Decoding Spatiotemporal Transcriptional Dynamics and Epithelial Fibroblast Crosstalk during Gastroesophageal Junction Development through Single Cell Analysis (Tissue-scRNAseq)

The gastroesophageal squamocolumnar junction (GE-SCJ) is a critical tissue interface between the esophagus and stomach, with significant relevance in the pathophysiology of gastrointestinal diseases. Despite this, the molecular mechanisms underlying GE-SCJ development remain unclear. Using single-cell transcriptomics, organoids, and spatial analysis, we examine the cellular heterogeneity and spatiotemporal dynamics of GE-SCJ development from embryonic to adult mice. We identify distinct transcriptional states and signaling pathways in the epithelial and mesenchymal compartments of the esophagus and stomach during development. Fibroblast-epithelial interactions are mediated by various signaling pathways, including WNT, BMP, TGF-ß, FGF, EGF, and PDGF. Our results suggest that fibroblasts predominantly send FGF and TGF-ß signals to the epithelia, while epithelial cells mainly send PDGF and EGF signals to fibroblasts. We observe differences in the ligands and receptors involved in cell-cell communication between the esophagus and stomach. Our findings provide insights into the molecular mechanisms underlying GE-SCJ development and fibroblast-epithelial crosstalk involved, paving the way to elucidate mechanisms during adaptive metaplasia development and carcinogenesis. Overall design: Single cells from esophagus, gastro-esophageal junction and stomach were isolated from embryonic day E15 (n=6), E19 (n=2), Newborn Day-2 (Pup, n=2) and 8-week-old mice (Adult, n=3). Replicates were pooled and cell multiplexed using Cell Multiplexing Oligos (CMOs) based on their tissue region and time points using 10x Genomics 3' CellPlex and Single-Cell 3' v3.1 RNA-seq kits.