Project description:The important contribution of glia to mechanisms of injury and repair of the nervous system is increasingly recognized. In stark contrast to the central nervous system (CNS), the peripheral nervous system (PNS) has a remarkable capacity for regeneration after injury. Schwann cells are recognized as key contributors to PNS regeneration but the molecular underpinnings of the Schwann cell response to injury remain incompletely understood. To gain insight into the acute SC injury response, we provide an RNAseq database of Schwann cells purified acutely from the naïve and injured rodent sciatic nerve at 3, 5, and 7 days post injury. Bioinformatic analysis provides validation of cell purity and dataset integrity as well as identification of discrete modules of genes that follow distinct patterns of regulation in the first days after injury and their corresponding molecular pathways. Our dataset provides a helpful resource for further deciphering the SC injury response and provides a depth of transcriptional data that can complement the findings of recent single cell sequencing approaches. In addition, as more data becomes available on the response of CNS glia to injury, we anticipate that this dataset will provide a valuable platform for understanding key differences in the PNS and CNS glial responses to injury and for designing approaches to ameliorate CNS regeneration.
Project description: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:Picornaviruses are a leading cause of central nervous system (CNS) infections. While genotypes such as Parechovirus A3 (PeV-A3) and echovirus 11 (E11) can elicit severe neurological disease, the highly prevalent PeV-A1 is not associated with CNS disease. Here, we expand our current understanding of these differences in PeV-A CNS disease using human brain organoids and clinical isolates of PeV-A genotypes. Our data indicates that PeV-A1 and A3 specific differences in neurological disease are not due to infectivity of CNS cells as both viruses productively infect brain organoids with a similar cell tropism. Proteomic analysis showed that PeV-A infection significantly alters the host cell metabolism. The inflammatory response following PeV-A3 (and E11 infection) was significantly more potent than that upon PeV-A1 infection. Collectively, our findings align with clinical observations and suggest a role for inflammatory-mediated neurology, rather than viral replication, in PeV-A3 (and E11) infection.
Project description:Diabetes is prevalent worldwide and associated with severe health complications, including blood vessel damage that leads to cardiovascular disease and death. We report the development of 3D blood vessel organoids from human embryonic and induced pluripotent stem cells. These human blood vessel organoids contain endothelium, perivascular pericytes, and basal membranes, and self-assemble into lumenized interconnected capillary networks. We treat these vascular organoids with hyperglycemia and inflammatory cytokines in vitro, which leads to basement membrane thickening, a structural hallmark of diabetic patient. To compare differential gene expression we performed RNAseq on endothelial cells, derived from control (NG) or diabetic (DI) vascular organoids.
Project description:The extracellular matrix (ECM), a key interface between the cerebrovasculature and other cells within the neuro-glial-vascular unit , provides structural stability and modulates cell behaviour and signalling. These functions are dependent on ECM protein composition. Since ECM defects may contribute to a broad range of disorders including cerebrovascular disease, it is crucially important to characterize ECM composition to better understand the mechanisms by which the ECM modulates brain health and disease. To date, molecular studies of the cerebrovascular ECM have been limited. We therefore generated ECM extracts from vascular enriched tissues of both mouse and human post-mortem brain. We used mass spectrometry with off-line high-pH reversed-phase fractionation to increase proteome depth and characterized the identified proteins. This identified a large number of proteins (>1000 mouse, >2000 human) in the ECM-enriched fractions, with > 66% of the identified proteins covered in both human and mouse samples. We now report 147 ECM proteins, including collagens (as major ECM components), laminins, fibronectin and nidogens, that can be considered as the core constituents of the human brain vascular matrisome. We also identified 12 novel proteins in the ECM-enriched fraction, that may have regulate or interact with the matrisome, including several key regulators of neurovascular interactions, such as BCAM, CDH5 and PARVB. Many of the identified brain vascular matrisome proteins are encoded by genes identified in stroke and cerebral small vessel disease genome-wide association studies, underscoring the importance of the vascular ECM in health and disease. This brain vascular matrisome represents a powerful resource for investigating the impact of aging and disease on the cerebrovasculature.
Project description:Age-related macular degeneration (AMD) is a common, blinding disease of the elderly in which macular photoreceptor cells, retinal pigment epithelium, and choriocapillaris endothelial cells ultimately degenerate. Recent studies have found that degeneration of the choriocapillaris occurs early in this disease and that this endothelial cell dropout is concomitant with increased deposition of the complement membrane attack complex (MAC) at the choroidal endothelium. However, the impact of MAC injury to choroidal endothelial cells is poorly understood. To model this event in vitro, and to study the downstream consequences of MAC injury, endothelial cells were exposed to complement from human serum, compared to heat inactivated serum which lacks complement components. Cells exposed to complement components in human serum showed increased labeling with antibodies directed against the MAC, time and dose dependent cell death as assessed by lactate dehydrogenase assay, and increased permeability. RNA-Seq analysis following complement injury revealed increased expression of genes associated with angiogenesis including matrix metalloproteases (MMPs) 3 and 9, and VEGF-A. The MAC-induced increase in MMP9 RNA expression was validated using C5 depleted serum compared to C5 reconstitited serum. Increased levels of MMP9 were also determined using Western blot and zymography. These data suggest that, in addition to cell lysis, complement attack on choroidal endothelial cells promotes an angiogenic phenotype in surviving cells. RNA-Seq of RF/6A (cultured choroidal endothelial cells from Rhesus macaque) treated with either 50% heat-inactivated human serum ([CONTROL], n=3) or 50% normal human serum (active complement membrane attack complex [MAC], n=3)
Project description:Blood-brain barrier (BBB) critically regulate the homeostasis of central nervous system (CNS). This barrier property allows cerebral vessels to meet the extremely high metabolic demand of neural activities and meanwhile protect sensitive neurons from toxic plasma components, blood immune cells and xenobiotics. Therefore, a comprehensive inventory of the molecular determinants of BBB would substantially facilitate understanding of the pathogenesis of neurological disorders involving BBB dysfunction and promote development of novel CNS drug delivery strategies. Here, we established the proteome activity landscapes of adult mouse brain, lung and liver ECs. In this study, we produced a comprehensive molecular atlas of adult mouse BBB and revealed novel insights into adult BBB in health and Alzheimer’s disease.
Project description:Cerebrovascular injuries can cause severe edema and inflammation that adversely affect human health. Here, we observed recanalization after successful endovascular thrombectomy for acute large vessel occlusion was associated with cerebral edema and poor clinical outcomes in patients who experienced hemorrhagic transformation. To understand this process, we developed a cerebrovascular injury model using transcranial ultrasound that enabled spatiotemporal evaluation of resident and peripheral myeloid cells. We discovered that injurious and reparative responses diverged based on time and cellular origin. Resident microglia initially stabilized damaged vessels in a purinergic receptor-dependent manner, which was followed by influx of myelomonocytic cells that caused severe edema. Prolonged blockade of myeloid cell recruitment with anti-adhesion molecule therapy prevented severe edema but also promoted neuronal destruction and fibrosis by interfering with vascular repair later orchestrated by pro-inflammatory monocytes and pro-angiogenic repair-associated microglia (RAM). These data demonstrate how temporally distinct myeloid cell responses can contain, exacerbate, and ultimately repair a cerebrovascular injury.
Project description:A broad range of brain pathologies critically relies on the vasculature, and cerebrovascular disease is a leading cause of death worldwide. However, the cellular and molecular architecture of the human brain vasculature remains incompletely understood. Here, we performed single-cell RNA sequencing of 606,380 freshly isolated endothelial, perivascular and other tissue-derived cells from 117 samples, from 68 human fetuses and adult patients to construct a molecular atlas of the developing fetal, adult control and diseased human brain vasculature. We uncover extensive molecular heterogeneity of the vasculature of healthy fetal and adult human brains and across eight vascular-dependent Central Nervous System (CNS) pathologies including brain tumors and brain vascular malformations. We identify alteration of arteriovenous differentiation and reactivated fetal as well as conserved dysregulated genes and pathways in the diseased vasculature. Pathological endothelial cells display a loss of CNS-specific properties and reveal an upregulation of MHC class II molecules, indicating atypical features of CNS endothelial cells. Cell-cell interaction analyses predict numerous endothelial-to-perivascular cell ligand-receptor crosstalk including immune-related and angiogenic pathways, thereby unraveling a central role for the endothelium within brain neurovascular unit signaling networks. Our single-cell brain atlas provides insight into the molecular architecture and heterogeneity of the developing, adult/control and diseased human brain vasculature and serves as a powerful reference for future studies
Project description:A broad range of brain pathologies critically relies on the vasculature, and cerebrovascular disease is a leading cause of death worldwide. However, the cellular and molecular architecture of the human brain vasculature remains incompletely understood. Here, we performed single-cell RNA sequencing of 606,380 freshly isolated endothelial, perivascular and other tissue-derived cells from 117 samples, from 68 human fetuses and adult patients to construct a molecular atlas of the developing fetal, adult control and diseased human brain vasculature. We uncover extensive molecular heterogeneity of the vasculature of healthy fetal and adult human brains and across eight vascular-dependent Central Nervous System (CNS) pathologies including brain tumors and brain vascular malformations. We identify alteration of arteriovenous differentiation and reactivated fetal as well as conserved dysregulated genes and pathways in the diseased vasculature. Pathological endothelial cells display a loss of CNS-specific properties and reveal an upregulation of MHC class II molecules, indicating atypical features of CNS endothelial cells. Cell-cell interaction analyses predict numerous endothelial-to-perivascular cell ligand-receptor crosstalk including immune-related and angiogenic pathways, thereby unraveling a central role for the endothelium within brain neurovascular unit signaling networks. Our single-cell brain atlas provides insight into the molecular architecture and heterogeneity of the developing, adult/control and diseased human brain vasculature and serves as a powerful reference for future studies