Intestinal epithelial cells (RYU) and Mesenchymal stromal cells (LONG)

The columnar epithelial cells comprising the intestinal tract, stomach, and uterus can be cultured in vitro as organoids or in adherent culture. However, the proliferation of these columnar epithelial cells in adherent culture is limited. Likewise, human pluripotent stem cell (hPSC)-derived intestinal epithelial cells do not show extensive or clonal propagation in vitro. In this study, we induced proliferation of hPSC-derived small intestinal epithelium for a longer time by utilizing mesenchymal stromal cells derived from self-organized intestinal organoids as feeders. The proliferating cells exhibited columnar form, microvilli and glycocalyx formation, and cell polarity, as well as expression of drug-metabolizing enzymes and transporters. It is noteworthy that small intestinal epithelial stem cells cannot be cultured in adherent culture alone, and the stromal cells cannot be replaced by other feeders. Organoid-derived mesenchymal stromal cells resemble the trophocytes essential for maintaining small intestinal epithelial stem cells and play a crucial role in adherent culture. The high proliferative expansion, productivity, and functionality of hPSC-derived small intestinal epithelial stem cells could have potential applications in pharmacokinetic and toxicity studies and regenerative medicine. Comprehensive gene expression analysis of small intestinal epithelial cells and mesenchymal stromal cells derived from human pluripotent stem cells (hESC or hiPSC) was performed. We also prepared a group where small intestinal epithelial cells were treated with puromycin to kill mesenchymal stromal cells used as feeder cells. Adult human small intestine and hESC cell-derived hepatocyte-like cells were used as controls.