Wnt Regulates Cellular Differentiation and Retinoid Metabolism Genes in the Mouse Stomach
Background & Aims: While recognized as a key regulator of gastrointestinal tissues, Wnt function in the stomach is poorly understood. This study aimed to define Wnt functions and identify Wnt-regulated genes in the stomach. Methods: Wnt signaling was localized using reporter mice. Proliferation was studied after canonical Wnt inhibition in vivo using Sox2-CreERT2; Ctnnb1fl/fl mice and in vitro using gastric organoid cultures. Wnt-regulated genes and potential effector pathways were identified by bulk RNAseq analysis of corpus and antral organoids 24 hours after Wnt inhibition. Retinoid metabolic components were localized in control and Wnt-inhibited mouse stomach by immunostaining. Results: Wnt signaling cells mapped to the base and proliferative region in both corpus and antrum. Wnt inhibition in vivo reduced proliferation and led to gastric stem cell loss. Cell signature analysis of RNAseq data revealed that gastric organoids adopt a surface cell transcriptional profile following Wnt inhibition instead of a basal cell profile. Furthermore, retinoid metabolism terms were differentially expressed after Wnt inhibition, with both corpus and antrum displaying decreased expression of retinoic acid target genes. Immunostaining mouse stomach showed differential localization of retinoid metabolic components in luminal pit cells (ALDH3A1), and basal chief/deep mucous cells (STRA6), with expression changes after b-catenin deletion consistent with the organoid analysis. Conclusion: Wnt signaling is required for gastric stem cell survival and promotes differentiation of cell types at the gland base. Retinoid metabolism is a candidate Wnt-regulated pathway, with cell-specific expression of key components. Localization of the retinol transporter STRA6 to high-Wnt cells at the gland base suggests that they are retinoic acid signaling cells. Overall design: RNA-seq of gastric corpus and antral organoids derived from WT mouse gastric glands. On day 4 post-passage, 3 independent gastric organoid lines were treated with the Wnt inhibitor XAV939 with removal of exogenous Wnt/Rspondin and supplementation with Noggin-1 , or with vehicle (DMSO) in normal culture media.