Reactive human iPSC astrocytes suppress oligodendrocyte precursor cell differentiation
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE196575 | GEO | 2022/02/14
REPOSITORIES: GEO
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Project description:Myelination depends on maintenance of oligodendrocytes that arise from oligodendrocyte precursor cells (OPCs). We show that OPC-specific proliferation, morphology, and BMAL1 are time-of-day dependent. Knock out of Bmal1 in OPCs during development disrupts expression of genes associated with circadian rhythms, proliferation, density, morphology, and migration, leading to changes in OPC dynamics in a spatio-temporal manner. Furthermore, these deficits translate into thinner myelin, dysregulated cognitive and motor function, and increased sleep fragmentation. OPC-specificBmal1loss in adulthood does not alter OPC density at baseline but impairs remyelination of a demyelinated lesion driven by changes in OPC morphology and migration. Lastly, we show sleep fragmentation is associated with increased prevalence of the demyelinating disorder multiple sclerosis (MS), suggesting a link between MS and sleep that requires further investigation. These findings have broad mechanistic and therapeutic implications for brain disorders that include both myelin and sleep phenotypes.
2023-09-07 | GSE239541 | GEO
Project description:Multiple sclerosis (MS) is characterized by demyelinated lesions in the central nervous system. Destruction of myelin and secondary damage to axons and neurons leads to significant disability, particularly in people with progressive MS. Accumulating evidence suggests that the potential for myelin repair exists in MS, although for unclear reasons this process fails. The cells responsible for producing myelin, the oligodendrocytes, and their progenitors oligodendrocyte precursor cells (OPCs) have been identified at the site of lesions, even in adults. Their presence suggests the possibility that endogenous remyelination without transplantation of donor stem cells may be a strategy for myelin repair in MS. Strategies to develop novel therapies have focused on induction of signaling pathways that stimulate OPCs to mature into myelin-producing oligodendrocytes that could then possibly remyelinate lesions. We have been investigating pharmacological approaches to enhance OPC differentiation, and have identified that the combination of two agents, triiodothyronine (T3) and quetiapine, leads to an additive effect on OPC differentiation and consequent myelin production via both overlapping and distinct signaling pathways. While the ultimate production of myelin requires cholesterol biosynthesis, we identified that quetiapine enhances gene expression in this pathway more potently than T3. Two blockers of cholesterol production, betulin and simvastatin, reduced OPC differentiation into myelin producing oligodendrocytes. Elucidating the nature of agents that lead to complementary and additive effects, possibly even synergistic effects, on oligodendrocyte differentiation and myelin production may pave the way for more efficient induction of remyelination in people with MS.
2019-07-30 | GSE125325 | GEO
Project description:Following CNS demyelination, adult oligodendrocyte progenitor cells (OPCs) can differentiate into new myelin-forming oligodendrocytes in a regenerative process called remyelination. While remyelination is very efficient in young adults, its efficiency declines progressively with ageing. Here we performed proteomic analysis on OPCs isolated acutely from the brains of neonate, young and aged female rats. Approximately 60% of the proteins are expressed at different levels in OPCs from neonates compared to their adult counterparts. The amount of myelin-associated and cell adhesion proteins are increased with age, while cholesterol-biosynthesis, transcription factors and cell cycle proteins decreased. Our experiments provide the first ageing OPC rodent proteome, revealing the distinct features of neonatal and adult OPCs, and providing new insights into why remyelination efficiency declines with ageing and potential roles for aged OPCs in other neurodegenerative diseases.
2022-02-25 | PXD010383 | Pride
Project description:Following CNS demyelination, adult oligodendrocyte progenitor cells (OPCs) can differentiate into new myelin-forming oligodendrocytes in a regenerative process called remyelination. While remyelination is very efficient in young adults, its efficiency declines progressively with ageing. Here we performed proteomic analysis on OPCs isolated acutely from the brains of neonate, young and aged female rats. Approximately 60% of the proteins are expressed at different levels in OPCs from neonates compared to their adult counterparts. The amount of myelin-associated and cell adhesion proteins are increased with age, while cholesterol-biosynthesis, transcription factors and cell cycle proteins decreased. Our experiments provide the first ageing OPC rodent proteome, revealing the distinct features of neonatal and adult OPCs, and providing new insights into why remyelination efficiency declines with ageing and potential roles for aged OPCs in other neurodegenerative diseases.
2020-05-22 | PXD013708 | Pride
Project description:Tissue progenitors maintain the integrity of organ systems through aging and stress. The brain’s white matter regions experience ischemic lesions and age-dependent degeneration. Brain white matter contains progenitors, oligodendrocyte precursor cells (OPCs), which can repair some insults. The response of OPCs to white matter ischemia and aging is not known. We characterized the response of OPCs to white matter stroke using OPC reporter mice, cell migration tracking, OPC specific RNA sequencing, and mechanistic studies in candidate biochemical pathways in the aged brain. White matter stroke induces initial proliferation of local OPCs but blocks differentiation, shunting a portion into astrocytes. Candidate signaling pathways for this differentiation block including novel interactions of inhibin and matrilin-2 and new roles of NgR1 ligands following white matter stroke. Stroke induces inhibin expression in astrocytes and downregulates OPC matrilin-2 that contributes into OPC differentiation block. Antagonism of NgR1 ligands promotes OPC differentiation by attenuating the OPC astrocytic transformation and enhances functional recovery from stroke in aged animals.
2019-11-15 | GSE53737 | GEO
Project description:A Transcriptome Database for Astrocytes, Neurons, and Oligodendrocytes: A New Resource for Understanding Brain Development and Function Understanding the cell-cell interactions that control CNS development and function has long been limited by the lack of methods to cleanly separate astrocytes, neurons, and oligodendrocytes. Here we describe the first method for the isolation and purification of developing and mature astrocytes from mouse forebrain. This method takes advantage of the expression of S100β by astrocytes. We used fluorescent activated cell sorting (FACS) to isolate EGFP positive cells from transgenic mice that express EGFP under the control of an S100β promoter. By depletion of astrocytes and oligodendrocytes we obtained purified populations of neurons, while by panning with oligodendrocyte-specific antibodies we obtained purified populations of oligodendrocytes. Using GeneChip Arrays we then created a transcriptome database of the expression levels of over 20,000 genes by gene profiling these three main CNS neural cell types at postnatal ages day 1 to 30. This database provides the first global characterization of the genes expressed by mammalian astrocytes in vivo and is the first direct comparison between the astrocyte, neuron, and oligodendrocyte transcriptomes. We demonstrate that Aldh1L1, a highly expressed astrocyte gene, is a highly specific antigenic marker for astrocytes with a substantially broader, and therefore potentially more useful, pattern of astrocyte expression than the traditional astrocyte marker GFAP. This transcriptome database of acutely isolated and highly pure populations of astrocytes, neurons and oligodendrocytes provides a resource to the neuroscience community by providing improved cell type specific markers and for better understanding of neural development, function, and disease. We acutely purified mouse astrocytes from early postnatal ages (P1) to later postnatal ages (P30), when astrocyte differentiation is morphologically complete (Bushong et al., 2004), and acutely purified mouse OL-lineage cells from stages ranging from OPCs to newly differentiated OLs to myelinating OLs. We extracted RNA from each of these highly purified, acutely isolated cell types and used GeneChip Arrays to determine the expression levels of over 20,000 genes and construct a comprehensive database of cell type specific gene expression in the mouse forebrain. Analysis of this database confirms cell type specific expression of many well characterized and functionally important genes. In addition, we have identified thousands of new cell type enriched genes, thereby providing important new information about astrocyte, OL, and neuron interactions, metabolism, development, and function. This database provides a comparison of the genome-wide transcriptional profiles of the main CNS cell types and is a resource to the neuroscience community for better understanding the development, physiology, and pathology of the CNS. Keywords: Developmental CNS Cell type comparision FACS purification of astrocytes: Dissociated forebrains from S100β-EGFP mice were resuspended in panning buffer (DBPS containing 0.02% BSA and 12.5 U/ml DNase) and sequentially incubated on the following panning plates: secondary antibody only plate to deplete microglia, O4 plate to deplete OLs, PDGFRα plate to deplete OPCs, and a second O4 plate to deplete any remaining OLs. This procedure was sufficient to deplete all OL-lineage cells from animals P8 and younger, however, in older animals that had begun to myelinate, additional depletion of OLs and myelin debris was accomplished as follows. The nonadherent cells from the last O4 dish were harvested by centrifugation, and the cells were resuspended in panning buffer containing GalC, MOG, and O1 supernatant and incubated for 15 minutes at room temperature. The cell suspension was washed and then resuspended in panning buffer containing 20 μg donkey anti-mouse APC for 15 minutes. The cells were washed and resuspended in panning buffer containing propidium iodide (PI). EGFP+ astrocytes were then purified by fluorescence activated cell sorting (FACS). Dead cells were gated out using high PI staining and forward light scatter. Astrocytes were identified based on high EGFP fluorescence and negative APC fluorescence from indirect immunostaining for OL markers GalC, MOG, and O1. Cells were sorted twice and routinely yielded >99.5% purity based on reanalysis of double sorted cells.; FACS purification of neurons: EGFP- cells were the remaining forebrain cells after microglia, OLs, and astrocytes had been removed, and were primarily composed of neurons, and to a lesser extent, endothelial cells (we estimate < 4% endothelial cells at P7 and < 20% endothelial cells at P17). EGFP- cells from S100β-EGFP dissociated forebrain were FACS purified in parallel with astrocyte purification and were sorted based on their negative EGFP fluorescence immunofluorescence. Cells were sorted twice and routinely yielded >99.9% purity. In independent preparations, the EGFP- cell population was additionally depleted of endothelial cells and pericytes by sequentially labeling with biotin-BSL1 lectin and streptavidin-APC while also labeling for OL markers as described above. Cells were sorted twice and routinely yielded >99.9% purity.; Panning purification of oligodendrocyte lineage cells: Dissociated mouse forebrains were resuspended in panning buffer. In order to deplete microglia, the single-cell suspension was sequentially panned on four BSL1 panning plates. The cell suspension was then sequentially incubated on two PDGFRα plates (to purify and deplete OPCs), one A2B5 plate (to deplete any remaining OPCs), two MOG plates (to purify and deplete myelinating OLs), and one GalC plate (to purify the remaining PDGFRα-, MOG-, OLs). The adherent cells on the first PDGFRα, MOG, and GalC plates were washed to remove all antigen-negative nonadherent cells. The cells were then lysed while still attached to the panning plate with Qiagen RLT lysis buffer, and total RNA was purified. Purified OPCs were >95% NG2 positive and 0% MOG positive. Purified Myelin OLs were 100% MOG positive, >95% MBP positive, and 0% NG2 positive. Purified GalC OLs depleted of OPCs and Myelin OLs were <10% MOG positive and ~50% weakly NG2 positive, a reflection of their recent development as early OLs.; Data normalization and analysis: Raw image files were processed using Affymetrix GCOS and the MAS 5.0 algorithm. Intensity data was normalized per chip to a target intensity TGT value of 500, and expression data and absent/present calls for individual probe sets were determined. Gene expression values were normalized and modeled across arrays using the dChip software package with invariant-set normalization and a PM model. (www.dchip.org, Li and Wong, 2001). The 29 samples were grouped into 9 sample types: Astros P7-P8, Astros P17, Astros P17-gray matter (P17g), Neurons P7, Neurons P17, Neurons-endothelial cell depleted (P7n, P17n), OPCs, GalC-OLs, and MOG-OLs. Gene filtering was performed to select probe sets that were consistently expressed in at least one cell type, where consistently expressed was defined as being called present and having a MAS 5.0 intensity level greater than 200 in at least two-thirds of the samples in the cell type. We identified 20,932 of the 45,037 probe sets that were consistently expressed in at least one of the nine cell types. The Significance Analysis of Microarrays (SAM) method (Tusher et al., 2001) was used to determine genes that were significantly differentially expressed between different cell types (see Supplemental Table S2 for SAM cell type groupings). Clustering was performed using the hclust method with complete linkage in R. Expression values were transformed for clustering by computing a mean expression value for the gene using those samples in the corresponding SAM statistical analysis, and then subtracting the mean from expression intensities. In order to preserve the log2 scale of the data, unless otherwise indicated, no normalization by variance was performed. Plots were created using the gplots package in R. The Bioconductor software package (Gentleman et al., 2004) was used throughout the expression analyses. Functional analyses were performed through the use of Ingenuity Pathways Analysis (Ingenuity® Systems, www.ingenuity.com).
2007-12-30 | E-GEOD-9566 | biostudies-arrayexpress
Project description:Remyelination in multiple sclerosis (MS) is from the start of the disease imperfect, and strongly decreases with its progression, mainly due to the harm on oligodendrocyte progenitor cells (OPCs), causing neurodegeneration and resulting in irreversible neurological deficits. Therapeutic strategies promoting remyelination are still very preliminary and lack within the current treatment panel of MS. In a previous study we have identified 21 microRNAs dysregulated mostly in the CSF of relapsing and/or remitting MS patients. Here, we observed that among these, the majority of 13 transfected microRNA mimics decreased the differentiation of an OPC cell line called CG-4. Herein we support by RNA sequencing and independent RT-qPCR analyses that miR-33-3p, miR-34c-5p, miR-124-5p, and miR-145-5p impede OPC differentiation as evidenced by the downregulation of premyelinating oligodendrocyte (OL) (Tcf7l2, Cnp [except for miR-145-5p]) and mature OL (Plp1, Mbp, Mobp) markers, whereas only miR-214-3p promotes OPC differentiation. We further propose a comprehensive exploration of their change in cell fate through Gene Ontology enrichment analysis. We finally confirm by RT-qPCR analysis the downregulation of several predicted mRNA targets for each microRNA that possibly support their effect on OPC differentiation by very distinctive mechanisms, of which some are still unexplored in OPC/OL physiology. We hereby open new perspectives in the research on OPC differentiation and the pathophysiology of demyelination/remyelination, and possibly even in the research on new remyelinating therapeutic strategies in the scope of MS.
2024-02-07 | GSE244479 | GEO
Project description:Remyelination after white matter injury (WMI) often fails in diseases such as multiple sclerosis due to improper recruitment and repopulation of oligodendrocyte precursor cells (OPCs) in lesions. How OPCs elicit specific intracellular programs in response to a chemically and mechanically diverse environment to properly regenerate myelin remains unclear. OPCs construct primary cilia, specialized signaling compartments that transduce Hh and GPCR signals. We investigated the role of primary cilia in the OPC response to WMI. Removing cilia from OPCs genetically via deletion of Ift88 results in OPCs failing to repopulate WMI lesions due to reduced proliferation. Interestingly, loss of cilia does not affect Hh signaling in OPCs or their responsiveness to Hh signals, but instead leads to dysfunctional cAMP-dependent CREB-mediated transcription. As inhibition of CREB activity in OPCs reduces proliferation, we propose that a GPCR/cAMP/CREB signaling axis initiated at OPC cilia orchestrates OPC proliferation during development and in response to WMI.
2023-09-29 | GSE243997 | GEO
Project description:Astrocyte activation is associated with progressive inflammatory demyelination in multiple sclerosis (MS). The molecular mechanisms underlying astrocyte activation remain incompletely understood. Recent studies have suggested that classical neurotransmitter receptors are implicated in the modulation of brain innate immunity. We investigated the role of dopamine signaling in the process of astrocyte activation. Here, we show the upregulation of dopamine D2 receptor (DRD2) in reactive astrocytes in MS brain and non-canonical role of astrocytic DRD2 in MS pathogenesis. Mice deficient in astrocytic Drd2 exhibit a remarkable suppression of reactive astrocytes and inflammation which are highly correlated with the amelioration of experimental autoimmune encephalomyelitis (EAE). Mechanistically, DRD2 regulates the expression of 6-pyruvoyl-tetrahydropterin synthase which modulates NF-κB activity through protein kinase C-δ. Pharmacological blockade of astrocytic DRD2 with a DRD2 antagonist dehydrocorybulbine remarkably inhibits the inflammatory response in mice lacking Drd2 in neurons. Together, our findings reveal previously uncharted roles for a DRD2 in astrocyte activation during EAE-associated CNS inflammation. Its therapeutic inhibition may provide a potent lever to alleviate autoimmune diseases.
2022-07-15 | GSE205291 | GEO
Project description:Astrocyte activation is associated with progressive inflammatory demyelination in multiple sclerosis (MS). The molecular mechanisms underlying astrocyte activation remain incompletely understood. Recent studies have suggested that classical neurotransmitter receptors are implicated in the modulation of brain innate immunity. We investigated the role of dopamine signaling in the process of astrocyte activation. Here, we show the upregulation of dopamine D2 receptor (DRD2) in reactive astrocytes in MS brain and non-canonical role of astrocytic DRD2 in MS pathogenesis. Mice deficient in astrocytic Drd2 exhibit a remarkable suppression of reactive astrocytes and inflammation which are highly correlated with the amelioration of experimental autoimmune encephalomyelitis (EAE). Mechanistically, DRD2 regulates the expression of 6-pyruvoyl-tetrahydropterin synthase which modulates NF-κB activity through protein kinase C-δ. Pharmacological blockade of astrocytic DRD2 with a DRD2 antagonist dehydrocorybulbine remarkably inhibits the inflammatory response in mice lacking Drd2 in neurons. Together, our findings reveal previously uncharted roles for a DRD2 in astrocyte activation during EAE-associated CNS inflammation. Its therapeutic inhibition may provide a potent lever to alleviate autoimmune diseases.
2022-07-15 | GSE205290 | GEO