Development and evolution of RORγt+ cells in a microbe's world

The nuclear hormone receptor retinoid-related orphan receptor γt (RORγt) induces a pro-inflammatory program in lymphoid cells, culminating in the expression of interleukin-6 (IL-6), IL-17, IL-22, granulocyte-macrophage colony-stimulating factor, and tumor necrosis factor. During ontogeny, the first type of cells expressing RORγt are lymphoid tissue inducer cells, a type of innate lymphoid cell (ILC) generated in mammalian fetuses to induce the development of lymph nodes and Peyer's patches. After birth, RORγt(+) ILCs and RORγt(+) T cells are involved in the defense of epithelial surfaces against extracellular microbes and play an important role in the intestinal homeostasis with symbiotic microbiota. The development and evolution of RORγt(+) cells is intimately associated with the construction of a stable host-microbe interface.     

Regulated expression of nuclear receptor RORγt confers distinct functional fates to NK cell receptor-expressing RORγt(+) innate lymphocytes

Whether the recently identified innate lymphocyte population coexpressing natural killer cell receptors (NKRs) and the nuclear receptor RORγt is part of the NK or lymphoid tissue inducer (LTi) cell lineage remains unclear. By using adoptive transfer of genetically tagged LTi-like cells, we demonstrate that NKR?RORγt(+) innate lymphocytes but not NK cells were direct progenitors to NKR(+)RORγt(+) cells in vivo. Genetic lineage tracing revealed that the differentiation of LTi-like cells was characterized by the stable upregulation of NKRs and a progressive loss of RORγt expression. Whereas interleukin-7 (IL-7) and intestinal microbiota stabilized RORγt expression within such NKR-LTi cells, IL-12 and IL-15 accelerated RORγt loss. RORγt(+) NKR-LTi cells produced IL-22, whereas RORγt? NKR-LTi cells released IFN-γ and were potent inducers of colitis. Thus, the RORγt gradient in NKR-LTi cells serves as a tunable rheostat for their functional program. Our data also define a previously unappreciated role of RORγt? NKR-LTi cells for the onset or maintenance of inflammatory bowel diseases.     

抑制RORγt活性及Th17分化的新型化合物研究

目的我们在实验中偶然发现了一种能抑制RORγt活性的新型化合物,拟通过鉴定该化合物对Th17体外分化及其细胞因子分泌的影响对其进行生物学评价,以确定该化合物是否可作为一种新型的自身免疫病候选药物。方法构建能在Jurkat细胞中稳定表达Gal4-RORγt的荧光素酶报告系统,通过该体系发现了一种对RORγt有显著抑制活性的新型化合物,并在小鼠Th17细胞分化实验中对该化合物的作用效果进行体外验证。在此基础上,对该化合物进行生物功能分析,包括EC50、CC50的测定以及T细胞特异性分析。结果通过体外验证实验,我们确定该化合物(编号为compound 2)能显著的抑制Th17分化及其细胞因子IL-17A、IL-17F的表达和分泌。在后续的生物学评价中,我们还发现该化合物对RORγt有较高的抑制效率,较低的细胞毒性和较强的T细胞特异性。结论本研究中得到的化合物compound 2可作为一种潜在的新型前导化合物应用于治疗自身免疫性疾病和一些炎症性疾病。     

甲氨蝶呤治疗对类风湿性关节炎患者RORγt基因甲基化水平和Th17/Treg平衡的影响

目的探讨RA患者MTX对视黄酸相关孤儿受体γt(RORγt)启动子甲基化水平和Th17/Treg平衡的影响。方法选择2016年8月至2017年7月于我院确诊并使用MTX进行治疗的RA患者89例,所有患者均接受小剂量MTX维持治疗。分别于治疗前及开始治疗后3个月抽取外周静脉血,使用流式细胞仪检测Th17及Treg含量,并分析Th17细胞RORγt甲基化水平,而后分析二者之间的相关性。结果患者治疗后Th17(2.75%±0.83%vs 3.39%±0.74%)及Th17/Treg(0.26±0.09 vs 0.41±0.13)较治疗前显著下降(P0.001),而Treg(7.29%±1.49%vs 6.13%±1.42%)较治疗前显著上升(P0.001)。RORγt(mRNA)(2.98±0.69 vs 4.15±0.78)表达水平较治疗前显著下降(P0.001)。患者治疗后RORγt序列1(32.21±4.71 vs 40.15±6.58)及序列2(20.73±3.3 vs 25.65±4.61)甲基化水平显著低于治疗前(P0.001)。Pearson相关性分析结果显示,患者治疗前后RORγt序列1及序列2甲基化水平均与Th17/Treg呈正相关(r=0.345,0.297;P0.05)。结论 MTX能有效抑制RORγt基因的甲基化水平,改善RA患者Th17/Treg失衡的作用,提示RORγt甲基化水平可能参与对Th17/Treg平衡的调节。     

Functionally distinct IFN-γ+IL-17A+ Th cells in experimental autoimmune uveitis: T-cell heterogeneity,migration, and steroid response

Immunopathogenic roles for both Th1 (CD4⁺IFN-γ⁺) and Th17 (CD4⁺IL-17A⁺) cells have been demonstrated in experimental autoimmune uveitis (EAU). However, the role for Th17/Th1 (CD4⁺ T cells co-expressing IFN-γ and IL-17A) cells in EAU is not yet understood. Using interphotoreceptor retinoid-binding protein peptide-induced EAU in mice, we found increased levels of Th17/Th1 cells in EAU retinae (mean 9.6 ± 4.2%) and draining LNs (mean 8.4 ± 3.9%; p = 0.01) relative to controls. Topical dexamethasone treatment effectively reduced EAU severity and decreased retinal Th1 cells (p = 0.01), but had no impact on retinal Th17/Th1 or Th17 cells compared to saline controls. Using in vitro migration assays with mouse CNS endothelium, we demonstrated that Th17/Th1 cells were significantly increased within the migrated population relative to controls (mean 15.6 ± 9.5% vs. 1.9 ± 1.5%; p = 0.01). Chemokine receptor profiles of Th17/Th1 cells (CXCR3 and CCR6) did not change throughout the transendothelial migration process and were unaffected by dexamethasone treatment. These findings support a role for Th17/Th1 cells in EAU and their resistance to steroid inhibition suggests the importance of targeting both Th17 and Th17/Th1 cells for improving therapy.     

RORγt+ innate lymphoid cells regulate intestinal homeostasis by integrating negative signals from the symbiotic microbiota

Lymphoid cells that express the nuclear hormone receptor RORγt are involved in containment of the large intestinal microbiota and defense against pathogens through the production of interleukin 17 (IL-17) and IL-22. They include adaptive IL-17-producing helper T cells (T(H)17 cells), as well as innate lymphoid cells (ILCs) such as lymphoid tissue-inducer (LTi) cells and IL-22-producing NKp46+ cells. Here we show that in contrast to T(H)17 cells, both types of RORγt+ ILCs constitutively produced most of the intestinal IL-22 and that the symbiotic microbiota repressed this function through epithelial expression of IL-25. This function was greater in the absence of adaptive immunity and was fully restored and required after epithelial damage, which demonstrates a central role for RORγt+ ILCs in intestinal homeostasis. Our data identify a finely tuned equilibrium among intestinal symbionts, adaptive immunity and RORγt+ ILCs.     

RORγt蛋白复合体在Th17细胞分化中的“阴-阳”调节

Th17细胞是诱导组织炎症反应的辅助性T细胞,而维甲酸相关孤核受体γt(RORγt)是Th17细胞的主要转录因子(TF)。目前普遍认为转录因子不会单独发挥功能,因此RORγt会形成介导Th17细胞分化的蛋白复合体。鉴于RORγt及其潜在复合体在Th17细胞发育和功能中的重要性,本综述总结了近期有关RORγt和潜在伙伴蛋白之间相互作用的进展,特别讨论了导致Th17细胞分化的机制及其炎症反应疾病中的潜在临床干预靶点。     

Th17 cells express interleukin-10 receptor and are controlled by Foxp3⁻ and Foxp3+ regulatory CD4+ T cells in an interleukin-10-dependent manner

1. Download : Download high-res image (287KB)
2. Download : Download full-size image

Highlights? IL-10 signaling in T cells controls the emergence of Th17 and Th17+Th1 cells ? IL-10Rα is highly expressed by IL-17A-producing CD4⁺ T cells in vivo ? Tr1 and Treg cells can both independently suppress IBD induced by Th17 and Th17+Th1 cells ? Suppression of Th17 and Th17+Th1 cells by Tr1 or Treg cells is dependent on IL-10     

洋地黄毒苷通过调控转录因子RORγt的表达抑制Th17细胞产生IL-17A

目的研究洋地黄毒苷(Digitoxin)在体外对人CD4+T细胞产生IL-17A和IFN-γ的影响及其机制。方法人PBMC用抗CD3和抗CD28抗体共刺激或在诱导Th17分化的条件下,同时加或不加Digitoxin,用酶联免疫吸附法检测培养上清中IL-17A和IFN-γ的产生。利用流式细胞术检测细胞因子IL-17A、IFN-γ以及转录因子RORγt的表达。利用Western blotting检测Digitoxin对RORγt表达的影响。结果 Digitoxin对人IL-17A有显著的剂量依赖性抑制作用,并且在刺激的早期(第0小时和第6小时)加入Digitoxin能显著地抑制IL-17A的产生,但不影响IFN-γ的产生,而晚期(第12小时和第24小时)加入Digitoxin对IL-17A的产生没有明显地抑制作用。进一步研究发现,Digitoxin抑制Th17细胞的分化,调控IL-17A的转录因子RORγt的表达。结论本实验表明,Digitoxin通过早期调控RORγt的表达来抑制IL-17A的产生,但不抑制IFN-γ的产生,提示Digitoxin除了对心血管作用外,也可以抑制炎症性疾病和自身免疫性疾病的发生发展。