Chemically-defined and scalable culture system for intestinal stem cells derived from human intestinal organoids.

Three-dimensional human intestinal organoids (hIO) are widely used as a platform for biological and biomedical research. However, reproducibility and challenges for large-scale expansion limit their applicability. Here, we establish a human intestinal stem cell (ISC) culture method expanded under feeder-free and fully defined conditions through selective enrichment of ISC populations (ISC) within hIO derived from human pluripotent stem cells. The intrinsic self-organisation property of ISC, combined with air-liquid interface culture in a minimally defined medium, forces ISC to differentiate into the intestinal epithelium with cellular diversity, villus-like structure, and barrier integrity. Notably, ISC is an ideal cell source for gene editing to study ISC biology and transplantation for intestinal diseases. We demonstrate the intestinal epithelium differentiated from ISC as a model system to study severe acute respiratory syndrome coronavirus 2 viral infection. ISC culture technology provides a biological tool for use in regenerative medicine and disease modelling.