title: Protein damage from electrophiles and oxidants in lungs of mice chronically exposed to the tumor promoter butylated hydroxytoluene.
abstract: The food additive butylated hydroxytoluene (BHT) promotes tumorigenesis in mouse lung. Chronic BHT exposure is accompanied by pulmonary inflammation and several studies indicate that elevated levels of reactive oxygen species (ROS) are involved in its promoting activity. The link between BHT and elevated ROS involves formation of quinone methide (QM) metabolites; these electrophiles form adducts with a variety of lung proteins including several enzymes that protect cells from oxidative stress. Studies in vitro demonstrated that QM alkylation of cytoprotective enzymes is accompanied by inactivation, so an objective of the present investigation was to determine if inactivation also occurs in vivo. Two groups of mice were exposed to BHT by intraperitoneal injection, one for 10 days and the other for 24 days, and proteins from lung cytosols were examined for damage. Analysis by Western blotting demonstrated that BHT treatment caused substantial increases in protein carbonylation, nitration and adduction by 4-hydroxynonenal, confirming the occurrence of sustained oxidative and nitrosative stress over the treatment period required for tumor promotion. Effects of BHT on the activities and/or levels of a representative group of antioxidant/protective enzymes in mouse lung also were assessed; NAD(P)H:quinone reductase and glutathione reductase were unaffected, however carbonyl reductase activity decreased 50-60%. Superoxide dismutase and glutathione peroxidase activities increased 2- and 1.5-fold, respectively, and glutamate-cysteine ligase catalytic subunit expression increased 32-39% relative to untreated mice. Glutathione S-transferase (GST) activity decreased 50-60% but concentrations of the predominant isoforms, GSTM1 and P1, were not affected. GSTP1 was substantially more susceptible than M1 to adduction and inhibition by treatment with BHT-QM in vitro, suggesting that lower GST activity in mice after BHT treatment is due to adduction of the P1 isoform. The results of this study provide additional insight into mechanisms of BHT-induced oxidative damage and further support a link between inflammation and tumor promotion in mouse lung.
authors.affiliation: Department of Pharmaceutical Sciences, University of Colorado Anschutz Medical Campus, 12850 E. Montview Blvd., Aurora, CO 80045, USA.
authors.forename: Colin T
authors.initials: CT
authors.lastname: Shearn
authors.forename: Kristofer S
authors.initials: KS
authors.lastname: Fritz
authors.forename: John A
authors.initials: JA
authors.lastname: Thompson
mesh_terms: D000818:Animals; D014408:Biomarkers, Tumor / Q000378:metabolism; D002084:Butylated Hydroxytoluene / Q000633:toxicity*; D005978:Glutathione / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D008168:Lung / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D008175:Lung Neoplasms / Q000378:metabolism / Q000473:pathology; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D016877:Oxidants / Q000378:metabolism*; D026361:Reactive Nitrogen Species / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D015262:Xenobiotics / Q000633:toxicity*
chemical_list: D014408:Biomarkers, Tumor; D016877:Oxidants; D026361:Reactive Nitrogen Species; D017382:Reactive Oxygen Species; D015262:Xenobiotics; D002084:Butylated Hydroxytoluene; D005982:Glutathione Transferase; D005978:Glutathione