Endocytosis of ultrafine particles by A549 cells

Alveolar epithelium's capacity to ingest inhaled ultrafine particles is not well characterized. The objectives of this study were to use an in vitro m odel of type II lung epithelium and evaluate the cells' ability to take up ultrafine particles (titanium dioxide [TiO2], 50 nm diameter). The human ep ithelial cell line A549 was grown on aclar substrates and exposed to 40 mug /ml TiO2 particles for 3, 6, and 24 h before imaging with energy-filtering transmission electron microscopy. Elemental mapping and electron energy los s spectroscopy were used to colocalize Ti/O with electron-dense particles. Particle endocytosis was compared in A549 cells with and without pretreatme nt with cytochalasin D (cyto a) (2 mug/ml). After 3 h of TiO2 exposure, cel ls internalized aggregates of the ultrafine particles which were observed i n cytosolic, membrane-bound vacuoles. After 24 h of exposure there were con siderably more intracellular aggregates of membrane-bound particles, and ag gregated particles were also enmeshed in loosely and tightly packed lamella r bodies. Throughout 24 h of exposure a preponderance of particles remained associated with the free surface of the cells and were not internalized. T he majority of membrane-bound vacuoles contained aggregates of particles an d only occasionally did they contain as few as two or three particles, desp ite the use of several different approaches to assure the possibility for i ndividual particles to be ingested and detected. There was morphologic evid ence of microfilament disturbance, but no evidence of a decrease in interna lized particles in cells pretreated with cyto D. Thus, this model of type I I epithelium is able to internalize aggregates of ultrafine particles.