Etheno adducts in spleen DNA of SJL mice stimulated to overproduce nitric oxide

Abstract

In order to investigate specific DNA damage caused by nitric oxide (NO) induced lipid peroxidation, levels of promutagenic etheno adducts 1,N6-ethenodeoxyadenosine (εdA) and 3,N4-ethenodeoxycytidine (εdC) were measured in spleen DNA of SJL mice induced to produce high levels of NO by injection of RcsX (pre-B-cell lymphoma) cells. εdA and εdC levels were quantified by an ultrasensitive immunoaffinity-32P-post-labeling method. Spleen DNA of control mice (n = 5) had background levels of 9.2 ± 5.4 εdA adducts per 109 dA and 13.1 ± 5.7 εdC adducts per 109 dC. In RcsX cell-injected mice (n = 7), levels of these adducts were elevated ~6-fold, i.e. 53.9 ± 39.4 εdA per 109 dA and 83.5 ± 57.8 εdC per 109 dC (P < 0.05). Mice injected with RcsX cells and also treated with N(G)-methyl-L-ginine (NMA), an inhibitor of inducible nitric oxide synthase (n = 6), had significantly reduced levels (P < 0.05) of both εdA and εdC (13.5 ± 5.7 εdA per 109 dA and 28.2 ± 15.7 εdC per 109 dC). These findings constitute the first available evidence of formation of etheno adducts associated with NO overproduction in vivo. The adducts were presumably formed from lipid peroxidation products such as trans-4-hydroxy-2-nonenal (HNE), generated via oxidation of lipids by peroxynitrite. The results suggest that etheno-DNA adducts, among other types of damage, may contribute to the etiology of cancers associated with chronic infection/inflammation in which NO is overproduced.