latelet-derived growth factor alpha-receptor (PDGFαR) mediated signaling plays a key role in the development of glial cells of the central nervous system. In vivo and in vitro studies show that PDGFαR is actively expressed in proliferative and motile oligodendrocyte type-2 astrocyte (O-2A) glial progenitor cells. However, PDGFαR expression is barely detectable in mature glial cells. The exact mechanism underlying the loss of PDGFαR expression is unknown. In this study, we employed a rat brain-derived O-2A glial progenitor cell line, CG4 as a culture model to investigate signals capable of inhibiting PDGFαR gene expression. PDGFαR mRNA levels decreased significantly as CG4 cells differentiated into both oligodendrocyte and astrocyte lineages. We showed that inhibition of PDGFαR expression was promoted by prostaglandin E2 via protein kinase A activation. Both cAMP analogs (db-cAMP and 8'bromo-cAMP) and adenylate cyclase activator (forskolin) were potent suppressors of PDGFαR expression in CG4 cells. This inhibitory effect resulted from an increased destabilization of PDGFαR mRNA instead of a decreased PDGFαR gene transcription. Importantly, db-cAMP failed to reduce PDGFαR mRNA levels in several PDGFαR over-expressing human glioma cell lines. Together, these results suggest that cAMP-dependent pathway played a key regulatory role in controlling PDGFαR mRNA levels during normal glial development, and that a breakdown in the cross talk between cAMP and PDGF pathways may underlie the uncontrolled proliferation and immature differentiation state in the glial tumors.
PDGF; cyclic AMP; mRNA turnover; glioma
Accumulations of hypertrophic, intensely glial fibrillary acidic protein positive (GFAP)+ astroglia, which also express immunoreactive nestin and vimentin, are prominent features of multiple sclerosis lesions. The issues of the cellular origin of hypertrophic GFAP+/vimentin+/nestin+ “reactive” astroglia and also the plasticities and lineage relationships among three macroglial progenitor populations - oligodendrocyte progenitor cells (OPCs), astrocytes and ependymal cells - during multiple sclerosis and other CNS diseases remain controversial. We employed genetic fate-mappings with a battery of inducible Cre drivers (Olig2-Cre-ERT2, GFAP-Cre-ERT2, FoxJ1-Cre-ERT2 and Nestin-Cre-ERT2) to explore these issues in adult mice with myelin oligodendrocyte glycoprotein peptide-induced experimental autoimmune encephalomyelitis (EAE). The proliferative rate of spinal cord OPCs rose five-fold above control levels during EAE, and numbers of oligodendroglia increased as well, but astrogenesis from OPCs was rare. Spinal cord ependymal cells, previously reported to be multipotent, did not augment their low proliferative rate, nor give rise to astroglia or OPCs. Instead, the hypertrophic, vimentin+/nestin+, reactive astroglia that accumulated in spinal cord in this multiple sclerosis model were derived by proliferation and phenotypic transformation of fibrous astroglia in white matter, and solely by phenotypic transformation of protoplasmic astroglia in gray matter. This comprehensive analysis of macroglial plasticity in EAE helps to clarify the origins of astrogliosis in CNS inflammatory demyelinative disorders.
experimental autoimmune encephalomyelitis; plasticity; reactive astrocytes; oligodendrocyte progenitor cells; ependymal cells; fate-mapping
The adult mammalian brain and spinal cord contain glial precursors that express platelet-derived growth factor receptors (alpha subunit, PDGFRA) and the NG2 proteoglycan. These “NG2 cells” descend from oligodendrocyte precursors in the perinatal CNS and continue to generate myelinating oligodendrocytes in the grey and white matter of the postnatal brain. It has been proposed that NG2 cells can also generate reactive astrocytes at sites of CNS injury or demyelination. To test this we examined the fates of PDGFRA/ NG2 cells in the mouse spinal cord during experimental autoimmune encephalomyelitis (EAE) - a demyelinating condition that models some aspects of multiple sclerosis in humans. We administered tamoxifen to Pdgfra-CreERT2: Rosa26R-YFP mice in order to induce yellow fluorescent protein (YFP) expression in PDGFRA/ NG2 cells and their differentiated progeny. We subsequently induced EAE and observed a large (>4-fold) increase in the local density of YFP+ cells, >90% of which were oligodendrocyte lineage cells. Many of these became CC1-positive, NG2-negative differentiated oligodendrocytes that expressed myelin markers CNP and Tmem10/ Opalin. PDGFRA/ NG2 cells generated very few GFAP+ reactive astrocytes (1-2% of all YFP+ cells) or NeuN+ neurons (<0.02%). Thus, PDGFRA/ NG2 cells act predominantly as a reservoir of new oligodendrocytes in the demyelinated spinal cord.
Demyelination; oligodendrocyte; neural precursor cell; Cre-lox; spinal cord; mouse
During development, multipotent neural precursors give rise to oligodendrocyte progenitor cells (OPCs), which migrate and divide to produce additional OPCs. Near the end of embryogenesis and during postnatal stages, many OPCs stop dividing and differentiate as myelinating oligodendrocytes, whereas others persist as nonmyelinating cells. Investigations of oligodendrocyte development in mice indicated that the Nkx2.2 transcription factor both limits the number of OPCs that are formed and subsequently promotes their differentiation, raising the possibility that Nkx2.2 plays a key role in determining myelinating versus nonmyelinating fate. We used in vivo time-lapse imaging and loss-of-function experiments in zebrafish to further explore formation and differentiation of oligodendrocyte lineage cells. Our data show that newly specified OPCs are heterogeneous with respect to gene expression and fate. Whereas some OPCs express the nkx2.2a gene and differentiate as oligodendrocytes, others that do not express nkx2.2a mostly remain as nonmyelinating OPCs. Similarly to mouse, loss of nkx2.2a function results in excess OPCs and delayed oligodendrocyte differentiation. Notably, excess OPCs are formed as a consequence of prolonged OPC production from neural precursor cells. We conclude that Nkx2.2 promotes timely specification and differentiation of myelinating oligodendrocyte lineage cells from species representing different vertebrate taxa.
Olig2; zebrafish; neural precursors; glia
Experimental models of myelin disorders can be treated by the transplantation of oligodendrocyte progenitor cells (OPCs) into the affected brain or spinal cord. OPCs express gangliosides recognized by MAb A2B5, but this marker also identifies lineage-restricted astrocytes and immature neurons. To establish a more efficient means of isolating myelinogenic OPCs, we asked if FACS could be used to sort PDGFα receptor+ cells from fetal human forebrain, based on expression of the PDGFRα epitope CD140a. CD140a+ isolates were maintained as mitotic bipotential progenitors that could be instructed to either oligodendrocyte or astrocyte fate. Transplanted CD140a+ cells were highly migratory, and rapidly and robustly myelinated the hypomyelinated shiverer mouse brain, more efficiently than did A2B5-sorted cells. Microarray analysis of CD140a+ cells revealed their differential expression of CD9, as well as of PTN-PTPRZ1, wnt, notch and BMP pathway components, indicating the dynamic interaction of self-renewal and fate-restricting pathways in these cells.
oligodendrocyte progenitor; PDGF receptor; myelin; remyelination
Cycling glial precursors - “NG2-glia” - are abundant in the developing and mature central nervous system (CNS). During development they generate oligodendrocytes. In culture, they can revert to a multipotent state, suggesting that they might have latent stem cell potential that could be harnessed to treat neurodegenerative disease. This hope has been subdued recently by a series of fate mapping studies that cast NG2-glia as dedicated oligodendrocyte precursors in the healthy adult CNS - though rare neuron production in the piriform cortex remains a possibility. Following CNS damage, the repertoire of NG2-glia expands to include Schwann cells and possibly astrocytes – but so far not neurons. This confirms the central role of NG2-glia in myelin repair. The realization that oligodendrocyte generation continues throughout normal adulthood has seeded the idea that myelin genesis might also be involved in neural plasticity. We review these developments, highlighting areas of current interest, contention and speculation.
oligodendrocyte; precursor/stem cell; cell fate; transgenic mice; Cre-lox; motor neuron disease; amyotrophic lateral sclerosis; multiple sclerosis; experimental autoimmune encephalomyelitis; EAE; spinal injury; axotomy; demyelination; motor skills; behaviour
A demyelinating disease induced in C57B1/6N mice by intracranial injection of a coronavirus (murine hepatitis virus strain A59) is followed by functional recovery and efficient CNS myelin repair. To study the biological properties of the cells involved in this repair process, glial cells were isolated and cultured from spinal cords of these young adult mice during demyelination and remyelination. Using three-color immunofluorescence combined with [3H]thymidine autoradiography, we have analyzed the antigenic phenotype and mitotic potential of individual glial cells. We identified oligodendrocytes with an antibody to galactocerebroside, astrocytes with an antibody to glial fibrillary acidic protein, and oligodendrocyte-type 2 astrocyte (O-2A) progenitor cells with the O4 antibody. Cultures from demyelinated tissue differed in several ways from those of age-matched controls: first, the total number of O-2A lineage cells was strikingly increased; second, the O-2A population consisted of a higher proportion of O4-positive astrocytes and cells of mixed oligodendrocyte-astrocyte phenotype; and third, all the cell types within the O-2A lineage showed enhanced proliferation. This proliferation was not further enhanced by adding PDGF, basic fibroblast growth factor (bFGF), or insulin-like growth factor I (IGF-I) to the defined medium. However, bFGF and IGF-I seemed to influence the fate of O-2A lineage cells in cultures of demyelinated tissue. Basic FGF decreased the percentage of cells expressing galactocerebroside. In contrast, IGF-I increased the relative proportion of oligodendrocytes. Thus, O-2A lineage cells from adult mice display greater phenotypic plasticity and enhanced mitotic potential in response to an episode of demyelination. These properties may be linked to the efficient remyelination achieved in this demyelinating disease.
Extracellular signals play essential roles in controlling the proliferation and differentiation of oligodendrocyte progenitor cells in the developing central nervous system. However, the intracellular pathways that transduce these extrinsic signals remain to be elucidated. In this study, we showed that conditional ablation of the nonreceptor tyrosine phosphatase Shp2 in Olig1-expressing oligodendrocyte lineage resulted in dramatic reduction in the generation and proliferation of oligodendrocyte progenitor cells in the spinal cord. Maturation and myelination of oligodendrocytes were also compromised in the Shp2 mutants. The deficits in oligodendrocyte development in Shp2 mutants nearly phenocopied those observed in PDGF-A mutants, suggesting that Shp2 is a crucial component in transducing PDGFRα signals in the control of oligodendrocyte proliferation and maturation.
Shp2; oligodendrocyte; spinal cord; proliferation; differentiation
Endogenous cell proliferation and gliogenesis have been extensively documented in spinal cord injury, particularly in terms of proliferating oligodendrocyte progenitor cells. Despite the characterization of different proliferating cell types in the intact and injured spinal cord, the exact sources of new glial cells have remained elusive. Most studies on cell fate within the spinal cord have focused on following the progeny of one specific population of dividing cells, thus making it difficult to understand the relative contributions of each mitotic cell population to the formation of new glia after spinal cord injury. A recent study from the Frisen laboratory is the first to quantitatively and qualitatively characterize the response of ependymal cells, oligodendrocyte progenitors, and astrocytes in parallel by using transgenic reporter mice corresponding to each cell type. The investigators characterize the distribution and phenotype of progeny, along with the quantitative contributions of each progenitor type to newly formed cells. Their findings provide valuable insight into the endogenous cell replacement response to spinal cord injury, thus paving the way for advances in modulating specific populations of progenitor cells with the goal of promoting structural and functional recovery after spinal cord injury.
Bone morphogenetic proteins have been implicated in the development of oligodendrocytes and astrocytes, however, a role for endogenous BMP signaling in glial development has not been demonstrated in a genetic model. Using mice in which signaling via type I BMP receptors Bmpr1a and Bmpr1b have been inactivated in the neural tube, we demonstrate that BMP signaling contributes to the maturation of glial cells in vivo. At P0, mutant mice exhibited a 25-40% decrease in GFAP+ or S100β+ astrocytes in the cervical spinal cord. The number of oligodendrocyte precursors and the timing of their emergence was unchanged in the mutant mice compared to the normals, however myelin protein expression and mature oligodendrocyte numbers were significantly reduced. These data indicate that BMP signaling promotes the generation of astrocytes and mature, myelinating oligodendrocytes in vivo but does not affect oligodendrocyte precursor development, thus suggesting tight regulation of BMP signaling to ensure proper gliogenesis.
Oligodendrocyte progenitor cells first proliferate to generate sufficient cell numbers and then differentiate into myelin-producing oligodendrocytes. The signal transduction mediators that underlie these events, however, remain poorly understood. The tyrosine phosphatase Shp1 has been linked to oligodendrocyte differentiation as Shp1-deficient mice show hypomyelination. The Shp1 homologue, Shp2, has recently been shown to regulate astrogliogenesis but its role in oligodendrocyte development remains unknown. Here we report that Shp2 protein levels were developmentally regulated in oligodendrocytes, with Shp2 phosphorylation being promoted by oligodendroglial mitogens but suppressed by laminin, an extracellular matrix protein that promotes oligodendroglial differentiation. In contrast, oligodendrocyte progenitors were found to be unresponsive to mitogens following Shp2, but not Shp1, depletion. In agreement with previous studies, Shp1 depletion led to decreased levels of myelin basic protein in differentiating oligodendrocytes, as well as reduced outgrowth of myelin membrane sheets. Shp2 depletion in contrast did not prevent oligodendrocyte differentiation but promoted expanded myelin membrane outgrowth. Taken together these data suggest that Shp1 and Shp2 have distinct functions in oligodendrocyte development: Shp2 regulates oligodendrocyte progenitor proliferation and Shp1 regulates oligodendrocyte differentiation. Adhesion to laminin may additionally provide extrinsic regulation of Shp2 activity and thus promote the transition from progenitor to differentiating oligodendrocyte.
Shp1; Shp2; oligodendrocyte; myelin; tyrosine phosphatase
Limited knowledge about human oligodendrogenesis prompted us to explore the lineage relationship between cortical radial glia (RG) cells and oligodendrocytes in the human fetal forebrain. RG cells were isolated from cortical ventricular/subventricular zone and their progeny was followed in vitro. One portion of RG cells differentiated into cells of oligodendrocyte lineage identified by cell-type specific antibodies, including PDGFRα, NG2, O4, MBP and MOG. Moreover, using Cre Lox fate mapping (BLBP-Cre/Floxed-YFP) we established a direct link between RG cells and oligodendrocyte progenitors. In vitro generation of RG-derived O4+ oligodendrocytes progenitors was enhanced by addition of Sonic hedgehog (SHH) and reduced by SHH inhibitor-cyclopamine, suggesting the role of SHH-signaling in this process. In summary, our in vitro experiments revealed that a portion of cortical RG cells isolated from human forebrain at the second trimester of gestation generate oligodendrocyte progenitors and suggest a role of SHH in this process.
oligodendrocyte progenitors; myelination; primate brain development; LeX immunocytochemistry; transfection; Cre-LoxP fate mapping
C57 BL/6N mice injected intracranially with the A59 strain of mouse hepatitis virus exhibit extensive viral replication in glial cells of the spinal cord and develop demyelinating lesions followed by virus clearing and remyelination. To study how different glial cell types are affected by the disease process, we combine three-color immunofluorescence labeling with tritiated thymidine autoradiography on 1-micron frozen sections of spinal cord. We use three different glial cell specific antibodies (a) to 2',3' cyclic-nucleotide 3' phosphohydrolase (CNP) expressed by oligodendrocytes, (b) to glial fibrillary acidic protein (GFAP) expressed by astrocytes, and (c) the O4 antibody which binds to O-2A progenitor cells in the rat. These progenitor cells, which give rise to oligodendrocytes and type 2 astrocytes and react with the O4 antibody in the adult central nervous system, were present but rare in the spinal cord of uninfected mice. In contrast, cells with the O-2A progenitor phenotype (O4 + only) were increased in number at one week post viral inoculation (1 WPI) and were the only immunostained cells labeled at that time by a 2-h in vivo pulse of tritiated thymidine. Both GFAP+ only and GFAP+, O4+ astrocytes were also increased in the spinal cord at 1 WPI. Between two and four WPI, the infected spinal cord was characterized by the loss of (CNP+, O4+) oligodendrocytes within demyelinating lesions and the presence of O-2A progenitor cells and O4+, GFAP+ astrocytes, both of which could be labeled with thymidine. As remyelination proceeded, CNP immunostaining returned to near normal and tritiated thymidine injected previously during the demyelinating phase now appeared in CNP+ oligodendrocytes. Thus O4 positive O-2A progenitor cells proliferate early in the course of the demyelinating disease, while CNP positive oligodendrocytes do not. The timing of events suggests that the O-2A progenitors may give rise to new oligodendrocytes and to type 2 astrocytes, both of which are likely to be instrumental in the remyelination process.
MicroRNAs (miRNAs) regulate various biological processes, but evidence for miRNAs that control the differentiation program of specific neural cell types has been elusive. To determine the role of miRNAs in the formation of myelinating oligodendrocytes, we selectively deleted a miRNA-processing enzyme Dicer1 in oligodendrocyte lineage cells. Mice lacking Dicer1 display severe myelinating deficits despite an expansion of oligodendrocyte progenitor pool. To search for miRNAs responsible for the induction of oligodendrocyte maturation, we identified miR-219 and miR-338 as oligodendrocyte-specific miRNAs in spinal cord. Overexpression of these miRNAs is sufficient to promote oligodendrocyte differentiation. Additionally, blockage of these miRNA activities in oligodendrocyte precursor culture and knockdown of miR-219 in zebrafish inhibit oligodendrocyte maturation. miR-219 and miR-338 function in part by directly repressing negative regulators of oligodendrocyte differentiation, including transcription factors Sox6 and Hes5. These findings illustrate that miRNAs are important regulators of oligodendrocyte differentiation, providing new targets for myelin repair.
Myelination; Dicer; differentiation inhibitors; oligodendrocytes; small non-coding RNAs; miR-219; miR-338; transcriptional regulation
Two recently generated targeted mouse alleles of the neurogenic gene Ascl1 were utilized in order to characterize cerebellum circuit formation. First, genetic inducible fate mapping (GIFM) with an Ascl1CreER allele was found to specifically mark all glial and neuron cell types that arise from the ventricular zone (vz). Moreover, each cell type has a unique temporal profile of marking with Ascl1CreER GIFM. Of great utility, Purkinje cells (Pcs), an early cohort of Bergmann glia, and four classes of GABAergic interneurons can be genetically birthdated during embryogenesis using Ascl1CreER GIFM. Astrocytes and oligodendrocytes in contrast express Ascl1CreER throughout their proliferative phase in the white matter. Interestingly, the final position each neuron type acquires differs depending on when it expresses Ascl1. Interneurons (including candelabrum) attain a more outside position the later they express Ascl1, whereas Pcs have distinct settling patterns each day they express Ascl1. Second, using a conditional Ascl1 allele we discovered that Ascl1 is differentially required for generation of most vz-derived cells. Mice lacking Ascl1 in the cerebellum have a major decrease in three types of interneurons with a tendency towards a loss of later born interneurons, as well as an imbalance of oligodendrocytes and astrocytes. Double mutant analysis indicates that a related helix-loop-helix protein, Ptf1a, functions with Ascl1 in generating interneurons and Pcs. By fate mapping vz-derived cells in Ascl1 mutants, we further discovered that Ascl1 plays a specific role during the time period when Pcs are generated in restricting vz progenitors from becoming rhombic lip progenitors.
GIFM; birth dating; Mash1; Purkinje cells; interneurons; candelabrum interneuron
The highly related transcription factors Sox4 and Sox11 are both expressed in oligodendrocyte precursors. Yet whether they have a function in oligodendrocyte development is unknown. By overexpressing Sox4 under the control of 3.1 kb of 5′ flanking sequences of the myelin basic protein gene in transgenic mice, we extended Sox4 expression in the oligodendrocyte lineage from oligodendrocyte precursors to cells undergoing terminal differentiation. As a consequence of transgene expression, mice develop the full spectrum of phenotypic traits associated with a severe hypomyelination during the first postnatal weeks. Myelin gene expression was severely reduced, and myelin dramatically thinned in several central nervous system (CNS) regions. Despite these disturbances in CNS myelination, the number of oligodendrocytic cells remained unaltered. Considering that apoptosis rates were normal and proliferation only slightly increased, oligodendrocytes likely persist in a premyelinating to early myelinating state. This shows that prolonged Sox4 expression in cells of the oligodendrocyte lineage is incompatible with the acquisition of a fully mature phenotype and argues that the presence of Sox4, and possibly Sox11, in oligodendrocyte precursors may normally prevent premature differentiation.
Adult multipotent neural progenitor cells can differentiate into neurons, astrocytes, and oligodendrocytes in the mammalian central nervous system, but the molecular mechanisms that control their differentiation are not yet well understood. Insulin-like growth factor I (IGF-I) can promote the differentiation of cells already committed to an oligodendroglial lineage during development. However, it is unclear whether IGF-I affects multipotent neural progenitor cells. Here, we show that IGF-I stimulates the differentiation of multipotent adult rat hippocampus-derived neural progenitor cells into oligodendrocytes. Modeling analysis indicates that the actions of IGF-I are instructive. Oligodendrocyte differentiation by IGF-I appears to be mediated through an inhibition of bone morphogenetic protein signaling. Furthermore, overexpression of IGF-I in the hippocampus leads to an increase in oligodendrocyte markers. These data demonstrate the existence of a single molecule, IGF-I, that can influence the fate choice of multipotent adult neural progenitor cells to an oligodendroglial lineage.
glia; neural stem cell; insulin; BMP; hippocampus
During development, progenitors that are committed to differentiate into oligodendrocytes, the myelinating cells of the central nervous system (CNS), are generated within discrete regions of the neuroepithelium. More specifically, within the developing spinal cord and hindbrain ventrally located progenitor cells that are characterized by the expression of the transcription factor olig2 give temporally rise to first motor neurons and then oligodendrocyte progenitors. The regulation of this temporal neuron-glial switch has been found complex and little is known about the extrinsic factors regulating it. Our studies described here identified a zebrafish ortholog to mammalian atx, which displays evolutionarily conserved expression pattern characteristics. Most interestingly, atx was found to be expressed by cells of the cephalic floor plate during a time period when ventrally-derived oligodendrocyte progenitors arise in the developing hindbrain of the zebrafish. Knock-down of atx expression resulted in a delay and/or inhibition of the timely appearance of oligodendrocyte progenitors and subsequent developmental stages of the oligodendrocyte lineage. This effect of atx knock-down was not accompanied by changes in the number of olig2-positive progenitor cells, the overall morphology of the axonal network or the number of somatic abducens motor neurons. Thus, our studies identified Atx as an extrinsic factor that is likely secreted by cells from the floor plate and that is involved in regulating specifically the progression of olig2-positive progenitor cells into lineage committed oligodendrocyte progenitors.
myelination; glia differentiation; CNS development; floor plate; zebrafish
In response to stroke, subpopulations of cortical reactive astrocytes proliferate and express proteins commonly associated with neural stem/progenitor cells, such as Glial Fibrillary Acidic Protein (GFAP) and Nestin. To examine the stem cell-related properties of cortical reactive astrocytes after injury, we generated GFAP-CreER™;tdRFP mice to permanently label reactive astrocytes. We isolated cells from the cortical peri-infarct area 3 days after stroke, and cultured them in neural stem cell medium containing EGF and bFGF. We observed tdRFP-positive neural spheres in culture, suggestive of tdRFP-positive reactive astrocyte-derived neural stem/progenitor cells (Rad-NSCs). Cultured Rad-NSCs self-renewed and differentiated into neurons, astrocytes, and oligodendrocytes. Pharmacological inhibition and conditional knockout mouse studies showed that Presenilin 1 and Notch 1 controlled neural sphere formation by Rad-NSCs after stroke. To examine the self-renewal and differentiation potential of Rad-NSCs in vivo, Rad-NSCs were transplanted into embryonic, neonatal and adult mouse brains. Transplanted Rad-NSCs were observed to persist in the subventricular zone and secondary Rad-NSCs were isolated from the host brain 28 days after transplantation. In contrast with neurogenic post-natal day 4 NSCs and adult NSCs from the subventricular zone, transplanted Rad-NSCs differentiated into astrocytes and oligodendrocytes, but not neurons, demonstrating that Rad-NSCs had restricted differentiation in vivo. Our results indicate that Rad-NSCs are unlikely to be suitable for neuronal replacement in the absence of genetic or epigenetic modification.
Neural stem/progenitor cells (NPCs) can differentiate into neurons, astrocytes and oligodendrocytes. NPCs are considered valuable for the cell therapy of injuries in the central nervous system (CNS). However, when NPCs are transplanted into the adult mammalian spinal cord, they mostly differentiate into glial lineage. The same results have been observed for endogenous NPCs during spinal cord injury. However, little is known about the mechanism of such fate decision of NPCs.
In the present study, we have found that myelin protein and Nogo-66 promoted the differentiation of NPCs into glial lineage. NgR and mTOR-Stat3 pathway were involved in this process. Releasing NgR from cell membranes or blocking mTOR-STAT3 could rescue the enhanced glial differentiation by Nogo-66.
These results revealed a novel function of Nogo-66 in the fate decision of NPCs. This discovery could have profound impact on the understanding of CNS development and could improve the therapy of CNS injuries.
Repair of myelin injury in multiple sclerosis may fail resulting in chronic demyelination, axonal loss and disease progression. As cellular pathways regulated by Phosphatase and tensin homologue deleted on chromosome 10 (PTEN; e.g., PI-3Kinase) have been reported to enhance axon regeneration and oligodendrocyte maturation, we investigated potentially beneficial effects of Pten loss-of-function in the oligodendrocyte lineage on remyelination.
We characterized oligodendrocyte numbers and myelin sheath thickness in mice with conditional inactivation of PTEN in oligodendrocytes, Olig2-cre, Ptenfl/fl mice. Utilizing a model of CNS demyelination, lysolecithin injection into the spinal cord white matter, we performed short and long-term lesioning experiments and quantified oligodendrocyte maturation and myelin sheath thickness in remyelinating lesions.
During development, we observed dramatic hypermyelination in the corpus callosum and spinal cord. Following white matter injury, however, there was no detectable improvement in myelin repair. Moreover, we observed progressive myelin sheath abnormalities and massive axon degeneration in the fasciculus gracilis of mutant animals, as indicated by ultrastructure and expression of SMI-32, APP and Caspase 6.
These studies indicate adverse effects of chronic PTEN inactivation (and by extension, activation PI-3K signaling) on myelinating oligodendrocytes and their axonal targets. We conclude that Pten function in oliogodendrocytes is required to regulate myelin thickness and preserve axon integrity. In contrast, Pten is dispensable during myelin repair and its inactivation confers not detectable benefit.
The transcriptional control of CNS myelin gene expression is poorly understood. Here we identify gene model 98, which we have named Myelin-gene Regulatory Factor (MRF), as a transcriptional regulator required for CNS myelination. Within the CNS, MRF is specifically expressed by postmitotic oligodendrocytes. MRF is a nuclear protein containing an evolutionarily conserved DNA binding domain homologous to a yeast transcription factor. Knockdown of MRF in oligodendrocytes by RNA interference prevents expression of most CNS myelin genes; conversely, overexpression of MRF within cultured oligodendrocyte progenitors or the chick spinal cord promotes expression of myelin genes. In mice lacking MRF within the oligodendrocyte lineage, pre-myelinating oligodendrocytes are generated but display severe deficits in myelin gene expression and fail to myelinate. These mice display severe neurological abnormalities, and die due to seizures during the third postnatal week. These findings establish MRF as a critical transcriptional regulator essential for oligodendrocyte maturation and CNS myelination.
CNS glia exhibit a variety of gap junctional interactions: between neighboring astrocytes, between neighboring oligodendrocytes, between astrocytes and oligodendrocytes, and as ‘reflexive’ structures between layers of myelin in oligodendrocytes. Together, these junctions are thought to form a network facilitating absorption and removal of extracellular K+ released during neuronal activity. In mice, loss of the two major oligodendrocyte connexins causes severe demyelination and early mortality, while loss of the two major astrocyte connexins causes mild dysmyelination and sensorimotor impairment, suggesting that reflexive and/or oligo-oligo coupling may be more important for the maintenance of myelin than other forms. To further explore the functional relationships between glial connexins, we generated double knockout mice lacking one oligodendrocyte and one astrocyte connexin. Cx32-Cx43 dKO animals develop white matter vacuolation without obvious ultrastructural abnormalities in myelin. Progressive loss of astrocytes but not oligodendrocytes or microglia accompanies sensorimotor impairment, seizure activity and early mortality at around 16 weeks of age. Our data reveal an unexpected role for connexins in the survival of white matter astrocytes, requiring the expression of particular isoforms in both oligodendrocytes and astrocytes.
Gap junction; connexin; myelin; astrocyte; oligodendrocytes; Cx47; Cx32; Cx43
Progenitors within the ventral telencephalon can generate GABAergic neurons and oligodendrocytes, but regulation of the neuron-glial switch is poorly understood. We investigated the combinatorial expression and function of Dlx1&2, Olig2, and Mash1 transcription factors in the ventral telencephalon. We show that Dlx homeobox transcription factors, required for GABAergic interneuron production, repress oligodendrocyte precursor cell (OPC) formation by acting on a common progenitor to determine neuronal versus oligodendroglial cell fate acquisition. We demonstrate that Dlx1&2 negatively regulate Olig2-dependant OPC formation and Mash1 promotes OPC formation by restricting the number of Dlx+ progenitors. Progenitors transplanted from Dlx1&2 mutant ventral telencephalon into newborn wild-type mice do not produce neurons but differentiate into myelinating oligodendrocytes that survive into adulthood. Our results identify a new role for Dlx genes as modulators of neuron versus oligodendrocyte development in the ventral embryonic forebrain.
Multiple sclerosis (MS) is a demyelinating disease in which blood-derived immune cells and activated microglia damage myelin in the central nervous system. While oligodendrocyte progenitor cells (OPCs) are essential for generating oligodendrocytes for myelin repair, other cell types also participate in the damage and repair processes. The NG2 proteoglycan is expressed by OPCs, pericytes, and macrophages/microglia. In this report we investigate the effects of NG2 on these cell types during spinal cord demyelination/remyelination.
Demyelinated lesions were created by microinjecting 1% lysolecithin into the lumbar spinal cord. Following demyelination, NG2 expression patterns in wild type mice were studied via immunostaining. Immunolabeling was also used in wild type and NG2 null mice to compare the extent of myelin damage, the kinetics of myelin repair, and the respective responses of OPCs, pericytes, and macrophages/microglia. Cell proliferation was quantified by studies of BrdU incorporation, and cytokine expression levels were evaluated using qRT-PCR.
The initial volume of spinal cord demyelination in wild type mice is twice as large as in NG2 null mice. However, over the ensuing 5 weeks there is a 6-fold improvement in myelination in wild type mice, versus only a 2-fold improvement in NG2 null mice. NG2 ablation also results in reduced numbers of each of the three affected cell types. BrdU incorporation studies reveal that reduced cell proliferation is an important factor underlying NG2-dependent decreases in each of the three key cell populations. In addition, NG2 ablation reduces macrophage/microglial cell migration and shifts cytokine expression from a pro-inflammatory to anti-inflammatory phenotype.
Loss of NG2 expression leads to decreased proliferation of OPCs, pericytes, and macrophages/microglia, reducing the abundance of all three cell types in demyelinated spinal cord lesions. As a result of these NG2-dependent changes, the course of demyelination and remyelination in NG2 null mice differs from that seen in wild type mice, with both myelin damage and repair being reduced in the NG2 null mouse. These studies identify NG2 as an important factor in regulating myelin processing, suggesting that therapeutic targeting of the proteoglycan might offer a means of manipulating cell behavior in demyelinating diseases.
Inflammation; myelin repair; NG2 ablation; oligodendrocyte progenitors; pericytes; macrophages