Please use this identifier to cite or link to this item: http://hdl.handle.net/2440/92814
Type: Thesis
Title: The effect of cannabinoids on cytokine evoked human colonic mucosal damage and Caco-2 epithelial permeability.
Author: Harvey, Benjamin Scott
Issue Date: 2014
School/Discipline: School of Medical Sciences
Abstract: Inflammatory bowel disease (IBD) is a disease characterised by two subtypes, ulcerative colitis (UC) and Crohn’s disease (CD). Both conditions can lead to inflammation and ulceration of the gastrointestinal mucosa. Treatments are available for IBD, however they can cause severe adverse effects and may not be useful in all patients. As a result, there is still an unmet need for novel IBD treatments. In animal models of colitis, cannabinoid (CB) agonists have shown efficacy in reducing inflammation. To further investigate this, we used a human colonic mucosal explant model to determine if CB agonists could attenuate mucosal damage. To induce damage in colonic mucosa, pro-inflammatory cytokines (that are elevated in IBD patients) were used. These included a combination of TNF-α + IL-1β and in other studies, IL-17A. Furthermore, we also tested if these cytokines modulated biochemical markers of inflammation. Immunohistochemistry was used to determine the identity of immune cells in the lamina propria of the mucosa and also localisation of IL-17A. Treatment of colonic mucosa with TNF-α + IL-1β induced damage characterised by luminal epithelial loss, crypt destruction and increased lymphocyte density. In addition, elevations in nitrite levels were found in TNF-α + IL-1β treated explants compared to controls. These damage parameters were attenuated by treatment with CB2R agonists. We found that PGE₂ concentration was significantly decreased after TNF-α + IL-1β incubation suggesting reductions in PGE₂ may partially mediate mucosal damage. IL-17A also induced a course of mucosal damage similar to that observed with TNF-α + IL-1β treatment, however no increase in lymphocyte density occurred. In this study, damage was reduced by the endocannabinoid anandamide as well as cannabidiol. We did not determine whether this effect was CB1R or CB2R mediated. Nitrite concentrations were not elevated after IL-17A treatment, however increased matrix metalloprotease activity was detected, suggesting this may mediate IL-17A induced mucosal damage. ELISA and western blotting was used to determine if the TNF-α + IL-1β combination we previously studied could influence IL-17A levels. There was no significant change in IL-17A expression, however basal expression of IL-17A was found in human colonic mucosa. This was confirmed by immunohistochemistry, showing extensive expression of IL-17A, particularly at the edge of the lumen. Therefore, IL-17A may also play a homeostatic or protective role against micro-organisms in the human colon. Cell culture studies examined the effects of cytokines and cannabinoids on Caco-2 epithelial permeability. In IBD, it has been established that increased mucosal permeability contributes to inflammation. TNF-α + IL-1β increased epithelial permeability; however this was not attenuated by CB ligands. IL-17A did not induce any significant increases in permeability. In conclusion, this thesis demonstrates that CB2R agonists may be useful in attenuating damage in human colonic mucosa induced by cytokines. Therefore, CB2R agonists may have utility as novel therapeutics in IBD. In addition, IL-17A which can be damaging in this model is also expressed in healthy human colonic mucosa, suggesting a homeostatic or protective role. It may be the case that excessive expression of IL-17A in IBD contributes to inflammation.
Advisor: Smid, Scott Darryl
Musgrave, Ian Francis
Dissertation Note: Thesis (Ph.D.) -- University of Adelaide, School of Medical Sciences, 2014
Keywords: Inflammatory bowel disease; Crohn's disease; ulcerative colitis; cannabinoids; epithelial permeability; cytokines
Provenance: This electronic version is made publicly available by the University of Adelaide in accordance with its open access policy for student theses. Copyright in this thesis remains with the author. This thesis may incorporate third party material which has been used by the author pursuant to Fair Dealing exceptions. If you are the owner of any included third party copyright material you wish to be removed from this electronic version, please complete the take down form located at: http://www.adelaide.edu.au/legals
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