Memory, Phenotype, and Function of TB-reactive Human MR1 Restricted T cells

项目摘要 粘膜相关不变T细胞(MAIT细胞)是受MR1(MR1T细胞)限制的T细胞亚群，是一种天然的T细胞。 像T细胞亚群一样，普遍存在于人类中，分布于黏膜部位。人类MAIT细胞被定义为 通过半不变的TcRα链TRAV1-2/TRAJ12/20/33的表示，非 多态主要组织相容性复合体(MHC)I类分子，MHC相关蛋白1(MR1)，以及 通过识别来自核黄素生物合成的小有机分子。我们发现MAIT细胞 两者都是“天生”的效应器，它们在胸腺中的功能证明了这一点，并且都有能力适应 他们的环境，从他们的效应器记忆细胞表面表型和选择性使用TCR来证明。 然而，目前还不清楚这些扩张是由持续的微生物或环境暴露驱动的，还是 MAIT细胞保留了选择性地应对未来抗原挑战的能力。如果MAIT细胞有 记忆，那么它们就可以被利用在未来的疫苗接种策略中。 我们提供了证据证明人类促炎MAIT细胞的寡克隆性扩增。 结核病感染者的呼吸道。我们还提供证据表明，接种卡介苗可以导致 MAIT细胞的功能性扩增，并提供证据表明MAIT细胞具有抗微生物、多细胞毒作用 表型。我们推测MR1T细胞可以促进结核分枝杆菌感染的控制，并将探索 这些细胞是否有免疫记忆。 特异性目的1：确定CD8+MR1T细胞是否具有免疫记忆功能。 通过在抗原暴露前确定MR1T细胞的TCR谱系(人脐血单核细胞 在婴儿接种卡介苗导致抗原暴露后，在结核病的情况下 (成人PBMC)，诊断时肺结核病患者的肺(BAL)和外周血 在治疗之后，我们将确定MAIT细胞是否表现出反映抗原的TCR用途 暴露，抗原暴露后的持久性，以及对抗原存在的依赖。具体来说， 我们预计我们将在人类脐带血中观察到MR1T细胞TCR的广泛谱系，并且在 成年后，我们将观察到以少克隆扩张为特征的较窄的曲目。接种疫苗后 在卡介苗和结核病中，我们希望观察到PBMC和BAL中的寡克隆扩张，并将 预测这些细胞将在治疗后恢复循环。最后，我们将询问这些TCR是否 与离散配基识别相关。 具体目的2：明确MR1T细胞的表型和功能。 体外MR1T细胞和MR1T细胞克隆的联合流式细胞术分析，表达数据 体外MR1T细胞的分析，以及从每个来源的MR1T细胞克隆的机制评估 描述的队列和时间点将被用来定义完整的功能和抗微生物表型 这些细胞。

Project Summary Mucosal associated invariant T (MAIT) cells, a subset of T cells restricted by MR1 (MR1T cells), are an innate- like T cell subset prevalent in humans and distributed throughout mucosal sites. Human MAIT cells are defined by the expression of the semi-invariant TCRα chain TRAV1-2/TRAJ12/20/33, restriction by the non- polymorphic major histocompatibility complex (MHC) class I-like molecule, MHC-related protein 1 (MR1), and by recognition of small organic molecules, derived from riboflavin biosynthesis. We have found that MAIT cells are both “innate” effectors as evidenced by their functionality in the thymus, and have the capacity to adapt to their environment as evidenced by their effector memory cell surface phenotype and selective TCR usage. However, it is not clear if these expansions are driven by ongoing microbial or environmental exposures, or if MAIT cells retain the capacity to respond selectively to future antigenic challenges. If MAIT cells have memory, then they could be harnessed in future vaccination strategies. We provide evidence demonstrating oligoclonal expansions of pro-inflammatory MAIT cells in the human airway of subjects infected with TB. We also provide evidence that vaccination with BCG can result in functional MAIT cell expansion, and provide evidence that MAIT cells have an anti-microbial, polycytotoxic phenotype. We postulate that MR1T cells can facilitate the control of infection with Mtb, and will explore whether or not these cells have immunologic memory. Specific Aim 1: To determine whether or not CD8+ MR1T cells have features of immunologic memory. By determining the TCR repertoire of MR1T cells prior to antigenic exposure (human cord blood mononuclear cells; CBMC), following antigenic exposure as a result of infant vaccination with BCG, in the setting of TB (adult PBMC), in the lungs (BAL) and peripheral blood of subjects with pulmonary TB at the time of diagnosis and following treatment, we will establish if MAIT cells demonstrate TCR usage that reflects antigenic exposure, persistence following antigenic exposure, and dependence on the presence of antigen. Specifically, we anticipate that we will observe a broad repertoire of MR1T cell TCRs in human cord blood, and that in adults we will observe a narrower repertoire characterized by oligocolonal expansions. Following vaccination with BCG and in TB, we would expect to observe oligoclonal expansions in the PBMC and BAL, and would predict that these cells will return to the circulation following treatment. Finally, we will ask if these TCRs are associated with discrete ligand recognition. Specific Aim 2: To define the phenotype and functional capacity of MR1T cells. A combination of flow cytometric analysis of both ex vivo MR1T cells and MR1T cell clones, expression data analysis of ex vivo MR1T cells, and mechanistic evaluation of MR1T cell clones derived from each of the cohorts and time points described will be used to define the full functional and anti-microbial phenotype of these cells.