The interaction of probiotic bacteria and an oligosaccharide-enriched fraction from goat whey on in vitro intestinal barrier function and mucin production : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy, Massey University, Manawatu, New Zealand

Abstract

Multiple interactions occur in the human large intestine between the host, the intestinal microbiota and fermentable carbohydrates which transit relatively intact through the small intestine. A major site at which many of these interactions occur is the intestinal epithelium, which is formed from a single layer of epithelial cells. The cellular composition of the epithelial layer in the human small and large intestine varies in respect to the numbers of absorptive enterocytes and mucus-secreting goblet cells. For the human intestine the proportion of goblet cells among epithelial cell types is thought to increase from the duodenum (4%) to the distal colon (16-24%). Epithelial cell co-culture models were developed containing absorptive enterocytes (Caco-2 cells) and mucus-secreting goblet cells (HT29-MTX cells) that more closely simulate the cell proportions found in the small (90:10) and large intestine (75:25). Trans-epithelial electrical resistance (TEER) of the co-cultures was more similar to reported values of ex vivo intestinal tissue of human small and large intestine than either of the two mono-cultures. Additionally, the mucus layer thickness present at the apical surface of 75:25 co-cultures (cellular composition representative of the large intestine) was similar to the reported thickness of the inner mucus layer of human large intestine. Introduction of an oligosaccharide-enriched fraction (OEF) from goat whey to the epithelial co-culture models was shown to modulate barrier integrity as measured by TEER, in a dose-dependent manner. Oligosaccharides (1 mg/mL) increased TEER and mucin gene/protein expression of epithelial co-cultures. Finally, the interaction between probiotic bacteria and the OEF and their individual or combined effects on intestinal epithelial barrier integrity and mucin gene/protein expression was investigated. The OEF supported the growth of selected probiotic strains, and enhanced the adhesion of defined strains to the epithelial co-cultures. When in combination with the OEF, Lactobacillus plantarum 299v enhanced TEER and mucin gene/protein expression, the increase of which was greater than that for either component alone. This suggests that an interaction between Lactobacillus plantarum 299v and the OEF exists which enhances barrier integrity through increased TEER and mucin gene/protein expression, all of which are essential components of the intestinal barrier. The research presented in this dissertation has indicated that in vitro epithelial co-cultures can be used as a model to improve our understanding of the mechanisms through which probiotic bacteria/food components and intestinal epithelial cells interact, and these key findings will assist in the development of strategies to improve intestinal barrier function using novel dietary components.