Mechanisms of Peripheral Induction of T-Cell Tolerance

性状（由申请方提供）：T淋巴细胞必须通过快速增殖和分化对特异性抗原作出反应，以产生有效的免疫应答。关键的是，这只发生在对外来抗原的反应中，这样自身抗原就不会诱导自身免疫反应。对自身抗原的耐受性部分是通过胸腺中的阴性选择实现的。然而，这并不完全，一些自身反应性T细胞逃逸到外周。存在使这些成熟T细胞耐受的机制，但对它们知之甚少。 该计划正在使用体外和体内模型解决成熟T细胞中这些机制的性质。在项目I（Jenkins）和II（Mueller）中研究了CD 4 T细胞的外周耐受性，在项目III（Hogquist）和IV（Mescher）中研究了CD 8 T细胞的外周耐受性。因此，T细胞的两个主要亚群都在研究中。此外，项目I和III使用新模型在生物学水平上检查CD 4和CD 8 T细胞耐受性，项目II和IV在分子水平上检查CD 4和CD 8 T细胞的耐受机制。计划的工作涉及调查人员之间的广泛协作互动。 预计这些研究中获得的结果将有助于更好地了解如何避免自身免疫。了解这些机制，从而如何操纵它们，有可能有助于改善移植和疾病治疗。诱导机制 对自身抗原的耐受性也可诱导对外来抗原（包括存在于肿瘤或病毒感染的细胞上的抗原）的耐受性，导致免疫系统不能产生保护性应答。最后，使用确定的肽抗原来诱导针对广泛疾病的保护性或治疗性免疫具有巨大潜力，并且目前大量的努力集中在这方面。然而，越来越清楚的是，必须非常谨慎地使用这些药物，因为它们也可能引起耐受性，从而可能导致保护作用减弱或疾病加重。因此，如本计划中所提出的，更好地理解可能导致T细胞耐受的机制，其意义远远超出了自身免疫性疾病。 项目1：体内外周耐受性分析（Jenkins，M.） 项目1描述（由申请方提供）：本项目的长期目标是了解导致CD 4 + T细胞对次级淋巴器官（而非胸腺）中呈递的抗原耐受的机制。以前，我们表明，幼稚的CD 4 + T细胞暴露于模型抗原在次级淋巴器官中的炎症的情况下增殖不良，然后大多数的后代死亡，和幸存者进入一个无能的状态，其特征在于穷人的淋巴因子生产。本应用程序的目标是确定一系列类似的事件是否解释了对某些天然自身蛋白的外周耐受性。自从发现AIRE转录因子驱动胸腺外基因产物的异位表达以来，这已经成为一个紧迫的问题。因此，外周耐受被认为发挥的生理作用仍然可能是由AIRE介导的胸腺内耐受发挥的。在这里，我们将测试外周耐受性是生理相关的假设，通过使用新的工具来确定适用于两种妊娠特异性蛋白质的耐受机制，一种似乎是由AIRE调节的，另一种则不是，还有一种精子特异性蛋白质。我们将确定这些蛋白质在从未在相关组织中表达过它们的小鼠中是否具有免疫原性（例如，未怀孕的雌性小鼠），并且在表达后在小鼠中变得无免疫原性（例如，怀孕的雌性小鼠）。相关的抗原肽将 鉴定并用于产生肽-MHC II多聚体。然后，多聚体将与灵敏的新富集方法一起使用，该方法能够检测每只小鼠少于100个细胞，以在多克隆抗体内的相关蛋白表达之前、期间和之后计数肽MHC II特异性CD 4 + T细胞。 正常小鼠的所有基因组。这种方法应该揭示相关的CD 4 + T细胞是否被删除，变成调节细胞，或者在这些发育调节的自身蛋白表达期间或之后变得无能。然后将这种方法用于基因靶向小鼠，以确定诸如Fas和Cbl-b以及由P01的其他成员鉴定的其他分子是否参与鉴定的耐受机制。成功将提供第一个明确的识别外周耐受机制，适用于天然自身抗原。这些信息应该有助于将未来的研究重点放在相关机制上，并阐明它可能失败并导致自身免疫的潜在方式。

DESCRIPTION (provided by applicant): T lymphocytes must respond to specific antigen by rapid proliferation and differentiation to mount an effective immune response. It is critical that this occur only in response to foreign antigen, so that self antigens do not induce autoimmune responses. Tolerance to self-antigen is achieved in part by negative selection in the thymus. This is not complete, however, and some self-reactive T cells escape into the periphery. Mechanisms exist for rendering these mature T cells tolerant, but they are poorly understood. This Program is addressing the nature of these mechanisms in mature T cells using both in vitro and in vivo models. Peripheral tolerance in CD4 T cells is being studied in Projects I (Jenkins) and II (Mueller), and in CD8 T cells in Projects III (Hogquist) and IV (Mescher). Thus, both major subsets of T cells are being studied. In addition, Projects I and III examine CD4 and CD8 T cell tolerance at the biological level using novel models, and Projects II and IV examine tolerance mechanisms in CD4 and CD8 T cells at the molecular levels. The planned work involves extensive collaborative interactions among the investigators. It is anticipated that the findings obtained in these studies will contribute to a better understanding of how autoimmunity is avoided. Understanding of these mechanisms, and hence how to manipulate them, has the potential to contribute to improvements in transplantation and disease therapy. Mechanisms that induce tolerance to self-antigens may also induce tolerance to foreign antigens, including those present on tumors or virus-infected cells, resulting in the immune system failing to mount a protective response. Finally, there is great potential for using defined peptide antigens to induce protective or therapeutic immunity for a broad range of diseases and a great deal of current effort is focusing on this. However, it is becoming increasingly clear that these must be used with great caution since they can also induce tolerance that may lead to lessened protection or exacerbated disease. Thus, developing a better understanding of the mechanisms that can lead to T cell tolerance, as proposed in this Program, has implications well beyond autoimmune diseases. PROJECT 1: Analysis of Peripheral Tolerance in vivo (Jenkins, M.) PROJECT 1 DESCRIPTION (provided by applicant): The long-term goal of this project is an understanding of the mechanisms that account for CD4+ T cell tolerance to antigens that are presented in the secondary lymphoid organs but not the thymus. Previously, we showed that naive CD4+ T cells that are exposed to model antigens in the secondary lymphoid organs in the absence of inflammation proliferate poorly, then most of the progeny die, and the survivors enter an anergic state characterized by poor lymphokine production. The goal of this application is to establish whether or not a similar series of events accounts for peripheral tolerance to certain natural self-proteins. This has become a pressing issue since the discovery that the AIRE transcription factor drives ectopic expression of extrathymic gene products in the thymus. Thus, it remains possible that the physiological role that peripheral tolerance was thought to play is really played by AIRE-mediated intrathymic tolerance. Here, we will test the hypothesis that peripheral tolerance is physiologically-relevant by using new tools to identify the tolerance mechanisms that apply to two pregnancy-specific proteins, one that appears to be regulated by AIRE and another that does not, and one sperm-specific protein. We will determine whether or not these proteins are immunogenic in mice that have never expressed them in the relevant tissue (e.g., non-pregnant female mice), and become non-immunogenic in mice after expression (e.g., pregnant female mice). The relevant antigenic peptides will be identified and used to produce peptide-MHC II multimers. The multimers will then be used with a sensitive new enrichment method capable of detecting fewer than 100 cells per mouse to enumerate peptide MHC II specific CD4+ T cells before, during, and after expression of the relevant protein within the polyclonal repertoires of normal mice. This approach should reveal whether or not the relevant CD4+ T cells are deleted, turn into regulatory cells, or become anergic during or after the period when these developmentally regulated self-proteins are expressed. This approach will then be used in gene-targeted mice to determine whether or not molecules such as Fas and Cbl-b, and others identified by other members of the P01, are involved in the identified tolerance mechanism. Success would provide the first definitive identification of the peripheral tolerance mechanism that applies to a natural self-antigen. This information should help focus future research on the relevant mechanism and shed light on the potential ways that it could fail and lead to autoimmunity.