Interaction between NK cells and DC induces an innate IFN-γ–IL-27 amplification loop that limits disease and is controlled by IL-10. (A–C) GKO mice depleted of NK cells and repleted with autologous or WT NK cells were immunized for EAU. (A) Il27 gene expression by RT PCR and (B) IL-27 protein expression by intracellular staining were determined in CD11c⁺ DCs isolated from draining LNs on d 5. (C) GKO mice repleted with autologous or WT NK cells were immunized for EAU and treated with anti-IL27 or isotype control antibody. Disease was monitored by funduscopy. (D) DCs from WT mice were incubated with LPS overnight and washed. The LPS-DCs were then co-cultured with WT NK cells in the presence of anti–IL-27, anti–IFN-γ, or isotype control. IL-27 and IFN-γ were quantitated by ELISA in 48 h supernatants. (E) CFSE-labeled WT or IL-27rα^−/− NK cells were transferred to the GKO mice at the day of immunization. Their IFN-γ expression in the draining LNs was determined by intracellular staining on day 5. (F) GKO mice were repleted with WT, GKO, or IL27Ra^−/− NK cells and were immunized for EAU. A, B, D, and E are representative of at least two independent experiments. Data in C and F are combined from two independent experiments with total of at least seven mice per group (G) IL-10 was measured in co-culture supernatant of LPS-DCs and NK cells, prepared as in D, or NK cells and DCs stimulated with IFN-γ or IL-27. IL-10 was measured in 48-h culture supernatants. (H and I) IL-10–sufficient or –deficient LPS-DCs and NK cells were co-cultured as in D in all possible permutations. Data in G–I are representative of at least three independent experiments. *, P < 0.05; **, P < 0.01, by linear regression (D–F).