The influence of suspension nebulization or instillation on particle uptake by guinea pig alveolar macrophages

Abstract

Phagocytosis represents a crucial event in the host defense against pathogens. Experimental methods are required that allow a range of particle doses to be delivered. However, it is not clear that these methods result in the same sites of deposition or mechanisms of clearance. The effect of particle administration by nebulization or instillation on the uptake by guinea pig alveolar macrophages ( AMs) has been studied. Suspensions of escalating quantities of 1-mum fluorescent polystyrene latex microspheres were delivered by 15 min of nebulization ( 1.4 x 10(7) -11.1 x 10(7) particles) or instillation (19 x 10(7) -152 x 10(7) particles) into the lungs of guinea pigs. These doses were selected to maximize delivery using each of these methods. Macrophages were collected by alveolar lavage 6 h postadministration. The total number of cells recovered was 3 x 10(6) and the cell viability was >97%, which was measured by trypan blue exclusion. Differential cell counts of lavaged cell suspensions were conducted and results showed no difference for the two methods of administration with various concentrations of latex particles and control samples. The uptake of particles was measured using epifluorescence, confocal microscopy, and flow cytometry. AMs showed a dose-dependent increase in associated particles measured by microscopy and flow cytometry. There was a direct correlation ( R-2 =.99) in the phagocytic indices ( PIs) measured by flow cytometry and fluorescence microscopy. The PI was 15 times higher after instillation than that obtained after particle nebulization. The percentage of AMs involved in phagocytosis observed after instillation was 55% and after nebulization 23%. The uptake of aerosolized particles by AMs and the number of cells involved in phagocytosis were dependent on the particle dose and the efficiency of aerosol delivery to the lungs