Figure 6. see also Figure S4. Phenotype, function and developmental requirements of murine intraepithelial ILC1.

(A) Small intestine IEL of Rag-1^-/- mice were analyzed for the expression of NKp46, NK1.1 and CD160. The majority of NKp46⁺NK1.1⁺ IEL (histogram, black line) express CD160, while NKp46⁺NK1.1⁻ ILC3 (histogram, gray profile) do not. Control staining of NKp46⁺NK1.1⁺ cells is indicated by a dotted line. Cells in the dot plots were gated on live lymphocytes. (B) IEL from Rag-1^-/- mice were stimulated in vitro with IL-12 and IL-15 and analyzed for intracellular IFN-γ. Cells were gated on CD45⁺NK1.1⁺ cells. (C) IEL from C57BL/6 mice were stimulated in vitro with a combination of IL-12 and IL-15, or with IL-23 and IL-1β, and analyzed for intracellular IFN-γ. Cells were gated on CD45⁺CD3⁻NKp46⁺ cells. (D) Frequencies of intraepithelial ILC1 in the small intestine of Nfil3^-/-, Ahr^-/-, Tbx21^-/- and Rorc^-/- mice and littermate controls. Cells were gated on CD45⁺CD3⁻CD19⁻. (E) Small intestinal IEL of conventionally housed (SPF) or germ-free C57BL/6 mice, as well as of adult and neonate C57BL/6 mice, were analyzed for the presence of NKp46⁺NK1.1⁺ cells. Cells were gated on CD45⁺CD3⁻CD19⁻. (F) ILC1 within IEL are largely preserved in Il15ra^-/- mice. Left, frequencies of NKp46⁺NK1.1⁺ cells within spleen and intestinal IEL (gated on CD45⁺CD3⁻CD19⁻) from wild-type and IL15ra^-/- mice from experiments with four mice each. Data are represented as mean +/- SD. Right, representative dot plots from IEL from wild-type and IL15ra^-/- mice. (G) ILC1 from small intestinal epithelium of wild-type (black line) and IL15ra^-/- mice (dotted line) express similar levels of CD160. Cells were gated on CD45⁺CD3⁻CD19⁻, followed by gating on NKp46⁺NK1.1⁺ cells. The gray profile indicates CD160 on wild-type splenic NKp46⁺NK1.1⁺ cells.