Fig. 1.
Analysis of expression of IL-22R subunits IL22R1 and IL10R2 demonstrated that iHOs generated from healthy control hIPSCs express the receptors for IL-22, IL22R1, and IL10R2, in contrast to iHOs generated from a patient with infantile IBD, which lack the IL10R2 subunit. To generate IL10RBComp, an isogenic control line for patient iPSCs (IL10RBPat), TALEN-mediated gene integration was used to integrate a copy of the IL10RB gene into the AAVS1 site in IL10RBPat iPSCs. (A) mRNA levels determined by RT-qPCR for IL22RA and IL10RB in three healthy control iHO lines (Yemz1, Lise1, and Kolf2), IL10RBPat iHOs, and control duodenal tissue. IL10RB is significantly up-regulated in control iHOs in comparison with IL10RBPat (P = 0.0005, P < 0.0001, and P < 0.0001, respectively; Student’s t tests). (B) RT-qPCR analysis shows significant up-regulation of IL10RB expression in IL10RBComp iHOs in comparison with IL10RBPat iHOs (P < 0.0001; unpaired, two-tailed Student’s t test). RT-qPCR was performed with TaqMan gene-expression assays and analyzed via the comparative cycle threshold (CT) method with GAPDH as an endogenous control. Data are presented from four technical replicates, with assays repeated at least three times from independent iHO batches. **P < 0.001; ***P < 0.0001. (C) Z-stacked immunostaining for IL-22 receptors IL10R2 or IL22R1 (green) and DAPI (blue) on healthy control Kolf2, IL10RBComp, and IL10RBPat iHOs showing localization of IL22R1 and IL10R2 on the basal IEC surface, with IL10R2 not detected in IL10RBPat iHOs. (Original magnification: 20×.)