Epigenetic modulation of WNT signalling in the ageing human intestinal epithelium: consequences for tissue homeostasis

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The human gut epithelium is one of the most dynamic tissues in the body, and the maintenance of its integrity is essential for the well-being of the individual throughout life. However, an age-related reduction in crypt number and crypt cell proliferation, and an impaired ability of the epithelium to recover from injury, have all been observed in rodents. The canonical Wnt signalling pathway is the master regulator of intestinal epithelial renewal. Constitutive activation of Wnt signals disrupts intestinal tissue homeostasis in epithelial neoplasia, but preliminary evidence indicates that it also occurs with age in the apparently normal gut. Our pilot studies show that the activity of Wnt signals, and the degree of CpG island methylation of Wnt inhibitor genes, an epigenetic mechanism for gene silencing, both increase with age and vary markedly from crypt-to-crypt in the same individual. We propose that the development of increased Wnt signals along the crypt-axis reflects this age-dependent methylation of Wnt inhibitor genes. To test this hypothesis and understand the consequences for tissue homeostasis we will apply bioimaging and functional genomic approaches to a unique native human colonic crypt culture system. We will measure levels of CpG island methylation of Wnt inhibitor genes, and Wnt signals, in single crypts derived from human volunteers of widely differing age. Age-related variations in levels of DNA methyltransferase activity, and their responses to imposed molecular and cellular stress, will be quantified. We will use RNAi techniques to silence Wnt inhibitors and study the functional consequences for tissue renewal. Finally, we will build a database that can be interrogated with multivariate statistics to derive and test mechanistic hypotheses on the relationship between loss of epithelial tissue homeostasis and levels of CpG island methylation in a panel of Wnt inhibitor genes.

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