T cell-derived iNOS switches off TH17 cell differentiation in inflammation

描述（由申请人提供）：虽然已经证明iNOS在宿主对微生物病原体的防御中起重要作用，但iNOS在T细胞和慢性炎症疾病中的确切功能尚未确定。分泌白细胞介素17 （IL-17）和IL-22的TH17细胞是最近发现的CD4+ T细胞亚群，不同于TH1和TH2亚群，RORgt已被确定为TH17细胞分化的关键转录因子。越来越多的证据表明，TH17免疫应答参与多种自身免疫性/炎症性疾病的发病机制。因此，阻断TH17细胞活化可能导致慢性炎症性疾病治疗新策略的发展。我们发现，iNOS敲除小鼠表现出更强大的TH17细胞分化，而对TH1或TH2细胞系没有重大影响。我们证明了iNOS蛋白在活化的CD4+ T细胞中被诱导，使用iNOS选择性抑制剂L-NIL显著增加WT细胞培养中产生IL-17的CD4+ T细胞的百分比，而NO供体SNAP在WT和iNOS-/- T细胞培养中剂量依赖性地抑制IL-17的产生。另外，对RORgt蛋白的酪氨酸残基进行硝化处理

DESCRIPTION (provided by applicant): Although it has been demonstrated that iNOS plays an important role in host defense against microbial pathogens, the exact function of iNOS in T cells and chronic inflammatory diseases has not been defined. TH17 cells, which secrete interleukin 17 (IL-17) and IL-22 comprise a recently identified subset of CD4+ T cells distinct from the TH1 and TH2 subsets and RORgt has been identified as a key transcription factor for TH17 cell differentiation. Increasing evidence indicates that TH17 immune responses are involved in the pathogenesis of various autoimmune/inflammatory diseases. Thus, blocking TH17 cell activation could lead to the development of novel strategies for the treatment of chronic inflammatory diseases. We show that iNOS knock out mice display more robust TH17 cell differentiation without major effects on either TH1 or TH2 cell lineages. We demonstrated that iNOS protein was induced in activated CD4+ T cells and the use of an iNOS selective inhibitor L-NIL significantly increased the percentage of IL-17-producing CD4+ T cells in WT cell cultures, while, an NO donor, SNAP, dose-dependently suppressed IL-17 production in WT and iNOS-/- T cell cultures. In addition, tyrosine residues of RORgt protein were nitrated resulting in the inhibition of RORgt-mediated IL-17 promoter activation. Finally, transfer of iNOS-/- CD4+CD45Rbhi cells into RAG-/- mice induces more severe colitis compared to control CD4+CD45Rbhi cells and mice reconstituted with iNOS-/- cells had a significantly higher percentage of IL-17-producing cells than control mice. The results suggest that T cell-derived iNOS negatively regulates the development of TH17 immune responses resulting in the control of inflammation. This proposal is structured around three aims: 1) We will characterize the molecular mechanisms involved in the regulation of TH17 cell differentiation by T cell-derived iNOS. 2) We will characterize the function of T cell- derived-iNOS on human TH17, TH1, TH2, and Treg cell development. 3) We will analyze the roles of iNOS in different cell compartments including macrophages, dendritic cells, and T cells in the development of colitis. The proposed studies will define a novel transcriptional inhibitor of TH17 cell differentiation and highlight th importance of T cell-derived iNOS as a novel therapeutic target for the treatment of chronic inflammatory diseases.