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Title: Therapeutic modulation of gastric secretion
Author: Lertkowit, Nantaporn
ISNI:       0000 0004 2737 9583
Awarding Body: University of Liverpool
Current Institution: University of Liverpool
Date of Award: 2011
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The pyloric antral hormone gastrin stimulates acid secretion and regulates proliferation, migration and differentiation of cells in the gastric epithelium. Hypergastrinaemia is associated with enterochromaffin-like (ECL) cell neuroendocrine tumours and may contribute to the progression of other tumours. Multiple active peptides may be derived from the gastrin precursor that are thought to act on different receptors. Therapy directed at suppressing the release of all forms of gastrin through the use of botulinum toxins (BoNTs) that specifically target secretion of G-cells, is therefore attractive. The aim of this study was to assess the feasibility of using retargeted BoNTs to inhibit gastrin release from human G-cells. Cultures of human antral gastric glands were shown by immunocytochemistry to express both t- and v-SNAREs (the proteins essential for regulated exocytosis which are cellular targets of BoNTs) in G-cells. In addition, G-cells in cultured glands were shown to secrete gastrin, basally and 'dose-dependently in response to gastrin releasing peptide (GRP). Acute treatment of glands with GRP-liganded BoNTs enhanced gastrin release, but two GRP receptor (GRPR) antagonists that attenuated the effect of GRP did not significantly inhibit gastrin secretion induced by GRP-liganded BoNTs. The prolonged treatment with GRP-liganded BoNTs, as well as other BoNT fusion proteins, did not suppress gastrin secretion. In further experiments, transfection of plasmids encoding active BoNT proteases and siRNA-mediated knockdown of SNAREs were employed. However, both methods failed to demonstrate the effect of disruption of SNARE complex formation on gastrin secretion, partly due to the low transfection efficiency attained. Treatment with either GRP or epidermal growth factor (EGF) stimulated proliferation of antral epithelial cells as indicated by EdU incorporation. However, no effect was observed with GRP-liganded BoNTs treatments. Taken together with the effect of GRPR antagonists the data suggest that these fusion proteins do not act on GRPR. Sub-epithelial cells such as myofibroblasts are known to release growth factors and other chemical mediators that regulate epithelial cell function. In pernicious anaemia patients there is hypergastrinaemia and it was considered possible that in this condition there was enhanced release of sub-epithelial gastrin secretagogues that could provide novel targets to direct BoNTs to G-cells. Microarray analysis of myofibroblasts from normal and pernicious anaemia subjects was performed and transcript profiles characterised to define putative novel paracrine stimulants of G-cells. The results indicated a possible role of keratinocyte growth factor (KGF) in controlling gastrin secretion, at least in the inflammatory condition. The expression of SNAREs in G-cells suggests selective targeting of BoNTs to these cells might be therapeutically useful. Further work is required to determine the feasibility of using GRP-retargeted BoNTs which appear to act on G-cells, though not via the GRPR. This work has also identified other possible retargeting strategies notably the use ofKGF-liganded BoNT fusion proteins.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available