Polyploidy in murine hepatocytes

• Main Author: Martin, Nicola Claire
• Published: University of Edinburgh 2002
• Subjects: 572.8
• Online Access: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.657376

This thesis has looked at various aspects of polyploidisation in the mouse liver to obtain more information about the function of polyploidisation: The effect of the non-genotoxic carcinogen, sodium phenobarbitone (PB) on ploidy and proliferation was determined over 21 days in p53 wild type (+/+), p53 +/- and p53 null (-/-) mice. PB induced a significant increase in the proportion of 8n nuclei and this increase occurred independently of p53. Whether polyploidisation occurs as a protective mechanism or acceleration of ageing in this case is unknown. However, the results confirm that the p53 +/- mouse model, often used in short-term bioassays, may not be suitable for the identification of non-genotoxic carcinogens. The function of polyploidy with respect to increasing cell size, receptor expression and susceptibility to apoptosis was also studied. The first accurate measure of volume of isolated hepatocytes differing in ploidy and nuclearity was carried out using confocal image analysis. The increase in volume associated with increasing DNA content was found to be proportional to intercellular adhesion molecule-1 (ICAM-1) surface expression, measured by flow cytometry. Apoptosis induced by interferon-gamma (IFN-γ) in culture did not occur preferentially in polyploid hepatocytes. This would suggest that the susceptibility of polyploid cells to apoptosis depends on the circumstances. Analysis of gene expression in the different hepatocyte populations also provides information regarding the function of polyploid cells. Evaluation of the effect of fluorescent activated cell sorting (FACS) and Hoechst 33342 on RNA from sorted hepatocytes found that the quality was not affected by the procedure and the RNA was suitable for subsequent gene expression analysis. Initial experiments using microarray and real-time PCR identified several genes that were induced or repressed in cells containing greater amounts of DNA. Further biological studies will hopefully determine whether these genes are important to the function or maintenance of polyploidy in hepatocytes.