Appropriate In Vitro Methods for Genotoxicity Testing of Silver Nanoparticles

Kim, Ha Ryong; Park, Yong Joo; Shin, Da Young; Oh, Seung Min; Chung, Kyu Hyuck

Abstract

OBJECTIVES
We investigated the genotoxic effects of 40-59 nm silver nanoparticles (Ag-NPs) by bacterial reverse mutation assay (Ames test), in vitro comet assay and micronucleus (MN) assay. In particular, we directly compared the effect of cytochalasin B (cytoB) and rat liver homogenate (S9 mix) in the formation of MN by Ag-NPs.
METHODS
Before testing, we confirmed that Ag-NPs were completely dispersed in the experimental medium by sonication (three times in 1 minute) and filtration (0.2 microm pore size filter), and then we measured their size in a zeta potential analyzer. After that the genotoxicity were measured and especially, S9 mix and with and without cytoB were compared one another in MN assay.
RESULTS
Ames test using Salmonella typhimurium TA98, TA100, TA1535 and TA1537 strains revealed that Ag-NPs with or without S9 mix did not display a mutagenic effect. The genotoxicity of Ag-NPs was also evaluated in a mammalian cell system using Chinese hamster ovary cells. The results revealed that Ag-NPs stimulated DNA breakage and MN formation with or without S9 mix in a dose-dependent manner (from 0.01 microg/mL to 10 microg/mL). In particular, MN induction was affected by cytoB.
CONCLUSIONS
All of our findings, with the exception of the Ames test results, indicate that Ag-NPs show genotoxic effects in mammalian cell system. In addition, present study suggests the potential error due to use of cytoB in genotoxic test of nanoparticles.