To examine the role of interleukin‐23 (IL‐23) in subgroup polarization of IL‐17A–positive and/or interferon‐γ (IFNγ)–positive T cells in autoimmune disease–prone DBA/1 mice with and without collagen‐induced arthritis.

A magnetic‐activated cell sorting system was used to isolate CD4+ T cells from the spleen of naive and type II collagen (CII)–immunized DBA/1 mice. These CD4+ T cells were stimulated in vitro under Th0, Th1, or different Th17 culture conditions. Intracellular staining for IL‐17A and IFNγ was evaluated by flow cytometry. In addition, Th17 cytokines and T helper–specific transcription factors were analyzed by enzyme‐linked immunosorbent assay and/or quantitative polymerase chain reaction.

In CD4+ T cells from naive DBA/1 mice, IL‐23 alone hardly induced retinoic acid–related orphan receptor γt (RORγt), Th17 polarization, and Th17 cytokines, but it inhibited T‐bet expression. In contrast, transforming growth factor β1 (TGFβ1)/IL‐6 was a potent inducer of RORγt, RORα, IL‐17A, IL‐17F, IL‐21, and FoxP3 in these cells. In contrast to TGFβ1/IL‐6, IL‐23 was critical for the induction of IL‐22 in CD4+ T cells from both naive and CII‐immunized DBA/1 mice. Consistent with these findings, IL‐23 showed a more pronounced induction of the IL‐17A+IFNγ– subset in CD4+ T cells from CII‐immunized mice. However, in CD4+ T cells from naive mice, IL‐23 significantly increased the TGFβ1/IL‐6–induced Th17 polarization, including elevated levels of IL‐17A and IL‐17F and decreased expression of T‐bet and FoxP3. Of note, the IL‐23–induced increase in IL‐17A and IL‐17F levels was prevented in T‐bet–deficient mice.

IL‐23 promotes Th17 differentiation by inhibiting T‐bet and FoxP3 and is required for elevation of IL‐22, but not IL‐21, levels in autoimmune arthritis. These data indicate different mechanisms for IL‐23 and TGFβ1/IL‐6 at the transcription factor level during Th17 differentiation in autoimmune experimental arthritis.