Pretreatment with magnesium sulfate attenuates white matter damage by preventing cell death of developing oligodendrocytes.

AIM: Antenatal maternal administration of magnesium sulfate (MgSO4 ) reduces cerebral palsy in preterm infants. However, it remains controversial as to whether it also reduces occurrence of white matter damage, or periventricular leukomalacia. We assessed the effect of MgSO4 against white matter damage induced by hypoxic-ischemic insult using a neonatal rat model and culture of premyelinating oligodendrocytes (pre-OL). METHODS: Rat pups at postnatal day (P) 6 were administered either MgSO4 or vehicle intraperitoneally before hypoxic-ischemic insult (unilateral ligation of the carotid artery followed by 6% oxygen for 1 h). The population of oligodendrocyte (OL) markers and CD-68-positive microglia at P11, and TdT-mediated biotin-16-dUTP nick-end labeling (TUNEL)-positive cells at P8 were evaluated in pericallosal white matter. Primary cultures of mouse pre-OL were subjected to oxygen glucose deprivation condition, and the lactate dehydrogenase release from culture cells was evaluated to assess cell viability. RESULTS: Pretreatment with MgSO4 attenuated the loss of OL markers, such as myelin basic protein and Olig2, in ipsilateral pericallosal white matter and decreased the number of CD-68-positive microglia and TUNEL-positive cells in vivo. Pretreatment with MgSO4 also inhibited lactate dehydrogenase release from pre-OL induced by oxygen glucose deprivation in vitro. CONCLUSION: Pretreatment with MgSO4 attenuates white matter damage by preventing cell death of pre-OL.

Effect of ninjin'yoeito, a Kampo (traditional Japanese) medicine, on cognitive impairment and depression in patients with Alzheimer's disease: 2 years of observation.

BACKGROUND: Only a few approved drugs are capable of alleviating the cognitive and behavioural symptoms of people living with Alzheimer's disease (AD). In recent years, however, the number of studies examining the clinical effects of herbal medicines on cognitive function in patients with AD has increased considerably. This study evaluated the long-term effects of a traditional Japanese medicine (Kampo medicine) known as ninjin'yoeito (NYT) on cognitive impairment and mood status in patients with AD over a 2-year period. METHODS: Twenty-three patients with mild-to-moderate probable AD according to the National Institute of Neurological and Communicative Disorders and Stroke and Alzheimer's Disease and Related Disorders Association criteria were included. Each participant had exhibited an insufficient response to treatment with donepezil alone before the start of the trial. Eleven patients received treatment with donepezil alone, and the remaining patients received a combined treatment of donepezil and NYT for 2 years. Patients were assessed by the Mini-Mental State Examination and the Alzheimer's Disease Assessment Scale-cognitive component-Japanese version for cognitive function, and the Neuropsychiatric Inventory was used to evaluate the patients' mood status at baseline and every 6 months for 2 years. RESULTS: The Mini-Mental State Examination results showed no significant differences between the two groups. Significant improvements were observed on the Alzheimer's Disease Assessment Scale-cognitive component-Japanese version and the Neuropsychiatric Inventory depression scores of patients who received the combined therapy with donepezil and NYT (Alzheimer's Disease Assessment Scale-cognitive component-Japanese version, 12 months: P < 0.01, 18 months: P = 0.04, 24 months: P < 0.01; Neuropsychiatric Inventory depression, 6 months: P < 0.05, 24 months: P < 0.05). CONCLUSIONS: A 2-year follow-up of patients receiving donepezil and NYT treatment showed an improved cognitive outcome and alleviation of AD-related depression.

Hypothermia attenuates apoptosis and protects contact between myelin basic protein-expressing oligodendroglial-lineage cells and neurons against hypoxia-ischemia.

Periventricular leukomalacia (PVL) is a major form of brain injury among preterm infants, which is characterized by extensive loss and dysfunction of premyelinating oligodendrocytes (pre-OLs) induced by hypoxia-ischemia (HI). Therapeutic hypothermia, which is a standard treatment for term infants with HI encephalopathy, is not indicated for preterm infants because its safety and effect have not been established. Here we investigate the effectiveness and mechanism of hypothermia for the inhibition of pre-OLs damage in PVL. For in vivo studies, 6-day-old rats underwent left carotid artery ligation, followed by exposure to 6% oxygen for 1 hr under hypothermic or normothermic conditions. The loss of myelin basic protein (MBP) was inhibited by hypothermia. For in vitro studies, primary pre-OLs cultures were subjected to oxygen-glucose deprivation (OGD) under normothermic or hypothermic conditions, and dorsal root ganglion neurons were subsequently added. Hypothermia inhibited apoptosis of pre-OLs, and, despite specific downregulation of 21.5- and 17-kDa MBP mRNA expression during hypothermia, recovery of the expression after OGD was superior compared with normothermia. OGD caused disarrangement of MBP distribution, decreased the levels of phosphorylated 21.5-kDa MBP, and disturbed the capacity to contact with neurons, all of which were restored by hypothermia. Pharmacological inhibition of ERK1/2 phosphorylation with U0126 during and after OGD significantly reduced the protective effects of hypothermia on apoptosis and myelination, respectively. These data suggest that phosphorylated exon 2-containing (21.5- and possibly 17-kDa) MBP isoforms may play critical roles in myelination and that hypothermia attenuates apoptosis and preserves the contact between OLs and neurons via ERK1/2 phosphorylation.

A DEAD-box RNA helicase Ddx54 protein in oligodendrocytes is indispensable for myelination in the central nervous system.

We recently reported that a new monoclonal antibody, 4F2, which labels oligodendroglial lineage cells, recognizes a DEAD-box RNA helicase Ddx54 and that Ddx54 binds to myelin basic protein (MBP) in brain and cultured oligodendrocytes. To elucidate the biological function of Ddx54, we generated a recombinant adenovirus, Ad-shRNA:Ddx54, expressing a short hairpin RNA to silence endogenous Ddx54 protein. The virus was intraventricularly injected into the brains of mice on postnatal day (PD) 2. The brains at PD 9 were then analyzed by immunohistochemistry. In untreated normal brain sections, as well as control brains that had been injected with Ad-ß-Gal, myelination of axons occurred in the corpus callosum with filamentous patterns of immunosignals of myelin-associated glycoprotein (MAG) and MBP. In Ad-shRNA:Ddx54-injected brain, substantial amounts of MAG and MBP immunosignals were present, but MBP immunosignals accumulated in the subplate layer and did not intrude into the emerging white matter. Immunoblot analysis revealed that Ddx54 knockdown caused a significant decrease in the level of 21.5 kDa MBP isoform and Ddx54, but the amount of Olig2; 2',3'-cyclic nucleotide 3' phosphodiesterase; MAG; three MBP isoforms (14, 17.5, and 18 kDa); and QKI-5, QKI-6, and QKI-7 proteins remained unchanged. Transfection of the Ddx54 expression vector into luciferase reporter-introduced neuroepithelial cells resulted in upregulated MBP promoter activity. Immunoprecipitation of Ddx54 protein in MBP-transfected HEK293 cells indicated that Ddx54 may directly interact with MBP mRNA. These results suggest that Ddx54 protein play an important role in central nervous system myelination, presumably in myelin sheath formation after the differentiation of oligodendrocytes.

An anti-sulfatide antibody O4 immunoprecipitates sulfatide rafts including Fyn, Lyn and the G protein α subunit in rat primary immature oligodendrocytes.

The association of sulfatide with specific proteins in oligodendrocytes was examined by co-immunoprecipitation with an anti-sulfatide antibody. Protein kinase activity was detected in precipitates with a monoclonal antibody to sulfatide (O4) from the rat primary immature oligodendrocytes. We conducted in vitro kinase assay of tyrosine phosphorylated proteins of 80, 59, 56, 53 and 40 kDa by gel electrophoresis. Of these proteins, the proteins of 59 kDa and 53/56 kDa were identified as the Src family tyrosine kinases Fyn and Lyn on the basis of their sequential immunoprecipitation with anti-Fyn and anti-Lyn antibodies, respectively. The 40 kDa protein was identified as the α subunit of the heterotrimeric G protein. These observations suggest that O4 immunoprecipitates sulfatide rafts including Fyn, Lyn and the α subunit of the heterotrimeric G protein.

Lethal adulthood myelin breakdown by oligodendrocyte-specific Ddx54 knockout.

Multiple sclerosis (MS) is a leading disease that causes disability in young adults. We have previously shown that a DEAD-box RNA helicase Ddx54 binds to mRNA and protein isoforms of myelin basic protein (MBP) and that Ddx54 siRNA blocking abrogates oligodendrocyte migration and myelination. Herein, we show that MBP-driven Ddx54 knockout mice (Ddx54 fl/fl;MBP-Cre), after the completion of normal postnatal myelination, gradually develop abnormalities in behavioral profiles and learning ability, inner myelin sheath breakdown, loss of myelinated axons, apoptosis of oligodendrocytes, astrocyte and microglia activation, and they die within 7 months but show minimal peripheral immune cell infiltration. Myelin in Ddx54fl/fl;MBP-Cre is highly vulnerable to the neurotoxicant cuprizone and Ddx54 knockdown greatly impairs myelination in vitro. Ddx54 expression in oligodendrocyte-lineage cells decreased in corpus callosum of MS patients. Our results demonstrate that Ddx54 is indispensable for myelin homeostasis, and they provide a demyelinating disease model based on intrinsic disintegration of adult myelin.

A new monoclonal antibody, 4F2, specific for the oligodendroglial cell lineage, recognizes ATP-dependent RNA helicase Ddx54: possible association with myelin basic protein.

Recent research in neural development has highlighted the importance of markers to discriminate phenotypic alterations of neural cells at various developmental stages. We isolated a new monoclonal antibody, 4F2, which was shown to be specific for an oligodendrocyte lineage. In primary cultures of oligodendroglial and mixed neural cells, the 4F2 antibody labeled a large proportion of Sox2(+) , Sox10(+) , A2B5(+) , NG2(+) , Olig2(+) , O4(+) , and myelin basic protein (MBP)(+) cells but did not label any GFAP(+) or NeuN(+) cells. In immunohistochemisty of rat embryos, the 4F2 antibody labeled a portion of neuroepithelial cells of the neural tube at embryonic day 9. The 4F2-positive cells were located initially in the ventricular zone as Musashi1(+) Tuj1(-) populations and distributed throughout the striatum; thereafter, they populated the whole brain and spinal cord. These cells showed ramified processes during embryonal development. The 4F2 antigen was associated with all four isoforms of MBP in coimmunoprecipitation experiments using brain homogenates or cell lysates of cultured oligodendrocytes. Immunoscreening of a brain cDNA library identified the antigen as DEAD (Asp-Glu-Ala-Asp) box polypeptide 54 (Ddx54), a member of the DEAD box family of RNA helicases involved in RNA metabolism, transcription, and translation. Cotransfection of the Ddx54 gene with MBP isoform genes increased the nuclear localization of the 21.5-kDa MBP isoform, which has been reported to function as a nuclear signal transduction molecule. These data indicate that Ddx54 might be not only a useful marker for investigating the ontogeny of oligodendrocytes but also an important factor in oligodendrocyte differentiation and myelination.

Administration of chinpi, a component of the herbal medicine ninjin-youei-to, reverses age-induced demyelination.

The disruption of myelin causes severe neurological diseases. An understanding of the mechanism of myelination and remyelination is essential for the development of therapeutic strategies for demyelination diseases. Our previous findings indicated that the FcRγ/Fyn cascade is a potential therapeutic target for remyelination caused by the Chinese/Japanese traditional herbal (Kampo) medicine ninjin'youeito (Ninjin-youei-to, NYT), which is a hot-water extract made from 12 medicinal herbs. To identify which constituents of NYT are involved in the reversal of demyelination and to examine the potential therapeutic effect, we tested several of the chemical constituents of NYT. Here, we report that Chinpi, a constituent of NYT, upregulates the FcRγ/Fyn signaling cascade resulting in a potentially therapeutic effect against age-induced demyelination. In addition, we observed that phosphorylated (activated) FcRγ/Fyn upregulated the expression of the 21.5 kDa isoform of myelin basic protein, inducing rapid morphological differentiation, when oligodendrocyte precursor cells (OPCs) were cultured in the presence of hesperidin and/or narirutin (the major active constituents of Chinpi). These results suggest that hesperidin and narirutin participate in the FcRγ/Fyn signaling pathway in OPCs causing these cells to differentiate into myelinating oligodendrocytes.

The Absence of Myelin Basic Protein Reduces Non-Amyloidogenic Processing of Amyloid Precursor Protein.

BACKGROUND: The accumulation of amyloid ß-protein (Aß) in the brain is a pathological feature of Alzheimer's disease (AD). Aß peptides originate from amyloid precursor protein (APP). APP can be proteolytically cleaved through amyloidogenic or non-amyloidogenic pathways. The molecular effects on APP metabolism/processing may be influenced by myelin and the breakdown of myelin basic protein (MBP) in AD patients and mouse models of AD pathology. METHODS: We directly tested whether MBP can alter influence APP processing in MBP-/- mice, known as Shiverer (shi/shi) mice, in which no functional MBP is produced due to gene breakage from the middle of MBP exon ll. RESULTS: A significant reduction of the cerebral sAPPα level in Shiverer (shi/shi) mice was found, although the levels of both total APP and sAPPß remain unchanged. The reduction of sAPPα was considered to be due to the changes in the expression levels of a disintegrin and metalloproteinase-9 (ADAM9) catalysis and non-amyloid genic processing of APP in the absence of MBP because it binds to ADAM9. MBP -/- mice exhibited increased Aß oligomer production. CONCLUSION: These findings suggest that in the absence of MBP, there is a marked reduction of nonamyloidogenic APP processing to sAPPα, and targeting myelin of oligodendrocytes may be a novel therapy for the prevention and treatment of AD.

Caspase-9 activation revealed by semaphorin 7A cleavage is independent of apoptosis in the aged olfactory bulb.

Caspases are essential in multicellular organisms for inducing cell death during normal development and in the immune system. However, caspases can also trigger the degenerative process under certain conditions such as pathophysiological conditions and aging. Here, we identified Semaphorin 7A (Sema7A) as a novel substrate for caspase-9 that can be used to monitor caspase-9 activity in mice, and found nonapoptotic caspase-9 activation in the aged olfactory bulb (OB). Immunostaining of the OB for the caspase-9-cleaved form of Sema7A revealed abundant caspase-9-activated cells in 2-year-old (aged) but not in 2-month-old (young) mice. In fact, various regions of the aged brain, including the OB, exhibited an increased level of caspase-9 activity. However, the number of dying cells in the aged OB was, intriguingly, much lower (<20%) than in the OB of young mice. Furthermore, we found that the lower number dying cells in the aged OB was accompanied by a decreased expression of procaspase-3. These results suggest a survival strategy for aged OB neurons, which can no longer regenerate, in which the central apoptotic machinery downstream of caspase-9 is inactivated.

Hypothermia-induced increase of oligodendrocyte precursor cells: Possible involvement of plasmalemmal voltage-dependent anion channel 1.

Hypothermia is believed to suppress cell proliferation by inducing apoptosis/necrosis and phase-specific/nonspecific cell cycle arrest, which are, directly or indirectly, related to a reduced energy supply. Intriguingly, hypothermia is known to improve neurological recovery of animals and humans exposed to focal brain hypoxic-ischemic injury. The underlying mechanism of the neuroprotective effect of hypothermia is unclear, although the prevention of neural cell apoptosis is thought to play a role. Herein we demonstrate that in vitro cell culture of oligodendrocyte precursor cells (OPCs) under conditions of mild hypothermia (31.5°C) results in an increase in cell number relative to cells cultured under normothermic conditions (37°C). Cell cycle analysis, immunoblotting of cyclins, TUNEL assay, and immunocytochemistry of OPC differentiation markers suggest that hypothermia shifts the balance between proliferation and apoptosis/differentiation toward proliferation. A combination of transcriptome analysis, pharmacological intervention, and immunoaffinity-based assays suggests a possible involvement of the Gα13-Rho GTPase Cdc42-ERK1/2 signaling cascade and voltage-dependent anion channel 1 (VDAC1), which associate or dissociate with Gα13 protein at 37°C and 31.5°C, respectively. Immunoelectron microscopy revealed the presence of VDAC1 in the plasma membrane of OPCs. Furthermore, the exogenous addition of impermeable VDAC1 inhibitors enhanced proliferation of OPCs at 37°C. These results may contribute to the elucidation of the mechanism of hypothermic neuroprotection as well as the possible novel role of plasmalemmal VDAC1.

Signaling via immunoglobulin Fc receptors induces oligodendrocyte precursor cell differentiation.

Dramatic changes in morphology and myelin protein expression take place during the differentiation of oligodendrocyte precursor cells (OPCs) into myelinating oligodendrocytes. Fyn tyrosine kinase was reported to play a central role in the differentiation process. Molecules that could induce Fyn signaling have not been studied. Such molecules are promising therapeutic targets in demyelinating diseases. We provide evidence that the common gamma chain of immunoglobulin Fc receptors (FcRgamma) is expressed in OPCs and has a role in triggering Fyn signaling. FcRgamma cross-linking by immunoglobulin G on OPCs promotes the activation of Fyn signaling and induces rapid morphological differentiation with upregulation of myelin basic protein (MBP) expression levels. Mice deficient in FcRgamma are hypomyelinated, and a significant reduction in MBP content is evident. Our findings indicate that the FcRgamma-Fyn-MBP cascade is pivotal during the differentiation of OPCs into myelinating oligodendrocytes, revealing an unexpected involvement of immunological molecules.

A new monoclonal antibody, A3B10, specific for astrocyte-lineage cells recognizes calmodulin-regulated spectrin-associated protein 1 (Camsap1).

Recent studies of adult neurogenesis of the mammalian central nervous system have suggested unexpected plasticity and complexity of neural cell ontogenesis. Redefinition and reconstitution of cell classification and lineage relationships, especially between glial and neural precursors, are an urgent and crucial concern. In the present study, we describe a new monoclonal antibody, A3B10, which was produced by immunizing mice with the membrane fraction prepared from astrocyte-enriched primary neural cell cultures. Immunohistochemistry of brain sections, including brains from glial fibrillary acidic protein (GFAP)-deficient mice and primary mixed neural cell cultures, as well as immunoblot analysis and immunoelectron microscopy, have revealed that 1) A3B10 recognizes a majority of cells in ependyma in neonatal and adult rats, 2) A3B10 stains almost all GFAP(+) cells and some S100beta(+) cells in the corpus callosum, 3) A3B10 specifically stains astrocytes in vitro in primary cultures of rat embryonic cerebral hemispheres, 4) A3B10 equally stains ependymal cells of wild-type and GFAP-deficient mice, and 5) A3B10 antigen might construct intermediate filament bundles with GFAP and/or vimentin. These data suggested that the antibody labels a wide array of astorcytic-lineage cells including astrocytes, astrocyte precursors, and neural stem cells. Screening a cDNA library derived from rat embryonic brain has revealed that the antibody recognizes calmodulin-regulated spectrin-associated protein 1 (Camsap1). Thus this antibody may provide not only a new marker to identify astrocyte-lineage cells but also a new target molecule to elucidate the ontogeny, development, and pathophysiological functions of astrocyte-lineage cells.

Osteopetrosis and thalamic hypomyelinosis with synaptic degeneration in DAP12-deficient mice.

Deletions in the DAP12 gene in humans result in Nasu-Hakola disease, characterized by a combination of bone fractures and psychotic symptoms similar to schizophrenia, rapidly progressing to presenile dementia. However, it is not known why these disorders develop upon deficiency in DAP12, an immunoreceptor signal activator protein initially identified in the immune system. Here we show that DAP12-deficient (DAP12(-/-)) mice develop an increased bone mass (osteopetrosis) and a reduction of myelin (hypomyelinosis) accentuated in the thalamus. In vitro osteoclast induction from DAP12(-/-) bone marrow cells yielded immature cells with attenuated bone resorption activity. Moreover, immature oligodendrocytes were arrested in the vicinity of the thalamus, suggesting that the primary defects in DAP12(-/-) mice are the developmental arrest of osteoclasts and oligodendrocytes. In addition, the mutant mice also showed synaptic degeneration, impaired prepulse inhibition, which is commonly observed in several neuropsychiatric diseases in humans including schizophrenia, and aberrant electrophysiological profiles in the thalami. These results provide a molecular basis for a unique combination of skeletal and psychotic characteristics of Nasu-Hakola disease as well as for schizophrenia and presenile dementia.

Identification of activated transcription factors from microarray gene expression data of Kampo medicine-treated mice.

We propose an approach to identify activated transcription factors from gene expression data using a statistical test. Applying the method, we can obtain a synoptic map of transcription factor activities which helps us to easily grasp the system's behavior. As a real data analysis, we use a case-control experiment data of mice treated by a drug of Kampo medicine remedying degraded myelin sheath of nerves in central nervous system. Kampo medicine is Japanese traditional herbal medicine. Since the drug is not a single chemical compound but extracts of multiple medicinal herb, the effector sites are possibly multiple. Thus it is hard to understand the action mechanism and the system's behavior by investigating only few highly expressed individual genes. Our method gives summary for the system's behavior with various functional annotations, e.g. TFAs and gene ontology, and thus offer clues to understand it in more holistic manner.

An Extract of Chinpi, the Dried Peel of the Citrus Fruit Unshiu, Enhances Axonal Remyelination via Promoting the Proliferation of Oligodendrocyte Progenitor Cells.

The aging-induced decrease in axonal myelination/remyelination is due to impaired recruitment and differentiation of oligodendrocyte progenitor cells (OPCs). Our previous studies have shown that a monoclonal antibody to DEAD (Asp-Glu-Ala-Asp) box polypeptide 54 (Ddx54), a member of the DEAD box family of RNA helicases, (1) specifically labels oligodendrocyte lineages, (2) binds to mRNA and protein isoforms of myelin basic proteins (MBP), and (3) regulates migration of OPCs from ventricular zone to corpus callosum in mice. It has also been demonstrated that specific loss of a 21.5 kDa MBP isoform (MBP21.5) reflects demyelination status, and oral administration of an extract of Chinpi, citrus unshiu peel, reversed the aging-induced demyelination. Here, we report that Chinpi treatment induced a specific increase in the MBP21.5, led to the reappearance of Ddx54-expressing cells in ventricular-subventricular zone and corpus callosum of aged mice, and promoted remyelination. Treatment of in vitro OPC cultures with Chinpi constituents, hesperidin plus narirutin, led to an increase in 5-bromo-2'-deoxyuridine incorporation in Ddx54-expressing OPCs, but not in NG2- or Olig2-expressing cell populations. The present study suggests that Ddx54 plays crucial role in remyelination. Furthermore, Chinpi and Chinpi-containing herbal medicines may be a therapeutic option for the aging-induced demyelination diseases.

Structure and biological function of ENPP6, a choline-specific glycerophosphodiester-phosphodiesterase.

Choline is an essential nutrient for all living cells and is produced extracellularly by sequential degradation of phosphatidylcholine (PC). However, little is known about how choline is produced extracellularly. Here, we report that ENPP6, a choline-specific phosphodiesterase, hydrolyzes glycerophosphocholine (GPC), a degradation product of PC, as a physiological substrate and participates in choline metabolism. ENPP6 is highly expressed in liver sinusoidal endothelial cells and developing oligodendrocytes, which actively incorporate choline and synthesize PC. ENPP6-deficient mice exhibited fatty liver and hypomyelination, well known choline-deficient phenotypes. The choline moiety of GPC was incorporated into PC in an ENPP6-dependent manner both in vivo and in vitro. The crystal structure of ENPP6 in complex with phosphocholine revealed that the choline moiety of the phosphocholine is recognized by a choline-binding pocket formed by conserved aromatic and acidic residues. The present study provides the molecular basis for ENPP6-mediated choline metabolism at atomic, cellular and tissue levels.

Advancement of differentiation of oligodendrocyte progenitor cells by a cascade including protein kinase A and cyclic AMP-response element binding protein.

A transcription factor, cyclic AMP-response element binding protein (CREB), which is phosphorylated by protein kinases (PKA and PKC), is known to be involved in the regulation of oligodendrocyte differentiation. However, it is still unclear whether protein kinase A (PKA) and protein kinase C (PKC) are used simultaneously or at different time points to phosphorylate CREB in oligodendrocytes and whether CREB phosphorylation advances oligodendrocyte differentiation or vise versa. Our previous experiments have shown that in the differentiation process from immature to mature cells, CREB phosphorylation depends on PKC activity and leads to the progression of differentiation. In order to gain a better understanding of the process of differentiation from progenitor to immature cells, we identified which protein kinase, i.e., PKA or PKC, regulates CREB phosphorylation and we determined whether CREB phosphorylation advances differentiation or the reverse. Our results showed that CREB phosphorylation is principally regulated by PKA activity in progenitor cells but not by PKC activity, and that this phosphorylation advances the differentiation of progenitor cells to immature cells in oligodendrocytes.

Specific expression of an HNK-1 carbohydrate epitope and NCAM on femoral nerve Schwann cells in mice.

Schwann cells are glial cells of the peripheral nervous system. There are two known subtypes of Schwann cells: those that are myelin-forming; and those that are non-myelin-forming. In this study, we looked at the expression of cell adhesion molecules in Schwann cells to determine whether other subtypes might exist. We used immunohistological analysis of femoral nerve segments containing sensory and motor fascicles, stained with anti-HNK-1, M6749 and anti-neural cell adhesion molecule (NCAM) monoclonal antibodies. Anti-HNK-1 and M6749 were positive in the motor fascicle, while anti-NCAM was positive in the sensory fascicle. Immunoblot analysis with the anti-HNK-1 and M6749 antibodies showed stronger immunoreactivity in the motor fraction than in the sensory fraction in the 100 kDa band. With the anti-NCAM antibody, the 140 and 120 kDa bands were seen in the sensory fascicle fraction, but not in the motor fascicle fraction. HNK-1-positive-cells were seen in motor fascicles 7 days after transection. However, the level of immunoreactivity diminished at 14 days, and no immunoreactivity was seen at 21 days. NCAM-positive cells were not observed 3 days after transection. In development, HNK-1-positive-cells and NCAM-positive cells were seen after P-21. These results suggest that the Schwann cells from the motor and the sensory fascicles have different subtypes. The motor and sensory Schwann cells may play different roles and function in a different way during peripheral nerve regeneration. In addition, there could be more stages of Schwann cell differentiation than previously thought; it is possible that myelin-forming Schwann cells differentiate into HNK-1-positive-cells (motor myelin-forming Schwann cells) and HNK-1-negative-cells (sensory myelin-forming Schwann cells), and non-myelin-forming Schwann cells differentiate into NCAM-positive cells (sensory non-myelin-forming Schwann cells) and NCAM-negative cells (autonomic non-myelin-forming Schwann cells).

Involvement of CD45 in central nervous system myelination.

Myelin is a multi-layered membranous lipid insulator surrounding axons that allows the rapid conduction of neuronal impulses. In the central nervous system (CNS), myelin is produced by oligodendrocytes. During development, morphologically immature oligodendrocyte precursor cells (OPCs) arise from neural stem cells before differentiating into myelinating oligodendrocytes shortly after birth. Fyn tyrosine kinase (Fyn) has been shown to play a central role during OPC differentiation, including inducing morphological changes in the cells and initiating the expression of myelin basic protein (MBP), a major structural protein required for the compaction of myelin sheaths. Recently, we have shown that signaling via the gamma chain of immunoglobulin Fc receptors (FcRgamma) induces the Fyn-MBP cascade and promotes the morphological differentiation of OPCs. The protein tyrosine phosphatases that are responsible for the positive regulation of Fyn tyrosine kinase activity during this cascade, however, remained unknown. Here we report that a protein tyrosine phosphatase, CD45, is involved in this process. Fyn co-immunoprecipitated with CD45 from differentiating wild-type OPCs in vitro, while CD45-deficient OPCs failed to differentiate. Additionally, dysmyelination was observed in CD45-deficient mice in vivo. Our findings suggest that CD45 is a key phosphatase involved in OPC differentiation and provide a preliminary explanation for the previously reported CD45 mutations observed in some multiple sclerosis (MS) patients.