Genetic and cytogenetic effects of 12-O-tetradecanoylphorbol-13-acetate in mammalian cells and lymphocyte stimulation by tumour-promoting phorbol esters.

  • Robert Steven Dewdney

Student thesis: Doctoral ThesisPhD

Abstract

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 Award1982
Original languageEnglish
Awarding Institution
  • University of Bath

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