Nrf2 overexpression protects against paraquat-induced A549 cell injury primarily by upregulating P-glycoprotein and reducing intracellular paraquat accumulation

Paraquat (PQ) intoxication causes thousands of mortalities every year, worldwide. Its pulmonary-targeted accumulation and the acute lung injury it subsequently causes, remain a challenge for detoxification treatment. A previous study has demonstrated that the upregulation of nuclear factor erythroid-2 related factor 2 (Nrf2) prevents PQ toxicity in cell line and murine models. As Nrf2 target genes include a group of membrane transporters, the current study assessed the protective mechanism exerted by Nrf2 against PQ toxicity and intracellular PQ accumulation via its effects on P-glycoprotein (P-gp), a downstream transporter of Nrf2. Adenovirus vectors containing the Nrf2 gene were transfected into A549 cells. Cell proliferation was assessed by Cell Counting Kit-8. The levels of LDH, MDA, SOD, TNF-α, IL-6 levels were detected using their respective ELISA kits. In addition, the levels of Nrf2 and P-gp protein expression were detected by western blot analysis. The concentration of PQ was measured by HPLC. The results revealed that overexpressed Nrf2 significantly increased P-gp protein levels, decreased the intracellular accumulation of PQ and attenuated PQ-induced toxicity. However, the protective effects of Nrf2 overexpression on PQ-challenged A549 cells were abrogated following cyclosporine A treatment, a competitive inhibitor of P-gp, which also increased intracellular PQ levels. These data indicated that Nrf2 gene overexpression prevented PQ toxicity in A549 cells, potentially via the upregulation of P-gp activity and the inhibition of intracellular PQ accumulation. Thus, Nrf2 and P-gp may serve as potential therapeutic targets for the treatment of PQ-induced injury.