NF-κB Signaling Is Increased in HD3 Cells Following Exposure to 1,4-Benzoquinone: Role of Reactive Oxygen Species and p38-MAPK

In utero exposure to benzene, a known environmental contaminant, is associated with increased risk of leukemia. We have previously shown that in utero benzene exposure can alter the redox-sensitive transcription factor NF-B, and we hypothesize that this is through benzene-induced reactive oxygen species (ROS) production interfering with the signaling pathway involving NF-B and p38-Mitogen Activated Protein Kinase (MAPK). As benzoquinone (BQ) is one of benzenes most toxic metabolites, the objectives of this study were to determine whether ROS and p38-MAPK-mediated BQ-induced increased NF-B activity. HD3 chicken erythroblast cells were transfected with an NF-B luciferase-linked reporter plasmid and exposed to BQ (25M) for 224h. NF-B activities were determined through luciferase assays; Western blotting was conducted to assess changes in protein levels in nontransfected cells; and the presence of ROS was determined via 2,7-dichlorodihydrofluorescein diacetate (DCFDA) flow cytometric assays. Results demonstrated that NF-B activity was significantly increased following exposure to BQ for 16 and 24h and DCFDA assays and pretreatment with antioxidants indicated that BQ-mediated ROS production was responsible for this increase. Furthermore, decreased inhibitor of kappaB-alpha (IB-) expression suggests that nuclear factor-kappaB (NF-B) translocates into the nucleus and that p38-MAPK activation through a ROS-dependent pathway mediates BQ-mediated increases in NF-B activity. Future studies investigating the role of p38-MAPK in this pathway are warranted. Evaluating the effects of toxicant exposure on cell signaling pathways is vital for understanding mechanisms of xenobiotic-induced toxicity.