Assessment of in vivo genotoxicity of citrated-coated silver nanoparticles via transcriptomic analysis of rabbit liver tissue

Yeo Jin Kim,1,2 Md Mujibur Rahman,1 Sang Min Lee,2 Jung Min Kim,3 Kwangsik Park,4 Joo-Hyon Kang,5 Young Rok Seo1,2 1Institute of Environmental Medicine for Green Chemistry, Dongguk University Biomedi Campus, Ilsandong-gu, Goyang-si, Republic of Korea; 2Department of Life Science, Dongguk University Biomedi Campus, Ilsandong-gu, Goyang-si, Republic of Korea; 3Genoplan Korea, Inc., Seocho-gu, Seoul, Republic of Korea; 4College of Pharmacy, Dongduk Women’s University, Seongbuk-gu, Seoul, Republic of Korea; 5Department of Civil & Environmental Engineering, Dongguk University, Jung-gu, Seoul, Republic of Korea Background: Silver nanoparticles (AgNPs) are widely used in industrial and household applications, arousing concern regarding their safety in humans. The risks posed by stabilizer-coated AgNPs continue to be unclear, and assessing their toxicity is for an understanding of the safety issues involved in their use in various applications. Purpose: We aimed to investigated the long-term toxicity of citrate-coated silver nanoparticles (cAgNPs) in liver tissue using several toxicity tests and transcriptomic analysis at 7 and 28 days after a single intravenous injection into rabbit ear veins (n=4). Materials and methods: The cAgNPs used in this study were in the form of a 20% (w/v) aqueous solution, and their size was 7.9±0.95 nm, measured using transmission electron microscopy. The animal experiments were performed based on the principles of good laboratory practice. Results: Our results showed that the structure and function of liver tissue were disrupted due to a single exposure to cAgNPs. In addition, in vivo comet assay showed unrepaired genotoxicity in liver tissue until 4 weeks after a single injection, suggesting a potential carcinogenic effect of cAgNPs. In our transcriptomic analysis, a total of 244 genes were found to have differential expression at 28 days after a single cAgNP injection. Carefully curated pathway analysis of these genes using Pathway Studio and Ingenuity Pathway Analysis tools revealed major molecular networks responding to cAgNP exposure and indicated a high correlation of the genes with inflammation, hepatotoxicity, and cancer. Molecular validation suggested potential biomarkers for assessing the toxicity of accumulated cAgNPs. Conclusion: Our investigation highlights the risk associated with a single cAgNP exposure with unrepaired damage persisting for at least a month. Keywords: nanotoxicity, liver toxicity, prolonged tissue damage, differentially expressed genes, molecular pathway analysis