28. Collaborative European Multicenter Study Group. Surfactant replacement therapy for severe neonatal respiratory distress syndrome: an international randomized clinical trial. Pediatrics 1988; 82:683-91. 29. Berry D, Ikegami M, Jobe A. Respiratory distress and surfactant inhibition following vagotomy in rabbits. J Appl Physiol 1986; 61:1741-8. 30. Matalon S, Holm BA, Notter RH. Mitigation of pulmonary hyperoxic injury by administration of exogenous surfactant. J Appl Physiol 1987; 67:756-61. 31. Harris JD, Jackson F Jr, Moxley MA, Longmore WJ. Effect of exogenous surfactant instillation on experimental acute lung injury. J Appl Physiol 1989; 66:1846-51. 32. Fujiwara T, Chida S, Watabe Y, Maeta H, Morita T, Abe T. Artificial surfactant therapy in hyaline-membrane disease. Lancet 1980; 1:55-9. 33. Hallman M, Merritt TA, Schneider H, et a/. Isolation of a human surfactant from amniotic fluid and a pilot study of its efficacy in respiratory distress syndrome. Pediatrics 1983; 171:473-82. 34. Oitlin JD, Soil FF, Parad RB. Randomized controlled trial of exogenous surfactant for the treatment of hyaline membrane disease. Pediatrics 1987; 79:31-;7. 35.. Davis JM, Veness-Meehan D, Notter RH, Bbutani VK, Kendig JW, Shapiro DL. Changes in pulmonary mechanics after the administration of surfactant to infants with respiratory distress syndrome. N Eng! J Med 1988; 319:476-9. 36. Jobe A, Ikegami M, Jacobs H, Jones S. Surfactant and pulmonary blood flow distributions following treatment of premature lambs with natural surfactant. J Clin Invest 1984; 73:848-56. 37. Ikegami M, Jobe A, Jacobs H, Jones SJ. Sequential treatments of premature lambs with an artificial surfactant and natural surfactant. J Clin Invest 1981; 68:491-6. 38. Jacobs HC, Berry D, Duane G, Ikegami M, Jobe AH, Jones S. Normalization of arterial blood gases after treatment of surfactant-deficient lambs with 1\veen 20. Am Rev Respir Dis 1985; 132:1313-8. 39. Hull WM, Whitsett JA. Immunologic analysis of infants receiving surfactant-TA (abstract). Pediatr Res 1988; 23(Suppl:411A).