Lung Lavage Phospholipid Research Articles

Alveolar type II cell abnormalities and peroxide formation in lungs of rats given IL-1 intratracheally.

Acute lung injury (ALI) is characterized by increased lung levels of proinflammatory cytokines, inflammation, oxidative stress, edema, and impaired gas exchange. Notably, ALI patients also exhibit pulmonary surfactant abnormalities, including increased levels of phospholipids in their lung lavages. In the present study, to assess early alterations of the lung surfactant system in ALI, we induced inflammation and acute lung injury in rats by administering interleukin-1alpha (IL-1) intratracheally. Five h after IL-1 instillation, we examined lung tissue ultrastructure by electron microscopy using both routine staining methods and cerium chloride staining to localize hydrogen peroxide (H2O2) histologically. We also measured lung lavage phospholipid levels, lung tissue gamma-glutamyl transpeptidase (GGT) activities (a marker of oxidative stress), and arterial blood oxygen tensions. We observed that lungs of rats given IL-1 intratracheally had increased neutrophil accumulation, increased H2O2 production, and increased alveolar type II (ATII) pneumocyte ultrastructural abnormalities compared to rats given saline intratracheally. Intratracheal instillation of IL-1 also increased phospholipid levels in the bronchoalveolar lavage (BAL), possibly as a consequence of the abnormal discharge of lamellar bodies into the alveolar lumen. In addition, IL-1-insuffated rats had increased lung GGT levels and impaired blood oxygenation compared to saline-insufflated rats. Treatment with mepacrine decreased lung neutrophil accumulation, ultrastructural lung abnormalities, lung lavage phospholipid levels, lung tissue GGT levels, and blood oxygenation impairment in rats given IL-1 intratracheally, suggesting a possible relationship between these events. Our results indicate that IL-1-induced acute lung injury in rats is marked by neutrophil-dependent oxidative stress, ATII cell defects, abnormal discharge of lamellar body phospholipids, and impaired blood oxygenation.

Mepacrine decreases lung leak in rats given interleukin-1 intratracheally.

We hypothesized that phospholipase A2 (PLA2) metabolites contribute to the acute, neutrophil-dependent, edematous lung leak that develops after administration of interleukin-1 (IL-1) intratracheally to rats and tested this premise by using mepacrine to inhibit PLA2 activity in vivo. We found that lung PLA2 activity, lung lavage phospholipid content, lung leak index, lung weight gain, and lung lavage protein concentrations were increased in rats given IL-1 intratracheally compared with sham-treated control rats. By comparison, lungs of mecaprine and IL-1-treated rats had decreased PLA2 activity, lavage phospholipid content, leak, weight gain, and lavage protein increases compared with rats given IL-1 intratracheally. Mepacrine treatment also decreased lung neutrophil accumulation, but not lung lavage cytokine-induced neutrophil chemoattractant (CINC) levels, in rats given IL-1 intratracheally. In parallel experiments, mepacrine treatment reduced the adhesion of human neutrophils to IL-1-treated human umbilical vein endothelial cells in vitro. Our results indicate that PLA2 activity participates in the lung neutrophil retention and pulmonary vascular leak that develops in rats given IL-1 intratracheally.

Impaired Regulation of Surfactant Phospholipid Metabolism in the Isolated Rat Lung after Nitrogen Dioxide Inhalation

Various drugs have been shown to stimulate surfactant phospholipid metabolism. Particularly β-adrenergic agonists play an important role under physiologic conditions. For the first time we have studied whether nitrogen dioxide (NO2) inhalation alters β-adrenergic regulation of surfactant phospholipid metabolism in the model of the isolated lung. Rats were continuously exposed in vivo to a 5 ppm NO2-containing atmosphere for 48 hr. The lungs were isolated and perfused in presence of the β-adrenergic agonist dopexamine and surfactant metabolism was studied in three lung compartments: (1) lung lavage, (2) lung tissue, and (3) lavagable free alveolar cells. We found that (1) in normal rat lungs dopexamine increased the incorporation of palmitate and choline from the perfusate into lung lavage phospholipids. In nitrogen dioxide exposed rat lungs β-adrenergic stimulation did not cause an increase in precursor incorporation. No significant difference in unstimulated precursor incorporation was found for normal and NO2-exposed rat lungs. (2) Lung tissue from rats exposed to NO2 showed a decreased precursor incorporation into disaturated phosphatidylcholine due to an augmented cellular pool size. (3) Lavagable alveolar cells showed an increased palmitate uptake after nitrogen dioxide inhalation and by β-adrenergic stimulation. From these data we conclude that nitrogen dioxide inhalation impairs the β-adrenergic regulation of surfactant phospholipid metabolism. Moreover these data underline the importance of β-adrenergic agonists in surfactant metabolism.

Lung surfactant replacement in premature lambs with extracted lipids from bovine lung lavage: effects of dose, dispersion technique, and gestational age.

Extracted bovine calf lung lipids (CLL) with minimal protein (approximately 1%) were instilled prior to ventilation in groups of premature lambs of average gestational ages of 127 and 133 days. Aqueous dispersions of CLL were prepared by two techniques prior to instillation: sonication in an ice bath (S) and mechanical vortexing at room temperature (V). A low surfactant dose (15 mg CLL/kg animal weight) and a high dose (100 mg/kg) were investigated for each dispersion technique. Following tracheal instillation of surfactant, lambs were ventilated with 100% oxygen for 2 h with umbilical circulation intact, and for up to an additional 10 h after separation. A clear improvement in blood oxygenation and lung compliance was found over controls for lambs given 15 mg/kg and 100 mg/kg CLL(V), and 100 mg/kg CLL(S). Lambs treated with 15 mg/kg CLL(S) failed to improve over controls. Experimental groups treated with equal doses of CLL(V) and CLL(S) had similar amounts of lung lavage phospholipid, with values progressively declining during ventilation. Analyses of in vitro surface properties showed that both vortexed and sonicated CLL dispersions adsorbed to equilibrium surface pressures of 45-47 dynes/cm in seconds at concentrations greater than or equal to 0.25 mg CLL/ml. Both dispersions also lowered surface tension to less than 1 dyne/cm under dynamic compression at 37 degrees C in 100% humidity, although CLL(V) showed some enhancement over CLL(S) in dynamic surface activity at low subphase concentration (0.5 mg/ml). Moreover, CLL(V) and CLL(S) differed markedly in their effects on pressure-volume mechanics in a surfactant-deficient excised rat lung model. Instilled CLL(V) dispersions improved excised lung pressure-volume mechanics at significantly lower concentrations than CLL(S) dispersions.

Vitamin E inhibition of the effects of hyperoxia on the pulmonary surfactant system of the newborn rabbit.

The effects of vitamin E on the neonatal surfactant system were studied in rabbits exposed to air or hyperoxia (greater than 95%) from birth through 48 h of life. Hyperoxia exposure resulted in lung lavage phospholipid content which reached only 74% of air-exposed controls, and static pressure:volume observations of decreased maximum distensibility and altered compliance. Treatment with vitamin E (100 mg/kg of dl-alpha-tocopherol S.Q.) at 1 and 24 h of life was shown to completely abolish these effects of hyperoxia. Morphometrically determined alterations in epithelial cell composition and erythrocyte-contaminated air spaces resulting from hyperoxia exposure were also absent in pups treated with vitamin E. These findings suggest that early vitamin E treatment in vitamin E-deprived newborns prevents hyperoxia-associated compromise to the pulmonary surfactant system and selected other aspects of oxygen-induced lung injury in the neonate.

Postnatal increase in airway surfactant in the premature rabbit exposed in utero to betamethasone.

In utero exposure to betamethasone on days 26 and 27 of gestation resulted in a significant increase in lung lavage phospholipid after 24 h of air breathing in rabbit pups delivered at 29 days of gestation. At 6 h of age, the proportion of type II epithelial cells in betamethasone-exposed newborns was nearly twice the saline-exposed control value. Phospholipid composition and pressure:volume analysis of compliance were both unaffected by betamethasone treatment. The betamethasone-associated increase in airway phospholipid content was observed only in female pups. These data support the conclusion that antenatal betamethasone treatment elicits an increase in airway phospholipid which is dependent on breathing and confined to the female newborn.

1715 EFFECT OF INSULIN (I) AND ISOXSUPRINE (X) ON LUNG SURFACTANT PRODUCTION IN THE FETAL RHESUS MONKEY

We examined the effect of chronic hyperinsulinemia on lung lavage phospholipid (PL) content and composition. I (19 U/d for 20 d) was administered to fetal rhesus monkeys with an Alzet minipump implanted in the hind leg (Diabetes 28: 1058, 1979). Shamand non-operated animals were used as controls (C). X (5-10 mg/d for 20 d) was administered to additional mothers. Fetuses were delivered by cesarean section at 134-147 d (term is 165 d). I levels were 49±14 and 4470±780 μU/ml and glucose levels were 43±6 and 21±6 mg/dl in the C and I-treated groups respectively. Results are shown in the table. Hyperinsulinemia did not inhibit surfactant production. This suggests that the increased incidence of RDS in infants of diabetic mothers may not be due to hyperinsulinemia but rather possibly to the concomitant hyperglycemia. X increased surfactant production. X may be useful in the prevention of RDS in humans. Supported by HD-11018 and HD-11343.

Thyrotropin-releasing hormone increases the amount of surfactant in lung lavage from fetal rabbits.

Administration of thyrotropin-releasing hormone (TRH) to pregnant rabbits at 25 and 26 days of gestation results in increased pulmonary surfactant production by the fetus at 27 days (full term is 31 days). There was 60% more total phospholipid and 150% more phosphatidylcholine (the major component of surfactant) in the lung lavage from the fetuses in the treated group than in that from the controls. Lung lavage from the fetuses in the treated litters contained 13.4 +/- 1.6 micrograms of total phospholipid phosphorus/g lung dry wt and 5.6 +/- 1.1 micrograms of phosphatidylcholine phosphorus while that from the fetuses in the control litters contained only 8.2 +/- 1.1 micrograms and 2.2 +/- 0.4 micrograms, respectively. The phosphatidylcholine/sphingomyelin ratio increased from 1.0 in the lavage from the controls to 2.2 in that from the treated group. These changes in lung lavage phospholipid content and composition are in the direction of increased lung maturation. TRH administration had no effect on the incorporation of choline into phosphatidylcholine in fetal lung slices. These data suggest that TRH stimulates surfactant release rather than synthesis. TRH has a physiologic role in fetal lung maturation and surfactant production. It may potentially be used in the prevention of the respiratory distress syndrome in humans.

Stimulation of Surfactant Production by Oxytocin-Induced Labor in the Rabbit

The respiratory distress syndrome is believed to be due to insufficient surfactant. It is known that there is a greater incidence of the respiratory distress syndrome among infants delivered by cesarean section before labor than among those delivered after labor at the same gestational age. The purpose of this study was to determine the effect of labor on the production of pulmonary surfactant. We measured the phospholipid content of lung lavage in newborn rabbits delivered by cesarean section before labor at 29, 30, and 31 (full-term) days gestation and after oxytocin-induced labor at 31 days. We also measured the activities of pulmonary cholinephosphate cytidylyltransferase and choline-phosphotransferase, enzymes involved in the de novo synthesis of phosphatidylcholine, the major component of surfactant. There was a two- to fourfold increase in the amount of lung lavage phospholipid during the first 6 h after birth. This was not dependent upon gestational age at delivery. There was a further two- to fourfold increase in the next 18 h which was, however, dependent upon gestational age. Labor increased the amount of lavage phospholipid from rabbits delivered at full term by 132%, 177%, and 50% at 3, 6, and 24 h after birth, respectively. There was a postnatal increase in the activity of cholinephosphate cytidylyltransferase. This was almost linear with time during the first 12 h, by which time essentially adult values were attained. Choline-phosphate cytidylyltransferase was not affected by labor. There was also a postnatal increase in the activity of cholinephosphotransferase but this was stimulated 86%, 59%, and 21% by labor at 0, 1, and 24 h after birth, respectively. These studies suggest that labor stimulates both the synthesis and secretion of surfactant in the immediate postnatal period and thus may be an important factor in the prevention of the respiratory distress syndrome of the newborn.

The influence of postnatal nutritional deprivation on the phospholipid content of developing rat lung

It has been previously reported that fasting may result in decreased lung surfactant production. In order to investigate this relationship and the role of nutrition in lung phospholipid synthesis, 21-day-old rats were exposed for 60 h to one of five dietary regimens: standard rat chow (controls), fasting, pure glucose, pure fat, or pure protein. After the period of fasting there was a 33% decrease in lung protein content, but there was no change in DNA content. Exposure to any of the experimental diets resulted in a decrease in tissue total phospholipid and phosphatidylcholine content per lung, but not per unit lung protein. Similarly lung lavage phospholipid and phosphatidylcholine content was decreased by 25% after fasting when expressed per lung or per unit DNA, but not per unit protein. Pulmonary cholinephosphotransferase (EC 2.7.8.2) activity was decreased in the fasted animals and those fed the protein diet, but not in the glucose or fatfed animals. The activities of acetyl-CoA carboxylase (EC 6.4.1.2) and microsomal fatty acid elongation were decreased in all the experimental groups except for the glucose-fed group. It is concluded that fasting results in a decrease in lung cell size but not in lung cell number. Total phospholipid and phosphatidylcholine content in lung tissue and lung lavage is decreased per cell but not per unit cell mass.