Therapeutic Immunoregulation Mediated by TGF-beta-induced iTregs in Autoimmune Ar

自身免疫 Ar 中 TGF-β 诱导的 iTreg 介导的治疗性免疫调节

• 批准号: 8721028
• 负责人: Song Guo Zheng
• 金额: $ 20.65万
• 依托单位: PENNSYLVANIA STATE UNIV HERSHEY MED CTR
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-07-01 至 2015-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8721028
• 关键词: Therapeutic; Immunoregulation; Mediated; TGF; beta

DESCRIPTION (provided by applicant): Present approaches are unable to cure rheumatoid arthritis (RA) and other chronic autoimmune diseases. The current proposal plans to develop a novel approach that tests the central hypothesis: CD4?? regulatory T cells (iTregs) generated with IL-2 and TGF-2 are stable in an inflammatory milieu and are able to treat collagen-induced arthritis (CIA). While thymus- derived, naturally-occurring CD4???? (nTregs) suppress Th1- or Th2-cell-mediated autoimmune diseases, these cells are less successful in controlling (IL-17-producing) Th17 cell- mediated diseases such as CIA. Additionally, unlike iTregs, nTregs have increased cell plasticity, and can be converted into Th1, Th2 or Th17 cells while losing their suppressive activities in the presence of pro-inflammatory cytokines. We and others have established TGF-2 is able to convert naove CD4? cells to iTregs that share similar phenotypic and functional characteristics with nTregs. Interestingly, our recent studies revealed that unlike nTregs, iTregs did not make conversion to Th17 and Th1 cells in the presence of pro-inflammatory cytokines. In addition, iTregs but not nTregs maintained the suppressive activity against T cell response in the presence of IL-6 in vitro. Moreover, iTregs but not nTregs even prevented other T cells from becoming Th17 cells in the presence of IL-6 and TGF-2. However, pretreated nTregs with IL- 2/TGF-2 or atRA alters the plasticity and restore functionality of nTregs. Based on these preliminary data, we anticipate that iTregs and pretreated nTregs are stable following adoptive transfer into the established autoimmune arthritis. We expect iTregs and pretreated nTregs can significantly ameliorate the clinical signs of the established arthritis following treatment. We also expect that combination of all-trans retinoic acid (atRA) and TGF-2, or antigen-specific iTregs can enhance the therapeutic effects of iTregs on the established arthritis. We believe that iTregs not only directly suppress T cell response, but also induce the formation of tolerogenic DCs and these DCs produce IL-27, atRA and/or IDO that eventually restrain Th17 cell differentiation and function. Accordingly, the project has three specific aims: 1) Determine the relative stability of nTreg and iTreg cells when adoptively transferred into established collagen-induced arthritis (CIA). nTreg or iTregs will be sorted or induced from DBA/1 Foxp3-GFP knock-in mice or Foxp3-GFP/IL-17-RFP double knock-in mice. nTreg or iTregs cells will be adoptively transferred into DBA/1 or C57BL/6 mice at day 14 or day 28 after immunization with collagen II (CII) and Complete Freund's Adjuvant (CFA). The migration, distribution, survival, phenotype (Foxp3) and conversion into T help cells (Th1, Th2 or Th17 cells) of Treg subsets will be monitored with GFP and RFP expression. 2) Compare the therapeutic effect of both nTreg and iTreg cells on development of collagen-induced arthritis. nTreg, iTreg or control cells will be administrated to DBA/1J mice on day 14 or day 28 after immunization with CII/CFA. The protective effect of these cells will be judged by arthritis incidence and severity, levels of anti-CII IgG2a antibodies in sera and histological examination of arthritic limbs. 3) Define the cellular and molecular mechanism(s) by which iTregs are resistant to Th17 cell conversion and regulate Th17 cell differentiation and function in the inflammatory milieu. We will examine whether the transcription factor T-bet and Th1 cytokine expression in iTregs are responsible for their resistance. We will also examine whether these factors or tolerogenic DCs induced by iTregs contribute to suppressing Th17 differentiation and function. The results from this study will promote the understanding of therapeutic effects of Treg cells in the prevention and cure of rheumatoid arthritis. If successful, this project will also have a direct clinical relevance and possibly provide a novel approach to treat RA and other autoimmune diseases which may not have the severe side effect(s) characteristic of current therapies.

描述（由申请人提供）： 目前的方法无法治愈类风湿性关节炎（RA）和其他慢性自身免疫性疾病。目前的提案计划开发一种新方法来测试中心假设：CD4??由 IL-2 和 TGF-2 产生的调节性 T 细胞 (iTreg) 在炎症环境中稳定，能够治疗胶原诱导的关节炎 (CIA)。而胸腺衍生的、天然存在的 CD4？？？ (nTreg) 抑制 Th1 或 Th2 细胞介导的自身免疫性疾病，但这些细胞在控制（产生 IL-17）Th17 细胞介导的疾病（如 CIA）方面不太成功。此外，与 iTreg 不同，nTreg 具有增加的细胞可塑性，并且可以转化为 Th1、Th2 或 Th17 细胞，同时在促炎细胞因子存在时失去其抑制活性。我们和其他人已经确定TGF-2能够转化naove CD4吗？细胞转化为与 nTreg 具有相似表型和功能特征的 iTreg。有趣的是，我们最近的研究表明，与 nTreg 不同，iTreg 在促炎细胞因子存在的情况下不会转化为 Th17 和 Th1 细胞。此外，在体外存在IL-6的情况下，iTregs（而非nTregs）保持了对T细胞反应的抑制活性。此外，在 IL-6 和 TGF-2 存在的情况下，iTregs（而不是 nTregs）甚至可以阻止其他 T 细胞变成 Th17 细胞。然而，用 IL-2/TGF-2 或 atRA 预处理的 nTreg 会改变 nTreg 的可塑性并恢复其功能。根据这些初步数据，我们预计 iTreg 和预处理的 nTreg 在过继转移至已建立的自身免疫性关节炎后是稳定的。我们预计 iTreg 和预处理的 nTreg 可以显着改善治疗后已确定的关节炎的临床症状。我们还期望全反式视黄酸（atRA）和TGF-2或抗原特异性iTregs的组合可以增强iTregs对已形成的关节炎的治疗效果。我们认为iTregs不仅直接抑制T细胞反应，还诱导耐受性DC的形成，这些DC产生IL-27、atRA和/或IDO，最终抑制Th17细胞的分化和功能。因此，该项目有三个具体目标：1) 确定 nTreg 和 iTreg 细胞过继转移到已建立的胶原诱导关节炎 (CIA) 中时的相对稳定性。 nTreg或iTreg将从DBA/​​1 Foxp3-GFP敲入小鼠或Foxp3-GFP/IL-17-RFP双敲入小鼠中分选或诱导。在用II型胶原(CII)和弗氏完全佐剂(CFA)免疫后第14天或第28天将nTreg或iTreg细胞过继转移至DBA/1或C57BL/6小鼠中。 Treg 亚群的迁移、分布、存活、表型 (Foxp3) 和转化为 T 辅助细胞（Th1、Th2 或 Th17 细胞）将通过 GFP 和 RFP 表达进行监测。 2）比较nTreg和iTreg细胞对胶原诱导性关节炎发展的治疗效果。在用CII/CFA免疫后第14天或第28天将nTreg、iTreg或对照细胞施用于DBA/1J小鼠。这些细胞的保护作用将根据关节炎的发病率和严重程度、血清中抗CII IgG2a抗体的水平以及关节炎肢体的组织学检查来判断。 3) 定义 iTregs 抵抗 Th17 细胞转化并在炎症环境中调节 Th17 细胞分化和功能的细胞和分子机制。我们将检查 iTreg 中转录因子 T-bet 和 Th1 细胞因子的表达是否与它们的耐药性有关。我们还将检查这些因素或 iTreg 诱导的耐受性 DC 是否有助于抑制 Th17 分化和功能。这项研究的结果将促进人们对Treg细胞预防和治疗类风湿性关节炎的治疗作用的了解。如果成功，该项目还将具有直接的临床意义，并可能提供一种治疗 RA 和其他自身免疫性疾病的新方法，这些疾病可能不具有当前疗法所特有的严重副作用。