The hypothesis to be examined is whether tumor promoters induce free radicals or oxidants, whether these oxidants are involved in the mechanism(s) of tumor promotion in mouse epidermis and whether the basis for the known resistance to TPA promotion in C57B1/6 mice as compared to the promotion sensitive SENCAR mouse resides in their differential ability to induce or respond to oxidant production. The rationale for this is based both on literature evidence that antioxidants inhibit promotion while certain organic peroxides have promoting activity and on studies from our laboratory demonstrating that TPA stimulates the production of an oxidant(s) in mouse epidermal cells. We propose to extend these observations through studies designed to answer the following questions, using epidermal cells from SENCAR and C57B1/6 mice treated with TPA, teleocidin, mezerein and A 23187. (1) What is the nature or identity of the tumor promoter-induced oxidant(s)? This will be studied using the following approaches: chemiluminescence, lipid peroxidation, oxygen consumption and electron spin resonance (spin trapping). (2) Do quantitative or qualitative differences exist between the membranes of SENCAR and C57B1/6 mice and is the membrane a target for TPA-induced oxidants? The following parameters will be measured in control and promoter stimulated cells: (a) phospholipid composition and turnover, (b) transmethylation and degradation of methylated phospholipids and (c) identification of the phospholipids that may undergo peroxidation in response to promoters. (3) Can specific radical generating systems mimic known TPA events? The markers to be measured are the induction of ornithine decarboxylase, transglutaminase and DNA synthesis. Additionally, the endogenous antioxidant levels, i.e., basal and promoter-induced levels of superoxide dismutase, catalase and glutathione peroxidase will be compared.
