Fig. 2.

Expression of IL10RB was required for iHO responses to IL-22. iHOs were treated with 100 ng/mL recombinant human IL-22 (rhIL-22) added to the iHO medium. (A) In iHO lines with a functional copy of IL10RB, transcripts for IL-22–regulated genes lipocalin 2 (LCN2) and dual oxidase 2 (DUOX2) are significantly up-regulated after the addition of 100 ng/mL rhIL-22 for 18 h, in comparison with unstimulated iHOs (**P < 0.001; ***P < 0.0001; unpaired, two-tailed, Student’s t tests; n.s., not significant). Data are presented from four technical replicates; assays were repeated with at least three biological replicates. RT-qPCR was performed with TaqMan gene-expression assays and analyzed via the comparative C_T method with GAPDH as an endogenous control. (B) Phospho-STAT3 level was detected by Western blot after stimulation of healthy control (Yemz1 and Kolf2), IL10RB^Pat, and IL10RB^Comp iHOs for 30 min with rhIL-22 and preparation of whole-cell extracts. To verify equal protein loading, the blot was stripped and reprobed with STAT3 antibody. Lysate from HeLa cells stimulated with IFNλ were used as a positive control. (C) Kolf2 iHOs challenged with IL-22 for 18 h or left untreated were examined for Mucin 4 (MUC4; green) and DAPI (blue) by immunofluorescence. (D) Z-stacked immunostaining for Mucin 4 in IL10RB^Pat and IL10RB^Comp iHOs challenged with IL-22 for 18 h or were left untreated. (Original magnification: 20×; L = iHO lumen.)