High levels of Wnt signaling reduce proliferation and promote deep glandular differentiation in human gastric corpus-derived organoids

Abstract

Introduction: The Wnt signaling pathway regulates gastrointestinal stem cell identity and function. Throughout the stomach, a gradient of Wnt signaling is established across the epithelium with highest levels at the gland base. However, the proximal (corpus) and distal (antrum) regions of the stomach have strikingly different epithelial architectures, with antral stem cells and proliferation localized to the gland base while corpus proliferation is localized higher-up in the glands where Wnt levels are diminished. It is unknown how Wnt may differentially regulate cell function and identity within these regions. Furthermore, activation of Wnt signaling through genetic diseases, such as familial adenomatous polyposis (FAP), results in abundant polyposis of the corpus while the antrum is mostly spared. This study investigated how activation of Wnt signaling in human gastric corpus- and antral-derived organoids regulates stem cell function, with the hypothesis that corpus stem/progenitor cells have a lower threshold for optimal Wnt signaling. Methods: Human patient-matched corpus and antral organoids from two individuals were grown in the presence of the Wnt agonist CHIR99021 to assess Wnt sensitivity in vitro­. Corpus organoids were then grown in high levels of CHIR99021 to assess how Wnt activation affected growth, proliferation, differentiation, and stem cell maintenance. Results: Treatment with CHIR99021 in media free of exogenous Wnt/R-spondin stimulated a maximal growth response in corpus organoids at lower concentrations than their respective patient-matched antral organoids. Increased Wnt signaling beyond the maximal growth concentration also suppressed proliferation at a rapid rate in corpus organoids. Chronic treatment with increasing levels of CHIR99021 reduced organoid size, suppressed surface cell differentiation, and promoted expression of markers for differentiated deep glandular cells, including neck and chief cells. Surprisingly, organoids retained in high levels of CHIR99021 had enhanced organoid formation ability when passaged, indicating that progenitor cell status was maintained in these non-proliferative, deep glandular cell-enriched organoids. Passaging these quiescent organoids into low Wnt conditions led to a rapid rescue of normal growth, organoid morphology, and surface cell differentiation. Conclusion: Our findings suggest that progenitor cells in human corpus organoids have a lower threshold for optimal Wnt signaling than progenitor cells in the antrum. We demonstrate that Wnt signaling in the corpus plays a central role in defining a bimodal axis of differentiation, and that high activation of Wnt signaling promotes stemness while simultaneously suppressing proliferation. Funding support for this research was provided through T32DK094775 (KPM), R25GM086262 (ED), and R01DK126451 (LCS). This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.