The Role of Calcium Ions in Lymphocyte Activation

描述(由申请人提供)：本实验室已率先使用 膜片钳和光学技术研究离子通道和 (CA)T淋巴细胞内的信号转导。T细胞启动免疫反应 与抗原提呈细胞直接接触后的外来抗原 (APC)，如树突状细胞。T细胞受体(TCR)的参与导致 细胞内钙信号导致钙依赖基因表达，细胞因子 分泌、增殖和分化。我们计划进行的实验 使用活细胞成像的方法将与这一系列事件平行。 双光子显微镜最近使T细胞和B细胞的深度可视化成为可能 在完整的淋巴结内，并显示了运动和运动的动态模式 细胞-细胞相互作用，为体内成像提供了独特的机会 免疫系统功能。因此，我们的方法将致力于巩固以下成果 抗原特异性原代T细胞的体外和体内实验 来自转基因小鼠和T细胞系。各种引进的和遗传的 编码的荧光记者将直接监测TCR，(Ca)，基因 刺激前后T细胞的表达和分泌反应。一个 该项目的另一个目标是比较 光中静止、激活和长期激活的自身反应性T细胞 它们不同的离子通道表型。

DESCRIPTION (provided by applicant):Our laboratory has pioneered the use of patch-clamp and optical techniques to investigate the role of ion channels and (Ca) signaling in T lymphocytes. T cells initiate the immune response to foreign antigens following direct contact with primed antigen presenting cells (APC) such as dendritic cells. T cell receptor (TCR) engagement results in a cytosolic Ca (Ca) signal that leads to Ca dependent gene expression, cytokine secretion, proliferation, and differentiation. Our proposed experimental approach using live-cell imaging will parallel this sequence of events. Two-photon microscopy has recently enable visualization of T and B cells deep within intact lymph node, and has revealed a dynamic pattern of motility and cell-cell interaction, offering a unique opportunity for in vivo imaging of immune system function. Thus, our approach will aim to consolidate results from in vitro and in vivo experiments performed on antigen-specific primary T cells from transgenic mice and T cell lines. A variety of introduced and genetically encoded fluorescent reporters will monitor the TCR directly, (Ca), gene expression, and secretory responses in T cells before and after stimulation. A further goal of this project is to compare the functional responses of quiescent, activated, and chronically activated auto reactive T cells in light of their differing ion channel phenotypes.