Investigating aging and membrane-associated T cell receptor signaling in human CD

描述(申请人提供)：随着年龄的增长，T细胞免疫功能发生变化，导致感染和恶性肿瘤的风险增加。T细胞在识别抗原提呈细胞(APC)通过T细胞受体(TCR)提呈的抗原肽时被激活。这种TCR的触发在T细胞中诱导了一系列称为信号转导的生化事件，这是T细胞功能所必需的。有趣的是，在小鼠和人类中已经报道了与年龄相关的TCR信号转导的改变，这与T细胞功能受损有关。然而，在大多数人类研究中，使用的是未分选的总CD4或CD8T细胞，尽管存在不同的CD4和CD8T细胞亚群，如NA和记忆性细胞，具有不同的功能，并且NAVE和记忆性CD4和CD8T细胞的比例随着年龄的变化而改变。最近，我的实验室测量了年轻人和老年人CD8 T细胞上IL-7受体α链(IL-7ra，CD127)的表达，它决定了对IL-7的反应，IL-7是一种关键的T细胞动态平衡的细胞因子。在本研究中，在记忆性CD8 T细胞中发现了高表达IL-7和低表达IL-7的细胞。令人惊讶的是，与年轻人(年龄=40岁)相比，老年人(65岁)的IL-7记忆性CD8 T细胞有较低的扩增。IL-7激活的记忆性CD8T细胞增殖较差，提示这些细胞存在TCR信号缺陷(S)，IL-7ra可能作为识别TCR反应受损的记忆性CD8T细胞的标志物。值得注意的是，在TCR触发和IL-15的共同作用下，IL-7低记忆性CD8 T细胞很好地增殖，IL-15是记忆性CD8 T细胞维持所必需的细胞因子。提示IL-15在恢复IL-7低记忆性CD8 T细胞受损的TCR信号转导中起一定作用。理解这些发现的机制(S)很重要，因为记忆CD8T细胞是宿主防御的关键，而IL-15被认为是细胞免疫的佐剂。这项拟议的研究将基于以下假设来解决这些问题：随着年龄的增长，扩增的IL-7低记忆性CD8T细胞在TCR信号转导方面存在缺陷(S)，该缺陷可以被IL-15拯救。这一假说将通过共聚焦显微镜和流式细胞术对一组与T细胞膜密切相关的TCR信号分子进行研究。具体地说，计划实现以下两个目标。首先，调查IL-7是否以及为什么随着年龄增长而扩张的低记忆性CD8T细胞在TCR信号转导方面存在缺陷(S)。此外，还将通过测量细胞膜胆固醇含量和T细胞表面分子的糖基化来研究其他CD8T细胞亚群TCR信号转导的变化(S)以及CD8T细胞亚群TCR信号转导缺陷的潜在机制。其次，将确定IL-15如何挽救IL-7低记忆性CD8 T细胞受损的TCR信号转导。这项研究的结果将提供重要的信息，通过增强细胞免疫来更好地预防老年人的肿瘤和感染。目前这项提议的目标是研究衰老如何通过CD8 T细胞的受体(T细胞受体)影响参与将刺激从外部传递到内部(信号转导)的分子。这项研究的结果将为增强老年人的免疫反应提供重要信息，从而改善对感染和肿瘤的保护。

DESCRIPTION (provided by applicant): Alterations in T cell immunity occur with aging, contributing to increased risk of infection and malignancy. T cells are activated as they recognize antigenic peptide presented by antigen-presenting cells (APCs) via T cell receptors (TCR). This TCR triggering induces a series of biochemical events called signal transduction in T cells which are essential for T cell functions. Of interest, age-associated alterations in TCR signal transduction have been reported in mice and humans, linking to impaired T cell functions. However, in most human studies unsorted total CD4+ or CD8+ T cells have been used although different CD4+ and CD8+ T cell subsets, such as na¿ve and memory cells, with distinct functions exist and the proportion of na¿ve and memory CD4+ and CD8+ T cells alters with aging. Recently, my lab measured expression of IL-7 receptor alpha chain (IL-7Ra, CD127) which dictates the response to IL-7, a cytokine critical for T cell homeostasis, on CD8+ T cells in young and elderly individuals. In this study, cells expressing IL-7Rahigh and low were found in memory CD8+ T cells. Surprisingly, the elderly (age=65) had expansion of IL-7Ralow memory CD8+ T cells compared to the young (age=40). IL-7Ralow memory CD8+ T cells poorly proliferated in response to TCR triggering, indicating a TCR signaling defect(s) in these cells and the potential role for IL-7Ra as a marker for identifying memory CD8+ T cells with impaired TCR responses. Of interest, IL-7Ralow memory CD8+ T cells nicely proliferate in response to a combination of TCR triggering and IL-15, a cytokine essential for memory CD8+ T cell maintenance. This suggests a role for IL-15 in recovering the impaired TCR signal transduction in IL-7Ralow memory CD8+ T cells. Understanding the mechanism(s) for these findings is important since memory CD8+ T cells are essential for host defense and IL-15 is considered as an adjuvant for cellular immunity. The proposed study will address these issues based on the hypothesis that expanded IL-7Ralow memory CD8+ T cells with aging have a defect(s) in TCR signal transduction which can be rescued by IL-15. This hypothesis will be investigated focusing on a group of TCR signaling molecules that are closely associated with T cell membrane using confocal microscopy and flow cytometry. Specifically, following two aims are planned. First, investigate whether and why IL-7Ralow memory CD8+ T cells that expand with aging have a defect(s) in TCR signal transduction. Plus, alteration(s) in TCR signal transduction of other CD8+ T cell subsets and the potential mechanisms for defective TCR signal transduction in the CD8+ T cell subsets will be studied by measuring cholesterol content of the cell membrane and glycosylation of T cell surface molecules that are known to affect TCR signal transduction with aging. Secondly, it will be determined how IL-15 rescues impaired TCR signal transduction in IL-7Ralow memory CD8+ T cells. The results of this study will provide essential information that leads to better protection against tumors and infections in the elderly by enhancing cellular immunity. The goal of the current proposal is to study how aging affects the molecules involved in transmitting stimulations from the outside to the inside (signal transduction) of CD8+ T cells through their receptors (T cell receptor). The results of this study will provide important information on enhancing immune responses in the aged populations, leading to improved protection against infections and tumors.