Therapeutic immunoregulation mediated by TGF-??-induced iTregs in autoimmune arth

自身免疫性关节炎中 TGF-β 诱导的 iTreg 介导的治疗性免疫调节

• 批准号: 8478044
• 负责人: Song Guo Zheng
• 金额: $ 11.38万
• 依托单位: UNIVERSITY OF SOUTHERN CALIFORNIA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-01 至 2013-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8478044
• 关键词: Accounting; Adoptive Transfer; Adverse effects; Affect; American; Animal Model; Animals; Anti-Inflammatory Agents; Anti-inflammatory; Antibodies; Antigens; Apoptosis; Arthritis; Autoantigens; Autoimmune Diseases; Autoimmune Process; Autologous; Back; Biological; C57BL/6 Mouse; CD4 Positive T Lymphocytes; Cell Differentiation process; Cell Therapy; Cell physiology; Cells; Characteristics; Chronic; Clinical; Code; Colitis; Collagen; Collagen Arthritis; Cytokine Receptors; DBA/1 Mouse; DBA/1J Mouse; Data; Development; Disease; Disease Progression; Disease model; Environment; Enzymes; Exons; Family member; Frequencies; Freund' s Adjuvant; Genes; Green Fluorescent Proteins; Helper-Inducer T-Lymphocyte; Human; IL2RA gene; Immune; Immune response; Immunization; Immunotherapy; In Vitro; Incidence; Inflammation; Inflammatory; Injection of therapeutic agent; Interleukin-17; Interleukin-2; Interleukin-6; Interleukins; Joints; Knock-in Mouse; Lead; Limb structure; Mediating; Modeling; Molecular; Monitor; Multiple Sclerosis; Mus; Nature; Organ; Pathogenesis; Patients; Phenotype; Prevention; Production; Protein Precursors; Regulatory T-Lymphocyte; Relative (related person); Research; Research Project Grants; Resistance; Rheumatoid Arthritis; Role; Safety; Serum; Severities; Signaling Molecule; Sorting - Cell Movement; Spleen; T cell response; T-Lymphocyte; T-bet protein; Testing; Th1 Cells; Th2 Cells; Therapeutic; Therapeutic Effect; Thymus Gland; Time; Transgenic Mice; Tretinoin; United States; Visual; Work; antiarthritic agent; autoimmune arthritis; base; chronic autoimmune disease; clinically relevant; cytokine; disability; disorder control; economic impact; functional restoration; immune function; immunoregulation; in vivo; lymph nodes; migration; novel; novel strategies; overexpression; peripheral blood; prevent; protective effect; protein expression; public health relevance; receptor expression; red fluorescent protein; research study; transcription factor

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关节炎（RA）和其他慢性自身免疫性疾病。当前的建议计划开发一种新的方法，该方法检验中心假设：CD4 ??由IL-2和TGF-2产生的调节性T细胞（ITREGS）在炎症环境中稳定，并且能够治疗胶原蛋白诱导的关节炎（CIA）。虽然胸腺衍生，自然呈现的CD4 ？？？ （NTREG）抑制Th1-或Th2细胞介导的自身免疫性疾病，这些细胞在控制（IL-17产生）TH17细胞介导的疾病（例如CIA）方面不太成功。此外，与ITREGS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，同时在存在促炎性细胞因子的情况下失去其抑制活性。我们和其他人已经建立了TGF-2可以转换NAOVE CD4？与NTREG共享类似表型和功能特征的ITREGS细胞。有趣的是，我们最近的研究表明，与Ntregs不同，ITREGS在存在促炎性细胞因子的情况下并未将其转化为Th17和Th1细胞。此外，在体外IL-6存在下，ITREGS但不能保持对T细胞反应的抑制活性。此外，在IL-6和TGF-2存在下，ITREGS但没有NTREG甚至阻止了其他T细胞成为Th17细胞。但是，用IL-2/TGF-2或ATRA预处理NTreg会改变NTREG的可塑性和恢复功能。基于这些初步数据，我们预计在继承转移到已建立的自身免疫性关节炎中后，ITREG和预处理的NTREG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建立的关节炎的临床体征。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对既定关节炎的治疗作用。我们认为，ITREG不仅可以直接抑制T细胞反应，还可以诱导耐受性DC的形成，这些DC会产生IL-27，ATRA和/或IDO，最终抑制TH17细胞分化和功能。因此，该项目具有三个特定的目的：1）确定NTREG和ITREG细胞的相对稳定性，当将NTREG和ITREG细胞转移到已建立的胶原蛋白诱导的关节炎（CIA）中。 NTREG或ITREGS将从DBA/​​1 FOXP3-GFP敲门小鼠或FOXP3-GFP/IL-17-RFP双重敲击小鼠中进行分类或诱导。 NTREG或ITREGS细胞将在用胶原II（CII）免疫后的第14天或第28天将DBA/1或C57BL/6小鼠转移到DBA/1或C57BL/6小鼠中。 Treg子集的迁移，分布，生存，表型（FOXP3）和转化为Treg子集的T帮助细胞（Th1，Th2或Th17细胞）将通过GFP和RFP表达进行监测。 2）比较NTREG和ITREG细胞对胶原蛋白诱导的关节炎的发育的治疗作用。 NTREG，ITREG或对照细胞将在使用CII/CFA免疫后第14天或第28天将DBA/1J小鼠施用。这些细胞的保护作用将由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四肢的组织学检查。 3）定义了ITREG对TH17细胞转化具有抗性并调节炎症环境中Th17细胞分化和功能的细胞和分子机制。我们将检查ITREG中的转录因子T-bet和Th1细胞因子的表达是否负责它们的抗性。我们还将检查ITREGS诱导的这些因素或耐受性DC是否有助于抑制Th17的分化和功能。这项研究的结果将促进对Treg细胞在预防和治愈类风湿关节炎中的治疗作用的理解。如果成功的话，该项目还将具有直接的临床相关性，并可能提供一种新颖的方法来治疗RA和其他自身免疫性疾病，这些疾病可能没有当前疗法的严重副作用。