Evaluation of N-acetyl-cysteine against tetrachlorobenzoquinone-induced genotoxicity and oxidative stress in HepG2 cells

Abstract

Tetrachlorobenzoquinone (TCBQ) is an active metabolite of pentachlorophenol (PCP). Although the genotoxic effect of PCP has been comprehensively investigated, there is little known about TCBQ's genotoxic effects. In the current study, TCBQ was tested for its genotoxicity using HepG2 cells as experimental model. To select the exposure concentration of interest, cell viability was measured and three concentrations were used for further investigation. In single cell gel electrophoresis (SCGE) assay, concentration-dependent increase in tail length, tail DNA percentage and tail moment were detected following TCBQ exposure. Micronucleus (MN) assay indicated TCBQ gradually increased MN frequency and decreased nuclear division index (NDI). Enzyme-linked immunosorbent assay (ELISA) and western blotting analyses both showed TCBQ caused histone H2AX phosphorylation (γ-H2AX). Furthermore, the elevation of 8-hydroxydeoxyguanosine (8-OHdG) and reactive oxygen species (ROS) level indicated TCBQ-induced genotoxicity is associated with oxidative stress. On the other hand, N-acetyl-cysteine (NAC) administration significantly protected cells from the genotoxic effect of TCBQ. Overall, our data suggested TCBQ exerted genotoxic effect possibly via an oxidative damage mechanism in HepG2 cells and this toxicity is prevented by pretreatment with NAC.

Keywords: 2′,7′-dichlorodihydrofluorescein diacetate; 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide; 4′,6-diamidino-2-phenylindole dihydrochloride; 8-OHdG; 8-hydroxydeoxyguanosine; Cytokinesis-block micronucleus assay; DAPI; DCFH-DA; ELISA; Genotoxic; MN; MTT; N-acetyl-cysteine; NAC; NDI; PCP; ROS; SCGE; Single cell gel electrophoresis assay; TCBQ; TCHQ; TCSQ; enzyme-linked immunosorbent assay; micronucleus; nuclear division index; pentachlorophenol; phosphorylation of histone H2AX; reactive oxygen species; single cell gel electrophoresis; tetrachlorobenzoquinone; tetrachlorohydroquinone; tetrachlorosemiquinone; γ-H2AX.