Toxicology Letters 222 (2013) 55–63 Contents lists available at ScienceDirect Toxicology Letters jou rn al hom epage: www.elsevier.com/locate/toxlet Multi-platform genotoxicity analysis of silver nanoparticles in the model cell line CHO-K1 Xiumei Jiang a,b , Rasmus Foldbjerg b , Teodora Miclaus c , Liming Wang a , Rajinder Singh d , Yuya Hayashi c , Duncan Sutherland c , Chunying Chen a,∗ , Herman Autrup b , Christiane Beer b,∗∗ a National Center for Nanoscience and Technology, Chinese Academy of Science, No. 11, Beiyitiao Zhongguancun, Beijing 100190, China b Department of Public Health, Aarhus University, Bartholins Alle 2, 8000 Aarhus C, Denmark c iNANO, Aarhus University, Gustav Wieds Vej 14, 8000 Aarhus C, Denmark d Cancer Biomarkers and Prevention Group, Department of Cancer Studies and Molecular Medicine, University of Leicester, Leicester LE1 7RH, UK a r t i c l e i n f o Article history: Received 28 May 2013 Received in revised form 9 July 2013 Accepted 9 July 2013 Available online 18 July 2013 Keywords: Genotoxicity Cytotoxicity Silver nanoparticle DNA adduct 8-oxodG Micronucleus assay a b s t r a c t Investigation of the genotoxic potential of nanomaterials is essential to evaluate if they pose a cancer risk for exposed workers and consumers. The Chinese hamster ovary cell line CHO-K1 is recommended by the OECD for use in the micronucleus assay and is commonly used for genotoxicity testing. However, studies investigating if this cell line is suitable for the genotoxic evaluation of nanomaterials, including induction of DNA adduct and micronuclei formation, are rare and for silver nanoparticles (Ag NPs) missing. There- fore, we here systematically investigated DNA and chromosomal damage induced by BSA coated Ag NPs (15.9 ± 7.6 nm) in CHO-K1 cells in relation to cellular uptake and intracellular localization, their effects on mitochondrial activity and production of reactive oxygen species (ROS), cell cycle, apoptosis and necro- sis. Ag NPs are taken up by CHO-K1 cells and are presumably translocated into endosomes/lysosomes. Our cytotoxicity studies demonstrated a concentration-dependent decrease of mitochondrial activity and increase of intracellular reactive oxygen species (ROS) in CHO-K1 cells following exposure to Ag NPs and Ag + (0–20 g/ml) for 24 h. Annexin V/propidium iodide assay showed that Ag NPs and Ag + induced apoptosis and necrosis, which is in agreement with an increased fraction of cells in subG1 phase of the cell cycle. Genotoxicity studies showed that Ag NPs but also silver ions (Ag + ) induced bulky-DNA adducts, 8-oxodG and micronuclei formation in a concentration-dependent manner, however, there were quan- titative and qualitative differences between the particulate and ionic form of silver. Taken together, our multi-platform genotoxicity and cytotoxicity analysis demonstrates that CHO-K1 cells are suitable for the investigation of genotoxicity of nanoparticles like Ag NPs. © 2013 Elsevier Ireland Ltd. All rights reserved. 1. Introduction Advances in nanoscience and technology prompt the pro- duction of various kinds of nanomaterials. Due to their unique physicochemical properties, nanomaterials show great potential for biomedical and industrial applications. Interest on interaction studies of nanomaterials and biological systems has been increas- ing in recent years (Cheng et al., 2013; Zhao et al., 2011). The toxicity of nanomaterials has also led to more and more concerns and a number of studies regarding the toxicity of nanomaterials have been performed (Li et al., 2012; Liu et al., 2011; Meng et al., 2012; Zhou et al., 2012). ∗ Corresponding author. Tel.: +86 10 8254 5560; fax: +86 10 6265 6765. ∗∗ Corresponding author. Tel.: +45 8716 8027. E-mail addresses: chenchy@nanoctr.cn (C. Chen), cbee@mil.au.dk (C. Beer). Among various kinds of nanomaterials, silver nanoparticles (Ag NPs) are one of the most widely applied nanomaterials in com- mercial products due to their effective antibacterial activity (Cohen et al., 2007; Fu et al., 2006; Xu et al., 2008). Widespread applications, especially in healthcare products, increase the chances of human exposure to Ag NPs, which has increased concerns regarding the potential adverse effects. Unlike gold nanoparticles, Ag NPs can release Ag + due to surface oxidation (Liu and Hurt, 2010). Kittler et al. (2010) showed that the toxicity of Ag NPs increases dur- ing storage due to the release of Ag + , that is considered to be readily bioactive. In accordance with this, we recently showed that the initial Ag + fraction of Ag NP suspensions is important as it contributes greatly to the toxicity of Ag NPs in vitro (Beer et al., 2011). Studies suggest that the toxicity of Ag NPs are due to a so-called Trojan-horse mechanism where Ag NPs are taken up by the cells accompanied with subsequent intracellular release of Ag + thereby leading to cell death (Lubick, 2008). The observed 0378-4274/$ – see front matter © 2013 Elsevier Ireland Ltd. All rights reserved. http://dx.doi.org/10.1016/j.toxlet.2013.07.011