Single-cell time-course transcriptomic analysis reveals epithelial remodeling in human gastric corpus organoids after WNT pathway perturbation

Background: WNT signaling is essential for gastric epithelial stem cell homeostasis. However, the temporal transcriptional dynamics following WNT/RSPO perturbation in human gastric organoids remain unclear.Methods: Human gastric corpus organoids derived from normal tissue were stabilized and then subjected to WNT/RSPO perturbation. Single-cell RNA sequencing was performed at 0, 6, 12, 24, 48, and 72 hours. Differential gene expression, pseudotime analysis, and pseudobulk profiling were conducted to characterize temporal changes.Results: WNT/RSPO perturbation induced progressive epithelial remodeling, characterized by a decline in stem cell populations and expansion of tuft and mucous cell populations. The number of differentially expressed genes increased markedly after 12 hours, indicating a major transcriptional transition. Stem cell–associated and canonical WNT target genes decreased over time, whereas differentiation-associated genes predominantly peaked after 48 hours. Early time points (≤24 hours) reflected primary WNT signaling responses, whereas later time points (≥48 hours) captured differentiation-associated programs.Conclusion: This temporal single-cell transcriptomic analysis delineates distinct phases of transcriptional remodeling following WNT/RSPO perturbation and distinguishes direct WNT signaling responses from subsequent differentiation-associated changes in human gastric corpus organoids.