Spatial transcriptomics defines segmental and temporal dynamics of pig intestinal epithelium

The pig ( Sus scrofa ) is widely studied as a large-animal model for human gastrointestinal biology, including gut physiology, pathology and disease. However, the spatial and developmental organisation of the pig intestine is not yet fully understood. Here, we generate a comprehensive spatiotemporal atlas of pig intestinal development by applying spatial transcriptomics to three intestinal segments across five developmental stages and an additional four intestinal segments at the adult stage. We identify spatial separation among enterocytes, plasma cells and stem cells, observing more pronounced separation in the small intestine compared to the large intestine. Pseudotime analysis of enterocyte-enriched regions reveals a gradient along the crypt-villus/crypt axis as early as post-conception day 70 in the jejunum, becoming fully evident across all intestinal segments by birth in the large intestine, with all three intestinal segments reaching a mature state by postnatal day 33. Our analysis indicates that the maturation of enterocytes during migration along the crypt-villus axis involves transcriptional programs similar to those governing enterocyte maturation across developmental stages, suggesting conserved molecular mechanisms governing both spatial and temporal differentiation. In the large intestine, nutrient transport-related gene expression declines progressively during development, transitioning from jejunum-like levels at early stages to markedly reduced levels in adulthood, consistent with functional specialization of distinct intestinal segments. Our study provides a high-resolution spatiotemporal framework of pig intestinal development and establishes a valuable resource for comparative biology and translational research.