Interrogating the Potential of Ccn1+ Astrocyte Niches to Drive Angiogenesis after Spinal Cord Injury

摘要 脊髓损伤(SCI)是一种毁灭性的神经疾病，其特征是上升和 下降的轴突网络，这可能会让人终身瘫痪。脊髓损伤后，白质中的轴突幸存 (Wm)区域试图经历短距离萌发，以恢复正常的神经功能。然而， 支配这种短程轴突萌发的神经元外在因素仍然知之甚少。在.期间 发育和损伤后，新的轴突在时空上跟随新的血管，因此紧密地联系在一起 轴突萌发到维管系统。中枢神经系统(CNS)的血管形成是由星形胶质细胞协调的 与血管内皮细胞和周细胞发生物理作用，形成神经胶质血管单位--这是一种不可或缺的组织利基 在调节血流和维持血脑屏障(BBB)方面的作用。星形胶质细胞也是主要的响应者 对任何中枢神经系统的侮辱，并在形态、基因表达和功能上经历高度上下文相关的变化 这一过程统称为“星形胶质细胞反应”。最近关于中风和缺氧的研究发现 反应性星形胶质细胞在恢复性血管生成中的必要作用，但星形胶质细胞分泌的特定分子 调解这些影响仍然是一个悬而未决的问题。我最近发现了一部受空间限制的小说 反应性星形胶质细胞亚群由强大的促血管生成分子持续上调所定义 CCN家庭成员1(CCN1)。在我的提案中，我将利用两个独立但互补的目标来测试 假设CCN1+星形胶质细胞划分了一个进化中的促血管生成组织生态位，这促进了功能 通过直接调控内皮细胞表型包括细胞增殖、成熟或 凹槽信号。在目标1中，我将分析SPARED的第一个纵向空间转录数据集 小鼠脊髓半横断模型(MhSCI)到A)的组织区域询问分子进化 脊髓内含有CCN1+星形胶质细胞的组织生态位，以及B)为脊髓损伤领域建立了一个强大的资源。从… 为此，我将了解CCN1+星形胶质细胞生态位的独特分子特征，并通过计算推断 相关生物过程、信号级联和转录调控因子的进化。在《目标2》中我会 利用新产生的星形胶质细胞特异性CCN1基因敲除补充来自Aim 1的计算数据 小鼠A)运动恢复的行为学检测和B)内皮细胞的组织学评估 MhSCI后的表型。从这个目的出发，我将询问CCN1对脊髓损伤的治疗潜力和直接的 它对血管内皮细胞的规格有影响。综上所述，这项研究将揭示血管生成的潜力 含有CCN1+星形胶质细胞的组织利基，开始提供一个潜在的作用机制的一瞥。 这些发现可能对开发新的疗法具有重要意义，这些疗法旨在 为脊髓损伤后再生轴突萌发提供更宽松的环境。

ABSTRACT Spinal Cord Injury (SCI) is a devastating neurological disorder, characterized by disruption to ascending and descending axonal networks, that can leave people paralyzed for life. After SCI, axons in spared white matter (WM) regions attempt to undergo short-range sprouting to restore normal neurological function. However, neuron-extrinsic factors that govern this short-range axon sprouting remain poorly understood. During development and after injury, new axons spatiotemporally follow new blood vessels, hence intimately linking axon sprouting to vasculature. Central Nervous System (CNS) vasculature is orchestrated by astrocytes, which physically interact with endothelia and pericytes to form the gliovascular unit- a tissue niche with indispensable roles in modulating blood flow and Blood Brain Barrier (BBB) maintenance. Astrocytes are also chief responders to any CNS insult and undergo highly context dependent changes in morphology, gene expression, and function in a process collectively referred to as “astrocyte reactivity”. Recent work in stroke and hypoxia have uncovered necessary roles for reactive astrocytes in restorative angiogenesis, but specific astrocyte-secreted molecules mediating these effects remain an outstanding question. I have recently identified a novel, spatially restricted subpopulation of reactive astrocytes defined by persistent upregulation of the powerful pro-angiogenic molecule CCN Family Member 1(CCN1). In my proposal I will utilize two independent, yet complimentary, aims to test the hypothesis that Ccn1+ astrocytes demarcate an evolving pro-angiogenic tissue niche, that promotes functional recovery after SCI by directly governing endothelial cell phenotype including cell proliferation, maturation, or Notch signaling. In Aim 1 I will analyze a first of its kind longitudinal Spatial Transcriptomics dataset of spared tissue regions in a mouse hemisection model of SCI (mhSCI) to A) interrogate the molecular evolution of intraspinal tissue niches harboring Ccn1+ astrocytes, and B) establish a powerful resource for the SCI field. From this aim I will understand the unique molecular features of Ccn1+ astrocyte niches and computationally infer the evolution of associated biological processes, signaling cascades, and transcriptional regulators. In Aim 2 I will complement the computational data from Aim 1 by utilizing a newly generated astrocyte specific CCN1 knockout mouse for A) behavioral assays of locomotor recovery and B) histological assessment of endothelial cell phenotype after mhSCI. From this aim I will interrogate the therapeutic potential of CCN1 for SCI and the direct effect it has on endothelial cell specification. Taken together, this study will uncover the angiogenic potential of tissue niches harboring Ccn1+ astrocytes and start to provide a glimpse into a potential mechanism of action. Such findings may have important implications in the development of new therapeutics that are aimed at providing a more permissive environment for regenerative axon sprouting after SCI.