Usefulness of Monitoring γ-H2AX and Cell Cycle Arrest in HepG2 Cells for Estimating Genotoxicity Using a High-Content Analysis System. (October 2014)

• Record Type: Journal Article
• Authors: Ando, Masamitsu; Yoshikawa, Keisuke; Iwase, Yumiko; Ishiura, Shoichi
• Is Part Of: Journal of biomolecular screening. Volume 19:Number 9(2014)
• Page Start: 1246
• Page End: 1254
• Publication Date: 2014-10
• Subjects: high-content screening -- genotoxicity -- γ-H2A.X -- cell cycle arrest; Drugs -- Analysis -- Periodicals; Drugs -- Testing -- Periodicals; Biomolecules -- Analysis -- Periodicals; 572.36
• DOI: 10.1177/1087057114541147
• Languages: English
• ISSNs: 1087-0571

Abstract

Formation of the phosphorylated protein γ-H2AX is a well-established marker of DNA strand breakage induced by DNA-damaging compounds. Many of these genotoxic compounds also inhibit cell division, leading to arrest at specific points in the cell cycle. Detection of γ-H2AX in combination with cell cycle arrest may therefore be useful for estimating the genotoxicity of experimental compounds. In this study, we examined γ-H2AX formation and cell cycle arrest using high-content screening (HCS) as a method for determining genotoxicity. HepG2 cells were treated with a panel of compounds and then stained with Hoechst 33342 and anti-γ-H2AX, anti-phospho-histone H3, and anti-tubulin antibodies. In total, 19 genotoxic and 7 nongenotoxic compounds were tested in this study. γ-H2AX production was observed within 1 h posttreatment for the majority of Ames-positive compounds, topoisomerase inhibitors, and DNA polymerase inhibitors. Cell cycle arrest in either the S or G2 phase was detected for all DNA-damaging compounds 24 h posttreatment, whereas tubulin-targeting compounds were shown to induce cell cycle arrest in the mitotic phase. Together, these results show that HCS is a simple, rapid, and effective tool for estimating the genotoxicity of compounds through detection of γ-H2AX production and cell cycle arrest.