The effect of vitamin E deficiency in male Sprague-Dawley rats upon lipid peroxidation in lung tissue was examined by measuring malonaldehyde and lipid epoxide production. In addition to controls, some animals were also exposed to 3 +/- 0.1 ppm NO2 continuously for 7 d in order to study the effects of oxidant stress on lung lipid peroxidation and vitamin E content. The observed changes in malonaldehyde and epoxide content could serve as good indices of lipid peroxidation, particularly under conditions of vitamin E deprivation. The responses measured indicated an inverse relation in the lung between tissue vitamin E content and quantity of lipid peroxidation products. Measurement of lipid epoxides served as a reliable indicator of lung tissue lipid peroxidation. Finally, NO2 inhalation appeared to elicit a response characterized by increased assimilation of vitamin E into lung tissue.

Phospholipids in plasma and erythrocytes of rats exposed to 100% oxygen for 3 days were analyzed. Exposed rats contained lower quantities of several phospholipids in both plasma and erythrocytes than in matched controls. Pronounced decreases were found in phosphatidylcholine and lysophosphatidyl choline. The proportion of disaturated species in erythrocyte phosphatidylcholine was significantly greater after oxygen exposure. There was an increase in percentage of saturated fatty acids with a proportionate decrease in percentage of unsaturated fatty acids. The increase in saturation was associated primarily with palmitate while decreased unsaturation involved primarily linoleic acid. In plasma the proportions of stearic acid increased while those of linoleic, arachidonic and docosahexaenoic acids decreased in phosphatidylcholine. Damage to cell lipids may account for increased hemolysis which occurs after exposure to high oxygen concentrations. Erythrocytes recovered following oxygen exposure contained phospholipid species commonly found in immature red blood cells.

Type II pneumocytes from fetal rabbit lungs were grown in an organotypic system and used to study surfactant phospholipid synthesis. This organotypic system was further employed as a means for isolating purified type II cells which were grown in monolayer cultures. Phospholipid synthetic properties for these purified type II cells at different stages of culture were studied using the radioactively labeled substrates: palmitate, choline and acetate. Additionally, fibroblasts were recovered from the organotypic system and grown in isolated cultures or in combination with type II cells. A comparison of the phospholipid composition and biosynthesis for these cell preparation indicated that in all instances, with the exception of fetal lung fibroblasts, the cultures were capable of producing saturated phosphatidylcholine as a major phospholipid product. The relative molar incorporation into phospholipids for the substrates studied differed depending on the type cell preparation being examined. The formation of surfactant phospholipids appeared most similar for the organotypic system and mixed fibroblast/type II cell preparations. Furthermore, a mixed culture of fibroblasts and type II cells produced larger proportions of surfactant phospholipids than type II cells alone. These observations indicate that the organotypic system is a useful model for examining fetal lung surfactant phospholipid synthesis and may also be employed as a simple means for isolating fetal type II pneumocytes.

Type-II (T-II) pneumocytes from 27 day fetal rabbit lungs were grown in an organotypic system (OS) and used to study phospholipid (PL) synthesis. Organotypic preparations were also used as a means for isolating purified T-II cells as well as lung fibroblasts. The PL synthetic properties of the various culture preparations were studied using palmitate, choline and acetate as precursors. In all cultures, excepting fibroblasts, saturated phosphatidylcholine (SPC) was a major PL product. The molar incorporation rate into PL for the substrates studied was: palmitate > choline > acetate. The proportion of these substrates incorporated into SPC versus total phosphatidylcholine (PC) differed between OS and monolayer cultures. In the OS the order of substrate incorporation into SPC as a percent of total PC formed, was acetate > choline > palmitate, whereas in T-II cultures the order was choline > acetate > palmitate. Among the culture preparations examined, the greatest similarity was found between the OS and the mixed fibroblast/T-II preparations. In this regard it was observed that a mixed culture of fibroblasts and T-II cells produced larger proportions of SPC and other surfaceactive PL than isolated T-II cells. From these observations it appears that the OS is a useful model for examining surfactant PL synthesis of T-II cells, and furthermore, may serve as an effective system for the isolation and study of T-II pneumocytes.

The activity of rat lung epoxide hydrolase (epoxide hydrolase, EC 3.3.2.3) was studied using two lipid epoxides which can be isolated from lung tissue. These epoxides displayed different Km,app and hydration rates. Methyl cis-9,10-epoxystearate was hydrated 20-times more rapidly than cholest-5α,6α-epoxy-3β-ol. The Km for the lung microsomal enzyme was variable and dependent on the microsome concentration in the medium. A soluble epoxide hydrolase was also detected in both lung and liver. This enzyme appears similar to the microsomal enzyme in its activity toward methyl epoxystearate. The measurement activities for liver microsomal epoxide hydrolase were over 8-times those for lung microsomes; activity against cholesterol epoxide was 40-times greater for liver. In spite of the slow rates measured with cholesterol epoxide in lung preparations, this compound was an effective competitive inhibitor against methyl epoxystearate over a wide concentration range. This suggests that cholesterol epoxide readily binds to epoxide hydrolase and is an effective competitive inhibitor against a much more actively metabolized substrate, methyl epoxystearate. Such circumstances indicate that cholesterol epoxide binds with a high degree of nonproductivity to lung microsomal epoxide hydrolase. This attribute of lung epoxide hydrolase may relate to the relatively high concentrations of cholesterol epoxide found in lung tissue.

Incorporation of labeled glucose and fatty acid residues into saturated phosphatidylcholine was significantly reduced in lung slices from 27.5 days of gestation fetal rabbits during 90 min incubation in the presence of 100 microU/ml insulin. When 14C-glucose was used as substrate, incorporation into both phosphatidylcholine and saturated phosphatidylcholine was reduced by insulin. This occurred despite an increase in overall glucose utilization by the lung from 11.3 +/- 3.9 to 16.3 +/- 5.2 nmole/g tissue in the presence of insulin (P less than 0.05). A decrease in incorporation of fatty acid residues into saturated phosphatidylcholine was also observed when 14C-palmitate was used as substrate, from 102 +/- 4 to 90 +/- 5 nmole palmitate/g tissue (P less than 0.01). In the presence of insulin, there were significant reductions of both substrates appearing in lysophosphatidylcholine, a precursor of saturated phosphatidylcholine. There was no significant change in incorporation of glucose residues into glycogen or lactate under these conditions.

Antepartum administration of aminophylline to pregnant rabbits resulted in accelerated formation of phospholipids regarded to be important components of pulmonary surfactant. Increases in the tissue content and synthesis of saturated phosphatidylcholine and phosphatidylinositol were found in lung slice preparations obtained from fetuses previously treated with aminophylline. Synthesis of phospholipids was measured using labeled palmitate, oleate, and glucose as precursors. The results showed increased de novo production of these lipids, and that glucose and glycogen may serve as important precursors. There was a significant reduction in triglyceride content while free fatty acids increased suggesting increased lipolysis in the aminophylline-treated groups, but this did not measurably affect the rate of labeled palmitate incorporation into total phospholipids.

Summary: Rabbits delivered at 27.0 days of gestation were studied after administration of cortisol (2 mg/kg/day), aminophylline (6.25 mg/kg/day), or sterile saline to the does on days 24–26 of gestation. Survival at 60 min was 52.9% in the aminophylline-treated group and 22.2% in the control and cortisol-treated groups with all animals being in a warm, oxygen-enriched environment and receiving frequent tactile stimulation. Lung volume at 30 cm H2O was lower in the cortisol-treated group than in the controls or aminophylline-treated group in animals surviving for 60 min (Table 2). The aminophylline-treated group retained significantly more gas at low pressures on the deflation curve (Table 2) and had significantly more phosphati-dylcholine recovered in lung lavage fluid (Table 3) than the other groups. Aminophylline appears to have enhanced lung maturation better than cortisol in this experimental model. Speculation: Improved survival of prematurely delivered rabbits after aminophylline administration (as compared with cortisol) may be due to a combination of factors including enhanced maturation of the lungs as well as stimulation of the respiratory center.

Fetal lung cyclic AMP phosphodiesterase, cyclic AMP, phosphatidyl choline, and incorporation of precursors into phosphatidyl choline were measured in rabbits after maternal administration of hydrocortisone phosphate and aminophylline. Both agents inhibited lung phosphodiesterase activity and augmented cyclic AMP concentrations (Table 1). Aminophylline administration was associated with a significant increase in lung saturated phosphatidyl choline (Table 2). Incorporation of [14C] choline and [3H] methionine was increased by both aminophylline and hydrocortisone (Table 3).