Figure 3.

IEC-derived IL-27 is required for optimal CD8αα⁺CD4⁺ IEL responses. (A) qPCR analyses for the expressions of Il27p28 and Ebi3 in CD45⁻EpCAM⁺ IECs from 8–12-wk-old WT and IEC-KO mice in the presence or absence of LPS stimulation. (B) ELISA analyses of the production of IL-27 by CD45⁻EpCAM⁺ IECs from 8–12-wk-old WT and IEC-KO mice in the presence or absence of Pam3CysSerLys4 (Pam3CSK4, a TLR2 agonist), LPS (TLR4 agonist), and CpG (TLR9 agonist) at different concentrations in vitro. (C and D) ELISA analyses of the production of IL-27 by CD45⁻EpCAM⁺ IECs, CD11c^hi cDCs, and CD11c⁻CD11b⁺ myeloid cells from 8–12-wk-old WT and IEC-KO mice in the presence or absence of LPS (1 µg/ml; C) or IFNγ (10 ng/ml; D) stimulation in vitro. (E–K) Frequencies of CD8α⁺ in TCRβ⁺TCRγδ⁻CD4⁺ cells (E) and TCRβ⁺TCRγδ⁻CD8α⁺CD4⁺ cells in total IELs (F), as well as frequencies of TCRαβ⁺CD8αβ⁺(G), TCRαβ⁺CD8αα⁺ (H), and TCRγδ⁺CD8αα⁺ (I) IEL subsets from old (>6 mo) IEC-KO mice and WT littermates. (J and K) FACS analysis and frequencies of Foxp3⁺ T reg cells in the intestinal epithelium of IEC-KO mice (J), T-rKO mice (K), and their corresponding WT littermates (>6 mo). Each symbol represents an individual mouse, and the bar represents the mean. Data are pooled from at least three independent experiments. Results of Student’s t test: *, P < 0.05; **, P < 0.01; ***, P < 0.001.