Contrasting macrophage activation by fine and ultrafine titanium dioxide particles is associated with different uptake mechanisms

ABSTRACT: Inhalation of (nano)particles may lead to pulmonary inflammation. However, the precise mechanisms of particle uptake and generation of inflammatory mediators by alveolar macrophages (AM) are still poorly understood. The aim of this study was to investigate the interactions between particles and AM and their associated pro-inflammatory effects in relation to particle size and physico-chemical properties. NR8383 rat lung AM were treated with ultrafine (uf), fine (f) TiO2 or fine crystalline silica (DQ12 quartz). Physico-chemical particle properties were investigated by transmission electron microscopy, elemental analysis and thermogravimetry. Aggregation and agglomeration tendency of the particles were determined in assay-specific suspensions by means of dynamic light scattering. All three particle types were rapidly taken up by AM. DQ12 and ufTiO2, but not fTiO2, caused increased extracellular reactive oxygen species (ROS), heme oxygenase 1 (HO-1) mRNA expression and tumor necrosis factor (TNF)-alpha release. Inducible nitric oxide synthase (iNOS) mRNA expression was increased most strongly by ufTiO2, while DQ12 exclusively triggered interleukin (IL) 1beta release. However, oscillations of intracellular calcium concentration and increased intracellular ROS were observed with all three samples. Uptake inhibition experiments with cytochalasin D, chlorpromazine and a Fcgamma receptor II (FcgammaRII) antibody revealed that the endocytosis of fTiO2 by the macrophages involves actin-dependent phagocytosis and macropinocytosis as well as clathrin-coated pit formation, whereas the uptake of ufTiO2 was dominated by FcgammaIIR. The uptake of DQ12 was found to be significantly reduced by all three inhibitors. Our findings suggest that the contrasting AM responses to fTiO2, ufTiO2 and DQ12 relate to differences in the involvement of specific uptake mechanisms.