Toxicological evaluation of textiles coated with antibacterial metal oxide nanoparticles by 2D and 3D in vitro skin model

Metal oxide (MeO) nanoparticles (NPs), such as copper (CuO) and zinc (ZnO) oxides, thanks to their antibacterial properties, are among the most eligible nanomaterials for the coating of textiles. Nevertheless, there is a general concern about their safety for human health. Within the EU funded H2020 project “PROTECT”, the skin toxicity of textiles coated with antibacterial CuO and ZnO nanoparticles was assessed by different in vitro approaches. CuO and ZnO NPs were produced by sonochemical process and characterized by TEM and DLS. Textiles, coated with ZnO and CuO NPs, were subjected to extraction procedure (ISO 10993-12), including 72 hours incubation in artificial sweat solution (AS, pH 4.7 and 6.3) at 37°C. The stability of the NPs at the different AS pHs, so as the release of NPs and/or ions from the textiles after the extraction procedure in AS, was evaluated through CPS and ICP-OES techniques. For the Skin Corrosion test, Epiderm™ 3D in vitro skin models (Mattek) were exposed to different concentration of CuO and ZnO in water, according to the OECD TG.431 protocol, while for the Skin Irritation test, the 3D models were exposed to the textile extracts, according to ISO/TC 194/WG 8 for Medical Devices. Balb/3T3 fibroblasts were also used to understand the mechanisms of action of the cytotoxicity trigged by CuO and ZnO NPs. Cell viability, inflammatory mediators release, morphological changes and wound healing process (scratch assay) were investigated. Data from the Corrosion test showed that CuO and ZnO NPs resulted non-corrosive up to 1000 ppm in water, according to the Globally Harmonized System (GHS) adopted by the OECD. For the Skin Irritation test, a significant reduction of tissue cells viability was induced by both metal oxide NPs extracted from textiles in AS pH 4.7, likely due to the high content of free Cu and Zn ions released in these conditions and detected by ICP-OES. At higher pH, the effects were observed at lower extent, due to the less solubility of NPs at these experimental conditions. Experiments on fibroblasts showed that ZnO NPs strongly affected cell viability and IL-8 release starting from the dose of 20 ppm. Moreover, the release of IL-8 resulted dose-dependent after the exposure to CuO NPs. Microscopy analyses showed that MeO NPs changed fibroblasts morphology and their ability to migrate during the wound healing process. All together, these data highlight that coated textiles seem to be safe on intact skin models, unless NPs dissolution in acid AS occurs. Nevertheless, further experiments are needed in order to understand the MeO NPs toxicity in case of wounded skin, as suggested by data on fibroblasts. In conclusion, NPs physical and chemical properties and appropriate in vitro tools are determinant parameters in order to assess NPs safety. Acknowledgements: This work was supported by EU funded H2020-720851 PROTECT project