Investigating the role of short-chain fatty acids in programming the differentiation and function of enteroendocrine L-cells

Enteroendocrine L-cells in the intestinal epithelium are central in initiating communication along the gut-brain axis by the secretion of the anorectic peptide hormones, Peptide YY (PYY) and Glucagon-like peptide 1 (GLP-1). Clinically, the action of these hormones has been harnessed and gut peptide hormone analogues are rising in popularity as anti-obesity therapies. However, there are outstanding limitations to these pharmacological agents, such as the incidence of associated gastrointestinal-related side effects and their inconvenient routes of administration. The G-protein coupled receptor (GPCR) Free fatty acid receptor 2 (FFAR2) has been implicated in mediating the release of GLP-1 and PYY in mouse L-cells by binding to the microbial metabolites, short chain fatty acids (SCFAs). This thesis aimed to characterise SCFA-activated signalling profiles of FFAR2 in human colonic L-cells, and the endocrine functions that these pathways underlie. Firstly, novel spatial- and context-dependent complexities to SCFA-mediated FFAR2 signalling in human L- cells were uncovered. In addition, the SCFA butyrate exerted prominent transcriptional and morphological effects of the in human L-cells which were indicative of the induction of Notch-sensitive enteroendocrine differentiation to a ‘PYY-bias’ phenotype. By developing a pH-sensitive fluorescent reporter system, this thesis also demonstrated an upregulatory effect of butyrate on PYY secretion at single-event resolution. The findings in this thesis also demonstrated a dependence on FFAR2- mediated activation of G𝛼i signalling in the SCFA-mediated effects on human L-cell enterohormone profiles and differentiation, and the use of selective receptor agonists further illuminated the functionalities of the FFAR2-G𝛼i axis in this system. In Notch-reporter and Lgr5-lineage tracer mouse colonic organoid cultures, butyrate modulated Notch activity and stem-cell like properties of heterogeneous populations in the intestinal epithelium. Together, the findings in this thesis uncovered novel effects of SCFAs and the SCFA-binding GPCR FFAR2 in supporting the nutrient-sensing functions and differentiation of human colonic L-cells.