PMC:2791889 / 5900-8905 JSONTXT

{"target":"https://pubannotation.org/docs/sourcedb/PMC/sourceid/2791889","sourcedb":"PMC","sourceid":"2791889","source_url":"http://www.ncbi.nlm.nih.gov/pmc/2791889","text":"IL-12 and High Doses of Antigen Induce the Development of Th1 Cells Producing IL-10\nTo study the differentiation of Th1 cells coproducing IFN-γ and IL-10, we cultured purified TCR-transgenic DO11.10 naive CD4+ T cells with purified DCs as APCs and increasing doses of ovalbumin peptide 323-339 (OVA). Culture with high doses of antigen for 7 days gave rise to Th1 cells expressing IFN-γ upon restimulation (Constant et al., 1995; Hosken et al., 1995), but not IL-10 (Figure S1A available online). Culture with low antigen doses under the same conditions led to the differentiation of Th2 cells, which expressed both IL-4 and IL-10 upon restimulation (Figure S1A). Culture of naive CD4+ T cells in an APC-free system by stimulation with anti-CD3 and anti-CD28 antibodies in the presence of IL-12 resulted in IFN-γ-producing Th1 cells, a proportion of which coproduced IL-10, as did Th2 cells resulting from culture in IL-4 (Figure S1B).\nTo investigate whether the lack of IL-10 produced by Th1 cells resulted from inhibition of IL-10 production by DCs and high antigen dose or alternatively required IL-12, we cultured naive CD4+ T cells with increasing doses of antigen presented by DC in the presence of IL-12. At low doses of antigen, IL-12 abrogated the development of Th2 cells and induced IFN-γ expression but only low levels of IL-10 expression, suggesting that IL-12 per se was not sufficient to induce significant IL-10 production in Th1 cells (Figure 1A). Strikingly, as the antigen dose was increased, Th1 populations driven with IL-12 now contained higher numbers of IL-10-producing cells (Figure 1A) and produced more IL-10 protein upon restimulation (Figures 1A and 1B). Thus, the development of Th1 cells producing IL-10 required both IL-12 and high doses of antigen. Th1 cells differentiated to produce large amounts of IL-10 and IFN-γ and lost their capacity to produce IL-2 (Figure 1B). Because the presence of IL-12 reduced the proliferation of CD4+ T cells at both high and low antigen doses (Table S1), and only the former showed IL-10 production, the development of high IL-10-producing cells is most likely not related to limited IL-2. Naive CD4+ T cells from DO11.10/recombination-activating gene 1 (Rag1)-deficient animals, cultured with high doses of antigen in the presence of IL-12, also resulted in IL-10 expression by Th1 cells, showing that this expression was not dependent on the presence of effector or memory T cells or Treg cells (Figure S2). Although it has been suggested that TGF-β can induce IL-10 in CD4+ T cells (Kitani et al., 2003; Schiott et al., 2000), we found that in developing Th1 and Th2 cells this was not the case (data not shown). In fact, neutralization of TGF-β led to increased IL-10 production by both T cell subsets (Figure S3). Thus, the development of IL-10-producing Th1 cells only depended on the presence of IL-12 together with high antigen dose and not on other soluble factors such as IL-2 or TGF-β or on the presence of other T cell 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