Effects of two typical organophosphate flame retardants on DNA damage of GT1-7 cells and relevant molecular mechanism.

The tris (2-chloroethyl) phosphate (TCEP) and tri-n-butyl phosphate (TNBP) are two typical organophosphate esters (OPEs), which have estrogenic activity. However, fewer studies have been conducted on their genotoxicity and mechanisms. In this study, we evaluated the effects of TCEP and TNBP on cellular DNA damage in GT1-7 cells and examined the role of estrogen signaling in regulating DNA damage. The results showed that TCEP and TNBP significantly inhibited the cell viability and elevated intracellular reactive oxygen species (ROS) levels at higher exposure concentrations. TCEP and TNBP at test concentrations significantly increased comet tail length, comet tail DNA percentage (Tail DNA%), and Olive tail moment (OTM) values, indicating that TCEP and TNBP induced cellular DNA damage. In addition, TCEP and TNBP significantly upregulated the mRNA levels of target genes related to DNA damage-repair pathways, and elevated the expression of ataxia-telangiectasia mutated (ATM) and γ-H2AX proteins associated with DNA double-strand breaks. Furthermore, TCEP and TNBP significantly up-regulated the expression of membrane bound G-protein coupled estrogen receptor 1 (GPER) protein and increased the phosphorylation of extracellular signal-regulated kinases 1/2 (ERK1/2) at 10 µM. While the pretreatment of GPER1 inhibitor G15 and ERK1/2 inhibitor U0126 significantly inhibited the up-regulation of mRNA expression of the target genes associated with the DNA damage-repair pathways and decreased the DNA damage induced by TCEP and TNBP. These findings indicate that TCEP and TNBP activate GPER1/ERK1/2 signaling pathway which plays an important role in regulating DNA damage. The study will provide a novel insight into the genotoxicity mechanism of OPEs.