Thesis (Ph. D.)--University of Wisconsin--Madison, 1966. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographies.

Thesis (M.S.)--University of Wisconsin--Madison, 1983. / Typescript. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

Genetic toxicology assesses the genotoxic potential of chemicals in consumer products, pharmaceuticals and from agricultural and industrial processes. Such assessment is integral in hazard identification and risk assessment to prevent unnecessary human exposure and limit cancer risk. Human risk assessments for genotoxic alkylating agents were based upon linear dose-response models where genotoxicity accrues proportionally with dose. Evidence is accumulating to support a non-linear dose-response at low doses of ethyl methanesulfonate (EMS), a model alkylating agent. For acceptance of non-linear dose responses, a strong explanatory mechanism of action needs to be elucidated. In the following work, low dose mutagenic effects of methyl nitorosurea (MNU), the most potent alkylating agent, have been examined in AHH-1 human lymphoblastoid cells using the HPRT assay. An increase in mutant frequency was not observed until 0.01pg/ml MNU (LOGEL, Lowest Observed Genotoxic Effect Level) with a No-Observed Genotoxic Effect Level (NOGEL) at 0.0075pg/ml MNU. Of interest, is the apparent hormesis induced at 0.0025pg/ml MNU. The principle adduct responsible for MNU mutagenesis is 0 6Methylguanine (06MeG) that miscodes during replication and becomes fixed as GC→AT transitions. Accordingly, the non-linear increase in mutant frequency is accompanied by a non-linear increase in GC→AT transitions. Furthermore, evidence is provided that implicates methlyguanine methyltransferase (MGMT) in protecting DNA from MNU induced mutagenesis by repairing 0 6MeG at low doses, thereby creating the NOGEL. AHH-1 cells treated with 0 6Benzylguanine (06BG), to inactivate MGMT, were hypersensitive to low dose MNU mutagenesis. At 0.0075pg/ml MNU, there was a three-fold increase in mutant frequency and an increase in proportion of GC-^AT transitions, from 28% to 48% in MGMT inactivated cells. This thesis presents a non-linear dose-response for MNU with a strong biological mechanism of action involving DNA repair.

Numerous studies on bacterial systems have shown that cell survival following exposure to various mutagens is greatly influenced by the capacity of the cells to repair the induced DNA lesions. Caffeine (1-3-7-trimethyI xanthine), has been shown to reduce this repair-capacity thereby producing mutagenic, and cytotoxic effects. The question was raised as to whether or not caffeine might reduce the repair-capacity of mammalian cells by interacting with the DNA lesions produced by either the carcinogenic, and oncogenic 4-nitroquinoIine l-oxide, or ultraviolet light (UV) irradiation. This was of interest due to the potential applications that synergistic relationships may hold for chemotherapy. An established line of Syrian-hamster cells (BHK-21, clone 13), was used throughout the experiments. Arginine deprivation was employed to arrest the cultured cells at G₁-phase. The mutagenic 4NQO, UV, and caffeine were applied to these non-dividing cells. The caffeine exposure was at varying time intervals prior to, during, and after 4NQO-induced DNA-repair synthesis (unscheduled DNA synthesis). Similarly, caffeine was added to cells immediately following UV-induced DNA-repair synthesis. Exposure of BHK-21 cells to caffeine, and 4NQO appeared to reduce their colony-forming ability to a greater extent than when the cells were exposed to either chemical alone. The addition of caffeine combined with 4NQO to cells, in G₁-phase, did not appear to significantly influence their rate of flow into S-phase, or through to metaphase. The effect of caffeine on 4NQO- (but not UV-) induced DNA-repair synthesis, using isotope labelling, and autoradiography suggested a reduction in the amount of DNA-repair synthesis. This reduction appeared to be dependent on the caffeine concentration, and on the time of exposure of the cells to this chemical. An initial inquiry was made into whether or not this synergistic effect between caffeine and 4NQO-induced DNA-repair synthesis could be detected in human lymphocytes in vitro. However, very low levels of 4NQO-induced DNA-repair synthesis were observed in these cells. It was therefore not conducive for the autoradiographic analysis of DNA-repair synthesis following 4NQO exposure. A model explaining the apparent caffeine suppression of 4NQO-induced DNA-repair synthesis is postulated. The potential medical implications are discussed. / Medicine, Faculty of / Medical Genetics, Department of / Graduate

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Two aspects of the biological activity of N-nitrosamines were studied. First, the effect of ascorbate on the mutagenicity of N-nitrosopiperidines was studied in the Ames Salmanella/ mammalian microsome mutagenicity test. The addition of ascorbate significantly enhanced the mutagenicity of these compounds. This enhancement was selective for N-nitrosamines suggesting a possible role of ascorbate in N-nitrosamine induced carcinogenicity. Second, the technique of velocity sedimentation in alkaline sucrose density gradients was applied to the detection of N-nitrosamine induced DNA damage in Balb/c 3T3 cells. This technique detected N-nitrosamine induced DNA damage when the cells were made permeable before treatment. This technique compares favorably with other test systems used to evaluate N-nitrosamines and should be useful in further studies of N-nitrosamines.

Thesis (doctoral)--Lund University, 1995. / Added t.p. with thesis statement inserted.

Thesis (doctoral)--Lund University, 1995. / Added t.p. with thesis statement inserted.