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In vivo localization and postmortem stability of benzo[a]pyrene‐DNA adducts

DNA adducts of carcinogenic polycyclic aromatic hydrocarbons (PAHs) play a critical role in the etiology of gastrointestinal tract cancers in humans and other species orally exposed to PAHs. Yet, the precise localization of PAH‐DNA adducts in the gastrointestinal tract, and the long‐term postmortem PAH‐DNA adduct stability are unknown. To address these issues, the following experiment was performed. Mice were injected intraperitoneally with the PAH carcinogen benzo[a]pyrene (BP) and euthanized at 24 h. Tissues were harvested either at euthanasia (0 time), or after 4, 8, 12, 24, 48, and 168 hr (7 days) of storage at 4°C. Portions of mouse tissues were formalin‐fixed, paraffin‐embedded, and immunohistochemically (IHC) evaluated by incubation with r7,t8‐dihydroxy‐t‐9,10‐epoxy‐7,8,9,10‐tetrahydrobenzo[a]pyrene (BPDE)‐DNA antiserum and H‐scoring. The remaining tissues were frozen, and DNA was extracted and assayed for the r7,t8,t9‐trihydroxy‐c‐10‐(N 2‐deoxyguanosyl)‐7,8,9,10‐tetrahydrobenzo[a]pyrene (BPdG) adduct using two quantitative assays, the BPDE‐DNA chemiluminescence immunoassay (CIA), and high‐performance liquid chromatography electrospray ionization tandem mass spectrometry (HPLC‐ES‐MS/MS). By IHC, which required intact nuclei, BPdG adducts were visualized in forestomach basal cells, which included gastric stem cells, for up to 7 days. In proximal small intestine villus epithelium BPdG adducts were visualized for up to 12 hr. By BPDE‐DNA CIA and HPLC‐ES‐MS/MS, both of which used DNA for analysis and correlated well (P= 0.0001), BPdG adducts were unchanged in small intestine, forestomach, and lung stored at 4°C for up to 7 days postmortem. In addition to localization of BPdG adducts, this study reveals the feasibility of examining PAH‐DNA adduct formation in wildlife species living in colder climates. Environ. Mol. Mutagen. 61:216–223, 2020. © 2019 Wiley Periodicals, Inc.