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

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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.