Nrf2/ARE pathway activation, HO-1 and NQO1 induction by polychlorinated biphenyl quinone is associated with reactive oxygen species and PI3K/AKT signaling

Abstract

Nrf2/ARE pathway plays an important role in adapt to oxidative stress caused by pro-oxidants and electrophiles through up-regulating phase II detoxifying enzymes. Our previous study has demonstrated that PCB quinone exposure causes severe cellular oxidative stress (Toxicology In Vitro 26 (2012) 841–848). There are no reports describing the ability of PCB quinone on Nrf2/ARE activation. In the present study, we found that exposure to PCB29-pQ resulted in a significant increase in Nrf2 and Keap1 expression in total protein, as well as the Nrf2 targeting genes, including NQO1 and HO-1. Next, immunocytochemistry analysis identified the accumulation of Nrf2 in nucleus subsequent to PCB29-pQ treatment. The increased Nrf2 and constant Keap1 expression in nucleus suggested the dissociation of Nrf2/Keap1 complex. Similarly, mRNA level of Nrf2 was elevated significantly with PCB29-pQ treatment, but not Keap1. Additionally, PCB29-pQ treatment led to significant up-regulation of the mRNA level of antioxidant enzymes, NQO1 and HO-1, in a concentration-dependent manner. Electrophoretic mobility shift assay and luciferase reporter assay further confirmed the formation of Nrf2–ARE complex. PCB29-pQ treatment has no effect on mitogen-activated protein kinase signaling, however, phospho-AKT was up-regulated and GSK-3β was down-regulated. Pretreatment with LY294002, a specific inhibitor of phosphatidylinositol 3-kinase (PI3K), suppressed the phosphorylation of AKT and inhibited PCB29-pQ induced Nrf2/HO-1 activation, meanwhile, GSK-3β expression was increased accordingly. At last, reactive oxygen species (ROS) scavengers inhibited PCB29-pQ induced Nrf2 activation partly. These results suggested that Nrf2 activation by PCB29-pQ in HepG2 cells is associated with ROS and AKT pathway but not MAPK signaling, the activation of Nrf2/ARE may be an adaptive response to oxidative stress.