Titanium dioxide nanoparticles induce COX-2 expression through ROS generation in human periodontal ligament cells

Titanium dioxide nanoparticles (TiO₂-NPs) are used to improve the aesthetic of toothpaste. While TiO₂-NPs have been used safely in toothpaste products for a long time, there haven't been studies to determine whether absorption of TiO₂-NPs by the mucous membranes in the mouth induces pathogenic conditions. Here, we assessed whether TiO₂-NPs induce cyclooxygenase-2 (COX-2) and investigated the molecular mechanisms underlying the pro-inflammatory effect of TiO₂-NPs on human periodontal ligament (PDL) cells. Treatment of PDL cells with TiO₂-NPs led to induction of both COX-2 mRNA and protein expression. TiO₂-NPs stimulated the nuclear translocation of nuclear factor-kappaB (NF-κB) as well as its DNA binding by inducing phosphorylation and subsequent degradation of the inhibitory protein IκBα in PDL cells. TiO₂-NPs treatment resulted in rapid activation of extracellular signal-regulated kinase (ERK)1/2 and Akt, which could be upstream of NF-κB. Treatment of PDL cells with both the MEK1/2 inhibitor U0126 and the PI3K inhibitor LY294002 strongly attenuated TiO₂-NPs-induced activation of NF-κB, and also the expression of COX-2. PDL cells treated with TiO₂-NPs exhibited increased accumulation of intracellular reactive oxygen species (ROS). Pretreatment of cells with ROS scavenger N-acetyl cysteine (NAC) abrogated the stimulatory effect of TiO₂-NPs on p65, p50, and COX-2 expression. In conclusion, ROS, concomitantly overproduced by TiO₂-NPs, induce COX-2 expression through activation of NF-κB signaling, which may contribute to the inflammatory effect of PDL cells.