Figure 1. OX40 inhibits induction of IL-17 in vivo and in vitro.

(A) CD4⁺Foxp3⁻ OT-II cells (CD45.1⁺) were transferred into wild-type B6 recipients (CD45.2⁺), and mice were immunized with OVA_(323–339) in CFA a day later. Groups of mice were given OX86 or IgG on days 0, 1, and 2 post immunization. CD4⁺ T cells in draining LN were analyzed on day 10 for IL-17A and IL-17F expression by flow cytometry. The FACS plots shown are cells gated on CD4⁺CD45.1⁺ OT-II cells.

(B and C) The percentage of IL-17A and IL-17F producing cells among gated CD45.1⁺ OT-II cells analyzed on days 5, 7, 10 and 12 post immunization (n = 3 per group; mean ± SD). ** p <0.01

(D, F) Naive CD4⁺ T cells sorted from WT Foxp3gfp and Tnfrsf4 ^−/− Foxp3gfp reporter mice were activated with anti-CD3 plus WT APCs or OX40L-TG APCs under Th17-polarizing conditions or non-polarizing conditions (Controls or Ctrl) for 3 days, and induction of IL-17A(D) and IL-17F (F) producing cells was analyzed and shown.

(E, G) The tabulated percentage of IL-17A (E) and IL-17F (G) producing cells from all experiments performed, and the graph depicts Mean ± SD of 5 experiments with triplicate cultures. ** p <0.01

(H) Volcano plot of RNA-seq data from CD4+ T cells activated with anti-CD3 plus WT-APC (WT) or OX40LTG-APC (OX40L) under Th17 conditions, concentrating on OX40 regulated genes. Black dots indicate differentially expressed genes (minimum two fold changes with a P <0.05), and Il17a and Il17f are shown in Red.

(I) Venn diagram of private and overlapping genes induced by RORγt and OX40, based on RNA-seq data.