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Title: Mechanisms of bacterial internalization in the stressed enterocyte
Author: Macutkiewicz, Christian
ISNI:       0000 0001 3616 0418
Awarding Body: University of Manchester
Current Institution: University of Manchester
Date of Award: 2008
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The 'leaky gut' hypothesiS proposes that disruption of the intestinal barrier is critical to the development of inflammatory bowel disease, and characterises the response to severe injury and infection, where local release of inflammatory mediators arid increased gut permeability lead to multiple organ failure. The mechanisms by which commensal gut bacteria enter entehocytes during inflammatory stress and the consequences for enterocyte function are unclear. The purpose of this study was to define the mechanisms by which inflammatory stimuli induce internalization of commensal bacteria within enterocytes, the signalling pathways involved, and the effects of internalized bacteria on the t~nterocyte. Caco-2 monolayers were exposed to an inflammatory stimulus in the form of IFN-y and then incubated with a variety of non-pathogenic commensal microflora. Bacterial internalization and translocation across these monolayers was .quantified by serial dilution culture, signalling mechanisms were investigated using signalling inhibitors, and US~l integrin antibody was used to determine the role of inte.grins. DNA was extracted from the different strains and screened for common bacterial adhesins. The effect of commensal E. coli strains on enterocyte IL-8 secretion was also investigated using ELISA. The translocation of bacteria appears to be strain-specific and depends on a change in the enterocyte brought about by exposure. to IFN-y. It involves interaction with US~l integrin, and utilises a Src-kinase and PI3-kinase dependent process. IFN·y does not appear to increase the expression of usn. integrin but may stimulate redistribution from the basolateral to apical membrane. Most bacterial strains possess the Type 1 fimbrial adhesin, but there appears to be nIJ difference between translocating and nontranslocating strains of E. coli. Most E. coli strains stimulate IL-8 secretion by enterocytes however, E. coli C25 inhi.bits the secretion of IL-8 in all enterocyte cell lines which is a novel finding and is worthy of further investigation. The in vitro model appears consistent and will allow the n1echanistic pathway of bacterial translocation to be elucidated.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available