Metabolic and ultrastructural characterization of guinea pig alveolar type II cells isolated by centrifugal elutriation

Abstract

Direct biochemical studies of the whole lung have been quite misleading because of the heterogeneity of the lung cell types. One of the advantages of studying the isolated cells is to be able to correlate specific metabolic functions with intracellular molecular events and to differentiate factors that affect the type II cell function directly. In the present study we have isolated type II cells from guinea pig lung with elastase and purified them by centrifugal elutriation. These cells fluoresce with phosphine 3R as the dye is specifically taken up by the lamellar bodies. In the electron micrographs, the type II cells display punctate villi, which underwent fragmentation in those cases where metrizamide density gradient was used. Mitochondria are scattered throughout the cytoplasm, and smooth endoplasmic reticulum is sparse. Type II cells possess large irregularly shaped nuclei with peripheral areas of dense hemochromatin and at least one prominent nucleolus. Ovoid lamellar bodies are the most prominent cellular inclusions. These bodies are present throughout the cytoplasm and contain a substructure of whorling and concentric laminations. Biochemical studies indicate that type II cells prepared by centrifugal elutriation are metabolically well preserved as seen from incorporation of [ 14C]leucine into cellular proteins, methyl- 14C]choline into cellular disaturated phosphatidylcholine and CDP[ methy- 14C]choline into mitochondrial and microsomal phosphatidylcholine. Superiority of centrifugal elutriation over the commonly employed combination of discontinuous metrizamide gradient and cell elutriation is evident from the present study.