Fig. 5

Barrier performance and transport capability of intestinal epithelial cells A Phase-contrast micrograph and immunocytochemistry of RYU on LONG. Immunocytochemistry was performed with antibodies to proteins with barrier functions (ECAD, CDH17, ZO-1, Claudin-2, Claudin-7) and CDX2. Nuclei were stained with DAPI. Scale bars: 500 (black) and 50 (yellow) µm. B Transmission electron microscopic analysis of epithelial lateral membrane. Tight junction (TJ) and adherens junction (AJ, white arrows) were observed. Scale bar: 500 nm. C Trans-epithelial electrical resistance measurements of Caco-2, LONG, and intestinal epithelial cells derived from human iPSCs (iPSC-RYU) and ESCs (ESC-RYU) on LONG. LONG and Caco-2 cells served as negative and positive controls, respectively. Results are expressed as mean ± SD (n = 3 triplicate biological experiments). Statistical significance was determined using Dunnett's test. D Lucifer Yellow permeability test of iPSC-RYU and ESC-RYU on LONG. Caco-2 and LONG were prepared as controls. Results are expressed as mean ± SD (n = 3 triplicate biological experiments). Statistical significance was determined using Dunnett's test. E Lucifer Yellow permeability test was performed for barrier performance. Barrier performance of RYU on LONG. Results are expressed as mean ± SD (n = 3 triplicate biological experiments). F P-glycoprotein (P-gp) gene expression by qRT-PCR analysis in iPSC-RYU and ESC-RYU on LONG in a Transwell. Caco-2 cells and cryopreserved human enterocytes served as controls. The expression level of adult small intestine whole tissue was set to 1.0. Results are expressed as mean ± SD (n = 3 triplicate biological experiments). G Permeability test with Rhodamine 123. The efflux ratio (ER) for Rhodamine 123 was derived from the Papp values associated with basal-to-apical transport (B to A) and apical-to-basal transport (A to B). iPSC-RYU and ESC-RYU on LONG. Mesenchymal stromal cells alone (ESC-LONG and iPSC-LONG) and Caco-2 cells served as negative and positive controls, respectively. Results are expressed as mean ± SD (n = 3 triplicate biological experiments). Statistical significance was determined using Student's t-test. H Expression of the genes for transporters (BCRP, PEPT1, OATP2B1, MRP2, and MRP3). The expression level of adult small intestine whole tissue was set to 1.0. Expression levels were calculated from the results of independent (biological) triplicate experiments. Results are expressed as mean ± SD (n = 3). ES-RYU: human ESC-derived intestinal epithelial cells, iPS-RYU: human iPSC-derived intestinal epithelial cells