Oligodendroglial Cell Line Research Articles

SOX17 transcription factor negatively regulates oligodendrocyte precursor cell differentiation.

Oligodendrocyte development is a critical process timely and spatially regulated to ensure proper myelination of the central nervous system. HMG-box transcription factors are key regulators of oligodendrocyte lineage progression. Among these factors, Sox17 was previously identified as a positive regulator of oligodendrocyte development. However, the role of Sox17 in oligodendroglial cell lineage progression and differentiation is still poorly understood. To define the functional role of Sox17, we generated new transgenic mouse models with inducible overexpression of Sox17, specifically in oligodendroglial cells. Here, we report that gain of Sox17 function has no effect on oligodendrocyte progenitor cells (OPCs) specification. During early postnatal development, Sox17 overexpression increases the pool of OPCs at the expense of differentiated oligodendrocytes. However, the oligodendroglial cell population, OPC proliferation and apoptosis remained unchanged in Sox17 transgenic mice. RNA sequencing, quantitative RT-PCR and immunohistochemical analysis showed that Sox17 represses the expression of the major myelin genes, resulting in a severe CNS hypomyelination. Overall, our data highlight an unexpected role for Sox17 as a negative regulator of OPC differentiation and myelination, suggesting stage specific functions for this factor during oligodendroglial cell lineage progression.

ProBDNF inhibits the proliferation and migration of OLN‑93 oligodendrocytes.

In contrast with mature brain-derived neurotrophic factor (mBDNF), proBDNF induces cell apoptosis. However, the function of proBDNF in oligodendrocytes remains unclear. In the present study, the OLN-93 oligodendroglia cell line was utilized as an in vitro model to analyse the functions of proBDNF in oligodendroglia. p75NTR, sortilin and proBDNF were expressed in cultured OLN-93 cells. It was indicated that proBDNF inhibited OLN-93 cell proliferation in a dose-dependent manner as determined using the MTT assay and BrdU staining. Furthermore, proBDNF suppressed the migration of OLN-93 cells as demonstrated using the scratch assay. proBDNF also decreased cell viability and promoted apoptosis as indicated by activated cysteine-aspartic acid protease-3 (caspase-3) immunocytochemistry. Notably, anti-proBDNF treatment neutralized the effect of proBDNF and resulted in increased cell proliferation and migration and decreased apoptosis. However, these effects were not observed in the presence of recombinant p75NTR extracellular domain-human FC fusion protein and p75NTR antibody, indicating that proBDNF imparts its inhibitory effects on oligodendrocytes through the p75NTR signal pathway.

Protective effect of 4-Phenylbutyrate against proteolipid protein mutation-induced endoplasmic reticulum stress and oligodendroglial cell death

Proteolipid protein (PLP) mutation causes oligodendrocyte degeneration and myelin disorders including Pelizaeus-Merzbacher Disease (PMD). As the pathophysiological mechanisms involved in PMD are poorly known, the development of therapies remains difficult. To elucidate the pathogenic pathways, an immortalized oligodendroglial cell line (158JP) expressing PLP mutation has been generated. Previous investigations revealed that 158JP oligodendrocytes exhibit several abnormalities including aberrant PLP insertion into the plasma membrane, cAMP, plasmalogen and cell cycle deficits. However, further clarifications of abnormal PLP-induced oligodendrocyte degeneration are required in order to identify relevant mechanisms to target for efficient protection against oligodendrocyte death. Because PLP overexpression may lead to its accumulation inside the endoplasmic reticulum (ER) and cause ER-stress, we explored whether ER-stress may pivotally determine 158JP cell survival/death. Viability assays, RT-qPCR, western blot and flow cytometry were combined to compare cell survival, ER-stress and apoptotic markers in 158JP and control (158N) oligodendrocytes. We observed a significant decreased viability/survival of 158JP compared to 158N cells. Consistently, ER-stress markers (BiP, caspase-12) increased in 158JP (+30%) compared to the controls. mRNA and protein ratios of apoptotic modulators (Bax/Bcl2) are higher in 158JP oligodendrocytes which are also more vulnerable than 158N cells to tunicamycin-induced ER-stress. Interestingly, 4-Phenylbutyrate (ER-stress inhibitor), which decreased ER-stress and apoptotic markers in 158JP cells, significantly increased their survival. Our results, which show a direct link between the viability and endogenous levels of ER-stress and apoptotic markers in 158JP cells, also suggest that 4-Phenylbutyrate-based strategy may contribute to develop effective strategies against oligodendrocyte dysfunctions/death and myelin disorders.

BDNF haploinsufficiency exerts a transient and regionally different influence upon oligodendroglial lineage cells during postnatal development

Brain-Derived Neurotrophic Factor (BDNF) plays important roles in promoting myelination in the developing central nervous system (CNS), however the influence it exerts on oligodendrocyte development in vivo remains unclear. As BDNF knockout mice die in the perinatal period, we undertook a systematic developmental analysis of oligodendroglial lineage cells within multiple CNS regions of BDNF heterozygous (HET) mice. Our data identify that BDNF heterozygosity results in transient reductions in oligodendroglial lineage cell density and progression that are largely restricted to the optic nerve, whereas the corpus callosum, cerebral cortex, basal forebrain and spinal cord white matter tracts are unaffected. In the first two postnatal weeks, BDNF HET mice exhibit reductions in the density of oligodendroglial lineage cells, oligodendrocyte precursor cells (OPCs) and postmitotic oligodendrocytes selectively in the optic nerve, but not in the brain or spinal cord white matter tracts. However, this normalizes later in development. The overall proportion of OPCs and mature oligodendrocytes remains unchanged from P9 to P30 in all CNS regions. This study identifies that BDNF exerts transient effects on oligodendroglial lineage cells selectively in the optic nerve during postnatal development. Taken together, this provides compelling evidence that BDNF haploinsufficiency exerts modest effects upon oligodendroglial cell density and lineage progression in vivo, suggesting its major role is restricted to promoting oligodendrocyte myelination.

Dynamic Calcium Release From Endoplasmic Reticulum Mediated by Ryanodine Receptor 3 Is Crucial for Oligodendroglial Differentiation.

Increased intracellular Ca2+ in oligodendrocyte progenitor cells (OPCs) is important to initiate their differentiation, but the intracellular Ca2+ channel involved in this process remains unclear. As a Ca2+-induced Ca2+ release (CICR) channel that mediates endoplasmic reticulum (ER) Ca2+ release, the role of ryanodine receptors (RyRs) in oligodendroglial development is unexplored. In the present study, we observed that among the three mammalian isoforms, oligodendroglial lineage cells selectively expressed RyR3. Strong RyR3-positive signal was distributed all over the cytoplasm and processes in OPCs and/or immature OLs (imOLs), whereas it gradually decreased and was located mainly around the perinuclear region in mature oligodendrocytes (OLs). In addition, RyR3-mediated intracellular Ca2+ waves following caffeine stimulation were correlated with the expression pattern of RyR3, in which high flat Ca2+ fluctuations and oscillatory Ca2+ waves were more frequently recorded in OPCs and/or imOLs than in OLs. Through further functional exploration, we demonstrated that pretreatment with the RyR antagonist ryanodine could neutralize the increase in intracellular Ca2+ induced by OPC differentiation and reduce the number of mature OLs. Moreover, gene-level knockdown of RyR3 by lentivirus in OPCs resulted in inhibition of OPC differentiation. Taken together, our results provide new insight into the crucial role of RyR3-mediated ER Ca2+ release in the regulation of OPC differentiation and/or myelination.

Characterization of a murine mixed neuron-glia model and cellular responses to regulatory T cell-derived factors

One of the unmet clinical needs in demyelinating diseases such as Multiple Sclerosis (MS) is to provide therapies that actively enhance the process of myelin regeneration (remyelination) in the central nervous system (CNS). Oligodendrocytes, the myelinating cells of the CNS, play a central role in remyelination and originate from oligodendrocyte progenitor cells (OPCs). We recently showed that depletion of regulatory T cells (Treg) impairs remyelination in vivo, and that Treg-secreted factors directly enhance oligodendrocyte differentiation. Here we aim to further characterize the dynamics of Treg-enhanced oligodendrocyte differentiation as well as elucidate the cellular components of a murine mixed neuron-glia model.Murine mixed neuron-glia cultures were generated from P2–7 C57BL/6 mice and characterized for percentage of neuronal and glial cell populations prior to treatment at 7 days in vitro (div) as well as after treatment with Treg-conditioned media at multiple timepoints up to 12 div.Mixed neuron-glia cultures consisted of approximately 30% oligodendroglial lineage cells, 20% neurons and 10% microglia. Furthermore, a full layer of astrocytes, that could not be quantified, was present. Treatment with Treg-conditioned media enhanced the proportion of MBP+ oligodendrocytes and decreased the proportion of PDGFRα+ OPCs, but did not affect OPC proliferation or survival. Treg-enhanced oligodendrocyte differentiation was not caused by Treg polarizing factors, was dependent on the number of activation cycles Treg underwent and was robustly achieved by using 5% conditioned media.These studies provide in-depth characterization of a murine mixed neuron-glia model as well as further insights into the dynamics of Treg-enhanced oligodendrocyte differentiation.

Role of Microvesicles in the Spread of Herpes Simplex Virus 1 in Oligodendrocytic Cells.

ABSTRACTHerpes simplex virus 1 (HSV-1) is a neurotropic pathogen that can infect many types of cells and establishes latent infections in the neurons of sensory ganglia. In some cases, the virus spreads into the central nervous system, causing encephalitis or meningitis. Cells infected with several different types of viruses may secrete microvesicles (MVs) containing viral proteins and RNAs. In some instances, extracellular microvesicles harboring infectious virus have been found. Here we describe the features of shedding microvesicles released by the human oligodendroglial HOG cell line infected with HSV-1 and their participation in the viral cycle. Using transmission electron microscopy, we detected for the first time microvesicles containing HSV-1 virions. Interestingly, the Chinese hamster ovary (CHO) cell line, which is resistant to infection by free HSV-1 virions, was susceptible to HSV-1 infection after being exposed to virus-containing microvesicles. Therefore, our results indicate for the first time that MVs released by infected cells contain virions, are endocytosed by naive cells, and lead to a productive infection. Furthermore, infection of CHO cells was not completely neutralized when virus-containing microvesicles were preincubated with neutralizing anti-HSV-1 antibodies. The lack of complete neutralization and the ability of MVs to infect nectin-1/HVEM-negative CHO-K1 cells suggest a novel way for HSV-1 to spread to and enter target cells. Taken together, our results suggest that HSV-1 could spread through microvesicles to expand its tropism and that microvesicles could shield the virus from neutralizing antibodies as a possible mechanism to escape the host immune response.IMPORTANCE Herpes simplex virus 1 (HSV-1) is a neurotropic pathogen that can infect many types of cells and establishes latent infections in neurons. Extracellular vesicles are a heterogeneous group of membrane vesicles secreted by most cell types. Microvesicles, which are extracellular vesicles which derive from the shedding of the plasma membrane, isolated from the supernatant of HSV-1-infected HOG cells were analyzed to find out whether they were involved in the viral cycle. The importance of our investigation lies in the detection, for the first time, of microvesicles containing HSV-1 virions. In addition, virus-containing microvesicles were endocytosed into CHO-K1 cells and were able to actively infect these otherwise nonpermissive cells. Finally, the infection of CHO cells with these virus-containing microvesicles was not completely neutralized by anti-HSV-1 antibodies, suggesting that these extracellular vesicles might shield the virus from neutralizing antibodies as a possible mechanism of immune evasion.

A single‐cell analysis reveals multiple roles of oligodendroglial lineage cells during post‐ischemic regeneration

NG2 cells represent precursors of oligodendrocytes under physiological conditions; however, following cerebral ischemia they play an important role in glial scar formation. Here, we compared the expression profiles of oligodendroglial lineage cells, after focal cerebral ischemia (FCI) and in Alzheimer's-like pathology using transgenic mice, which enables genetic fate-mapping of Cspg4-positive NG2 cells and their progeny, based on the expression of red fluorescent protein tdTomato. tdTomato-positive cells possessed the expression profile of NG2 cells and oligodendrocytes; however, based on the expression of cell type-specific genes, we were able to distinguish between them. To shed light on the changes in the expression patterns caused by FCI, we employed self-organizing Kohonen maps, enabling the division of NG2 cells and oligodendrocytes into subpopulations based on similarities in the expression profiles of individual cells. We identified three subpopulations of NG2 cells emerging after FCI: proliferative; astrocyte-like and oligodendrocyte-like NG2 cells; such phenotypes were further confirmed by immunohistochemistry. Oligodendrocytes themselves formed four subpopulations, which reflected the process of oligodendrocytes maturation. Finally, we used 5-ethynyl-2' deoxyuridine (EdU) labeling to reveal that NG2 cells can differentiate directly into reactive astrocytes without preceding proliferation. In contrast, in Alzheimer's-like pathology we failed to identify these subpopulations. Collectively, here we identified several yet unknown differences between the expression profiles of NG2 cells and oligodendrocytes, and characterized specific genes contributing to oligodendrocyte maturation and phenotypical changes of NG2 cells after FCI. Moreover, our results suggest that, unlike in Alzheimer's-like pathology, NG2 cells acquire a multipotent phenotype following FCI.

Sox2 Is Essential for Oligodendroglial Proliferation and Differentiation during Postnatal Brain Myelination and CNS Remyelination.

In the CNS, myelination and remyelination depend on the successful progression and maturation of oligodendroglial lineage cells, including proliferation and differentiation of oligodendroglial progenitor cells (OPCs). Previous studies have reported that Sox2 transiently regulates oligodendrocyte (OL) differentiation in the embryonic and perinatal spinal cord and appears dispensable for myelination in the postnatal spinal cord. However, the role of Sox2 in OL development in the brain has yet to be defined. We now report that Sox2 is an essential positive regulator of developmental myelination in the postnatal murine brain of both sexes. Stage-specific paradigms of genetic disruption demonstrated that Sox2 regulated brain myelination by coordinating upstream OPC population supply and downstream OL differentiation. Transcriptomic analyses further supported a crucial role of Sox2 in brain developmental myelination. Consistently, oligodendroglial Sox2-deficient mice developed severe tremors and ataxia, typical phenotypes indicative of hypomyelination, and displayed severe impairment of motor function and prominent deficits of brain OL differentiation and myelination persisting into the later CNS developmental stages. We also found that Sox2 was required for efficient OPC proliferation and expansion and OL regeneration during remyelination in the adult brain and spinal cord. Together, our genetic evidence reveals an essential role of Sox2 in brain myelination and CNS remyelination, and suggests that manipulation of Sox2 and/or Sox2-mediated downstream pathways may be therapeutic in promoting CNS myelin repair.SIGNIFICANCE STATEMENT Promoting myelin formation and repair has translational significance in treating myelin-related neurological disorders, such as periventricular leukomalacia and multiple sclerosis in which brain developmental myelin formation and myelin repair are severely affected, respectively. In this report, analyses of a series of genetic conditional knock-out systems targeting different oligodendrocyte stages reveal a previously unappreciated role of Sox2 in coordinating upstream proliferation and downstream differentiation of oligodendroglial lineage cells in the mouse brain during developmental myelination and CNS remyelination. Our study points to the potential of manipulating Sox2 and its downstream pathways to promote oligodendrocyte regeneration and CNS myelin repair.

Ion Mobility-Enhanced Data-Independent Acquisitions Enable a Deep Proteomic Landscape of Oligodendrocytes.

Oligodendrocytes are a type of neuroglia that provide trophic support and insulation to axons in the central nervous system. The genesis and maturation of oligodendrocytes are essential processes for myelination and the course of CNS development. Using ion mobility-enhanced, data-independent acquisitions and 2D-nanoUPLC fractionation operating at nanoscale flow rates, we established a comprehensive data set of proteins expressed by the human oligodendroglia cell line MO3.13. The final dataset incorporating all fractions comprised 223 531 identified peptides assigned to 10 390 protein hits, an improvement of 4.5 times on identified proteins described previously by our group using the same cell line. Identified proteins play pivotal roles in many biological processes such as cell growth and development and energy metabolism, providing a rich resource for future studies on oligodendrocyte development, myelination, axonal support, and the regulation of such process. Our results can help further studies that use MO3.13 cells as a tool of investigation, not only in relation to oligodendrocyte maturation, but also to diseases that have oligodendrocytes as key players. All MS data have been deposited in the ProteomeXchange with identifier PXD004696.

Ion Mobility-Enhanced Data-Independent Acquisitions Enable a Deep Proteomic Landscape of Oligodendrocytes.

Oligodendrocytes are a type of neuroglia that provide trophic support and insulation to axons in the central nervous system. The genesis and maturation of oligodendrocytes are essential processes for myelination and the course of CNS development. Using ion mobility-enhanced, data-independent acquisitions and 2D-nanoUPLC fractionation operating at nanoscale flow rates, we established a comprehensive data set of proteins expressed by the human oligodendroglia cell line MO3.13. The final dataset incorporating all fractions comprised 223 531 identified peptides assigned to 10 390 protein hits, an improvement of 4.5 times on identified proteins described previously by our group using the same cell line. Identified proteins play pivotal roles in many biological processes such as cell growth and development and energy metabolism, providing a rich resource for future studies on oligodendrocyte development, myelination, axonal support, and the regulation of such process. Our results can help further studies that use MO3.13 cells as a tool of investigation, not only in relation to oligodendrocyte maturation, but also to diseases that have oligodendrocytes as key players. All MS data have been deposited in the ProteomeXchange with identifier PXD004696.

Human miR-26a-5p regulates the glutamate transporter SLC1A1 (EAAT3) expression. Relevance in multiple sclerosis

Multiple sclerosis (MS) is an autoimmune disease of the central nervous system, characterized by chronic inflammation, demyelination and scarring as well as a broad spectrum of signs and symptoms. MicroRNA plays pivotal roles in cellular and developmental processes by regulating gene expression at the post-transcriptional level. Increasing evidence suggests the involvement of microRNAs in the pathogenesis of neurodegenerative diseases, including MS.We have already found that the expression of a specific miRNA, hsa-mir-26a-5p (miR-26a), changed during INF-β treatment in responder Relapsing-Remitting MS patients. Functional annotations of mir-26a targets revealed that a number of genes were implicated in Glutamate Receptor Signaling pathway, which is notoriously altered in neurodegenerative diseases as MS. In this study, the different potential targets were subjected to a validation test based on luciferase reporter constructs transfected in an oligodendroglial cell line. In this functional screening, miR-26a was able to interact with SLC1A1 3′ UTR suppressing the reporter activity. Transfection of a miR-26a mimic was then shown to decrease the endogenous SLC1A1 mRNA. Afterward, we have evaluated in blood platelets from interferon-β treated Multiple Sclerosis patients the expression of miR-26a and SLC1A1, finding not only their converse expression, but also a responsiveness to interferon-β therapy.Overall, these data suggest that mir-26a and SLC1A1 may play a role in the MS pathogenesis, and may be potential targets for the development of new biomarkers and/or therapeutic tools.

Exercise in Adulthood after Irradiation of the Juvenile Brain Ameliorates Long-Term Depletion of Oligodendroglial Cells.

Cranial radiation severely affects brain health and function, including glial cell production and myelination. Recent studies indicate that voluntary exercise has beneficial effects on oligodendrogenesis and myelination. Here, we hypothesized that voluntary running would increase oligodendrocyte numbers in the corpus callosum after irradiation of the juvenile mouse brain. The brains of C57Bl/6J male mice were 6 Gy irradiated on postnatal day 9 during the main gliogenic developmental phase, resulting in a loss of oligodendrocyte precursor cells. Upon adulthood, the mice were injected with bromodeoxyuridine and allowed to exercise on a running wheel for four weeks. Cell proliferation and survival, Ascl1+ oligodendrocyte precursor and Olig2+ oligodendrocyte cell numbers as well as CC1+ mature oligodendrocytes were quantified using immunohistology. Radiation induced a reduction in the number of Olig2+ oligodendrocytes by nearly 50% without affecting production or survival of new Olig2+ cells. Ascl1+ cells earlier in the oligodendroglial cell lineage were also profoundly affected, with numbers reduced by half. By three weeks of age, Olig2+ cell numbers had not recovered, and this was paralleled by a volumetric loss in the corpus callosum. The deficiency of Olig2+ oligodendrocytes persisted into adulthood. Additionally, the depletion of Ascl1+ progenitor cells was irreversible, and was even more pronounced at 12 weeks postirradiation compared to day 2 postirradiation. Furthermore, the overall number of CC1+ mature oligodendrocytes decreased by 28%. The depletion of Olig2+ cells in irradiated animals was reversed by 4 weeks of voluntary exercise. Moreover, voluntary exercise also increased the number of Ascl1+ progenitor cells in irradiated animals. Taken together, these results demonstrate that exercise in adulthood significantly ameliorates the profound and long-lasting effects of moderate exposure to immature oligodendrocytes during postnatal development.

Issue Cover (August 2017)

Front cover: Rat spinal cord oligodendrocytes differentiating in vitro. A pure oligodendroglial lineage cell (OLC) culture from the rat neonatal spinal cord is stained for oligodendrocyte markers, O4 antigen (red) and galactocerebroside (green) by immunocytochemistry. Nuclei are stained with DAPI (Blue). By comparing pure oligodendrocyte cultures from rat spinal cords and forebrains, we demonstrated differing intrinsic biological properties between these two OLC groups originated from distinct CNS regions. Further understanding the regional differences of oligodendrocytes will help to improve treatments for demyelination in different CNS regions and to facilitate the development of stem cell-derived OLCs for cell transplantation therapies for demyelination. Read the full article ‘Differing intrinsic biological properties between forebrain and spinal oligodendroglial lineage cells’ by M. Horiuchi, Y. Suzuki-Horiuchi, T. Akiyama, A. Itoh, D. Pleasure, E. Carstens and T. Itoh (J. Neurochem. 2017, vol. 142 (3), pp. 378–391) on doi: 10.1111/jnc.14074

Differing intrinsic biological properties between forebrain and spinal oligodendroglial lineage cells.

Differentiation of oligodendroglial progenitor cells (OPCs) into myelinating oligodendrocytes is known to be regulated by the microenvironment where they differentiate. However, current research has not verified whether or not oligodendroglial lineage cells (OLCs) derived from different anatomical regions of the central nervous system (CNS) respond to microenvironmental cues in the same manner. Here, we isolated pure OPCs from rat neonatal forebrain (FB) and spinal cord (SC) and compared their phenotypes in the same invitro conditions. We found that although FB and SC OLCs responded differently to the same external factors; they were distinct in proliferation response to mitogens, oligodendrocyte phenotype after differentiation, and cytotoxic responses to α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate-type glutamate receptor-mediated excitotoxicity at immature stages of differentiation in a cell-intrinsic manner. Moreover, transcriptome analysis identified genes differentially expressed between these OPC populations, including those encoding transcription factors (TFs), cell surface molecules, and signaling molecules. Particularly, FB and SC OPCs retained the expression of FB- or SC-specific TFs, such as Foxg1 and Hoxc8, respectively, even after serial passaging invitro. Given the essential role of these TFs in the regional identities of CNS cells along the rostrocaudal axis, our results suggest that CNS region-specific gene regulation by these TFs may cause cell-intrinsic differences in cellular responses between FB and SC OLCs to extracellular molecules. Further understanding of the regional differences among OPC populations will help to improve treatments for demyelination in different CNS regions and to facilitate the development of stem cell-derived OPCs for cell transplantation therapies for demyelination. Cover Image for this issue: doi. 10.1111/jnc.13809.

NCAM1 is the Target of miRNA-572: Validation in the Human Oligodendroglial Cell Line.

The neural cell adhesion molecule 1 (NCAM1) is a fundamental protein in cell-cell interaction and in cellular developmental processes, and its dysregulation is involved in a number of diseases including multiple sclerosis. Studies in rats suggest that the modulation of NCAM1 expression is regulated by miRNA-572, but no data are available confirming such interaction in the human system. We analyzed whether this is the case using a human oligodendroglial cell line (MO3.13). MO3.13 cells were transfected with miRNA-572 mimic and inhibitor separately; NCAM1 mRNA and protein expression levels were analyzed at different time points after transfection. Results indicated that NCAM1 expression is increased after transfection with miRNA-572 inhibitor, whereas it is decreased after transfection with the mimic (p<0.005). The interaction between NCAM1 and miRNA-572 was subsequently confirmed in a Vero cell line that does not express NCAM1, by luciferase assay after transfection with NCAM1. These results confirm that miRNA-572 regulates NCAM1 and for the first time demonstrate that this interaction regulates NCAM1 expression in human cells. Data herein also support the hypothesis that miRNA-572 is involved in diseases associated with NCAM1 deregulation, suggesting its possible use as a biomarker in these diseases.

Cuprizone Intoxication Induces Cell Intrinsic Alterations in Oligodendrocyte Metabolism Independent of Copper Chelation.

Cuprizone intoxication is a common animal model used to test myelin regenerative therapies for the treatment of diseases such as multiple sclerosis. Mice fed this copper chelator develop reversible, region-specific oligodendrocyte loss and demyelination. While the cellular changes influencing the demyelinating process have been explored in this model, there is no consensus about the biochemical mechanisms of toxicity in oligodendrocytes and about whether this damage arises from the chelation of copper in vivo. Here we have identified an oligodendroglial cell line that displays sensitivity to cuprizone toxicity and performed global metabolomic profiling to determine biochemical pathways altered by this treatment. We link these changes with alterations in brain metabolism in mice fed cuprizone for 2 and 6 weeks. We find that cuprizone induces widespread changes in one-carbon and amino acid metabolism as well as alterations in small molecules that are important for energy generation. We used mass spectrometry to examine chemical interactions that are important for copper chelation and toxicity. Our results indicate that cuprizone induces global perturbations in cellular metabolism that may be independent of its copper chelating ability and potentially related to its interactions with pyridoxal 5'-phosphate, a coenzyme essential for amino acid metabolism.

DNA methylation in oligodendroglial cells during developmental myelination and in disease.

Oligodendrocyte progenitor cells (OPC) are the myelinating cells of the central nervous system (CNS). During development, they differentiate into mature oligodendrocytes (OL) and ensheath axons, providing trophic and functional support to the neurons. This process is regulated by the dynamic expression of specific transcription factors, which, in turn, is controlled by epigenetic marks such as DNA methylation. Here we discuss recent findings showing that DNA methylation levels are differentially regulated in the oligodendrocyte lineage during developmental myelination, affecting both genes expression and alternative splicing events. Based on the phenotypic characterization of mice with genetic ablation of DNA methyltransferase 1 (Dnmt1) we conclude that DNA methylation is critical for efficient OPC expansion and for developmental myelination. Previous work suggests that in the context of diseases such as multiple sclerosis (MS) or gliomas, DNA methylation is differentially regulated in the CNS of affected individuals compared with healthy controls. In this commentary, based on the results of previous work, we propose the potential role of DNA methylation in adult oligodendroglial lineage cells in physiologic and pathological conditions, and delineate potential research approaches to be undertaken to test this hypothesis. A better understanding of this epigenetic modification in adult oligodendrocyte progenitor cells is essential, as it can potentially result in the design of new therapeutic strategies to enhance remyelination in MS patients or reduce proliferation in glioma patients.

Neonatal Dexamethasone Treatment Exacerbates Hypoxia/Ischemia-Induced White Matter Injury.

Dexamethasone, a synthetic glucocorticoid, has been widely used to prevent or ameliorate morbidity of chronic lung disease in preterm infants with respiratory distress syndrome. Despite its beneficial effect on neonatal lung function, growing concern has arisen about adverse effects of this clinical practice on fetal brain development. We demonstrated previously that neonatal dexamethasone (DEX) treatment may render the newborn brain to be more vulnerable to hypoxia/ischemia (HI)-induced gray matter injury. Here, we examined whether neonatal DEX treatment may also affect the extent of HI-induced subcortical white matter (WM) injury in the developing rat brain. Using a HI model of premature brain injury, we demonstrated that a 3-day tapering course (0.5, 0.3, and 0.1mg/kg) of DEX treatment in rat pups on postnatal days 1-3 (P1-3) significantly reduced the number of all stages of the oligodendroglial lineage cells on P7 and exacerbated HI-induced WM injury. Neonatal DEX treatment also enhanced HI-induced oligodendroglial apoptosis and astrocyte activation in the developing WM on P14. Likewise, HI-induced reductions in myelin thickness, axon caliber, and function during WM development were exacerbated by neonatal DEX treatment. Furthermore, neonatal DEX treatment further aggravated HI-induced motor deficits as assessed in the rotarod test. We also found that the administration of β-lactam antibiotic ceftriaxone increased glutamate transporter-1 protein expression and significantly reduced HI-induced WM injury in neonatal DEX-treated rats. These results suggest that neonatal DEX treatment may lead the developing brain to be more vulnerable to subsequent HI-induced WM injury, which can be ameliorated by ceftriaxone administration.

Auraptene induces oligodendrocyte lineage precursor cells in a cuprizone-induced animal model of demyelination

We investigated the effects of auraptene on mouse oligodendroglial cell lineage in an animal model of demyelination induced by cuprizone. Auraptene, a citrus coumarin, was intraperitoneally administered to mice fed the demyelinating agent cuprizone. Immunohistochemical analysis of the corpus callosum and/or Western blotting analysis of brain extracts revealed that cuprizone reduced immunoreactivity for myelin-basic protein, a marker of myelin, whereas it increased immunoreactivity to platelet derived-growth factor receptor-α, a marker of oligodendrocyte precursor cells. Administration of auraptene enhanced the immunoreactivity to oligodendrocyte transcription factor 2, a marker of oligodendrocyte precursor cells and oligodendrocyte lineage precursor cells, but had no effect on immunoreactivity to myelin-basic protein or platelet-derived growth factor receptor-α. These findings suggest that auraptene promotes the production of oligodendrocyte lineage precursor cells in an animal model of demyelination and may be useful for individuals with demyelinating diseases.