Modeling intestinal epithelial function and polarity using human iPSCs under Air-Liquid interface culture

The human intestinal epithelium is essential for nutrient absorption, barrier function, and immune system regulation. Although organoid cultures have advanced the field of intestinal biology, limitations regarding restricted access to the apical side remain, impeding drug administration and epithelial secretion analysis. To address these limitations, we developed an air-liquid interface (ALI) culture system using human induced pluripotent stem cells (iPSCs) to generate intestinal epithelium and evaluate epithelial polarity by analyzing extracellular vesicle (EV) secretion profiles.

Human iPSCs were sequentially differentiated into intestinal stem cells and cultured under ALI conditions. The resulting epithelial tissues were characterized using immunohistochemistry, gene expression analysis, and transepithelial electrical resistance (TEER) measurements. Side-specific responses to indomethacin exposure were evaluated by inflammatory gene expression and apoptosis. EVs collected from the apical and basal compartments were characterized by particle analysis and miRNA profiling.

The ALI culture system successfully generated a polarized monolayer of intestinal epithelium with key features of the human intestine, including absorptive and secretory cell types. Indomethacin treatment from the basal side induced significant mucosal injury and inflammatory gene expression compared to the apical side. Furthermore, EVs secreted from each side displayed distinct miRNA profiles, reflecting polarized secretion patterns and functional compartmentalization.

The human iPSC-derived ALI culture system provides a physiologically polarized epithelial model for studying intestinal biology, drug responses, and polarized EV secretion, with potential applications in personalized medicine.