Reactive oxygen species (ROS) have been implicated as playing an important role in tumor promotion and in progression of benign to malignant tumors. ROS produced by tumor promoter-activated polymorphonuclear leukocytes (PMNs) are mutagenic and carcinogenic, and cause formation of the oxidized bases thymidine glycol, 5-hydroxymethyl-2'-deoxyuridine (HMdU) and 8- hydroxyl-2'-deoxyguanosine (8-OHdG) in DNA, which are lethal, mutagenic and miscoding. The long-term objective of this work is to elucidate the mechanism by which ROS are induced in tumor promotion and the role of oxidative DNA damage in carcinogenesis. The main hypothesis to be tested is that the formation of ROS and oxidative DNA damage constitutes a common denominator among tumor promoters and that they are tumor-promoting equivalents in the action of complete carcinogens, which require oxidative metabolism. This hypothesis is based on our recent findings, which show that in vivo treatment of SINCERE mice with tumor promoters of the phorbol ester type results in the production of H2O2 and oxidized bases in the DNA of epidermal cells. We also showed that a treatment of mouse skin with 7,12-dimethylbenz(a)anthracene (DMBA), which led to tumor development, also caused prolonged inflammation, H2O2 production and formation of oxidized bases in the epidermal DNA. Their levels were comparable to those mediated by the potent tumor promoter TPA (12-O-tetradecanoylphorbol-13-acetate), but apparently with different rates of formation and disappearance.
Aims of this study are: 1) To determine whether TPA and non-TPA type tumor promoters, and polycyclic aromatic hydrocarbons induce PMN infiltration, H2O2. Hmdu and 8-Ohdg formation in epidermis of SINCERE mice; 2) To find whether anti-tumor promoters suppress DMBA-mediated inflammatory responses, formation of oxidized DNA bases and chemotactic factors (IL-1 and IL-8), and have an effect on chemotaxis of PMNs; 3) To establish whether TPA- and DMBA-induced Hmdu and 8-Ohdg disappear from epidermal DNA with different rates and effects of anti-tumor promoters on that disappearance; 4) To determine whether DMBA-mediated formation of H2O2, Hmdu and 8OHdG are comparable in SINCERE and C57BL/6J mice; and 5) To establish whether oxidative DNA damage is required for the TPA-mediated expression of the competence genes c-fos and c-myc. Hmdu and 8-OHdC will be assayed by HPLC and 3H-postlabeling with [3H]acetic anhydride, cytokines in sera by binding of antibodies labeled with a fluorochrome, and gene expression by Northern blot analysis and nuclear run-off transcription.
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