INVESTIGATION OF THE ROLE OF OXIDATIVE DNA DAMAGE IN AFLATOXIN B1-INDUCED PULMONARY CARCINOGENESIS
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Studies described in this thesis were aimed at characterizing the mechanism(s) of aflatoxin B1 (AFB1) pulmonary carcinogenesis by addressing the formation, prevention, and repair of AFB1-induced oxidative DNA damage. The ability of AFB1 to cause oxidative DNA damage in different lung cell types of the A/J mouse was examined. The formation of 8-hydroxy-2’-deoxyguanosine (8-OHdG) in freshly isolated mouse lung alveolar macrophages, alveolar type II cells, and nonciliated bronchial epithelial (Clara) cells, was assessed by high performance liquid chromatography with electrochemical detection. An increase in 8-OHdG formation occurred in macrophage and Clara cell preparations isolated from A/J mice two hours following in vivo treatment with a single tumourigenic dose of AFB1. Prior treatment with polyethylene glycol-conjugated catalase (PEG-CAT) prevented the AFB1-induced increase in 8-OHdG levels in all mouse lung cell preparations. These results support the possibility that oxidative DNA damage in mouse lung cells contributes to AFB1 carcinogenicity. Mouse lung tumourigenesis was assessed following treatment of A/J mice with PEG-CAT and/or AFB1. Unexpectedly, the mean number of tumours per mouse and tumour size in the PEG-CAT + AFB1 group were greater than those of the group treated with AFB1 alone. There was no difference in K-ras exon 1 mutation spectrum or in the histological diagnosis of tumours between treatment groups. In vitro incubation with mouse liver catalase (CAT) resulted in conversion of [3H]AFB1 into a DNA-binding species, a possible explanation for the results observed in vivo. These results demonstrate that PEG-CAT is not protective against AFB1 carcinogenicity in mouse lung despite preventing DNA oxidation. The effect of in vivo treatment of mice with AFB1 on pulmonary and hepatic base excision repair (BER) activities and levels of 8-oxoguanine DNA glycosylase (OGG1) was investigated. AFB1 treatment increased 8-OHdG levels and BER activity in mouse lung, but did not significantly affect either in liver. Levels of OGG1 immunoreactive protein were increased in both mouse lung and liver. These results indicate that oxidative DNA damage may be an important mechanism in the carcinogenicity of AFB1. However, BER activity is increased by AFB1 treatment, possibly representing a compensatory response to the production of oxidative DNA damage.