Spatial and temporal mapping of cell fate within lymphoid tissue

摘要：单细胞基因组学的最新进展提供了高分辨率的信息 关于细胞身份，然而，将分子信息与空间和时间背景联系起来仍然是一个难题。 重大挑战。这与免疫系统特别相关，因为免疫应答发生在 高度组织化的组织环境，并且涉及细胞状态随时间的严格控制的变化。这里 我们建议开发新的方法来提高单细胞的空间和时间分辨率， 分析，并使用这些方法来生成，然后传播一个4维（时间和3D 空间坐标）图的T细胞发育和相关的微环境中的胸腺。在 目的1，我们将同时测量单个细胞上的mRNA和表面蛋白， 计算和实验方法来开发T细胞期间细胞状态转换的时间图 胸腺的发育。在目标2中，我们将使用相干拉曼和多光子显微镜的生活， 胸腺组织切片，连同激光显微切割，以分离组织的功能相关区域。 然后，我们将对这些定义区域内的单个细胞进行单细胞分析，并使用 计算分析来定义细胞类型和解决细胞串扰。在目标3中，我们将增加 为了提高这一资源对科学界的价值，我们将使研究人员能够随时访问这些数据。 通过用户友好的界面。 !

Summary: Spectacular recent advances in single cell genomics have provided high-resolution information about cell identity, however relating molecular information to the spatial and temporal context remains a major challenge. This is particularly relevant for the immune system, since immune responses occur in highly organized tissue environments, and involve tightly-controlled changes in cell state over time. Here we propose to develop new approaches to increase the spatial and temporal resolution of single cell analyses, and to use these methods to generate and then disseminate a 4-dimensional (time and 3D spatial coordinates) map of T cell development and the associated microenvironment in the thymus. In Aim 1, we will use simultaneous measurement of mRNA and surface proteins on single cells, together with computation and experimental approaches to develop a temporal map of cell state transitions during T cell development in the thymus. In Aim 2, we will use coherent Raman and multiphoton microscopy of living thymic tissue slices, together with laser microdissection, to isolate functionally relevant regions of tissues. We will then perform single cell analyses of individual cells within these defined regions and use computational analyses to define cell types and resolve cellular cross-talk. In Aim 3, we will increase the value of this resource to the scientific community, we will make the data readily accessible to researchers via a user-friendly interface. !