The cancer stem cell hypothesis suggests that tumour growth is initiated and maintained by a subset of tumour cells that display stem cell- like properties. Stem cell markers such as Leucine-rich repeat-containing G-protein coupled receptor 5 (LGRS), a regulator and downstream target of Wnt signalling, have been identified in the normal intestinal stem cells and also in colorectal tumours. This project aimed to investigate the regulation of LGRS by epigenetic mechanisms and in particular by sodium butyrate, a dietary fibre derivative, histone deacetylase (HDAC) inhibitor and candidate chemopreventive agent. It is important to study dietary compounds as many have been reported to play a role in carcinogenesis. This investigation may therefore lead to improved chemoprevention and the development of new cancer therapies. Initially the role of DNA methylation in regulating LGRS expression in normal t issue and cancer cell lines was investigated, however, although LGRS methylation was observed in some cell lines, no evidence for the functional regulation of LGRS by methylation was found. This study hypothesised that butyrate acts as a chemopreventive agent by targeting cancer stem cells and therefore aimed to investigate the effect of butyrate on the expression of LGRS and other cancer stem cell markers. Butyrate at physiological concentrations was found to downregulate LGRS expression in colorectal cancer cell lines through inhibition of LGR5 gene transcription. This effect occurred within four hours of treatment and was reversible upon removal of butyrate. LGRS expression was also decreased by other pan and class I HDAC inhibitors such as Trichostatin A and Mocetinostat, leading to the conclusion that HDAC inhibit ion was the critical mechanism by which butyrate regulated LGRS. siRNA knockdown of individual HDACs showed that it was the inhibition of HDACl specifically that mediated this effect. Additional work in to the mechanism of butyrate action suggested that Wnt signalling is not involved in the downregulation of LGRS and further work in to other potential intermediate factors will be interesting. Finally the role of LGRS overexpression on cell growth and response to butyrate was investigated. These results suggested that in some cell lines and culture conditions LGRS may enhance cell growth and also reduce the growth inhibitory effects of butyrate. These results present a novel mechanism for the regulation of LGRS, an important cancer stem cell marker, by a well-known dietary derived chemopreventive agent and hint at a role for this effect in regulating stem cell dynamics
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:681727 |
| Date | January 2015 |
| Creators | Jones, Rosanna Frances |
| Publisher | University of Bristol |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
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