Role of necroptosis of alveolar macrophages in acute lung inflammation of mice exposed to titanium dioxide nanoparticles

Titanium dioxide (TiO₂) nanoparticles are indispensable for daily life but induce acute inflammation, mainly via inhalation exposure. TiO₂ nanoparticles can be phagocytosed by alveolar macrophages (AMs) <i>in vivo</i> and cause necroptosis of exposed cells <i>in vitro</i>. However, the relationship between localization of TiO₂ nanoparticles in the lungs after exposure and their biological responses including cell death and inflammation remains unclear. This study was conducted to investigate the intra/extracellular localization of TiO₂ nanoparticles in murine lungs at 24 h after intratracheal exposure to rutile TiO₂ nanoparticles and subsequent local biological reactions, specifically necroptosis of AMs and lung inflammation. We found that TiO₂ exposure induced leukocyte migration into the alveolar region and increased the secretion of C-C motif ligand (CCL) 3 in the bronchoalveolar lavage (BAL) fluid. A combination of Raman spectroscopy and staining of cell and tissue samples confirmed that AMs phagocytose TiO₂. AMs that phagocytosed TiO₂ nanoparticles showed necroptosis, characterized by the expression of phosphorylated mixed lineage kinase domain-like protein and translocation of high mobility group box-1 from the cell nucleus to the cytoplasm. In primary cultured AMs, TiO₂ also induced necroptosis and increased the secretion of CCL3. Necroptosis inhibitors suppressed the increase in CCL3 secretion in both the BAL fluid and culture supernatant of AMs and suppressed the increase in leukocytes in the BAL fluid. These data suggest that necroptosis of AMs that phagocytose TiO₂ nanoparticles is involved as part of the mechanism by which TiO₂ induces acute lung inflammation.