Identification of proteins adducted by reactive metabolites of naphthalene and 1-nitronaphthalene in dissected airways of rhesus macaques

Naphthalene and 1‐nitronaphthalene are ambient air pollutants, which undergo P450‐dependent bioactivation in the lung. Reactive metabolites of naphthalene and 1‐nitronaphthalene covalently bind to proteins, and the formation of covalent adducts correlates with airway epithelial cell injury in rodent models. These studies were designed to identify protein adducts generated from these reactive metabolites within distal respiratory airways. Distal bronchioles and parenchyma from rhesus monkeys were incubated with [14C]naphthalene or [14C]1‐nitronaphthalene. Proteins were separated by 2‐DE, blotted to PVDF membranes, and adducted proteins imaged by storage phosphor analysis. MS of in‐gel tryptic digests identified numerous adducted proteins including: eight cytoskeletal proteins, two chaperone proteins, seven metabolic enzymes, one redox protein, two proteins involved in ion balance and cell signaling, and two extracellular proteins. While many proteins are adducted by both naphthalene and 1‐nitronaphthalene, some are unique to the individual toxicant and airway subcompartment. Although the role which adduction of these proteins plays in cytotoxicity was not evaluated, these studies provide candidate proteins for future work designed to determine the importance of protein adducts in the mechanisms of toxicity and for developing biomarkers useful in determining the relevance of findings in animal models to exposed human populations.