Investigation of the in vitro genotoxicity of two rutile TiO2 nanomaterials in human intestinal and hepatic cells and evaluation of their interference with toxicity assays

TiO2 nanomaterials (NM) have a wide range of industrial applications, including their use in food products. The incorporation of these NMs in consumer products represents a clear concern for public health safety agencies and consumers, and further investigation of the potential impact of these products on human health is necessary. Indeed, since human oral exposure to TiO2 NMs is expected to increase in the years to come, there exist legitimate concerns about the risk assessment of these nanomaterials present in food products. A considerable amount of studies investigating the adverse effects of TiO2 NMs have focused on the genotoxic effects of these NMs, and more recently they have been classified by the International Agency for Research on Cancer (IARC) as carcinogen group 2B following inhalation studies (IARC, 2010). While numerous data are available for anatase or mixes of anatase/rutile forms, the toxicity and the genotoxicity of rutile TiO2 NMs have been rarely investigated. The aim of our study was therefore to investigate the cytotoxic and genotoxic effects of two rutile TiO2 NMs, differing in surface coating, NM103 (hydrophobic) and NM104 (hydrophilic), on intestinal and hepatic cell models. Following 3 or 24 h treatments with concentrations of TiO2 NMs from 1.2 to 80 μg/cm2, we have assessed the genotoxicity of these NMs with γH2AX, alkaline comet assay and micronucleus (MN) assays. Cellular viability and effects on oxidative stress were also evaluated. Although TEM imaging demonstrated the presence of the two TiO2 NMs within the cytoplasm, no significant cytotoxic or genotoxic were observed in either cell model. We have also evaluated and taken into account a variety of potential sources of interference of NMs with cellular assays. TiO2 NMs present in the cytoplasm introduce uncertainty in the scoring of micronuclei, and therefore this assay is not recommended for the evaluation of the genotoxicity of TiO2 NMs, or other NMs demonstrating similar interference. The unique properties of TiO2 NMs introduce additional complexity for genotoxicity testing, and caution must be taken in order to obtain reliable results necessary for accurate hazard assessment.