A gut Feeling: Investigating Gut epithelial adaptations to metabolic disease.

Roux-en-Y gastric bypass resolves diabetes, but it is not practical for all obese patients. Remission can also be achieved with non-surgical interventions which exclude nutrients from the duodenum. This highlights the important glucoregulatory role of the small intestine, placing it front and centre in the pathophysiology and treatment of metabolic disease. This project will answer the following key questions:

1) What are the pathological changes in the gut epithelium in response to obesity with a focus on the immunological landscape?

2) Are the changes stem-cell driven and how is gut epithelial cell fate and function altered, with a focus on endocrine function?

3) How does duodenal exclusion improve gut physiology and glucose homeostasis?

To determine the interaction between high fat diet and the small intestinal mucosa (duodenum, jejunum, ileum) we will use an in vitro organoid model. High fat diet will be modelled either by exposure to palmitate or a mixed lipid solution. Gross observations of organoids will be made e.g seeding and developmental capacity, budding structure size and number per organoid, rate of proliferation and apoptosis. The effect of fatty acid exposure on epithelial cell fate, including enteroendocrine cell types, will be determined. As will their effects on enteroendocrine function including gut hormone secretion. Transcriptional analysis of key epithelial compartments (FACs sorting from fluorescent reporter organoids eg. LGR5-GFP, CCK-GFP, NGN3-RFP) or at the single cell level will provide a detailed unbiased view of the effect of high fat diet on the mucosa. This data will be correlated with bulk RNA-seq data already collected from a DIO mouse study, this will allow the student to unpick the effect of diet versus obesity.

a) Recapitulate key experiments in human derived small intestinal organoids.

b) The student will collect lean and obese duodenal and terminal ileum samples from patients attending endoscopy screening clinics. Additionally, Terminal ileum samples will be obtained from pre and post Roux en Y gastric bypass surgeries. These samples will be transcriptionally and morphologically interrogated and organoids generated for downstream analysis of epithelial function using similar experiments as described above. These experiments will allow the student to understand the effect of obesity on the epithelium in man and to interrogate how duodenal exclusion via roux en Y may influence the pathology of the epithelium and thereby improve metabolic disease.