Titanium dioxide nanoparticles (TiO2 NPs) are used in paints, plastics, papers, inks, foods, toothpaste, pharmaceuticals and cosmetics. However, TiO2 NPs cause inflammation, pulmonary damage, fibrosis and lung tumours in animals and are possibly carcinogenic to humans. Although there are a large number of studies on the toxicities of TiO2 NPs, the data are inconclusive and the mechanisms underlying the toxicity are not clear. In this study, we used the Comet assay to evaluate genotoxicity and whole-genome microarray technology to analyse gene expression pattern in vivo to explore the possible mechanisms for toxicity and genotoxicity of TiO2 NPs. Mice were treated with three daily i.p. injections of 50 mg/kg 10 nm anatase TiO2 NPs and sacrificed 4 h after the last treatment. The livers and lungs were then isolated for the Comet assay and whole genome microarray analysis of gene expression. The NPs were heavily accumulated in liver and lung tissues. However, the treatment was positive for DNA strand breaks only in liver measured with the standard Comet assay, but positive for oxidative DNA adducts in both liver and lung as determined with the enzyme-modified Comet assay. The genotoxicity results suggest that DNA damage mainly resulted from oxidised nucleotides. Gene expression profiles and functional analyses revealed that exposure to TiO2 NPs triggered distinct gene expression patterns in both liver and lung tissues. The gene expression results suggest that TiO2 NPs impair DNA and cells by interrupting metabolic homeostasis in liver and by inducing oxidative stress, inflammatory responses and apoptosis in lung. These findings have broad implications when evaluating the safety of TiO2 NPs used in numerous consumer products.