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Genotoxicity of citrate-coated silver nanoparticles to human keratinocytes assessed by the comet assay and cytokinesis blocked micronucleus assay

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Abstract

Silver nanoparticles (AgNPs) are widely used in industrial, cosmetic, and biomedical products, and humans are frequently exposed to these products through the skin. It is widely recognized that the characteristics of AgNPs (e.g., size, coating) may influence their cytotoxic effects, but their correlation with DNA damage and mitotic disorders remains poorly explored. In this study, human keratinocytes (HaCaT cell line) were exposed to well-characterized 30 nm AgNPs coated with citrate, and their effects on viability, DNA fragmentation (assessed by the comet assay), and micronuclei (MNi) induction (assessed by the cytokinesis-block micronucleus cytome assays, CBMN) were investigated. The results showed that 10 and 40 μg/mL AgNPs decreased cell proliferation and viability, and induced a significant genetic damage. This was observed by an increase of DNA amount in comet tail, which linearly correlated with dose and time of exposure. Also, cytostaticity (increase of mononucleated cells) and MNi rates increased in treated cells. In contrast, no significant changes were observed in nucleoplasmatic bridges (NPBs) or nuclear buds (NBUDs), although NBUDs tended to increase in all conditions and periods. The cytostatic effects on HaCaT cells were also shown by the decrease of their nuclear division index. Thus, both comet and CBMN assays supported the observation that citrate-AgNPs induced genotoxic effects on HaCaT cells. Considering that AgNPs are present in a vast number of consumer products and also in multiple nanomedicine skin applications and formulations, more research is needed to determine the properties that confer less toxicity of AgNPs to different cell lines.

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Acknowledgements

This work has been funded by the European Regional Development Fund (FEDER) through the Competitive Factors Thematic Operational Programme (COMPETE) and by National Funds through the Foundation for Science and Technology (FCT), under the projects CICECO - FCOMP-01-0124-FEDER-037271 (Refª. FCT PEst-C/CTM/LA0011/2013) and FCOMP-01-0124-FEDER-021456 (Refª. FCT PTDC/SAU-TOX/120953/2010). The grants awarded by FCT to V.B. (SFRH/BD/81792/2011) and H.O. (SFRH/BPD/111736/2015) are also acknowledged. I.F.D acknowledges FCT/MCTES for a research contract under the Program ‘Investigador FCT’ 2014.

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  1. CESAM & Laboratory of Biotechnology and Cytomics, Department of Biology, University of Aveiro, 3810-193, Aveiro, Portugal

    V. Bastos & H. Oliveira

  2. CICECO, Aveiro Institute of Materials, Department of Chemistry, University of Aveiro, 3810-193, Aveiro, Portugal

    I. F. Duarte

  3. Department of Biology, Faculty of Sciences, University of Porto, 4169-007, Porto, Portugal

    C. Santos

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  1. V. Bastos
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Correspondence to C. Santos or H. Oliveira.

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Communicated by: Markus Hecker

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Bastos, V., Duarte, I.F., Santos, C. et al. Genotoxicity of citrate-coated silver nanoparticles to human keratinocytes assessed by the comet assay and cytokinesis blocked micronucleus assay. Environ Sci Pollut Res 24, 5039–5048 (2017). https://doi.org/10.1007/s11356-016-8240-6

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