Spatiotemporal Molecular Substrates of TBI at Single Cell Resolution

摘要 创伤性脑损伤具有复杂的神经病理改变，涉及大脑中心的进行性改变。 它处理认知和情感行为，并由不同的细胞群组成。情结 在进行性脑损伤病理基础上的时空细胞和分子电路可以演变为 慢性创伤性脑病和创伤后应激障碍等疾病仍有待于 明白了。全面了解脑外伤复杂性的分子机制 由于缺乏有效的方法来检查单个脑细胞中的分子事件，这一点受到了阻碍 这推动了整个病理过程。我们最近对海马体进行了一项单细胞分辨率研究 应用单细胞RNA测序(ScRNAseq)和揭示细胞类型特异性的脑损伤急性期(24小时) 创伤性脑损伤的信号转导途径和调节因子。特别是，我们发现抑制细胞新陈代谢是一个关键 脑损伤急性期海马区的致病成分。这一发现表明，追踪 细胞的代谢状态可以用来解决空间和时间依赖的关键知识差距 脑外伤的关键病理驱动因素的研究进展。在这里，我们建议检验细胞代谢的假设 调节器通过利用现代的力量来确定脑损伤的动态和空间致病途径 高通量技术。我们提出了一个高度整合的团队方法，以从以下方面的最新进展中获利 单细胞RNA测序(ScRNAseq)和多重错误稳健荧光原位杂交 (MerFish)以及先进的基因-基因和细胞-细胞网络建模，以告知干预目标 在特定的时间点或大脑部位，这是脑损伤领域一个基本尚未解决的问题。在目标1中，我们建议 利用scRNAseq、MerFish和网络建模方法的独特组合来评估和 验证每种细胞类型在多个时间段的多个脑区对脑损伤的时空易损性 以数据驱动、不偏不倚的方式点位，这可以让我们了解脑外伤发病机制的隐藏调控因素。 我们将关注脑外伤进展过程中细胞代谢途径的空间和时间变化。我们的 初步结果支持mt-Rnr2，编码一种线粒体多肽Human in，并参与细胞 代谢，是脑损伤的主要部位和时间依赖的驱动因素。在目标2中，我们将从功能上评估 调节mt-Rnr2(人素)具有减轻脑损伤病理和防止进展的治疗潜力。 我们还将探索细胞类型的特定机制，特别是新陈代谢的作用，潜在的 人类的行为。该提案的总体目标是详细阐述一种创新战略，该战略可以提供 全面理解脑外伤病理和发现的时空细胞底物 新的靶点和机制，以改变脑损伤的进程，以克服随后的神经障碍。

Abstract Traumatic brain injury (TBI) has a complex neuropathology involving progressive alterations in brain centers that process cognitive and emotional behaviors and consist of heterogeneous cell populations. The complex spatiotemporal cell and molecular circuits underlying progressive TBI pathologies that can evolve into other disorders such as chronic traumatic encephalopathy and posttraumatic stress disorder remain to be understood. A comprehensive understanding of the molecular mechanisms underlying the complexity of TBI has been hindered by the lack of effective approaches to examine molecular events in individual brain cells that drive the overall pathology. We recently conducted a single cell resolution study of the hippocampus at the acute phase (24hr) of TBI using single cell RNA sequencing (scRNAseq) and revealed cell-type specific pathways and regulators of TBI. In particular, we found that depression of cell metabolism to be a key pathogenic component in the hippocampus at the acute phase of TBI. This finding suggests that tracking metabolic state of cells can be used to address key knowledge gaps on the spatial and time dependent progression of key pathologic drivers of TBI. Here we propose to test the hypothesis that cell metabolic regulators determine dynamic and spatial pathogenic pathways of TBI by harnessing the power of modern high-throughput technologies. We propose a highly integrative team approach to profit from recent advances in single cell RNA sequencing (scRNAseq) and multiplexed error robust fluorescent in situ hybridization (MERFISH) along with advanced gene-gene and cell-cell network modeling to inform on targets for intervention at specific time points or brain sites, a fundamental unsolved question in the TBI field. In Aim 1, we propose to utilize a unique combination of scRNAseq, MERFISH, and network modeling approaches to assess and validate the spatial and temporal vulnerability of each cell type to TBI in multiple brain regions at multiple time points in a data-driven, unbiased manner, which can inform us about hidden regulators of TBI pathogenesis. We will focus on the spatial and temporal changes in cellular metabolic pathways during TBI progression. Our preliminary results support that mt-Rnr2, encoding a mitochondrial peptide humanin and involved in cell metabolism, is a major site- and time-dependent driver of TBI. In Aim 2, we will functionally assess whether modulating mt-Rnr2 (humanin) has therapeutic potential to mitigate TBI pathology and prevent progression. We will also explore the cell-type specific mechanisms, especially the role of metabolism, underlying the actions of humanin. The overall goal of the proposal is to elaborate on an innovative strategy that can offer a comprehensive mechanistic understanding of the spatiotemporal cell substrates of TBI pathology and uncover novel targets and mechanisms to redirect the courses of TBI to overcome subsequent neurological disorders.