CONFOCAL IMAGING OF TIME-DEPENDENT INTERNALIZATION AND LOCALIZATION OF NBD-PC IN INTACT RAT LUNGS

Morphological localization of a fluorescent analogue of phosphatidylch oline (PC), C-12-NBD-PC palmitoyl-2-[12-(7-nitro-2-1,3-benzoxadiazol-4 -yl) amino] dodecanoyl}PC) was evaluated in adult rat lung. Fluorescen ce was observed at various times after intratracheal liposome instilla tion. Fluorometric analysis of C-12-NBD-PC in lung homogenate showed a n initial rapid uptake followed by a more gradual uptake phase. Micros copic observation of fixed lung slices showed C-12-NBD-PC uptake in al veolar type II cells (identified by surfactant protein A and Maclura p omifera agglutinin colocalization), nonciliated distal airway epitheli al cells, but not in subepithelial tissue. Macrophages showed an initi al uptake that did not increase with time. Fluorescence measured in ty pe II cells was diffuse at 5 min, became punctate, and intensified gra dually by 2 h. Isolation of type II cells and lamellar bodies from ins tilled lung confirmed localization of fluorophore in them. This study shows time-dependent uptake of C-12-NBD-PC by type II and bronchiolar cells and preferential internalization to lung lamellar bodies as prev iously described for surfactant PC.