TNFα promotes cytokine production by ILC3 and ILC3²³ formation <i>in vivo</i>, and synergizes with IL-23 to directly activate ILC3 <i>in vitro</i>.

Abstract

<p>(A-C) Mice were injected with sham mc (open symbols) or IL-23 mc (filled symbols), with some groups receiving anti-TNFα (or PBS) as indicated. (A) SI length 2 weeks post mc injection (left), and cytokine secretion from SI explants harvested 3 days post mc injection (right) are shown. (B) Flow cytometry was performed on LP cells from the proximal SI three days post mc injection. Absolute numbers of cytokine producing lymphocytes are shown. (C) Flow cytometry was performed on LP cells from the proximal SI 2 weeks post mc injection. Top: absolute numbers are shown. Bottom: representative staining shows the frequency of ILC3²³ amongst CCR6- NCR- ILC3. (D-E) FACS purified ILCs enriched for ILC3 (~90% RORγt+) were sorted from the proximal SI LP of RAG KO mice and cultured overnight with the indicated cytokines prior to flow cytometry. (D) Protein expression by ILC3 is shown for triplicate wells with staining intensity depicted as geometric MFI (top), and representative IL-7R and T-bet staining is shown for total ILC3 (bottom). (E) Cytokine expression by ILC3 is shown for triplicate wells treated with protein inhibitors for the last 3 hours of culture. Scatter plots show means ± SEM for all mice from one of 2–3 similar experiments, 4–5 mice per group, with each symbol representative of a single mouse. Bar graphs show means ± SEM for triplicate wells from one of two similar experiments.</p