Spatiotemporal Molecular Substrates of TBI at Single Cell Resolution

摘要 创伤性脑损伤 (TBI) 具有复杂的神经病理学，涉及大脑中枢的进行性改变 处理认知和情感行为并由异质细胞群组成。综合体 进行性 TBI 病理学背后的时空细胞和分子回路，可以演变成其他疾病 慢性创伤性脑病和创伤后应激障碍等疾病仍有待解决 明白了。全面了解 TBI 复杂性背后的分子机制 由于缺乏有效的方法来检查单个脑细胞中的分子事件而受到阻碍 驱动整体病理学。我们最近对海马体进行了单细胞分辨率研究 使用单细胞 RNA 测序 (scRNAseq) 分析 TBI 急性期（24 小时）并揭示细胞类型特异性 TBI 的途径和调节因素。特别是，我们发现细胞代谢的抑制是一个关键 TBI 急性期海马中的致病成分。这一发现表明跟踪 细胞的代谢状态可用于解决空间和时间依赖性的关键知识差距 TBI 关键病理驱动因素的进展。在这里，我们建议检验细胞代谢的假设 监管机构利用现代技术的力量来确定 TBI 的动态和空间致病途径 高通量技术。我们提出了一种高度整合的团队方法，以从最近的进展中获利 单细胞 RNA 测序 (scRNAseq) 和多重误差稳健荧光原位杂交 (MERFISH) 以及先进的基因-基因和细胞网络模型，以告知干预目标 在特定时间点或大脑部位，这是 TBI 领域尚未解决的基本问题。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 多次验证多个大脑区域中每种细胞类型对 TBI 的空间和时间脆弱性 以数据驱动、公正的方式分析点，这可以让我们了解 TBI 发病机制的隐藏调节因素。 我们将重点关注 TBI 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，编码线粒体肽人素并参与细胞 新陈代谢是 TBI 的主要位点和时间依赖性驱动因素。在目标 2 中，我们将从功能上评估是否 调节 mt-Rnr2（护脑素）具有减轻 TBI 病理和预防进展的治疗潜力。 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， 护脑素的作用。该提案的总体目标是详细阐述一项创新战略，该战略可以提供 对 TBI 病理学时空细胞基质的全面机制理解并揭示 重新定向 TBI 进程以克服随后的神经系统疾病的新目标和机制。

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.