Wnt Signaling Regulation of the Human Gastric Corpus Epithelium

The gastrointestinal epithelium is one of the most proliferative tissues within the body and consistently undergoes complete cellular turnover. This renewal is fueled by resident populations of adult stem cells which asymmetrically divide to establish differentiated cells that carry out functions required for digestion, absorption, and barrier integrity. Wnt signaling is an essential pathway regulating stem cell identity throughout the gastrointestinal tract, and its dysregulation rapidly leads to disease. Interestingly, there appear to be regional differences in Wnt function along the gastrointestinal tract, including between the proximal corpus and distal antral regions of the human stomach. Despite similar gradients of Wnt signaling throughout each region, the corpus and antrum have strikingly different glandular architectures and proliferative zones. Furthermore, while Wnt target genes label active antral progenitors as demonstrated in the intestines, these markers localize to differentiated cells within the corpus. Currently, little is known about the role of Wnt signaling in regulating corpus epithelial cell homeostasis. Gastric Wnt regionality is further suggested by the pathogenesis of Wnt activation diseases such as familial adenomatous polyposis (FAP). FAP is caused by germline loss-of-function mutation to APC, a negative regulator of the Wnt pathway. These patients develop abundant yet benign fundic gland polyps (FGPs) within the corpus but experience little polyp burden in the antrum. It remains poorly understood how activation of Wnt signaling underscores FGP formation, why they are regionally restricted, and what mechanisms underly their benign nature. In this thesis, I sought to investigate the mechanisms through which activation of Wnt signaling regulates the human corpus epithelium to understand the role of Wnt during homeostasis and disease. Throughout my studies, I utilized cultures of three-dimensional, self-renewing organoids derived from primary human tissue. Through comparative experiments, I demonstrated that peak growth of corpus organoids was induced by a lower concentration of GSK3β inhibitor and Wnt pathway activator CHIR99021 than in patient-matched antral organoids. In-depth analysis revealed region-specific, Wnt-dependent regulation of proliferation and that supramaximal CHIR99021 dose-dependently suppressed corpus organoid growth. Further, Wnt signaling regulated a bimodal axis of differentiation within corpus organoids, with activation promoting deep glandular cell differentiation. Paradoxically, deep glandular-enriched, slow-growing organoids established new organoids at enhanced rates. My findings suggest that high Wnt induces differentiated, quiescent cells that are poised to function as proliferative progenitors upon Wnt reduction. To investigate the role of Wnt signaling activation in gastric disease, our lab established a biobank of patient-matched FGP and surrounding non-polyp sample pairs from FAP patients. I determined that FGP biopsies and organoids were associated with increased levels of Wnt signaling yet exhibited reduced tolerance for additional pathway upregulation. This aligns with a theory established to explain the association of specific APC mutations with the development of colon cancer, termed the ‘just-right’ hypothesis. Genomic sequencing demonstrated that in contrast to colon polyposis, second-hit somatic APC mutations were infrequent in FGPs, indicating that most polyps arose without the need for loss-of-heterozygosity. Ultimately, these findings were translated into Apc-deficient mouse models where heterozygous deletion of Apc increased corpus proliferation, while homozygous deletion was required for antral hyperproliferation. In conclusion, this work elucidates a role for Wnt signaling to drive growth and differentiation within the human gastric corpus. These studies provide translatable findings to the clinical manifestations of disease and offer insight for the continued study of human tissue in ex vivo systems.PhDMolecular and Integrative PhysiologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/177687/1/kpmcg_1.pd