Characterization of meningeal type 2 innate lymphocytes and their response to CNS injury
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ABSTRACT: The meningeal space is occupied by a diverse repertoire of innate and adaptive immune cells. CNS injury elicits a rapid immune response that affects neuronal survival and recovery, but the role of meningeal inflammation in CNS injury remains poorly understood. Here we describe group 2 innate lymphoid cells (ILC2s) as a novel cell type resident in the healthy meninges that is activated following CNS injury. ILC2s are present throughout the naïve mouse meninges, though are concentrated around the dural sinuses, and have a unique transcriptional profile relative to lung ILC2s. After spinal cord injury, meningeal ILC2s are activated in an IL-33 dependent manner, producing type 2 cytokines. Using RNAseq, we characterized the gene programs that underlie the ILC2 activation state. Finally, addition of wild type lung-derived ILC2s into the meningeal space of IL-33R-/- animals improves recovery following spinal cord injury. These data characterize ILC2s as a novel meningeal cell type that responds to and functionally affects outcome after spinal cord injury, and could lead to new therapeutic insights for CNS injury or other neuroinflammatory conditions.
Project description:Tertiary lymphoid structures (TLS) are organized aggregates of B and T cells formed ectopically during different life periods in response to inflammation, infection, or cancer. Here, we describe formation of structures reminiscence of TLS in the spinal cord meninges under several central nervous system (CNS) pathologies. Following acute spinal cord injury, B and T lymphocytes locally aggregate within the meninges to form TLS, which continue to accumulate during the late phase of repair, with a negative impact on subsequent pathological conditions, such as experimental autoimmune encephalomyelitis. Using a chronic model of spinal cord pathology, the mSOD1 mouse model of amyotrophic lateral sclerosis, we further showed by single cell RNA-sequencing that a meningeal lymphocyte niche forms, with a unique organization and activation state, including accumulation of pre-B cells in the spinal cord meninges. Such a response was not found in the CNS-draining cervical lymph nodes. The present findings suggest that a unique immune response develops in the meninges during various neurological pathologies in the CNS, a reflection of its immune privileged nature.
Project description:The meninges are a tripartite system of membranous tissue comprised of pial, arachnoid and dural layers that cover both the brain and spinal cord. Between the arachnoid and pial layers is the subarachnoid space filled with cerebrospinal fluid (CSF). Though the meninges have long been thought to provide largely a protective function, a growing number of studies highlight this tissue to be a nest of immune activity, harboring T cells, B cells, macrophages, and a variety of other myeloid cell types during health and disease. But despite the burgeoning prospect that the meninges might play a decidedly more active role in immune and other regulatory processes than previously thought, little is known about their structural makeup. Contributing to this void, considerable technical challenges prevent sophisticated analysis of the meninges at cellular and molecular levels. In addition, removal of the brain and spinal cord from their bony encasement leads to tearing of the tenuous meninges and significant disruption of the delicate inter-membrane arrangements. Also lacking are sufficient molecular targets to identify the various meningeal structural elements, only hinted at so far by scanning electron microscopy. Accordingly, we developed a method to allow removal of brain and spinal meninges while minimizing risk of parenchymal contamination and performed shotgun proteomics on the two meningeal domains. While the vast majority of proteins at both locales overlapped, several proteins – including those of structural nature – were exclusive to brain or spinal meninges. Targeting proteins revealed in the proteomic data, the cellular and extracellular elements that provide the meninges’ structure were identified using confocal and electron microscopy, and were observed for the first time in high-resolution
Project description:2nd generation sequencing was used to compare expression profiles of MBP-specific T cells retrieved from blood, CSF, spinal cord meninges and parenchyma. The overall expression profiles were found to be very similar.However, genes regulated during T cell activation were found to be upregulated in T cells from spinal cord meninges and parenchyma compared to blood and CSF. 2nd generation sequencing of MBP-specific T cells retrieved from blood and CNS compartments during experimental autoimmune encephalomyelitis
Project description:We used single-cell RNA sequencing to profile immune cells in the injured spinal cord parenchyma and lymphatic endothelial cells in the spinal cord meninges from young and aged mice. This help us understand the heterogeneity of the immune response after injury and how it is altered in aging. Moreover, the data obtained from the spinal cord meninges provides novel molecular insights into how the meninges may contribute to the repair process.
Project description:We used single-cell RNA sequencing to profile immune cells in the injured spinal cord parenchyma and lymphatic endothelial cells in the spinal cord meninges from young and aged mice. This help us understand the heterogeneity of the immune response after injury and how it is altered in aging. Moreover, the data obtained from the spinal cord meninges provides novel molecular insights into how the meninges may contribute to the repair process.
Project description:We used single-cell RNA sequencing to profile immune cells in the injured spinal cord parenchyma and lymphatic endothelial cells in the spinal cord meninges from young and aged mice. This help us understand the heterogeneity of the immune response after injury and how it is altered in aging. Moreover, the data obtained from the spinal cord meninges provides novel molecular insights into how the meninges may contribute to the repair process.
Project description:In homeostasis, because of the blood-brain barrier, immune cells rarely infiltrate the central nervous system (CNS). However, after spinal cord injury (SCI), many cells, including both myeloid and T cells, infiltrate the spinal cord. However, the role immune cells play in SCI remains controversial. We are curious whether after SCI there are self-peptides that are released and sensed by T cells that then modulate response to CNS injury.
Project description:Spinal cord injury (SCI) leads to fibrotic scar formation at the lesion site, which finally affects axon regeneration and motor functional recovery. Myofibroblasts have been regarded as the main cell types that filled in the fibrotic scar, however, the cell source of myofibroblasts in transection and crush SCI model remain to be elusive. Here we used lineage tracing or single cell transcription sequencing to investigate the cell origin of fibrotic scar. We found fibrotic scars were filled from PDGFRβ+ daughter cells in spinal cord in crush SCI or transection SCI. The parenchyma perivascular-derived and meninges-derived PDGFRβ+ fibroblasts, but not PDGFRβ+ pericytes, proliferated and contributed to fibrotic cells in the lesion core. The percentage of meninges-derived fibroblasts specifically was higher than parenchyma perivascular-derived fibroblasts in transection model, which might contribute to the more fibrotic scar in transection model than crush model. These findings may provide theoretical support for the treatment of spinal cord injury.
Project description:Mice lacking the developmental axon guidance molecule EphA4 have previously been shown to exhibit extensive axonal regeneration and functional recovery following spinal cord injury. To assess mechanisms by which EphA4 may modify the response to neural injury, a microarray was performed on spinal cord tissue from mice with spinal cord injury and sham injured controls. RNA was purified from spinal cords of adult EphA4 knockout and wild-type mice four days following lumbar spinal cord hemisection or laminectomy only and was hybridised to Affymetrix All-Exon Array 1.0 GeneChips. While subsequent analyses indicated that several pathways were altered in EphA4 knockout mice, of particular interest was the attenuated or otherwise altered expression of a number of inflammatory genes, including Arginase 1, expression of which was lower in injured EphA4 knockout compared to wild-type mice. Immunohistological analyses of different cellular components of the immune response were then performed in injured EphA4 knockout and wild-type spinal cords. While numbers of infiltrating CD3+ T cells were low in the hemisection model, a robust CD11b+ macrophage / microglial response was observed post-injury. There was no difference in the overall number or spread of macrophages / activated microglia in injured EphA4 knockout compared to wild-type spinal cords at two, four or fourteen days post-injury, however a lower proportion of Arginase-1 immunoreactive macrophages / activated microglia was observed in EphA4 knockout spinal cords at four days post-injury. Subtle alterations in the neuroinflammatory response in injured EphA4 knockout spinal cords may contribute to the regeneration and recovery observed in these mice following injury. Comparison was made between gene expression in wild-type and knockout samples both before and after injury. 3 replicates per group.
Project description:Patients carrying one or two ApoE4 alleles suffer from worse functional recovery after spinal cord injury. Using transgenic mice expression human ApoE3 or ApoE4 we investigated potential cellular mechanisms of reduced recovery after spinal cord injury. Bulk RNA sequencing of the spinal cord lesion site followed by pathway enrichment analysis predicts that ApoE4 mice have a higher inflammatory and extracellular matrix remodeling activity 7 days after spinal cord injury. Contrary, higher activities for neuronal projection and action potential patways were predicted in the ApoE3 mice at 21 days after injury.