The introduction to this thesis examines examples and characteristics of two-stage chemical carcinogenesis, with particular reference to the promotion of mouse skin tumours by 12-O-tetradecanoylphorbol-13-acetate (TPA). The effects of TPA, TPA analogues and other tumour promoters on mammalian cells in culture are also reviewed. The experimental section is in five parts. Chapter 2 lists materials and methods basic to the cell culture procedures used in the succeeding chapters. Chapter 3 is concerned with the established Chinese hamster ovary cell line, CHO-K1. Procedures for the maintenance, manipulation and storage of CHO-K1 cells are detailed. Investigations, including karyological studies, certifying cells as CHO-K1 are reported, along with experiments to determine the lethal effects of three mutagens/carcinogens and TPA towards these cells. In Chapter 4 the design and validation of a CHO-K1/ouabain-resistance mutation assay system is described. Subsequently, the effect of TPA on the frequency of both spontaneous and mutagen-induced ouabain-resistant mutants is determined, the promoter being added to cultures for various intervals during mutation expression and selection. In similar experiments the effect of TPA on spontaneous and ethyl methanesul-phonate-induced mutation to thioguanine-resistance is also determined. The hypothesis that tumour promoters act by inducing mitotic recombination is examined in Chapter 5 by ascertaining if TPA is capable of inducing or enhancing sister chromatid exchanges, or of inducing metaphase quadriradial chromosome structures. Chapter 6 initially confirms that TPA induces blast transformation of human lymphocytes. The possibility of utilising a microculture assay of lymphocyte stimulation as a short term screening test for environmental tumour promoters is then studied by investigating an apparent correlation between lymphocyte mitogenicity and tumour-promoting potency. Chapter 7 summarises the findings of the experimental sections and relates them to recent data reported by other workers and to established effects of tumour promoters.
|Date of Award||1982|