IL‐4 is a differentiation factor for transforming growth factor‐β secreting Th3 cells and oral administration of IL‐4 enhances oral tolerance in experimental allergic encephalomyelitis

We have previously shown that following oral administration of myelin basic protein (MBP), regulatory T cells are generated from gut-associated lymphoid tissue and that these cells suppress experimental allergic encephalomyelitis (EAE). These regulatory T cells produce transforming growth factor-β (TGF-β) with various amounts of IL-4 and IL-10 and these TGF-β-secreting T cells have been termed Th3 cells. T cells in lymphoid organs drained by mucosal sites secrete IL-4 as a primary T cell growth factor. In the present study, we examined the role of IL-4 on oral tolerance and in the generation of TGF-β secreting cells. Treatment of (PLJ × SJL)F1 mice with intraperitoneal (i.  p.) IL-4 and low-dose oral MBP (0.5 mg) given three times reduced the severity of EAE, whereas i.  p. injection of IL-4 alone or oral MBP alone given in these suboptimal doses, showed no protection. Spleen cells from protected mice produced increased amounts of TGF-β and reduced IFN-γ upon stimulation with MBP in vitro. Mucosal MBP-specific IgA production was significantly increased in IL-4 plus MBP fed animals. Moreover, oral administration of IL-4 (1 μg per feeding) also enhanced the suppression of EAE by oral MBP and this protective effect was reversed by administration of anti-TGF-β antibody in vivo. Reverse transcription-PCR showed enhanced suppression of IFN-γ in Peyer's patch in animals fed MBP and IL-4 versus those fed MBP alone. We then investigated the role of IL-4 in the generation of TGF-β-secreting cells using MBP Ac1-11 TCR transgenic animals. Cells were cultured with IL-2, IL-4, or IFN-γ in the presence of MBP and limiting dilution analysis for cytokine-secreting cells performed. We found that IL-4, but not IL-2 or IFN-γ, generated TGF-β-secreting T cells from naive splenic T cells and that these cells provided help for IgA production. These findings demonstrate that IL-4 is a differentiation factor for TGF-β-secreting Th3 cells and oral IL-4 has a synergistic effect on low-dose oral tolerance that is associated with increased TGF-β secretion.