Effects of lipid-specific antibodies (sulphatide, O4) on dissociated myelinating cultures
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PROVIDER: PRJNA631776 | ENA |
REPOSITORIES: ENA
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Project description:Analysis of the effects of sulphatide specific antibody (O4, 20microg/ml) on myelinating cultures generated from dissociated embryonic rat spinal cord after 24 hours treatment to give insights into effects of lipid-specific antibodies and its implication in demyelinating diseases.
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Project description:Adult oligodendrocyte progenitors (aOPCs) share with their neonatal counterpart (nOPC) the ability to give rise to myelinating oligodendrocytes, but they also display unique functional features. This study addresses the molecular mechanisms underlying the intrinsic differences between these two populations. Using RNA-sequencing and unbiased histone proteomics analysis performed on PDGFRa+ OPCs sorted from a reporter mouse, we define the unique transcriptome and histone marks of aOPCs. We describe the higher levels of genes related to lipid metabolism and myelin sheath, and lower expression of genes related to cycle and proliferation in aOPCs compared to nOPCs. We also identify greater levels of activating H4K8ac and repressive H4K20me3 histone marks in aOPCs compared to nOPCs, and increased occupancy of H4K8ac mark at chromatin locations corresponding to oligodendrocyte-specific genes. Pharmacological inhibition of histone acetylation in neonatal O4+OPCs (nO4+OPCs) and adult O4+OPCs (aO4+OPCs) followed by RNA-sequencing showed a moderate change in the transcriptome of these cells resulting in the decrease of proliferation capacity in aO4+OPCs but not nO4+OPCs. Pharmacological inhibition of H4K20me3 mark was not sufficient to change the transcriptome or functional properties of nO4+OPCs and aO4+OPC. Overall, this study identifies histone acetylation as key in regulating the proliferation of aO4+OPCs.
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Project description:We report a method for deriving oligodendrocyte lineage cells from human pluripotent stem cells (hPSCs) in three-dimensional (3D) culture called human oligodendrocyte spheroids (hOLS). To characterize oligodendrocyte-lineage cells in hOLS, we isolated O4+ cells by immunopanning and performed deep single cell RNA sequencing. We sequenced 295 cells and compared their profiles to unsorted cells isolated from primary human fetal cortex, primary human adult cortex, and hCS. Clustering of all cells using the t-distributed stochastic neighbor embedding (tSNE) approach revealed a distinct populations of SOX10+ oligodendrocytes, within which the O4+ cells derived from hOLS clustered most closely to oligodendrocyte progenitor cells (OPCs) and mature oligodendrocytes from the primary human adult cortical tissue. Additionally, subpopulations of OPCs, newly formed oligodendrocytes, and myelinating oligodendrocytes derived were observed in the hOLS-derived cluster. To further assess the state of oligodendrocyte-lineage cells in hOLS, we performed a Monocle analysis which revealed a spectrum of oligodendrocyte-lineage stages in hOLS ranging from dividing cells that closely resembled primary OPCs to mature cells that closely resembled primary oligodendrocytes.
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Project description:Human fetal tissue dissociates between 18-22wk gestational age were FACS sorted on the basis of CD140a (PDGFRA) and O4 antigens. Each population was profiled immediately following FACS. 20 samples, 4 groups (FACS-sorted cell populations), between 3 and 6 replicates in each group, each replicate is from a separate patient sample
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