Date of Award

2021

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Cell & Molecular Biology

First Advisor

Klaus H. Kaestner

Abstract

The intestine is a highly dynamic organ with diverse functions that are essential for human health, and disruption of the homeostatic processes involved in intestinal function causes a number of diseases, such as colorectal cancer, celiac disease, and inflammatory bowel disease (IBD). However, there are still many gaps in our knowledge of intestinal biology, ranging from the mechanisms of intestinal development, the signaling networks involved in maintaining intestinal epithelial homeostasis, and the mechanisms of complex diseases such as IBD. Thus, there is still a lot of work that needs to be done to better understand the biology of the intestine, which would ultimately benefit intestinal disease research and therapeutic development.

This thesis contains two distinct studies that address knowledge gaps in intestinal research. First, I investigated the mechanisms by which the structure of the intestine is refined during embryonic development, using a conditional genetic ablation mouse model for a critical signaling molecule. Specifically, FOXL1+ expressing mesenchymal cells, which constitute the sub-epithelial layer of the intestine from the stomach to the colon, express key components of the Wnt signaling pathway and are important for maintaining the intestinal stem cell niche in adult mice. Interestingly, these cells also express WNT5A, a major non-canonical Wnt ligand that is essential for embryonic intestinal development but whose cell type of action was previously undetermined. For my thesis work, I derived Foxl1-Cre; Wnt5af/f mice for cell-type specific deletion of Wnt5a solely in epithelium-proximal mesenchymal cells and assessed the consequences to embryonic development. My results showed that although epithelial structural integrity was not lost, ablation of WNT5A in FOXL1+ cells is sufficient to cause shortening of the midgut due to increased epithelial apoptosis during fetal development.

My second project utilized a multiplexed imaging technique called Imaging Mass Cytometry (IMC) to study the cellular and proteomic profiles of IBD tissues. IBD is a highly heterogeneous disease that leads to disruption of homeostasis of the immune response, causing chronic inflammation of the intestinal mucosa. While there are several therapies available for IBD patients, there is still a large unmet need for those individuals who do not respond to the currently available treatments. Thus, development of novel therapeutics requires improved disease stratification based on deeper characterization of the inflamed mucosa. IMC can quantify multiple antigens and cell types of the intestine while maintaining spatial resolution for assessment of cell-to-cell interactions within disease tissue. I performed multiplexed image analysis of 12 IBD and 12 healthy control samples, for which I was able to identify and quantify major immune cell types, their expression markers, and the cell-cell interaction enrichment between regulatory T cells (Tregs) and other immune cells. This study is an important steppingstone for future spatially resolved analysis of IBD.

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