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Title: Signalling tissue renewal and crypt survival in the human colonic epithelium and Barrett's oesophagus
Author: Wharton, Natalia
Awarding Body: University of East Anglia
Current Institution: University of East Anglia
Date of Award: 2013
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Stem cell driven tissue renewal in the intestinal epithelium is a tightly regulated and controlled process. The colonic epithelium is organised into millions of invaginations called crypts, each of which represents the self-renewing unit of the tissue. In the mouse, renewal of the intestinal epithelium is regulated by signalling cross-talk between the Wnt, Notch, EGF and TGFβ/BMP pathways. The molecular mechanisms that regulate the processes of tissue renewal in the human are of great interest because they are disrupted in colorectal cancer and inflammatory diseases. Barrett’s oesophagus is an intestinal metaplasia arising in response to inflammation and ulceration provoked by gastroesophageal reflux. Detailed knowledge of the processes and signalling pathways involved in tissue renewal in Barrett’s oesophagus is still lacking and is required to understand more fully the risk and pathogenesis of this metaplasia and oesophageal adenocarcinoma. Intact human colonic crypts were isolated and placed into 3D tissue culture conditions optimised for steady-state tissue renewal. The role of Wnt and TGFβ/BMP signalling pathways in tissue renewal was investigated. Native human colonic crypts exhibited distinct activation profiles for canonical Wnt, TGFβ and BMP pathways. A population of intestinal Lgr5/OLFM4+ stem cells were found to be interspersed between goblet cells at the base of the crypt. Exogenous Wnt signals maintained Lgr5/OLFM4+ stem cells, whilst BMP and TGFβ inhibited and caused complete loss of stem cells. Wnt signals also rescued the inhibitory effects of Dkk1, IWP2 and dnTCF4 on Wnt target gene expression, cell proliferation and crypt length. BMP and TGFβ inhibited Wnt target gene expression, cell proliferation and crypt length. A near-native human Barrett’s oesophagus ex vivo culture model was developed similar to the colonic model which was amenable to real-time time-lapse microscopy and imaging techniques. The Wnt and NFκB signalling pathways exhibited distinct activation profiles. A population of OLFM4+ stem cells were found to reside in the lower third of the Barrett’s crypt. Steady-state tissue renewal in the human colonic epithelium is dependant on Wnt signals combined with suppressed TGFβ/BMP pathways. The human colonic crypt model and the Barrett’s oesophagus crypt model will permit functional interrogation of the mechanisms underlying tissue renewal and risk of inflammatory diseases and adenocarcinoma.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available