Enhanced Utilization of Induced Pluripotent Stem Cell-Derived Human Intestinal Organoids Using Microengineered Chips

The 3-dimensional structure of human intestinal organoids makes them challenging to use. Here we describe how organoids, derived from induced pluripotent stem cells, can be incorporated into small microengineered Chips making them more amenable for study. BACKGROUND AND AIMS: Human intestinal organoids derived from induced pluripotent stem cells have tremendous potential to elucidate the intestinal epithelium's role in health and disease, but it is difficult to directly assay these complex structures. This study sought to make this technology more amenable for study by obtaining epithelial cells from induced pluripotent stem cell-derived human intestinal organoids and incorporating them into small microengineered Chips. We then investigated if these cells within the Chip were polarized, had the 4 major intestinal epithelial subtypes, and were biologically responsive to exogenous stimuli. METHODS: Epithelial cells were positively selected from human intestinal organoids and were incorporated into the Chip. The effect of continuous media flow was examined. Immunocytochemistry and in situ hybridization were used to demonstrate that the epithelial cells were polarized and possessed the major intestinal epithelial subtypes. To assess if the incorporated cells were biologically responsive, Western blot analysis and quantitative polymerase chain reaction were used to assess the effects of interferon (IFN)-gamma, and fluorescein isothiocyanate-dextran 4 kDa permeation was used to assess the effects of IFN-gamma and tumor necrosis factor-alpha on barrier function. RESULTS: The optimal cell seeding density and flow rate were established. The continuous administration of flow resulted in the formation of polarized intestinal folds that contained Paneth cells, goblet cells, enterocytes, and enteroendocrine cells along with transit-amplifying and LGR5(+) stem cells. Administration of IFN-gamma for 1 hour resulted in the phosphorylation of STAT1, whereas exposure for 3 days resulted in a significant upregulation of IFN-gamma related genes. Administration of IFN-gamma and tumor necrosis factor-alpha for 3 days resulted in an increase in intestinal permeability. CONCLUSIONS: We demonstrate that the Intestine-Chip is polarized, contains all the intestinal epithelial subtypes, and is biologically responsive to exogenous stimuli. This represents a more amenable platform to use organoid technology and will be highly applicable to personalized medicine and a wide range of gastrointestinal conditions.