myelin-basic-protein and Demyelinating-Diseases

Rapid nerve conduction requires the coating of axons by a tightly packed multilayered myelin membrane. In the central nervous system, myelin is formed from cellular processes that extend from oligodendrocytes and wrap in a spiral fashion around an axon, resulting in the close apposition of adjacent myelin membrane bilayers. In this review, we discuss the physical principles underlying the zippering of the plasma membrane of oligodendrocytes at the cytoplasmic and extracellular leaflet. We propose that the interaction of the myelin basic protein with the cytoplasmic leaflet of the myelin bilayer triggers its polymerization into a fibrous network that drives membrane zippering and protein extrusion. In contrast, the adhesion of the extracellular surfaces of myelin requires the down-regulation of repulsive components of the glycocalyx, in order to uncover weak and unspecific attractive forces that bring the extracellular surfaces into close contact. Unveiling the mechanisms of myelin membrane assembly at the cytoplasmic and extracelluar sites may help to understand how the myelin bilayers are disrupted and destabilized in the different demyelinating diseases.

Topics: Amino Acid Sequence; Cell Membrane; Demyelinating Diseases; Models, Biological; Molecular Sequence Data; Myelin Basic Protein; Myelin Proteolipid Protein; Myelin Sheath; Oligodendroglia; Sequence Alignment

Multiple sclerosis (MS) is a disease of the human CNS, characterized by perivascular inflammation, demyelination and axonal damage. Although the etiology of MS is unknown, it is believed that the disease results from destructive autoimmune mechanisms, presumably initiated by abnormal activation of potentially pathogenic autoimmune T-cells recognizing CNS components. The myelin-associated oligodendrocyte basic protein (MOBP), a relatively abundant CNS-specific myelin protein, which plays a role in stabilizing the myelin sheath in the CNS, has recently been implicated in the pathogenesis of MS. Here we review studies showing that MOBP is as an important candidate target antigen in MS as the other widely studied target antigens, myelin basic protein (MBP), proteolipid protein (PLP), and myelin oligodendrocyte glycoprotein (MOG). The studies summarized below indicate that T-cell autoimmunity against MOBP can be detected in MS patients; T-cells reactive against MOBP can be pathogenic in several mouse strains as well as in the "humanized" HLA-DR15-Tg mice; and, that the HLA-DQ6-restricted, but not HLA-DR15-restricted, MOBP-reactive T-cells cause in HLA-DR15-Tg mice MS-like clinical disease associated with perivascular and parenchymal infiltration, demyelination, axonal loss, and optic neuritis. Accordingly, the MOBP should be considered a bona fide primary target antigen in MS, in addition to MBP, PLP, and MOG.

Topics: Animals; Autoantigens; Demyelinating Diseases; Disease Models, Animal; HLA-DR Antigens; HLA-DR Serological Subtypes; Lymphocyte Activation; Mice; Mice, Transgenic; Multiple Sclerosis; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Optic Neuritis; T-Lymphocytes

Myelin basic protein (MBP) binds to negatively charged lipids on the cytosolic surface of oligodendrocyte membranes and is responsible for adhesion of these surfaces in the multilayered myelin sheath. The pattern of extensive posttranslational modifications of MBP is dynamic during normal central nervous system development and during myelin degeneration in multiple sclerosis (MS), affecting its interactions with the myelin membranes and other proteins. In particular, the degree of deimination (or citrullination) of MBP is correlated with the severity of MS, and may represent a primary defect that precedes neurodegeneration due to autoimmune attack. That MBP deimination also affects topological accessibility of an otherwise partially buried immunodominant epitope of the protein indicates that this modification may play a major role in the autoimmune pathogenesis of the disease. In this chapter, we describe the structural and functional consequences of MBP deimination in healthy and diseased myelin.

Topics: Animals; Autoimmunity; Demyelinating Diseases; Humans; Immunodominant Epitopes; Multiple Sclerosis; Myelin Basic Protein; Myelin Sheath; Protein Conformation; Protein Processing, Post-Translational

Topics: Animals; Cell Differentiation; Demyelinating Diseases; Disease Models, Animal; Gene Expression Regulation, Developmental; Mice; Mice, Neurologic Mutants; Mutation; Myelin Basic Protein; Myelin Sheath; Nerve Tissue Proteins; Oligodendroglia; Transcription Factors

Early recognition of whether a product has potential as a new therapy for treating multiple sclerosis (MS) relies upon the quality of the animal models used in the preclinical trials. The promising effects of new treatments in rodent models of experimental autoimmune encephalomyelitis (EAE) have rarely been reproduced in patients suffering from MS. EAE in outbred marmoset monkeys, Callithrix jacchus, is a valid new model, and might provide an experimental link between EAE in rodent models and human MS. Using magnetic resonance imaging techniques similar to those used in patients suffering from MS pathological abnormalities in the brain, white matter of the animal can be visualized and quantified. Moreover, NMR spectroscopy, in combination with pattern recognition, offers an advanced uroscopic technique for the identification of biomarkers of inflammatory demyelination.

Topics: Animals; Animals, Outbred Strains; Antigens, CD; Biomarkers; Brain; Callithrix; CD4-Positive T-Lymphocytes; Chronic Disease; Demyelinating Diseases; Disease Models, Animal; Encephalomyelitis, Autoimmune, Experimental; Genes, MHC Class II; Humans; Magnetic Resonance Imaging; Magnetic Resonance Spectroscopy; Mice; Multiple Sclerosis; Myelin Basic Protein; Myelin Proteins; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Radiography; T-Lymphocytes, Cytotoxic; Th2 Cells

We report here an infant with 18q deletion syndrome, and intractable apneic seizures. He had intrauterine growth retardation and dysmorphic features. Chromosomal analysis demonstrated mosaicism of 18q interstitial deletion (q12.3-q22.3). From the age of 3 months, apneic attacks occurred from once a week to over 10 times a day despite many oral antiepileptic agents, and were diagnosed as complex partial seizures. Ictal electroencephalogram and 18F-fluorodeoxyglucose-positron emission tomography at the age of 10 months identified the epileptic focus in the right parieto-temporal region. He also had severe psychomotor retardation. Head MRI examination revealed diffuse cerebral atrophy and severe white matter dysmyelination, which was caused by the deletion of myelin basic protein gene at the locus of 18q22.3. This locus may be responsible for the clinical manifestations of 18q deletion syndrome. Detailed description of the onset, seizure types, and prognosis of epilepsy associated with 18q deletion syndrome is rare. It was suggested that the locus of 18q21.3-q22.3 was responsible for autonomic seizures in 18q deletion syndrome.

Topics: Apnea; Brain Diseases; Chromosome Deletion; Chromosomes, Human, Pair 18; Demyelinating Diseases; Epilepsy, Complex Partial; Gene Deletion; Humans; Infant; Infant, Newborn; Male; Myelin Basic Protein; Syndrome

These results support a model of epitope spreading (Figure 4) wherein localized virus-specific T cell-mediated inflammatory processes lead to the recruitment/activation of CNS-resident APCs which can serve both as effector cells for myelin destruction and as APCs which efficiently process/present endogenous self epitopes to autoreactive T cells. Thus, inflammatory responses induced by viruses which trigger pro-inflammatory Th1 responses and have the ability to persist in genetically susceptible hosts, may lead to chronic organ-specific autoimmune disease via epitope spreading. Regardless of the specificity of the T cells (myelin peptides in R-EAE or TMEV epitopes in TMEV-IDD) responsible for initiating myelin destruction, epitope spreading plays an important contributory role in the chronic disease process in genetically susceptible SJL mice. Epitope spreading has obvious important implications to the design of antigen-specific therapies for the potential treatment of MS and other autoimmune diseases. This process indicates that autoimmune diseases are evolving pathologies and that the specificity of the effector autoantigen-specific T cells varies during the chronic disease process.

Topics: Animals; Autoimmune Diseases; Demyelinating Diseases; Disease Models, Animal; Encephalomyelitis, Autoimmune, Experimental; Epitopes; Humans; Mice; Multiple Sclerosis; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Theilovirus

Inborn errors of the single-carbon transfer pathway are rare disorders of folate and cobalamin metabolism. They may be complicated by demyelination resembling subacute combined degeneration of the cord and brain. The study of CSF metabolites in children with serial errors affecting the single-carbon transfer pathway has suggested that S-adenosylmethionine deficiency is a cause of the demyelination. This deficiency is corrected by treatment that causes clinical improvement and remyelination. Some treatments can only have an indirect effect on the brain and this is discussed with other evidence that the liver may produce factors that are necessary for the maintenance of central myelin.

Topics: Demyelinating Diseases; Humans; Liver; Metabolism, Inborn Errors; Methionine Adenosyltransferase; Methylation; Myelin Basic Protein; One-Carbon Group Transferases; S-Adenosylmethionine

Topics: Animals; Antigen-Presenting Cells; Autoantigens; Autoimmunity; CD4-Positive T-Lymphocytes; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Gene Rearrangement; Histocompatibility Antigens Class II; Humans; Lymphocyte Activation; Macrophage Activation; Mice; Microglia; Models, Biological; Multiple Sclerosis; Myelin Basic Protein; Receptors, Antigen, T-Cell, alpha-beta; T-Lymphocytes

A previously healthy 19-month-old boy developed acute encephalopathy, thrombocytopenia and hepatic dysfunction. Human herpesvirus-6 (HHV-6) DNA was found in his CSF during the acute stage of the disease by means of the polymerase chain reaction. T2-weighted MRI revealed high signal intensity in the left thalamus and left parieto-occipital deep white matter. The myelin basic protein concentration in the CSF was elevated suggesting acute demyelination. The patient is now 2.5 years old and has no sequelae.. Since clinical course and neuroimaging after HHV-6 infection are similar to those in acute disseminated encephalomyelitis, clinicians must pay attention to primary HHV-6 infection in patients under 2 years old with white matter lesions.

Topics: Acute Disease; Brain; Brain Diseases; Child, Preschool; Demyelinating Diseases; DNA, Viral; Dominance, Cerebral; Encephalitis, Viral; Follow-Up Studies; Herpesviridae Infections; Herpesvirus 6, Human; Humans; Infant; Liver Function Tests; Magnetic Resonance Imaging; Male; Myelin Basic Protein

The cause of multiple sclerosis (MS) is unknown, but the pathology is consistent with an immunological etiology. Studies conducted with the animal model of MS, experimental autoimmune encephalomyelitis (EAE), have provided insight into how the immune system can provoke an immunopathological response characteristic of that seen in MS. The use of inbred rats and mice for studies of EAE has been especially rewarding with respect to the identification of the epitopes of encephalitogenic antigens responsible for the induction of this autoimmune disease and in elucidating the effector mechanism underlying EAE. Moreover, it has also been possible to ascertain how EAE can be regulated, leading to therapeutic modalities which have been applied in clinical studies of MS patients. This review briefly summarizes studies of EAE in rodents, drawing comparisons with immunological findings reported in patients with MS. It is clear that important lessons can be learned from the detailed investigation of animal models that may be applicable to human immunological disorders.

Topics: Amino Acid Sequence; Animals; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Humans; Mice; Molecular Sequence Data; Myelin Basic Protein; Myelin Proteolipid Protein; Peptides; Rats

Multiple sclerosis is a chronic inflammatory disease characterized by multifocal damage of the central nervous system myelin. Both humoral and cell-mediated immune abnormalities have been observed in patients with multiple sclerosis, but their relation to the demyelination process is not understood. The etiology of the disease is still unknown; however, evidence exists for an interplay between environmental and genetic factors. Several genes are involved in determining the disease susceptibility, at least one of them encoded within human leukocyte antigen gene complex. Other genomic regions coding for components of the immune system or myelin have also been suggested. Clinical, immunological and genetic data suggest that multiple sclerosis may turn out to be a heterogeneous disease. Therefore, molecular genetic dissection of this complex disease should provide important clues to its pathogenesis as well as unravel metabolic pathways for potential therapeutic or preventive strategies. This review will give an overview of recent progress and future challenges in identifying susceptibility genes for multiple sclerosis.

Topics: Adult; Animals; Demyelinating Diseases; Disease Models, Animal; Family Health; Female; Genes, Immunoglobulin; Genetic Linkage; Genetic Predisposition to Disease; HLA Antigens; Humans; Lod Score; Male; Multiple Sclerosis; Myelin Basic Protein; Polymorphism, Genetic; Receptors, Antigen, T-Cell

The myelin sheath is an extension of a plasma membrane tightly wrapped around axons. It facilitates conduction while conserving space and energy. Myelin is characterized by a high lipid content (80% of dry weight). Most myelin proteins are unique to that structure and some of them are restricted to the central or peripheral nervous system. In this review a few examples of inherited metabolic disorders affecting the oligodendrocyte and/or the Schwann cells are presented. Emphasis is placed on mutations in animals that represent invaluable models for investigating the molecular mechanisms of inherited myelin diseases in humans.

Topics: Animals; Demyelinating Diseases; Humans; Lipids; Mutation; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin Sheath

Primary demyelination in the central nervous system results from damage to the myelin sheath or oligodendroglia and can be produced by a variety of mechanisms, including metabolic disturbances, toxicities, infection, and autoimmunity. The major human demyelinating disease affecting the central nervous system is multiple sclerosis (MS). Although the etiology of MS is not known, existing data indicate that both genetic and environmental factors contribute to pathogenesis. Experimental allergic encephalomyelitis (EAE) is induced by immunization of genetically susceptible animals with myelin proteins. This is mediated by autoimmune T cells. Characterization of MHC restriction, fine specificity of antigen recognition, and T cell receptor (TCR) usage by encephalitogenic T cells has resulted in highly specific immunotherapies. Both HLA and TCR genes have been linked to susceptibility for MS which is widely believed to be mediated by T cells that recognize an as yet unidentified autoantigen. Because of the advances in the understanding and treatment of EAE, recent research in MS has been focused on the characterization of cellular immune responses against myelin components. The results of these studies are reviewed and the potential implications of these findings for the pathogenesis and future therapy of MS are examined.

Topics: Animals; Autoantigens; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Humans; Multiple Sclerosis; Myelin Basic Protein; T-Lymphocytes

The genomic organization of the T-cell receptor (TCR) gene complexes accounts for many central aspects of T-cell immunobiology, including specificity and diversity. Recent data indicate that polymorphism of TCR genes is present within a species and may influence the immune phenotype of an individual. Such polymorphism has been detected by RFLP, by the presence of large regions of insertion or deletion of germline DNA, and by allelic variability of individual gene segments that are expressed. In addition to allelic variation of TCR genes, immune responses may also be influenced by the repertoire of the TCR molecules that are expressed by responding T-cell populations. In some situations, pathogenic T-cell responses may involve expression of limited numbers of TCR gene families. This is true, for example, in experimental allergic encephalomyelitis, an autoimmune nervous system disease mediated by T-cells reactive to myelin basic protein. In the human disease counterpart, multiple sclerosis, a more complex pattern of T-cell recognition to the putative autoantigen is generally present, although in some individuals a restricted response may occur. Specific therapies targeted to certain TCR molecules represents a promising approach to chronic inflammatory diseases in humans. The efficacy of such therapies will be determined in part by the TCR repertoire expressed in individual disease situations and by the potential for plasticity in the pathogenic T-cell response that may exist.

Topics: Alleles; Animals; Autoimmune Diseases; Chromosomes, Human, Pair 14; Chromosomes, Human, Pair 7; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Gene Rearrangement, T-Lymphocyte; Humans; Mice; Multiple Sclerosis; Myelin Basic Protein; Polymorphism, Genetic; Polymorphism, Restriction Fragment Length; Prevalence; Receptors, Antigen, T-Cell

The mechanism by which the myelin sheath is degraded in demyelinating diseases is unknown. The demonstration of increased activities of both acid (cathepsins B, D, A) and neutral proteinases in tissue from experimental allergic encephalomyelitis (EAE) in animals and multiple sclerosis (MS, plaques) and the disappearance of myelin proteins implicate a role for proteolytic enzyme in myelin breakdown. The degradation of myelin basic protein (MBP) by proteinase yields encephalitogenic peptides and its loss has been found to cause structural alteration of the myelin sheath. This suggests that MBP degradation is an initial step in the breakdown of myelin in demyelinating diseases. A calcium-activated neutral proteinase (calpain), which degrades MBP, was found to increase in activity in MS tissue and cerebrospinal fluid (CSF), and its presence in myelin suggests that myelin may be autodigested in demyelinating disease. The source of increased proteinase activity has been indicated as macrophages, lymphocytes, and proliferative astrocytes (reactive cells). Increased proteinase activity is found in Schwann cells in Wallerian degeneration, and the presence of calpain in myelin-forming oligodendrocytes and Schwann cells suggests that these cells are likely sources of degradative enzymes. The involvement of proteolytic enzymes in the mechanism of myelin breakdown indicates the possible intervention with proteinase inhibitors for beneficial effect.

Topics: Animals; Brain; Demyelinating Diseases; Endopeptidases; Humans; Models, Neurological; Myelin Basic Protein; Myelin Sheath; Peptide Hydrolases

Multiple sclerosis (MS) and a number of related distinctive diseases are characterized by the active degradation of central nervous system (CNS) myelin, an axonal sheath comprised essentially of proteins and lipids. These demyelinating diseases appear to arise from complex interactions of genetic, immunological, infective, and biochemical mechanisms. While circumstances of MS etiology remain hypothetical, one persistent theme involves recognition by the immune system of myelin-specific antigens derived from myelin basic protein (MBP), the most abundant extrinsic myelin membrane protein, and/or another equally susceptible myelin protein or lipid component. Knowledge of the biochemical and physical-chemical properties of myelin proteins and lipids, particularly their composition, organization, structure, and accessibility with respect to the compacted myelin multilayers, thus becomes central to the understanding of how and why these antigens become selected during the development of MS. This review focuses on current understanding of the molecular basis underlying demyelinating disease as it may relate to the impact of the various protein and lipid components on myelin morphology; the precise molecular architecture of this membrane as dictated by protein-lipid and lipid-lipid interactions; and the relationship, if any, between the protein/lipid components and the destruction of myelin in pathological situations.

Topics: Amino Acid Sequence; Animals; Central Nervous System; Demyelinating Diseases; Humans; Lipid Metabolism; Lipids; Molecular Sequence Data; Myelin Basic Protein; Myelin Proteins; Myelin Sheath; Proteolipids; Uteroglobin

Topics: Animals; Antibodies; Antibodies, Monoclonal; Antigen-Antibody Reactions; Biological Evolution; Cross Reactions; Demyelinating Diseases; Humans; Immunologic Techniques; Models, Molecular; Myelin Basic Protein; Peptide Fragments; Species Specificity

Shiverer (shi/shi) is an autosomal recessive mouse mutation that produces a shivering phenotype in affected mice. A shivering gait can be seen from a few weeks after birth until their early death, which occurs between 50 and 100 days. The central nervous system of the mutant mouse is hypomyelinated but the peripheral nervous system appears normal. The myelin of the CNS, wherever present, is not well compacted and lacks the major dense line. Myelin basic protein (MBP), which is associated with the major dense line, is absent, and this is due to a deletion of the major part of the gene encoding MBP. Transgenic shiverer mice that have integrated and express the wild-type mouse MBP transgene no longer shiver and have normal life spans. Conversely, normal mice that have integrated an antisense MBP transgene, shiver. Myelin deficient shimld/shimld is allelic to shiverer (shi/shi) but the mutant mouse is less severely affected. Although MBP is present in the CNS, it is low in quantity and is not developmentally regulated. The gene encoding MBP has been both duplicated and inverted. Transgenic shimld/shimld mice with the wild-type MBP transgene have normal phenotypes.

Topics: Animals; Demyelinating Diseases; Mice; Mice, Jimpy; Mice, Neurologic Mutants; Mice, Transgenic; Mutation; Myelin Basic Protein; Myelin Proteins; Myelin Sheath

Oligodendrocyte development and myelinogenesis, both in vivo and in vitro, are characterized by the sequential and coordinate expression of markers which participate in the differentiation of oligodendrocytes as a prerequisite for myelination. The myelin deficient (md) rat shows greatly reduced mRNA expression for several oligodendrocyte markers: glycerol phosphate dehydrogenase (GPDH), myelin basic protein (MBP) and proteolipid protein (PLP). Brain GPDH mRNA levels are initially equivalent in md and unaffected littermates, but the mutant rats fail to display the normal developmental increase in gene expression. Immunostaining of brain tissue sections also reveals decreased expression of these oligodendrocyte markers. The number of oligodendrocytes containing GPDH-like immunoreactivity is reduced in mutant rats, and in general these cells appear morphologically less complex with shorter processes. However, the intensity of staining in many oligodendrocytes appears equivalent to that observed in unaffected rats. Expression of the neuronal marker, glutamic acid decarboxylase, and the astrocyte markers, glutamine synthetase and glial fibrillary acidic protein, are largely unaffected at either the mRNA or protein level. Mixed glial cultures prepared from the brains of neonatal male md rats possess fewer oligodendrocytes compared to cultures derived from unaffected littermates, and the temporal sequence of marker development is delayed. Although an abnormality in the PLP gene is suspected in the md rat, these findings document profound deficits in many oligodendrocyte gene products.

Topics: Animals; Demyelinating Diseases; Gene Expression Regulation; Glycerolphosphate Dehydrogenase; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Oligodendroglia; Rats; Rats, Mutant Strains

Topics: Demyelinating Diseases; Humans; Myelin Basic Protein; Myelin Sheath

The precise role of T cells in multiple sclerosis (MS) remains to be defined. No MS-specific antigen has been found. The autoimmune hypothesis for MS rests on immune changes seen in the spinal fluid and brain and on the demonstration, in an experimental animal model, that T cells raised to myelin basic protein transfer demyelination. In this review, Virginia Calder and colleagues focus on recent studies suggesting that in MS, the initial T-cell response occurs within the central nervous system and that the blood poorly reflects this immune activity. This contrasts with the animal model, experimental allergic encephalomyelitis, where the initial immune response is peripheral.

Topics: Central Nervous System; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Humans; Multiple Sclerosis; Myelin Basic Protein; T-Lymphocytes

Topics: Animals; Antigens, Viral; Autoimmune Diseases; Central Nervous System; Cross Reactions; Demyelinating Diseases; Galactosidases; Galactosylceramidase; Humans; Myelin Basic Protein

Topics: Amino Acid Sequence; Animals; Autoimmune Diseases; Demyelinating Diseases; Disease Models, Animal; Disease Susceptibility; Encephalomyelitis, Autoimmune, Experimental; Epitopes; Molecular Sequence Data; Myelin Basic Protein; Primates; Rabbits; Rodentia; Species Specificity; T-Lymphocytes

Experimental allergic encephalomyelitis (EAE) is an autoimmune disease of the central nervous system (CNS). Its immune mechanism is well understood at the cellular and molecular levels, which is herein reviewed. Susceptibility to EAE is under the control of the genes partially inside and partially outside the H-2 complex. There are two myelin constituents known to be encephalitogenic, myelin basic protein and proteolipid apoprotein. EAE is mediated by effector T cells sensitized to the encephalitogen. Effector T cells bear surface phenotypes of Lyt1+2-, L3T4+, and they are activated by the encephalitogen/self Ia complex or certain alloantigens and acquire encephalitogenic activity. By unknown homing mechanisms, the effector T cells invade the CNS and induce the target phase phenomena, which include Ia-antigen expression in the local tissue, activation of procoagulant activity, breakdown of the blood-brain barrier, and excretion of lymphokines which induce inflammation and demyelination, resulting in functional alteration. Possibility of specific immune therapy is postulated as a model for human autoimmune disease.

Topics: Animals; Brain; Cell Movement; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Immunotherapy; Mice; Myelin Basic Protein; Recurrence; Remission, Spontaneous; T-Lymphocytes

During postnatal development of the higher vertebrate CNS, large populations of oligodendroglia are generated from precursor cells in a very dependable way. In adult lesioned CNS tissues, local populations of oligodendroglia are replenished by proliferation of this replenishment varies from one species to another and also from one lesion type another. Studies on the developmental generation of oligodendroglia are reviewed here, delineating what is known of the early relationships between the CNS glial lineages and of what regulates this development. Contributions from recent cell biological work are considered against the background of morphological and radioautographic results. The quiescent condition of extremely slow turnover in the normal adult CNS is noted, and the dramatic effects of lesions on the neural cell environment are considered. Lesions can trigger proliferation at a much greater rate in the mature oligodendroglial population, as observed both in situ and in tissue culture; in addition to persisting stem cells, the mature cells participate in replenishing the local oligodendroglial population. This regeneration from cells already committed to the oligodendroglial lineage may minimise such disturbing effects of the lesion environment as might distort replenishment of the population from precursor cells.

Topics: Animals; Blood-Brain Barrier; Cell Differentiation; Cell Separation; Cell Survival; Cells, Cultured; Central Nervous System; Demyelinating Diseases; Galactosylceramides; Glial Fibrillary Acidic Protein; Glucose; Humans; Insulin; Mice; Mice, Neurologic Mutants; Mitosis; Myelin Basic Protein; Myelin Proteins; Myelin Sheath; Neuroglia; Oligodendroglia; Rats

Topics: Animals; Animals, Newborn; Antigens, Surface; Brain; Cell Differentiation; Cell Division; Cell Movement; Cells, Cultured; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Galactosylceramides; Humans; Mice; Myelin Basic Protein; Myelin Sheath; Neuroglia; Oligodendroglia

Topics: Animals; Autoantigens; Demyelinating Diseases; Disease Models, Animal; Encephalomyelitis, Autoimmune, Experimental; Humans; Immunity, Cellular; Multiple Sclerosis; Myelin Basic Protein

Topics: Animals; Autoimmune Diseases; Demyelinating Diseases; Disease Models, Animal; Encephalomyelitis, Autoimmune, Experimental; Galactosylceramides; Guinea Pigs; Humans; Mice; Microscopy, Electron; Multiple Sclerosis; Myelin Basic Protein; Rats

Topics: Amino Acid Sequence; Amino Acids; Animals; Antigens; Chemical Phenomena; Chemistry; Demyelinating Diseases; Humans; Myelin Basic Protein; Neuritis; Peripheral Nervous System Diseases; Polyradiculoneuropathy; Sciatic Nerve

Topics: Antibody-Producing Cells; Autoantibodies; Brain; Cross Reactions; Demyelinating Diseases; Hemolytic Plaque Technique; Humans; Hydrolases; Immunoglobulin G; Lipid Metabolism; Multiple Sclerosis; Myelin Basic Protein; Myelin Sheath; Neuroglia; Nucleotidases; Subacute Sclerosing Panencephalitis

Myelin instability and citrullinated myelin basic protein have been demonstrated in the brains of patients with chronic and fulminating forms of multiple sclerosis (MS). Our aim was to trace citrulline in the brains of patients with early-onset MS by using proton MR spectroscopy ((1)H-MR spectroscopy).. A short-echo single-voxel (1)H-MR spectroscopy by using the point-resolved proton spectroscopy sequence was performed in 27 patients with MS and 23 healthy subjects. Voxels of interest were chronic demyelinating lesions (CDLs, n = 25) and normal-appearing white matter (NAWM, n = 25) on T2-weighted imaging, and when available in patients with MS, enhancing demyelinating lesions (EDLs, n = 8). Frontal white matter (WM) was studied in control subjects. N-acetylaspartate, choline, and myo-inositol (mIns)-creatine (Cr) ratios and the presence of a citrulline peak were noted.. Citrulline peaks were more frequently observed in patients with MS than in control subjects (P = .035), located in the NAWM in 8/25 (32%), in CDLs in 7/25 (28%), and in EDLs of 1/8 (12.5%) patients with MS. The presence of citrulline and measured metabolite/Cr ratios was not related to age at imaging, age at disease onset, duration of disease, or number of relapses. There was no significant metabolic difference between the NAWM of patients with MS and the WM of the control subjects. mIns/Cr was significantly greater in CDLs compared with the NAWM of patients with MS and the WM of healthy subjects.. Citrulline was more frequently identified in the brains of patients with early-onset MS than in healthy subjects by (1)H-MR spectroscopy, suggesting an association of increased citrullination of myelin proteins with demyelinating diseases.

Topics: Adolescent; Age of Onset; Aspartic Acid; Child; Child, Preschool; Choline; Citrulline; Creatine; Demyelinating Diseases; Female; Humans; Inositol; Magnetic Resonance Spectroscopy; Male; Multiple Sclerosis, Relapsing-Remitting; Myelin Basic Protein; Myelin Sheath; Protons; Young Adult

To investigate the hypothesis that methotrexate causes demyelination due to a deficiency in S-adenosylmethionine (SAM) during the treatment of acute lymphoblastic leukemia (ALL).. Twenty-four patients treated on the Medical Research Council United Kingdom ALL trial no. 11 (MRC UKALL XI) were studied. The trial randomized patients at the presymptomatic CNS treatment (PCNS) phase to receive (1) intrathecal methotrexate and cranial radiotherapy (CRTX); (2) high-dose intravenous methotrexate with folinic acid rescue and continuing intrathecal methotrexate (HDMTX); and (3) continuing intrathecal methotrexate alone (ITMTX). Serial CSF samples were collected throughout treatment and concentrations of 5-methyltetrahydrofolate (MTF), methionine (MET), SAM, and myelin basic protein (MBP) were measured. The results were grouped into treatment milestones and compared with an age-matched reference population.. There was a highly significant effect of both treatment milestones and trial arm on the metabolite and MBP concentrations. CSF MTF reached a nadir during the induction phase of treatment, while SAM and MET reached their nadir during the consolidation phase. CSF MBP mirrored SAM concentration and there was a significant inverse relationship between the two. MTF, SAM, and MBP returned to normal values by the end of treatment, while MET was increased significantly. The effect of treatment was decremental across the ITMTX, HDMTX, and CRTX groups.. Treatment of ALL causes marked abnormalities in the single-carbon transfer pathway and subclinical demyelination. Methotrexate is one cause of this. Whether these abnormalities contribute to the late cognitive deficits requires further study.

Topics: Antidotes; Antimetabolites, Antineoplastic; Central Nervous System; Child; Child, Preschool; Combined Modality Therapy; Demyelinating Diseases; Drug Interactions; Humans; Infant; Leucovorin; Methionine; Methotrexate; Myelin Basic Protein; Precursor Cell Lymphoblastic Leukemia-Lymphoma; S-Adenosylmethionine; Tetrahydrofolates

Demyelination disorder is an unusual pathologic event, which occurs in the central nervous system (CNS). Multiple sclerosis (MS) is an inflammatory demyelinating disease that affects the CNS, and it is the leading cause of disability in young adults. Lysolecithin (LPC) is one of the best toxin-induced demyelination models. In this study, a suitable model is created, and the effect of fluoxetine treatment is examined on this model. In this case, it was assumed that daily fluoxetine treatment had increased the endogenous remyelination in the LPC model. This study was focused on investigating the influence of the fluoxetine dose of 5 or 10 mg/kg per day for 1 and 4 weeks on LPC-induced neurotoxicity in the corpus callosum region. It was performed as a demyelinating model in male Wistar rats. After 3 days, fluoxetine was injected intraperitoneally (5 or 10 mg/kg per day) for 1 and 4 weeks in each group. After completing the treatment course, the corpus callosum was removed to examine the gene expression and histological analysis was performed. The results of the histopathological study of hematoxylin and eosin staining of the corpus callosum showed that in 1 and 4-week treatment groups, fluoxetine has reduced the level of inflammation at the LPC injection site (5 and 10 mg/kg per day). Fluoxetine treatment in the luxol fast blue (LFB) staining of the corpus callosum has been led to an increase in myelination capacity in all doses and times. The results of the genetic study showed that the fluoxetine has significantly reduced the expression level of tumor necrosis factor-α, nuclear factor κβ, and induced nitric oxide synthase in comparison with the untreated LPC group. Also, the fluoxetine treatment has enhanced the expression level of the forkhead box P3 (

Topics: Animals; Brain-Derived Neurotrophic Factor; Corpus Callosum; Demyelinating Diseases; Disease Models, Animal; Fluoxetine; Forkhead Transcription Factors; Gene Expression; Lysophosphatidylcholines; Male; Myelin Basic Protein; NF-kappa B; Nitric Oxide Synthase; Oligodendrocyte Transcription Factor 2; Rats, Wistar; Tumor Necrosis Factor-alpha

1,2-Dichloroethane (1,2-DCE) is a pervasive environmental pollutant, and overexposure to this hazardous material causes brain edema and demyelination in humans. We found that 1,2-DCE inhibits aquaporin 4 (AQP4) and is a primary pathogenic effector of 1,2-DCE-induced brain edema in animals. However, AQP4 down-regulation's link with cortex demyelination after 1,2-DCE exposure remains unclear. Thus, we exposed wild-type (WT) CD-1 mice and AQP4 knockout (AQP4-KO) mice to 0, 100, 350 and 700 mg/m

Topics: Animals; Aquaporin 4; Demyelinating Diseases; Ethylene Dichlorides; Mice; Myelin Basic Protein

The main pathological characteristics of demyelinating diseases are central nervous system (CNS) myelin damage, and the differentiation of oligodendrocyte precursor cells is the therapeutic target of myelin repair. Previous studies have found that a large number of platelet-derived growth factor receptor α(PDGFRα) positive oligodendrocyte progenitor cells (OPCs) accumulate in the lesion area of myelin injury, and differentiation is blocked. However, the therapeutic effects of drugs currently used clinically on OPCs differentiation and myelin repair are limited. The main reason is that it is difficult to reach the effective concentration of the drug in the lesion area. Therefore, efficiently delivering into the CNS lesion area is of great significance for the treatment of MS. Natural exosomes have good biocompatibility and are ideal drug carriers. The delivery of drugs to lesion areas can be achieved by giving the exosomes armed targeting ligand. Therefore, in this study, combining exosomes with PDGFA helps them accumulate in OPCs in vitro and in vivo. Further, load montelukast into exosomes to achieve targeted therapy for cuprizone-induced demyelination animal model. The implementation of this research will help provide effective treatments for demyelinating diseases and lay a theoretical foundation for its application in the clinical treatment of different demyelinating diseases.

Topics: Acetates; Animals; Cell Differentiation; Cell Lineage; Cuprizone; Cyclopropanes; Demyelinating Diseases; Disease Models, Animal; Drug Delivery Systems; Exosomes; Extracellular Vesicles; In Vitro Techniques; Ligands; Male; Mice; Mice, Inbred C57BL; Myelin Basic Protein; Myelin Sheath; Neurons; Oligodendrocyte Precursor Cells; Oligodendroglia; Phagocytosis; Quinolines; Receptor, Platelet-Derived Growth Factor alpha; Regeneration; Stem Cells; Sulfides

Multiple sclerosis (MS) is a chronic demyelinating disease of the central nervous system (CNS). Most exercise studies concentrate on the impact of exercise on cardiovascular system; this study aims to present the effects of exercise of varying intensity on the nervous system. Most recently in MS, positive outcomes were obtained with resistance and high-intensity exercises. This study also analyzes the effects of a prior conditioning program before the induction of demyelination and subsequent neuroprotective effects of such program.. To study and determine the neuroprotective and remyelinating effects of different intensity of aquatic exercise and a preconditioning exercise program on demyelination induced by oral administration of cuprizone (Cup).. Six groups of animals, each containing 6 rats, were used in the study. The groups were as follows: group I - control group; group II - Cup group; group III - treated with methylprednisolone (MP); group IV - treated with low-intensity exercise (LIE), free swimming for 40 min and high-intensity exercise (HIE); group V - treated with a resistance of 9% body weight and free swimming for 40 min; group VI - treated with preconditioning exercise (free swimming for 40 min for 3 weeks) before Cup administration followed by the same exercise protocol as for group V. All data were analyzed using one-way analysis of variance (ANOVA) with Tukey's test, by means of SigmaPlot v. 14.5 software.. Similarly to the MP group, group VI showed a positive outcome. A value of p < 0.001 was considered statistically significant. Also, group VI showed improved areas of remyelination in histopathology, an increased expression of myelin basic protein (MBP), reduced expression of glial fibrillary acidic protein (GFAP) in corpus callosum, and improved gene expression of brain-derived neurotrophic factor (BDNF) in the hippocampus region.. General fitness achieved through a preconditioning program combined with HIE showed neuroprotective effects, as evidenced by increased areas of remyelination and improved neuronal plasticity, observed mostly in group VI (conditioning+HIE).

Topics: Animals; Brain-Derived Neurotrophic Factor; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Glial Fibrillary Acidic Protein; Male; Methylprednisolone; Mice; Mice, Inbred C57BL; Multiple Sclerosis; Myelin Basic Protein; Neuronal Plasticity; Neuroprotective Agents; Rats; Rats, Wistar; Remyelination

Multiple sclerosis (MS) is a chronic neurodegenerative disease marked by oligodendrocyte loss, which results in central neuronal demyelination. AC/cAMP/CREB signaling dysregulation is involved in the progression of MS, including mitochondrial dysfunctions, reduction in nerve growth factors, neuronal inflammation, apoptosis, and white matter degeneration. Our previous research has shown that Forskolin (FSK), a naturally occurring direct adenylyl cyclase (AC)/cAMP/CREB activator, has neuroprotective potential to alleviate pathogenic factors linked with numerous neurological abnormalities. The current study intends to explore the neuroprotective potential of FSK at doses of 40 mg/kg and 60 mg/kg alone, as well as in combination with conventional medicines, such as Fingolimod (FNG), Donepezil (DON), Memantine (MEM), and Simvastatin (SIM) in EB-induced demyelinated experimental MS rats. Adult Wistar rats were divided into nine groups, and EB was infused stereotaxically in the rat brain's intracerebropeduncle (ICP) area. Chronic gliotoxin EB treatment results in demyelination as well as motor and cognitive dysfunctions. FSK, combined with standard medications, improves behavioral dysfunctions, such as neuromuscular and motor deficits and memory and cognitive abnormalities. Following pharmacological treatments improved remyelination by enhancing myelin basic protein and increasing AC, cAMP, and CREB levels in brain homogenates. Furthermore, FSK therapy restored brain mitochondrial-ETC complex enzymes and neurotransmitter levels while decreasing inflammatory cytokines and oxidative stress markers. The Luxol fast blue (LFB) stain results further indicate FSK's neuroprotective potential in preventing oligodendrocyte death. Therefore, the results of these studies contribute to a better understanding of the possible role that natural phytochemicals FSK could have in preventing motor neuron diseases, such as multiple sclerosis.

Topics: Adenylyl Cyclases; Animals; Colforsin; Cytokines; Demyelinating Diseases; Donepezil; Ethidium; Fingolimod Hydrochloride; Gliotoxin; Memantine; Multiple Sclerosis; Myelin Basic Protein; Myelin Sheath; Nerve Growth Factors; Neurodegenerative Diseases; Oligodendroglia; Rats; Rats, Wistar; Simvastatin

Microglia are the primary phagocytes of the central nervous system and are responsible for removing damaged myelin following demyelination. Previous investigations exploring the consequences of myelin phagocytosis on microglial activation overlooked the biochemical modifications present on myelin debris. Such modifications, including citrullination, are increased within the inflammatory environment of multiple sclerosis lesions.. Mouse cortical myelin isolated by ultracentrifugation was citrullinated ex vivo by incubation with the calcium-dependent peptidyl arginine deiminase PAD2. Demyelination was induced by 6 weeks of cuprizone (0.3%) treatment and spontaneous repair was initiated by reversion to normal chow. Citrullinated or unmodified myelin was injected into the primary motor cortex above the cingulum bundle at the time of reversion to normal chow and the consequent impact on remyelination was assessed by measuring the surface area of myelin basic protein-positive fibers in the cortex 3 weeks later. Microglial responses to myelin were characterized by measuring cytokine release, assessing flow cytometric markers of microglial activation, and RNAseq profiling of transcriptional changes.. Citrullinated myelin induced a unique microglial response marked by increased tumor necrosis factor α (TNFα) production both in vitro and in vivo. This response was not induced by unmodified myelin. Injection of citrullinated myelin but not unmodified myelin into the cortex of cuprizone-demyelinated mice significantly inhibited spontaneous remyelination. Antibody-mediated neutralization of TNFα blocked this effect and restored remyelination to normal levels.. These findings highlight the role of post-translation modifications such as citrullination in the determination of microglial activation in response to myelin during demyelination. The inhibition of endogenous repair induced by citrullinated myelin and the reversal of this effect by neutralization of TNFα may have implications for therapeutic approaches to patients with inflammatory demyelinating disorders.

Topics: Animals; Cells, Cultured; Chelating Agents; Citrulline; Cuprizone; Cytokines; Demyelinating Diseases; Mice; Mice, Inbred C57BL; Microglia; Microinjections; Motor Cortex; Myelin Basic Protein; Myelin Sheath; Tumor Necrosis Factor-alpha

Spinal cord injury (SCI) can cause a variety of cells apoptosis, neurodegeneration, and eventually permanent paralysis. This study aimed to examine whether transplanting human umbilical cord mesenchymal stem cells (hucMSCs) can promote locomotor function recovery, reduce apoptosis and inhibit demyelination in SCI models.. Rats were allocated into Sham group (spinal cord exposure only), SCI + PBS group (spinal cord impact plus phosphate-buffered saline (PBS) injections), SCI + hucMSCs group (spinal cord impact plus hucMSCs injections) groups. Behavioral tests, Basso-Beattie-Bresnahan locomotion scores (BBB scores), were carried out at 0, 3, 7, 14, 21, 28 days after SCI surgery. Hematoxylin-eosin staining observed spinal cord morphology. Nissl staining detected the number of nissl bodies. Myelin basic protein (MBP) and oligodendrocyte (CNPase) were examed by immunohistochemical staining. The apoptosis of oligodendrocyte and neurons were detected by immunofluorescence.. The 28-day behavioral test showed that the BBB score of rats in the SCI + hucMSCs group increased significantly, comparing to the SCI + PBS group. The numbers of nissl bodies and myelin sheath in the damaged area of SCI + hucMSCs group were also significantly increased compared to the SCI + PBS group. HucMSCs transplanting decreased the expression of protein level of Caspase-3 and Bax and increased the Bcl-2, MBP and CNPase, rescued the apoptosis of neurons and the oligodendrocyte.. These results showed that hucMSCs can improve motor function, tissue repairing and reducing apoptosis in SCI rats.

Topics: Animals; Apoptosis; Blood-Brain Barrier; Demyelinating Diseases; Female; Gene Expression Regulation; Humans; Locomotion; Male; Mesenchymal Stem Cell Transplantation; Myelin Basic Protein; Neurons; Oligodendroglia; Rats; Rats, Sprague-Dawley; Recovery of Function; Spinal Cord Injuries

Multiple sclerosis (MS) is a neuroinflammatory and neurodegenerative disease characterized by myelin damage followed by axonal and ultimately neuronal loss. The etiology and physiopathology of MS are still elusive, and no fully effective therapy is yet available. We investigated the role in MS of autophagy (physiologically, a controlled intracellular pathway regulating the degradation of cellular components) and of mitophagy (a specific form of autophagy that removes dysfunctional mitochondria). We found that the levels of autophagy and mitophagy markers are significantly increased in the biofluids of MS patients during the active phase of the disease, indicating activation of these processes. In keeping with this idea, in vitro and in vivo MS models (induced by proinflammatory cytokines, lysolecithin, and cuprizone) are associated with strongly impaired mitochondrial activity, inducing a lactic acid metabolism and prompting an increase in the autophagic flux and in mitophagy. Multiple structurally and mechanistically unrelated inhibitors of autophagy improved myelin production and normalized axonal myelination, and two such inhibitors, the widely used antipsychotic drugs haloperidol and clozapine, also significantly improved cuprizone-induced motor impairment. These data suggest that autophagy has a causal role in MS; its inhibition strongly attenuates behavioral signs in an experimental model of the disease. Therefore, haloperidol and clozapine may represent additional therapeutic tools against MS.

Topics: Animals; Antipsychotic Agents; Autophagy; Autophagy-Related Proteins; Axons; Biomarkers; Clozapine; Cytokines; Demyelinating Diseases; Disease Models, Animal; Glucose; Haloperidol; Inflammation; Interleukin-1beta; Mitochondria; Mitophagy; Models, Biological; Motor Activity; Multiple Sclerosis; Myelin Basic Protein; Myelin Sheath; Stress, Physiological; Tumor Necrosis Factor-alpha

Incomplete remyelination is frequent in multiple sclerosis (MS)-lesions, but there is no established marker for recent remyelination. We investigated the role of the oligodendrocyte/myelin protein ermin in de- and remyelination in the cuprizone (CPZ) mouse model, and in MS. The density of ermin+ oligodendrocytes in the brain was significantly decreased after one week of CPZ exposure (p < 0.02). The relative proportion of ermin+ cells compared to cells positive for the late-stage oligodendrocyte marker Nogo-A increased at the onset of remyelination in the corpus callosum (p < 0.02). The density of ermin-positive cells increased in the corpus callosum during the CPZ-phase of extensive remyelination (p < 0.0001). In MS, the density of ermin+ cells was higher in remyelinated lesion areas compared to non-remyelinated areas both in white- (p < 0.0001) and grey matter (p < 0.0001) and compared to normal-appearing white matter (p < 0.001). Ermin immunopositive cells in MS-lesions were not immunopositive for the early-stage oligodendrocyte markers O4 and O1, but a subpopulation was immunopositive for Nogo-A. The data suggest a relatively higher proportion of ermin immunopositivity in oligodendrocytes compared to Nogo-A indicates recent or ongoing remyelination.

Topics: Animals; Brain; Cerebral Cortex; Corpus Callosum; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Female; Gray Matter; Mice; Mice, Inbred C57BL; Multiple Sclerosis; Myelin Basic Protein; Myelin Proteins; Myelin Sheath; Oligodendroglia; Remyelination; White Matter

Cognitive decline remains an unaddressed problem for the elderly. We show that myelination is highly active in young mice and greatly inhibited in aged mice, coinciding with spatial memory deficits. Inhibiting myelination by deletion of Olig2 in oligodendrocyte precursor cells impairs spatial memory in young mice, while enhancing myelination by deleting the muscarinic acetylcholine receptor 1 in oligodendrocyte precursor cells, or promoting oligodendroglial differentiation and myelination via clemastine treatment, rescues spatial memory decline during aging.

Topics: Aging; Animals; Demyelinating Diseases; Memory Disorders; Mice; Mice, Transgenic; Myelin Basic Protein; Myelin Sheath; Oligodendrocyte Precursor Cells; Oligodendrocyte Transcription Factor 2; Oligodendroglia

Lead is a toxic metal found in environment with great neurotoxic potential. The main effect is associated with impairments in hippocampus and cerebellum, driving to cognitive and motor dysfunctions, however, there is a lack of evidences about the effects over the spinal cord. In this way, we aimed to investigate in vivo the effects of long-term exposure to lead acetate in oxidative biochemistry and morphology of rats' spinal cord. For this, 36 male Wistar rats (Rattus norvegicus) were divided into the group exposed to 50 mg/kg of lead acetate and control group, which received only distilled water, both groups through intragastric gavage, for 55 days. After the exposure period, the animals were euthanized and the spinal cords were collected to perform the analyses of lead levels quantification, oxidative biochemistry evaluation by levels of malondialdehyde (MDA), nitrites and the antioxidant capacity against peroxyl radicals (ACAP). Besides, morphological evaluation with quantitative analysis of mature and motor neurons and reactivity to myelin basic protein (MBP). Our results showed high levels of lead in spinal cord after long-term exposure; there was a reduction on ACAP level; however, there was no difference observed in MDA and nitrite levels. Moreover, there was a reduction of mature and motor neurons in all three regions, and a reduction of immunolabeling of MBP in the thoracic and lumbar segments. Therefore, we conclude that long-term exposure to lead is able of increasing the levels of the metal in spinal cord, affecting the antioxidant capacity and inducing morphological impairments in spinal cord parenchyma. Our results also suggest that the tissue impairments triggered by lead may be resultant from others molecular mechanisms besides the oxidative stress.

Topics: Animals; Antioxidants; Demyelinating Diseases; Hazardous Substances; Hippocampus; Lead; Male; Malondialdehyde; Motor Neurons; Myelin Basic Protein; Nitrites; Oxidation-Reduction; Oxidative Stress; Peroxides; Rats; Rats, Wistar; Spinal Cord; Toxicity Tests

Inorganic arsenic is among the major contaminants of groundwater in the world. Worldwide population-based studies demonstrate that chronic arsenic exposure is associated with poor cognitive performance among children and adults, while research in animal models confirms learning and memory deficits after arsenic exposure. The aim of this study was to investigate the long-term effects of environmentally relevant arsenic exposure in the myelination process of the prefrontal cortex (PFC) and corpus callosum (CC). A longitudinal study with repeated follow-up assessments was performed in male Wistar rats exposed to 3 ppm sodium arsenite in drinking water. Animals received the treatment from gestation until 2, 4, 6, or 12 months of postnatal age. The levels of myelin basic protein (MBP) were evaluated by immunohistochemistry/histology and immunoblotting from the PFC and CC. As plausible alterations associated with demyelination, we considered mitochondrial mass (VDAC) and two axonal damage markers: amyloid precursor protein (APP) level and phosphorylated neurofilaments. To analyze the microstructure of the CC in vivo, we acquired diffusion-weighted images at the same ages, from which we derived metrics using the tensor model. Significantly decreased levels of MBP were found in both regions together with significant increases of mitochondrial mass and slight axonal damage at 12 months in the PFC. Ultrastructural imaging demonstrated arsenic-associated decreases of white matter volume, water diffusion anisotropy, and increases in radial diffusivity. This study indicates that arsenic exposure is associated with a significant and persistent negative impact on microstructural features of white matter tracts.

Topics: Aging; Amyloid beta-Protein Precursor; Animals; Arsenic Poisoning; Arsenites; Axons; Corpus Callosum; Demyelinating Diseases; Diffusion Tensor Imaging; Drinking Water; Immunohistochemistry; Male; Mitochondria; Myelin Basic Protein; Neurofilament Proteins; Prefrontal Cortex; Rats; Rats, Wistar; Sodium Compounds; White Matter

Ketogenic diet (KD) is defined as a high-fat, low-carbohydrate diet with appropriate amounts of protein, which has broad neuroprotective effects. However, the mechanisms of ameliorating the demyelination and of the neuroprotective effects of KD have not yet been completely elucidated. Therefore, the present study investigated the protection mechanism of KD treatment in the cuprizone (bis-cyclohexanone oxalydihydrazone, CPZ)-induced demyelination mice model, with special emphasis on neuroinflammation. After the KD treatment, an increased ketone body level in the blood of mice was detected, and a significant increase in the distance traveled within the central area was observed in the open field test, which reflected the increased exploration and decreased anxiety of mice that received CPZ. The results of Luxol fast blue and myelin basic protein (MBP) immunohistochemistry staining for the evaluation of the myelin content within the corpus callosum revealed a noticeable increase in the number of myelinated fibers and myelin score after KD administration in these animals. Concomitant, the protein expressions of glial fibrillary acidic protein (GFAP, an astrocyte marker), ionized calcium-binding adaptor molecule 1 (Iba-1, a microglial marker), CD68 (an activated microglia marker) and CD16/32 (a M1 microglial marker) were down-regulated, while the expression of oligodendrocyte lineage transcription factor 2 (OLIG2, an oligodendrocyte precursor cells marker) was up-regulated by the KD treatment. In addition, the KD treatment not only reduced the level of the C-X-C motif chemokine 10 (CXCL10), which is correlated to the recruitment of activated microglia, but also inhibited the production of proinflammatory cytokines, including interleukin 1β (IL-1β) and tumor necrosis factor-α (TNF-α), which are closely correlated to the M1 phenotype microglia. It is noteworthy, that the expression levels of histone deacetylase 3 (HADC3) and nod-like receptor pyrin domain containing 3 (NLRP3) significantly decreased after KD administration. In conclusion, these data demonstrate that KD decreased the reactive astrocytes and activated the microglia in the corpus callosum, and that KD inhibited the HADC3 and NLRP3 inflammasome signaling pathway in CPZ-treated mice. This suggests that the inhibition of the HADC3 and NLRP3 signaling pathway may be a novel mechanism by which KD exerts its protective actions for the treatment of demyelinating diseases.

Topics: Alanine Transaminase; Animals; Aspartate Aminotransferases; Astrocytes; Body Weight; Brain; Chemokine CXCL10; Cuprizone; Cytokines; Demyelinating Diseases; Diet, Ketogenic; Disease Models, Animal; Down-Regulation; Glial Fibrillary Acidic Protein; Histone Deacetylases; Inflammasomes; Male; Mice; Mice, Inbred C57BL; Microglia; Myelin Basic Protein; NLR Family, Pyrin Domain-Containing 3 Protein; Signal Transduction; Weight Loss

Accumulating evidence suggests that platelet-activating factor (PAF) increases the inflammatory response in demyelinating diseases such as multiple sclerosis. However, PAF receptor (PAFR) antagonists do not show therapeutic efficacy for MS, and its underlying mechanisms remain poorly understood. In the present study, we investigated the effects of PAF on an ex vivo demyelination cerebellar model following lysophosphatidylcholine (LPC, 0.5 mg/mL) application using wild-type and PAFR conventional knockout (PAFR-KO) mice. Demyelination was induced in cerebellar slices that were cultured with LPC for 18 h. Exogenous PAF (1 μM) acting on cerebellar slices alone did not cause demyelination but increased the severity of LPC-induced demyelination in both wild-type and PAFR-KO mice. LPC inhibited the expression of PAF-AH, MBP, TNF-α, and TGF-β1 but facilitated the expression of IL-1β and IL-6 in wild-type preparations. Of note, exogenous PAF stimulated microglial activation in both wild-type and PAFR-KO mice. The subsequent inflammatory cytokines TNFα, IL-1β, and IL-6 as well as the anti-inflammatory cytokine TGF-β1 demonstrated a diverse transcriptional profile with or without LPC treatment. PAF promoted TNF-α expression and suppressed TGF-β1 expression indiscriminately in wild-type and knockout slices; however, transcription of IL-1β and IL-6 was not significantly affected in both slices. The syntheses of IL-1β and IL-6 were significantly increased in LPC-induced demyelination preparations without PAF but showed a redundancy in PAF-treated wild-type and knockout slices. These data suggest that PAF can play a detrimental role in LPC-induced demyelination probably due to a redundant response of PAFR-dependent and PAFR-independent effects on inflammatory cytokines.

Topics: Animals; Cerebellum; Cytokines; Demyelinating Diseases; Gene Deletion; Inflammation Mediators; Lysophosphatidylcholines; Mice, Inbred C57BL; Mice, Knockout; Microglia; Myelin Basic Protein; Neurofilament Proteins; Platelet Activating Factor; Platelet Membrane Glycoproteins; Receptors, G-Protein-Coupled; Transcription, Genetic

Topics: Anacardic Acids; Animals; Central Nervous System; Demyelinating Diseases; Female; Interleukin-33; Male; Mice; Mice, Inbred C57BL; Myelin Basic Protein; Myelin Proteins; Myelin Sheath; Oligodendrocyte Precursor Cells; Oligodendroglia; Remyelination; Stem Cells

Macromolecular proton fraction (MPF) has been established as a quantitative clinically-targeted MRI myelin biomarker based on recent demyelination studies. This study aimed to assess the capability of MPF to quantify remyelination using the murine cuprizone-induced reversible demyelination model. MPF was measured in vivo using the fast single-point method in three animal groups (control, cuprizone-induced demyelination, and remyelination after cuprizone withdrawal) and compared to quantitative immunohistochemistry for myelin basic protein (MBP), myelinating oligodendrocytes (CNP-positive cells), and oligodendrocyte precursor cells (OPC, NG2-positive cells) in the corpus callosum, caudate putamen, hippocampus, and cortex. In the demyelination group, MPF, MBP-stained area, and oligodendrocyte count were significantly reduced, while OPC count was significantly increased as compared to both control and remyelination groups in all anatomic structures (

Topics: Animals; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Gray Matter; Magnetic Resonance Imaging; Male; Mesothelin; Mice; Myelin Basic Protein; Oligodendrocyte Precursor Cells; Oligodendroglia; Remyelination; White Matter

Demyelination in white matter is the end product of numerous pathological processes. This study was designed to evaluate the neuroprotective effect of l-serine and the underlying mechanisms against the demyelinating injury of white matter. A model of focal demyelinating lesions (FDL) was established using the two-point stereotactic injection of 0.25% lysophosphatidylcholine (LPC, 10 μg per point) into the corpus callosum of mice. Mice were then intraperitoneally injected with one of three doses of l-serine (114, 342, or 1026 mg/kg) 2 h after FDL, and then twice daily for the next five days. Behavior tests and histological analysis were assessed for up to twenty-eight days post-FDL induction. Electron microscopy was used for ultrastructural investigation. In vitro, we applied primary co-cultures of microglia and oligodendrocytes for oxygen glucose deprivation (OGD). After establishing FDL, l-serine treatment: 1) improved spatial learning, memory and cognitive ability in mice, and relieved anxiety for 4 weeks post-FDL induction; 2) reduced abnormally dephosphorylated neurofilament proteins, increased myelin basic protein, and preserved anatomic myelinated axons; 3) inhibited microglia activation and reduced the release of inflammatory factors; 4) promoted recruitment and proliferation of oligodendrocyte progenitor cells, and the efficiency of subsequent remyelination on day twenty-eight post-FDL induction. In vitro experiments, showed that l-serine not only directly protected against oligodendrocytes from OGD damage, but also provided an indirect protective effect by regulating microglia. In our study, l-serine offered long-lasting behavioral and oligodendrocyte protection and promoted remyelination. Therefore, l-serine may be an effective clinical treatment aganist white matter injury.

Topics: Animals; Anxiety; Axons; Calcium-Binding Proteins; Corpus Callosum; Demyelinating Diseases; Exploratory Behavior; Inflammation; L-Lactate Dehydrogenase (Cytochrome); Lysophosphatidylcholines; Male; Mice; Mice, Inbred C57BL; Microfilament Proteins; Microglia; Myelin Basic Protein; Myelin Sheath; Oligodendroglia; Serine; Spatial Learning; Spatial Memory

Multiple sclerosis (MS) is one of the most common causes of progressive disability affecting young people with very few therapeutic options available for its progressive forms. Its pathophysiology involves demyelination and neurodegeneration apparently driven by microglial activation, which is physiologically dependent on colony-stimulating factor-1 receptor (CSF-1R) signaling. In the present work, we used microglial modulation through oral administration of brain-penetrant CSF-1R inhibitor BLZ945 in acute and chronic cuprizone (CPZ)-induced demyelination to evaluate preventive and therapeutic effects on de/remyelination and neurodegeneration. Our results show that BLZ945 induced a significant reduction in the number of microglia. Preventive BLZ945 treatment attenuated demyelination in the acute CPZ model, mainly in cortex and external capsule. In contrast, BLZ945 treatment in the acute CPZ model failed to protect myelin or foster remyelination in myelin-rich areas, which may respond to a loss in microglial phagocytic capacity and the consequent impairment in oligodendroglial differentiation. Preventive and therapeutic BLZ945 treatment promoted remyelination and neuroprotection in the chronic model. These results could be potentially transferred to the treatment of progressive forms of MS.

Topics: Amyloid beta-Peptides; Animals; Benzothiazoles; Brain; Bromodeoxyuridine; Cuprizone; Cytokines; Demyelinating Diseases; Disease Models, Animal; Male; Mice; Mice, Inbred C57BL; Microglia; Microscopy, Electron, Transmission; Myelin Basic Protein; Nerve Tissue Proteins; Nitric Oxide Synthase Type II; Picolinic Acids; Receptors, Colony-Stimulating Factor; Time Factors

Organophosphorus compound (OP)-induced delayed neuropathy (OPIDN) is characterized by distal axonal degeneration and demyelination of the central and peripheral axons, which leads to progressive muscle weakness, ataxia and paralysis in several days after OP intoxication. This study aimed to investigate the possible use of an imidazole fungicide miconazole as a novel therapy for OPIDN. Adult hens, the most commonly used animal models in OPIDN studies, were orally given tri-o-cresyl phosphate (TOCP). We showed that miconazole, which was administered daily to hens beginning on the 7th day after TOCP exposure, drastically ameliorated the neurotoxic symptoms and histopathological damages in spinal cord and sciatic nerves. Mechanistically, miconazole inhibited the TOCP-induced activation of ErbB/Akt signaling, and enhanced the myelin basic protein (MBP) expression. In a glial cell model sNF96.2 cells, miconazole restored the TOCP-inhibited MBP expression, and promoted cell differentiation as well as cell migration by inhibiting the activation of ErbB/Akt signaling pathway. In sum, miconazole, a synthetic imidazole fungicide, could ameliorate the symptoms and histopathological changes of OPIDN, probably by promoting glial cell differentiation and migration to enhance myelination via inhibiting the activation of ErbB/Akt. Thus, miconazole is a promising candidate therapy for the clinical treatment of OPIDN.

Topics: Animals; Cell Differentiation; Cell Movement; Cells, Cultured; Chickens; Demyelinating Diseases; Female; Humans; Miconazole; Myelin Basic Protein; Neurotoxicity Syndromes; Oncogene Proteins v-erbB; Proto-Oncogene Proteins c-akt; Sciatic Nerve; Signal Transduction; Spinal Cord; Tritolyl Phosphates

In multiple sclerosis patients, chronic clinical deficits are known to result from axonal degeneration which is triggered by inadequate remyelination. The underlying molecular mechanisms of remyelination and its failure remain currently unclear. In vivo models, among the cuprizone model, are valuable tools to study underlying mechanisms of remyelination and its failure. Since complete and reproducible demyelination of the analyzed brain region is an indispensable prerequisite for efficient remyelination experiments, in this study we systematically addressed which part of the corpus callosum is reliably and consistently demyelinated after acute cuprizone-induced demyelination. Following a novel evaluation strategy, we can show that at the level of the rostral hippocampus, the most medial sectors of the corpus callosum (spanning 500 μm in the horizontal plane) are consistently demyelinated, whereas more lateral sectors show inconsistent and incomplete demyelination. These results precisely define a part of the corpus callosum which should be used as a region of interest during remyelination experiments.

Topics: Animals; Brain; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Male; Mice, Inbred C57BL; Myelin Basic Protein; Myelin Sheath; Oligodendroglia

Demyelination occurs following many neurological insults, most notably in multiple sclerosis (MS). Therapeutics that promote remyelination could slow the neurological decline associated with chronic demyelination; however, in vivo testing of candidate small molecule drugs and signaling cascades known to impact myelination is expensive and labor intensive. Here, we describe the development of a novel zebrafish line which uses the putative promoter of Myelin Protein Zero (mpz), a major structural protein in myelin, to drive expression of Enhanced Green Fluorescent Protein (mEGFP) specifically in the processes and nascent internodes of myelinating glia. We observe that changes in fluorescence intensity in Tg(mpz:mEGFP) larvae are a reliable surrogate for changes in myelin membrane production per se in live larvae following bath application of drugs. These changes in fluorescence are strongly predictive of changes in myelin-specific mRNAs [mpz, 36K and myelin basic protein (mbp)] and protein production (Mbp). Finally, we observe that certain drugs alter nascent internode number and length, impacting the overall amount of myelin membrane synthesized and a number of axons myelinated without significantly changing the number of myelinating oligodendrocytes. These studies demonstrate that the Tg(mpz:mEGFP) reporter line responds effectively to positive and negative small molecule regulators of myelination, and could be useful for identifying candidate drugs that specifically target myelin membrane production in vivo. Combined with high throughput cell-based screening of large chemical libraries and automated imaging systems, this transgenic line is useful for rapid large scale whole animal screening to identify novel myelinating small molecule compounds in vivo.

Topics: Animals; Animals, Genetically Modified; Culture Media, Conditioned; Demyelinating Diseases; Disease Models, Animal; Embryo, Nonmammalian; Embryonic Stem Cells; Gene Expression Regulation, Developmental; Green Fluorescent Proteins; Immunosuppressive Agents; Larva; Luminescent Proteins; Myelin Basic Protein; Myelin P0 Protein; Myelin Sheath; Neuroglia; Oligodendroglia; Red Fluorescent Protein; Sirolimus; SOXE Transcription Factors; Spinal Cord; Zebrafish; Zebrafish Proteins

The obstacle to successful remyelination in demyelinating diseases, such as multiple sclerosis, mainly lies in the inability of oligodendrocyte precursor cells (OPCs) to differentiate, since OPCs and oligodendrocyte-lineage cells that are unable to fully differentiate are found in the areas of demyelination. Thus, promoting the differentiation of OPCs is vital for the treatment of demyelinating diseases. Shikimic acid (SA) is mainly derived from star anise, and is reported to have anti-influenza, anti-oxidation, and anti-tumor effects. In the present study, we found that SA significantly promoted the differentiation of cultured rat OPCs without affecting their proliferation and apoptosis. In mice, SA exerted therapeutic effects on experimental autoimmune encephalomyelitis (EAE), such as alleviating clinical EAE scores, inhibiting inflammation, and reducing demyelination in the CNS. SA also promoted the differentiation of OPCs as well as their remyelination after lysolecithin-induced demyelination. Furthermore, we showed that the promotion effect of SA on OPC differentiation was associated with the up-regulation of phosphorylated mTOR. Taken together, our results demonstrated that SA could act as a potential drug candidate for the treatment of demyelinating diseases.

Topics: Animals; Apoptosis; Cell Differentiation; Cell Proliferation; Cells, Cultured; Demyelinating Diseases; Encephalitis; Encephalomyelitis, Autoimmune, Experimental; Female; Mice, Inbred C57BL; Myelin Basic Protein; Neuroprotective Agents; Oligodendrocyte Precursor Cells; Rats; Remyelination; Shikimic Acid; TOR Serine-Threonine Kinases

Multiple sclerosis is among the most common causes of neurological disabilities in young adults. Over the past decade, several therapeutic strategies have emerged as having potential neuroprotective and neuroregenerative properties. We investigated the effect of intranasal administration of LINGO-1-directed siRNA-loaded chitosan nanoparticles on demyelination and remyelination processes in a rat model of demyelination. Adult male Wistar rats were randomly assigned to one of 6 groups (n = 10 each) and subjected to intrapontine stereotaxic injection of ethidium bromide (EB) to induce demyelination. EB-treated rats were either left untreated or received intranasal LINGO-1-directed siRNA-chitosan nanoparticles from day 1 to day 7 (demyelination group) or from day 7 to day 21 (remyelination group) after EB injection. Chitosan nanoparticle (50 μl) was given alone after EB stereotaxic injection for both demyelination and remyelination groups. Two additional groups received 10 μl of saline by stereotaxic injection, followed by intranasal saline as controls for demyelination and remyelination groups (n = 10/group). Behavioural testing was conducted for all rats, as well as terminal biochemical assays and pathological examination of pontine tissues were done. After EB injection, rats had compromised motor performance and coordination. Pathological evidence of demyelination was observed in pontine tissue and higher levels of caspase-3 activity were detected compared to control rats. With LINGO-1-directed siRNA-chitosan nanoparticle treatment, animals performed better than controls. Remyelination-treated group showed better motor performance than demyelination group. LINGO-1 downregulation was associated with signs of repair in histopathological sections, higher expression of pontine myelin basic protein (MBP) mRNA and protein and lower levels of caspase-3 activity indicating neuroprotection and remyelination enhancement.

Topics: Administration, Intranasal; Animals; Ataxia; Caspase 3; Chitosan; Demyelinating Diseases; Disease Models, Animal; Drug Carriers; Drug Compounding; Ethidium; Gene Expression Regulation; Male; Membrane Proteins; Myelin Basic Protein; Myelin Sheath; Nanoparticles; Nerve Tissue Proteins; Neuroprotective Agents; Nucleus Raphe Magnus; Rats; Rats, Wistar; Remyelination; RNA, Small Interfering; Stereotaxic Techniques

To study cellular and molecular mechanisms of demyelination and remyelination in vivo, we developed a transgenic zebrafish line, Tg(mbp:mCherry-NTR), in which expression of the bacterial enzyme nitroreductase (NTR) is driven under the myelin basic protein promoter (mbp) and thus is expressed in myelinating glia. When NTR-expressing larvae are treated with the prodrug metronidazole, the reaction between NTR and Mtz results in a toxic metabolite which selectively kills NTR-expressing cells. Using the Tg(mbp:mCherry-NTR) line, we can ablate two-thirds of oligodendrocytes following a 2-day MTZ treatment. Demyelination is evident seven days later, and remyelination is observed 16 days after Mtz treatment. The Tg(mbp:mCherry-NTR) model can be used to image cell behavior during, and to test how genetic manipulations or chemical compounds regulate, demyelination and remyelination. In this chapter, we describe the methods we used to characterize the oligodendrocyte loss, demyelination and remyelination in the Tg(mbp:mCherry-NTR) model.

Topics: Animals; Animals, Genetically Modified; Demyelinating Diseases; Disease Models, Animal; Gene Expression Regulation, Developmental; Metronidazole; Myelin Basic Protein; Neuroglia; Nitroreductases; Oligodendroglia; Promoter Regions, Genetic; Remyelination; Zebrafish

Longstanding psychological stress has been associated with increased risk of neurodegenerative disorders, such as dementia and Alzheimer's disease. In a prospective population study of women (n = 81), we tested if midlife stress (mean age 49 years) was associated with late-life biomarkers of neurodegeneration in cerebrospinal fluid (CSF) (mean age 74 years) in linear regression models. It was found that women who report of stress at baseline (n = 20) had higher levels of CSF visinin-like protein-1 (VILIP-1) (age adjusted β = 0.113, p = 0.017) and CSF myelin basic protein (β = 0.060, p = 0.030) compared with women without midlife stress (n = 61). There was also a trend observed for higher CSF neurofilament light (β = 0.133, p = 0.056). In addition, longer periods of stress (i.e., stress at 2-3 midlife examinations) were associated with higher levels of CSF VILIP-1. The results suggest that longstanding stress might be associated with neurodegenerative processes in the brain, as CSF VILIP-1 is an unspecific marker for neuronal injury and CSF myelin basic protein reflects neuroaxonal demyelination.

Topics: Aged; Aging; Alzheimer Disease; Axons; Biomarkers; Brain; Dementia; Demyelinating Diseases; Female; Follow-Up Studies; Humans; Middle Aged; Myelin Basic Protein; Nerve Degeneration; Neurocalcin; Neurodegenerative Diseases; Neurofilament Proteins; Risk; Stress, Psychological; Time Factors

Topics: Alzheimer Disease; Amyloid beta-Peptides; Animals; Calcium; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Cell Differentiation; Cell Proliferation; Cell Survival; Cells, Cultured; Demyelinating Diseases; Integrin beta1; Myelin Basic Protein; Oligodendroglia; Organoids; Proto-Oncogene Proteins c-fyn; Rats; Rats, Sprague-Dawley; Signal Transduction

Myelin imaging in the central nervous system is essential for monitoring pathologies involving white matter alterations. Various quantitative MRI protocols relying on the modeling of the interactions of water protons with myelinated tissues have shown sensitivities in case of myelin disruption. Some extracted model parameters are more sensitive to demyelination, such as the bound pool fraction (f) in quantitative magnetization transfer imaging (qMTI), the radial diffusivity in diffusion tensor imaging (DTI), and the myelin water fraction (MWF) in myelin water imaging (MWI). A 3D ultrashort echo time (UTE) sequence within an appropriate water suppression condition (Diff-UTE) is also considered for the direct visualization of the myelin semi-solid matrix (Diff-UTE normalized signal; rSPF). In this paper, we aimed at assessing the sensitivities and correlations of the parameters mentioned above to an immuno-histological study of the myelin basic protein (MBP) in a murine model of demyelination at 7 T. We demonstrated a high sensitivity of the MRI metrics to demyelination, and strong Spearman correlations in the corpus callosum between histology, macromolecular proton fraction (ρ>0.87) and Diff-UTE signal (ρ>0.76), but moderate ones with radial diffusivity and MWF (|ρ|<0.70).

Topics: Animals; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Fluorescence; Magnetic Resonance Imaging; Mice, Inbred C57BL; Myelin Basic Protein

Myelin basic protein (MBP) citrullination by peptidylarginine deiminase (PAD) enzymes leads to incomplete protein-lipid bilayer interactions and vulnerability to proteolytic enzymes, resulting in disorganization of the myelin sheath in the central nervous system. Therefore, citrullinated MBP (citMBP) has been suggested as a hallmark of demyelination, but how citMBP is implicated in prion diseases remains unknown. For the first time, we developed mouse monoclonal anti-citMBP IgG1 (clones 1B8, 1H1, and 3C6) and IgM (clone 3G5) antibodies that recognize human citMBP at its R25, R122, and R130 residues and at its C-terminal region (or the corresponding sites in mouse MBP), respectively. Using a biochemical, immunohistochemical, and immunogold-silver staining for electron microscopy techniques, we found that MBP residue R23 (corresponding to human R25) was specifically citrullinated, was stained as intense punctae in the corpus callosum, the striatum, and the cerebellar white matter, and was predominantly localized in disorganized myelin in the brains of scrapie-infected mice. In the brains of Creutzfeldt-Jakob disease (CJD) patients, MBP residues R25, R122, and R130 were markedly citrullinated and were stained as fibrils and punctae. In particular, white matter regions, such as the midbrain and the medulla, exhibited high levels of citMBP compared to other regions. However, the high levels of citMBP were not correlated with PAD2 expression. The clone 3G5 recognized significantly increased expression of the 18.5 kDa and/or 21.5 kDa variants of MBP in prion disease. Our findings suggest that significantly increased levels of citMBP may reflect demyelinating neuropathology, and that these newly developed antibodies may be useful for identifying demyelination.

Topics: Aged; Aged, 80 and over; Animals; Antibodies, Monoclonal; Brain; Central Nervous System; Citrullination; Creutzfeldt-Jakob Syndrome; Demyelinating Diseases; Female; Humans; Male; Mice; Middle Aged; Myelin Basic Protein; Myelin Sheath; Prion Diseases; Scrapie; White Matter

Myelin abnormalities, oligodendrocyte damage, and concomitant glia activation are common in demyelinating diseases of the central nervous system (CNS). Increasing evidence has demonstrated that the inflammatory response triggers demyelination and gliosis in demyelinating disorders. Numerous clinical interventions, including those used to treat multiple sclerosis (MS), have confirmed prednisone (PDN) as a powerful anti-inflammatory drug that reduces the inflammatory response and promotes tissue repair in multiple inflammation sites. However, the underlying mechanism of PDN in ameliorating myelin damage is not well understood. In our study, a cuprizone (CPZ)-induced demyelinated mouse model was used to explore the mechanism of the protection provided by PDN. Open-field tests showed that CPZ-treated mice exhibited significantly increased anxiety and decreased exploration. However, PDN improved emotional behavior, as evidenced by an increase in the total distance traveled, and central distance traveled as well as the mean amount of time spent in the central area. CPZ-induced demyelination was observed to be alleviated in PDN-treated mice based on luxol fast blue (LFB) staining and myelin basic protein (MBP) expression analyses. In addition, PDN reduced astrocyte and microglia activation in the corpus callosum. Furthermore, we demonstrated that PDN inhibited the Nod-like receptor pyrin domain containing 3 (NLRP3) inflammasome signaling pathway and related inflammatory cytokines and chemokines, including TNF-α, CCL8, CXCL10 and CXCL16. PDN also reduced the serum corticosterone levels in the CPZ-treated mice. Taken together, these results suggest that inhibition of the NLRP3 signaling pathway may be a novel mechanism by which PDN exerts its protective actions in demyelinating diseases.

Topics: Animals; Astrocytes; Corpus Callosum; Cuprizone; Cytokines; Demyelinating Diseases; Disease Models, Animal; Inflammasomes; Male; Mice; Mice, Inbred C57BL; Multiple Sclerosis; Myelin Basic Protein; Myelin Sheath; NLR Family, Pyrin Domain-Containing 3 Protein; Oligodendroglia; Prednisone; Signal Transduction

Sciatic nerve chronic constriction injury (CCI) in rodents produces nerve demyelination via proteolysis of myelin basic protein (MBP), the major component of myelin sheath. Proteolysis releases the cryptic MBP epitope, a demyelination marker, which is hidden in the native MBP fold. It has never been established if the proteolytic release of this cryptic MBP autoantigen stimulates the post-injury increase in the respective circulating autoantibodies. To measure these autoantibodies, we developed the ELISA that employed the cryptic 84-104 MBP sequence (MBP84-104) as bait. This allowed us, for the first time, to quantify the circulating anti-MBP84-104 autoantibodies in rat serum post-CCI. The circulating IgM (but not IgG) autoantibodies were detectable as soon as day 7 post-CCI. The IgM autoantibody level continually increased between days 7 and 28 post-injury. Using the rat serum samples, we established that the ELISA intra-assay (precision) and inter-assay (repeatability) variability parameters were 2.87% and 4.58%, respectively. We also demonstrated the ELISA specificity by recording the autoantibodies to the liberated MBP84-104 epitope alone, but not to intact MBP in which the 84-104 region is hidden. Because the 84-104 sequence is conserved among mammals, we tested if the ELISA was applicable to detect demyelination and quantify the respective autoantibodies in humans. Our limited pilot study that involved 16 female multiple sclerosis and fibromyalgia syndrome patients demonstrated that the ELISA was efficient in measuring both the circulating IgG- and IgM-type autoantibodies in patients exhibiting demyelination. We believe that the ELISA measurements of the circulating autoantibodies against the pathogenic MBP84-104 peptide may facilitate the identification of demyelination in both experimental and clinical settings. In clinic, these measurements may assist neurologists to recognize patients with painful neuropathy and demyelinating diseases, and as a result, to personalize their treatment regimens.

Topics: Animals; Autoantibodies; Autoantigens; Biomarkers; Demyelinating Diseases; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Epitopes; Female; Humans; Multiple Sclerosis; Myelin Basic Protein; Peptide Fragments; Polyradiculoneuropathy; Rats; Rats, Sprague-Dawley; Sciatic Nerve; Sensitivity and Specificity

Animal models of scopolamine-induced amnesia are widely used to study underlying mechanisms and treatment of cognitive impairment in neurodegenerative diseases such as Alzheimer's disease (AD). Previous studies have identified that melatonin improves cognitive dysfunction in animal models. In this study, using a mouse model of scopolamine-induced amnesia, we assessed spatial and short-term memory functions for 4 weeks, investigated the expression of myelin-basic protein (MBP) in the dentate gyrus, and examined whether melatonin and scopolamine cotreatment could keep cognitive function and MBP expression. In addition, to study functions of melatonin for keeping cognitive function and MBP expression, we examined expressions of brain-derived neurotrophic factor (BDNF) and tropomycin receptor kinase B (TrkB) in the mouse dentate gyrus. Scopolamine (1 mg/kg) and melatonin (10 mg/kg) were intraperitoneally treated for 2 and 4 weeks. Two and 4 weeks after scopolamine treatment, mice showed significant cognitive impairment; however, melatonin and scopolamine cotreatment recovered cognitive impairment. Two and 4 weeks of scopolamine treatment, the density of MBP immunoreactive myelinated nerve fibers was significantly decreased in the dentate gyrus; however, scopolamine and melatonin cotreatment significantly increased the scopolamine-induced reduction of MBP expression in the dentate gyrus. Furthermore, the cotreatment of scopolamine and melatonin significantly increased the scopolamine-induced decrease of BDNF and TrKB immunoreactivity in the dentate gyrus. Taken together, our results indicate that melatonin treatment exerts anti-amnesic effect and restores the scopolamine-induced reduction of MBP expression through increasing BDNF and TrkB expressions in the mouse dentate gyrus.

Topics: Animals; Antioxidants; Brain-Derived Neurotrophic Factor; Cognition; Cognitive Dysfunction; Demyelinating Diseases; Dentate Gyrus; Gene Expression Regulation; Male; Melatonin; Membrane Glycoproteins; Mice; Myelin Basic Protein; Receptor, trkB; Scopolamine; Signal Transduction

Oligodendrocytes (OLs) are myelinating glial cells that form myelin sheaths around axons to ensure rapid and focal conduction of action potentials. Here, we found that an axonal outgrowth regulatory molecule, AATYK (apoptosis-associated tyrosine kinase), was up-regulated with OL differentiation and remyelination. We therefore studied its role in OL differentiation. The results showed that AATYK knockdown inhibited OL differentiation and the expression of myelin genes in vitro. Moreover, AATYK-deficiency maintained the proliferation status of OLs but did not affect their survival. Thus, AATYK is essential for the differentiation of OLs.

Topics: Animals; Animals, Newborn; Apoptosis Regulatory Proteins; Cell Differentiation; Cell Proliferation; Cells, Cultured; Cuprizone; Demyelinating Diseases; Embryo, Mammalian; Gene Expression Regulation, Developmental; Ki-67 Antigen; Mice; Mice, Inbred C57BL; Myelin Basic Protein; Myelin Proteolipid Protein; Myelin Sheath; Oligodendroglia; Protein-Tyrosine Kinases; Rats; Rats, Sprague-Dawley; RNA, Small Interfering

BackgroundPelizaeus Merzbacher disease (PMD) is a dysmyelinating disorder of the central nervous system caused by impaired differentiation of oligodendrocytes. This study was prompted by findings that antimuscarinic compounds enhance oligodendrocyte differentiation and remyelination in vitro. One of these compounds, clemastine fumarate, is licensed for treatment of allergic conditions. We tested whether clemastine fumarate can promote myelination in two rodent PMD models, the myelin-deficient and the PLP transgenic rat.MethodsPups were treated with daily injections of clemastine (10-30 mg/kg/day) on postnatal days 1-21. Neurologic phenotypes and myelination patterns in the brain, optic nerves, and spinal cords were assessed using histological techniques.ResultsNo changes in neurological phenotype or survival were observed even at the highest dose of clemastine. Postmortem staining with Luxol fast blue and myelin basic protein immunohistochemistry revealed no evidence for improved myelination in the CNS of treated rats compared to vehicle-treated littermates. Populations of mature oligodendrocytes were unaffected by the treatment.ConclusionThese results demonstrate lack of therapeutic effect of clemastine in two rat PMD models. Both models have rapid disease progression consistent with the connatal form of the disease. Further studies are necessary to determine whether clemastine bears a therapeutic potential in milder forms of PMD.

Topics: Animals; Animals, Genetically Modified; Animals, Newborn; Blood-Brain Barrier; Brain; Cell Differentiation; Central Nervous System; Central Nervous System Diseases; Clemastine; Demyelinating Diseases; Disease Models, Animal; Genotype; Injections, Subcutaneous; Male; Myelin Basic Protein; Myelin Sheath; Oligodendroglia; Optic Nerve; Phenotype; Rats; Spinal Cord

Methods to promote myelin regeneration in response to central myelin loss are essential to prevent the progression of clinical disability in demyelinating diseases. The neurotrophin brain-derived neurotrophic factor (BDNF) is known to promote myelination during development via oligodendrocyte expressed TrkB receptors. Here, we use a structural mimetic of BDNF to promote myelin regeneration in a preclinical mouse model of central demyelination. In female mice, we show that selective targeting of TrkB with the BDNF-mimetic enhances remyelination, increasing oligodendrocyte differentiation, the frequency of myelinated axons, and myelin sheath thickness after a demyelinating insult. Treatment with exogenous BDNF exerted an attenuated effect, increasing myelin sheath thickness only. Further, following conditional deletion of TrkB from premyelinating oligodendrocytes, we show the effects of the BDNF-mimetic on oligodendrocyte differentiation and remyelination are lost, indicating these are dependent on oligodendrocyte expression of TrkB. Overall, these studies demonstrate that targeting oligodendrocyte TrkB promotes

Topics: Animals; Brain; Brain-Derived Neurotrophic Factor; Cell Division; Corpus Callosum; Cuprizone; Demyelinating Diseases; Female; Infusion Pumps, Implantable; Infusions, Intraventricular; Male; Membrane Glycoproteins; Mice; Mice, Inbred C57BL; Mice, Knockout; Molecular Targeted Therapy; Myelin Basic Protein; Myelin Sheath; Neural Stem Cells; Oligodendroglia; Peptides, Cyclic; Phosphorylation; Protein Processing, Post-Translational; Protein-Tyrosine Kinases; Remyelination; Specific Pathogen-Free Organisms

Multiple sclerosis (MS) is a chronic autoimmune disease of the central nervous system (CNS) for which several new treatment options were recently introduced. Among them is the monoclonal anti-CD52 antibody alemtuzumab that depletes mainly B cells and T cells in the immune periphery. Considering the ongoing controversy about the involvement of B cells and in particular the formation of B cell aggregates in the brains of progressive MS patients, an in-depth understanding of the effects of anti-CD52 antibody treatment on the B cell compartment in the CNS itself is desirable.. We used myelin basic protein (MBP)-proteolipid protein (PLP)-induced experimental autoimmune encephalomyelitis (EAE) in C57BL/6 (B6) mice as B cell-dependent model of MS. Mice were treated intraperitoneally either at the peak of EAE or at 60 days after onset with 200 μg murine anti-CD52 vs. IgG2a isotype control antibody for five consecutive days. Disease was subsequently monitored for 10 days. The antigen-specific B cell/antibody response was measured by ELISPOT and ELISA. Effects on CNS infiltration and B cell aggregation were determined by immunohistochemistry. Neurodegeneration was evaluated by Luxol Fast Blue, SMI-32, and Olig2/APC staining as well as by electron microscopy and phosphorylated heavy neurofilament serum ELISA.. Treatment with anti-CD52 antibody attenuated EAE only when administered at the peak of disease. While there was no effect on the production of MP4-specific IgG, the treatment almost completely depleted CNS infiltrates and B cell aggregates even when given as late as 60 days after onset. On the ultrastructural level, we observed significantly less axonal damage in the spinal cord and cerebellum in chronic EAE after anti-CD52 treatment.. Anti-CD52 treatment abrogated B cell infiltration and disrupted existing B cell aggregates in the CNS.

Topics: Animals; Antibodies; Antigen-Presenting Cells; Antigens, CD; B-Lymphocytes; CD52 Antigen; Central Nervous System; Demyelinating Diseases; Disease Models, Animal; Encephalomyelitis, Autoimmune, Experimental; Female; Flow Cytometry; Freund's Adjuvant; Leukocyte Common Antigens; Mice; Mice, Inbred C57BL; Myelin Basic Protein; Myelin Proteolipid Protein; Neurofilament Proteins; Oligodendrocyte Transcription Factor 2; Time Factors

Oligodendrocytes are a type of glial cells that ensheath multiple neuronal axons and form myelin. Under pathological conditions, such as multiple sclerosis (MS), inflammatory damage to myelin and oligodendrocytes leads to demyelination. Although the demyelinated regions can partially resolve functional deficits through remyelination, however, as the disease progresses, remyelination typically becomes incomplete and ultimately fails. One possible explanation for this failure is the activation of the Notch pathway in MS lesions, which impedes oligodendrocyte precursor cells (OPCs) at maturation. This leads to a potential target for remyelination. Here, we have identified a compound Yhhu4952 that promoted the maturation of cultured OPCs in a dose-dependent and time-dependent manner. Neonatal rats showed a significant increase in the expression of myelin basic protein (MBP) and the prevalence of mature oligodendrocytes in the corpus callosum after Yhhu4952 treatment. The compound was also effective in promoting remyelination in cuprizone-induced demyelination model and improving severity scores in experimental autoimmune encephalomyelitis (EAE) model. Mechanism studies revealed that Yhhu4952 promotes OPC differentiation through the inhibition of the Jagged1-Notch1 pathway. These findings suggest Yhhu4952 is potentially useful for proceeding oligodendrocyte differentiation and remyelination.

Topics: Animals; Cell Differentiation; Cells, Cultured; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Dose-Response Relationship, Drug; Myelin Basic Protein; Myelin Sheath; Oligodendroglia; Quinazolines; Rats; Remyelination; Signal Transduction; Time Factors

Intraventricular hemorrhage (IVH) is a frequent complication of preterm newborns, resulting in cerebral palsy and cognitive handicap as well as hypoxic ischemic encephalopathy and periventricular leukomalacia. In this study, we investigated the restorative effect on neonatal IVH by umbilical cord-derived mesenchymal stromal cells (UC-MSCs) cultured in serum-free medium (RM medium) for clinical application. UC-MSCs were cultured with αMEM medium supplemented with FBS or RM. A neonatal IVH mouse model at postnatal day 5 was generated by intraventricular injection of autologous blood, and mice were intravenously administered 1×10

Topics: Adipocytes; Animals; Animals, Newborn; Cell Differentiation; Cerebral Intraventricular Hemorrhage; Culture Media, Serum-Free; Demyelinating Diseases; Disease Models, Animal; Female; Gene Expression Regulation, Developmental; Gliosis; Intercellular Signaling Peptides and Proteins; Lung; Male; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Mice; Myelin Basic Protein; Nerve Tissue Proteins; Umbilical Cord

Failure of oligodendrocyte precursor cells (OPCs) to differentiate and remyelinate axons is thought to be a major cause of the limited ability of the central nervous system to repair plaques of immune-mediated demyelination in multiple sclerosis (MS). Current therapies for MS aim to lessen the immune response in order to reduce the frequency and severity of attacks, but these existing therapies do not target remyelination or stimulate repair of the damaged tissue. Thus, the promotion of OPC differentiation and remyelination is potentially an important therapeutic goal. Previous studies have shown that a recombinant human-derived monoclonal IgM antibody, designated rHIgM22, promotes remyelination, particularly of the spinal cord in rodent models of demyelination. Here, we examined the effects of rHIgM22 in remyelination in the brain using the mouse model of cuprizone-induced demyelination, which is characterized by spontaneous remyelination. The myelination state of the corpus callosum of cuprizone-fed mice treated with rHIgM22 was examined immediately after the end of the cuprizone diet as well as at different time points during the recovery period with regular food, and compared with that of cuprizone-fed animals treated with either vehicle or human IgM isotype control antibody. Mice fed only regular food were used as controls. We demonstrate that treatment with rHIgM22 accelerated remyelination of the demyelinated corpus callosum. The remyelination-enhancing effects of rHIgM22 were found across different, anatomically distinct regions of the corpus callosum, and followed a spatiotemporal pattern that was similar to that of the spontaneous remyelination process. These enhancing effects were also accompanied by increased differentiation of OPCs into mature oligodendrocytes. Our data indicate strong remyelination-promoting capabilities of rHIgM22 and further support its therapeutic potential in MS.

Topics: Analysis of Variance; Animals; Cell Differentiation; Corpus Callosum; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Immunoglobulin M; Male; Mice; Mice, Inbred C57BL; Monoamine Oxidase Inhibitors; Myelin Basic Protein; Myelin Sheath; Oligodendrocyte Transcription Factor 2; Oligodendroglia; Optic Nerve; Receptor, Platelet-Derived Growth Factor alpha; Remyelination; Time Factors

Cerebellar pathology is a frequent feature of multiple sclerosis (MS), a demyelinating and neuroinflammatory disease of the central nervous system (CNS). Interleukin (IL)-6 is a multifunctional cytokine with a potential role in MS. Here we studied cuprizone-induced cerebellar pathology in transgenic mice with astrocyte-targeted production of IL-6 (GFAP-IL6), specifically focusing on demyelination, oligodendrocyte depletion and microglial cell response.. Over the course of cuprizone treatment, when compared with WT mice, GFAP-IL6Tg showed a reduced demyelination in the deep lateral cerebellar nuclei (LCN). The oligodendrocyte numbers in the LCN were comparable between WT and GFAP-IL6Tg mice after 4-6weeks of cuprizone treatment, however after the chronic cuprizone treatment (12weeks) we detected higher numbers of oligodendrocytes in GFAP-IL6Tg mice. Contrary to strong cuprizone-induced microglial activation in the LCN of WT mice, GFAP-IL6Tg mice had minimal cuprizone-induced microglial changes, despite an already existing reactive microgliosis in control GFAP-IL6Tg not present in control WT mice.. Our results show that chronic transgenic production of IL-6 reduced cuprizone-induced cerebellar demyelination and induced a specific activation state of the resident microglia population (Iba1

Topics: Animals; Astrocytes; Calcium-Binding Proteins; Cerebellum; Cuprizone; Cytokines; Demyelinating Diseases; Disease Models, Animal; Gene Expression Regulation; Glial Fibrillary Acidic Protein; Interleukin-6; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Microfilament Proteins; Microglia; Myelin Basic Protein; Time Factors

Fractalkine/CX3CR1 signalling has been implicated in many neurodegenerative and neurological diseases of the central nervous system (CNS). This signalling pathway plays an important role in regulating reactive oxygen species (ROS), as well as itself being altered in conditions of oxidative stress. Here, we investigated the effects of recombinant fractalkine (rCX3CL1) in models of hydrogen peroxide (H. Organotypic cerebellar slice cultures were generated from postnatal day 10 C57BL/6J mice to assess myelination. Immunohistochemistry was used to measure the degree of myelination. Fluorescent images were obtained using a leica SP8 confocal microscope and data analysed using ImageJ software.. We show here, for the first time, that rCX3CL1 significantly attenuated bolus H. This data indicates possible protective effects of fractalkine signalling in oxidative stress-induced demyelination in the central nervous system. This opens up the possibility of fractalkine receptor (CX3CR1) modulation as a potential new target for protecting against oxidative stress-induced demyelination in both inflammatory and non-inflammatory nervous system disorders.

Topics: Animals; Animals, Newborn; Catalase; Cell Death; Cerebellum; Chemokine CX3CL1; CX3C Chemokine Receptor 1; Demyelinating Diseases; Female; Gliosis; Glucose Oxidase; Hydrogen Peroxide; Male; Mice; Mice, Inbred C57BL; Myelin Basic Protein; Myelin-Oligodendrocyte Glycoprotein; Neurofilament Proteins; Organ Culture Techniques; Reactive Oxygen Species

Myelin is a specialized multilamellar structure involved in various functions of the nervous system. Oligodendrocytes are responsible for myelin formation in the central nervous system. Motor proteins play important roles in differentiation and myelin formation of the oligodendrocyte lineage. Recently, we revealed that one of the unconventional myosins, myosin ID (Myo1d), is expressed in mature oligodendrocytes and is required for myelin-like membrane formation in vitro. Previously, Cahoy et al. (J Neurosci 28:264-278, 2008) reported that another unconventional myosin VI (Myo6) is upregulated in transcriptome data of differentiated oligodendrocytes. However, it is uncertain whether Myo6 protein is present in oligodendrocytes. In this study, to analyze expression of Myo6 in oligodendrocytes, we performed immunofluorescence analysis on brains of adult normal and cuprizone-induced demyelination mice. Myo6 expression was detected in mature oligodendrocytes and oligodendrocyte progenitor cells in the cerebellum and corpus callosum. To compare temporal expression patterns of myosin superfamily members in vitro, double immunostainings using anti-Myo6, myosin Va (Myo5a), or Myo1d with each stage-specific oligodendrocyte marker antibody were performed. In cultured oligodendrocytes, although Myo1d was found only in mature oligodendrocytes, Myo6 and Myo5a signals were detected in all stages of differentiation, from oligodendrocyte progenitor cells to mature oligodendrocytes. Additionally, similar to Myo5a, Myo6-positive signals were confined to the cell body and processes. These results showed that Myo6 is one of the unconventional myosins in oligodendrocyte lineage cells, which could play a role in clathrin-related endocytosis.

Topics: Animals; Cell Differentiation; Cell Lineage; Central Nervous System; Cuprizone; Demyelinating Diseases; Female; Mice, Inbred C57BL; Myelin Basic Protein; Myelin Sheath; Neurogenesis; Neurons; Oligodendroglia

Multiple sclerosis (MS) is a severe autoimmune disease characterized by inflammatory, demyelinating and neurodegenerative components causing motor, sensory, visual and/or cognitive symptoms. The relapsing-remitting MS affecting 85% of patients is reliably mimicked by the proteolipid-protein (PLP)-induced experimental autoimmune encephalomyelitis (EAE) SJL/J-mouse model. Significant progress was made for MS treatment but the development of effective therapies devoid of severe side-effects remains a great challenge. Here, we combine clinical, behavioral, histopathological, biochemical and molecular approaches to demonstrate that low and well tolerated doses (10-20mg/kg) of TSPO ligand XBD173 (Emapunil) efficiently ameliorate clinical signs and neuropathology of PLP-EAE mice. In addition to the conventional clinical scoring of symptoms, we applied the robust behavioral Catwalk-method to confirm that XBD173 (10mg/kg) increases the maximum contact area parameter at EAE-disease peak, indicating an improvement/recovery of motor functions. Consistently, histopathological studies coupled with microscope-cellSens quantification and RT-qPCR analyzes showed that XBD173 prevented demyelination by restoring normal protein and mRNA levels of myelin basic protein that was significantly repressed in PLP-EAE mice spinal cord and brain. Interestingly, ELISA-based measurement revealed that XBD173 increased allopregnanolone concentrations in PLP-EAE mice spinal and brain tissues. Furthermore, flow cytometry assessment demonstrated that XBD173 therapy decreased serum level of pro-inflammatory cytokines, including interleukin-17A, Interleukin-6 and tumor-necrosis-factor alpha in PLP-EAE mice. As the optimal XBD173 dosing exerting the maximal beneficial action in EAE mice is the lower 10mg/kg dose, the paper opens interesting perspectives for the development of efficient and safe therapies against MS with slight or no side-effects.

Topics: Animals; Biomarkers; Brain; Cytokines; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Female; Ligands; Mice; Mice, Inbred Strains; Multiple Sclerosis, Relapsing-Remitting; Myelin Basic Protein; Neurotransmitter Agents; Pregnanolone; Purines; Receptors, GABA; RNA, Messenger; Spinal Cord

A 59-year-old man had been admitted to another hospital because of diplopia and thirst at the beginning of March and was diagnosed with diabetic ketoacidosis. He was referred to our hospital because he had limb weakness, dysarthria, and bilateral sensory impairment of the upper limbs, which worsened rapidly from the middle of March, although plasma glucose had been well controlled after the initiation of insulin therapy in the previous hospital. Contrast spinal MRI in our hospital revealed hyperintense lesions at the level of C4 to C5 and T10. The level of myelin basic protein was high (1,260 pg/ml) in the cerebrospinal fluid and serum anti-neurofascin antibody was negative. Nerve conduction study showed typical findings of demyelination at least 2 regions. Although anti-neurofascin antibody was negative, he was diagnosed with combined central and peripheral demyelination (CCPD) based on these clinical findings. After the repeated methylprednisolone pulse therapy for five times, the hyperintense lesions of the spinal cord disappeared gradually. He was bedridden at the beginning of his hospitalization but could ambulate with a cane on discharge 2 months after the admission. Then we received the result of anti-galactocerebroside antibody test as positive. This case suggested that high-dose steroid pulse therapy is safe and may be effective for anti-galactocerebroside antibody-positive CCPD.

Topics: Autoantibodies; Biomarkers; Cell Adhesion Molecules; Central Nervous System Diseases; Demyelinating Diseases; Galactosylceramides; Humans; Immunoglobulin G; Magnetic Resonance Imaging; Male; Methylprednisolone; Middle Aged; Myelin Basic Protein; Nerve Growth Factors; Peripheral Nervous System Diseases; Pulse Therapy, Drug; Treatment Outcome

The G protein-coupled receptor EBI2 (Epstein-Barr virus-induced gene 2) is activated by 7α, 25-dihydroxycholesterol (7α25HC) and plays a role in T cell-dependant antibody response and B cell migration. Abnormal EBI2 signaling is implicated in a range of autoimmune disorders; however, its role in the CNS remains poorly understood.. Here we characterize the role of EBI2 in myelination under normal and pathophysiological conditions using organotypic cerebellar slice cultures and EBI2 knock-out (KO) animals.. We find that MBP expression in brains taken from EBI2 KO mice is delayed compared to those taken from wild type (WT) mice. In agreement with these in vivo findings, we show that antagonism of EBI2 reduces MBP expression in vitro. Importantly, we demonstrate that EBI2 activation attenuates lysolecithin (LPC)-induced demyelination in mouse organotypic slice cultures. Moreover, EBI2 activation also inhibits LPC-mediated release of pro-inflammatory cytokines such as IL6 and IL1β in cerebellar slices.. These results, for the first time, display a role for EBI2 in myelin development and protection from demyelination under pathophysiological conditions and suggest that modulation of this receptor may be beneficial in neuroinflammatory and demyelinating disorders such as multiple sclerosis.

Topics: Animals; Cerebellum; Demyelinating Diseases; Lysophosphatidylcholines; Mice; Mice, Knockout; Myelin Basic Protein; Myelin Sheath; Organ Culture Techniques; Receptors, G-Protein-Coupled

Adverse early-life experience such as depriving the relationship between parents and children induces permanent phenotypic changes, and impairs the cognitive functions associated with the prefrontal cortex (PFC). However, the underlying mechanism remains unclear. In this work, we used rat neonatal maternal separation (NMS) model to illuminate whether and how NMS in early life affects cognitive functions, and what the underlying cellular and molecular mechanism is. We showed that rat pups separated from their dam 3 h daily during the first 3 postnatal weeks alters medial prefrontal cortex (mPFC) myelination and impairs mPFC-dependent behaviors. Myelination appears necessary for mPFC-dependent behaviors, as blockade of oligodendrocytes (OLs) differentiation or lysolecithin-induced demyelination, impairs mPFC functions. We further demonstrate that histone deacetylases 1/2 (HDAC1/2) are drastically reduced in NMS rats. Inhibition of HDAC1/2 promotes Wnt activation, which negatively regulates OLs development. Conversely, selective inhibition of Wnt signaling by XAV939 partly rescue myelination arrestment and behavior deficiency caused by NMS. These findings indicate that NMS impairs mPFC cognitive functions, at least in part, through modulation of oligodendrogenesis and myelination. Understanding the mechanism of NMS on mPFC-dependent behaviors is critical for developing pharmacological and psychological interventions for child neglect and abuse.

Topics: Animals; Animals, Newborn; Anxiety; Cognition Disorders; Demyelinating Diseases; Enzyme Inhibitors; Exploratory Behavior; Gene Expression Regulation, Developmental; Heterocyclic Compounds, 3-Ring; Histone Deacetylases; Lipopolysaccharides; Maternal Deprivation; Maze Learning; Myelin Basic Protein; Neurogenesis; Prefrontal Cortex; Rats; Rats, Sprague-Dawley; Valproic Acid; Wnt Proteins; Wnt Signaling Pathway

There is compelling evidence that microglial activation negatively impacts neurogenesis. However, microglia have also been shown to promote recruitment of newly born neurons to injured areas of the gray matter. In the present study, we explored whether demyelination-triggered inflammation alters the process of neurogenesis in the white matter. A 2-μl solution of 0.04 % ethidium bromide was stereotaxically injected into the corpus callosum of adult male rats. Brain inflammation was dampened by daily injections of progesterone (5 mg/kg, s.c.) for 14 days. Control rats received oil (s.c.). Newly born neurons (DCX and Tbr2), microglia (Iba-1), astrocytes (vimentin or GFAP), oligodendrocyte progenitor cells (OPCs; NG2), and mature oligodendrocytes (CC-1) were monitored in the vicinity of demyelination site using immunofluorescent staining. Western blot was used to explore microglial polarization using M1 (iNOS) and M2 (arginase-1) markers. Focal demyelination elicited strong microglial and astroglial activation and reduced the number of OPCs at the site of demyelination. This inflammatory response was associated with enhanced number of newly born neurons in the white matter and the subventricular zone (SVZ). A proportion of newly born neurons within the white matter showed features of OPCs. Interestingly, blunting brain inflammation led to reduced neurogenesis around the demyelination area and in the SVZ. These data suggest that the white matter inflammation creates a conducive environment for the recruitment of newly born neurons. The fact that a sizable fraction of these newly born neurons adopt OPC features suggests that they could contribute to the remyelination process.

Topics: Animals; Astrocytes; Cell Differentiation; Corpus Callosum; Demyelinating Diseases; Doublecortin Protein; Inflammation; Male; Microglia; Myelin Basic Protein; Myelin Sheath; Neurons; Oligodendroglia; Progesterone; Rats, Sprague-Dawley; White Matter

The conditional Cre/lox system has recently emerged as a valuable tool for studies on both embryonic and adult Zebrafish. Temporal control and site-specific recombination are achieved by using the ligand-inducible CreER. Here we report the generation of a transgenic Zebrafish line, which expresses an mCherry-tagged variant of CreER. The transgenic Zebrafish line Tg(mbpa:mCherry-T2A-CreER

Topics: Animals; Animals, Genetically Modified; Demyelinating Diseases; Gene Expression Regulation, Developmental; Genes, Reporter; Integrases; Myelin Basic Protein; Myelin Sheath; Oligodendroglia; Promoter Regions, Genetic; Recombination, Genetic; Transgenes; Zebrafish

It is now accepted that white matter abnormalities play an important role in demyelinating diseases and a wide range of psychiatric disorders. Experimental demyelination (especially induced by cuprizone) has been investigated extensively. However, details regarding demyelination and ultrastructural changes of myelinated fibers have not been previously reported. Therefore, we determined the extent of demyelination using quantitative stereology. Mice exposed to cuprizone in the current study showed abnormal anxiety-like behavior without impaired spatial learning or memory. The myelinated fibers in whole corpus callosum of mice exposed to cuprizone showed extensive myelin deficiencies and occasional axonal injuries. The total length of the myelinated fibers in whole corpus callosum of mice exposed to cuprizone was significantly decreased by 45% compared with control mice. The loss of myelinated fibers was mainly due to the marked loss of the fibers with a diameter of 0.4 to 0.8 μm. The g-ratio of the myelinated fibers in the corpus callosum of mice exposed to cuprizone (0.69 ± 0.02) was significantly decreased compared with control mice (0.76 ± 0.02). These results might help us to further understand the role of white matter abnormalities in demyelinating diseases or a wide range of psychiatric disorders. © 2016 Wiley Periodicals, Inc.

Topics: Animals; Chelating Agents; Corpus Callosum; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Exploratory Behavior; Male; Maze Learning; Mice; Mice, Inbred C57BL; Microscopy, Electron, Transmission; Myelin Basic Protein; Nerve Fibers, Myelinated; Rotarod Performance Test

Animal models, such as cuprizone (bis-cyclohexanone oxaldihydrazone) feeding, are helpful to study experimental demyelination and remyelination in the context of diseases like multiple sclerosis. Cuprizone is a copper chelator, which when supplemented to the normal food of C57BL/6J mice in a concentration of 0.2% leads to oligodendroglial loss, subsequent microglia and astrocyte activation, resulting in demyelination. Termination of the cuprizone diet results in remyelination, promoted by newly formed mature oligodendrocytes. The exact mode of cuprizone's action is not well understood, and information about its inactivation and cleavage are still not available. The knowledge of these processes could lead to a better understanding of cuprizone's mode of action, as well as a safer handling of this toxin. We therefore performed experiments with the aim to inactivate cuprizone by thermal heating, since it was suggested in the past that cuprizone is heat sensitive. C57BL/6J mice were fed for 4 weeks with 0.2% cuprizone, either thermally pretreated (60, 80, 105, 121 °C) or not heated. In addition, primary rat oligodendrocytes, as a known selective toxic target of cuprizone, were incubated with 350 μM cuprizone solutions, which were either thermally pretreated or not. Our results demonstrate that none of the tested thermal pretreatment conditions could abrogate or restrict the toxic and demyelinating effects of cuprizone, neither in vitro nor in vivo. In conclusion, the current study rebuts the hypothesis of cuprizone as a heat-sensitive compound, as well as the assumption that heat exposure is a reason for an insufficient demyelination of cuprizone-containing pellets.

Topics: Adenomatous Polyposis Coli Protein; Agglutinins; Animals; Cell Differentiation; Cerebral Cortex; Corpus Callosum; Cuprizone; Demyelinating Diseases; Glial Fibrillary Acidic Protein; Hot Temperature; Mice; Myelin Basic Protein; Oligodendroglia; Primary Cell Culture; Rats

The therapeutic effect of glial progenitor transplantation in diseases of dysmyelination is currently attributed to the formation of new myelin. Using magnetic resonance imaging (MRI), we show that the therapeutic outcome in dysmyelinated shiverer mice is dependent on the extent of cell migration but not the presence of mature and compact myelin. Human or mouse glial restricted progenitors (GRPs) were transplanted into rag2

Topics: Animals; Basic Helix-Loop-Helix Transcription Factors; Brain; Cell Differentiation; Cell Movement; Cell Survival; Demyelinating Diseases; Disease Models, Animal; DNA-Binding Proteins; Gangliosides; Glial Fibrillary Acidic Protein; Humans; Mice; Mice, Transgenic; Myelin Basic Protein; Myelin Proteolipid Protein; Myelin Sheath; Nerve Tissue Proteins; Neuroglia; Oligodendrocyte Transcription Factor 2; Spinal Cord; Stem Cell Transplantation; Time Factors; Tubulin

The abnormal expression and function of myelin-related proteins contribute to nervous system dysfunction associated with neuropsychiatric disorders; however, the underlying mechanism of this remains unclear. We found here that breast carcinoma amplified sequence 1 (BCAS1), a basic protein abundant in the brain, was expressed specifically in oligodendrocytes and Schwann cells, and that its expression level was decreased by demyelination. This suggests that BCAS1 is a novel myelin-associated protein. BCAS1 knockout mice displayed schizophrenia-like behavioral abnormalities and a tendency toward reduced anxiety-like behaviors. Moreover, we found that the loss of BCAS1 specifically induced hypomyelination and the expression of inflammation-related genes in the brain. These observations provide a novel insight into the functional link between oligodendrocytes and inflammation and/or abnormal behaviors.

Topics: Animals; Brain; Demyelinating Diseases; Inflammation; Mice, Knockout; Myelin Basic Protein; Myelin Sheath; Neoplasm Proteins; Oligodendroglia; Schizophrenia; Transcriptional Activation; Up-Regulation

Rapid and efficient protocols to generate oligodendrocytes (OL) from human induced pluripotent stem cells (iPSC) are currently lacking, but may be a key technology to understand the biology of myelin diseases and to develop treatments for such disorders. Here, we demonstrate that the induction of three transcription factors (SOX10, OLIG2, NKX6.2) in iPSC-derived neural progenitor cells is sufficient to rapidly generate O4

Topics: Animals; Biomarkers; Brain; Cell Death; Cell Differentiation; Cell Lineage; Cells, Cultured; Cluster Analysis; Demyelinating Diseases; Disease Models, Animal; Ectopic Gene Expression; Gene Expression Profiling; Humans; Induced Pluripotent Stem Cells; Mice; Mutation; Myelin Basic Protein; Myelin Sheath; Neural Stem Cells; Oligodendroglia; Oxidative Stress; Spinal Cord; tau Proteins; Transcription Factors; Transcriptome

Oligodendrocyte progenitor cell (OPC) differentiation is an important therapeutic target to promote remyelination in multiple sclerosis (MS). We previously reported hyperphosphorylated and aggregated microtubule-associated protein tau in MS lesions, suggesting its involvement in axonal degeneration. However, the influence of pathological tau-induced axonal damage on the potential for remyelination is unknown. Therefore, we investigated OPC differentiation in human P301S tau (P301S-htau) transgenic mice, both in vitro and in vivo following focal demyelination. In 2-month-old P301S-htau mice, which show hyperphosphorylated tau in neurons, we found atrophic axons in the spinal cord in the absence of prominent axonal degeneration. These signs of early axonal damage were associated with microgliosis and an upregulation of IL-1β and TNFα. Following in vivo focal white matter demyelination we found that OPCs differentiated more efficiently in P301S-htau mice than wild type (Wt) mice. We also found an increased level of myelin basic protein within the lesions, which however did not translate into increased remyelination due to higher susceptibility of P301S-htau axons to demyelination-induced degeneration compared to Wt axons. In vitro experiments confirmed higher differentiation capacity of OPCs from P301S-htau mice compared with Wt mice-derived OPCs. Because the OPCs from P301S-htau mice do not ectopically express the transgene, and when isolated from newborn mice behave like Wt mice-derived OPCs, we infer that their enhanced differentiation capacity must have been acquired through microenvironmental priming. Our data suggest the intriguing concept that damaged axons may signal to OPCs and promote their differentiation in the attempt at rescue by remyelination.

Topics: Animals; Basic Helix-Loop-Helix Transcription Factors; CD11b Antigen; Cell Death; Cell Differentiation; Cell Movement; Cell Proliferation; Demyelinating Diseases; Disease Models, Animal; Gene Expression Regulation; Humans; Mice; Mice, Inbred C57BL; Mice, Transgenic; Mutation; Myelin Basic Protein; Nerve Tissue Proteins; Neurons; Oligodendrocyte Transcription Factor 2; Oligodendroglia; Spinal Cord Injuries; Stem Cells; tau Proteins

Different types of insults to the CNS lead to axon demyelination. Remyelination occurs when the CNS attempts to recover from myelin loss and requires the activation of oligodendrocyte precursor cells. With the rationale that CB1 receptor is expressed in oligodendrocytes and marijuana consumption alters CNS myelination, we study the effects of the cannabinoid agonist WIN55212.2 in (1) an in vitro model of oligodendrocyte differentiation and (2) the cuprizone model for demyelination.. The synthetic cannabinoid agonist WIN55212.2 at 1 μM increased the myelin basic protein mRNA and protein expression in vitro. During cuprizone-induced acute demyelination, the administration of 0.5 mg/kg WIN55212.2 confers more myelinated axons, increased the expression of retinoid X receptor alpha, and declined nogo receptor expression. Controversially, 1 mg/kg of the drug increased the number of demyelinated axons and reduced the expression of nerve growth factor inducible, calreticulin and myelin-related genes coupling specifically with a decrease in 2',3'-cyclic nucleotide 3' phosphodiesterase expression.. The cannabinoid agonist WIN55212.2 promotes oligodendrocyte differentiation in vitro. Moreover, 0.5 mg/kg of the drug confers neuroprotection during cuprizone-induced demyelination, while 1 mg/kg aggravates the demyelination process.

Topics: Animals; Benzoxazines; Cell Differentiation; Cell Line, Transformed; Central Nervous System; Chelating Agents; Corpus Callosum; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Dose-Response Relationship, Drug; Gene Expression Regulation; Male; Mice; Mice, Inbred C57BL; Morpholines; Myelin Basic Protein; Naphthalenes; Neuroprotective Agents; Oligodendroglia; Prepulse Inhibition; Transcriptome

BAF312 (Siponimod) is a dual agonist at the sphingosine-1 phosphate receptors, S1PR1 and S1PR5. This drug is currently undergoing clinical trials for the treatment of secondary progressive multiple sclerosis (MS). Here, we investigated the effects of BAF312 on isolated astrocyte and microglia cultures as well as in slice culture models of demyelination.. Mouse and human astrocytes were treated with S1PR modulators and changes in the levels of pERK, pAkt, and calcium signalling as well as S1PR1 internalization and cytokine levels was investigated using Western blotting, immunochemistry, ELISA and confocal microscopy. Organotypic slice cultures were prepared from the cerebellum of 10-day-old mice and treated with lysophosphatidylcholine (LPC), psychosine and/or S1PR modulators, and changes in myelination states were measured by fluorescence of myelin basic protein and neurofilament H.. BAF312 treatment of human and mouse astrocytes activated pERK, pAKT and Ca(2+) signalling as well as inducing S1PR1 internalization. Notably, activation of S1PR1 increased pERK and pAKT in mouse astrocytes while both S1PR1 and S1PR3 equally increased pERK and pAKT in human astrocytes, suggesting that the coupling of S1PR1 and S1PR3 to pERK and pAKT differ in mouse and human astrocytes. We also observed that BAF312 moderately attenuated lipopolysaccharide (LPS)- or TNFα/IL17-induced levels of IL6 in both astrocyte and microglia cell cultures. In organotypic slice cultures, BAF312 reduced LPC-induced levels of IL6 and attenuated LPC-mediated demyelination. We have shown previously that the toxic lipid metabolite psychosine induces demyelination in organotypic slice cultures, without altering the levels of cytokines, such as IL6. Importantly, psychosine-induced demyelination was also attenuated by BAF312.. Overall, this study suggests that BAF312 can modulate glial cell function and attenuate demyelination, highlighting this drug as a further potential therapy in demyelinating disorders, beyond MS.

Topics: Animals; Animals, Newborn; Anti-Inflammatory Agents; Astrocytes; Azetidines; Benzyl Compounds; beta-Alanine; Calcium Signaling; Cerebellum; Demyelinating Diseases; eIF-2 Kinase; Humans; Immunosuppressive Agents; In Vitro Techniques; Indans; Interleukin-6; Lysophosphatidylcholines; Mice; Myelin Basic Protein; Organ Culture Techniques; Oxadiazoles; Protein Transport; Receptors, Lysosphingolipid; Thiophenes; Time Factors

White matter abnormalities in the CNS have been reported recently in various neurological and psychiatric disorders. Quantitation of non-Gaussianity for water diffusion by q-space diffusional MRI (QSI) renders biological diffusion barriers such as myelin sheaths; however, the time-consuming nature of this method hinders its clinical application. In the current study, we aimed to refine QSI protocols to enable their clinical application and to visualize myelin signals in a clinical setting. For this purpose, animal studies were first performed to optimize the acquisition protocol of a non-Gaussian QSI metric. The heat map of standardized kurtosis values derived from optimal QSI (myelin map) was then created. Histological validation of the myelin map was performed in myelin-deficient mice and in a nonhuman primate by monitoring its variation during demyelination and remyelination after chemical spinal cord injury. The results demonstrated that it was sensitive enough to depict dysmyelination, demyelination, and remyelination in animal models. Finally, its utility in clinical practice was assessed by a pilot clinical study in a selected group of patients with multiple sclerosis (MS). The human myelin map could be obtained within 10 min with a 3 T MR scanner. Use of the myelin map was practical for visualizing white matter and it sensitively detected reappearance of myelin signals after demyelination, possibly reflecting remyelination in MS patients. Our results together suggest that the myelin map, a kurtosis-related heat map obtainable with time-saving QSI, may be a novel and clinically useful means of visualizing myelin in the human CNS.. Myelin abnormalities in the CNS have been gaining increasing attention in various neurological and psychiatric diseases. However, appropriate methods with which to monitor CNS myelin in daily clinical practice have been lacking. In the current study, we introduced a novel MRI modality that produces the "myelin map." The myelin map accurately depicted myelin status in mice and nonhuman primates and in a pilot clinical study of multiple sclerosis patients, suggesting that it is useful in detecting possibly remyelinated lesions. A myelin map of the human brain could be obtained in <10 min using a 3 T scanner and it therefore promises to be a powerful tool for researchers and clinicians examining myelin-related diseases.

Topics: Adult; Animals; Brain Mapping; Callithrix; Demyelinating Diseases; Diffusion Magnetic Resonance Imaging; Disease Models, Animal; Female; Humans; Image Processing, Computer-Assisted; Lysophosphatidylcholines; Male; Mice; Mice, Jimpy; Mice, Mutant Strains; Multiple Sclerosis; Mutation; Myelin Basic Protein; Myelin Sheath; Spinal Cord; White Matter

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.

Topics: Animals; Basic Helix-Loop-Helix Transcription Factors; Brain; Coumarins; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Drug Evaluation, Preclinical; Gene Expression; Injections, Intraperitoneal; Male; Mice, 129 Strain; Mice, Inbred C57BL; Microglia; Myelin Basic Protein; Nerve Tissue Proteins; Neuroprotective Agents; Oligodendrocyte Transcription Factor 2; Oligodendroglia; Receptor, Platelet-Derived Growth Factor alpha; Stem Cells

Hes3 is a component of the STAT3-Ser/Hes3 Signaling Axis controlling the growth and survival of neural stem cells and other plastic cells. Pharmacological activation of this pathway promotes neuronal rescue and behavioral recovery in models of ischemic stroke and Parkinson's disease. Here we provide initial observations implicating Hes3 in the cuprizone model of demyelination and remyelination. We focus on the subpial motor cortex of mice because we detected high Hes3 expression. This area is of interest as it is impacted both in human demyelinating diseases and in the cuprizone model. We report that Hes3 expression is reduced at peak demyelination and is partially restored within 1 week after cuprizone withdrawal. This raises the possibility of Hes3 involvement in demyelination/remyelination that may warrant additional research. Supporting a possible role of Hes3 in the maintenance of oligodendrocyte markers, a Hes3 null mouse strain shows lower levels of myelin basic protein in undamaged adult mice, compared to wild-type controls. We also present a novel method for culturing the established oligodendrocyte progenitor cell line oli-neu in a manner that maintains Hes3 expression as well as its self-renewal and differentiation potential, offering an experimental tool to study Hes3. Based upon this approach, we identify a Janus kinase inhibitor and dbcAMP as powerful inducers of Hes3 gene expression. We provide a new biomarker and cell culture method that may be of interest in demyelination/remyelination research.

Topics: Animals; Basic Helix-Loop-Helix Transcription Factors; Cell Culture Techniques; Culture Media, Conditioned; Cuprizone; Demyelinating Diseases; Gene Expression Regulation; Male; Mice; Mice, Inbred C57BL; Motor Cortex; Myelin Basic Protein; Myelin Sheath; Nerve Tissue Proteins; Repressor Proteins; RNA, Messenger

Both multiple sclerosis (MS) and Alzheimer's disease (AD) are progressive neurological disorders with myelin injury and memory impairment. However, whether myelin impairment could cause AD-like neurological pathology remains unclear. To explore neurological pathology following myelin injury, we assessed cognitive function, the expression of myelin proteins, axonal transport-associated proteins, axonal structural proteins, synapse-associated proteins, tau and beta amyloid and the status of neurons, using the cuprizone mouse model of demyelination. We found the mild impairment of learning ability in cuprizone-fed mice and the decreased expression of myelin basic protein (MBP) in the hippocampus. And anti-LINGO-1 improved learning ability and partly restored MBP level. Furthermore, we also found kinesin light chain (KLC), neurofilament light chain (NFL) and neurofilament heavy chain (NF200) were declined in demyelinated hippocampus, which could be partly improved by treatment with anti-LINGO-1. However, we did not observe the increased expression of beta amyloid, hyperphosphorylation of tau and loss of neurons in demyelinated hippocampus. Our results suggest that demyelination might lead to the impairment of neuronal transport, but not cause increased level of hyperphosphorylated tau and beta amyloid. Our research demonstrates remyelination might be an effective pathway to recover the function of neuronal axons and cognition in MS.

Topics: Alzheimer Disease; Amyloid beta-Peptides; Animals; Antibodies, Monoclonal; Axonal Transport; Axons; Cognition; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Fluorescent Antibody Technique; Hippocampus; Humans; Kinesins; Male; Maze Learning; Membrane Proteins; Mice; Mice, Inbred C57BL; Microtubule-Associated Proteins; Myelin Basic Protein; Myelin Sheath; Nerve Tissue Proteins; Neurofilament Proteins; Phosphorylation; tau Proteins

Multiple sclerosis (MS) is a chronic demyelinating disease of the central nervous system. Interleukin (IL)-6 is a pleiotropic cytokine with a potential role in MS. Here we used transgenic mice with astrocyte-targeted production of IL-6 (GFAP-IL6Tg) to study the effect of IL-6 in the cuprizone-induced demyelination paradigm, which is an experimental model of de- and re-myelination, both hallmarks of MS. Our results demonstrated that cuprizone-treated GFAP-IL6Tg mice showed a significant reduction in astroglial and especially microglial activation/accumulation in the corpus callosum in comparison with the corresponding cuprizone-treated wild type (WT). Production of a key microglial attracting chemokine CXCL10, as well as CXCL1 and CCL4 was lower in cuprizone-treated GFAP-IL6Tg mice compared with cuprizone-treated WT. Reduced microglial cell accumulation was associated with inefficient removal of degraded myelin and axonal protection in cuprizone-treated GFAP-IL6Tg mice, compared with WT mice at the peak of demyelination. In addition, transgenic production of IL-6 did not alter initial oligodendrocyte (OL) apoptosis and oligodendrocyte precursor cell recruitment to the lesion site, but it impaired early OL differentiation, possibly due to impaired removal of degraded myelin. Indeed, a microglial receptor involved in myelin phagocytosis, TREM2, as well as the phagolysosomal protein CD68 were lower in cuprizone-treated GFAP-IL6Tg compared with WT mice. Our results show for the first time that astrocyte-targeted production of IL-6 may play a role in modulating experimental demyelination induced by cuprizone. Further understanding of the IL-6-mediated molecular mechanisms involved in the regulation of demyelination is needed, and may have implications for the development of future therapeutic strategies for the treatment of MS. GLIA 2016;64:2104-2119.

Topics: Amyloid beta-Protein Precursor; Animals; Astrocytes; Calcium-Binding Proteins; Caspase 3; Cuprizone; Cytokines; Demyelinating Diseases; Disease Models, Animal; Green Fluorescent Proteins; Interleukin-6; Male; Membrane Glycoproteins; Mice; Mice, Inbred C57BL; Mice, Transgenic; Microfilament Proteins; Microglia; Monoamine Oxidase Inhibitors; Myelin Basic Protein; Myelin Sheath; Nerve Tissue Proteins; Receptors, Immunologic

Multiple sclerosis is a chronic inflammatory, demyelinating degenerative disease of the central nervous system. Current treatments target pathological immune responses to counteract the inflammatory processes. However, these drugs do not restrain the long-term progression of clinical disability. For this reason, new therapeutic approaches and identification of novel target molecules are needed to prevent demyelination or promote repair mechanisms. Transient Receptor Potential Ankyrin 1 (TRPA1) is a nonselective cation channel with relatively high Ca

Topics: Adenomatous Polyposis Coli; Animals; Apoptosis; bcl-2 Homologous Antagonist-Killer Protein; Body Weight; Brain; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Fibroblast Growth Factor 2; Gene Expression Regulation; Gliosis; Mice; Mice, Knockout; Monoamine Oxidase Inhibitors; Myelin Basic Protein; Nerve Tissue Proteins; Oligodendroglia; Signal Transduction; TRPA1 Cation Channel

In the central nervous system (CNS) the main proteins of myelin are proteolipid protein (PLP), myelin basic protein (MBP), myelin oligodendrocyte glycoprotein (MOG) and CNPase. Myelin oligodendrocyte glycoprotein is a minor component of the myelin sheath, but is an important autoantigen linked to the pathogenesis of multiple sclerosis (MS). CNPase is expressed exclusively by oligodendrocytes in the CNS, and the appearance of CNPase seems to be one of the earliest events of oligodendrocyte differentiation and myelination. In this study the effects of vitamin D on total protein concentration, CNPase and MOG expression in the cerebral cortex of the murine model of cuprizone-induced demyelination was investigated. The mice were treated by cuprizone for five weeks in order to induce demyelination. The mice were then divided into 3 groups. The first group was injected intraperitoneally (IP) with vitamin D diluted in olive oil in the amount of 5 µg/kg/daily body weight. The second group (SHAM) was injected IP with olive oil and the third group was left without any injection as the control group (n = 11 for each group). After five weeks the mice were killed and the cerebral cortex was collected and the expression of CNPase and MOG was studied by Western blot. Total protein concentration in the vitamin D injected, SHAM and control groups were 0.918 ± 0.003, 0917 ± 0.004 and 0.916 ± 0.004 g/l, respectively (p > 0.05). However, a significant increase in the MOG and CNPase expression was seen in vitamin D injected group as compared to SHAM and control groups. It is concluded that vitamin D plays a role in the process of remyelination by increasing MOG and CNPase expression in the cortex.

Topics: Animals; Cerebral Cortex; Cholecalciferol; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Mice; Mice, Inbred BALB C; Myelin Basic Protein; Myelin-Oligodendrocyte Glycoprotein; Oligodendroglia

The occurrence of epilepsy is associated with myelin sheath injury; oligodendrocyte (OL) is the main cell of myelin sheath; In this study, we observed the changes of OL, demyelination, and myelin associated protein in different stages of intractable epilepsy (IE) at the epileptic foci of patients, and provide useful information for the pathophysiology of IE.. IE patients who received epileptogenic focus resection were recruited as the experimental group, their medical records were collected and postoperative follow-up was performed. The brain tissues from10 cases with non-brain disorders were obtained as controls. Immunofluorescence double staining was used to observe OL expression. The demyelination in epileptic foci was observed by luxol fast blue (LFB) staining method. Real-time fluorescent quantitative (RT) PCR, Western blot methods were used to detect the expressions of myelin-related proteins.. We observed increased OL precursor cells, former OL and decreased mature OL in experimental group when compared with controls (both p < 0.05). The demyelination was obviously higher in experimental group when compared with controls (p < 0.01). We also observed significantly decreasing of myelin basic protein (MBP), oligodendrocytes myelin glycoprotein (OMgp), myelin associated glycoprotein (MAG) and other MAG associated proteins-Nogo receptors (NgRs) and GD1α (p < 0.01) in epileptic foci.. OL is differentiated abnormally at epileptic foci of patients with IE; the demyelination, decreasing of demyelination and myelin associated protein are related to the occurrence of IE.

Topics: Demyelinating Diseases; Drug Resistant Epilepsy; Humans; Myelin Basic Protein; Myelin Sheath; Neuroglia

Multiple Sclerosis affects mainly women and consists in intermittent or chronic damages to the myelin sheaths, focal inflammation, and axonal degeneration. Current therapies are limited to immunomodulators and antiinflammatory drugs, but there is no efficient treatment for stimulating the endogenous capacity of myelin repair. Progesterone and synthetic progestins have been shown in animal models of demyelination to attenuate myelin loss, reduce clinical symptoms severity, modulate inflammatory responses and partially reverse the age-dependent decline in remyelination. Moreover, progesterone has been demonstrated to promote myelin formation in organotypic cultures of cerebellar slices. In the present study, we show that progesterone and the synthetic 19-nor-progesterone derivative Nestorone® promote the repair of severe chronic demyelinating lesions induced by feeding cuprizone to female mice for up to 12 weeks. Progesterone and Nestorone increase the density of NG2(+) oligodendrocyte progenitor cells and CA II(+) mature oligodendrocytes and enhance the formation of myelin basic protein (MBP)- and proteolipid protein (PLP)-immunoreactive myelin. However, while demyelination in response to cuprizone was less marked in corpus callosum than in cerebral cortex, remyelination appeared earlier in the former. The remyelinating effect of progesterone was progesterone receptor (PR)-dependent, as it was absent in PR-knockout mice. Progesterone and Nestorone also decreased (but did not suppress) neuroinflammatory responses, specifically astrocyte and microglial cell activation. Therefore, some progestogens are promising therapeutic candidates for promoting the regeneration of myelin.

Topics: Animals; Cerebral Cortex; Corpus Callosum; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Female; Mice, Inbred C57BL; Mice, Knockout; Multiple Sclerosis; Myelin Basic Protein; Myelin Proteolipid Protein; Myelin Sheath; Oligodendroglia; Progesterone; Stem Cells

Deciphering the molecular pathways involved in myelin gene expression is a major point of interest to better understand re/myelination processes. In this study, we investigated the role of Lithium Chloride (LiCl), a drug largely used for the treatment of neurological disorders, on the two major central myelin gene expression (PLP and MBP) in mouse oligodendrocytes. We show that LiCl enhances the expression of both PLP and MBP, by increasing mRNA amount and promoter activities. We investigated whether Wnt/β-catenin and/or Akt/CREB pathways are modulated by LiCl to regulate myelin gene expression. We showed that β-catenin is required both for PLP and MBP basal promoter activities and for LiCl-induced myelin gene stimulation. Furthermore, while CREB functionality does not influence PLP expression, MBP promoter activity depends on Akt/CREB activation. Finally, we show that LiCl can stimulate oligodendrocyte morphological maturation, and promote remyelination after lysolecithin-induced demyelination of organotypic cerebellar slice cultures. Our data provide mechanistic evidences that Akt/CREB together with β-catenin participate in the transcriptional control of PLP and MBP exerted by LiCl. Therefore, the use of LiCl to balance between β-catenin and CREB effectors could be considered as an efficient remyelinating strategy.

Topics: Animals; beta Catenin; Brain; Cell Line; Cells, Cultured; Cyclic AMP Response Element-Binding Protein; Demyelinating Diseases; Disease Models, Animal; Gene Expression; Lithium Chloride; Mice, Inbred C57BL; Mice, Transgenic; Myelin Basic Protein; Myelin Proteolipid Protein; Neuroprotective Agents; Oligodendroglia; Proto-Oncogene Proteins c-akt; RNA, Messenger; Signal Transduction; Tissue Culture Techniques; Wnt Proteins

Electroacupuncture (EA) has been used worldwide to treat demyelinating diseases, but its therapeutic mechanism is poorly understood. In this study, a custom-designed model of compressed spinal cord injury (CSCI) was used to induce demyelination. Zusanli (ST36) and Taixi (KI3) acupoints of adult rats were stimulated by EA to demonstrate its protective effect. At 14 days after EA, both locomotor skills and ultrastructural features of myelin sheath were significantly improved. Phenotypes of proliferating cells were identified by double immunolabeling of 5-ethynyl-2'-deoxyuridine with antibodies to cell markers: NG2 [oligodendrocyte precursor cell (OPC) marker], 2',3'-cyclic-nucleotide 3'-phosphodiesterase (CNPase) (oligodendrocyte marker), and glial fibrillary acidic protein (GFAP) (astrocyte marker). EA enhanced the proliferation of OPCs and CNPase, as well as the differentiation of OPCs by promoting Olig2 (the basic helix-loop-helix protein) and attenuating Id2 (the inhibitor of DNA binding 2). EA could also improve myelin basic protein (MBP) and protect existing oligodendrocytes from apoptosis by inhibiting caspase-12 (a representative of endoplasmic reticulum stress) and cytochrome c (an apoptotic factor and hallmark of mitochondria). Therefore, our results indicate that the protective effect of EA on neural myelin sheaths is mediated via promotion of oligodendrocyte proliferation and inhibition of oligodendrocyte death after CSCI.

Topics: Animals; Astrocytes; Cell Death; Cell Differentiation; Demyelinating Diseases; Electroacupuncture; Glial Fibrillary Acidic Protein; Myelin Basic Protein; Myelin Sheath; Nerve Regeneration; Oligodendroglia; Rats, Sprague-Dawley; Spinal Cord Injuries; Stem Cells

Autoantibodies against various components of host are known to occur in leprosy. Nerve damage is the primary cause of disability associated with leprosy. The aim of this study was to detect the level of autoantibodies and lympho-proliferative response against myelin basic protein (MBP) in leprosy patients (LPs) and their correlation with clinical phenotypes of LPs. Further, probable role of molecular mimicry in nerve damage of LPs was investigated. We observed significantly high level of anti-MBP antibodies in LPs across the spectrum and a positive significant correlation between the level of anti-MBP antibodies and the number of nerves involved in LPs. We report here that 4 B cell epitopes of myelin A1 and Mycobacterium leprae proteins, 50S ribosomal L2 and lysyl tRNA synthetase are cross-reactive. Further, M. leprae sonicated antigen hyperimmunization was responsible for induction of autoantibody response in mice which could be adoptively transferred to naive mice. For the first time our findings suggest the role of molecular mimicry in nerve damage in leprosy.

Topics: Animals; Demyelinating Diseases; Humans; Leprosy; Lysine-tRNA Ligase; Mice; Mice, Inbred BALB C; Molecular Mimicry; Mycobacterium leprae; Myelin Basic Protein; Rabbits; Ribosomal Proteins

Experimental evidence has shown that the adrenal steroid hormone, androstenediol, dampens the symptoms of demyelination. However, the cellular and molecular effects of androstenediol are not yet known. In the present study, we investigated the cellular and subcellular effects of this hormone in a gliotoxin-induced demyelination model.. Male Sprague Dawley rats received 2 μl of either saline or the gliotoxin ethidium bromide (EB, 0.04%) into the corpus callosum. These rats received daily subcutaneous injections of either oil or androstenediol (5 mg/kg). Their brains were collected at 2, 7, 14 and 28 days post-EB injection. Demyelinated lesions were assessed using Luxol fast blue staining. Immunofluorescent staining was used to investigate the number of oligodendrocyte progenitor cells, their maturation and microglial activation at the lesion site. Remyelination was further explored using transmission electron microscopy. The expression levels of total and phosphorylated MBP isoforms and CNPase were explored using western blot.. Androstenediol decreased the size of demyelinated lesions in the corpus callosum at 7 and 14 days post-EB injection. It enhanced the number of oligodendrocyte precursor cells, promoted an increase in the number of mature oligodendrocytes and reduced microglial activation. Androstenediol also stimulated the phosphorylation of MBP at the site of the lesion and promoted remyelination of the affected axons.. These data strongly suggest that androstenediol is endowed with promyelinating properties in a model of focal gliotoxin-induced demyelination. It induces its promyelinating effects by enhancing the number of oligodendrocyte precursor cells and their maturation at the lesion site.

Topics: Androstenediol; Animals; Corpus Callosum; Demyelinating Diseases; Gliotoxin; Male; Myelin Basic Protein; Myelin Sheath; Phosphorylation; Rats; Rats, Sprague-Dawley

The inflammatory mediator lipopolysaccharide (LPS) has been shown to induce acute gliosis in neonatal mice. However, the progressive effects on the murine neurodevelopmental program over the week that follows systemic inflammation are not known. Thus, we investigated the effects of repeated LPS administration in the first postnatal week in mice, a condition mimicking sepsis in late preterm infants, on the developing central nervous system (CNS).. Systemic inflammation was induced by daily intraperitoneal administration (i.p.) of LPS (6 mg/kg) in newborn mice from postnatal day (PND) 4 to PND6. The effects on neurodevelopment were examined by staining the white matter and neurons with Luxol Fast Blue and Cresyl Violet, respectively. The inflammatory response was assessed by quantifying the expression/activity of matrix metalloproteinases (MMP), toll-like receptor (TLR)-4, high mobility group box (HMGB)-1, and autotaxin (ATX). In addition, B6 CX3CR1(gfp/+) mice combined with cryo-immunofluorescence were used to determine the acute, delayed, and lasting effects on myelination, microglia, and astrocytes.. LPS administration led to acute body and brain weight loss as well as overt structural changes in the brain such as cerebellar hypoplasia, neuronal loss/shrinkage, and delayed myelination. The impaired myelination was associated with alterations in the proliferation and differentiation of NG2 progenitor cells early after LPS administration, rather than with excessive phagocytosis by CNS myeloid cells. In addition to disruptions in brain architecture, a robust inflammatory response to LPS was observed. Quantification of inflammatory biomarkers revealed decreased expression of ATX with concurrent increases in HMGB1, TLR-4, and MMP-9 expression levels. Acute astrogliosis (GFAP(+) cells) in the brain parenchyma and at the microvasculature interface together with parenchymal microgliosis (CX3CR1(+) cells) were also observed. These changes preceded the migration/proliferation of CX3CR1(+) cells around the vessels at later time points and the subsequent loss of GFAP(+) astrocytes.. Collectively, our study has uncovered a complex innate inflammatory reaction and associated structural changes in the brains of neonatal mice challenged peripherally with LPS. These findings may explain some of the neurobehavioral abnormalities that develop following neonatal sepsis.

Topics: Age Factors; Anethole Trithione; Animals; Animals, Newborn; Body Weight; Cerebellum; CX3C Chemokine Receptor 1; Demyelinating Diseases; Developmental Disabilities; Gene Expression Regulation; Green Fluorescent Proteins; HMGB1 Protein; Inflammation; Lipopolysaccharides; Mice; Mice, Inbred C57BL; Mice, Transgenic; Microglia; Myelin Basic Protein; Nervous System Malformations; Neurodegenerative Diseases; Receptors, Chemokine; Time Factors; Toll-Like Receptor 4

Here we studied the autoantibody specificity elicited by proteolipid protein (PLP) in MP4-induced experimental autoimmune encephalomyelitis, a mouse model of multiple sclerosis (MS). In C57BL/6 (B6) mice, antibodies were induced by immunization with one of the two extracellular and by the intracellular PLP domain. Antibodies against extracellular PLP were myelin-reactive in oligodendrocyte cultures and induced mild spinal cord demyelination upon transfer into B cell-deficient J(H)T mice. Remarkably, also antibodies against intracellular PLP showed binding to intact oligodendrocytes and were capable of inducing myelin pathology upon transfer into J(H)T mice. In MP4-immunized mice peptide-specific T(H)1/T(H)17 responses were mainly directed against the extracellular PLP domains, but also involved the intracellular epitopes. These data suggest that both extracellular and intracellular epitopes of PLP contribute to the pathogenesis of MP4-induced EAE already in the setting of intact myelin. It remains to be elucidated if this concept also applies to MS itself.

Topics: Animals; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Epitopes; Mice; Myelin Basic Protein; Myelin Proteolipid Protein; Oligodendroglia; Peptide Fragments; Protein Structure, Tertiary; Recombinant Fusion Proteins; Th1 Cells; Th17 Cells

NADPH oxidase (NOX)-dependent reactive oxygen species (ROS) production in inflammatory cells including microglia plays an important role in demyelination and free radical-mediated tissue injury in multiple sclerosis (MS). However, the mechanism underlying microglial ROS production and demyelination remains largely unknown. The voltage-gated proton channel, Hv1, is selectively expressed in microglia and is required for NOX-dependent ROS generation in the brain. In the present study, we sought to determine the role of microglial Hv1 proton channels in a mouse model of cuprizone-induced demyelination, a model for MS. Following cuprizone exposure, wild-type mice presented obvious demyelination, decreased myelin basic protein expression, loss of mature oligodendrocytes, and impaired motor coordination in comparison to mice on a normal chow diet. However, mice lacking Hv1 (Hv1(-/-) ) are partially protected from demyelination and motor deficits compared with those in wild-type mice. These rescued phenotypes in Hv1(-/-) mice in cuprizone-induced demyelination is accompanied by reduced ROS production, ameliorated microglial activation, increased oligodendrocyte progenitor cell (NG2) proliferation, and increased number of mature oligodendrocytes. These results demonstrate that the Hv1 proton channel is required for cuprizone-induced microglial oxidative damage and subsequent demyelination. Our study suggests that the microglial Hv1 proton channel is a unique target for controlling NOX-dependent ROS production in the pathogenesis of MS.

Topics: Animals; Chelating Agents; Cuprizone; Demyelinating Diseases; Ion Channels; Macrophage Activation; Mice; Mice, Inbred C57BL; Mice, Knockout; Microglia; Multiple Sclerosis; Myelin Basic Protein; NADPH Oxidases; Neural Stem Cells; Oxidative Stress; Postural Balance; Reactive Oxygen Species

Multiple sclerosis (MS) is believed to be initiated by myelin-reactive CD4(+) Th cells. IL-12-polarized Th1 cells, IL-23-polarized Th17 cells, and Th17 cells that acquire Th1 characteristics were each implicated in autoimmune pathogenesis. It is debated whether Th cells that can drive the development of demyelinating lesions are phenotypically diverse or arise from a single lineage. In the current study, we assessed the requirement of IL-12 or IL-23 stimulation, as well as Th plasticity, for the differentiation of T cells capable of inducing CNS axon damage. We found that stable murine Th1 and Th17 cells independently transfer experimental autoimmune encephalomyelitis (widely used as an animal model of MS) in the absence of IL-23 and IL-12, respectively. Plastic Th17 cells are particularly potent mediators of demyelination and axonopathy. In parallel studies, we identified MS patients who consistently mount either IFN-γ- or IL-17-skewed responses to myelin basic protein over the course of a year. Brain magnetic resonance imaging revealed that patients with mixed IFN-γ and IL-17 responses have relatively high T1 lesion burden, a measure of permanent axon damage. Our data challenge the dogma that IL-23 and Th17 plasticity are universally required for the development of experimental autoimmune encephalomyelitis. This study definitively demonstrates that autoimmune demyelinating disease can be driven by distinct Th-polarizing factors and effector subsets, underscoring the importance of a customized approach to the pharmaceutical management of MS.

Topics: Adoptive Transfer; Animals; Autoimmunity; Brain; Cell Differentiation; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Humans; Interferon-gamma; Interleukin-12; Interleukin-17; Interleukin-23; Magnetic Resonance Imaging; Mice; Mice, Inbred C57BL; Mice, Knockout; Multiple Sclerosis; Myelin Basic Protein; Optic Nerve; Radiography; Th1 Cells; Th17 Cells

Increasing evidence suggests that white matter disorders based on myelin sheath impairment may underlie the neuropathological changes in schizophrenia. But it is unknown whether enhancing remyelination is a beneficial approach to schizophrenia. To investigate this hypothesis, we used clemastine, an FDA-approved drug with high potency in promoting oligodendroglial differentiation and myelination, on a cuprizone-induced mouse model of demyelination. The mice exposed to cuprizone (0.2% in chow) for 6 weeks displayed schizophrenia-like behavioral changes, including decreased exploration of the center in the open field test and increased entries into the arms of the Y-maze, as well as evident demyelination in the cortex and corpus callosum. Clemastine treatment was initiated upon cuprizone withdrawal at 10 mg/kg per day for 3 weeks. As expected, myelin repair was greatly enhanced in the demyelinated regions with increased mature oligodendrocytes (APC-positive) and myelin basic protein. More importantly, the clemastine treatment rescued the schizophrenia-like behavioral changes in the open field test and the Y-maze compared to vehicle, suggesting a beneficial effect via promoting myelin repair. Our findings indicate that enhancing remyelination may be a potential therapy for schizophrenia.

Topics: Animals; Behavior, Animal; Brain; Cell Differentiation; Clemastine; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Exploratory Behavior; Male; Mice; Mice, Inbred C57BL; Motor Activity; Myelin Basic Protein; Myelin Sheath; Oligodendroglia; Schizophrenia

Optic neuritis associated with multiple sclerosis and its animal model, experimental autoimmune optic neuritis (EAON), is characterized by inflammation, T cell activation, demyelination, and neuronal damage, which might induce permanent vision loss. Elucidating the chronological relationship among the features is critical for treatment of demyelinating optic neuritis. EAON was induced in C57BL/6 mice immunized with myelin oligodendrocyte glycoprotein subcutaneously, and visual function was assessed by flash-visual evoked potential (F-VEP) at days 7, 11, 14, 19, 23, 28 post-immunization. Retinal ganglion cell (RGC) apoptosis was measured by terminal-deoxynucleotidyl transferase-mediated nick-end labeling. Demyelination and axonal damage were verified with myelin basic protein (MBP) and β-amyloid precursor protein staining, respectively. Real-time polymerase chain reaction quantified IL-17, IL-1β, TGF-β, FoxP3, IL-6, and IL-10 mRNA expression in the optic nerve, as well as FoxP3 and IL-17 staining. Systemic changes of Th17 and Treg cells were tested by flow cytometry in spleen. F-VEP latency was prolonged at 11 days and peaked at 23 days commensurate with demyelination. However, F-VEP amplitude was reduced at 11 days, preceding axon damage, and was exacerbated at 23 days when a peak in RGC apoptosis was detected. Th17 cells up-regulated as early as 7 days and peaked at 11 days, while Treg cells down-regulated inversely compared to Th17 cells change as verified by IL-17 and FoxP3 expression; spleen cell samples were slightly different, demonstrating marked changed at 14 days. Treg/Th17 cell imbalance in the optic nerve precedes and may initiate neuronal damage of axons and RGCs. These changes are commensurate with the appearances of visual dysfunction reflected in F-VEP and hence may offer a novel therapeutic avenue for vision preservation.

Topics: Amyloid beta-Protein Precursor; Animals; Apoptosis; Axons; Demyelinating Diseases; Evoked Potentials, Visual; Female; Forkhead Transcription Factors; Interleukins; Lymphocyte Count; Lymphotoxin-alpha; Mice; Mice, Inbred C57BL; Myelin Basic Protein; Nerve Tissue Proteins; Nervous System Autoimmune Disease, Experimental; Optic Nerve; Optic Neuritis; Retinal Ganglion Cells; T-Lymphocytes, Regulatory; Th17 Cells

Multiple sclerosis (MS) is a progressive neurological disorder associated with myelin destruction and neurodegeneration. Oligodendrocyte precursor cells (OPCs) present in demyelinated lesions gradually fail to differentiate properly, so remyelination becomes incomplete. Protein tyrosine phosphatase receptor type Z (PTPRZ), one of the most abundant protein tyrosine phosphatases expressed in OPCs, is known to suppress oligodendrocyte differentiation and maintain their precursor cell stage. In the present study, we examined the in vivo mechanisms for remyelination using a cuprizone-induced demyelination model. Ptprz-deficient and wild-type mice both exhibited severe demyelination and axonal damage in the corpus callosum after cuprizone feeding. The similar accumulation of OPCs was observed in the lesioned area in both mice; however, remyelination was significantly accelerated in Ptprz-deficient mice after the removal of cuprizone. After demyelination, the expression of pleiotrophin (PTN), an inhibitory ligand for PTPRZ, was transiently increased in mouse brains, particularly in the neurons involved, suggesting its role in promoting remyelination by inactivating PTPRZ activity. In support of this view, oligodendrocyte differentiation was augmented in a primary culture of oligodendrocyte-lineage cells from wild-type mice in response to PTN. In contrast, these cells from Ptprz-deficient mice showed higher oligodendrocyte differentiation without PTN and differentiation was not enhanced by its addition. We further demonstrated that PTN treatment increased the tyrosine phosphorylation of p190 RhoGAP, a PTPRZ substrate, using an established line of OPCs. Therefore, PTPRZ inactivation in OPCs by PTN, which is secreted from demyelinated axons, may be the mechanism responsible for oligodendrocyte differentiation during reparative remyelination in the CNS.. Multiple sclerosis (MS) is an inflammatory disease of the CNS that destroys myelin, the insulation that surrounds axons. Associated damages to oligodendrocytes (the cells that produce myelin) and nerve fibers produce neurological disability. Most patients with MS have an initial relapsing-remitting course for 5-15 years. Remyelination during the early stages of the disease process has been documented; however, the molecular mechanism underlying remyelination has not been understood. Protein tyrosine phosphatase receptor type Z (PTPRZ) is a receptor-like protein tyrosine phosphatase preferentially expressed in the CNS. This study shows that pleiotrophin, an inhibitory ligand for PTPRZ, is transiently expressed and released from demyelinated neurons to inactivate PTPRZ in oligodendrocyte precursor cells present in the lesioned part, thereby allowing their differentiation for remyelination.

Topics: Amyloid beta-Protein Precursor; Animals; Antigens; Carrier Proteins; Cell Differentiation; Cells, Cultured; Corpus Callosum; Cuprizone; Cytokines; Demyelinating Diseases; Disease Models, Animal; Gene Expression Regulation; Mice; Mice, Inbred C57BL; Mice, Transgenic; Myelin Basic Protein; Nerve Tissue Proteins; Oligodendroglia; Proteoglycans; Receptor-Like Protein Tyrosine Phosphatases, Class 5; Stem Cells; Tumor Suppressor Protein p53

During mammalian development, myelin-forming oligodendrocytes are generated and axons ensheathed according to a tightly regulated sequence of events. Excess premyelinating oligodendrocytes are eliminated by apoptosis and the timing of the onset of myelination in any specific CNS region is highly reproducible. Although the developing CNS recovers more effectively than the adult CNS from similar insults, it is unknown whether early loss of oligodendrocyte lineage cells leads to long-term functional deficits. To directly assess whether the loss of oligodendrocytes during early postnatal spinal cord development impacted oligodendrogenesis, myelination, and remyelination, transgenic mouse lines were generated in which a modified caspase-9 molecule allowed spatial and temporal control of the apoptotic pathway specifically in mature, myelin basic protein expressing oligodendrocytes (MBP-iCP9). Activating apoptosis in MBP(+) cells of the developing spinal cord during the first postnatal week inhibited myelination. This inhibition was transient, and the levels of myelination largely returned to normal after 2 weeks. Despite robust developmental plasticity, MBP-iCP9-induced oligodendrocyte apoptosis compromised the rate and extent of adult remyelination. Remyelination failure correlated with a truncated proliferative response of oligodendrocyte progenitor cells, suggesting that depleting the oligodendrocyte pool during critical developmental periods compromises the regenerative response to subsequent demyelinating lesions.. This manuscript demonstrates that early insults leading to oligodendrocyte apoptosis result in the impairment of recovery from demyelinating diseases in the adult. These studies begin to provide an initial understanding of the potential failure of recovery in insults, such as periventricular leukomalacia and multiple sclerosis.

Topics: Age Factors; Animals; Animals, Newborn; Apoptosis; Caspase 9; Cells, Cultured; Demyelinating Diseases; Dimerization; Disease Models, Animal; Female; Glial Fibrillary Acidic Protein; Lysophosphatidylcholines; Male; Mice; Mice, Transgenic; Myelin Basic Protein; Oligodendroglia; Platelet-Derived Growth Factor; Spinal Cord; Tubulin

Ιn multiple sclerosis (MS), axonal damage leads to permanent neurological disabilities and the spreading of the autoimmune response to axonal antigens is implicated in disease progression. Experimental autoimmune encephalomyelitis (EAE) provides an animal model that mimics MS. Using different EAE models, we investigated the pathophysiological basis of epitope spreading to neurofascin, a protein localized at the node of Ranvier and its regulation by non-MHC genes.. We used two different EAE models in DA rat; one which is induced with myelin oligodendrocyte glycoprotein (MOG) which leads to disease characterized by profound demyelination, and the second which is induced with myelin basic protein (MBP) peptide 63-88 which results in severe central nervous system (CNS) inflammation but little or no demyelination. We determined anti-neurofascin antibody levels during the course of disease. Furthermore, the anti-neurofascin IgG response was correlated with clinical parameters in 333 (DAxPVG.1AV1) x DA rats on which we performed linkage analysis to determine if epitope spreading to neurofascin was affected by non-MHC genes.. Spreading of the antibody response to neurofascin occurred in demyelinating MOG-induced EAE but not in EAE induced with MBP peptide 63-88. Anti-neurofascin IgG levels correlated with disease severity in (DAxPVG.1AV1) x DA rats, and a genomic region on chromosome 3 was found to influence this response.. Inter-molecular epitope spreading to neurofascin correlates with disease severity in MOG-EAE is dependent on extensive demyelination and is influenced by non-MHC genes. The findings presented here may shed light on factors involved in the severity of MS and its genetics.

Topics: Animals; Cell Adhesion Molecules; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Epitopes; Female; Immunoglobulin G; Inflammation; Male; Myelin Basic Protein; Myelin-Oligodendrocyte Glycoprotein; Nerve Growth Factors; Peptides; Rats

Multiple sclerosis (MS) is an autoimmune disease of the central nervous system (CNS) characterized by inflammation, demyelination and axonal pathology. Myelin basic protein/proteolipid protein (MBP-PLP) fusion protein MP4 is capable of inducing chronic experimental autoimmune encephalomyelitis (EAE) in susceptible mouse strains mirroring diverse histopathological and immunological hallmarks of MS. Lack of human tissue underscores the importance of animal models to study the pathology of MS.. Twenty-two female C57BL/6 (B6) mice were immunized with MP4 and the clinical development of experimental autoimmune encephalomyelitis (EAE) was observed. Methylene blue-stained semi-thin and ultra-thin sections of the lumbar spinal cord were assessed at the peak of acute EAE, three months (chronic EAE) and six months after onset of EAE (long-term EAE). The extent of lesional area and inflammation were analyzed in semi-thin sections on a light microscopic level. The magnitude of demyelination and axonal damage were determined using electron microscopy. Emphasis was put on the ventrolateral tract (VLT) of the spinal cord.. B6 mice demonstrated increasing demyelination and severe axonal pathology in the course of MP4-induced EAE. Additionally, mitochondrial swelling and a decrease in the nearest neighbor neurofilament distance (NNND) as early signs of axonal damage were evident with the onset of EAE. In semi-thin sections we observed the maximum of lesional area in the chronic state of EAE while inflammation was found to a similar extent in acute and chronic EAE. In contrast to the well-established myelin oligodendrocyte glycoprotein (MOG) model, disease stages of MP4-induced EAE could not be distinguished by assessing the extent of parenchymal edema or the grade of inflammation.. Our results complement our previous ultrastructural studies of B6 EAE models and suggest that B6 mice immunized with different antigens constitute useful instruments to study the diverse histopathological aspects of MS.

Topics: Animals; Axons; Demyelinating Diseases; Disease Models, Animal; Disease Progression; Encephalomyelitis, Autoimmune, Experimental; Female; Humans; Immunization; Lumbar Vertebrae; Mice; Mice, Inbred C57BL; Microtomy; Mitochondria; Mitochondrial Swelling; Multiple Sclerosis; Myelin Basic Protein; Myelin Proteolipid Protein; Myelin Sheath; Recombinant Fusion Proteins; Severity of Illness Index; Time Factors

Demyelination is one of the most important pathological factors of spinal cord injury. Oligodendrocyte apoptosis is involved in triggering demyelination. However, fewer reports on pathological changes and mechanism of demyelination have been presented from compressed spinal cord injury (CSCI). The relative effect of oligodendrocyte apoptosis on CSCI-induced demyelination and the mechanism of apoptosis remain unclear.. In this study, a custom-designed model of CSCI was used to determine whether or not demyelination and oligodendrocyte apoptosis occur after CSCI. The pathological changes in axonal myelinated fibers were investigated by osmic acid staining and transmission electron microscopy. Myelin basic protein (MBP), which is used in myelin formation in the central nervous system, was detected by immunofluorescence and Western blot assays. Oligodendrocyte apoptosis was revealed by in situ terminal-deoxytransferase-mediated dUTP nick-end labeling. To analyze the mechanism of oligodendrocyte apoptosis, we detected caspase-12 [a representative of endoplasmic reticulum (ER) stress], cytochrome c (an apoptotic factor and hallmark of mitochondria), and inhibitor of DNA binding 2 (Id2, an oligodendrocyte lineage gene) by immunofluorescence and Western blot assays.. The custom-designed model of CSCI was successfully established. The rats were spastic, paralyzed, and incontinent. The Basso, Beattie, and Bresnahan (BBB) locomotor rating scale scores were decreased as time passed. The compressed spinal cord slices were ischemic. Myelin sheaths became swollen and degenerative; these sheaths were broken down as time passed after CSCI. MBP expression was downregulated after CSCI and consistent with the degree of demyelination. Oligodendrocyte apoptosis occurred at 1 day after CSCI and increased as caspase-12 expression was enhanced and cytochrome c was released. Id2 was distributed widely in the white matter. Id2 expression increased with time after CSCI.. Demyelination occurred after CSCI and might be partly caused by oligodendrocyte apoptosis, which was positively correlated with ER-mitochondria interactions and enhanced Id2 expression after CSCI in rats.

Topics: Animals; Apoptosis; Axons; Blotting, Western; Caspase 12; Cytochromes c; Demyelinating Diseases; Endoplasmic Reticulum; Fluorescent Antibody Technique; In Situ Nick-End Labeling; Inhibitor of Differentiation Protein 2; Ischemia; Lumbar Vertebrae; Microscopy, Electron, Transmission; Mitochondria; Myelin Basic Protein; Oligodendroglia; Organelles; Rats; Rats, Sprague-Dawley; Spinal Cord Compression

Accumulating evidence indicates that the medial prefrontal cortex (mPFC) is a site of myelin and oligodendrocyte abnormalities that contribute to psychotic symptoms of schizophrenia. The development of therapeutic approaches to enhance remyelination, a regenerative process in which new myelin sheaths are formed on demyelinated axons, may be an attractive remedial strategy. Geissoschizine methyl ether (GM) in the Uncaria hook, a galenical constituent of the traditional Japanese medicine yokukansan (Yi-gan san), is one of the active components responsible for the psychotropic effects of yokukansan, though little is known about the mechanisms underlying the effects of either that medicine or GM itself. In the present study, we employed a cuprizone (CPZ)-induced demyelination model and examined the cellular changes in response to GM administration during the remyelination phase in the mPFC of adult mice. Using the mitotic marker 5-bromo-2'-deoxyuridine (BrdU), we demonstrated that CPZ treatment significantly increased the number of BrdU-positive NG2 cells, as well as microglia and mature oligodendrocytes in the mPFC. Newly formed oligodendrocytes were increased by GM administration after CPZ exposure. In addition, GM attenuated a decrease in myelin basic protein immunoreactivity caused by CPZ administration. Taken together, our findings suggest that GM administration ameliorated the myelin deficit by mature oligodendrocyte formation and remyelination in the mPFC of CPZ-fed mice. The present findings provide experimental evidence supporting the role for GM and its possible use as a remedy for schizophrenia symptoms by promoting the differentiation of progenitor cells to and myelination by oligodendrocytes.

Topics: Animals; Cuprizone; Demyelinating Diseases; Indole Alkaloids; Indoles; Mice; Myelin Basic Protein; Prefrontal Cortex; Regeneration

Although multiple sclerosis (MS) has been associated with the coagulation system, the temporal and spatial regulation of coagulation activity in neuroinflammatory lesions is unknown. Using a novel molecular probe, we characterized the activity pattern of thrombin, the central protease of the coagulation cascade, in experimental autoimmune encephalomyelitis. Thrombin activity preceded onset of neurological signs, increased at disease peak, and correlated with fibrin deposition, microglial activation, demyelination, axonal damage, and clinical severity. Mice with a genetic deficit in prothrombin confirmed the specificity of the thrombin probe. Thrombin activity might be exploited for developing sensitive probes for preclinical detection and monitoring of neuroinflammation and MS progression.

Topics: Animals; Axons; Blood Coagulation Factors; Connexin 30; Connexins; Demyelinating Diseases; Disease Models, Animal; Disease Progression; Encephalomyelitis, Autoimmune, Experimental; Fibrin; Green Fluorescent Proteins; Mice; Mice, Inbred C57BL; Mice, Transgenic; Myelin Basic Protein; Myelin-Oligodendrocyte Glycoprotein; Peptide Fragments; Poly I-C; Thrombin

Clinicopathological paradox has hampered significantly the effective assessment of the efficacy of therapeutic intervention for multiple sclerosis. Neuroimaging biomarkers of tissue injury could guide more effective treatment by accurately reflecting the underlying subclinical pathologies. Diffusion tensor imaging-derived directional diffusivity and anisotropy indices have been applied to characterize white matter disorders. However, these biomarkers are sometimes confounded by the complex pathologies seen in multiple sclerosis and its animal models. Recently, a novel technique of diffusion basis spectrum imaging has been developed to quantitatively assess axonal injury, demyelination and inflammation in a mouse model of inflammatory demyelination. Lenaldekar, which inhibits T-cell expansion in a non-cytolytic manner, has been shown to suppress relapses and preserve white matter integrity in mice with experimental autoimmune encephalomyelitis. In this study, relapsing-remitting experimental autoimmune encephalomyelitis was induced through active immunization of SJL/J mice with a myelin proteolipid protein peptide. The therapeutic efficacy of Lenaldekar treatment was evaluated via daily clinical score, cross-sectional ex vivo diffusion basis spectrum imaging examination and histological analysis. Lenaldekar greatly reduced relapse severity and protected white matter integrity in these experimental autoimmune encephalomyelitis mice. Diffusion basis spectrum imaging-derived axial diffusivity, radial diffusivity and restricted diffusion tensor fraction accurately reflected axonal injury, myelin integrity and inflammation-associated cellularity change, respectively. These results support the potential use of diffusion basis spectrum imaging as an effective outcome measure for preclinical drug evaluation.

Topics: Animals; Axons; Biomarkers; Demyelinating Diseases; Diffusion Tensor Imaging; Encephalomyelitis, Autoimmune, Experimental; Female; Hydrazones; Indoles; Inflammation; Mice; Myelin Basic Protein; Quinolines; Recurrence

Myelin loss is a widespread neuropathological hallmark of the atypical parkinsonian disorder multiple system atrophy (MSA). On a cellular level, MSA is characterized by alpha-synuclein (aSyn)-positive glial cytoplasmic inclusions (GCIs) within mature oligodendrocytes leading to demyelination as well as axonal and neuronal loss. Oligodendrocyte progenitor cells (OPCs) represent a proliferative cell population distributed throughout the adult mammalian central nervous system. During remyelination, OPCs are recruited to sites of demyelination, differentiate, and finally replace dysfunctional mature oligodendrocytes. However, comprehensive studies investigating OPCs and remyelination processes in MSA are lacking. In the present study, we therefore investigate the effect of human aSyn (h-aSyn) on early primary rat OPC maturation. Upon lentiviral transduction, h-aSyn expressing OPCs exhibit fewer and shorter primary processes at the initiation of differentiation. Until day 4 of a 6day differentiation paradigm, h-aSyn expressing OPCs further show a severely delayed maturation evidenced by reduced myelin gene expression and increased levels of the progenitor marker platelet derived growth factor receptor-alpha (PDGFRα). Matching these results, OPCs that take up extracellular recombinant h-aSyn exhibit a similar delayed differentiation. In both experimental setups however, myelin gene expression is restored at day 6 of differentiation paralleled by decreased intracellular h-aSyn levels indicating a reverse correlation of h-aSyn and the differentiation potential of OPCs. Taken together, these findings suggest a tight link between the intracellular level of h-aSyn and maturation capacity of primary OPCs.

Topics: alpha-Synuclein; Animals; Axons; Cell Differentiation; Cells, Cultured; Demyelinating Diseases; Intracellular Space; Myelin Basic Protein; Myelin Sheath; Neural Stem Cells; Oligodendroglia; Rats, Wistar

Dedicator of cytokinesis 3 (Dock3) belongs to an atypical family of the guanine nucleotide exchange factors. It is predominantly expressed in the neural tissues and causes cellular morphological changes by activating the small GTPase Rac1. We previously reported that Dock3 overexpression protects retinal ganglion cells from excitotoxic cell death. Oligodendrocytes are the myelinating cells of axons in the central nervous system and these cells are damaged in demyelinating disorders including multiple sclerosis (MS) and optic neuritis. In this study, we examined if Dock3 is expressed in oligodendrocytes and if increasing Dock3 signals can suppress demyelination in a cuprizone-induced demyelination model, an animal model of MS. We demonstrate that Dock3 is expressed in oligodendrocytes and Dock3 overexpression protects myelin in the corpus callosum following cuprizone treatment. Furthermore, we show that cuprizone demyelinates optic nerves and the extent of demyelination is ameliorated in mice overexpressing Dock3. Cuprizone treatment impairs visual function, which was demonstrated by multifocal electroretinograms, an established non-invasive method, and Dock3 overexpression prevented this effect. In mice overexpressing Dock3, Erk activation is increased, suggesting this may at least partly explain the observed protective effects. Our findings suggest that Dock3 may be a therapeutic target for demyelinating disorders including optic neuritis.Cell Death and Disease (2014) 5, e1395; doi:10.1038/cddis.2014.357; published online 28 August 2014.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Carrier Proteins; Cells, Cultured; Chelating Agents; Corpus Callosum; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Electroretinography; Extracellular Signal-Regulated MAP Kinases; Guanine Nucleotide Exchange Factors; Mice; Mice, Inbred C57BL; Mice, Transgenic; Myelin Basic Protein; Nerve Tissue Proteins; Oligodendroglia; Optic Nerve; Retina; Signal Transduction

Rapid and efficient axon remyelination aids in restoring strong electrochemical communication with end organs and in preventing axonal degeneration often observed in demyelinating neuropathies. The signals from axons that can trigger more effective remyelination in vivo are still being elucidated. Here we report the remarkable effect of delayed brief electrical nerve stimulation (ES; 1 hour @ 20 Hz 5 days post-demyelination) on ensuing reparative events in a focally demyelinated adult rat peripheral nerve. ES impacted many parameters underlying successful remyelination. It effected increased neurofilament expression and phosphorylation, both implicated in axon protection. ES increased expression of myelin basic protein (MBP) and promoted node of Ranvier re-organization, both of which coincided with the early reappearance of remyelinated axons, effects not observed at the same time points in non-stimulated demyelinated nerves. The improved ES-associated remyelination was accompanied by enhanced clearance of ED-1 positive macrophages and attenuation of glial fibrillary acidic protein expression in accompanying Schwann cells, suggesting a more rapid clearance of myelin debris and return of Schwann cells to a nonreactive myelinating state. These benefits of ES correlated with increased levels of brain derived neurotrophic factor (BDNF) in the acute demyelination zone, a key molecule in the initiation of the myelination program. In conclusion, the tremendous impact of delayed brief nerve stimulation on enhancement of the innate capacity of a focally demyelinated nerve to successfully remyelinate identifies manipulation of this axis as a novel therapeutic target for demyelinating pathologies.

Topics: Animals; Axons; Brain-Derived Neurotrophic Factor; Demyelinating Diseases; Electric Stimulation; Intermediate Filaments; Macrophages; Male; Myelin Basic Protein; Myelin Sheath; Nerve Tissue; Phosphorylation; Ranvier's Nodes; Rats, Wistar; Schwann Cells; Staining and Labeling; Stilbamidines

Multiple sclerosis (MS) is an autoimmune disease characterized by inflammatory cell infiltration of the central nervous system (CNS) and multifocal demyelination. Clinical data and clinical indicators demonstrate that estrogen improves the relapse-remittance of MS patients. This study aimed to investigate the anti-inflammatory effects and the underlying mechanism(s) of action of estrogen and estrogen receptor α (ERα) in an experimental autoimmune encephalomyelitis (EAE) mouse model of MS. An ERα recombinant lentivirus was constructed. Mouse neurons were cultured in serum-free culture medium, and ERα recombinant lentivirus with a multiplicity of infection (MOI) of 5 was used to infect the neurons. Furthermore, neuronal ERα mRNA and protein expression were detected using real-time quantitative PCR and western blot analysis. We sterotaxically injected ERα recombinant lentivirus into the lateral ventricle of mouse brains, and successfully identified infected neurons using Flag immunofluorescence staining to determine the optimal dose. A total of 75 C57BL/6 mice were ovariectomized. After 2 weeks, EAE was induced with myelin oligodendrocyte glycoprotein (MOG) 35-55 peptide. The EAE mice were divided into 5 groups: the estrogen group (treatment with estradiol), the ERα agonist group (treatment with raloxifene), the ERα recombinant lentivirus group (ERα group, treatment with ERα recombinant lentivirus), the empty virus group and the normal saline (NS) group; clinical symptoms and body weight were compared among the groups. We assessed EAE-related parameters, detected pathological changes with immunohistochemistry and quantified the expression of myelin basic protein (MBP), matrix metalloproteinase-9 (MMP-9), and a subset of EAE-related cytokines using enzyme-linked immunosorbent assay (ELISA). We successfully constructed an ERα recombinant lentivirus. C57BL/6 mouse neurons can survive in culture for at least 8 weeks. During that period, the recombinant lentivirus was able to infect the neurons, while sustaining green fluorescence protein (GFP) expression. ERα recombinant lentivirus also infected the neurons at a MOI of 5. The ERα mRNA and protein expression levels were higher in the infected neurons compared to the uninfected ones. We successfully infected the CNS of C57BL/6 mice by stereotaxically injecting ERα recombinant lentivirus into the lateral ventricle of the mouse brains and induced EAE. The lentivirus-mediated overexpression of ERα reduced the inciden

Topics: Animals; Body Weight; Brain; Central Nervous System; Cytokines; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Estrogen Receptor alpha; Green Fluorescent Proteins; HEK293 Cells; Humans; Immunohistochemistry; Inflammation; Lentivirus; Matrix Metalloproteinase 9; Mice; Mice, Inbred C57BL; Myelin Basic Protein; Nerve Fibers; Recombination, Genetic; RNA, Messenger; Spinal Cord; Transfection

Schizophrenia is a mental disease that mainly affects young individuals (15 to 35 years old) but its etiology remains largely undefined. Recently, accumulating evidence indicated that demyelination and/or dysfunction of oligodendrocytes is an important feature of its pathogenesis. We hypothesized that the vulnerability of young individuals to demyelination may contribute to the onset of schizophrenia. In the present study, three different age cohorts of mice, i.e. juvenile (3 weeks), young-adult (6 weeks) and middle-aged (8 months), were subjected to a 6-week diet containing 0.2% cuprizone (CPZ) to create an animal model of acute demyelination. Then, age-related vulnerability to CPZ-induced demyelination, behavioral outcomes, and myelination-related molecular biological changes were assessed. We demonstrated: (1) CPZ treatment led to more severe demyelination in juvenile and young-adult mice than in middle-aged mice in the corpus callosum, a region closely associated with the pathophysiology of schizophrenia; (2) the higher levels of demyelination in juvenile and young-adult mice were correlated with a greater reduction of myelin basic protein, more loss of CC-1-positive mature oligodendrocytes, and higher levels of astrocyte activation; and (3) CPZ treatment resulted in a more prominent exploratory behavior deficit in juvenile and young-adult mice than in middle-aged mice. Together, our data demonstrate an age-related vulnerability to demyelination with a concurrent behavioral deficit, providing supporting evidence for better understanding the susceptibility of the young to the onset of schizophrenia.

Topics: Age Factors; Analysis of Variance; Animals; Antigens, CD; Cell Adhesion Molecules; Chelating Agents; Cohort Studies; Corpus Callosum; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Exploratory Behavior; Gene Expression Regulation; Glial Fibrillary Acidic Protein; Mental Disorders; Mice; Mice, Inbred C57BL; Myelin Basic Protein; Oligodendroglia

The Long-Evans shaker (les) rat has a mutation in myelin basic protein that results in severe CNS dysmyelination and subsequent demyelination during development. During this time, les oligodendrocytes accumulate cytoplasmic vesicles, including lysosomes and membrane-bound organelles. However, the mechanism and functional relevance behind these oligodendrocyte abnormalities in les have not been investigated. Using high-magnification electron microscopy, we identified the accumulations in les oligodendrocytes as early and late autophagosomes. Additionally, immunohistochemistry and Western blots showed an increase in autophagy markers in les. However, autophagy did not precede the death of les oligodendrocytes. Instead, upregulating autophagy promoted membrane extensions in les oligodendrocytes in vitro. Furthermore, upregulating autophagy in les rats via intermittent fasting increased the proportion of myelinated axons as well as myelin sheath thickness in les and control rats. Overall, this study provides insight into the abnormalities described in les as well as identifying a novel mechanism that promotes the survival and function of oligodendrocytes.

Topics: Age Factors; Animals; Animals, Newborn; Autophagy; Bromodeoxyuridine; Cell Survival; Cells, Cultured; Chloroquine; Cytoplasm; Demyelinating Diseases; Fasting; Female; Male; Microtubule-Associated Proteins; Mutation; Myelin Basic Protein; Myelin Proteolipid Protein; Myelin Sheath; Oligodendroglia; Peptides; Rats; Rats, Long-Evans; Rats, Mutant Strains; Spinal Cord; Up-Regulation

Remyelination is disrupted in demyelinating diseases such as multiple sclerosis, but the underlying pathogenetic mechanisms are unclear. In this study, we employed the murine cuprizone model of demyelination, in which remyelination occurs after removal of the toxin from the diet, to examine the cellular and molecular changes during demyelination and remyelination. Microglia accumulated in the corpus callosum during weeks 2-4 of the cuprizone diet, and these cells remained activated 2 weeks after the change to the normal diet. To examine the role of microglia in remyelination, mice were treated with minocycline to inactivate these cells after cuprizone-induced demyelination. Minocycline treatment reduced the number of CC1-positive oligodendrocytes, as well as levels of myelin basic protein (MBP) and CNPase in the remyelination phase. The expression of CNTF mRNA in the corpus callosum increased after 4 weeks on the cuprizone diet and remained high 2 weeks after the change to the normal diet. Minocycline suppressed CNTF expression during the remyelination phase on the normal diet. Primary culture experiments showed that CNTF was produced by microglia in addition to astrocytes. In vitro, CNTF directly affected the differentiation of oligodendrocytic cells. These findings suggest that minocycline reduces remyelination by suppressing CNTF expression by microglia after cuprizone-induced demyelination.

Topics: Animals; Anti-Bacterial Agents; Blotting, Western; Cells, Cultured; Ciliary Neurotrophic Factor; Corpus Callosum; Cuprizone; Demyelinating Diseases; Female; Immunohistochemistry; Mice; Mice, Inbred C57BL; Microglia; Minocycline; Monoamine Oxidase Inhibitors; Myelin Basic Protein; Myelin Sheath; Oligodendroglia; Real-Time Polymerase Chain Reaction

The temporal and spatial patterning involved in the specification, differentiation, and myelination by oligodendroglia is coordinated in part by the activation and repression of various transcriptional programs. Olig2 is a basic helix-loop-helix transcription factor necessary for oligodendroglial development and expressed continuously throughout the lineage. Despite evidence for the critical role of Olig2 in oligodendroglial specification and differentiation, the function for Olig2 during later stages of oligodendroglial development, namely, the transition into mature oligodendrocytes (OLs) and the formation of the myelin sheath, remains unclear. To address the possibility for a stage-specific role, we deleted Olig2 in oligodendrocyte precursor cells (OPCs) under the control of the CNPase-promoter or in immature OLs under the inducible proteolipid protein promoter. As expected, ablation of Olig2 in OPCs significantly inhibits differentiation, resulting in hypomyelination. However, deletion of the Olig2 gene in immature OLs significantly enhances the maturation process and accelerates the kinetics of myelination/remyelination. Underlying the stage-specific roles for Olig2 is the compensatory expression and function of Olig1, a transcription factor that promotes OL maturation and (re)myelination. Olig1 expression is significantly reduced upon Olig2 deletion in OPCs but is dramatically increased by nearly threefold when deleted in immature OLs. By enforcing expression of Olig1 into OPCs in a null Olig2 background, we demonstrate that overexpression of Olig1 is sufficient to rescue the differentiation phenotype and partially compensates for the Olig2 deletion in vitro. Our results suggest a stage-specific regulatory role for Olig2, mediated by Olig1 that conveys opposing functions on the differentiation and maturation of oligodendrocytes.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Animals, Newborn; Arabidopsis Proteins; Autophagy-Related Proteins; Basic Helix-Loop-Helix Transcription Factors; Bromodeoxyuridine; Cell Differentiation; Cuprizone; Demyelinating Diseases; Gene Expression Regulation, Developmental; Green Fluorescent Proteins; In Situ Nick-End Labeling; Intracellular Signaling Peptides and Proteins; Intramolecular Transferases; Male; Mice; Mice, Transgenic; Microscopy, Electron, Transmission; Monoamine Oxidase Inhibitors; Myelin Basic Protein; Myelin Proteolipid Protein; Nerve Tissue Proteins; Oligodendrocyte Transcription Factor 2; Oligodendroglia; ran GTP-Binding Protein; Transfection

Multiple sclerosis (MS) is characterized by the presence of inflammatory demyelinating foci throughout the brain and spinal cord, accompanied by axonal and neuronal damage. Although inflammatory processes are thought to underlie the pathological changes, the individual mediators of this damage are unclear. In order to study the role of pro-inflammatory cytokines in demyelination in the central nervous system, we have utilized a hyperbranched poly(2-dimethyl-aminoethylmethacrylate) based non-viral gene transfection system to establish an inflammatory demyelinating model of MS in an ex-vivo environment. The synthesized non-viral gene transfection system was optimized for efficient transfection with minimal cytotoxicity. Organotypic brain slices were then successfully transfected with the TNF or IFNγ genes. TNF and IFNγ expression and release in cerebellar slices via non-viral gene delivery approach resulted in inflammation mediated myelin loss, thus making it a promising ex-vivo approach for studying the underlying mechanisms of demyelination in myelin-related diseases such as MS.

Topics: Animals; Cerebral Cortex; Demyelinating Diseases; Humans; Inflammation; Interferon-gamma; Methacrylates; Models, Biological; Multiple Sclerosis; Myelin Basic Protein; Myelin Sheath; Neurofilament Proteins; Polymers; Rats; Rats, Sprague-Dawley; Transfection; Tumor Necrosis Factor-alpha

In multiple sclerosis (MS) demyelination occurs in both white and gray matter. Here we introduce an electrophysiological approach for studying functional demyelination and myelin repair in rat hippocampus, a gray matter structure, which is frequently affected in patients suffering from MS.. Using a stereotaxic approach, bipolar stimulating and monopolar recording electrodes were respectively implanted into the perforant path and the dentate gyrus of the hippocampus of the adult male Sprague-Dawley rats weighing 280-320 g. Animals received intra-hippocampal injections of saline or lysolecithin (LPC) and afterward, changes in the parameters of field potentials recorded from the dentate gyrus granular cells in response to electrical stimulation of perforant path were investigated on days 7, 14, and 28, post-lesion. Changes in the electrophysiological parameters were compared with changes in the molecular markers of myelination (myelin basic protein, MBP) and repairing cells (Olig2).. On day 2, a significant decrease in the slope of the population excitatory postsynaptic potential (pEPSP) and the amplitude of population spike (PS) was observed. However, during days 7-28, these parameters were increased toward control. Decreased expression of MBP and increased expression of Olig2 were observed on days 2 and 7 while the expression levels were partially reversed toward control on day 28.. Our data showed the efficacy of field potential recording for studying demyelination and endogenous myelin repair in hippocampus. Changes in electrophysiological parameters were concomitant with the level of molecular markers. This recording method provides an opportunity for functional evaluation of myelin loss and repair and the effect of potential therapies.

Topics: Animals; Basic Helix-Loop-Helix Transcription Factors; Demyelinating Diseases; Evoked Potentials; Gene Expression; Hippocampus; Lysophosphatidylcholines; Male; Myelin Basic Protein; Myelin Sheath; Nerve Tissue Proteins; Oligodendrocyte Transcription Factor 2; Rats

Stem cell repair shows substantial translational potential for neurological injury, but the mechanisms of action remain unclear. This study aimed to investigate whether transplanted stem cells could induce comprehensive functional remyelination. Subventricular zone (SVZ)-derived adult neural precursor cells (aNPCs) were injected bilaterally into major cerebral white matter tracts of myelin-deficient shiverer mice on postnatal day (P) 0, P7, and P21. Tripotential NPCs, when transplanted in vivo, integrated anatomically and functionally into local white matter and preferentially became Olig2+, Myelin Associated Glycoprotein-positive, Myelin Basic Protein-positive oligodendrocytes, rather than Glial Fibrillary Acidic Protein-positive astrocytes or Neurofiliment 200-positive neurons. Processes interacted with axons and transmission electron microscopy showed multilamellar axonal ensheathment. Nodal architecture was restored and by quantifying these anatomical parameters a computer model was generated that accurately predicted action potential velocity, determined by ex vivo slice recordings. Although there was no obvious phenotypic improvement in transplanted shi/shis, myelinated axons exhibited faster conduction, lower activation threshold, less refractoriness, and improved response to high-frequency stimulation than dysmyelinated counterparts. Furthermore, they showed improved resilience to ischemic insult, a promising finding in the context of perinatal brain injury. This study describes, for the first time mechanistically, the functional characteristics and anatomical integration of nonimmortalized donor SVZ-derived murine aNPCs in the dysmyelinated brain at key developmental time points.

Topics: Adult Stem Cells; Animals; Axons; Basic Helix-Loop-Helix Transcription Factors; Cerebral Ventricles; Demyelinating Diseases; Disease Models, Animal; Mice; Myelin Basic Protein; Myelin Sheath; Myelin-Associated Glycoprotein; Nerve Tissue Proteins; Neural Stem Cells; Oligodendrocyte Transcription Factor 2; Oligodendroglia; Stem Cell Transplantation

We have previously demonstrated that Sox17 expression is prominent at developmental stages corresponding to oligodendrocyte progenitor cell (OPC) cycle exit and onset of differentiation, and that Sox17 promotes initiation of OPC differentiation. In this study, we examined Sox17 expression and regulation under pathological conditions, particularly in two animal models of demyelination/remyelination and in post-mortem multiple sclerosis (MS) brain lesions. We found that the number of Sox17 expressing cells was significantly increased in lysolecithin (LPC)-induced lesions of the mouse spinal cord between 7 and 30 days post-injection, as compared with controls. Sox17 immunoreactivity was predominantly detected in Olig2(+) and CC1(+) oligodendrocytes and rarely in NG2(+) OPCs. The highest density of Sox17(+) oligodendrocytes was observed at 2 weeks after LPC injection, coinciding with OPC differentiation. Consistent with these findings, in cuprizone-treated mice, Sox17 expression was highest in newly generated and in maturing CC1(+) oligodendrocytes, but low in NG2(+) OPCs during the demyelination and remyelination phases. In MS tissue, Sox17 was primarily detected in actively demyelinating lesions and periplaque white matter. Sox17 immunoreactivity was co-localized with NOGO-A+ post-mitotic oligodendrocytes both in active MS lesions and periplaque white matter. Taken together, our data: (i) demonstrate that Sox17 expression is highest in newly generated oligodendrocytes under pathological conditions and could be used as a marker of oligodendrocyte regeneration, and (ii) are suggestive of Sox17 playing a critical role in oligodendrocyte differentiation and lesion repair.

Topics: Aged; Animals; Antigens; Autophagy-Related Proteins; Basic Helix-Loop-Helix Transcription Factors; Brain; Bromodeoxyuridine; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Female; Humans; Intracellular Signaling Peptides and Proteins; Leukocyte Common Antigens; Lipopolysaccharides; Male; Mice; Mice, Inbred C57BL; Middle Aged; Monoamine Oxidase Inhibitors; Multiple Sclerosis; Myelin Basic Protein; Nerve Tissue Proteins; Oligodendrocyte Transcription Factor 2; Oligodendroglia; Proteoglycans; SOXF Transcription Factors; Time Factors; Up-Regulation

Hippocampus learning disturbance is a major symptom of patients with seizure, hence hippocampal dysfunction has essential role in worsening the disease. Hippocampal formation includes neurons and myelinated fibers that are necessary for acquisition and consolidation of memory, long-term potentiation and learning activity. The exact mechanism by which seizure can decrease memory and learning activity of hippocampus remains unknown. In the present study, electrical kindling-induced learning deficit in rats was evaluated by Morris water maze (MWM) test. The hippocampus was removed and changes in neurons and myelin sheaths around hippocampal fibers were investigated using histological and immunohistochemical methods. Demyelination was assessed by luxol fast blue staining, and immunohistological staining of myelin-binding protein (MBP). The TUNEL assay was used for evaluation of neuronal apoptosis and the glial fibriliary acetic protein (GFAP) was used for assessment of inflammatory reaction. The results indicated that electrical kindling of hippocampus could induce deficiency in spatial learning and memory as compared to control group. In addition, electrical kindling caused damage to the myelin sheath around hippocampal fibers and produced vast demyelination. Furthermore, an increase in the number of apoptotic cells in hippocampal slices was observed. In addition, inflammatory response was higher in kindled animals as compared to the control group. The results suggested that the decrease in learning and memory in kindled animals is likely due to demyelination and augmentation in apoptosis rate accompanied by inflammatory reaction in hippocampal neurons of kindled rats.

Topics: Animals; Apoptosis; Avoidance Learning; Demyelinating Diseases; Disease Models, Animal; Electron Transport Complex IV; Excitatory Amino Acid Antagonists; Gene Expression Regulation; Glial Fibrillary Acidic Protein; Hippocampus; Indoles; Ketamine; Kindling, Neurologic; Learning Disabilities; Locomotion; Male; Maze Learning; Myelin Basic Protein; Rats; Rats, Wistar; Reaction Time; Seizures

In stroke, a common cause of neurological disability in adults is that the myelin sheaths are lost through the injury or death of mature oligodendrocytes, and the failure of remyelination may be often due to insufficient proliferation and differentiation of oligodendroglial progenitors. In the current study, we used middle cerebral artery occlusion (MCAO) to induced transient focal cerebral ischemia, and found that WIN55, 212-2 augmented actively proliferating oligodendrocytes measured by CC1 immunoreactive cells within the peri-infarct areas. To establish whether these effects were associated with changes in myelin formation, we analyzed the expression of myelin basic protein (MBP) and myelin ultrastructure. We found that WIN55, 212-2 showed more extensive remyelination than vehicle at 14 days post injection (dpi). The extracellular signal-regulated kinase/mitogen-activated protein kinase (ERK/MAPK) signaling pathway may be involved in OPCs differentiation. To determine the regulatory effect of WIN55, 212-2 post-treatment on phospho-ERK 1/2 (p-ERK 1/2) after ischemia/reperfusion, Western blot analysis was performed. We found that WIN55, 212-2 regulated the phosphorylation level of the ERK 1/2 to promote OPCs survival and differentiation. Notably, cannabinoid receptor 1 is coupled to the activation of the ERK cascade. Following rimonabant combined treatment, the effect of WIN55, 212-2 on regulating the phosphorylation level of the ERK 1/2 was reversed, and the effect of accelerated myelin formation was partially inhibited. Together, we first found that WIN55, 212-2 promoted OPCs differentiation and remyelination through regulation of the level of the p-ERK 1/2 via cannabinoid receptor 1.

Topics: Animals; Antimetabolites; Benzoxazines; Blotting, Western; Brain Ischemia; Bromodeoxyuridine; Cannabinoid Receptor Antagonists; Cell Differentiation; Cell Survival; Demyelinating Diseases; Immunohistochemistry; Indicators and Reagents; Male; MAP Kinase Signaling System; Microscopy, Electron; Morpholines; Myelin Basic Protein; Myelin Sheath; Naphthalenes; Oligodendroglia; Phosphorylation; Piperidines; Pyrazoles; Rats; Rats, Sprague-Dawley; Receptor, Cannabinoid, CB1; Rimonabant; Stroke

In chronic demyelinating lesions of the central nervous system, insufficient generation of oligodendrocytes (OLs) is not due to a lack of oligodendrocyte precursor cells (OPCs), because the accumulation of OPCs and premyelinating OLs can be observed within these lesions. Here we sought to identify the basis for the failure of OLs to achieve terminal differentiation in chronic demyelinating lesions through the utilization of plp1-overexpressing (Plp(tg/-)) mice. These mice are characterized by progressive demyelination in young adults and chronic demyelinating lesions at more mature stages. We show that neural stem cells, which are the precursors of OL-lineage cells, are present in the Plp(tg/-) mouse brain and that their multipotentiality and ability to self-renew are comparable to those of wild-type adults in culture. Lineage-tracing experiments using a transgenic mouse line, in which an inducible Cre recombinase is knocked in at the Olig2 locus, revealed that Olig2-lineage cells preferentially differentiated into OPCs and premyelinating OLs, but not into astrocytes, in the Plp(tg/-) mouse brain. These Olig2-lineage cells matured to express myelin basic protein but after that their processes degenerated in the chronic demyelinating lesions of the Plp(tg/-) brain. These results indicate that in chronic demyelinated lesions more OL-lineage cells are produced as part of the repair process, but their processes degenerate after maturation.

Topics: Age Factors; Animals; Antineoplastic Agents, Hormonal; Basic Helix-Loop-Helix Transcription Factors; Brain; Cell Differentiation; Cell Lineage; Demyelinating Diseases; Disease Models, Animal; Gene Expression Regulation, Developmental; Green Fluorescent Proteins; Mice; Mice, Transgenic; Myelin Basic Protein; Myelin Proteolipid Protein; Nerve Tissue Proteins; Neural Stem Cells; Oligodendrocyte Transcription Factor 2; Oligodendroglia; Tamoxifen

After peripheral nerve injury, axons regenerate and become remyelinated by resident Schwann cells. However, myelin repair never results in the original myelin thickness, suggesting insufficient stimulation by neuronal growth factors. Upon testing this hypothesis, we found that axonal neuregulin-1 (NRG1) type III and, unexpectedly, also NRG1 type I restored normal myelination when overexpressed in transgenic mice. This led to the observation that Wallerian degeneration induced de novo NRG1 type I expression in Schwann cells themselves. Mutant mice lacking a functional Nrg1 gene in Schwann cells are fully myelinated but exhibit impaired remyelination in adult life. We suggest a model in which loss of axonal contact triggers denervated Schwann cells to transiently express NRG1 as an autocrine/paracrine signal that promotes Schwann cell differentiation and remyelination.

Topics: Animals; Animals, Newborn; Axons; Cell Proliferation; Cells, Cultured; Culture Media, Conditioned; Demyelinating Diseases; Disease Models, Animal; Early Growth Response Protein 2; Electric Stimulation; Enzyme Inhibitors; Evoked Potentials, Motor; Ganglia, Spinal; Gene Expression Regulation; Hedgehog Proteins; Ki-67 Antigen; Locomotion; Mice; Mice, Inbred C57BL; Mice, Transgenic; Microscopy, Electron, Transmission; Myelin Basic Protein; Myelin P0 Protein; Myelin Sheath; NAV1.6 Voltage-Gated Sodium Channel; Neuregulin-1; Neurons; Octamer Transcription Factor-6; Rats; Recovery of Function; RNA, Messenger; S100 Proteins; Schwann Cells; Sciatic Nerve; Sciatic Neuropathy; Signal Transduction; Statistics, Nonparametric; Time Factors

The rat dorsal root ganglia (DRG) model is a long-standing in vitro model for analysis of myelination in the peripheral nervous system. For performing systematic, high throughput analysis with transgenic animals, a simplified BL6 mouse protocol is indispensable. Here we present a stable and reliable protocol for myelinating co-cultures producing a high myelin ratio using cells from C57BL/6 mice. As an easy accessible and operable method, Sudan staining proved to be efficient in myelin detection for fixed cultures. Green fatty acid stain turned out to be highly reliable for analysis of the dynamic biological processes of myelination in vital cultures. Once myelinated we were able to induce demyelination by the addition of forskolin into the model system. In addition, we provide an optimised rat DRG protocol with significantly improved myelin ratio and a comparison of the protocols presented. Our results strengthen the value of ex vivo myelination models in neurobiology.

Topics: Animals; Azo Compounds; Blotting, Western; Boron Compounds; Cells, Cultured; Colforsin; Coloring Agents; Demyelinating Diseases; Fluorescent Dyes; Ganglia, Spinal; Mice; Mice, Inbred C57BL; Microscopy, Electron; Microscopy, Fluorescence; Myelin Basic Protein; Myelin P0 Protein; Myelin Sheath; Naphthalenes; Palmitic Acids; Primary Cell Culture; Rats; Rats, Inbred Lew; Rats, Wistar; Real-Time Polymerase Chain Reaction; Schwann Cells; Sciatic Nerve; Species Specificity; Staining and Labeling

Multiple sclerosis (MS) is a debilitating neurodegenerative disease characterized by axonal/neuronal damage that may be caused by defective remyelination. Current therapies aim to slow the rate of degeneration, however there are no treatment options that can stop or reverse the myelin sheath damage. Bone marrow mesenchymal stem cells (MSCs) are a potential candidate for the cell implantation-targeted therapeutic strategies, but the pro-remyelination effects of MSCs when directly injected into a demyelinated cord lesion have been questioned. Neurotrophin-3 (NT-3) has been shown to serve a crucial role in the proliferation, differentiation and maturation of oligodendrocyte lineages. Here, we showed that implantation of NT-3 gene-modified MSCs via a recombinant adenoviral vector (Adv) into a region of ethidium bromide (EB)-induced demyelination in the spinal cord resulted in significant improvement of locomotor function and restoration of electrophysiological properties in rats. The morphological basis of this recovery was evidenced by robust myelin basic protein (MBP) expression and the extensive remyelination. AdvNT-3-MSC implants promote the endogenous remyelinating cells to participate directly in myelination, which was confirmed under light and electron microscopy. Our study suggested that genetically modified MSCs could be a potential therapeutic avenue for improving the efficacy of stem cell treatment for neurodegenerative diseases such as MS.

Topics: Animals; Demyelinating Diseases; Female; Genetic Vectors; Mesenchymal Stem Cell Transplantation; Motor Activity; Myelin Basic Protein; Myelin Sheath; Neurotrophin 3; Rats; Rats, Sprague-Dawley; Recovery of Function; Spinal Cord Injuries; Thoracic Vertebrae

Sphingosine-1-phosphate receptors (S1PRs) are drug targets for the compound FTY720, which is the first oral therapy developed for treatment of relapsing-remitting multiple sclerosis. S1PRs play a variety of functional roles in the differentiation, proliferation, survival and/or migration of neurons and glia. In this study, rat organotypic cerebellar slice cultures were used to assess whether S1PRs play a role in demyelination induced by lysolecithin (LPC). The data demonstrated that FTY720 and SEW2871 (a S1P1R-specific agonist) inhibited LPC-induced demyelination as assessed by myelin basic protein (MBP) immunofluorescence. Treatment with both drugs for 48 h also induced an increase in S1P1R expression in astrocytes. Moreover, FTY720 and SEW2871 inhibited the release of several chemokines in conditions of LPC-induced demyelination, including LIX (CXCL5), MIP-1alpha, and MIP-3alpha. Taken together, the data suggest that activation of S1P1Rs prevents LPC-induced demyelination via a mechanism involving a reduction of chemotactic chemokine release. The study supports the concept that FTY720 attenuates demyelination by not only preventing S1PR-mediated T cell migration into the CNS but also by limiting cytokine communication between cells of the immune system and the CNS.

Topics: Animals; Animals, Newborn; Astrocytes; Cell Movement; Cerebellum; Chemokine CCL20; Chemokine CCL3; Chemokines; Demyelinating Diseases; Fingolimod Hydrochloride; Gene Expression Regulation; Immunosuppressive Agents; Lysophosphatidylcholines; Myelin Basic Protein; Neurofilament Proteins; Organ Culture Techniques; Oxadiazoles; Propylene Glycols; Rats; Rats, Wistar; Receptors, Lysosphingolipid; Sphingosine; Thiophenes

The significance of cerebral white matter (WM) demyelination in the cognitive decline of elderly humans is disputed. Cognitive decline also occurs in aged dogs, although the age-related changes that occur in the canine cerebral WM are yet to be studied, particularly with regard to their relevance to the WM alterations of elderly humans. The present study revealed age-dependent myelin loss in the frontal lobe WM of canine brains. The accumulation of ceroid-lipofuscin-laden phagocytes was observed in the perivascular spaces of the WM and was correlated with the decrease in myelination. Also, myelin basic protein was detected in some of the vacuoles of these phagocytes. In the WM, beta-amyloid (Aβ) was deposited focally in capillary walls, and colocalized with apolipoprotein E (Apo E). Note that the dog is homozygous for Apo E4, which genotype is related to capillary CAA in humans. These findings indicate that WM demyelination occurs in aged dogs as well as in aged humans, hence WM alterations may account for age-related behavioral changes of the dog. In conclusion, dogs are useful for chronological studies of age-related WM changes.

Topics: Aging; Amyloid beta-Peptides; Animals; Apolipoproteins E; Demyelinating Diseases; Disease Models, Animal; Dog Diseases; Dogs; Frontal Lobe; Leukoencephalopathies; Microscopy, Confocal; Myelin Basic Protein; Myelin Sheath; Nerve Fibers, Myelinated; Phagocytes

Chronic hypoxia in rodents induces white matter (WM) injury similar to that in human preterm infants. We used diffusion tensor imaging (DTI) and immunohistochemistry to study the impact of hypoxia in the immature ferret at two developmental time points relevant to the preterm and term brain.. On ex vivo imaging, the apparent diffusion coefficient (ADC) was decreased throughout the WM after 10 days of hypoxia (hypoxia from postnatal day 10 (P10) to P20 and killed at P20 (early hypoxia P20)), corresponding to increased astrocytosis and decreased myelination. Diffusion values normalized after 10 days of normoxia (hypoxia from P10 to P20 and killed at P30 (early hypoxia P30)), but immunohistochemistry revealed significant astrocytosis and hypomyelination. In contrast, ADC and anisotropy were increased after 10 days of hypoxia at a later developmental time point (hypoxia from P20 to P30 and killed at P30 (late hypoxia P30)), with less astrocytosis and more prominent myelination.. The patterns of alteration in imaging and histology varied in relation to the developmental time at which hypoxia occurred. Normalization of diffusion measures did not correspond to the normalization of underlying histopathology.. Ferrets were subjected to 10% hypoxia and divided into three groups: early hypoxia P20, early hypoxia P30, and late hypoxia P30.

Topics: Age Factors; Animals; Astrocytes; Biomarkers; Brain; Chronic Disease; Demyelinating Diseases; Diffusion Tensor Imaging; Disease Models, Animal; Ferrets; Glial Fibrillary Acidic Protein; Hypoxia, Brain; Immunohistochemistry; Leukoencephalopathies; Myelin Basic Protein

Multiple Sclerosis (MS) is an inflammatory demyelinating disease of the Central Nervous System which is characterized by multifocal demyelinated lesions dispersed throughout the brain. Although white matter lesions have been the most extensively studied, cortical demyelinaton lesions are also detected in MS brains. Cuprizone (CPZ)-induced demyelination in rodents has been widely used as a model for MS. Most of these studies focus on oligodendrocyte-rich structures, such as the corpus callosum (CC) and the cerebellar peduncles. However, it has been recently described that CPZ administration in mice also produces cortical demyelination, resembling some of the lesions found in MS patients. In this work we used CPZ-demyelinating model in Wistar rats to study demyelination in cortical forebrain areas. At the ultrastructural level, demyelination in the cortex was observed before detectable myelin loss in the subcortical white matter. During the course of CPZ intoxication Myelin Basic Protein immunodetection was decreased in cortical layers I-III due to a reduction in the number of cortical oligodendrocytes (OL). Oligodendroglial loss in CPZ-intoxicated rats correlated with an increase in the number of Glial Fibrillary Acidic Protein positive astrocytes and a shift in the location of Carbonic Anhydrase II from OL to astrocytes. After removal of CPZ from the diet, we evaluate intranasal Thyroid hormone (TH) effects on the progression of cortical lesions. As previously reported in the CC, TH treatment also accelerates remyelination rate in the cortex compared to rats undergoing spontaneous remyelination. Our results suggest that manipulation of TH levels could be considered as a strategy to promote remyelination process in the cortex and to prevent neuronal irreversible damage in patients suffering from MS.

Topics: Administration, Intranasal; Animals; Axons; Cerebral Cortex; Cuprizone; Demyelinating Diseases; Myelin Basic Protein; Myelin Sheath; Nerve Fibers, Myelinated; Oligodendroglia; Rats; Rats, Wistar; Thyroid Hormones

Golli myelin basic proteins are necessary for normal myelination, acting via voltage and store-dependent Ca(2+) entry at multiple steps during oligodendrocyte progenitor cell (OPC) development. To date nothing is known regarding the role of golli proteins in demyelination or remyelination events. Here the effects of golli ablation and overexpression in myelin loss and recovery were examined using the cuprizone (CPZ) model of demyelination/remyelination. We found severe demyelination in the corpus callosum (CC) of golli-overexpressing mice (JOE) during the CPZ treatment, which was accompanied by an increased number of reactive astrocytes and activation of microglia/macrophages. During demyelination of JOE brains, a significant increase in the number of proliferating OPCs was found in the CC as well as in the subventricular zone, and our data indicate that these progenitors matured and fully remyelinated the CC of JOE animals after CPZ withdrawal. In contrast, in the absence of golli (golli-KO mice) delayed myelin loss associated with a smaller immune response, and a lower number of OPCs was found in these mice during the CPZ treatment. Furthermore, incomplete remyelination was observed after CPZ removal in large areas of the CC of golli-KO mice, reflecting irregular recovery of the oligodendrocyte population and subsequent myelin sheath formation. Our findings demonstrate that golli proteins sensitize mature oligodendrocytes to CPZ-induced demyelination, while at the same time stimulate the proliferation/recruitment of OPCs during demyelination, resulting in accelerated remyelination.

Topics: Animals; Astrocytes; Calcium; Cell Differentiation; Cell Proliferation; Corpus Callosum; Cuprizone; Demyelinating Diseases; Macrophages; Mice; Mice, Knockout; Microglia; Myelin Basic Protein; Myelin Sheath; Neural Stem Cells; Oligodendroglia

To minimize as much as possible the neurological consequences from hypoxic-ischemic (HI) brain injury, neuroprotective strategies are urgently required. In this sense, there is growing interest in the neuroprotective potential of melatonin after perinatal asphyxia, due to its high efficacy, low toxicity and ready cross through the blood-brain barrier. Twenty six Wistar rats at postnatal day 7 were randomly assigned to: two hypoxic-ischemic groups: pups with the left common carotid artery ligated and then submitted to hypoxia (HI group) and animals that received a dose of 15 mg/kg melatonin just after the hypoxic-ischemic event and repeated twice with an interval of 24 hours (HI+MEL group). Pups without ischemia or hypoxia were used as controls (Sham group). Seven days after surgery, brains were collected and coronal sections Nissl-stained, TUNEL-labeled, or MBP- and GFAP-immunolabeled prior to determining brain infarct area, quantify surviving neurons and evaluate oligodendroglial injury and reactive astrogliosis. The number of surviving neurons showing a well preserved architecture in HI+MEL group was similar to that observed in the Sham group. Moreover, TUNEL-positive cells only appeared in the HI group. The ratio of left-to-right hemispheric MBP immunostaining showed a significant decrease in the HI group in comparison with Sham pups, which was restored after melatonin administration. Melatonin also reduced reactive gliosis. Thus, our results suggest that treatment with melatonin after neonatal hypoxia-ischemia led to a neuroprotective effect reducing cell death, white matter demyelination and reactive astrogliosis.

Topics: Animals; Animals, Newborn; Apoptosis; Biomarkers; Brain; Brain Infarction; Demyelinating Diseases; Disease Models, Animal; Glial Fibrillary Acidic Protein; Gliosis; Hypoxia-Ischemia, Brain; Immunohistochemistry; In Situ Nick-End Labeling; Leukoencephalopathies; Melatonin; Myelin Basic Protein; Neurons; Neuroprotective Agents; Rats; Rats, Wistar; Time Factors

Increasing evidence supports an important role of oligodendrocytes and myelination in the pathogenesis of schizophrenia. Oligodendrocytes are the myelin-producing cells in the central nervous system. To test the myelination dysfunction hypothesis of schizophrenia, possible myelination dysfunction was evaluated in a phencyclidine (PCP)-induced neurodevelopmental model of schizophrenia. On postnatal day (PND) 2, rat pups were treated with a total 14 subcutaneous daily injections of PCP (10mg/kg) or saline. PCP-injected rats showed schizophrenia-like behaviors including hyper-locomotor activity on PND 30 and prepulse inhibition deficit on PND 31. Cerebral myelination was measured by the expression of myelin basic protein (MBP), and cerebral mature oligodendrocytes were measured by the expression of glutathione S-transferase (GST)-π in rats. The results indicate that the expressions of MBP on PND 16, 22 and 32 and GST-π on PND 22 decreased in the frontal cortex of PCP-injected rats. Our results suggest that there was myelination impairment in the phencyclidine-induced schizophrenia animal model, and indicate that myelination may play an important role in the pathogenesis of schizophrenia.

Topics: Animals; Behavior, Animal; Brain; Demyelinating Diseases; Disease Models, Animal; Motor Activity; Myelin Basic Protein; Myelin Sheath; Oligodendroglia; Phencyclidine; Rats; Rats, Sprague-Dawley; Schizophrenia

Myelination is regulated by extracellular proteins, which control interactions between oligodendrocytes and axons. Semaphorins are repulsive axon guidance molecules, which control the migration of oligodendrocyte precursors during normal development and possibly in demyelinating diseases. We show here that the transmembrane semaphorin 6A (Sema6A) is highly expressed by myelinating oligodendrocytes in the postnatal mouse brain. In adult mice, Sema6A expression is upregulated in demyelinating lesions in cuprizone-treated mice. The analysis of the optic nerve and anterior commissure of Sema6A-deficient mice revealed a marked delay of oligodendrocyte differentiation. Accordingly, the development of the nodes of Ranvier is also transiently delayed. We also observed an arrest in the in vitro differentiation of purified oligodendrocytes lacking Sema6A, with a reduction of the expression level of Myelin Basic Protein. Their morphology is also abnormal, with less complex and ramified processes than wild-type oligodendrocytes. In myelinating co-cultures of dorsal root ganglion neurons and purified oligodendrocytes we found that myelination is perturbed in absence of Sema6A. These results suggest that Sema6A might have a role in myelination by controlling oligodendrocyte differentiation.

Topics: Age Factors; Animals; Animals, Newborn; Antigens, Differentiation; Basic Helix-Loop-Helix Transcription Factors; Brain; Bromodeoxyuridine; Cell Differentiation; Cells, Cultured; Coculture Techniques; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Embryo, Mammalian; Female; Ganglia, Spinal; Gene Expression Regulation, Developmental; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Monoamine Oxidase Inhibitors; Mutation; Myelin Basic Protein; Myelin Proteolipid Protein; Myelin Sheath; Nerve Tissue Proteins; Neurons; Oligodendrocyte Transcription Factor 2; Oligodendroglia; Pregnancy; Ranvier's Nodes; Rats; Rats, Sprague-Dawley; Rats, Wistar; Receptors, Cell Surface; RNA, Messenger; Semaphorins; Stem Cells; Time Factors; Transcription Factors

Encephalitis/encephalopathy is a rare, but severe, complication of pertussis. Here, we report a case of an unimmunized 7-year-old boy with confirmed pertussis complicated by acute encephalitis/encephalopathy. Eighteen days after the onset of pertussis, generalized seizures began. Magnetic resonance imaging (MRI) indicated that marked demyelination without cytotoxic edema may have occurred to the patient. Notably, this is the first report to show precise MRI findings of pertussis-associated encephalitis/encephalopathy. Markedly increased myelin basic protein levels in the cerebrospinal fluid were consistent with the MRI findings. There was no evidence of direct invasion of the causative bacterium or its products into the central nervous system. The levels of interleukin-6 and -10 in the cerebrospinal fluid were higher than those in serum. Taken together, we conclude that indirect immune-mediated mechanisms may have contributed to the pathogenesis of the encephalitis/encephalopathy.

Topics: Child; Demyelinating Diseases; Encephalitis; Humans; Immunization; Interleukin-10; Interleukin-6; Magnetic Resonance Imaging; Male; Myelin Basic Protein; Neuroimaging; Whooping Cough

Myelination is essential for proper functioning of the CNS. In this study, we have identified a mouse mutation, designated furue, which causes tremors and hypomyelination in the CNS, particularly in the spinal cord, but not in the sciatic nerve of the PNS. In the spinal cord of the furue mice, myelination of small-diameter axons was dramatically reduced, and differentiation of oligodendrocytes, the myelin-forming cells in the CNS, was inhibited. We subsequently found that the furue mutation was associated with a transgene insertion into the teneurin-4 (Ten-4, Ten-m4/Odz4) gene, encoding a transmembrane protein of unknown function. Ten-4 was strongly expressed in the spinal cord of wild-type mice and was induced during normal oligodendrocyte differentiation. In contrast, in the furue mice, the expression of Ten-4 was absent. Differentiation and cellular process formation of oligodendrocytes were inhibited in primary cell culture from the furue mice. Cell differentiation and process formation were also inhibited in the oligodendrocyte progenitor cell line CG-4 after suppression of Ten-4 expression by shRNA. Furthermore, Ten-4 positively regulated focal adhesion kinase, an essential signaling molecule for oligodendrocyte process formation and myelination of small-diameter axons. These findings suggest that Ten-4 is a novel regulator of oligodendrocyte differentiation and that it plays a critical role in the myelination of small-diameter axons in the CNS.

Topics: 2',3'-Cyclic Nucleotide 3'-Phosphodiesterase; Adenomatous Polyposis Coli Protein; Age Factors; Animals; Animals, Newborn; Antigens; Axons; Brain; Cell Differentiation; Cell Size; Central Nervous System; Demyelinating Diseases; Focal Adhesion Kinase 1; Galactosylceramidase; Gene Expression Regulation, Developmental; Humans; Membrane Proteins; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Mice, Transgenic; Microscopy, Electron, Transmission; Myelin Basic Protein; Neuroglia; Nuclear Proteins; Oligodendroglia; Organogenesis; Proteoglycans; Receptors, Interleukin-2; RNA, Messenger; RNA, Small Interfering; Transfection

Multiple sclerosis is an inflammatory demyelinating and neurodegenerative disorder of the central nervous system (CNS). The primary cause of the disease remains unknown, but an altered immune regulation with features of autoimmunity has generally been considered to play a critical role in the pathogenesis. Historically, lesion development has been attributed to activation of CD4 and CD8 T lymphocytes, B lymphocytes, and monocytes in the peripheral circulation and the migration of these cells through the blood-brain barrier to exert direct or indirect cytotoxic effects on myelin, oligodendrocytes and neuronal processes in the CNS. This broadly accepted concept was significantly influenced by the experimental autoimmune encephalitis (EAE) model, in which either immunization with myelin antigens or injection of a myelin antigen-specific T cell line into a recipient results in inflammatory demyelination in the CNS. More recent studies reveal that the loss of oligodendrocytes and neurons begins in the earliest stages of the disease and may not always be associated with blood-derived inflammatory cells. The pathology affects both the white and the gray matters and the clinical disability best correlates with the overall neurodegenerative process. These newer observations prompted several revisions of the classical concept of MS and facilitated a shift from using EAE to using other model systems. This chapter summarizes the classical and more contemporary concepts of MS, and provides methodologies for employing the cuprizone model for further explorations of the pathogenesis and treatment of the disease.

Topics: Animals; Apoptosis; Cuprizone; Demyelinating Diseases; Gene Expression Regulation; Humans; Immunohistochemistry; Immunologic Techniques; Mice; Mice, Inbred C57BL; Models, Biological; Multiple Sclerosis; Myelin Basic Protein; Oligodendroglia

In multiple sclerosis, microglia/macrophage activation and astrocyte reactivity are important components of the lesion environment that can impact remyelination. The current study characterizes these glial populations relative to expression of candidate regulatory molecules in cuprizone demyelinated corpus callosum. Importantly, periods of recovery after acute or chronic cuprizone demyelination are examined to compare conditions of efficient versus limited remyelination, respectively. Microglial activation attenuates after early demyelination. In contrast, astrocyte reactivity persists throughout demyelination and a 6-week recovery period following either acute or chronic demyelination. This astrocyte reaction is characterized by (a) early proliferation, (b) increased expression of GFAP (glial fibrillary acidic protein), Vim (vimentin), Fn1 (fibronectin) and CSPGs (chondroitin sulphate proteoglycans) and (c) elaboration of a dense network of processes. Glial processes elongated in the axonal plane persist throughout lesion areas during both the robust remyelination that follows acute demyelination and the partial remyelination that follows chronic demyelination. However, prolonged astrocyte reactivity with chronic cuprizone treatment does not progress to barrier formation, i.e. dense compaction of astrocyte processes to wall off the lesion area. Multiple candidate growth factors and inflammatory signals in the lesion environment show strong correlations with GFAP across the acute cuprizone demyelination and recovery time course, yet there is more divergence across the progression of chronic cuprizone demyelination and recovery. However, differential glial scar formation does not appear to be responsible for differential remyelination during recovery in the cuprizone model. The astrocyte phenotype and lesion characteristics in this demyelination model inform studies to identify triggers of non-remyelinating sclerosis in chronic multiple sclerosis lesions.

Topics: Animals; Cell Proliferation; Chondroitin Sulfate Proteoglycans; Cicatrix; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Gene Expression Regulation; Glial Fibrillary Acidic Protein; Gliosis; Male; Mice; Mice, Inbred C57BL; Monoamine Oxidase Inhibitors; Myelin Basic Protein; Statistics as Topic; Time Factors

Multiple sclerosis (MS) is a demyelinating disease of the central nervous system that presents with variable pathologies that may reflect different disease-causing mechanisms. Existing animal models of MS induce pathology using either local injection of gliotoxins or stimulation of the immune system with myelin-related peptides. In none of these models is the primary cellular target well characterized, and although demyelination is a hallmark pathological feature in MS, it is unclear to what extent this reflects local oligodendrocyte loss. To unambiguously identify the effects of oligodendrocyte death in the absence of inflammatory stimulation, we developed a method for experimentally inducing programmed cell death selectively in mature oligodendrocytes and assessed the effects on demyelination, immunological stimulation, and gliosis. The resulting pathology is discussed relative to observed MS pathologies.. Oligodendrocyte apoptosis was induced in the adult rat brain using a lentivirus to express experimentally inducible caspase 9 (iCP9) cDNA under transcriptional control of the promoter for myelin basic protein, which is oligodendrocyte-specific. Activation of iCP9 was achieved by distal injection of a small molecule dimerizer into the lateral ventricle resulting in localized, acute oligodendrocyte apoptosis.. Induced oligodendrocyte apoptosis resulted in rapid demyelination and robust, localized microglial activation in the absence of peripheral immune cell infiltration. Lesion borders showed layers of preserved and degraded myelin, whereas lesion cores were demyelinated but only partially cleared of myelin debris. This resulted in local proliferation and mobilization of the oligodendrocyte progenitor pool.. This approach provides a novel model to understand the pathological changes that follow from localized apoptosis of myelinating oligodendrocytes. It provides the first direct proof that initiation of apoptosis in oligodendrocytes is sufficient to cause rapid demyelination, gliosis, and a microglial response that result in lesions sharing some pathological characteristics with a subset of MS lesions.

Topics: Animals; Apoptosis; Apoptotic Protease-Activating Factor 1; Basic Helix-Loop-Helix Transcription Factors; Brain; Caspase 9; Cell Count; Cells, Cultured; Demyelinating Diseases; Disease Models, Animal; Enzyme Activation; Gangliosides; Glial Fibrillary Acidic Protein; Gliotoxin; Green Fluorescent Proteins; Immunosuppressive Agents; Myelin Basic Protein; Nerve Tissue Proteins; O Antigens; Oligodendrocyte Transcription Factor 2; Oligodendroglia; Protein Multimerization; Rats; Tacrolimus; Transduction, Genetic; Transfection

Failure of remyelination is a critical impediment to recovery in multiple sclerosis (MS). Chondroitin sulfate proteoglycans (CSPGs) have been reported to accumulate in MS lesions, and we thus examined the functional roles of CSPGs on oligodendrocyte precursor cells (OPCs), oligodendrocytes, and remyelination.. We evaluated the expression of CSPGs in lysolecithin-injected mouse spinal cord, an animal model of demyelination and spontaneous remyelination. The functional impact of CSPGs on OPCs and remyelination was investigated using cultured adult murine and human OPCs and by treating demyelinated mice with xyloside to reduce the CSPG deposition that occurred following injury.. Early and robust upregulation of CSPGs following lysolecithin-induced demyelination was cleared during remyelination. In culture, CSPGs anchored onto the substratum reduced the adhesion of mouse and human OPCs and their subsequent morphological differentiation into process-bearing oligodendrocytes. Soluble CSPGs added to already adherent OPCs reduced the development of processes, whereas the acquisition of mature myelin proteins was unimpeded. Stripe assays of alternating CSPG and control substrata confirmed the nonpermissive nature of CSPGs for OPC adhesion and morphological differentiation. Enzymatic degradation of CSPGs with chondroitinase ABC was sufficient to overcome CSPG-dependent inhibition of human oligodendrocytes. Finally, in vivo xyloside treatment to reduce CSPG synthesis in lysolecithin-demyelinated mice increased numbers of OPCs and oligodendrocytes in lesions, and culminated in improved remyelination.. These results identify CSPGs as a nonpermissive substrate for OPCs and oligodendrocytes, and as a prominent impediment to remyelination. The data suggest the requirement for the neutralization of CSPGs for repair after demyelination.

Topics: Analysis of Variance; Animals; Calcium-Binding Proteins; Cell Adhesion; Cell Differentiation; Cell Line, Transformed; Chondroitin ABC Lyase; Chondroitin Sulfate Proteoglycans; Demyelinating Diseases; Disease Models, Animal; Female; Glial Fibrillary Acidic Protein; Glycosides; Humans; In Vitro Techniques; Indoles; Lysophosphatidylcholines; Mice; Microfilament Proteins; Myelin Basic Protein; Myelin Proteins; Nerve Regeneration; Platelet-Derived Growth Factor; Spinal Cord; Stem Cells; Time Factors; Up-Regulation

Interaction of Schwann cells with axons triggers signal transduction that drives expression of Pou3f1 and Egr2 transcription factors, which in turn promote myelination. Signal transduction appears to be mediated, at least in part, by cyclic adenosine monophosphate (cAMP) because elevation of cAMP levels can stimulate myelination in the absence of axon contact. The mechanisms by which the myelinating signal is conveyed remain unclear.. By analyzing mutations that disrupt myelination in zebrafish, we learned that Dynein cytoplasmic 1 heavy chain 1 (Dync1h1), which functions as a motor for intracellular molecular trafficking, is required for peripheral myelination. In dync1h1 mutants, Schwann cell progenitors migrated to peripheral nerves but then failed to express Pou3f1 and Egr2 or make myelin membrane. Genetic mosaic experiments revealed that robust Myelin Basic Protein expression required Dync1h1 function within both Schwann cells and axons. Finally, treatment of dync1h1 mutants with a drug to elevate cAMP levels stimulated myelin gene expression.. Dync1h1 is required for retrograde transport in axons and mutations of Dync1h1 have been implicated in axon disease. Our data now provide evidence that Dync1h1 is also required for efficient myelination of peripheral axons by Schwann cells, perhaps by facilitating signal transduction necessary for myelination.

Topics: Animals; Animals, Genetically Modified; Cell Differentiation; Cell Movement; Central Nervous System; Colforsin; Cyclic AMP; Cytoplasmic Dyneins; Demyelinating Diseases; Embryo, Nonmammalian; Gene Expression Regulation, Developmental; Green Fluorescent Proteins; Larva; Microscopy, Electron, Transmission; Morpholinos; Mutation; Myelin Basic Protein; Myelin Sheath; Peripheral Nerves; RNA; Schwann Cells; Zebrafish; Zebrafish Proteins

Human umbilical cord blood is a rich source of pluripotent mesenchymal stem cells and possesses significant advantages over other stem cell sources such as the embryo and bone marrow. In the present study, we aimed to investigate the potential of human umbilical cord blood-derived pluripotent mesenchymal stem cells (hUCB) to myelinate the axons of shiverer mice brains. We also investigated the effect of hUCB treatment on regulation of myelin basic protein in vitro in PC-12 cells, which are normally not myelinated. The results of our study clearly demonstrated that hUCB survive and migrate in vivo and has the potential to myelinate shiverer mice brains. The expression level of myelin basic protein, a major component of the myelin sheath, has been significantly increased in vivo and in vitro as revealed by Western blot, reverse transcription (RT)-polymerase chain reaction, immunohistochemistry, immunocytochemistry, and fluorescent in situ hybridization results. Further, transmission electron microscopic images of hUCB-treated shiverer mice brains showed several layers of myelin around the axons compared with a thin and fragmented layer of myelin in untreated animals. Moreover, the frequency of shivering was diminished 1 month after hUCB treatment in shiverer mice. Our results strongly indicated that hUCB transplantation could be an effective means of treating demyelinating or hypomyelinating disorders.

Topics: Animals; Axons; Blotting, Western; Brain; Cell Differentiation; Cell Movement; Cells, Cultured; Cord Blood Stem Cell Transplantation; Demyelinating Diseases; Female; Fetal Blood; Gene Expression; Humans; Immunohistochemistry; Mesenchymal Stem Cells; Mice; Mice, Transgenic; Microscopy, Electron, Transmission; Myelin Basic Protein; Myelin Sheath; Rats; Recovery of Function; Reverse Transcriptase Polymerase Chain Reaction; Shivering; Up-Regulation

Myelin oligodendrocyte glycoprotein (MOG) may be implicated in the immunopathogenesis of multiple sclerosis (MS) inducing demyelination in the animal model of MS. In adults reported anti-MOG antibody frequencies have been variable across a number of studies and can also be detected in controls.. To measure antibodies against MOG in paediatric patients with demyelinating disorders of the central nervous system and in controls.. Serum antibodies against MOG and myelin basic protein were measured by ELISA, flow cytometry (FACS) and in the liquid phase in 11 children with acute disseminated encephalomyelitis (ADEM), 22 children with MS, seven children with acute viral encephalitis and 13 healthy controls. The serostatus of Epstein-Barr virus (EBV) infections were assessed.. Anti-MOG antibodies, measured either by ELISA or FACS were exclusively detected in children with demyelination. In ADEM these antibodies were highly reactive. Anti-MBP reactivity was detectable equally in all groups. The presence of either autoantibodies did not associate with EBV serostatus, age, gender or disease course.. This study independently corroborates recently published results of seroprevalence and specificity of the assay. Due to their low sensitivity anti-MOG antibodies will not serve as disease-specific biomarkers, but could help to support the diagnosis of ADEM in difficult cases.

Topics: Adolescent; Autoantibodies; Biomarkers; Chi-Square Distribution; Child; Child, Preschool; Demyelinating Diseases; Diagnosis, Differential; Encephalitis, Viral; Encephalomyelitis, Acute Disseminated; Enzyme-Linked Immunosorbent Assay; Epstein-Barr Virus Infections; Female; Flow Cytometry; France; Germany; Humans; Immunity, Humoral; Male; Myelin Basic Protein; Myelin Proteins; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Nerve Tissue Proteins; Predictive Value of Tests; Retrospective Studies; Transcription Factors

The authors examined whether (1)H-magnetic resonance spectroscopy (MRS) can identify damage to the centrum semiovale in the subacute phase after CO exposure.. Subjects comprised 29 adult patients who were treated with hyperbaric oxygenation within a range of 4-95 h (mean 18.7 h) after CO exposure. Subjects were classified into three groups according to clinical behaviours: Group A, patients with transit acute symptoms only; Group P, patients with persistent neurological symptoms; and Group D, patients with 'delayed neuropsychiatric sequelae' occurring after a lucid interval. MRS of bilateral centrum semiovale was performed 2 weeks after CO inhalation for all patients and 13 healthy volunteers. The mean ratios of choline-containing compounds/creatine ((mean)Cho/Cr) and N-acetylaspartate/Cr ((mean)NAA/Cr) for bilateral centrum semiovale were calculated and compared between the three CO groups and controls. Myelin basic protein (MBP) concentration in cerebrospinal fluid was examined at 2 weeks to evaluate the degree of demyelination in patients.. MBP concentration was abnormal for almost all patients in Groups P and D, but was not abnormal for any Group A patients. The (mean)Cho/Cr ratios were significantly higher in Groups P and D than in Group A. No significant difference in (mean)NAA/Cr ratio was seen between the three pathological groups and controls. A significant correlation was identified between MBP and (mean)Cho/Cr ratio.. These results suggest that the Cho/Cr ratio in the subacute phase after CO intoxication represents early demyelination in the centrum semiovale, and can predict chronic neurological symptoms.

Topics: Brain; Brain Diseases, Metabolic; Carbon Monoxide Poisoning; Choline; Creatine; Demyelinating Diseases; Female; Follow-Up Studies; Humans; Hyperbaric Oxygenation; Magnetic Resonance Spectroscopy; Male; Middle Aged; Myelin Basic Protein; Nerve Fibers, Myelinated; Protons; Time Factors

Myelination is one of the fundamental biological processes in the development of vertebrate nervous system. Disturbance of myelination is found to be associated with progression in many neurological diseases such as multiple sclerosis. Tremendous efforts have been made to develop novel therapeutic agents that prevent demyelination and/or promote remyelination. These efforts need to be accompanied by the development of imaging tools that permit direct quantification of myelination in vivo. In this work, we describe a novel near-infrared fluorescence imaging technique that is capable of direct quantification of myelination in vivo. This technique is developed based on a near-infrared fluorescent probe, 3,3'-diethylthiatricarbocyanine iodide (DBT) that readily enters the brain and specifically binds to myelinated fibers. In vivo imaging studies were first conducted in two animal models of hypermyelination and hypomyelination followed by longitudinal studies in the cuprizone-induced demyelination/remyelination mouse model. Quantitative analysis suggests that DBT is a sensitive and specific imaging probe of myelination, which complements other current myelin-imaging modalities and is of low cost.

Topics: Analysis of Variance; Animals; Benzothiazoles; Carbocyanines; Corpus Callosum; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Longitudinal Studies; Mice; Mice, Inbred C57BL; Mice, Neurologic Mutants; Mice, Transgenic; Myelin Basic Protein; Myelin Proteolipid Protein; Myelin Sheath; Permeability; Protein Binding; Proto-Oncogene Proteins c-akt; Spectroscopy, Near-Infrared; Time Factors

Multiple studies indicate that T-cells play a major role in the pathogenesis of experimental autoimmune encephalomyelitis (EAE), the animal model of multiple sclerosis, but recently an involvement of antibodies has also been discussed. The aim of our study was to examine the effects of myelin basic protein (MBP) immunization on survival of neurons, alteration of antibody reactivity, and microglia in the retinal ganglion cell layer.. EAE was induced in rats by immunization with MBP. Intraocular pressure (IOP) measurements and funduscopies were performed regularly. Neuron cell density was evaluated on cresyl-stained retinal flatmounts. IgG antibody deposition and activated microglia were detected in retina and optic nerve sections via immunohistology. The intensity of autoreactive IgG antibodies was quantified in successive serum samples via tissue arrays.. Significant loss of neurons was detected 6 weeks after immunization (p < 0.05). At the same time, IgG antibody deposits accumulated in the retina and the optic nerve of EAE animals and a significant microglia turn-over to activation was observed. The level of IgG antibody reactivity against retina and optic nerve tissue continuously increased (p < 0.05). While clinical parameters indicated typical EAE progression, we observed no changes in IOP (p > 0.9) or abnormalities in fundi.. Immunization with MBP not only causes neuron loss in the retinal ganglion cell layer, but also triggers antibody reactivity against ocular tissue. Possibly some of these antibodies are involved in the induction of neuronal apoptosis. This study suggests that, apart from T-cell mediation, alteration of antibody reactivity and activated microglia do also influence the ocular pathomechanisms in the EAE model.

Topics: Animals; Apoptosis; Autoantibodies; Cell Count; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Fluorescent Antibody Technique, Indirect; Immunoenzyme Techniques; Immunoglobulin G; Intraocular Pressure; Male; Microglia; Myelin Basic Protein; Ophthalmoscopy; Optic Nerve; Rats; Rats, Inbred Lew; Retina; Retinal Ganglion Cells

Multiple sclerosis is a demyelinating disease of the central nervous system which only affects humans. This makes it difficult to study at a molecular level, and to develop and test potential therapies that may change the course of the disease. The development of therapies to promote remyelination in multiple sclerosis is a key research aim, to both aid restoration of electrical impulse conduction in nerves and provide neuroprotection, reducing disability in patients. Testing a remyelination therapy in the many and various in vivo models of multiple sclerosis is expensive in terms of time, animals and money. We report the development and characterisation of an ex vivo slice culture system using mouse brain and spinal cord, allowing investigation of myelination, demyelination and remyelination, which can be used as an initial reliable screen to select the most promising remyelination strategies. We have automated the quantification of myelin to provide a high content and moderately-high-throughput screen for testing therapies for remyelination both by endogenous and exogenous means and as an invaluable way of studying the biology of remyelination.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Analysis of Variance; Animals; Animals, Newborn; Antigens; Bromodeoxyuridine; Cell Proliferation; Central Nervous System; Demyelinating Diseases; Green Fluorescent Proteins; Lysophosphatidylcholines; Mice; Mice, Transgenic; Models, Biological; Myelin Basic Protein; Myelin Sheath; Neurofilament Proteins; Oligodendroglia; Organ Culture Techniques; Proteoglycans; RNA, Messenger; Stem Cells

Neural stem cell-based approaches to repair damaged white matter in the central nervous system have shown great promise; however, the optimal cell population to employ in these therapies remains undetermined. A default mechanism of neural induction may function during development, and in embryonic stem cells (ESCs) neural differentiation is elicited in the absence of any extrinsic signaling in minimal, serum-free culture conditions. The default mechanism can be used to derive clonal neurosphere-forming populations of neural stem cells that have been termed leukemia inhibitory factor-dependent primitive neural stem cells (pNSCs), which subsequently give rise to fibroblast growth factor 2-dependent definitive NSCs (dNSCs). Here we characterized the neural differentiation pattern of these two cell types in vitro and in vivo when transplanted into the dysmyelinated spinal cords of shiverer mice. We compared the differentiation pattern to that observed for neural stem/progenitor cells derived from the adult forebrain subependymal zone [adult neural precursor cells (aNPCs)]. dNSCs produced a differentiation pattern similar to that of aNPCs in vitro and in the shiverer model in vivo, where both cell types produced terminally differentiated oligodendrocytes that associated with host axons and expressed myelin basic protein. This is the first demonstration of the in vivo differentiation of NSCs, derived from ESCs through the default mechanism, into the oligodendrocyte lineage. We conclude that dNSCs derived through the default pathway of neural induction are a similar cell population to aNPCs and that the default mechanism is a promising approach to generate NSCs from pluripotent cell populations for use in cell therapy or other research applications.

Topics: Animals; Cell Count; Cell Differentiation; Cell Line; Cell Lineage; Coculture Techniques; Demyelinating Diseases; Embryonic Stem Cells; Gene Expression Profiling; Mice; Mice, Knockout; Microscopy, Fluorescence; Myelin Basic Protein; Neural Stem Cells; Spheroids, Cellular; Spinal Cord

Mitochondrial content within axons increases following demyelination in the central nervous system, presumably as a response to the changes in energy needs of axons imposed by redistribution of sodium channels. Myelin sheaths can be restored in demyelinated axons and remyelination in some multiple sclerosis lesions is extensive, while in others it is incomplete or absent. The effects of remyelination on axonal mitochondrial content in multiple sclerosis, particularly whether remyelination completely reverses the mitochondrial changes that follow demyelination, are currently unknown. In this study, we analysed axonal mitochondria within demyelinated, remyelinated and myelinated axons in post-mortem tissue from patients with multiple sclerosis and controls, as well as in experimental models of demyelination and remyelination, in vivo and in vitro. Immunofluorescent labelling of mitochondria (porin, a voltage-dependent anion channel expressed on all mitochondria) and axons (neurofilament), and ultrastructural imaging showed that in both multiple sclerosis and experimental demyelination, mitochondrial content within remyelinated axons was significantly less than in acutely and chronically demyelinated axons but more numerous than in myelinated axons. The greater mitochondrial content within remyelinated, compared with myelinated, axons was due to an increase in density of porin elements whereas increase in size accounted for the change observed in demyelinated axons. The increase in mitochondrial content in remyelinated axons was associated with an increase in mitochondrial respiratory chain complex IV activity. In vitro studies showed a significant increase in the number of stationary mitochondria in remyelinated compared with myelinated and demyelinated axons. The number of mobile mitochondria in remyelinated axons did not significantly differ from myelinated axons, although significantly greater than in demyelinated axons. Our neuropathological data and findings in experimental demyelination and remyelination in vivo and in vitro are consistent with a partial amelioration of the supposed increase in energy demand of demyelinated axons by remyelination.

Topics: Adult; Aged; Aged, 80 and over; Animals; Antigens, CD; Antigens, Differentiation, Myelomonocytic; Axons; Brain; Cells, Cultured; Coculture Techniques; Demyelinating Diseases; Disease Models, Animal; Ethidium; Female; Ganglia, Spinal; HLA Antigens; Humans; Leukocyte Common Antigens; Lysophosphatidylcholines; Male; Microscopy, Electron, Transmission; Middle Aged; Mitochondria; Multiple Sclerosis; Myelin Basic Protein; Neurofilament Proteins; Rats; Rats, Sprague-Dawley; Schwann Cells; Voltage-Dependent Anion Channels

The peroxisome-proliferator-activated receptor gamma coactivator 1 α (PGC1α) has been implicated in the pathogenesis of several neurodegenerative disorders, including Huntington's disease (HD). Recent data demonstrating white matter abnormalities in PGC1α knock-out (KO) mice prompted us to examine the role of PGC1α in CNS myelination and its relevance to HD pathogenesis. We found deficient postnatal myelination in the striatum of PGC1α KO mice, accompanied by a decrease in myelin basic protein (MBP). In addition, brain cholesterol, its precursors, and the rate-limiting enzymes for cholesterol synthesis, HMG CoA synthase (HMGCS1) and HMG CoA reductase (HMGCR), were also reduced in PGC1α KO mice. Moreover, knockdown of PGC1α in oligodendrocytes by lentiviral shRNA led to a decrease in MBP, HMGCS1, and Hmgcr mRNAs. Chromatin immunoprecipitations revealed the recruitment of PGC1α to MBP promoter in mouse brain, and PGC1α over-expression increased MBP and SREBP-2 promoter activity, suggesting that PGC1α regulates MBP and cholesterol synthesis at the transcriptional level. Importantly, expression of mutant huntingtin (Htt) in primary oligodendrocytes resulted in decreased expression of PGC1α and its targets HmgcS1, Hmgcr, and MBP. Decreased expression of MBP and deficient myelination were found postnatally and in adult R6/2 mouse model of HD. Diffusion tensor imaging detected white matter abnormalities in the corpus callosum of R6/2 mice, and electron microscopy revealed thinner myelin sheaths and increased myelin periodicity in BACHD [bacterial artificial chromosome (BAC)-mediated transgenic model for Huntington's disease] mice expressing full-length mutant Htt. Together, these data suggest that PGC1α plays a role in postnatal myelination and that deficient PGC1α activity in oligodendrocytes may contribute to abnormal myelination in HD.

Topics: Analysis of Variance; Animals; Blotting, Western; Brain; Chromatin Immunoprecipitation; Demyelinating Diseases; Diffusion Tensor Imaging; Disease Models, Animal; Huntington Disease; Hydroxymethylglutaryl CoA Reductases; Hydroxymethylglutaryl-CoA Synthase; Immunohistochemistry; Mice; Mice, Knockout; Myelin Basic Protein; Myelin Sheath; Oligodendroglia; Reverse Transcriptase Polymerase Chain Reaction; Transcription Factors

Microglial activation in multiple sclerosis has been postulated to contribute to long-term neurodegeneration during disease. Fingolimod has been shown to impact on the relapsing remitting phase of disease by modulating autoreactive T-cell egress from lymph organs. In addition, it is brain penetrant and has been shown to exert multiple effects on nervous system cells.. In this study, the impact of fingolimod and other sphingosine-1-phosphate receptor active molecules following lysophosphotidyl choline-induced demyelination was examined in the rat telencephalon reaggregate, spheroid cell culture system. The lack of immune system components allowed elucidation of the direct effects of fingolimod on CNS cell types in an organotypic situation.. Following demyelination, fingolimod significantly augmented expression of myelin basic protein in the remyelination phase. This increase was not associated with changes in neurofilament levels, indicating de novo myelin protein expression not associated with axonal branching. Myelin wrapping was confirmed morphologically using confocal and electron microscopy. Increased remyelination was associated with down-regulation of microglial ferritin, tumor necrosis factor alpha and interleukin 1 during demyelination when fingolimod was present. In addition, nitric oxide metabolites and apoptotic effectors caspase 3 and caspase 7 were reduced during demyelination in the presence of fingolimod. The sphingosine-1-phosphate receptor 1 and 5 agonist BAF312 also increased myelin basic protein levels, whereas the sphingosine-1-phosphate receptor 1 agonist AUY954 failed to replicate this effect on remyelination.. The results presented indicate that modulation of S1P receptors can ameliorate pathological effectors associated with microglial activation leading to a subsequent increase in protein and morphological markers of remyelination. In addition, sphingosine-1-phosphate receptor 5 is implicated in promoting remyelination in vitro. This knowledge may be of benefit for treatment of chronic microglial inflammation in multiple sclerosis.

Topics: Animals; Biomarkers; Cell Culture Techniques; Cells, Cultured; Demyelinating Diseases; Female; Fingolimod Hydrochloride; Humans; Immunosuppressive Agents; Lysophosphatidylcholines; Microglia; Multiple Sclerosis, Relapsing-Remitting; Myelin Basic Protein; Myelin Sheath; Nerve Degeneration; Pregnancy; Propylene Glycols; Rats; Rats, Sprague-Dawley; Receptors, Lysosphingolipid; Spheroids, Cellular; Sphingosine

Central nervous system hypomyelination is a feature common to a number of transgenic (Tg) mouse lines that express a variety of unrelated exogenous (i.e. non-central nervous system) transgenes. In this report, we document hypomyelination structurally by immunocytochemistry and functionally in the Tg line MBP-JE, which over expresses the chemokine CCL2 (MCP-1) within oligodendrocytes targeted by a myelin basic protein (MBP) promoter. Analysis of hypomyelinated optic nerves of Tg mice revealed progressive decrease in oligodendrocyte numbers with age (p < 0.01). Although molecular mechanisms underlying hypomyelination in this and other Tg models remain largely unknown, we present preliminary findings on oligodendrocyte progenitor cell (OPC) cultures in which, although OPC expressed CCR2, the receptor for CCL2, treatment with CCL2 had no significant effect on OPC proliferation, differentiation, or apoptosis. We suggest that hypomyelination in the MBP-JE model might not be due to CCL2 expression but rather the result of transcriptional dysfunction related to random insertion of the MBP promoter that disrupts myelinogenesis and leads to oligodendrocyte demise. Because an MBP promoter is a common denominator in most Tg lines displaying hypomyelination, we hypothesize that use of myelin gene sequences in the regulator region of Tg constructs might underlie this perturbation of myelination in such models.

Topics: Animals; Demyelinating Diseases; Disease Models, Animal; Mice; Mice, Transgenic; Myelin Basic Protein; Myelin Sheath; Neural Stem Cells; Oligodendroglia; Promoter Regions, Genetic

Cognitive deficits have been observed in patients with multiple sclerosis (MS) due to hippocampal insults. Antioxidant vitamins D and E are suggested for patients suffering from neurodegenerative diseases like MS, while their mechanisms of action are not well understood. Here, we have tried to study the effects of these vitamins on demyelination, cell death, and remyelination of rat hippocampus following local ethidium bromide (EB) injection. Animals received 100 mg/kg vitamin E or 5 microg/kg of vitamin D3 for 2, 7, or 28 days. The extent of demyelination, myelin staining intensity, and expression of myelin basic protein and caspase-3 were investigated using histological and immunoblotting verification. Administration of EB alone caused demyelination, cell death, and afterward an endogenous repair. Vitamins E and D3 reduced the EB-induced damage and increased the endogenous remyelination of hippocampus. Although the anti-apoptotic effect of these vitamins and protection against demyelination were predictable based on their antioxidant effect, our results indicated the positive effect of vitamins E and D3 on process of remyelination by endogenous progenitor cells and supported their possible therapeutic effects in the context of demyelinating diseases like MS.

Topics: Animals; Antioxidants; Apoptosis; Caspase 3; Cholecalciferol; Demyelinating Diseases; Enzyme Inhibitors; Ethidium; Hippocampus; Humans; Male; Myelin Basic Protein; Nerve Regeneration; Rats; Rats, Sprague-Dawley; Vitamin E; Vitamins

The myelin vacuolation (mv) rat exhibits hypomyelination and vacuole formation in the myelin throughout the CNS, caused by a null mutation in the attractin gene. Myelin alterations in the spinal cord of mv rats progress during postnatal development and are more prominent in the white matter. In contrast, microglial activation is confined to the gray matter of mv rats. We here investigate the distribution and expression patterns of major CNS myelin proteins in the spinal cord of mv rats during the development of the myelin lesions. Immunohistochemical and Western blot analyses demonstrated a considerable reduction in the expression of major CNS myelin proteins both in the white and gray matter of mv rats, which was consistent with the morphological alterations of myelin sheaths. Real-time PCR analysis revealed a significant decrease in expression of proteolipid protein (PLP) mRNA both in the white and gray matter of mv rats. However, there was no significant difference between control and mv rats in the cell number of PLP mRNA-positive oligodendrocytes either in the white or gray matter, suggesting an impairment of myelin protein production by oligodendrocytes. Our results indicate that myelinogenesis but not oligodendrogenesis is severely altered both in the white and gray matter of mv rats.

Topics: Animals; Central Nervous System; Demyelinating Diseases; Disease Models, Animal; Gene Expression Regulation; Membrane Proteins; Microglia; Mutation; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin Sheath; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Oligodendroglia; Rats; Rats, Sprague-Dawley; Rats, Transgenic; RNA, Messenger; Vacuoles

Between November, 2006, and May, 2008, a subacute neurological syndrome affected workers from two swine abattoirs in Minnesota and Indiana who had occupational exposure to aerosolised porcine brain. We aimed to describe the pathogenic and immunological characteristics of this illness.. All patients from two abattoirs who presented or were referred to the Mayo Clinic (Rochester, MN, USA) with neurological symptoms were included. We recorded details of exposure to aerosolised brain tissue and did comprehensive neurological, laboratory, neuroimaging, electrophysiological, pathological, and autoimmune serological assessments. Healthy controls were recruited from the community and from workers at the plant in Minnesota.. 24 patients were identified (21 from Minnesota, three from Indiana). The shortest duration from first exposure to symptom onset was 4 weeks. No infectious agent that could trigger disease was identified. All patients developed polyradiculoneuropathy, which was usually sensory predominant and painful. Two patients had initial CNS manifestations: transverse myelitis and meningoencephalitis. Nerve conduction studies localised abnormalities to the most proximal and distal nerve segments. Quantitative sensory and autonomic testing revealed involvement of large and small sensory fibres and sweat fibres. MRI showed prominent abnormalities of roots and ganglia. Nerve biopsies identified mild demyelination, axonal degeneration, and perivascular inflammation. Protein concentrations were high in the CSF of 18 (86%) of 21 patients. Sera from all patients and 29 (34%) of 85 unaffected workplace controls (but none of 178 community controls) had a distinctive neural-reactive IgG; 75% of patients' sera contained an IgG specific to myelin basic protein. Seropositivity correlated directly with exposure risk in patients and controls. 17 patients required immunomodulatory therapies, six improved spontaneously, and one was lost to follow-up after exposure stopped.. The neurological disorder described is autoimmune in origin and is related to occupational exposure to multiple aerosolised porcine brain tissue antigens. The pattern of nerve involvement suggests vulnerability of nerve roots and terminals where the blood-nerve barrier is most permeable.. Mayo Clinic Foundation; Minnesota Department of Health; Centers for Disease Control and Prevention.

Topics: Abattoirs; Adult; Aerosols; Air Pollutants, Occupational; Animals; Autoantibodies; Autoimmune Diseases of the Nervous System; Brain; Demyelinating Diseases; Disease Outbreaks; Female; Follow-Up Studies; Humans; Indiana; Inhalation Exposure; Male; Mice; Middle Aged; Minnesota; Myelin Basic Protein; Occupational Diseases; Occupational Exposure; Polyradiculoneuropathy; Swine; Young Adult

IL-15 initially identified as a T proliferating cytokine has several structural and biological similarities with IL-2 and has been associated with a number of autoimmune diseases. Because of the scarcity of information available on the role of IL-15 in MS pathogenesis, we have investigated how the absence of IL-15 affected the development of experimental autoimmune encephalomyelitis, a mouse model of MS. Following immunization of IL-15(-/-) and C57BL/6 mice with MOG(35-55), we observed a more severe neurological impairment in the IL-15 knockout mice than in the wild-type group. The enhanced disease severity in IL-15(-/-) mice was associated with greater demyelination in the spinal cord, increased immune cell infiltration and inflammation. These events may be related to the higher CD4/CD8 ratio and the almost absent NK cell activity, congenital immune features of IL-15KO mice. Moreover, we found that the fractalkine receptor CX3CR1 was overexpressed in the spinal cord of IL-15(-/-) mice, mainly localized on infiltrating CD8(+) T cells. How these findings are contributing to the aggravated EAE development in IL-15 KO mice remain unclear and need to be further investigated.

Topics: Analysis of Variance; Animals; CD4 Antigens; CD8 Antigens; CX3C Chemokine Receptor 1; Cytokines; Demyelinating Diseases; Eliminative Behavior, Animal; Encephalomyelitis, Autoimmune, Experimental; Flow Cytometry; Gene Expression Regulation; Glycoproteins; Interleukin-15; Killer Cells, Natural; Mice; Mice, Inbred C57BL; Mice, Knockout; Myelin Basic Protein; Myelin-Oligodendrocyte Glycoprotein; Neutrophil Infiltration; Peptide Fragments; Receptors, Chemokine; Spinal Cord

In rats, iron deficiency produces an alteration in myelin formation. However, there is limited information on the effects of this condition on oligodendroglial cell (OLGc) proliferation and maturation. In the present study, we further analyzed the hypomyelination associated with iron deficiency by studying the dynamics of oligodendrogenesis. Rats were fed control (40 mg Fe/kg) or iron-deficient (4 mg Fe/kg) diets from gestation day 5 until postnatal day 3 (P3) or 11 (P11). OLGc proliferation, migration and differentiation were investigated before and after an intracranial injection of apotransferrin at 3 days of age (P3). The proliferating cell population was evaluated at P3. Iron-deficient (ID) animals showed an increase in the oligodendrocyte precursors cell (OPC) population in comparison with controls. The overall pattern of migration of cells labeled with BrdU was investigated at P11. Iron deficiency increased the amount of BrdU(+) cells in the corpus callosum (CC) and decreased OLGc maturation and myelin formation. Changes in nerve conduction were analyzed by measuring visual evoked potentials. Latency and amplitude were significantly disturbed in ID rats compared with controls. Both parameters were substantially normalized when animals were treated with a single intracranial injection of 350 ng apotransferrin (aTf). The current results give support to the idea that iron deficiency increases the number of proliferating and undifferentiated cells in the CC compared with the control. Treatment with aTf almost completely reverted the effects of iron deficiency, both changing the migration pattern and increasing the number of mature cells in the CC and myelin formation.

Topics: Age Factors; Analysis of Variance; Animals; Animals, Newborn; Apoproteins; Basic Helix-Loop-Helix Transcription Factors; Body Weight; Brain; Bromodeoxyuridine; Cell Count; Cell Differentiation; Cell Proliferation; Corpus Callosum; Demyelinating Diseases; Electroencephalography; Evoked Potentials, Visual; Female; Gene Expression Regulation, Developmental; Hematocrit; Iron Deficiencies; Myelin Basic Protein; Nerve Tissue Proteins; Neural Cell Adhesion Molecule L1; Oligodendroglia; Photic Stimulation; Pregnancy; Prenatal Exposure Delayed Effects; Proliferating Cell Nuclear Antigen; Rats; Sialic Acids; Transferrin

Two types of interventions to remyelinate the adult demyelinated central nervous system were investigated in heterozygous transgenic mice overexpressing the proteolipid protein gene. 1) A cocktail of trophic factors, "TS1," was directed toward the activation of the endogenous pool of neural progenitors to increase the number of myelinating oligodendrocytes (OL) in the brain. 2) A combinatorial approach in which OL progenitors were coinjected with TS1 into the corpus callosum of wild-type and He4e transgenic mice that displayed hindlimb paralysis. The levels of locomotor ability in these mice were evaluated after a single treatment. The data showed that a single administration of either one of the interventions had similar therapeutic effects, alleviating the symptoms of demyelination and leading to the recovery of hindlimb function. Histological and immunofluorescent examination of brain sections showed extensive remyelination that was sufficient to reverse hindlimb paralysis in transgenic mice. When the interventions were administered prior to hindlimb paralysis, He4e mice were able to walk up to 1 year of age without paralysis.

Topics: Amidines; Animals; Cell Movement; Cells, Cultured; Central Nervous System; Corpus Callosum; Culture Media, Conditioned; Demyelinating Diseases; Disease Models, Animal; Dose-Response Relationship, Drug; Exploratory Behavior; Gangliosides; Gene Expression Regulation; Green Fluorescent Proteins; Hindlimb; Intercellular Signaling Peptides and Proteins; Intermediate Filament Proteins; Mice; Mice, Transgenic; Microscopy, Confocal; Myelin Basic Protein; Myelin Proteolipid Protein; Nerve Tissue Proteins; Nestin; Neurofilament Proteins; Neuroglia; Recovery of Function; Time Factors

White matter disorders can involve injury to myelin or axons but the respective contribution of each to clinical course is difficult to evaluate non-invasively. Here, to develop a paradigm for further investigations of axonal pathology by MRI, we compared two genetic mouse models exhibiting relatively selective axonal or myelin deficits using quantitative MRI relaxography of the transverse relaxation times (T2) in vivo and ultrastructural morphometry. In HM-DKO mice, which lack genes encoding the heavy (NF-H) and medium (NF-M) subunits of neurofilaments, neurofilament content of large myelinated axons of the central nervous system (CNS) is markedly reduced in the absence of changes in myelin thickness and volume. In shiverer mutant mice, which lack functional myelin basic protein, CNS myelin sheath formation is markedly reduced but neurofilament content is normal. We observed increases in T2 in nearly all white matter in shiverer mice compared to their wild type, while more subtle increases in T2 were observed in HM-DKO in the corpus callosum. White matter T2 was generally greater in shiverer mice than HM-DKO mice. Ultrastructural morphometry of the corpus callosum, which exhibited the greatest T2 differences, confirmed that total cross-sectional area occupied by axons was similar in the two mouse models and that the major ultrastructural differences, determined by morphometry, were an absence of myelin and larger unmyelinated axons in shiverer mice and absence of neurofilaments in HM-DKO mice. Our findings indicate that T2 is strongly influenced by myelination state and axonal volume, while neurofilament structure within the intra-axonal compartment has a lesser effect upon single compartment T2 estimates.

Topics: Animals; Brain; Demyelinating Diseases; Disease Models, Animal; Humans; Magnetic Resonance Imaging; Mice; Mice, Inbred C3H; Mice, Inbred C57BL; Mice, Knockout; Mice, Neurologic Mutants; Myelin Basic Protein; Neurofilament Proteins

Angiostrongylus cantonensis causes eosinophilic meningitis or meningoencephalitis, yet little is known about demyelination caused by this parasite. To define the course of demyelination caused by A. cantonensis, we analyzed the expression of myelin proteins including myelin-associated glycoprotein (MAG), myelin basic protein (MBP), myelin-associated oligodendrocytic basic protein (MOBP), and proteolipid protein (PLP) in brain and cerebrospinal fluid (CSF)-like fluid of infected and uninfected BALB/c mice. In A. cantonensis-infected mice, the expression of MAG, MBP, MOBP, and PLP mRNAs in brain tissue was decreased, while expression of the corresponding proteins was significantly increased in CSF-like fluid. Light microscopy revealed perivascular infiltrates in the brain during meningoencephalitis, suggesting that the cause of demyelination in angiostrongyliasis was immune system attack on the oligodendrocytic myelin sheath and subsequent release of myelin proteins into the CSF. Thus, intracerebral myelin breakdown in angiostrongyliasis may be a response to inflammatory mediators and the cause of increased myelin proteins in the CSF-like fluid.

Topics: Angiostrongylus cantonensis; Animals; Blotting, Western; Demyelinating Diseases; Histocytochemistry; Male; Meningoencephalitis; Mice; Mice, Inbred BALB C; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Random Allocation; Reverse Transcriptase Polymerase Chain Reaction; RNA; Statistics, Nonparametric; Strongylida Infections

The mechanisms that regulate the developmental potential of adult neural progenitor populations under physiological and pathological conditions remain poorly defined. Glutamic acid decarboxylase 65 (GAD65)- and Doublecortin (Dcx)-expressing cells constitute major progenitor populations in the adult mouse subventricular zone (SVZ). Under normal physiological conditions, SVZ-derived GAD65-positive and Dcx-positive cells expressed the transcription factor Pax6 and migrated along the rostral migratory stream to the olfactory bulb to generate interneurons. After lysolecithin-induced demyelination of corpus callosum, however, these cells altered their molecular and cellular properties and migratory path. Demyelination upregulated chordin in the SVZ, which redirected GAD65-positive and Dcx-positive progenitors from neuronal to glial fates, generating new oligodendrocytes in the corpus callosum. Our findings suggest that the lineage plasticity of SVZ progenitor cells could be a potential therapeutic strategy for diseased or injured brain.

Topics: Adult Stem Cells; Animals; Bromodeoxyuridine; Cell Count; Cell Differentiation; Cell Lineage; Cell Movement; Cerebral Ventricles; Corpus Callosum; Demyelinating Diseases; Doublecortin Domain Proteins; Doublecortin Protein; Gene Expression Regulation; Glutamate Decarboxylase; Glycoproteins; Green Fluorescent Proteins; Intercellular Signaling Peptides and Proteins; Lysophosphatidylcholines; Mice; Mice, Transgenic; Microtubule-Associated Proteins; Myelin Basic Protein; Nerve Tissue Proteins; Neurogenesis; Neuronal Plasticity; Neurons; Neuropeptides; Oligodendroglia

Simvastatin, the most widely used cholesterol-lowering drug, has been reported to protect the adult brain from ischemia. Nevertheless, little is known about its action on developing brain after stroke. Although a few reports have found recently that simvastatin displays anti-inflammation and anti-apoptosis properties and improves the cognitive and morphological consequences in the neonatal rats after hypoxia-ischemia (HI) damage, to our best knowledge, there has been no study of the effect of it on myelin formation after neonatal brain damage. Therefore, we investigated whether simvastatin could promote the myelination of oligodendrocytes in the neonatal rats after HI and explored the possible role of microglial responses in this process.. Postnatal day 7 Sprague-Dawley rats were subjected to HI. White matter integrity and myelination were evaluated by the densitometry of myelin basic protein (MBP) immunostaining. OX-42 immunoreactivity and nissl staining were used for identifying microglial responses and the structure changes of white matter and adjacent gray matter after HI. Simvastatin was administrated prophylactically to rats.. HI induced serious hypomyelination especially in external and internal capsules 3 and 7 days after HI, accompanying with microglial activation remarkably. Simvastatin treatment greatly increased the densities of MBP immunostaining, inhibited microglial activation and reduced the numbers of pyknotic cells and neuronal loss.. The present study shows that simvastatin treatment in neonatal rats attenuates HI-induced developing oligodendrocytes injury and hypomyelination. Its anti-inflammatory properties via suppression of microglial activation are likely to contribute to this action. It provides experimental evidence to support the neuroprotective effects of statins in neonatal ischemic stroke.

Topics: Age Factors; Analysis of Variance; Animals; Animals, Newborn; Body Weight; CD11b Antigen; Demyelinating Diseases; Disease Models, Animal; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Hypoxia-Ischemia, Brain; Myelin Basic Protein; Rats; Rats, Sprague-Dawley; Simvastatin

Topics: Brain; Child; Demyelinating Diseases; Female; Humans; Leukocytes; Magnetic Resonance Imaging; Myelin Basic Protein; Spinal Cord; Spinal Puncture

Cuprizone (bis-cyclohexanone oxaldihydrazone) was previously shown to induce demyelination in white matter enriched brain structures. In the present study we used the cuprizone demyelination model in transgenic mice expressing the enhanced green fluorescent protein (GFP) under the 2'-3'-cyclic nucleotide 3'-phosphodiesterase (CNPase) promoter. The use of these particular transgenic mice allows easy detection of cells belonging to the entire oligodendroglial (OLG) lineage, ranging from OLG precursors to mature myelinating OLGs. We were able to evaluate the precise extent of oligodendroglial cell damage and recovery within the murine adult central nervous system (CNS) after inducing demyelination by acute cuprizone intoxication. A generalized loss of GFP+ cells was observed after cuprizone exposure and correlated with a decline in myelin basic protein (MBP) expression. OLGs were depleted in many brain areas that were previously thought to be unaffected by cuprizone treatment. Thus, in addition to the well-known cuprizone effects on the medial corpus callosum, we also found a loss of GFP+ cells in most brain structures, particularly in the caudatus putamen, cortex, anterior commissure, olfactory bulb, hippocampus, optic chiasm, brainstem, and cingulum. Loss of GFP+ cells was accompanied by extensive astrogliosis and microglial activation, although neurons were not affected. Interestingly, cuprizone-treated animals showed both activation of GFAP expression and a higher proliferation rate in subventricular zone cells. A week after cuprizone removal from the diet, GFP+ oligodendroglial cells began repopulating the damaged structures. GFP expression precedes that of MBP and allows OLG detection before myelin restoration.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Astrocytes; Brain; Cell Count; Cell Proliferation; Cuprizone; Demyelinating Diseases; Glial Fibrillary Acidic Protein; Gliosis; Green Fluorescent Proteins; Mice; Mice, Inbred C57BL; Mice, Transgenic; Microglia; Myelin Basic Protein; Neurons; Oligodendroglia; Promoter Regions, Genetic

The study shows the dynamic expression of connexin47 (Cx47) in oligodendrocytes and myelin of mice, either in myelinogenesis occurring in early development or in an experimental model of new-myelinogenesis of adult mice. Cx47 first appeared in the embryonic mouse brain at E10.5 successively the expression increased, principally in regions populated by developing oligodendrocytes. The expression declined postnatally toward adulthood and immunoreactivity was restricted to a few specific areas, such as the corpus callosum, the striatum, the cerebellum, and the spinal cord. Since the expression of Cx47 in developing oligodendrocytes preceded those of Cx32 and Cx29, a role of Cx47 in myelinogenesis was postulated. This hypothesis was tested in a model of re-myelination, which principally involved the corpus callosum, occurring in adult mice by treatment with cuprizone. Cx47 was upregulated during demyelination and recovered during the remyelination phase. During demyelination, Cx47 was first over-expressed in the corpus callosum and later, when the myelin virtually disappeared in the injured areas, Cx47 was expressed in astrocytes located inside and closely around the demyelinated areas. The remyelination of injured areas occurred after stopping the administration of cuprizone and continued to complete recovery. In this period the expression of Cx47 shifted from astrocytes to newly-formed myelin. Thus, Cx47 exhibits in this model a transient and de novo expression in astrocytes with a topographic segregation in the injured areas, only when oligodendrocytes and the myelin were most severely affected. Taken as a whole the evidence suggests that Cx47 play a key role in myelination.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Animals, Newborn; Brain; Connexins; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Embryo, Mammalian; Enzyme-Linked Immunosorbent Assay; Female; Gene Expression Regulation, Developmental; Gliosis; HeLa Cells; Humans; Immunoprecipitation; Mice; Mice, Inbred C57BL; Myelin Basic Protein; Myelin Sheath; Neuronal Plasticity; Oligodendroglia; Transfection

Multiple sclerosis (MS) is an inflammatory demyelinating disease of the CNS and remyelination in MS ultimately fails. Although strategies to promote myelin repair are eagerly sought, mechanisms underlying remyelination in vivo have been elusive. CXCR2 is expressed on neutrophils and oligodendrocyte lineage cells in the CNS. CXCR2-positive neutrophils facilitate inflammatory demyelination in demyelination models such as experimental autoimmune encephalomyelitis (EAE) and cuprizone intoxication. Systemic injection of a small molecule CXCR2 antagonist at the onset of EAE decreased demyelinated lesions. These results left the cellular target of the CXCR2 antagonist uncertain and did not clarify whether CXCR2 blockade prevented demyelination or promoted remyelination. Here, we show that the actions of CXCR2 on nonhematopoietic cells unexpectedly delay myelin repair. Bone marrow chimeric mice (Cxcr2(+/-)-->Cxcr2(-/-) and Cxcr2(+/-)-->Cxcr2(+/+)) were subjected to two distinct models of myelin injury. In all cases, myelin repair was more efficient in Cxcr2(+/-)-->Cxcr2(-/-) animals. Oligodendrocyte progenitor cells (OPCs) in demyelinated lesions of Cxcr2(+/-)-->Cxcr2(-/-) mice proliferated earlier and more vigorously than in tissues from Cxcr2(+/-)--> Cxcr2(+/+) animals. In vitro demyelinated CNS slice cultures also showed better myelin repair when CXCR2 was blocked with neutralizing antibodies or was genetically deleted. Our results suggest that CXCR2 inactivation permits optimal spatiotemporal positioning of OPCs in demyelinating lesions to receive local proliferative and differentiating signals. Given that CXCR2 exerts dual functions that promote demyelination and decrease remyelination by actions toward hematopoietic cells and nonhematopoietic cells, respectively, our findings identify CXCR2 as a promising drug target for clinical demyelinating disorders.

Topics: Animals; Animals, Newborn; Antibodies; Bone Marrow; Cell Differentiation; Central Nervous System Stimulants; Cerebellum; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Encephalomyelitis, Autoimmune, Experimental; Flow Cytometry; Freund's Adjuvant; Glycoproteins; In Vitro Techniques; Leukocyte Common Antigens; Lipopolysaccharides; Mice; Mice, Inbred C57BL; Mice, Knockout; Microscopy, Electron, Transmission; Myelin Basic Protein; Myelin Proteolipid Protein; Myelin Sheath; Myelin-Oligodendrocyte Glycoprotein; Nerve Regeneration; Neurologic Examination; Oligodendroglia; Peptide Fragments; Picrotoxin; Proliferating Cell Nuclear Antigen; Receptors, Interleukin-8B; Recovery of Function; Severity of Illness Index; Stem Cells; Time Factors

Multiple sclerosis (MS) patients may suffer from optic disturbances. Toxin-induced demyelinations have frequently been developed to investigate the cellular and structural aspects of demyelination and remyelination processes, separately. The present study describes functional consequence of lysolecithin (LPC)-induced lesion in the adult rat optic nerves and chiasm by recording the visual evoked potentials (VEPs) from the visual cortex and its correlation with myelin basic protein (MBP) expression in lesion site. Records of VEP were obtained at 2, 7, 14 and 28 days post-injection. We observed that the VEPs generated by light stimuli progressively changed in both amplitude and latency after the lesion as well as in comparison with those generated in control animals. These observations were confirmed through measurement of mRNA expression level for MBP which is one of the important genes expressed in mature oligodendrocytes and Schwann cells. The level of MBP mRNAs in demyelinated chiasm and optic nerves decreased following lysolecithin injection with its least value on day 7, and then it increased to the control level 14 days post-lesion. However, it continued to increase even after that and reached a maximum level 28 days post lesion. Results of the present paper show that, LPC injection in the chiasm share functional and molecular alterations which are found in demyelinating disorders in both the optic nerves and chiasm and also these alterations were coming back to level of control animal on 28 days post lesion, which is typically seen in myelin repair process. The present paper provides new insights into the experimental toxin-induced models that may be useful for evaluating the functional recovery of demyelinated optic nerves and chiasm following various repairing strategies. It also seems to be useful for studying the protective or remyelinating effects of different therapies in e.g. optic apparatus which is more affected by MS.

Topics: Age Factors; Animals; Demyelinating Diseases; Evoked Potentials, Visual; Female; Gene Expression Regulation; Lysophosphatidylcholines; Myelin Basic Protein; Myelin Sheath; Optic Chiasm; Optic Nerve; Rats; Rats, Sprague-Dawley; Transcription Factors

To study inflammatory reactions occurring in relation to demyelination, aggregating rat brain cell cultures were subjected to three different demyelinating insults, i.e., (i) lysophosphatidylcholine (LPC), (ii) interferon-gamma combined with lipopolysaccharide (IFN-gamma+LPS), and (iii) anti-MOG antibodies plus complement (alpha-MOG+C). Demyelination was assessed by measuring the expression of myelin basic protein (MBP) and myelin oligodendrocyte glycoprotein (MOG), and the activity of 2',3'-cyclic nucleotide 3'-phosphohydrolase (CNP). The accompanying inflammatory reactions were examined by the quantification of microglia-specific staining, by immunostaining for glial fibrillary acidic protein (GFAP), and by measuring the mRNA expression of a panel of inflammation-related genes. It was found that all three demyelinating insults decreased the expression of MBP and MOG, and induced microglial reactivity. LPC and alpha-MOG+C, but not IFN-gamma+LPS, decreased CNP activity; they also caused the appearance of macrophagic microglia, and increased GFAP staining indicating astrogliosis. LPC affected also the integrity of neurons and astrocytes. LPC and IFN-gamma+LPS upregulated the expression of the inflammation-related genes IL-6, TNF-alpha, Ccl5, Cxcl1, and iNOS, although to different degrees. Other inflammatory markers were upregulated by only one of the three insults, e.g., Cxcl2 by LPC; IL-1beta and IL-15 by IFN-gamma+LPS; and IFN-gamma by alpha-MOG+C. These findings indicate that each of the three demyelinating insults caused distinct patterns of demyelination and inflammatory reactivity, and that of the demyelinating agents tested only LPC exhibited general toxicity.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Antibodies; Brain; Cells, Cultured; Choline O-Acetyltransferase; Cytokines; Demyelinating Diseases; Drug Interactions; Embryo, Mammalian; Glial Fibrillary Acidic Protein; Glutamate Decarboxylase; Immunologic Factors; Interferon-gamma; Lectins; Lysophosphatidylcholines; Myelin Basic Protein; Myelin Proteins; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Neurons; Polysaccharides; Rats; RNA, Messenger; Tumor Necrosis Factor-alpha; Up-Regulation

Loss of aquaporin 4 and glial fibrillary acidic protein (GFAP) with necrosis and demyelination is a prominent pathologic feature of neuromyelitis optica (NMO). However, the clinicopathologic significance of astrocytic damage and its relation with demyelination are unknown.. To analyze clinical and pathologic values of a CSF biomarker of astrocytic damage in NMO.. We measured the levels of GFAP, S100B, myelin basic protein (MBP), and neurofilament H (NF-H) in CSF obtained from patients with NMO (n = 33), multiple sclerosis (MS) (n = 27), acute disseminated encephalomyelitis (ADEM), ischemia, meningitis, and other neurologic disease controls (OND).. The CSF-GFAP levels during relapse in NMO (2,476.6 +/- 8,815.0 ng/mL) were significantly higher than those in MS (0.8 +/- 0.4 ng/mL) and OND (0.7 +/- 0.5 ng/mL), and much beyond those in ADEM (14.1 +/- 27.4 ng/mL). The sensitivity and specificity of CSF-GFAP for NMO was 90.9% and 76.9% in all, but the specificity improved above 90% in cases limited to demyelinating diseases. CSF-S100B showed a similar trend but was less remarkable. In contrast, MBP and NF-H are not different between NMO and MS. Following treatments, the CSF-GFAP rapidly decreased to a normal level, but CSF-MBP remained high. There were strong correlations between the CSF-GFAP, CSF-S100B, or CSF-MBP levels and Expanded Disability Status Scale (EDSS) or spinal lesion length in the acute phase (r > 0.6). Only CSF-GFAP correlated with EDSS at 6-month follow-up (r = 0.51) in NMO.. Astrocytic damage reflected by elevated CSF glial fibrillary acidic protein is a clinically relevant, primary pathologic process in neuromyelitis optica, and is far more severe than demyelination.

Topics: Adult; Aged; Antibodies; Aquaporin 4; Astrocytes; Biomarkers; Central Nervous System; Demyelinating Diseases; Disability Evaluation; Encephalomyelitis, Acute Disseminated; Female; Follow-Up Studies; Glial Fibrillary Acidic Protein; Humans; Male; Methylprednisolone; Middle Aged; Multiple Sclerosis; Myelin Basic Protein; Nerve Growth Factors; Nervous System Diseases; Neurofilament Proteins; Neuromyelitis Optica; Neuroprotective Agents; S100 Calcium Binding Protein beta Subunit; S100 Proteins; Statistics, Nonparametric

A link between diffuse axonal loss and diffuse inflammation has been established in the brain of patients with progressive multiple sclerosis (MS). In the present paper, we sought to determine whether such a link could be similarly demonstrated in the spinal cord of patients with progressive MS.. A neuropathological quantitative assessment of inflammation and axonal loss was performed in the cervical spinal cord of 18 patients with progressive MS and 5 control subjects.. As previously reported, we found a mean 25% decrease of axonal density in the normal-appearing white matter (NAWM) of MS versus control spinal cords. T-cell perivascular infiltrates were rare, but a robust diffuse inflammation was observed in both the normal-appearing parenchyma and the meninges. The extent of diffuse axonal loss in the NAWM correlated with both the density of major histocompatibility complex (MHC) class II(+) microglia in the NAWM and, surprisingly, the density of CD3(+) T cells in the meninges. Interestingly, close interactions between T cells and MHC class II(+) macrophages were observed in the meninges of spinal cords from MS patients.. Recent studies assigned a major role to meningeal B-cell follicles in the pathophysiology of secondary progressive MS. The present work also emphasizes the link between meningeal inflammation and parenchymal lesions and points to a specific role exerted by both meningeal T cells and activated microglia in diffuse axonal loss in the spinal cord.

Topics: Adult; Antigens, CD; Axons; Cytokines; Demyelinating Diseases; Disease Progression; Female; Humans; Male; Meninges; Middle Aged; Multiple Sclerosis; Myelin Basic Protein; Myelitis; Neutrophil Infiltration; Postmortem Changes; Spinal Cord; Statistics, Nonparametric; T-Lymphocytes

Polysialic acid (PSA) is a carbohydrate polymer added post-translationally on the neural cell adhesion molecule (NCAM) affecting its adhesion properties. It has been suggested that the presence of PSA in demyelinated lesions in multiple sclerosis could prevent axon-glia interactions inhibiting spontaneous remyelination. The enzyme St8siaIV is one of the two polysialyltransferases responsible for PSA synthesis, and it is predominantly active during adult life. Here we treated 8-10-weeks old St8siaIV deficient and wild-type mice for 5 weeks with cuprizone, which is a reliable model for de- and remyelination in the corpus callosum and cortex. Developmental myelination of the St8siaIV knock-out mice was not disturbed and adult mice showed normal myelin protein expression. Demyelination did not differ between transgenic and wild-type mice but early myelin protein re-expression and thus remyelination were accelerated in St8siaIV knock-out mice during the first week after withdrawal of the toxin. This was mainly due to enhanced oligodendrocyte precursor cells (OPC) differentiation and to a lesser extent to OPC recruitment. These data are proof of principle that PSA expression interferes at least to some extent with remyelination in vivo.

Topics: Age Factors; Analysis of Variance; Animals; Animals, Newborn; Antigens; Antigens, Differentiation; Cell Count; Cell Differentiation; Cerebral Cortex; Corpus Callosum; Cuprizone; Demyelinating Diseases; Gene Expression Regulation; Glial Fibrillary Acidic Protein; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Monoamine Oxidase Inhibitors; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Neural Cell Adhesion Molecules; Neuroglia; Neurons; Nogo Proteins; Proteoglycans; Regeneration; Sialic Acids; Sialyltransferases

Transplantation of neural precursor cells has been proposed as a possible approach for replacing missing or damaged central nervous system myelin. Neonatal and adult myelin-deficient shiverer (shi) mice, bearing a mutation of the myelin basic protein (MBP) gene, have been used extensively as hosts for testing cell engraftment, migration, and myelination, but relatively little progress has been made in reversing shi motor deficits. Here we describe a prenatal cell replacement strategy, showing that embryonic stem cells injected into shi blastocyst embryos can generate chimeric mice with strong and widespread immunoreactive MBP expression throughout the brain and a behavioral (motor) phenotype that appears essentially rescued.

Topics: Animals; Ataxia; Basic Helix-Loop-Helix Transcription Factors; Demyelinating Diseases; Embryo, Mammalian; Embryonic Stem Cells; Genotype; Glial Fibrillary Acidic Protein; Mice; Mice, Neurologic Mutants; Mutation; Myelin Basic Protein; Phosphopyruvate Hydratase; Tremor

The process of demyelination occurring in diseases as multiple sclerosis is usually investigated in animal models. A major drawback of animal models is that only one condition can be tested per animal, necessitating many animals and systemic effects are factors to be considered. The aim of the study was to develop a reproducible in vitro model for de- and remyelination using whole brain spheroid cultures and lysophosphatidyl choline (LPC). In spheroid cultures, single cell suspensions of embryonic day 15 rodent brain cells reaggregate under constant rotation. Three-dimensional contacts form between the central nervous system cell types present. Multilayered myelin is maximal in four-week old cultures. A week of repeated exposure to LPC led to 30% loss of MBP protein concentration and 2',3'-cyclic nucleotide 3'-phosphodiesterase activity measurements in both rat and mouse spheroids and 56% loss in the number of myelin sheets, with partial remyelination after a week of recovery. The number of dividing cells was increased after LPC exposure and oligodendrocytes were shown to be among the dividing cells. Microglia and astrocytes were not affected and neurons were relatively spared. This suggests that LPC toxicity is specific for myelin and oligodendrocytes. LPC toxicity could be decreased using cholesterol and simvastatin, suggesting that LPC works through altering membrane composition. Thus, in different rodent species and using different read-outs, we could reproducibly show de- and remyelination in spheroid cultures after LPC exposure. This model for demyelination with potential for remyelination offers possibilities for testing novel therapies and studying mechanisms of remyelination.

Topics: 2',3'-Cyclic Nucleotide 3'-Phosphodiesterase; Animals; Brain; Cell Proliferation; Central Nervous System Agents; Cholesterol; Demyelinating Diseases; Lysophosphatidylcholines; Mice; Mice, Inbred C57BL; Models, Neurological; Myelin Basic Protein; Myelin Sheath; Nerve Tissue Proteins; Neuroglia; Neurons; Phosphoric Diester Hydrolases; Rats; Rats, Wistar; Simvastatin; Spheroids, Cellular; Time Factors; Transcription Factors

Mononuclear cell infiltrates, deposits of immunoglobulin and complement as well as demyelination and axonal damage are neuropathological hallmarks of Multiple Sclerosis (MS) lesions. An involvement of antibodies is further suggested by the presence of oligoclonal immunoglobulins in the cerebrospinal fluid of almost all MS patients. However, which mechanisms are most relevant for de- and remyelination and axonal loss in MS lesions is poorly understood. To characterize the regenerative abilities of demyelinated CNS tissue, we utilized murine organotypic cerebellar slice cultures expressing GFP in oligodendrocytes. The addition of a demyelinating monoclonal antibody specific for myelin oligodendrocyte glycoprotein and complement induced complete myelin destruction and oligodendrocyte loss, as demonstrated by confocal live imaging and staining for different myelin proteins. After removal of antibodies and complement we visualized the stages of remyelination, presumably originating from proliferating oligodendrocyte precursor cells and guided by morphologically intact appearing axons. Allowing for the detailed live imaging of de- and remyelination in an ex vivo situation closely resembling the three dimensional cytoarchitecture of the CNS, we provide a useful experimental system for the evaluation of new therapeutic strategies to enhance remyelination and repair in MS.

Topics: Animals; Animals, Newborn; Bromodeoxyuridine; Cerebral Cortex; Complement System Proteins; Demyelinating Diseases; Enzyme-Linked Immunosorbent Assay; Green Fluorescent Proteins; Immunoglobulins; Mice; Mice, Transgenic; Microscopy, Confocal; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin Sheath; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Oligodendroglia; Organ Culture Techniques; Specific Pathogen-Free Organisms; Time Factors

C57BL/6 mice were given 0.2% cuprizone (CPZ) for 2 to 6 weeks while controls ate the same diet without CPZ. At various time points the animals were subjected to behavioral tests and their brains were analyzed. Mice exposed to CPZ for 2 and 3 weeks displayed more climbing behavior and lower prepulse inhibition, suggesting an increase in central nervous system activity and impaired sensorimotor gating. In addition, they showed lower activities of monoamine oxidase and dopamine beta hydroxylase in the hippocampus and prefrontal cortex, and had higher dopamine but lower norepinephrine levels in the prefrontal cortex. Mice exposed to CPZ for 4 to 6 weeks had less social interaction, which is an animal correlate of social withdrawal of patients with schizophrenia. Also, these CPZ-exposed mice showed evident brain demyelination, myelin break down, and loss of oligodendrocytes. At all time points the CPZ-exposed mice spent more time in the open arms of an elevated plus maze and exhibited spatial working memory impairment. These data are in line with evidence from human studies suggesting a putative role of white matter abnormality in the pathophysiology of schizophrenia.

Topics: Analysis of Variance; Animals; Behavior, Animal; Brain; Brain Chemistry; Chromatography, High Pressure Liquid; Cuprizone; Demyelinating Diseases; Dopamine beta-Hydroxylase; Electrochemistry; Exploratory Behavior; Interpersonal Relations; Male; Maze Learning; Mice; Mice, Inbred C57BL; Monoamine Oxidase; Monoamine Oxidase Inhibitors; Myelin Basic Protein; Neural Inhibition; Time Factors

There are several diverse rat models of experimental autoimmune encephalomyelitis (EAE) that can be used to investigate the pathogenesis and regulation of autoimmunity against CNS myelin. The disease course of these models ranges from an acute monophasic disease with limited demyelination to a chronic relapsing or chronic progressive course marked by severe demyelination. These models enable the study of encephalitogenic T cells and demyelinating antibody specific for major neuroantigens such as myelin basic protein (MBP), myelin oligodendrocyte glycoprotein (MOG), or proteolipid protein (PLP), among other important CNS autoantigens. Overall, this unit provides an overview of common methods for induction of active and passive EAE, assessment and analysis of clinical disease, preparation and purification of myelin basic protein, and derivation of neuroantigen-specific rat T cell lines. This unit also provides a brief discussion of the basic characteristics of these models.

Topics: Adoptive Transfer; Animals; Autoantigens; Autoimmunity; Biomedical Research; Cell Extracts; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Guinea Pigs; Immunization; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Rats; Rats, Inbred Strains; Spinal Cord; T-Cell Antigen Receptor Specificity; T-Lymphocytes

Brain inflammation plays a central role in numerous brain pathologies, including multiple sclerosis (MS). Microglial cells and astrocytes are the effector cells of neuroinflammation. They can be activated also by agents such as interferon-gamma (IFN-gamma) and lipopolysaccharide (LPS). Peroxisome proliferator-associated receptor (PPAR) pathways are involved in the control of the inflammatory processes, and PPAR-beta seems to play an important role in the regulation of central inflammation. In addition, PPAR-beta agonists were shown to have trophic effects on oligodendrocytes in vitro, and to confer partial protection in experimental autoimmune encephalomyelitis (EAE), an animal model of MS. In the present work, a three-dimensional brain cell culture system was used as in vitro model to study antibody-induced demyelination and inflammatory responses. GW 501516, a specific PPAR-beta agonist, was examined for its capacity to protect from antibody-mediated demyelination and to prevent inflammatory responses induced by IFN-gamma and LPS.. Aggregating brain cells cultures were prepared from embryonal rat brain, and used to study the inflammatory responses triggered by IFN-gamma and LPS and by antibody-mediated demyelination induced by antibodies directed against myelin-oligodendrocyte glycoprotein (MOG). The effects of GW 501516 on cellular responses were characterized by the quantification of the mRNA expression of tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), inducible NO synthase (i-NOS), PPAR-beta, PPAR-gamma, glial fibrillary acidic protein (GFAP), myelin basic protein (MBP), and high molecular weight neurofilament protein (NF-H). GFAP expression was also examined by immunocytochemistry, and microglial cells were visualized by isolectin B4 (IB4) and ED1 labeling.. GW 501516 decreased the IFN-gamma-induced up-regulation of TNF-alpha and iNOS in accord with the proposed anti-inflammatory effects of this PPAR-beta agonist. However, it increased IL-6 m-RNA expression. In demyelinating cultures, reactivity of both microglial cells and astrocytes was observed, while the expression of the inflammatory cytokines and iNOS remained unaffected. Furthermore, GW 501516 did not protect against the demyelination-induced changes in gene expression.. Although GW 501516 showed anti-inflammatory activity, it did not protect against antibody-mediated demyelination. This suggests that the protective effects of PPAR-beta agonists observed in vivo can be attributed to their anti-inflammatory properties rather than to a direct protective or trophic effect on oligodendrocytes.

Topics: Animals; Anti-Inflammatory Agents; Antibodies; Astrocytes; Brain; Cells, Cultured; Demyelinating Diseases; Encephalitis; Encephalomyelitis, Autoimmune, Experimental; Glial Fibrillary Acidic Protein; Interferon-gamma; Interleukin-6; Lipopolysaccharides; Microglia; Myelin Basic Protein; Neurofilament Proteins; Nitric Oxide Synthase Type II; PPAR gamma; PPAR-beta; Rats; Thiazoles; Tumor Necrosis Factor-alpha

Gray matter (GM) lesions are recognized as important components of the pathology of multiple sclerosis (MS), and involvement of the deep gray matter (DGM) is suggested by magnetic resonance imaging. The aims of this study were to determine the frequency and distribution of lesions and characterize the inflammatory and neurodegenerative changes in DGM of MS patients. Histochemistry, immunohistochemistry, and morphometry were performed on whole coronal sections of 14 MS and 12 control (6 normal, 6 from amyotrophic lateral sclerosis patients) brains. Demyelinating lesions were frequent in MS DGM; most often in the thalamus and caudate, but they were also seen in the putamen, pallidum, claustrum, amygdala, hypothalamus, and substantia nigra. Most DGM lesions involved both GM and white matter. Inflammation in active DGM lesions was similar to that in lesions only in white matter but was less intense, and there was a preponderance of activated microglia, scarce myelin-laden macrophages, and a lesser extent of axonal damage. Neuronal loss was observed both in DGM lesions and nondemyelinated DGM with neuron atrophy in nondemyelinated DGM. In conclusion, demyelination and neurodegenerative changes are common in MS DGM and may contribute to clinical impairment. Inflammation in DGM lesions is intermediate between the destructive inflammation of white matter lesions and the minimal inflammation of cortical lesions. We hypothesize that alterations of glutamate reuptake mechanisms may contribute to these differences.

Topics: Adult; Aged; Amyotrophic Lateral Sclerosis; Antigens, CD; Brain; Demyelinating Diseases; Female; Fibrinogen; HLA-DR Antigens; Humans; Inflammation; Male; Middle Aged; Multiple Sclerosis; Myelin Basic Protein; Nerve Tissue Proteins; Neurodegenerative Diseases; Neuroglia; Neurons; Staining and Labeling

Astrocytes are coupled via gap junctions (GJs) comprising connexin 43 (Cx43) (Gja1) and Cx30 (Gjb6), which facilitate intercellular exchange of ions. Astrocyte connexins also form heterotypic GJs with oligodendrocytic somata and lamellae. Loss of oligodendrocyte gap junctions results in oligodendrocyte and myelin pathology. However, whether loss of astrocyte GJs affects oligodendrocytes and myelin is not known. To address this question, mice with astrocyte-targeted deletion of Cx43 and global loss of Cx30 [double knock-out (dKO)] were studied using Western blotting, immunohistochemistry, electron microscopy, and functional assays. Commencing around postnatal day 23 and persisting into old age, we found widespread pathology of white matter tracts comprising vacuolated oligodendrocytes and intramyelinic edema. In contrast, gray matter pathology was restricted to the CA1 region of the hippocampus, and consisted of edematous astrocytes. No differences were observed in synaptic density or total NeuN(+) cells in the hippocampus, or olig2(+) cells in the corpus callosum. However, in dKO mice, fewer CC1-positive mature oligodendrocytes were detected, and Western blotting indicated reduced myelin basic protein. Pathology was not noted in mice expressing a single allele of either Cx43 or Cx30. When compared with single connexin knock-outs, dKO mice were impaired in sensorimotor (rotarod, balance beam assays) and spatial memory tasks (object recognition assays). We conclude that loss of astrocytic GJs can result in white matter pathology that has functional consequences.

Topics: Animals; Animals, Newborn; Astrocytes; Basic Helix-Loop-Helix Transcription Factors; Calcium-Binding Proteins; Cell Proliferation; Connexin 30; Connexin 43; Connexins; Demyelinating Diseases; Disease Models, Animal; DNA-Binding Proteins; Gap Junctions; Glial Fibrillary Acidic Protein; Hippocampus; In Situ Nick-End Labeling; Memory Disorders; Mice; Mice, Inbred C57BL; Mice, Knockout; Microfilament Proteins; Microscopy, Electron; Myelin Basic Protein; Myelin Sheath; Nerve Tissue Proteins; Oligodendrocyte Transcription Factor 2; Phenotype

The expression of sodium channels (NaCh(s)) change after inflammatory and nerve lesions, and this change has been implicated in the generation of pain states. Here we examine NaCh expression within nerve fibers from normal and painful extracted human teeth with special emphasis on their localization within large accumulations, like those seen at nodes of Ranvier. Pulpal tissue sections from normal wisdom teeth and from teeth with large carious lesions associated with severe and spontaneous pain were double-stained with pan-specific NaCh antibody and caspr (paranodal protein used to visualize nodes of Ranvier) antibody, while additional sections were triple-stained with NaCh, caspr and myelin basic protein (MBP) antibodies. Z-series of images were obtained with the confocal microscope and evaluated with NIH ImageJ software to quantify the density and size of NaCh accumulations, and to characterize NaCh localization at caspr-identified typical and atypical nodal sites. Although the results showed variability in the overall density and size of NaCh accumulations in painful samples, a common finding included the remodeling of NaChs at atypical nodal sites. This remodeling of NaChs included prominent NaCh expression within nerve regions that showed a selective loss of MBP staining in a pattern consistent with a demyelinating process.. This study identifies the remodeling of NaChs at demyelinated sites within the painful human dental pulp and suggests that the contribution of NaChs to spontaneous pulpal pain generation may be dependant not only on total NaCh density but may also be related to NaCh expression at atypical nodal sites.

Topics: Adolescent; Adult; Axons; Cell Adhesion Molecules, Neuronal; Contactins; Demyelinating Diseases; Dental Pulp; Female; Humans; Immunohistochemistry; Male; Microscopy, Confocal; Molar, Third; Myelin Basic Protein; Myelin Sheath; Nerve Fibers; Pain; Ranvier's Nodes; Sodium Channels; Young Adult

Multiple sclerosis (MS) is a T-cell mediated autoimmune disease of the CNS, possessing both immune and neurodegenerative events that lead to disability. Adoptive transfer (AT) of myelin basic protein (MBP)-specific T cells into naïve female SJL/J mice results in a relapsing-remitting (RR) form of experimental autoimmune encephalomyelitis (EAE). Blocking the mechanisms by which MBP-specific T cells are activated before AT may help characterize the immune arm of MS and offer novel targets for therapy. One such target is calpain, which is involved in activation of T cells, migration of immune cells into the CNS, degradation of axonal and myelin proteins, and neuronal apoptosis. Thus, the hypothesis that inhibiting calpain in MBP-specific T cells would diminish their encephalitogenicity in RR-EAE mice was tested. Incubating MBP-specific T cells with the calpain inhibitor SJA6017 before AT markedly suppressed the ability of these T cells to induce clinical symptoms of RR-EAE. These reductions correlated with decreases in demyelination, inflammation, axonal damage, and loss of oligodendrocytes and neurons. Also, calpain : calpastatin ratio, production of truncated Bid, and Bax : Bcl-2 ratio, and activities of calpain and caspases, and internucleosomal DNA fragmentation were attenuated. Thus, these data suggest calpain as a promising target for treating EAE and MS.

Topics: Animals; Axons; Boron Compounds; Calcium-Binding Proteins; Calpain; Cell Survival; Demyelinating Diseases; Dipeptides; Disease Models, Animal; DNA Fragmentation; Dose-Response Relationship, Drug; Encephalomyelitis, Autoimmune, Experimental; Female; In Situ Nick-End Labeling; L-Lactate Dehydrogenase; Mice; Myelin Basic Protein; Statistics, Nonparametric; T-Lymphocytes; Time Factors

Although periventricular white matter injury is a leading cause of major neurologic disability in premature infants, the relationship between myelination deficiency and long-term cognitive dysfunction is not well understood. The purpose of this study was to investigate oligodendrocytes myelination and long-term spatial cognitive function in rats with perinatal hypoxia-ischemia (HI). Postnatal day 3 (P3) rats were subjected to right carotid artery ligation followed by 2.5 h of hypoxia (6% oxygen). Brain injury during the early and late phases was evaluated by immunostaining at P6 (72 h after the injury) and P47. Spatial cognitive function was evaluated at P42 using the Morris Water Maze test followed by histologic evaluation. HI caused an increase in pre-oligodendrocytes, astrocytes, and microglia in the ipsilateral white matter 72 h after the insult compared to contralateral regions and sham-operated controls (both p<0.05). There were significant decreases in myelin basic protein (MBP)and 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase)-labeled oligodendrocytes with glial fibrillary acidic protein (GFAP)-labeled glial scarring in the ipsilateral periventricular white matter at P47 compared to contralateral regions and sham-operated controls (all p<0.05). The rats with HI had spatial learning deficits in navigation trials (longer escape latency and swimming distance) and memory dysfunction in probe trials (fewer number of platform crossings and percentage of time in the target quadrant) compared with sham-operated controls (p<0.05). In this neonatal rat model of HI, myelination deficiency induced by activated astrocytes and microglia during the early phase with subsequent glial scarring was associated with long-term spatial learning and memory dysfunction.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Analysis of Variance; Animals; Animals, Newborn; Antigens, CD; Antigens, Differentiation, Myelomonocytic; CA1 Region, Hippocampal; Cell Count; Cognition Disorders; Demyelinating Diseases; Exploratory Behavior; Female; Gliosis; Hypoxia-Ischemia, Brain; Immunohistochemistry; Male; Maze Learning; Myelin Basic Protein; Nerve Fibers, Myelinated; Neuroglia; Rats; Space Perception; Time Factors

Cortical myelin can be severely affected in patients with demyelinating disorders of the central nervous system. However, the functional implication of cortical demyelination remains elusive. In this study, we investigated whether cortical myelin influences cortical spreading depression (CSD).. CSD measurements were performed in rodent models of toxic and autoimmune induced cortical demyelination, in neuregulin-1 type I transgenic mice displaying cortical hypermyelination, and in glial fibrillary acidic protein-transgenic mice exhibiting pronounced astrogliosis.. Cortical demyelination, but not astrogliosis or inflammation per se, was associated with accelerated CSD. In contrast, hypermyelinated neuregulin-1 type I transgenic mice displayed a decelerated CSD propagation.. Cortical myelin may be crucially involved in the stabilization and buffering of extracellular ion content that is decisive for CSD propagation velocity and cortical excitability, respectively. Our data thus indicate that cortical involvement in human demyelinating diseases may lead to relevant alterations of cortical function.

Topics: Animals; Astrocytes; Cerebral Cortex; Cortical Spreading Depression; Cuprizone; Demyelinating Diseases; Electroencephalography; Encephalomyelitis, Autoimmune, Experimental; Female; Functional Laterality; Glial Fibrillary Acidic Protein; Gliosis; Humans; Male; Mice; Mice, Inbred BALB C; Mice, Transgenic; Multiple Sclerosis; Myelin Basic Protein; Myelin Sheath; Neuregulin-1; Rats; Rats, Inbred Lew

In the present work we analyzed the capacity of thyroid hormones (THs) to improve remyelination using a rat model of cuprizone-induced demyelination previously described in our laboratories. Twenty one days old Wistar rats were fed a diet containing 0.6% cuprizone for two weeks to induce demyelination. After cuprizone withdrawal, rats were injected with triiodothyronine (T3). Histological studies carried out in these animals revealed that remyelination in the corpus callosum (CC) of T3-treated rats improved markedly when compared to saline treated animals. The cellular events occurring in the CC and in the subventricular zone (SVZ) during the first week of remyelination were analyzed using specific oligodendroglial cell (OLGc) markers. In the CC of saline treated demyelinated animals, mature OLGcs decreased and oligodendroglial precursor cells (OPCs) increased after one week of spontaneous remyelination. Furthermore, the SVZ of these animals showed an increase in early progenitor cell numbers, dispersion of OPCs and inhibition of Olig and Shh expression compared to non-demyelinated animals. The changes triggered by demyelination were reverted after T3 administration, suggesting that THs could be regulating the emergence of remyelinating oligodendrocytes from the pool of proliferating cells residing in the SVZ. Our results also suggest that THs receptor beta mediates T3 effects on remyelination. These results support a potential role for THs in the remyelination process that could be used to develop new therapeutic approaches for demyelinating diseases.

Topics: Animals; Animals, Newborn; Antigens; Blood Coagulation Factors; Cell Differentiation; Cuprizone; Demyelinating Diseases; Gangliosides; Glial Fibrillary Acidic Protein; Hedgehog Proteins; Myelin Basic Protein; O Antigens; Oligodendroglia; Proteoglycans; Rats; Rats, Wistar; Receptor-Interacting Protein Serine-Threonine Kinases; Receptors, Cell Surface; Stem Cells; Thyroid Hormones; Time Factors; Triiodothyronine

The aim of this study was to assess quantitatively structural changes in myelin content occurring during demyelination and remyelination by magnetization transfer imaging (MTI).. In a reversible model of demyelination with no axonal loss, mice intoxicated by cuprizone were studied by MTI in vivo at 9.4 T. MRI data were compared to histopathological examinations.. Data revealed that the magnetization transfer ratio (MTR) decreased significantly during demyelination and increased during remyelination with strong correlation to the myelin content (r=0.79, P=0.01).. This study demonstrated that MTR is a sensitive and reproducible quantitative marker to assess myelin loss and repair. This may lead to in vivo monitoring of therapeutic strategies promoting remyelination.

Topics: Animals; Brain; Cuprizone; Demyelinating Diseases; Glial Fibrillary Acidic Protein; Magnetic Resonance Imaging; Magnetics; Mice; Mice, Inbred C57BL; Myelin Basic Protein; Myelin Sheath; Nerve Tissue Proteins

Experimental studies of intracerebral hemorrhage (ICH) have focused on neuron death, with little or no information on axonal and myelin damage outside the hematoma. Because development of effective therapies will require an understanding of white matter injury, we examined white matter injury and its spatial and temporal relationship with microglial/macrophage activation in a collagenase model of rat striatal ICH. The hematoma and parenchyma surrounding the hematoma were assessed in young and aged animals at 6 h, 1, 3 and 28 days after ICH onset. Demyelination occurred inside and at the edge of the hematoma; regions where we have shown substantial neuron death. In contrast, there was axonal damage without demyelination at the edge of the hematoma, and by 3 days this damage had spread to the surrounding parenchyma, a region where we have shown there is no neuron death. Because the axonal damage preceded infiltration of activated microglia into the white matter tracts (seen at 3 days), our results support the hypothesis that these cells respond to, rather than perpetrate the damage. Importantly, axonal damage was worse in aged animals, which provides a plausible explanation for the poorer functional recovery of older animals after ICH, despite a similar loss of grey matter. Our findings support strategies that target white matter injury to reduce neurological impairment after ICH.

Topics: Age Factors; Aging; Amyloid beta-Protein Precursor; Animals; Brain Injuries; Cerebral Hemorrhage; Demyelinating Diseases; Disease Models, Animal; Male; Microglia; Myelin Basic Protein; Nerve Fibers, Myelinated; Neuritis; Rats; Rats, Sprague-Dawley; Stroke

A dynamic equilibrium between proliferation and programmed cell death (PCD) of auto-reactive T lymphocytes plays a pivotal role in the prevention of autoimmune diseases. We analyzed T lymphocytes myelin basic protein (MBP)-specific PCD and proliferation in demyelinating diseases. Results showed that MBP-specific PCD was significantly decreased in CD4+ and CD8+ T lymphocytes of progressive multifocal leukoencephalopathy (PML), not determined leukoencephalopathy (NDLE), and acute MS (AMS) patients compared to patients with stable MS (SMS) and healthy controls. MBP-specific proliferation/PCD rates were high in CD4+ T lymphocytes of PML, NDLE, and AMS patients, and in CD8+ T cells of PML and AMS individuals alone. Alterations of the balance between MBP-specific proliferation and PCD are present in demyelinating diseases and could play a major role in the pathogenesis of these diseases.

Topics: Adult; Amino Acid Sequence; Apoptosis; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Cell Proliferation; Cytokines; Demyelinating Diseases; DNA, Viral; Female; Flow Cytometry; Humans; JC Virus; Leukocytes, Mononuclear; Lymphocyte Activation; Male; Middle Aged; Molecular Sequence Data; Myelin Basic Protein; Peptide Fragments; Polymerase Chain Reaction; Young Adult

Transgenic and disease model mice have been used to investigate the molecular mechanisms of demyelinating diseases. However, less attention has been given to elucidating changes in nerve conduction in these mice. We established an experimental system to measure the response latency of cortical neurons and examined changes in nerve conduction in cuprizone-induced demyelinating mice and in myelin basic protein-deficient shiverer mice. Stimulating and recording electrodes were placed in the right and left sensori-motor cortices, respectively. Electrical stimulation of the right cortex evoked antidromic responses in left cortical neurons with a latency of 9.38 +/- 0.31 ms (n = 107; mean +/- SEM). While response latency was longer in mice at 7 days and 4 weeks of cuprizone treatment (12.35 +/- 0.35 ms, n = 102; 11.72 +/- 0.29 ms, n = 103, respectively), response latency at 7 days and 4 weeks after removal of cuprizone was partially restored (10.72 +/- 0.45 ms, n = 106; 10.27 +/- 0.34 ms, n = 107, respectively). Likewise, electron microscopy showed cuprizone-induced demyelination in the corpus callosum and nearly complete remyelination after cuprizone removal. We also examined whether the myelin abnormalities in shiverer mice affected their response latencies. But there were no significant differences in response latencies in shiverer (9.83 +/- 0.24 ms, n = 103) and wild-type (9.33 +/- 0.22 ms, n = 112) mice. The results of these electrophysiological assessments imply that different demyelinating mechanisms, differentially affecting axon conduction, are present in the cuprizone-treated and shiverer mice, and may provide new insights to understanding the pathophysiology of demyelination in animal models in the CNS.

Topics: Animals; Axons; Central Nervous System; Chelating Agents; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Electric Stimulation; Mice; Mice, Inbred BALB C; Mice, Knockout; Mice, Neurologic Mutants; Motor Cortex; Myelin Basic Protein; Nerve Fibers, Myelinated; Neural Conduction; Neural Pathways; Reaction Time; Wallerian Degeneration

The aim of this study was to determine the relation between changes in myelin basic protein (MBP) levels during the acute and subacute phases of central nervous system injury following whole-brain radiation and delayed demyelination in the radiation-injured brain tissue.. Adult Sprague-Dawley rats were treated with single fractions of 2, 10, or 30 Gy of whole-brain radiation. The authors measured MBP gene expression and protein levels in the brain tissue by using reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay at 1 week and 1-3 months following irradiation to monitor myelin changes in the brain. Demyelination was determined with Luxol fast blue myelin staining and routine histopathological and electron microscopy examination of injured brain tissue. The changes in MBP levels in the different animal groups at specific time points were correlated with demyelination in corresponding dose groups.. At 1 month after applying the 10 and 30 Gy of radiation, MBP mRNA expression showed a transient but significant decrease, followed by recovery to baseline levels at 3 months after treatment. The MBP levels were decreased by only 70-75% at 1 month after 10 and 30 Gy of radiation. At 2-3 months after applying the higher dose of 30 Gy, however, the MBP levels continued to decline, and typical demyelination changes were observed with myelin staining and ultrastructural examination.. The authors' results suggest that the early radiation-induced MBP changes between 1 and 3 months after single treatments of 10 and 30 Gy of radiation to the whole brain are indicative of permanent injury shown as demyelination of irradiated brain tissue.

Topics: Animals; Brain; Cranial Irradiation; Demyelinating Diseases; Dose-Response Relationship, Radiation; Models, Animal; Myelin Basic Protein; Nerve Tissue Proteins; Radiation Injuries, Experimental; Rats; Rats, Sprague-Dawley; RNA, Messenger; Transcription Factors

Following acute and chronic neurodegenerative disorders, a cascade of pathological events including inflammatory response, excitotoxicity and oxidative stress induces secondary tissue loss in both gray and white matter. Axonal damage and demyelination are important components of the white matter demise during these diseases. In spite of this, a few studies have addressed the patterns of inflammatory response, axonal damage and demyelination following focal ischemic damage to the central nervous system (CNS). In the present study, we describe the patterns of inflammatory response, axonal damage and myelin impairment following microinjections of 10 pmol of endothelin-1 into the rat striatum. Animals were perfused at 1 day, 3 days and 7 days after injection. 20 mum sections were stained by hematoxylin and immunolabeled for neutrophils (anti-MBS-1), activated macrophages/microglia (anti-ED1), damaged axons (anti-betaAPP) and myelin (anti-MBP). The evolution of acute inflammation was quantitatively assessed by cell counts in different survival times. There was recruitment of both neutrophils and macrophages to the damaged striatal parenchyma with maximum recruitment at 1 day and 7 days, respectively. Progressive myelin impairment in the striatal white matter tracts has been observed mainly at later survival times. beta-APP+ endbulbs were not present in all evaluated time points. These results suggest that progress myelin impairment in the absence of damage to axonal cylinder is a feature of white matter pathology following endothelin-1-induced focal striatal ischemia.

Topics: Amyloid beta-Peptides; Animals; Axons; Biomarkers; Brain Ischemia; Cerebral Arteries; Chemotaxis, Leukocyte; Corpus Striatum; Demyelinating Diseases; Disease Progression; Encephalitis; Endothelin-1; Male; Microcirculation; Microglia; Microinjections; Myelin Basic Protein; Nerve Fibers, Myelinated; Neutrophils; Rats; Rats, Wistar

Lysophosphatidic acid (LPA) causes neuropathic pain with demyelination in sensory fibers. In dorsal root (DR) ex vivo culture, the addition of 0.1 microM LPA caused a characteristic demyelination at 24h in scanning and transmission electron microscopy analyses. Moreover, direct contact between C-fibers due to loss of partition by Schwann cell in Remak bundles was observed. LPA-induced demyelination of DR was concentration-dependent in the range between 0.01 and 1M, and was abolished by BoNT/C3 and Y-27632, a RhoA and Rho kinase inhibitor, respectively. The demyelination was equivalent between the preparations with and without dorsal root ganglion. LPA also caused a down-regulation of myelin proteins, such as myelin basic protein (MBP) and myelin protein zero (MPZ) to approximately 70% of control. All these findings suggest that the demyelination observed in the neuropathic pain due to nerve injury occurs through a direct action of LPA on Schwann cells.

Topics: Animals; Blotting, Western; Cells, Cultured; Demyelinating Diseases; Hyperalgesia; Lysophospholipids; Male; Mice; Mice, Knockout; Microscopy, Electron, Scanning; Microscopy, Electron, Transmission; Myelin Basic Protein; Myelin P0 Protein; Myelin Sheath; Nerve Fibers; Pain; Receptors, Lysophosphatidic Acid; Schwann Cells; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Spinal Nerve Roots; Tolonium Chloride

In attempts to produce mature oligodendrocytes from human embryonic stem (huES) cells, we searched conditions inducing transcription factors Olig1/2, as well as Nkx2.2 and Sox10, which are needed for maturation. This was obtained by retinoic acid treatment followed by noggin, an antagonist of bone morphogenetic proteins (BMPs). We found that retinoic acid induces BMPs in huES cells. Addition of noggin at a specific step was essential to form numerous mature oligodendrocytes with ramified branches and producing myelin basic protein (MBP). We describe a procedure converting huES cells into enriched populations of oligodendrocyte precursors that can be expanded and passaged repeatedly and subsequently differentiated into mature cells. Transplantation of such precursors showed that pretreatment by noggin markedly stimulates their capacity to myelinate in the brain of MBP-deficient shiverer mice in organotypic cultures and in living animals. Arrays of numerous long MBP+ fibers were generated over extended areas in the brain, with evidence of cell migration after transplantation and with formation of compact myelin sheaths.

Topics: Animals; Animals, Newborn; Carrier Proteins; Cell Differentiation; Cell Line; Demyelinating Diseases; Drug Interactions; Embryonic Stem Cells; Fetus; Gene Expression Profiling; Gene Expression Regulation; Homeobox Protein Nkx-2.2; Homeodomain Proteins; Humans; Intercellular Signaling Peptides and Proteins; Mice, Neurologic Mutants; Microscopy, Electron, Transmission; Myelin Basic Protein; Nerve Tissue Proteins; Nuclear Proteins; Oligodendroglia; Organogenesis; Stem Cell Transplantation; Transcription Factors; Tretinoin

Histopathologic studies have shown that subpial cortical demyelination is extensive in chronic multiple sclerosis (MS).. To study whether subpial cortical demyelination in MS is associated with focal or diffuse white matter (WM) pathologic features on magnetic resonance imaging (MR imaging).. Comparison of postmortem MR imaging findings with histopathologic findings.. Brain donations from a general community.. Three patients with MS with extensive cortical demyelination and 3 patients with minor cortical demyelination were selected from an MS autopsy data set. The postmortem MR imaging and histopathologic data of the patients were compared.. Two observers blinded to the results of each other assessed the presence, extent, and distribution of focal and diffuse pathologic changes in WM by MR imaging and by histopathology.. Extensive subpial demyelination was not associated with a significant increase in the area of focal and diffuse WM pathologic changes as assessed by Luxol fast blue histochemistry or by MR imaging or with the presence or extent of juxtacortical abnormalities on MR imaging.. The lack of association of MS gray matter demyelination with diffuse or focal WM changes indicates that gray matter demyelination in MS occurs largely independent of WM pathologic changes. The extent or distribution of WM abnormalities cannot be used to identify extensive cortical demyelination in the clinical setting.

Topics: Aged; Aged, 80 and over; Autopsy; Cerebral Cortex; Demyelinating Diseases; Female; Humans; Immunohistochemistry; Magnetic Resonance Imaging; Male; Middle Aged; Multiple Sclerosis; Myelin Basic Protein; Postmortem Changes; Statistics, Nonparametric

Dichloroacetate (DCA) is an investigational drug for genetic mitochondrial diseases whose use has been mitigated by reversible peripheral neuropathy. We investigated the mechanism of DCA neurotoxicity using cultured rat Schwann cells (SCs) and dorsal root ganglia (DRG) neurons. Myelinating SC-DRG neuron co-cultures, isolated SCs and DRG neurons were exposed to 1-20 mm DCA for up to 12 days. In myelinating co-cultures, DCA caused a dose- and exposure-dependent decrease of myelination, as determined by immunolabeling and immunoblotting for myelin basic protein (MBP), protein zero (P0), myelin-associated glycoprotein (MAG) and peripheral myelin protein 22 (PMP22). Partial recovery of myelination occurred following a 10-day washout of DCA. DCA did not affect the steady-state levels of intermediate filament proteins, but promoted the formation of anti-neurofilament antibody reactive whirls. In isolated SC cultures, DCA decreased the expression of P0 and PMP22, while it increased the levels of p75(NTR) (neurotrophin receptor), as compared with non-DCA-treated samples. DCA had modest adverse effects on neuronal and glial cell vitality, as determined by the release of lactate dehydrogenase. These results demonstrate that DCA induces a reversible inhibition of myelin-related proteins that may account, at least in part, for its clinical peripheral neuropathic effects.

Topics: Animals; Animals, Newborn; Blotting, Western; Cell Death; Cells, Cultured; Coculture Techniques; Demyelinating Diseases; Dichloroacetic Acid; Dose-Response Relationship, Drug; Ganglia, Spinal; Gene Expression Regulation; Immunohistochemistry; Myelin Basic Protein; Myelin P0 Protein; Myelin Proteins; Myelin-Associated Glycoprotein; Neurons; Rats; Schwann Cells; Sciatic Nerve

Our groups have reported that tumor necrosis factor-alpha (TNF-alpha) causes myelin damage and apoptosis of oligodendrocytes and their precursors in vitro and in vivo. We also have reported that insulin-like growth factor-I (IGF-I) can protect cultured oligodendrocytes and their precursors from TNF-alpha-induced damage. In this study, we investigated whether IGF-I can protect oligodendrocytes and myelination from TNF-alpha-induced damage in vivo by cross-breeding TNF-alpha transgenic (Tg) mice with IGF-I Tg mice that overexpress IGF-I exclusively in brain. At 8 weeks of age, compared with those of wild-type (WT) mice, the brain weights of TNF-alpha Tg mice were decreased by approximately 20%, and those of IGF-I Tg mice were increased by approximately 20%. The brain weights of mice that carry both TNF-alpha and IGF-I transgenes (TNF-alpha/IGF-I Tg mice) did not differ from those of WT mice. As judged by histochemical staining and immunostaining, myelin content in the cerebellum of TNF-alpha/IGF-I Tg mice was similar to that in WT mice and much more than that in TNF-alpha Tg mice. Consistently, Western immunoblot analysis showed that myelin basic protein (MBP) abundance in the cerebellum of TNF-alpha/IGF-I Tg mice was double that in TNF-alpha Tg mice. In comparison with WT mice, the number of oligodendrocytes was decreased by approximately 36% in TNF-alpha Tg mice, whereas it was increased in IGF-I Tg mice by approximately 40%. Oligodendrocyte number in TNF-alpha/IGF-I Tg mice was almost twice that in TNF-alpha Tg mice. Furthermore, IGF-I overexpression significantly reduced TNF-alpha-induced increases in apoptotic cell number, active caspase-3 abundance, and degradaion of MBP. Our results indicate that IGF-I is capable of protecting myelin and oligodendrocytes from TNF-alpha-induced damage in vivo.

Topics: Animals; Antigens; Caspase 3; Cell Count; Cerebellum; Demyelinating Diseases; Female; Gene Expression Regulation; Glutathione S-Transferase pi; Immunohistochemistry; In Situ Nick-End Labeling; Insulin-Like Growth Factor I; Male; Mice; Mice, Transgenic; Myelin Basic Protein; Organ Size; Proteoglycans; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tumor Necrosis Factor-alpha

Thyroid hormone (T3) deficiency impairs the development of the CNS, particularly myelination. We have previously described an increase in the frequency of morphological abnormalities in the central myelin sheath in a hypothyroidism model, which reinforced the hypothesis of a role for T3 in myelin compaction. However, there are no data concerning the cellular distribution of myelin proteins in hypothyroid animals. In the present work, we describe the distribution of 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase), myelin basic protein (MBP) and proteolipid protein (PLP) throughout the central myelin sheath of a hypothyroidism model. We used euthyroid and hypothyroid adult rats at 90 days of age. In order to induce hypothyroid status, animals received 0.02% methimazol from the 19th gestation day onwards. After perfusion with a fixative mixture, small pieces of corpus callosum were obtained, dehydrated and embedded in LR White resin. Ultrathin sections were immunoreacted, using specific antibodies revealed by a secondary antibody coupled to colloidal gold particles of 10nm. Gold particle density per region of myelin sheath for each one of these proteins was obtained. In normal animals, CNPase, PLP and MBP were identified in sites that had already been described in previous studies. In hypothyroid animals, CNPase was identified in the region corresponding to compact lamellae, which normally does not contain this protein, while, in this same region, PLP and MBP immunolabeling were decreased. These results suggest that thyroid hormone deficiency impairs the distribution of the major oligodendrocyte/myelin markers. This effect may justify the reduction in myelin sheath compaction previously demonstrated in a similar model of hypothyroidism.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Antithyroid Agents; Biomarkers; Cell Membrane; Corpus Callosum; Demyelinating Diseases; Hypothyroidism; Methimazole; Microscopy, Immunoelectron; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin Sheath; Oligodendroglia; Rats; Rats, Wistar

Disruption of myelin causes severe neurological diseases. An understanding of the mechanisms that control myelination and remyelination is needed to develop therapeutic strategies for demyelinating diseases such as multiple sclerosis (MS). Our previous finding indicating the critical involvement of the gamma chain of immunogloblin Fc receptors (FcRgamma) and Fyn signaling in oligodendrocyte differentiaion and myelination demands a fundamental revision of the strategies used for MS therapy, because antigen-antibody complexes in MS patients may induce the direct dysregulation of myelination process as well as the inflammatory destruction of myelin sheath. Here we show that the FcRgamma/Fyn signaling cascade is critically involved in cuprizone-induced demyelination/remyelination, with no lymphocytic response. The levels of phosphorylated myelin basic proteins (p-MBPs), especially the 21.5-kDa isoform, but not the levels of total MBPs, decreased markedly during demyelination induced by aging, cuprizone treatment, and double knockout of FcRgamma/Fyn genes. We also showed that the recovery from demyelination in cuprizone-treated and aged mice is achieved after administration of the herbal medicine Ninjin'yoeito, an effective therapy targeting the FcRgamma/Fyn-Rho (Rac1)-MAPK (P38 MAPK)-p-MBPs signaling cascade. These results suggest that the restoration of FcRgamma/Fyn signaling represents a new approach for the treatment of demyelinating diseases.

Topics: Aging; Animals; Anti-Inflammatory Agents; Central Nervous System; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Drugs, Chinese Herbal; Female; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Monoamine Oxidase Inhibitors; Multiple Sclerosis; Myelin Basic Protein; Myelin Sheath; Nerve Regeneration; Neuroprotective Agents; Neurotoxins; Proto-Oncogene Proteins c-fyn; Receptors, IgG; Recovery of Function; Signal Transduction

The neurotoxicant cuprizone has been used extensively to create a mouse model of demyelination. However, the effects on behavior of cuprizone treatment have not been previously reported. We have analyzed the behavioral changes of mice given a diet containing 0.2% cuprizone for 6 weeks followed by 6 weeks of recovery. Behavior was assessed using a range of tests: the functional observation battery, the open-field test and the rota-rod test. Concurrent with the start of demyelination, at 3 and 4 weeks of 0.2% cuprizone treatment, the animals exhibited an increase in central nervous system activity and an inhibited anxiogenic response to the novelty challenge test. At 5 weeks of treatment (the period of maximal demyelination) equilibrium was altered and sensorimotor reactivity was also affected. Further, rota-rod analysis demonstrated that the treated group had poorer motor co-ordination than control animals. This effect was not reversed 6 weeks after cuprizone withdrawal. The animals in the recovery period also exhibited difficulties in the rota-rod progressive learning task. Our results indicate that behavioral deficits follow the course of demyelination-remyelination induced by administration of 0.2% cuprizone, and that some of the changes persist even after 6 weeks on normal diet.

Topics: Animals; Behavior, Animal; Brain; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Immunohistochemistry; Male; Mice; Mice, Inbred C57BL; Myelin Basic Protein

Emerging evidence suggests that cell-based remyelination strategies may be a feasible therapeutic approach for CNS diseases characterized by myelin deficiency as a result of trauma, congenital anomalies, or diseases. Although experimental demyelination models targeted at the transient elimination of oligodendrocytes have suggested that transplantation-based remyelination can partially restore axonal molecular structure and function, it is not clear whether such therapeutic approaches can be used to achieve functional remyelination in models associated with long-term, irreversible myelin deficiency. In this study, we transplanted adult neural precursor cells (aNPCs) from the brain of adult transgenic mice into the spinal cords of adult Shiverer (shi/shi) mice, which lack compact CNS myelin. Six weeks after transplantation, the transplanted aNPCs expressed oligodendrocyte markers, including MBP, migrated extensively along the white matter tracts of the spinal cord, and formed compact myelin. Conventional and three-dimensional confocal and electron microscopy revealed axonal ensheathment, establishment of paranodal junctional complexes leading to de novo formation of nodes of Ranvier, and partial reconstruction of the juxtaparanodal and paranodal molecular regions of axons based on Kv1.2 and Caspr (contactin-associated protein) expression by the transplanted aNPCs. Electrophysiological recordings revealed improved axonal conduction along the transplanted segments of spinal cords. We conclude that myelination of congenitally dysmyelinated adult CNS axons by grafted aNPCs results in the formation of compact myelin, reconstruction of nodes of Ranvier, and enhanced axonal conduction. These data suggest the therapeutic potential of aNPCs to promote functionally significant myelination in CNS disorders characterized by longstanding myelin deficiency.

Topics: Animals; Axons; Cell Differentiation; Demyelinating Diseases; Disease Models, Animal; Evoked Potentials; Mice; Mice, Mutant Strains; Myelin Basic Protein; Nerve Fibers, Myelinated; Neural Conduction; Neuroglia; Neurons; Ranvier's Nodes; Spinal Cord Diseases; Stem Cell Transplantation

Chronic exposure to the copper-chelating agent, cuprizone (CPZ), is an increasingly popular model for producing demyelination. More importantly, cessation of cuprizone exposure allows for full remyelination, which represents a window of opportunity for determining the influence of environmental factors on regenerative processes. In the present study, CPZ-treated animals were assessed for functional status of systemic and central cytokine responsiveness to LPS, as well as assessment for signs of body weight changes. Exposure of male C57BL/6J mice to 5 weeks of 0.2% CPZ in the diet was optimal in producing demyelination and microglial activation, as measured by myelin basic protein, CD11b, and CD45 immunohistochemistry. Acute challenge with LPS at the end of 5 weeks CPZ treatment did not alter IL-1beta, IL-6, nor TNFalpha responses in the spleen and corpus callosum. Similarly, repeated exposure to LPS during the remyelination phase (CPZ removal) did not influence these measures to LPS. Plasma corticosterone was unaffected following acute challenge of CPZ-pretreated animals, but after repeated LPS treatment, there was a significant augmentation of the corticosterone response in CPZ-pretreated mice. Interestingly, the basal concentration of IL-1beta in the corpus callosum of CPZ treated animals was significantly increased, which was in keeping with the increase in activated microglial cells. In conclusion, the cuprizone model of demyelination and remyelination does not appear to influence the systemic nor central IL-1, IL-6, and TNF responses to acute nor repeated LPS. This opens up the possibility for studying the contribution of systemic inflammatory processes on remyelination after cessation of CPZ treatment.

Topics: Acute Disease; Animals; Body Weight; CD11b Antigen; Chelating Agents; Chronic Disease; Corpus Callosum; Corticosterone; Cuprizone; Cytokines; Demyelinating Diseases; Disease Models, Animal; Leukocyte Common Antigens; Lipopolysaccharides; Male; Mice; Mice, Inbred C57BL; Microglia; Myelin Basic Protein; Myelin Sheath; Nerve Regeneration; Spleen

Progressive demyelination in multiple sclerosis (MS) reflects the negative balance between myelin damage and repair due to physical and molecular barriers, such as astrocytic glial scars, between oligodendrocytes and target neurons. In this paper, we show that combination therapy with paclitaxel (Taxol) plus the universal methyl-donor, vitamin B12CN (B12CN), dramatically limits progressive demyelination, and enhances remyelination in several independent, immune and nonimmune, in vivo and in vitro model systems. Combination therapy significantly reduced clinical signs of EAE in SJL mice, as well as the spontaneously demyelinating ND4 transgenic mouse. Astrocytosis was normalised in parallel to ultrastructural and biochemical evidence of remyelination. The combination therapy suppressed T cell expansion, reduced IFN-gamma, while enhancing IFN-beta and STAT-1 expression, STAT-1 phosphorylation and methylation of STAT-1 and MBP in the brain. Paclitaxel/B12CN has nearly identical effects to the previously described combination of IFN-beta/ B12CN, whose clinical usefulness is transient because of IFN-neutralising antibodies, not observed (or expected) with the present drug combination. This report provides a mechanistic foundation for the development of a new therapeutic strategy in humans with MS.

Topics: Animals; Antineoplastic Agents, Phytogenic; Autoimmune Diseases; Demyelinating Diseases; Drug Synergism; Gliosis; Methylation; Mice; Mice, Inbred Strains; Mice, Transgenic; Myelin Basic Protein; Nerve Fibers, Myelinated; Nerve Regeneration; Paclitaxel; STAT1 Transcription Factor; T-Lymphocytes; Vitamin B 12; Vitamin B Complex

Metachromatic leukodystrophy (MLD) is a lysosomal storage disorder caused by the deficiency of arylsulfatase A (ASA). This results in accumulation of sulfated glycosphingolipids, mainly 3-O-sulfogalactosylceramide (sulfatide), in the nervous system and various other organs. In patients, lipid storage causes a progressive loss of myelin leading to various neurological symptoms. The sulfatide storage pattern in ASA-deficient [ASA(-/-)] mice is comparable to humans, but regrettably, the mice do not mimic the myelin pathology. We reasoned that increasing sulfatide storage in this animal model might provoke demyelination. Therefore, we generated transgenic ASA(-/-) [tg/ASA(-/-)] mice overexpressing the sulfatide-synthesizing enzyme galactose-3-O-sulfotransferase-1 in myelinating cells. Indeed, these tg/ASA(-/-) mice displayed a significant increase in sulfatide storage in brain and peripheral nerves. Mice older than 1 year developed severe neurological symptoms. Nerve conduction velocity was significantly reduced in tg/ASA(-/-) mice because of a peripheral neuropathy characterized by hypomyelinated and demyelinated axons. Inhomogeneous myelin thickness in the corpus callosum, increased frequency of hypomyelinated and demyelinated axons in corpus callosum and optic nerve, and substantially reduced myelin basic protein levels are in accordance with loss of myelin in the CNS. Thus, increasing sulfatide storage in ASA(-/-) mice leads to neurological symptoms and morphological alterations that are reminiscent of human MLD. The approach described here may also be applicable to improve other mouse models of lysosomal as well as nonlysosomal disorders.

Topics: Age Factors; Animals; Cerebroside-Sulfatase; Demyelinating Diseases; Disease Models, Animal; Electromyography; Hindlimb Suspension; Humans; Leukodystrophy, Metachromatic; Lipids; Mice; Mice, Knockout; Microscopy, Electron, Transmission; Motor Activity; Myelin Basic Protein; Myelin Sheath; Neural Conduction; Peripheral Nerves; Rotarod Performance Test; Sciatic Nerve; Spinal Cord; Sulfoglycosphingolipids

Inflammatory response, axonal damage and demyelination are important components of the pathophysiology of acute neurodegenerative diseases. We have investigated the outcome of these pathological events following an excitotoxic or an ischemic damage to the spinal nucleus of adult rats at 1 and 7 days postinjury. Microinjections of 80 nmol of NMDA or 40 pmol of endothelin-1 into the rat spinal nucleus induced differential histopathological events. NMDA injection induced intense tissue loss in the gray matter (GM) without significant tissue loss in the white matter (WM). There was a mild inflammatory response, with recruitment of a few neutrophils and macrophages. Axonal damage was present in the GM following NMDA injection, with negligible axonal damage in the WM. Myelin impairment was apparent at 7 days. Microinjections of endothelin-1 into the same region induced lesser tissue loss than NMDA injections, concomitant with an intense inflammatory response characterized by recruitment of macrophages, but not of neutrophils. There were more axonal damage and early myelin impairment after endothelin-1 injection. These results were confirmed by quantitative analysis. Microcysts were present in the WM of the trigeminothalamic tract at 7 days following injection of endothelin-1. These results show that an ischemic damage to the spinal nucleus affects both GM and WM with more bystander inflammation, axonal damage and myelin impairment, while excitotoxic damage induces effects more restricted to the GM. These pathological events may occur following acute damage to the human brain stem and can be an important contributing factor to the underlying functional deficits.

Topics: Amyloid beta-Protein Precursor; Analysis of Variance; Animals; Axons; Brain Ischemia; Cell Count; Demyelinating Diseases; Ectodysplasins; Endothelin-1; Inflammation; Male; Myelin Basic Protein; N-Methylaspartate; Neurotoxins; Rats; Rats, Wistar; Time Factors; Trigeminal Nucleus, Spinal

Diffusion tensor magnetic resonance imaging (DT-MRI) was applied for in vivo quantification of myelin loss and regeneration. A transgenic mouse line (Oligo-TTK) expressing a truncated form of the herpes simplex virus 1 thymidine kinase gene (hsv1-tk) in oligodendrocytes was studied along with two induced phenotypes of myelin pathology. Myelin loss and axonal abnormalities differentially affect values of DT-MRI parameters in the brain of transgenic mice. Changes in the anisotropy of the white matter were assessed by calculating and mapping the radial (D perpendicular) and axial (D parallel) water diffusion to axonal tracts and fractional anisotropy (FA). A significant increase in D perpendicular attributed to the lack of myelin was observed in all selected brain white matter tracts in dysmyelinated mice. Lower D parallel values were consistent with the histological observation of axonal modifications, including reduced axonal caliber and overexpression of neurofilaments and III beta-tubulin. We show clearly that myelination and axonal changes play a role in the degree of diffusion anisotropy, because FA was significantly decreased in dysmyelinated brain. Importantly, myelin reparation during brain postnatal development induced a decrease in the magnitude of D( perpendicular) and an increase in FA compared with the same brain before recovery. The progressive increase in D parallel values was attributed to the gain in normal axonal morphology. This regeneration was confirmed by the detection of enlarged oligodendrocyte population, newly formed myelin sheaths around additional axons, and a gradual increase in axonal caliber.

Topics: Animals; Animals, Newborn; Anisotropy; Antiviral Agents; Brain; Brain Diseases; Brain Mapping; Demyelinating Diseases; Diffusion Magnetic Resonance Imaging; Ganciclovir; Image Processing, Computer-Assisted; Immunohistochemistry; Mice; Mice, Transgenic; Microscopy, Electron, Transmission; Myelin Basic Protein; Nerve Fibers, Myelinated; Oligodendroglia; Protein Serine-Threonine Kinases; Protein-Tyrosine Kinases; Recovery of Function; Time Factors

Twenty-one-day-old Wistar rats were fed a diet containing 0.6% cuprizone for 2 weeks. Studies carried out after withdrawal of cuprizone showed histological evidences of marked demyelination in the corpus callosum. Biochemical studies of isolated myelin showed a marked decrease in myelin proteins, phospholipids, and galactocerebrosides as well as a marked decrease in myelin yield. Treatment of these animals with a single intracranial injection of 350 ng of apotransferrin at the time of withdrawal of cuprizone induced a marked increase in myelin deposition resulting in a significantly improved remyelination, evaluated by histological, immunocytochemical, and biochemical parameters, in comparison to what was observed in spontaneous recovery. Immunocytochemical studies of cryotome sections to analyze developmental parameters of the oligodendroglial cell population at the time of termination of cuprizone and at different times thereafter showed that in the untreated animals, there was a marked increase in the number of NG2-BrdU-positive precursor cells together with a marked decrease in MBP expression at the peak of cuprizone-induced demyelination. As expected, the amount of precursor cells decreased markedly during spontaneous remyelination and was accompanied by an increase in MBP reactivity. In the apotransferrin-treated animals, these phenomena occurred much faster, and remyelination was much more efficient than in the untreated controls. The results of this study suggest that apotransferrin is a very active promyelinating agent which could be important for the treatment of certain demyelinating conditions.

Topics: Analysis of Variance; Animals; Animals, Newborn; Antigens; Apoproteins; Brain; Bromodeoxyuridine; CD11b Antigen; Cell Count; Cuprizone; Cytoskeletal Proteins; Demyelinating Diseases; Drug Interactions; Galactolipids; Glial Fibrillary Acidic Protein; Immunohistochemistry; Indoles; Myelin Basic Protein; Myelin Sheath; Proteoglycans; Rats; Rats, Wistar; Recovery of Function; Regeneration; Time Factors; Transferrin

Axonal degeneration contributes to the transient and permanent neurological deficits seen in multiple sclerosis, an inflammatory disease of the central nervous system. To study the immunological mechanisms causing axonal degeneration, we induced experimental autoimmune encephalomyelitis (EAE) in wildtype Lewis rats and Lewis rats with a slowly progressive myelin degeneration due to proteolipid protein (PLP) overexpression. EAE was triggered either by the transfer of encephalitogenic T-cells alone or by the co-transfer of T-cells with demyelinating antibodies. Inducible nitric oxide synthase (iNOS) expression in perivascular macrophages was associated with a transient functional disturbance of axons, reflected by the focal and reversible accumulation of amyloid precursor protein. Clinical disease correlated with the numbers of APP positive axon spheroids. Demyelination was associated with a further increase of iNOS expression in macrophages and with a higher degree of axonal injury. Our studies suggest that nitric oxide and its metabolites contribute to axonal pathology and possibly also to subsequent neurological dysfunction in EAE.

Topics: Amyloid beta-Protein Precursor; Animals; Animals, Genetically Modified; Axons; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Immunohistochemistry; Microscopy, Confocal; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Nitric Oxide; Nitric Oxide Synthase Type II; Proteolipids; Rats; Rats, Inbred Lew; T-Lymphocytes

Recent studies revealed an important involvement of the cerebral cortex in multiple sclerosis (MS) patients. Cortical lesions in MS were reported to be less inflammatory and to show less structural damage than white matter lesions. Animal models reflecting the histopathological hallmarks of cortical demyelinated lesions in MS are sparse. Induction of experimental autoimmune encephalomyelitis (EAE) in the common marmoset has turned out to be an attractive non-human-primate model for MS. In the present study we investigated the presence and detailed cellular composition of cortical inflammatory demyelinating pathology in the common marmoset upon immunization with myelin oligodendrocyte glycoprotein (MOG). Extensive cortical demyelination reflecting the topographically distinct cortical lesion types in MS patients was revealed by immunohistochemistry for myelin basic protein (MBP). We explored the density of T- and B-lymphocytes, MHC-II expressing macrophages/microglia cells and early activated macrophages (MRP14) at perivascular and parenchymal lesions sites in neocortex and subcortical white matter. Despite a similar density of perivascular inflammatory infiltrates in the demyelinated neocortex, a considerable lower fraction of macrophages was found to express MRP14 in the neocortex indicating a different activation pattern in cortical compared with white matter lesions. Furthermore, cortical EAE lesions in marmoset monkeys revealed immunoglobulin leakage and complement component C9 deposition in intracortical but not subpial demyelination. Our findings indicate that the inflammatory response, especially macrophage and microglia activation, may be regulated differently in gray matter areas in primate brain.

Topics: Animals; Callithrix; Demyelinating Diseases; Disease Models, Animal; Encephalomyelitis, Autoimmune, Experimental; Female; Immunization; Macrophages; Male; Microglia; Multiple Sclerosis; Myelin Basic Protein; Myelin Proteins; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Neocortex; Neutrophil Infiltration; Tissue Distribution

Although neural stem cells hold considerable promise for treatment of the injured or degenerating nervous system, their current human sources are embryonic stem cells and fetally derived neural tissue. Here, we asked whether rodent and human skin-derived precursors (SKPs), neural crest-related precursors found in neonatal dermis, represent a source of functional, myelinating Schwann cells. Specifically, cultured SKPs responded to neural crest cues such as neuregulins to generate Schwann cells, and these Schwann cells proliferated and induced myelin proteins when in contact with sensory neuron axons in culture. Similar results were obtained in vivo; 6 weeks after transplantation of naive SKPs or SKP-derived Schwann cells into the injured peripheral nerve of wild-type or shiverer mutant mice (which are genetically deficient in myelin basic protein), the majority of SKP-derived cells had associated with and myelinated axons. Naive rodent or human SKPs also generated Schwann cells that myelinated CNS axons when transplanted into the dysmyelinated brain of neonatal shiverer mice. Thus, neonatal SKPs generate functional neural progeny in response to appropriate neural crest cues and, in so doing, provide a highly accessible source of myelinating cells for treatment of nervous system injury, congenital leukodystrophies, and dysmyelinating disorders.

Topics: Animals; Animals, Newborn; Cell Differentiation; Cells, Cultured; Cerebellum; Coculture Techniques; Demyelinating Diseases; Embryo, Mammalian; Ganglia, Spinal; Humans; Immunohistochemistry; Mice; Mice, Neurologic Mutants; Microscopy, Electron; Multipotent Stem Cells; Myelin Basic Protein; Nerve Tissue Proteins; Neurons; Peripheral Nervous System Diseases; Rats; Schwann Cells; Skin; Stem Cell Transplantation

Protease M/neurosin is a serine protease expressed by oligodendrocytes (OLGs) in the central nervous system (CNS). To investigate the role of protease M/neurosin during experimental demyelination and remyelination, mice were fed cuprizone (bis-cyclohexanon oxaldihydrazone). Semi-quantitative RT-PCR analysis and immunohistochemistry revealed that the expressions of protease M/neurosin mRNA and protein were rapidly reduced in demyelination, whereas the expression of protease M/neurosin was increased in pi form of glutathione-S-transferases (GST-pi)-positive OLGs during remyelination. Cultured primary OLGs displayed a strong correlation between protease M/neurosin and myelin basic protein (MBP). After tumor necrosis factor-alpha (TNF-alpha) and IFN-gamma stimulation, these proteins showed colocalization in the oligodendroglial process. The suppression of protease M/neurosin using RNAi reduced the level of MBP mRNA in cultured OLGs. In contrast, the reduced level of protease M/neurosin was not associated with oligodendroglial cell death or differentiation in cultured OLGs. This study identifies that protease M/neurosin in OLGs is closely associated with the expression of the MBP and the PLP gene. Our data emphasize that the maintenance of myelination is an important function of protease M/neurosin in OLGs, suggesting its relation to the oligodendroglial response to myelin disorders.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Cells, Cultured; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Glutathione S-Transferase pi; Immunohistochemistry; Interferon-gamma; Kallikreins; Mice; Mice, Inbred BALB C; Monoamine Oxidase Inhibitors; Myelin Basic Protein; Myelin Sheath; Oligodendroglia; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; RNA, Small Interfering; Time Factors; Tumor Necrosis Factor-alpha

As a consequence of secondary pathophysiological mechanisms elicited after spinal cord injury (SCI), oligodendrocytes die by waves of apoptosis. This ultimately results in demyelination of intact axons leading to a loss of their conducting properties. Preservation of as few as 5% to 10% of myelinated axons in individual tracts can confer locomotor recovery. Thus, strategies aimed at rescuing mature oligodendrocytes ensheathing viable axons are likely to be of therapeutic significance. We report that leukemia inhibitory factor (LIF) can prevent oligodendrocyte apoptosis, notably contralateral to the spinal cord lesion, through the induction of the JAK/STAT and Akt signaling pathways as well as by potentiating the expression of the antiapoptotic molecule, cIAP2. Reduced oligodendrocyte apoptosis after SCI with LIF administration resulted in a substantial decrease in demyelination shown by the preservation of lamellated myelin surrounding viable axons and deposition of the degraded myelin basic protein. The data suggest that LIF signals survival in oligodendrocytes after SCI, prevents the secondary wave of demyelination, and thereby reduces inhibitory myelin deposits.

Topics: Animals; Axotomy; Baculoviral IAP Repeat-Containing 3 Protein; Cell Death; Demyelinating Diseases; Disease Models, Animal; Female; Gene Expression; Immunoprecipitation; In Situ Nick-End Labeling; Inhibitor of Apoptosis Proteins; Interleukin-6; Leukemia Inhibitory Factor; Leukemia Inhibitory Factor Receptor alpha Subunit; Mice; Mice, Inbred C57BL; Myelin Basic Protein; Oligodendroglia; Receptors, Cytokine; Receptors, OSM-LIF; Severity of Illness Index; Spinal Cord Injuries; STAT Transcription Factors; Ubiquitin-Protein Ligases

Remyelination is a critical step for recovery of function after demyelination and defines the ability to generate new myelin. This repair process is dependent on the presence of resident oligodendrocyte progenitors (OLPs) that have been shown to remyelinate axons after demyelination. We have previously shown that the levels of the cell cycle inhibitor p27Kip-1 modulate the number of neonatal cortical OLPs. We now asked whether this cell cycle molecule plays also a role in regulating the number of adult OLP in the spinal cord after demyelination induced by lysolecithin injection. The proliferative response of OLP in the spinal cord of injected wild-type (wt) and p27Kip-1 null mice was evaluated 3 days after lesion. In vivo labeling with bromodeoxyuridine (BrdU) was used to identify cells in S phase. Double immunofluorescence for the OLP marker NG2, and for BrdU was used to count the number of proliferating progenitors. Consistent with a role of p27Kip-1 in regulating the number of adult OLP in the injured spinal cord, a larger number of proliferating OLPs was observed in p27Kip-1null mice compared with wild-type controls. These cells were able to differentiate as assessed by the presence of MBP+ cells in the spinal cord 14 days after injury. We conclude that the cellular levels of the cell cycle inhibitor p27Kip-1 modulate the repair response of OLPs to injury in the adult spinal cord.

Topics: Animals; Bromodeoxyuridine; Cell Count; Cell Cycle Proteins; Cell Differentiation; Cell Division; Cell Proliferation; Cyclin-Dependent Kinase Inhibitor p27; Demyelinating Diseases; Female; Gene Expression Regulation; Lysophosphatidylcholines; Male; Mice; Mice, Knockout; Myelin Basic Protein; Nerve Regeneration; Oligodendroglia; S Phase; Spinal Cord Diseases; Stem Cells; Tumor Suppressor Proteins

Proliferation of oligodendrocyte progenitor cells (OPCs) is important for initial myelination as well as for remyelination in demyelinating diseases. Previously, we showed that numerous OPCs and activated microglia, are present around multiple sclerosis lesions, and that they accumulate Golli proteins. Golli proteins, present in both neuronal and immune cells, might have a role in the immune processes, as well as in development of neurons and oligodendrocytes. We hypothesize that Golli proteins, generated by microglia in response to inflammation, promote proliferation of OPCs. To test this hypothesis, we induced inflammation in neonatal mouse brain slice culture with bacterial endotoxin lipopolysaccharide (LPS). Treated slices showed an increase in the number of OPCs. Several results support the notion that this effect of LPS is conveyed through activation of microglia and upregulation of Golli proteins. First, LPS-treated brain slices have increased expression of Golli proteins observed by immunofluorescence and Western blot analysis. Second, Golli proteins were demonstrated only in the conditioned medium from LPS-treated microglial cell cultures (LPS-MCM), and were absent in either the conditioned media from LPS-treated astrocytes or the control media. Third, proliferation of purified OPCs was promoted with LPS-MCM or Golli proteins, but not with LPS alone. Taken together, these results demonstrate that microglia and/or microglia secreted factors, are necessary for the LPS-promoted proliferation of OPCs and suggest possible involvement of Golli proteins as one of mediators in this process.

Topics: Animals; Cell Communication; Cell Differentiation; Cell Proliferation; Cells, Cultured; Culture Media, Conditioned; Demyelinating Diseases; Encephalitis; Fluorescent Antibody Technique; Lipopolysaccharides; Mice; Mice, Inbred ICR; Microglia; Myelin Basic Protein; Nerve Regeneration; Nerve Tissue Proteins; Oligodendroglia; Organ Culture Techniques; Stem Cells; Transcription Factors; Up-Regulation

Although macrophages are mediators of CNS demyelination, they are also implicated in remyelination. To examine the role of macrophages in CNS remyelination, adult rats were depleted of monocytes using clodronate liposomes and demyelination induced in the spinal cord white matter using lysolecithin. In situ hybridization for scavenger receptor-B and myelin basic protein (MBP) revealed a transiently impaired macrophage response associated with delayed remyelination in liposome-treated animals. Macrophage reduction corresponded with delayed recruitment of PDGFRalpha+ oligodendrocyte progenitor cells (OPCs), which preceded changes in myelin phagocytosis, indicating a macrophage effect on OPCs independent of myelin debris clearance. Macrophage-depletion induced changes in the mRNA expression of insulin-like growth factor-1 and transforming growth factor beta1, but not platelet-derived growth factor-A and fibroblast growth factor-2. These data suggest that the macrophage response to toxin-induced demyelination influences the growth factor environment, thereby affecting the behavior of OPCs and hence the efficiency of remyelination.

Topics: Animals; Central Nervous System; Demyelinating Diseases; Disease Models, Animal; Down-Regulation; Female; Growth Substances; Macrophages; Myelin Basic Protein; Myelin Sheath; Nerve Regeneration; Oligodendroglia; Phagocytosis; Rats; Rats, Sprague-Dawley; Receptor, Platelet-Derived Growth Factor alpha; Receptors, Immunologic; Receptors, Scavenger; RNA; Stem Cells

We have previously shown that fibroblast growth factor 2 (FGF2) disrupts myelin formation by oligodendrocytes in vivo. Here, we have investigated the possibility that this is associated with changes in the expression of tau, a major microtubule-associated protein (MAP) involved in the production of myelin membranes by oligodendrocytes. FGF2, or saline vehicle in controls, was delivered into the brain ventricles of deeply anaesthetised young rats, and their actions were examined on the anterior medullary velum (AMV), a thin sheet of tissue that roofs part of the ventricular system underlying the cerebellum. The results show that the FGF2-induced loss of myelin is associated with increased immunostaining for tau within oligodendrocyte somata. Immunohistochemical and Western blot analyses demonstrate a 50% decrease in myelin-forming oligodendrocytes, axonal myelin sheaths, and levels of myelin-related proteins, with a correlative 100% increase in the level of tau. The results identify a potential mechanism by which FGF2-mediated accumulation of tau disrupts the transport of myelin-related gene products, resulting in disruption and eventual loss of oligodendrocytes and myelin, which are features of ischemia and a variety of demyelinating and neurodegenerative diseases.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Analysis of Variance; Animals; Animals, Newborn; Blotting, Western; Demyelinating Diseases; Drug Administration Schedule; Fibroblast Growth Factor 2; Gene Expression Regulation, Developmental; Immunohistochemistry; Medulla Oblongata; Myelin Basic Protein; Neurofilament Proteins; Oligodendroglia; Oligopeptides; Rats; Rats, Wistar; tau Proteins

The natural history of post-traumatic demyelination and myelin repair in the human spinal cord is largely unknown and has remained a matter of speculation. A wealth of experimental studies indicate that mild to moderate contusive injuries to the mammalian spinal cord evolve into a cavity with a preserved rim of white matter in which a population of segmentally demyelinated axons persists. It is believed that such injured axons have abnormal conduction properties. Theoretically, such axons might show improved function if myelin repair occurred. Schwann cells can remyelinate axons affected by multiple sclerosis, but little evidence exists that such repair can occur spontaneously following traumatic human SCI. Therefore, it is important to determine if chronic demyelination is present following human spinal cord injury. There are no previous reports that have conclusively demonstrated demyelination in the human spinal cord following traumatic spinal cord injury using immunohistochemical techniques. Immunohistochemical methods were used to study the distribution of peripheral and central myelin proteins as well as axonal neurofilament at the injury epicenter in 13 postmortem chronically injured human spinal cords 1-22 years following injury. Of these seven could be assessed by our methods. We found that some axonal demyelination can be detected even a decade following human SCI and indirect evidence that invading Schwann cells contributed to restoration of myelin sheaths around some spinal axons.

Topics: Adult; Aged; Aged, 80 and over; Demyelinating Diseases; Diagnostic Imaging; Female; Glial Fibrillary Acidic Protein; Humans; Immunohistochemistry; Male; Middle Aged; Myelin Basic Protein; Myelin Proteolipid Protein; Neurofilament Proteins; Postmortem Changes; Schwann Cells; Spinal Cord Injuries; Survival Rate; Time Factors

A transgenic mouse line (Oligo-TTK) was established to monitor oligodendrocyte cell death and myelin formation in the CNS. The expression of a conditionally toxic gene, the herpes simplex virus-1 thymidine kinase (HSV1-TK), was made under control of the MBP (myelin basic protein) gene promoter. A truncated form of the HSV1-TK (TTK) gene was used to avoid both bystander effect resulting from leaking in thymidine kinase activity and sterility in transgenic males observed in previous transgenic mice. The transgene was expressed in the CNS with a restricted localization in oligodendrocytes. Oligodendrocyte proliferation and myelin formation are therefore tightly controlled experimentally by administration of ganciclovir (GCV) via the induction of oligodendrocyte cell death. The most severe and irreversible hypomyelination was obtained when GCV was given daily from postnatal day 1 (P1) to P30. Oligodendrocyte plasticity and myelin recovery were analyzed in another phenotype generated by GCV treatment from P1 to P15. In this model, after dysmyelination, an apparent normal behavior was restored with no visible pathological symptoms by P30. Proliferating cells, which may be implicated in myelin repair in this model, are detected primarily in myelin tracts expressing the oligodendrocyte phenotype. Therefore, the endogenous potential of oligodendrocytes to remyelinate was clearly demonstrated in the mice of this study.

Topics: Age Factors; Analysis of Variance; Animals; Animals, Newborn; Antiviral Agents; Brain; Bromodeoxyuridine; Cell Death; Demyelinating Diseases; Disease Models, Animal; Ganciclovir; Gene Expression Regulation, Developmental; Herpesvirus 1, Human; Immunohistochemistry; In Situ Hybridization; Male; Mice; Mice, Transgenic; Microscopy, Electron, Transmission; Myelin Basic Protein; Myelin Sheath; Oligodendroglia; Promoter Regions, Genetic; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Thymidine Kinase

How do myelinated axons signal to the nuclei of cells that enwrap them? The cell bodies of oligodendrocytes and Schwann cells are segregated from axons by multiple layers of bimolecular lipid leaflet and myelin proteins. Conventional signal transduction strategies would seem inadequate to the challenge without special adaptations. Wallerian degeneration provides a model to study axon-to-Schwann cell signaling in the context of nerve injury. We show a hitherto undetected rapid, but transient, activation of the receptor tyrosine kinase erbB2 in myelinating Schwann cells after sciatic nerve axotomy. Deconvolving microscopy using phosphorylation state-specific antibodies shows that erbB2 activation emanates from within the microvilli of Schwann cells, in direct contact with the axons they enwrap. To define the functional role of this transient activation, we used a small molecule antagonist of erbB2 activation (PKI166). The response of myelinating Schwann cells to axotomy is inhibited by PKI166 in vivo. Using neuron/Schwann cell cocultures prepared in compartmentalized cell culture chambers, we show that even transient activation of erbB2 is sufficient to initiate Schwann cell demyelination and that the initiating functions of erbB2 are localized to Schwann cells.

Topics: Analysis of Variance; Animals; Axotomy; Blotting, Western; Bromodeoxyuridine; Cell Proliferation; Cells, Cultured; Coculture Techniques; Demyelinating Diseases; Disease Models, Animal; Embryo, Mammalian; Female; Fluorescent Antibody Technique; Ganglia, Spinal; Gene Expression; Glycoproteins; Immunoprecipitation; Mitogen-Activated Protein Kinase Kinases; Myelin Basic Protein; Myelin Sheath; Neuregulins; Neuroglia; Neurons; Platelet-Derived Growth Factor; Pyrimidines; Pyrroles; Rats; Rats, Sprague-Dawley; Receptor Protein-Tyrosine Kinases; Receptor, ErbB-2; Schwann Cells; Sciatic Neuropathy; Signal Transduction; Sodium Channels; Time Factors; Wallerian Degeneration

Inflammation, demyelination, and axonal damage of the central nervous system (CNS) are major pathological features of multiple sclerosis (MS). Proteolytic digestion of the blood-brain barrier and myelin protein by serine proteases is known to contribute to the development and progression of MS. Neuropsin, a serine protease, has a role in neuronal plasticity, and its expression has been shown to be upregulated in response to injury to the CNS. To determine the possible involvement of neuropsin in demyelinating diseases of the CNS, we examined its expression in myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalomyelitis (EAE), a recognized animal model for MS. Neuropsin mRNA expression was induced in the spinal cord white matter of mice with EAE. Combined in situ hybridization and immunohistochemistry demonstrated that most of the cells expressing neuropsin mRNA showed immunoreactivity for CNPase, a cell-specific marker for oligodendrocytes. Mice lacking neuropsin (neuropsin-/-) exhibited an altered EAE progression characterized by delayed onset and progression of clinical symptoms as compared to wild-type mice. Neuropsin-/- mice also showed attenuated demyelination and delayed oligodendroglial death early during the course of EAE. These observations suggest that neuropsin is involved in the pathogenesis of EAE mediated by demyelination and oligodendroglial death.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Demyelinating Diseases; Disease Progression; Encephalomyelitis, Autoimmune, Experimental; Fluorescent Antibody Technique; Glial Fibrillary Acidic Protein; Immunohistochemistry; In Situ Hybridization; In Situ Nick-End Labeling; Kallikreins; Mice; Mice, Inbred C57BL; Mice, Knockout; Myelin Basic Protein; Myelin Proteins; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Peptide Fragments; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Serine Endopeptidases; Tumor Necrosis Factor-alpha

Glutamate excitotoxicity is increasingly being recognized as a pathogenic mechanism in autoimmune inflammatory disorders of the central nervous system (CNS). Astrocytes are the predominant players in clearing the extracellular space from glutamate and normally have extensive spare capacities in terms of glutamate uptake. We asked what might be the basis of glutamate accumulation in T cell triggered autoimmune inflammation. In vitro, coculture of primary rat astrocytes with activated myelin basic protein (MBP)-specific T cells resulted in a decrease of astrocytic glutamate uptake rates (Vmax). In parallel, the amount of the Na+-dependent glutamate transporter GLAST was reduced within 48-60 h. Significant decreases of GLAST protein were observed in astrocytes harvested after incubation with T cells activated by MBP during coculture or after incubation with T cell blasts preactivated in the presence of splenocytes beforehand. Since exposure of astrocytes to cell-free supernatant of MBP-activated T cells also resulted in reduced expression of GLAST, a humoral factor appeared to be the driving agent. In blocking experiments using neutralizing antibodies and by incubation of astrocytes with recombinant cytokines, tumor necrosis factor-alpha (TNF-alpha) was identified as being responsible for the down-modulation of GLAST. GLAST was also down-regulated in the CNS of autoimmune encephalomyelitic rats but not in animals suffering from systemic inflammation. Since the loss of GLAST was not confined to inflammatory infiltrates, here too, a humoral factor seemed to be causative. In conclusion, T cell derived TNF-alpha impairs glutamate clearance capacity of astrocytes in vitro and probably also in vivo providing a pathogenic link to glutamate excitotoxicity that may contribute to early axonal dysfunction remote from active autoimmune inflammatory demyelination.

Topics: Amino Acid Transport System X-AG; Animals; Astrocytes; Autoantigens; Binding Sites; Central Nervous System; Coculture Techniques; Cytokines; Demyelinating Diseases; Dose-Response Relationship, Drug; Down-Regulation; Encephalomyelitis, Autoimmune, Experimental; Flow Cytometry; Glutamates; Immunoblotting; Inflammation; Kinetics; Models, Biological; Myelin Basic Protein; Rats; Sodium; T-Lymphocytes; Time Factors; Tumor Necrosis Factor-alpha

Adult macaque Schwann cells were infected using adeno-associated virus type-2-derived vectors expressing the green fluorescent protein reporter gene under the control of the cytomegalovirus, the hybrid cytomegalovirus-betaactin, the myelin basic protein or the tetracycline-inducible promoters. On the basis of green fluorescent protein expression, gene transfer efficiency was compared in resting and dividing conditions following or not following hydroxyurea or etoposide treatment. Hydroxyurea allowed promoter-dependent expression of green fluorescent protein in infected Schwann cells. Etoposide treatment led to a high percentage of green fluorescent protein expressing cells (over 50%) with all promoters tested. When infected cells were grafted into demyelinated nude mice spinal cord, green fluorescent protein expression was only observed with the cytomegalovirus-betaactin and tetracycline-inducible promoters. In addition, adeno-associated virus type-2 infection reduced the grafted cell survival but increased their differentiation.

Topics: Analysis of Variance; Animals; Cell Count; Cell Proliferation; Cell Transplantation; Cells, Cultured; Cytomegalovirus; Demyelinating Diseases; Dependovirus; Etoposide; Fibroblasts; Gene Expression Regulation; Genetic Vectors; Green Fluorescent Proteins; Hydroxyurea; Immunohistochemistry; Macaca fascicularis; Mice; Myelin Basic Protein; Nucleic Acid Synthesis Inhibitors; Promoter Regions, Genetic; Schwann Cells; Time Factors; Transduction, Genetic

Cyclic AMP response element (CRE) is a specific DNA sequence, which mediates transcriptional activation in the response to the cyclic AMP-activated and protein kinase A dependent signaling pathway. In the present study, phosphorylated CRE binding protein (CREB) immunoreactivity was mainly localized in the white matter of chick central nervous system (CNS). We have further confirmed the specificity of phospho-CREB immunoreaction in myelin using demyelinated optic nerve induced by lysophophatidylcholine (LPC), which is known to produce demyelination with little axonal damage. Double immunofluorescent analyses with myelin basic protein (MBP) and transferrin binding protein (TfBP), oligodendrocyte marker showed that phospho-CREB recognized a myelin-related protein in chick. Immunoblot analyses showed that phospho-CREB recognized a protein with molecular weights of approximately 70 kDa. Our data suggest that the antigen recognized by phospho-CREB is a myelin-associated protein in the chick CNS.

Topics: Animals; Brain; Chickens; Cyclic AMP Response Element-Binding Protein; Demyelinating Diseases; Humans; Immunohistochemistry; Lysophosphatidylcholines; Myelin Basic Protein; Myelin Sheath; Optic Nerve; Phosphorylation; Rats; Transferrin-Binding Proteins

The role played by oligodendrocytes (OLs), the myelinating cells of the CNS, during brain development has not been fully explored. We have addressed this question by inducing a temporal and reversible ablation of OLs on postnatal CNS development. OL ablation in newborn mice leads to a profound alteration in the structure of the cerebellar cortex, which can be progressively rescued by newly generated cells, leading to a delayed myelination. Nevertheless, the temporal shift of the OL proliferation and myelinating program cannot completely compensate for developmental defects, resulting in impaired motor functions in the adult. Strikingly, we show that, despite these abnormalities, epigenetic factors, such as motor training, are able to fully rescue cerebellar-directed motor skills.

Topics: Animals; Animals, Newborn; Cerebellar Cortex; Demyelinating Diseases; Membrane Proteins; Mice; Mice, Inbred C57BL; Mice, Transgenic; Microscopy, Electron; Motor Skills; Myelin Basic Protein; Myelin Sheath; Oligodendroglia; Physical Conditioning, Animal; R-SNARE Proteins; Synaptic Vesicles; Thymidine Kinase

Excitotoxic oligodendroglial death is one of the mechanisms which has been proposed to underlie demyelinating diseases of the CNS. We describe here functional consequences of excitotoxic lesions to the rabbit optic nerve by studying the visual evoked potentials (VEPs) measured in the visual cortex. Nerves were slowly infused with the excitotoxin kainate a subcutaneously implanted osmotic pump which delivered the toxin through a cannula onto the optic nerve. Records of VEPs were obtained before pump implantation and at 1, 3 and 7 days post-implantation, and weekly evaluated thereafter for up to 4 months. We observed that the VEPs generated by light stimuli progressively changed in both amplitude and profile after the lesion as well as in comparison to those generated in control animals infused with vehicle. Histological examination of the damage caused by the excitotoxic insult showed that large areas of the optic nerve were demyelinated and their axons distorted. These observations were confirmed and extended by immunohistochemical analyses using markers to neurofilaments, myelin basic protein and the oligodendrocyte marker APC. The results of the present paper indicate that the consequences of excitotoxicity in the optic nerve share functional and morphological alterations which are found in demyelinating disorders. In addition, this experimental paradigm may be useful to evaluate the functional recovery of demyelinated optic nerves following various repair strategies.

Topics: Animals; Axons; Demyelinating Diseases; Electrodes, Implanted; Evoked Potentials, Visual; Excitatory Amino Acid Agonists; Image Processing, Computer-Assisted; Immunohistochemistry; Kainic Acid; Myelin Basic Protein; Neurofilament Proteins; Oligodendroglia; Optic Nerve Injuries; Photic Stimulation; Rabbits; Visual Cortex

Both late-gestation and adult human forebrain contain large numbers of oligodendrocyte progenitor cells (OPCs). These cells may be identified by their A2B5(+)PSA-NCAM(-) phenotype (positive for the early oligodendrocyte marker A2B5 and negative for the polysialylated neural cell adhesion molecule). We used dual-color fluorescence-activated cell sorting (FACS) to extract OPCs from 21- to 23-week-old fetal human forebrain, and A2B5 selection to extract these cells from adult white matter. When xenografted to the forebrains of newborn shiverer mice, fetal OPCs dispersed throughout the white matter and developed into oligodendrocytes and astrocytes. By 12 weeks, the host brains showed extensive myelin production, compaction and axonal myelination. Isolates of OPCs derived from adult human white matter also myelinated shiverer mouse brain, but much more rapidly than their fetal counterparts, achieving widespread and dense myelin basic protein (MBP) expression by 4 weeks after grafting. Adult OPCs generated oligodendrocytes more efficiently than fetal OPCs, and ensheathed more host axons per donor cell than fetal cells. Both fetal and adult OPC phenotypes mediated the extensive and robust myelination of congenitally dysmyelinated host brain, although their differences suggested their use for different disease targets.

Topics: Adult; Animals; Demyelinating Diseases; Fetus; Humans; Immunohistochemistry; Mice; Microscopy, Confocal; Microscopy, Electron; Myelin Basic Protein; Oligodendroglia; Stem Cells

Multiple sclerosis (MS) is a progressive immune-mediated disease characterized by the loss of the oligodendrocytes that constitute the myelin sheath. Recent reports show that glutamate-mediated excitotoxic death of oligodendrocytes contributes to pathogenesis in demyelinating disease. A link between the immune-mediated inflammatory response and glutamate-mediated excitotoxicity of oligodendrocytes could involve the interaction of inducible isoforms of nitric oxide synthase (iNOS) and cyclooxygenase (COX-2). Both enzymes are tightly coupled to neuronal excitotoxic death. We examined tissue from two controls and seven MS patients with chronic active lesions to determine the extent of COX-2 and iNOS expression. In contrast to the lack of expression in controls, there was a marked induction of COX-2 in all these MS lesions. COX-2 was frequently expressed in association with iNOS. COX-2 was found in areas that contained catabolites of myelin basic protein, indicating recent demyelination. COX-2 expression was found near damaged oligodendrocytes in cells that expressed the macrophage/microglia marker CD64, indicating that a substantial portion of the COX-2 in the lesions was expressed in immune-derived cells. We discuss these findings in the context of how COX-2 could be coupled with iNOS to contribute to excitotoxic death and damage of oligodendrocytes.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Adult; Brain; Cell Count; Cyclooxygenase 2; Demyelinating Diseases; Female; Fluorescent Antibody Technique; Humans; Inflammation; Isoenzymes; Male; Membrane Proteins; Microscopy, Confocal; Middle Aged; Models, Neurological; Multiple Sclerosis; Myelin Basic Protein; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Oligodendroglia; Prostaglandin-Endoperoxide Synthases; Protein Isoforms; Receptors, IgG; Staining and Labeling

Oligodendrocyte precursor cells require exogenous neurotrophin-3 (NT-3) for differentiation into oligodendrocytes. We transfected precursor cells with the gene for NT-3 and looked for changes in their development into myelin-forming cells. The expression of NT-3 in transfected cells was demonstrated by reverse transcription followed by PCR as well as by Northern blots. Direct synthesis of the neurotrophin product and its release to the culture supernatants were also shown by specific ELISA. Transfection converts precursor cells into actively dividing cells that can incorporate 3H-thymidine into DNA. In the absence of growth factors, a parallel increase in the survival of the transfected cultures was also demonstrated by the MTT test. The final demonstration of biological changes in transfected versus untreated cells was a 10-fold increase in myelin basic protein production observed in Western blots and the direct observation by phase-contrast and electron microscopy of myelin membranes in cocultures with hippocampal neurons. We discuss the future use of this transfected cells in regeneration and functional recovery in experimental models of multiple sclerosis.

Topics: Animals; Animals, Newborn; Cell Differentiation; Cell Division; Cell Survival; Cells, Cultured; Demyelinating Diseases; Genetic Vectors; Microscopy, Electron; Myelin Basic Protein; Myelin Sheath; Nerve Regeneration; Neurotrophin 3; Oligodendroglia; Rats; Rats, Wistar; RNA, Messenger; Stem Cells; Transfection; Up-Regulation

In order to investigate a role for insulin-like growth factor-1 (IGF-1) in ameliorating the effects of demyelinating events and potentiating remyelination, we have generated transgenic (tg) mice expressing IGF-1 under the control of the myelin basic protein promoter. Heterozygous tg mice expressed the highest levels of IGF-1 in brain during the most active periods of myelination as determined by Western and Northern blotting. A high level of expression was found throughout the lives of the tg mice. There was no increased expression of IGF-1 in other organs. The brains of heterozygous mice were larger than those of normal mice by 2 weeks of age, and they continued to increase in size for several months. Light and electron microscopy showed extensive myelination of axons. Behavioral studies of the older heterozygous mice documented difficulty with balance. This new tg mouse model can be bred to mice that are heterozygous for genetic leukodystrophies to produce eventually mice that are affected with a given leukodystrophy but overexpress IGF-1 during myelination and remyelination. It will be interesting to see if overexpression of IGF-1 can modulate the pathological and clinical features of the inherited leukodystrophies with or without supplemental therapies.

Topics: Animals; Blotting, Northern; Blotting, Western; Demyelinating Diseases; Immunohistochemistry; Insulin-Like Growth Factor I; Mice; Mice, Inbred C57BL; Mice, Transgenic; Myelin Basic Protein; Promoter Regions, Genetic; RNA, Messenger

We have shown previously that myelin-forming oligodendrocytes express the protein tyrosine phosphatase SHP-1 and that myelin formation was decreased in SHP-1-deficient motheaten mice compared to that in normal littermates. These studies suggested a potential importance for SHP-1 in oligodendrocyte and myelin development. To address further this possibility, we analyzed myelin formation by microscopy and myelin basic protein (MBP) gene expression in motheaten mice at ages when myelination occurs in the developing central nervous system (CNS). Furthermore, we correlate these findings with MBP gene expression in oligodendrocytes grown in vitro. We have found that CNS myelination was significantly reduced in SHP-1-deficient mice relative to their normal littermates at multiple times during the active period of myelination. Under electron microscopy, greater numbers of axons in spinal cords of motheaten mice were either unmyelinated or had thinner myelin sheathes compared to those in matched areas of normal littermates. Accordingly, MBP protein and mRNA levels were reduced in SHP-1-deficient mice compared to that in the CNS of normal littermates. In vitro, O1(+) oligodendrocytes from motheaten mice expressed much less MBP than O1(+) oligodendrocytes of normal littermates indicating an alteration in oligodendrocyte differentiation. The latter correlated with reduced MBP mRNA relative to cerebroside galactosyl transferase (CGT) gene mRNA in SHP-1-deficient oligodendrocytes in purified cultures. We propose that SHP-1 is a critical regulator of developmental signals leading to terminal differentiation and myelin sheath formation by oligodendrocytes.

Topics: Animals; Cell Differentiation; Cells, Cultured; Cerebrosides; Demyelinating Diseases; Disease Models, Animal; Down-Regulation; Female; Galactosyltransferases; Gene Expression Regulation, Developmental; Intracellular Signaling Peptides and Proteins; Male; Mice; Mice, Neurologic Mutants; Microscopy, Electron; Myelin Basic Protein; Myelin Sheath; Nerve Fibers, Myelinated; Oligodendroglia; Protein Tyrosine Phosphatase, Non-Receptor Type 6; Protein Tyrosine Phosphatases; RNA, Messenger

Voltage-gated potassium channels (Kv channels) are ion channels, openings of which provide an outward flow of potassium ions repolarising the cell. In neurons, Kv channels play a crucial role in action potential repolarisation and in shaping neuronal excitability. In non-excitable cells, such as T lymphocytes, Kv channels and calcium-activated K+ channels (KCa channels) determine the driving force for Ca2+ entry. During T cell activation the calcium entry depolarises the cell and increases the cytosolic calcium concentration, which in return activates Kv and KCa channels. K+ channel opening repolarises the cell and drives the membrane potential to a negative voltage. The roles of Kv channels in nervous and immune systems have been investigated here by means of a rat experimental autoimmune disease of the central nervous system, the experimental autoimmune encephalomyelitis (EAE). EAE is characterised clinically by paralysis, and pathologically by inflammatory cell infiltrations into the brain and the spinal cord. Among the inflammatory cells, T lymphocytes play a major role. Hence, EAE can be adoptively transferred into syngenic animals by the injection of T cells reactive to myelin antigens. During adoptive-EAE, somato-sensory evoked potentials recorded along the spinal tracts decrease in amplitude and axonal propagation is disrupted. We have analysed the consequences of Kv channels blockade by peptidyl toxins on central nerve conduction, on T cell activation and on the time course of EAE. In rat optic nerves, Kv channels have been identified up from postnatal day 1. Their blockade by kaliotoxin (a scorpion toxin) or by dendrotoxin-I (a snake toxin) enlarges the compound action potentials, demonstrating the participation of Kv channels to spike repolarisation. This effect disappears at adult age due to the sequestration of Kv channels under the myelin, in the paranodal regions. During acute demyelination by lysophosphatidyl-choline, the surface area of compound action potential decreased probably because conduction block occurred. Demyelination unmasked Kv channels, which are again accessible to toxins. Their blockade by dendrotoxin-I or kaliotoxin favoured a slow delayed conduction suggesting that those Kv channel blockers exert a neurological benefit during demyelinating diseases. In a T-cell line reactive to myelin basic protein antigen, which is used to adoptively transfer experimental autoimmune encephalomyelitis, Kv1.3 channels are constitutively e

Topics: Action Potentials; Adoptive Transfer; Aging; Animals; Animals, Newborn; Calcium; Cytokines; Demyelinating Diseases; Elapid Venoms; Encephalomyelitis, Autoimmune, Experimental; Ion Channel Gating; Ion Channels; Lymphocyte Activation; Myelin Basic Protein; Myelin Sheath; Neural Conduction; Neurotoxins; Optic Nerve; Potassium Channel Blockers; Rats; Rats, Wistar; Scorpion Venoms; T-Lymphocytes

Experimental autoimmune encephalomyelitis (EAE) and Theiler's murine encephalitis virus-induced demyelinating disease (TMEV-IDD) are two clinically relevant murine models of multiple sclerosis (MS). Like MS, both are characterized by mononuclear cell infiltrate into the central nervous system and demyelination. EAE is induced by either the administration of protein or peptide in adjuvant or by the adoptive transfer of encephalitogenic T-cell blasts into naïve recipients. The relative merits of each of these protocols are compared. Depending on the type of question asked, different mouse strains and peptides are used. Different disease courses are observed with different strains and different peptides in active EAE. These variations are addressed, and grading of mice in EAE is discussed. In addition to EAE induction, useful references for other disease indicators, such as delayed-type hypersensitivity, in vitro proliferation, and immunohistochemistry, are provided. TMEV-IDD is a useful model for understanding the potential viral etiology of MS. This chapter provides detailed information on the preparation of viral stocks and subsequent intracerebral infection of mice. In addition, virus plaque assay and disease assessment are discussed. Recombinant TMEV strains have been created for the study of molecular mimicry; these strains incorporate 30 various amino acid myelin epitopes within the leader region of TMEV.

Topics: Adoptive Transfer; Amino Acid Sequence; Animals; Cardiovirus Infections; Cell Line; Cricetinae; Demyelinating Diseases; Disease Models, Animal; Encephalomyelitis, Autoimmune, Experimental; Mice; Mice, Inbred Strains; Molecular Sequence Data; Multiple Sclerosis; Myelin Basic Protein; Myelin Proteolipid Protein; Peptide Fragments; Theilovirus; Virus Cultivation

Multiple sclerosis (MS) and experimental autoimmune encephalomyelitis (EAE), an animal model of MS, are inflammatory demyelinating diseases of the central nervous system. The inflammatory attacks lead to glial dysfunction and death, axonal damage, and neurological deficits. Numerous studies in rat suggest that extracellular calcium influx, via voltage-gated calcium channels (VGCC), contributes to white matter damage in acute spinal cord injury and stroke. Our immunohistochemical finding that mouse spinal cord axons display subunits of L-type VGCC also supports this hypothesis. Furthermore, we hypothesized that VGCC also play a role in EAE, and possibly, MS. In our study, administration of the calcium channel blockers (CCB) bepridil and nitrendipine significantly ameliorated EAE in mice, compared with vehicle-treated controls. Spinal cord samples showed reduced inflammation and axonal pathology in bepridil-treated animals. Our data support the hypothesis that calcium influx via VGCC plays a significant role in the development of neurological disability and white matter damage in EAE and MS.

Topics: Animals; Axons; Bepridil; Calcium Channel Blockers; Calcium Channels, L-Type; Demyelinating Diseases; Disease Models, Animal; Encephalomyelitis, Autoimmune, Experimental; Female; Immunohistochemistry; Mice; Multiple Sclerosis; Myelin Basic Protein; Neutrophil Infiltration; Nitrendipine; Spinal Cord; Time Factors; Wallerian Degeneration

The demyelination (dmy) rat is a unique mutant exhibiting severe myelin breakdown in the central nervous system (CNS). In this study, we conducted immunohistochemical and morphometrical investigations in the dmy rat. From around 6 weeks of age, the affected rats developed ataxia especially in the hindlimbs. Afterwards, ataxia worsened rapidly, resulting in complete paralysis of the hindlimbs and recumbency. Histopathology at 7 to 10 weeks of age revealed myelin destruction throughout the white matter of the CNS in the dmy rats. The most severely affected lesions were distributed in the corpus callosum, capsula interna, striatum, subcortical white matter, cerebellar peduncle, and ventral and lateral parts of the spinal cord. Immunohistochemistry demonstrated prominent astrogliosis and many ED-1 positive macrophages in the myelin-destructed areas. Until the 4th week, no significant differences in myelin thickness and fiber diameter were found between dmy and control rats. However, from 5 weeks of age, myelin thickness of residual myelinated fibers in dmy rats became significantly less than that in controls. These data indicated that the dmy phenotype shows a prolonged period of myelin destruction, suggesting that dmy mutation affects the adequate maintenance of myelin.

Topics: Age Factors; Animals; Axons; Ciliary Neurotrophic Factor; Demyelinating Diseases; Disease Models, Animal; Glial Fibrillary Acidic Protein; Histological Techniques; Immunohistochemistry; Microscopy, Electron, Transmission; Myelin Basic Protein; Myelin Sheath; Rats; Rats, Mutant Strains; Rats, Sprague-Dawley; Spinal Cord

Postresuscitation cerebral hypothermia is consistently neuroprotective in experimental preparations; however, its effects on white matter injury are poorly understood. Using a model of reversible cerebral ischemia in unanesthetized near-term fetal sheep, we examined the effects of cerebral hypothermia (fetal extradural temperature reduced from 39.4 +/- 0.1 degrees C to between 30 and 33 degrees C), induced at different times after reperfusion and continued for 72 hours after ischemia, on injury in the parasagittal white matter 5 days after ischemia. Cooling started within 90 minutes of reperfusion was associated with a significant increase in bioactive oligodendrocytes in the intragyral white matter compared with sham cooling (41 +/- 20 vs 18 +/- 11 per field, P < 0.05), increased myelin basic protein density and reduced expression of activated caspase-3 (14 +/- 12 vs 91 +/- 51, P < 0.05). Reactive microglia were profoundly suppressed compared with sham cooling (4 +/- 6 vs 38 +/- 18 per field, P < 0.05) with no effect on numbers of astrocytes. When cooling was delayed until 5.5 hours after reperfusion there was no significant effect on loss of oligodendrocytes (24 +/- 12 per field). In conclusion, hypothermia can effectively protect white matter after ischemia, but only if initiated early after the insult. Protection was closely associated with reduced expression of both activated caspase-3 and of reactive microglia.

Topics: Animals; Caspase 3; Caspases; Cerebral Cortex; Demyelinating Diseases; Enzyme Activation; Female; Fetus; Hypothermia, Induced; Hypoxia-Ischemia, Brain; Immunohistochemistry; In Situ Hybridization; Microglia; Myelin Basic Protein; Myelin Proteolipid Protein; Oligodendroglia; Pregnancy; Proliferating Cell Nuclear Antigen; Reperfusion Injury; RNA, Messenger; Sheep; Time Factors

Demyelination is a hallmark of several human diseases, including multiple sclerosis. To understand better the process of demyelination and remyelination, we explored the use of an in vitro organotypic cerebellar slice culture system. Parasagittal slices of postnatal Day 10 (P10) rat cerebella cultured in vitro demonstrated significant myelination after 1 week in culture. Treatment of the cultures at 7 days in vitro (DIV) with the bioactive lipid lysolecithin (lysophosphatidylcholine) for 15-17 hr in vitro produced marked demyelination. This demyelination was observed by immunostaining for the myelin components myelin basic protein (MBP), myelin oligodendrocyte glycoprotein (MOG), and 2', 3'-cyclic nucleotide 3'-phosphodiesterase (CNPase). After a transient demyelinating insult with lysolecithin in vitro, the cultures recovered with oligodendrocyte differentiation recapitulating a normal time course; there was initially re-expression of CNPase and MBP during this recovery, and this was followed by MOG. In addition, there seemed to be some limited remyelination during the recovery phase. Lysolecithin thus induces demyelination in an in vitro organotypic cerebellar slice culture system, providing a model system for studying myelination, demyelination, and remyelination in vitro.

Topics: 2',3'-Cyclic Nucleotide 3'-Phosphodiesterase; Animals; Calbindins; Cerebellum; Culture Techniques; Demyelinating Diseases; Lysophosphatidylcholines; Myelin Basic Protein; Myelin Proteins; Myelin Sheath; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Nerve Tissue Proteins; Neurofilament Proteins; Neurons; Phosphoric Diester Hydrolases; Rats; Rats, Sprague-Dawley; Regeneration; S100 Calcium Binding Protein G

The adult CNS has the capacity to remyelinate following metabolic, toxic and autoimmune demyelinating insults. In cuprizone-induced demyelination, spontaneous remyelination occurs after the cessation of cuprizone diet. We used the cuprizone model to investigate the role of glial K(+) channels in oligodendroglial (OLG) regeneration and remyelination in vivo. We found that treatment with 4-aminopyridine (4-AP), a broad-spectrum K(+) channel antagonist, results in: (1) decreased number of oligodendroglial progenitors (OP) and OLGs; (2) diminished astrogliosis; and (3) decreased remyelination in the corpus callosum based on the immunoreactivity to myelin basic protein (MBP), Rip monoclonal antibody, and by electron microscopy. Our findings support the concept that glial K(+) channels play an important role during OLG regeneration and remyelination, a crucial factor to be considered during the development of therapeutic strategies to facilitate recovery in demyelinating diseases and spinal cord injury.

Topics: 4-Aminopyridine; Animals; Astrocytes; Chelating Agents; Corpus Callosum; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Gliosis; Male; Mice; Mice, Inbred C57BL; Microscopy, Electron, Transmission; Myelin Basic Protein; Myelin Sheath; Nerve Regeneration; Neuroglia; Oligodendroglia; Potassium Channel Blockers; Potassium Channels; Stem Cells

Periventricular leukomalacia (PVL), the dominant form of brain injury in premature infants, is characterized by diffuse white matter injury and is associated with cerebral palsy (CP). Maternal and placental infections are major causes of prematurity and identifiable etiology of PVL and CP. Here we have evaluated the therapeutic efficacy of N-acetylcysteine (NAC), a potent antioxidant and precursor of glutathione, to attenuate lipopolysaccharide (LPS)-induced white matter injury and hypomyelination in the developing rat brain, an animal model of PVL. Intraperitoneal pretreatment of pregnant female rats with NAC (50 mg/kg), 2 hr prior to administration of LPS at embryonic day 18 (E18), attenuated the LPS-induced expression of inflammatory cytokines such as tumor necrosis factor-alpha, interleukin-1beta, and inducible nitric oxide synthase in fetal rat brains. There were significantly reduced numbers of TUNEL(+) nuclei coimmunostained for platelet-derived growth factor-alphaR(+) [a surface marker for oligodendrocyte progenitor cells (OPCs)] at E20 in the subventricular zone of fetal rat brain in the NAC + LPS group compared with the untreated LPS group. Interestingly, immunostaining for O4 and O1 as markers for late OPCs and immature oligodendrocytes demonstrated fewer O4(+) and O1(+) cells in the LPS group compared with the NAC + LPS and control groups. Consistent with O4(+)/O1(+) cell counts, the expression of myelin proteins such as myelin basic protein, proteolipid protein, and 2'3'-cyclic nucleotide phosphodiesterase, including transcription factors such as MyT1 and Gtx, was less in the LPS group at late postnatal days, indicating severe hypomyelination in the developing rat brain when compared with NAC + LPS and control groups. Collectively, these data support the hypothesis that NAC may provide neuroprotection and attenuate the degeneration of OPCs against LPS evoked inflammatory response and white matter injury in developing rat brain. Moreover, these data suggest the possible use of NAC as a treatment for pregnant women with maternal or placental infection as a means of minimizing the risk of PVL and CP.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Acetylcysteine; Age Factors; Analysis of Variance; Animals; Animals, Newborn; Antigens; CD11b Antigen; Cell Count; Cell Death; Cytokines; Demyelinating Diseases; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Interactions; Embryo, Mammalian; Female; Humans; Immunohistochemistry; In Situ Nick-End Labeling; Infant, Newborn; Leukomalacia, Periventricular; Lipopolysaccharides; Male; Myelin Basic Protein; Nerve Degeneration; Neuroprotective Agents; O Antigens; Oligodendroglia; Pregnancy; Proteoglycans; Rats; Rats, Sprague-Dawley; Receptor, Platelet-Derived Growth Factor alpha; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Stem Cells; Survival Rate; Time Factors; Transcription Factors

By using an in vitro model of antibody-mediated demyelination, we investigated the relationship between tumor necrosis factor-alpha (TNF-alpha) and heat shock protein (HSP) induction with respect to oligodendrocyte survival. Differentiated aggregate cultures of rat telencephalon were subjected to demyelination by exposure to antibodies against myelin oligodendrocyte glycoprotein (MOG) and complement. Cultures were analyzed 48 hr after exposure. Myelin basic protein (MBP) expression was greatly decreased, but no evidence was found for either necrosis or apoptosis. TNF-alpha was significantly up-regulated. It was localized predominantly in neurons and to a lesser extent in astrocytes and oligodendrocytes, and it was not detectable in microglial cells. Among the different HSPs examined, HSP32 and alphaB-crystallin were up-regulated; they may confer protection from oxidative stress and from apoptotic death, respectively. These results suggest that TNF-alpha, often regarded as a promoter of oligodendroglial death, could alternatively mediate a protective pathway through alphaB-crystallin up-regulation.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Antibodies; Apoptosis; Blotting, Western; Cells, Cultured; Complement System Proteins; Crystallins; Demyelinating Diseases; Disease Models, Animal; Drug Interactions; Embryo, Mammalian; Glial Fibrillary Acidic Protein; Heme Oxygenase (Decyclizing); Heme Oxygenase-1; Immunohistochemistry; In Situ Hybridization; Myelin Basic Protein; Myelin Proteins; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Necrosis; Neurons; Oligodendroglia; Rats; Telencephalon; Tumor Necrosis Factor-alpha; Up-Regulation

In human central nervous system (CNS) demyelinating diseases, spontaneous remyelination is often incomplete. Therefore, we have tested whether neutrotrophin-3 (NT-3) accelerates CNS myelin repair after a chemically-induced demyelination. One group of adult rats was injected in the corpus callosum (CC) with 1 microl of 1% lysophosphatidylcholine (LPC) and 1 microl of NT-3 (1 microg/microl), and 15 days after injury (D15) remyelination was compared to control rats (receiving 1 microl of LPC+1 microl of vehicle buffer of NT-3). The demyelinated volume decreased by 56% in NT-3-treated rats at D15, and immunohistochemistry showed an increase in mature MBP(+) oligodendrocytes (OL) (+66%) in treated animals (whereas less mature (CNP(+)) OL were unchanged). Since less than 3% axons degenerate in this model, and as astrocytic gliosis was not modified, these data suggest that NT-3 acts directly on cells of the OL lineage to enhance remyelination in vivo.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Bromodeoxyuridine; Central Nervous System Diseases; Corpus Callosum; Demyelinating Diseases; Disease Models, Animal; Glial Fibrillary Acidic Protein; Immunohistochemistry; Lysophosphatidylcholines; Male; Myelin Basic Protein; Nerve Regeneration; Neurotrophin 3; Oligodendroglia; Rats; Rats, Wistar; Stem Cells; Time Factors

Myelin basic protein (MBP) is required for normal myelin compaction and is implicated in both experimental and human demyelinating diseases. In this study, as an initial step in defining the regulatory network controlling MBP transcription, we located and characterized the function of evolutionarily conserved regulatory sequences. Long-range human-mouse sequence comparison revealed over 1 kb of conserved noncoding MBP 5' flanking sequence distributed into four widely spaced modules ranging from 0.1 to 0.4 kb. We demonstrate first that a controlled strategy of transgenesis provides an effective means to assign and compare qualitative and quantitative in vivo regulatory programs. Using this strategy, single-copy reporter constructs, designed to evaluate the regulatory significance of modular and intermodular sequences, were introduced by homologous recombination into the mouse hprt (hypoxanthine-guanine phosphoribosyltransferase) locus. The proximal modules M1 and M2 confer comparatively low-level oligodendrocyte expression primarily limited to early postnatal development, whereas the upstream M3 confers high-level oligodendrocyte expression extending throughout maturity. Furthermore, constructs devoid of M3 fail to target expression to newly myelinating oligodendrocytes in the mature CNS. Mutation of putative Nkx6.2/Gtx sites within M3, although not eliminating oligodendrocyte targeting, significantly decreases transgene expression levels. High-level and continuous expression is conferred to myelinating or remyelinating Schwann cells by M4. In addition, when isolated from surrounding MBP sequences, M3 confers transient expression to Schwann cells elaborating myelin. These observations define the in vivo regulatory roles played by conserved noncoding MBP sequences and lead to a combinatorial model in which different regulatory modules are engaged during primary myelination, myelin maintenance, and remyelination.

Topics: Animals; Base Sequence; Cell Differentiation; Cells, Cultured; Cholera Toxin; Conserved Sequence; Demyelinating Diseases; Female; Gene Expression; Gene Targeting; Genes, Regulator; Genes, Reporter; Humans; Hypoxanthine Phosphoribosyltransferase; Mice; Mice, Inbred C57BL; Mice, Transgenic; Molecular Sequence Data; Myelin Basic Protein; Neuroglia; Oligodendroglia; Phenotype; Plant Proteins; Ribosome Inactivating Proteins, Type 1; Saporins; Sequence Homology, Nucleic Acid

A distinct population of white matter progenitor cells (WMPCs), competent but not committed to generate oligodendrocytes, remains ubiquitous in the adult human subcortical white matter. These cells are present in both sexes and into senescence and may constitute as much as 4% of the cells of adult human capsular white matter. Transduction of adult human white matter dissociates with plasmids bearing early oligodendrocytic promoters driving fluorescent reporters permits the separation of these cells at high yield and purity, as does separation based on their expression of A2B5 immunoreactivity. Isolates of these cells survive xenograft to lysolecithin-demyelinated brain and migrate rapidly to infiltrate these lesions, without extending into normal white matter. Within several weeks, implanted progenitors mature as oligodendrocytes, and develop myelin-associated antigens. Lentiviral tagging with green fluorescent protein confirmed that A2B5-sorted progenitors develop myelin basic protein expression within regions of demyelination and that they fail to migrate when implanted into normal brain. Adult human white matter progenitor cells can thus disperse widely through regions of experimental demyelination and are able to differentiate as myelinating oligodendrocytes. This being the case, they may constitute appropriate vectors for cell-based remyelination strategies.

Topics: Adult; Age Factors; Animals; Brain Tissue Transplantation; Cell Differentiation; Cell Movement; Cell Survival; Demyelinating Diseases; Female; Gene Expression; Genetic Vectors; Graft Survival; Green Fluorescent Proteins; Humans; Immunomagnetic Separation; Indicators and Reagents; Lentivirus; Luminescent Proteins; Lysophosphatidylcholines; Male; Middle Aged; Myelin Basic Protein; Myelin Sheath; Neurons; Oligodendroglia; Rats; Stem Cell Transplantation; Stem Cells; Transplantation, Heterologous

The Long Evans shaker (les) rat is a myelin basic protein (MBP) mutant that exhibits severe central nervous system (CNS) dysmyelination. We used a combination of immunohistochemistry, immunoblot and Northern blot analyses to determine the effect of MBP deficits on the expression of other CNS myelin genes in this mutant. Immunohistochemistry revealed a marked reduction in all major myelin proteins and differences in their intracellular distribution. Immunoblots confirmed the decreased expression of these proteins and indicated that relative levels of proteolipid protein (PLP) and DM20 were altered in this mutant. Quantitation of mRNA levels indicated that decreases in PLP and DM20 were a result of changes in mRNA levels but detected no change in other myelin gene transcripts.

Topics: Animals; Demyelinating Diseases; Gene Expression Regulation; Mutation; Myelin Basic Protein; Myelin Proteins; Myelin Sheath; Oligodendroglia; Rats; Rats, Long-Evans; Rats, Mutant Strains; RNA, Messenger

Relative to the gray matter, there is a paucity of information regarding white matter biochemical alterations and their contribution to Alzheimer's disease (AD). Biochemical analyses of AD white matter combining size-exclusion, normal phase, and gas chromatography, immunoassays, and Western blotting revealed increased quantities of Abeta40 and Abeta42 in AD white matter accompanied by significant decreases in the amounts of myelin basic protein, myelin proteolipid protein, and 2',3'-cyclic nucleotide 3'-phosphodiesterase. In addition, the AD white matter cholesterol levels were significantly decreased while total fatty acid content was increased. In some instances, these white matter biochemical alterations were correlated with patient apolipoprotein E genotype, Braak stage, and gender. Our observations suggest that extensive white matter axonal demyelination underlies Alzheimer's pathology, resulting in loss of capacitance and serious disturbances in nerve conduction, severely damaging brain function. These white matter alterations undoubtedly contribute to AD pathogenesis and may represent the combined effects of neuronal degeneration, microgliosis, oligodendrocyte injury, microcirculatory disease, and interstitial fluid stasis. To accurately assess the success of future therapeutic interventions, it is necessary to have a complete appreciation of the full scope and extent of AD pathology.

Topics: 2',3'-Cyclic Nucleotide 3'-Phosphodiesterase; Aged; Aged, 80 and over; Aging; Alzheimer Disease; Amyloid beta-Peptides; Cerebral Cortex; Cholesterol; Demyelinating Diseases; Fatty Acids; Female; Humans; Male; Middle Aged; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin Sheath; Oligodendroglia; Peptide Fragments; Phosphoric Diester Hydrolases

Pigmentary type of orthochromatic leukodystrophy (POLD) is an adult-onset leukodystrophy, characterized pathologically by the presence of glial and microglial cytoplasmic pigment inclusions. The complete phenotype, genotype and pathogenetic mechanisms in POLD have not been elucidated. We followed for 18 years a woman with autopsy-proven POLD, who presented with 'frontal' dementia and spasticity. Her further course was marked by progressive mutism, apraxia and seizures. Her sister had died of the same disease after a much more rapidly progressing course. These sisters had primary infertility with pathologic evidence of streak ovaries. Diagnosis was confirmed in both cases by post-mortem examination. POLD is a rare cause of adult-onset leukodystrophy presenting with dementia. Ovarian dysgenesis is extremely rare in the absence of demonstrable chromosomal abnormalities and extends the clinical spectrum of POLD.

Topics: Adult; Brain; Brain Diseases; Demyelinating Diseases; Female; Humans; Immunohistochemistry; Microscopy, Electron; Middle Aged; Myelin Basic Protein; Myelin Proteolipid Protein; Ovary; Pedigree

Quaking viable (qk(v)) mice fail to properly compact myelin in their central nervous systems. Although the defect in the qk(v) mice involves a mutation affecting the expression of the alternatively spliced qk gene products, their roles in myelination are unknown. We show that the QKI RNA binding proteins regulate the nuclear export of MBP mRNAs. Disruption of the QKI nucleocytoplasmic equilibrium in oligodendrocytes results in nuclear and perikaryal retention of the MBP mRNAs and lack of export to cytoplasmic processes, as it occurs in qk(v) mice. MBP mRNA export defect leads to a reduction in the MBP levels and their improper cellular targeting to the periphery. Our findings suggest that QKI participates in myelination by regulating the mRNA export of key protein components.

Topics: 3' Untranslated Regions; Active Transport, Cell Nucleus; Alternative Splicing; Animals; Binding Sites; Brain; Cells, Cultured; Demyelinating Diseases; Exons; Humans; Mice; Mice, Quaking; Myelin Basic Protein; Oligodendroglia; Point Mutation; Protein Binding; Protein Isoforms; Rats; Recombinant Fusion Proteins; RNA-Binding Proteins; RNA, Messenger

Cerebral white matter (WM) lesions are observed frequently in human ischemic cerebrovascular disease and have been thought to contribute to cognitive impairment. This type of lesion can be experimentally induced in rat brains under chronic cerebral hypoperfusion by the permanent occlusion of both common carotid arteries. However, it remains uncertain whether chronic ischemia can damage both the gray and white matter, and whether it can induce demyelination with or without axonal damage. Therefore, we examined axonal damage using immunohistochemistry for the amyloid beta/A4 precursor protein (APP), chromogranin A (CgA) and demyelination using immunohistochemistry for the encephalitogenic peptide (EP) in this model. Severe WM lesions such as vacuolation and the loss of nerve fibers appeared in the optic nerve and optic tract after 3 days of ligation, and less intense changes were observed in the corpus callosum, internal capsule, and fiber bundles of the caudoputamen after 7 days with Klüver-Barrera and Bielschowsky staining. These WM lesions persisted even after 30 days. The APP, CgA, and EP-immunopositive fibers increased in number from 1 to 30 days after the ligation in the following WM regions: the optic nerve, optic tract, corpus callosum, internal capsule, and fiber bundles of the caudoputamen. In contrast, only a few APP, CgA, or EP-immunopositive fibers were detected in the gray matter regions, including the cerebral cortex and hippocampus. These results indicate that the WM is more susceptible to chronic cerebral hypoperfusion than the gray matter, with an involvement of both axonal and myelin components. Furthermore, immunohistochemistry for APP, CgA, and EP is far superior to routine histological staining in sensitivity and may become a useful tool to investigate WM lesions caused by various pathoetiologies.

Topics: Amyloid beta-Protein Precursor; Animals; Axons; Brain Ischemia; Cerebral Infarction; Chromogranin A; Chromogranins; Chronic Disease; Demyelinating Diseases; Immunohistochemistry; Male; Myelin Basic Protein; Nerve Fibers, Myelinated; Peptide Fragments; Prosencephalon; Rats; Rats, Wistar; Wallerian Degeneration

Multiple sclerosis (MS) is characterized by inflammation within the CNS. This inflammatory response is associated with production of nitric oxide (NO) and NO-related species that nitrosylate thiols. We postulated that MS patients would exhibit an antibody (Ab) response directed against proteins containing S-nitrosocysteine (SNO-cysteine) and showed that anti-NO-cysteine Abs of the IgM isotype are in fact present in the sera of some MS patients (Boullerne et al., 1995). We report here the presence of a seemingly identical Ab response directed against SNO-cysteine in an acute model of MS, experimental autoimmune encephalomyelitis (EAE) induced in Lewis rats with the 68-84 peptide of guinea pig myelin basic protein (MBP(68-84)). Serum levels of anti-SNO-cysteine Abs peaked 1 week before the onset of clinical signs and well before the appearance of anti-MBP(68-84) Abs. The anti-SNO-cysteine Ab peak titer correlated with the extent of subsequent CNS demyelination, suggesting a link between Ab level and CNS lesion formation. In relapsing-remitting MS patients, we found elevated anti-SNO-cysteine Ab at times of relapse and normal values in most patients judged to be in remission. Two-thirds of patients with secondary progressive MS had elevated anti-SNO-cysteine Ab levels, including those receiving interferon beta-1b. The data show that a rise in circulating anti-SNO-cysteine Ab levels precedes onset of EAE. Anti-SNO-cysteine Abs are also elevated at times of MS attacks and in progressive disease, suggesting a possible role for these Abs, measurable in blood, as a biological marker for clinical activity.

Topics: Animals; Antibody Specificity; Autoantibodies; Biomarkers; Cysteine; Demyelinating Diseases; Disease Models, Animal; Disease Progression; Encephalomyelitis, Autoimmune, Experimental; Female; Humans; Immunoglobulin M; Multiple Sclerosis; Myelin Basic Protein; Nitroso Compounds; Peptide Fragments; Predictive Value of Tests; Rats; Rats, Inbred Lew; Recurrence; Remission, Spontaneous; S-Nitrosothiols; Serum Albumin, Bovine; Spinal Cord

Cell cultures prepared from oligodendrocytes directly obtained from adult rat brain are composed of mature cells that lose their cell processes and myelin membrane during their isolation and therefore represent a very useful model to investigate the factors that could stimulate their recovery. We have observed that mature oligodendrocytes isolated from adult animals remain as round cells that lack processes for the first 3-4 days in culture. At the end of this lag period, however, the majority of the adult oligodendrocytes show a remarkable recovery, rapidly growing complex and extensive cell processes. Interestingly, the end of this lag period is accompanied by a dramatic upregulation in the expression of thyroid hormone (T(3)) receptor (TR). The functional importance of this increase in TR levels is supported by the observation that the majority of the cells cultured in the presence of T(3) show significantly more extensive and complex process outgrowth than the control cells in cultures lacking this hormone. In addition, this reactivation of the adult cells was also preceded by an increased expression of glucocorticoid receptor (GR) and cyclic AMP-response element binding protein (CREB), two transcription factors that together with TR appear to play important roles in the control of neonatal oligodendrocyte development. Thus, it is possible to hypothesize that upregulation of these proteins may be part of the metabolic changes that occur during the lag period required for recovery of the adult oligodendrocytes. These observations raise the question of whether these transcription factors may play any significant role during remyelination after demyelinating lesions of adult CNS.

Topics: Animals; Animals, Newborn; Blotting, Western; Cell Differentiation; Cells, Cultured; Central Nervous System; Cyclic AMP Response Element-Binding Protein; Demyelinating Diseases; Female; Gene Expression Regulation; Immunohistochemistry; Myelin Basic Protein; Nerve Regeneration; Oligodendroglia; Rats; Rats, Sprague-Dawley; Receptors, Glucocorticoid; Receptors, Thyroid Hormone; Recovery of Function; Time Factors; Transcription Factors; Triiodothyronine; Up-Regulation

To study the biological role of the chemokine ligands CCL19 and CCL21, we generated transgenic mice expressing either gene in oligodendrocytes of the CNS. While all transgenic mice expressing CCL19 in the CNS developed normally, most (18 of 26) of the CCL21 founder mice developed a neurological disease that was characterized by loss of landing reflex, tremor, and ataxia. These neurological signs were observed as early as postnatal day 9 and were associated with weight loss and death during the first 4 wk of life. Microscopic examination of the brain and spinal cord of CCL21 transgenic mice revealed scattered leukocytic infiltrates that consisted primarily of neutrophils and eosinophils. Additional findings included hypomyelination, spongiform myelinopathy with evidence of myelin breakdown, and reactive gliosis. Thus, ectopic expression of the CC chemokine CCL21, but not CCL19, induced a significant inflammatory response in the CNS. However, neither chemokine was sufficient to recruit lymphocytes into the CNS. These observations are in striking contrast to the reported activities of these molecules in vitro and may indicate specific requirements for their biological activity in vivo.

Topics: Animals; Brain; Cell Movement; Central Nervous System Diseases; Cerebellum; Chemokine CCL19; Chemokine CCL21; Chemokines; Chemokines, CC; Cytokines; Demyelinating Diseases; Gliosis; Leukocytes; Medulla Oblongata; Mice; Mice, Inbred C57BL; Mice, Inbred DBA; Mice, Transgenic; Myelin Basic Protein; Neurodegenerative Diseases; Oligodendroglia; Phenotype; Spinal Cord

C4d-immunoreactive complement-activated oligodendrocytes (C4d-CAOs) have been described in several neurodegenerative diseases but have not been studied in multiple sclerosis (MS). Here we report that such CAOs delineate miniature MS plaques of 300-500 mum diameter. They are devoid of myelin and are surrounded by a rim of activated microglia intermingled with the C4d-CAOs. Although C4d-immunostained periaxonal oligodendroglial processes are often swollen, the axons of passage appear undamaged and extend through the demyelinated plaque area. No immunostaining with other components of the complement cascade (C1q-C9) was observed in association with these miniature plaques. However, in large MS lesions, C1q-C9 immunoreactive fibers were present, indicating complete activation of the complement cascade in these more developed lesions. It is possible that the miniature plaques, bordered by C4d-CAOs, represent the earliest stage of plaque development, preceding even the larger, transient plaques frequently observed in serial MRI studies. The association of CAOs with miniature areas of demyelination suggests a direct attack on oligodendroglial cells by the early complement components as an initiating event in MS. Incomplete complement activation indicates that this step may be reversible, whereas full and persistent activation as observed in large MS lesions may lead to death of oligodendroglia with permanent axonal damage.

Topics: Aged; Aged, 80 and over; Cerebral Cortex; Complement C4; Complement C4b; Complement System Proteins; Demyelinating Diseases; HLA-DR Antigens; Humans; Immunohistochemistry; Microglia; Middle Aged; Multiple Sclerosis; Myelin Basic Protein; Oligodendroglia; Peptide Fragments; Supranuclear Palsy, Progressive

Theiler's murine encephalomyelitis virus-induced demyelinating disease (TMEV-IDD), a multiple sclerosis (MS) model, is a central nervous system (CNS) demyelinating disease characterized by early peripheral T cell responses to virus epitopes which spreads to myelin epitopes during chronic disease. We show that CD4(+) T cells isolated from the spinal cords of chronically infected SJL mice proliferate and secrete pro-inflammatory cytokines upon in vitro challenge with both TMEV epitopes and proteolipid protein (PLP(139-151)). Importantly, myelin-specific tolerance induced by intravenous administration of MP4, a fusion of the myelin proteins myelin basic protein (MBP) and PLP, to SJL mice with ongoing TMEV-IDD attenuated disease progression and resulted in significantly less demyelination and decreased inflammatory cell infiltration in the CNS. Paradoxically, peptide-specific splenic T cell proliferative and IFN-gamma responses were enhanced in the tolerized mice. Collectively, these results indicate that myelin-specific T cell responses contribute to chronic disease progression in this virus-induced model of MS, and suggest caution in the use of antigen-specific tolerance for treatment of ongoing autoimmune disease.

Topics: Amino Acid Sequence; Animals; Cardiovirus Infections; Cytokines; Demyelinating Diseases; Epitopes; Female; Hypersensitivity, Delayed; Immune Tolerance; Mice; Molecular Sequence Data; Multiple Sclerosis; Myelin Basic Protein; Myelin Proteolipid Protein; Myelin Sheath; Peptide Fragments; Recombinant Fusion Proteins; T-Lymphocytes; Theilovirus

Proteomic diversity is frequently achieved by alternative RNA-splicing events that can be fine-tuned in tissue-specific and developmentally regulated ways. Understanding this type of genetic regulation is compelling because of the extensive complexity of alternative splicing found in the nervous system. quaking (qk), one of the classical mouse dysmyelination mutants, is defective for the expression of myelin-associated glycoprotein (MAG), and the misregulation of MAG pre-mRNA alternative splicing is implicated as a causal factor. The qk locus encodes several RNA-binding proteins with heterogeneous nuclear ribonucleoprotein K-type homology, a characteristic of several known alternative splicing regulators. Here we test the nuclear-localized qk isoform (QKI-5) for its ability to regulate alternative splicing of MAG pre-mRNA in transient coexpression assays. QKI-5 exhibits properties of a negative regulator of MAG exon 12 alternative splicing. An intronic sequence element required for the repressive function and binding of QKI-5 is also identified. Direct evidence for irregularities in alternative splicing of MAG and other myelin protein transcripts in the qk mouse is demonstrated.

Topics: Alternative Splicing; Animals; Base Sequence; COS Cells; Demyelinating Diseases; Mice; Mice, Quaking; Molecular Sequence Data; Myelin Basic Protein; Myelin Proteolipid Protein; Myelin Sheath; Myelin-Associated Glycoprotein; RNA Precursors; RNA-Binding Proteins

Glial cells from neonatal MbetaP5 transgenic mice, which express bacterial beta-galactosidase (lacZ) under control of the myelin basic protein (MBP) promoter (Gow et al, 1992), were transplanted into the spinal cord or cerebral hemisphere of immunosuppressed normal and myelin-deficient (md) rats in order to assess the ability of the donor cells to survive, migrate, and differentiate within normal compared with myelin-deficient central nervous system (CNS). LacZ+ cells were detected as early as 6-7 days after transplantation into the low thoracic cord and by 10 days had spread rostrally to the brainstem and caudally to the sacral spinal cord. Initially, compact lacZ+ cells, lacking processes, were found associated with small blood vessels and with the glia limitans. Cells of this type persisted throughout the experiment. Later, lacZ+ cells with processes were seen along fiber tracts in the dorsal columns and, after intracerebral injection, subjacent to ventricular ependyma, as well as scattered in cerebral white and gray parenchyma. The extent of spread was comparable in md and normal rats, but in the md group, the success rate was higher, and more cells differentiated into process-bearing oligodendrocytes. Acceptance of xenografts in immunosuppressed recipients equaled that of allografts. The overall spread of grafted cells exceeded that of injected charcoal, indicating active migration. In contrast to earlier studies that identified oligodendrocytes based on morphology alone, this study has allowed us to identify and track oligodendrocytes based on myelin gene expression. We show some oligodendrocytes whose morphology is consistent with classical morphological descriptions, some that resemble astrocytes, and a class of compact perivascular oligodendrocyte-lineage cells that we suggest are migratory.

Topics: Animals; Brain; Brain Tissue Transplantation; Cell Differentiation; Cell Lineage; Cell Movement; Cell Size; Cerebral Cortex; Demyelinating Diseases; Female; Genes, Reporter; Graft Survival; Male; Mice; Mice, Transgenic; Myelin Basic Protein; Myelin Sheath; Oligodendroglia; Rats; Rats, Mutant Strains; Spinal Cord; Stem Cell Transplantation; Stem Cells

The age-associated decrease in the efficiency of CNS remyelination has clear implications for recovery from demyelinating diseases such as multiple sclerosis (MS) that may last for several decades. Developing strategies to reverse the age-associated decline requires the identification of how the regenerative process is impaired. We addressed whether remyelination becomes slower because of an impairment of recruitment of oligodendrocyte progenitors (OPs) or, as is the case in some MS lesions, an impairment of OP differentiation into remyelinating oligodendrocytes. The OP response during remyelination of focal, toxin-induced CNS demyelination in young and old rats was compared by in situ hybridization using probes to two OP-expressed mRNA species: platelet-derived growth factor-alpha receptor and the OP transcription factor myelin transcription factor 1 (MyT1). We found that the expression patterns for the two OP markers are very similar and reveal a delay in the colonization of the demyelinated focus with OPs in the old animals compared with the young animals. By comparing the mRNA expression pattern of MyT1 with that of the myelin proteins myelin basic protein and Gtx, we found that in the old animals there is also a delay in OP differentiation that increases with longer survival times. These results indicate that the age-associated decrease in remyelination efficiency occurs because of an impairment of OP recruitment and the subsequent differentiation of the OPs into remyelinating oligodendrocytes, and that strategies aimed at ameliorating the age-associated decline in remyelination efficiency will therefore need to promote both components of the regenerative process.

Topics: Age Factors; Aging; Animals; Cell Differentiation; Cell Division; Cerebellum; Demyelinating Diseases; Ethidium; Female; Homeodomain Proteins; In Situ Hybridization; Myelin Basic Protein; Nerve Fibers, Myelinated; Oligodendroglia; Protein Serine-Threonine Kinases; Protein-Tyrosine Kinases; Rats; Receptor, Platelet-Derived Growth Factor alpha; Regeneration; RNA, Messenger; Stem Cells; Time Factors; Transcription Factors

Neurological deficit in experimental allergic encephalomyelitis (EAE) and multiple sclerosis (MS) is probably a consequence of synergy between T and B cell responses to CNS antigens. During the demyelinating phase of chronic relapsing EAE in ABH mice, anti-myelin oligodendrocyte glycoprotein (MOG) responses were increased compared to the inflammatory acute phase, but such levels did not correlate with the severity of clinical disease. The pathogenicity of antibodies (Ab) to MOG, myelin basic protein (MBP), proteolipid protein (PLP) and galactocerebroside (GalC) was investigated in vivo following injection at the onset of EAE. An IgG2a monoclonal Ab (mAb), clone Z12, directed to MOG augmented clinical disease and demyelination in ABH and C57BL/6 mice, but not MOG knock-out mice. No effect was observed with F(ab(2))' fragments of Z12 or with the anti-MOG IgG1 mAbs, clones Y10 or 8-18C5. Cobra venom factor partially reduced the augmenting effect of mAb Z12 suggesting a role for complement. The pathogenic effect of anti-myelin Abs was not restricted to MOG since an anti-GalC mAb exacerbated inflammation in the CNS while an MBP mAb (clone 22) reduced clinical disease. Taken together, these data provide further evidence that myelin-reactive Abs generated during autoimmune neurological disease may play an important role not only in the pathogenesis of disease but also the regulation of myelin-targeted autoimmune disease.

Topics: Animals; Autoantibodies; Complement System Proteins; Demyelinating Diseases; Disease Models, Animal; Elapid Venoms; Encephalomyelitis, Autoimmune, Experimental; Female; Galactosylceramides; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Recurrence; Spinal Cord

Experimental allergic encephalomyelitis (EAE) is a CD4(+) Th1 cell-mediated inflammatory demyelinating autoimmune disease of the CNS that serves as an animal model for multiple sclerosis (MS). IL-12 is a proinflammatory cytokine that plays a crucial role in the induction of neural Ag-specific Th1 differentiation and pathogenesis of CNS demyelination in EAE and MS. Curcumin (1,7-Bis(4-hydroxy-3-methoxyphenyl)-1,6-heptadiene-3,5-dione) is a naturally occurring polyphenolic phytochemical isolated from the rhizome of the medicinal plant Curcuma longa. It has profound anti-inflammatory activity and been traditionally used to treat inflammatory disorders. In this study we have examined the effect and mechanism of action of curcumin on the pathogenesis of CNS demyelination in EAE. In vivo treatment of SJL/J mice with curcumin significantly reduced the duration and clinical severity of active immunization and adoptive transfer EAE. Curcumin inhibited EAE in association with a decrease in IL-12 production from macrophage/microglial cells and differentiation of neural Ag-specific Th1 cells. In vitro treatment of activated T cells with curcumin inhibited IL-12-induced tyrosine phosphorylation of Janus kinase 2, tyrosine kinase 2, and STAT3 and STAT4 transcription factors. The inhibition of Janus kinase-STAT pathway by curcumin resulted in a decrease in IL-12-induced T cell proliferation and Th1 differentiation. These findings highlight the fact that curcumin inhibits EAE by blocking IL-12 signaling in T cells and suggest its use in the treatment of MS and other Th1 cell-mediated inflammatory diseases.

Topics: Adoptive Transfer; Animals; Anti-Inflammatory Agents, Non-Steroidal; Cells, Cultured; Central Nervous System; Curcumin; Demyelinating Diseases; DNA-Binding Proteins; Encephalomyelitis, Autoimmune, Experimental; Epitopes, T-Lymphocyte; Female; Immunosuppressive Agents; Injections, Intraperitoneal; Interleukin-12; Janus Kinase 1; Macrophages; Mice; Mice, Inbred Strains; Microglia; Myelin Basic Protein; Phosphorylation; Protein-Tyrosine Kinases; Signal Transduction; Spleen; STAT3 Transcription Factor; STAT4 Transcription Factor; T-Lymphocytes; Trans-Activators; Tyrosine; Vaccination

In earlier studies we demonstrated that an increase in the relative amounts of citrullinated myelin basic protein (MBP) was found in multiple sclerosis (Moscarello et al. 1994). To determine the temporal relationship between the citrullinated MBP and peptidylarginine deiminase (PAD), the enzyme responsible for deiminating arginyl residues in proteins, we studied enzyme activity, enzyme protein, PAD mRNA in a spontaneously demyelinating transgenic mouse model and we correlated the amount of PAD with citrullinated MBP. Both PAD protein as measured in an immunoslot blot method and PAD RNA were elevated. In fractionation studies we showed that the increase in PAD enzyme was due to an increase in the PAD found in membrane fractions and not the soluble PAD (PADII). From our data we concluded that up-regulation of myelin-associated PAD was responsible for the increase in citrullinated MBP in our transgenic mice prior to onset of clinical or pathological signs of demyelination. We postulate that a similar mechanism may be responsible for the increase in citrullinated MBP in multiple sclerosis.

Topics: Age of Onset; Animals; Brain; Brain Chemistry; Citrulline; Demyelinating Diseases; Disease Models, Animal; Disease Progression; Enzyme Activation; Gene Dosage; Hydrolases; Mice; Mice, Transgenic; Myelin Basic Protein; Myelin Sheath; Protein-Arginine Deiminase Type 4; Protein-Arginine Deiminases; RNA, Messenger; Solubility; Transgenes; Up-Regulation

Exposure of mice to the copper chelator, cuprizone, results in CNS demyelination. There is remyelination after removal of the metabolic insult. We present brain regional studies identifying corpus callosum as particularly severely affected; 65% of cerebroside is lost after 6 weeks of exposure. We examined recovery of cerebroside and ability to synthesize cerebroside and cholesterol following removal of the toxicant. The temporal pattern for concentration of myelin basic protein resembled that of cerebroside. We applied Affymetrix GeneChip technology to corpus callosum to identify temporal changes in levels of mRNAs during demyelination and remyelination. Genes coding for myelin structural components were greatly down-regulated during demyelination and up-regulated during remyelination. Genes related to microglia/macrophages appeared in a time-course (peaking at 6 weeks) correlating with phagocytosis of myelin and repair of lesions. mRNAs coding for many cytokines had peak expression at 4 weeks, compatible with intercellular signaling roles. Of interest were other genes with temporal patterns correlating with one of the three above patterns, but of function not obviously related to demyelination/remyelination. The ability to correlate gene expression with known pathophysiological events should help in elucidating further function of such genes as related to demyelination/remyelination.

Topics: Animals; Biomarkers; Brain; Brain Stem; Cerebellum; Cerebrosides; Chelating Agents; Cholesterol; Corpus Callosum; Cuprizone; Demyelinating Diseases; Disease Models, Animal; Disease Progression; Galactosyltransferases; Gene Expression Profiling; Gene Expression Regulation; Mice; Mice, Inbred C3H; Myelin Basic Protein; Myelin Sheath; N-Acylsphingosine Galactosyltransferase; Oligonucleotide Array Sequence Analysis; RNA, Messenger

Peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors of the nuclear hormone receptor superfamily that have been described as master genes that switch cells from an undifferentiated phenotype to a differentiated phenotype. In the present investigation, we examined the possibility that ligands for PPARs are potent activators of oligodendrocyte (OL) differentiation and/or proliferation. Primary glial cultures and enriched OL cultures of neonatal mouse cerebra were treated with three different PPAR agonists: a PPAR gamma-selective agonist, a PPAR delta-selective agonist, and a pan agonist selective for both PPAR gamma and delta. Treatment with PPAR gamma agonist does not have an effect on the differentiation of OLs; however, PPAR delta agonist and the pan agonist treatment accelerates the differentiation of OLs within 24 h of application in mixed glial cultures. The number of OLs with processes and huge membrane sheets increases two- to threefold in both groups. The increase in the size of the sheets is also mirrored by changes in the intensity and distribution of myelin basic protein (MBP) and proteolipid protein (PLP) mRNAs. As compared to controls, the PPAR delta agonist-treated groups contain more OLs that have MBP and PLP mRNA extending into distal processes. These results indicate that PPAR delta plays a significant role in the maturation of OLs and regulates the size of OL sheets. BrdU immunostaining reveals that these agonists do not significantly stimulate proliferation of OLs expressing glycolipids. The studies in enriched OL cultures reproduce the effects of the PPAR agonists seen in the mixed glial cultures, indicating that the effect of the PPAR agonists is directly on the OLs and not via astrocytes. In the enriched cultures, the total number of OLs increases significantly in the PPAR delta agonist-treated groups, but BrdU immunostaining does not show an increased proliferation of cells. These findings suggest that PPAR delta increases the survival of cells and/or prevents cell death in enriched cultures. Although PPAR delta is expressed in various cell types, its role as a factor in the transcriptional regulation of OL differentiation has not been explored. We show for the first time that a ligand that serves as an agonist for PPAR delta activates the program of OL differentiation in primary and enriched OL cultures.

Topics: Animals; Brain; Cell Count; Cell Differentiation; Cells, Cultured; Demyelinating Diseases; Gene Expression; Mice; Mice, Inbred CBA; Myelin Basic Protein; Myelin Proteolipid Protein; Oligodendroglia; Phenotype; Receptors, Cytoplasmic and Nuclear; RNA, Messenger; Transcription Factors

We analyzed the role of Fyn tyrosine kinase in CNS myelination by using fyn(-/-) null mutant mice, which express no Fyn protein. We found a severe myelin deficit in forebrain at all ages from 14 d to 1 year. The deficit was maximal at 1 month of age and was similar regardless of mouse strain background or whether it was determined by bulk isolation of myelin or by quantitation of myelin basic protein. To determine the cellular basis of the myelin deficit, we counted oligodendrocytes in tissue sections of mice expressing oligodendrocyte-targeted beta-galactosidase, and we used light and electron microscopy to examine the number and morphology of myelinated fibers and size of myelinated CNS structures. All of these parameters were reduced in fyn(-/-) mice. Unexpectedly, there were regional differences in the myelin deficit; in contrast to forebrain, fyn(-/-) cervical spinal cord exhibited no reduction in myelin content, number of oligodendrocytes, or number of myelinated fibers, nor was myelination delayed developmentally. We found that oligodendrocytes express Src, but there was no significant reduction of myelin content in null mutants lacking the Fyn-related kinases Src, Yes, or Lyn. Finally, we investigated the molecular features of Fyn that are required for myelination and found that a single amino acid substitution, which abolishes the tyrosine kinase activity of Fyn, resulted in a myelin deficit as great as that observed in the complete absence of Fyn protein. These results demonstrate that Fyn plays a unique role in myelination, one that requires its kinase activity.

Topics: Animals; Blotting, Western; Cell Count; Cells, Cultured; Central Nervous System; Corpus Callosum; Demyelinating Diseases; Genes, Reporter; Mice; Mice, Inbred C57BL; Mice, Knockout; Myelin Basic Protein; Myelin Sheath; Nerve Fibers, Myelinated; Prosencephalon; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-fyn; Proto-Oncogene Proteins c-yes; Spinal Cord; src-Family Kinases

The genetic lesion of quakingviable (qk(v)) causes diminished expression of the QKI RNA-binding protein in myelin producing cells. Consequently, several structural myelin proteins are severely reduced. Among these affected proteins, the reduction of the myelin basic protein (MBP) results from post-transcriptional abnormalities of the MBP mRNA, presumably due to the lack of interactions with QKI. However, whether this is the common mechanism for reduced expression of other myelin proteins in qk(v) dysmyelination remains unclear. Here we report that distinct molecular mechanisms underlie the reduction of MBP and the 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNP) in qk(v) dysmyelination. MBP transcripts bind QKI strongly and are markedly reduced in the qk(v)/qk(v) oligodendrocytes in which QKI is almost completely lost. In contrast, CNP transcripts bind QKI weakly and are only slightly affected by the lack of QKI. None the less, CNP proteins are severely reduced in the qk(v)/qk(v) brain. Since CNP transcripts are predominantly associated with translating polyribosomes, diminished CNP expression in qk(v) dysmyelination is unlikely to be due to translational failures, but more likely results from accelerated protein degradation.

Topics: 2',3'-Cyclic Nucleotide 3'-Phosphodiesterase; 2',3'-Cyclic-Nucleotide Phosphodiesterases; 3' Untranslated Regions; Animals; Binding Sites; Brain; Demyelinating Diseases; Male; Mice; Mice, Quaking; Myelin Basic Protein; Phosphoric Diester Hydrolases; Polyribosomes; Protein Processing, Post-Translational; RNA-Binding Proteins; RNA, Messenger

Bone marrow transplantation (BMT) has therapeutic value for twitcher (globoid cell leukodystrophy) mice, which suffer from a genetic deficiency of the lysosomal enzyme galactosylceramidase that leads to progressive demyelination and early death. Preliminary investigations indicated that a semiallogeneic BMT resulted in graft vs. host disease (GVHD) in twitcher mice but not normal mice. Increased production of the cytokine IL-6 has been demonstrated in twitcher mice, and it has been linked with induction of GVHD. We investigated the effects of BMT in twitcher/IL-6 deficient mice and compared these findings with those from transplanted twitcher and control mice. After a semiallogeneic BMT, 11.4% of controls died within few weeks while the rest survived >100 days without GVHD. In contrast, 85% of the transplanted twitcher mice died by 70 days and 65% developed clinical signs of GVHD, e.g., alopecia and weight loss. In transplanted twitcher/IL-6 deficient mice, only 21% died by Day 70, none had alopecia, and 23% had weight loss. There was no difference in the onset day and severity of twitching between twitcher and twitcher/IL-6 deficient mice after BMT. In transplanted twitcher/IL-6 deficient mice, there was improvement of BBB integrity and a decrease in globoid cell number compared with nontransplanted twitcher/IL-6 deficient mice. In summary, these results demonstrate that an underlying pathology like globoid cell leukodystrophy leads to activation of GVHD responses in a donor-host combination that would not normally induce GVHD. Furthermore, IL-6 seems to play a key role because a deficiency of IL-6 results in a better prognosis.

Topics: Animals; Astrocytes; beta-Galactosidase; Blood-Brain Barrier; Body Weight; Bone Marrow Transplantation; Brain; Demyelinating Diseases; Female; Gliosis; Graft vs Host Disease; Immunohistochemistry; Interleukin-6; Lectins; Leukodystrophy, Globoid Cell; Macrophages; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Neurologic Mutants; Minor Histocompatibility Antigens; Myelin Basic Protein; Serum Albumin; Survival Rate

In previous work we found that mezerein, a C kinase activator, as well as basic fibroblast growth factor (FGF-2) induce demyelination and partial oligodendrocyte dedifferentiation in highly differentiated aggregating brain cell cultures. Here we show that following protein kinase C activator-induced demyelination, effective remyelination occurs. We found that mezerein or FGF-2 caused a transient increase in DNA synthesis following a pronounced decrease of the myelin markers myelin basic protein and 2',3'-cyclic nucleotide 3'-phosphohydrolase. Both oligodendrocytes and astrocytes were involved in this mitogenic response. Within 17 days after demyelination, myelin was restored to the level of the untreated controls. Transient mitotic activity was indispensable for remyelination. The present results suggest that myelinating oligodendrocytes retain the capacity to reenter the cell cycle, and that this plasticity is important for the regeneration of the oligodendrocyte lineage and remyelination. Although it cannot be excluded that a quiescent population of oligodendrocyte precursor cells was present in the aggregates and able to proliferate, differentiate and remyelinate, we could not find evidence supporting this view.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Becaplermin; Biomarkers; Brain; Cell Division; Cytarabine; Demyelinating Diseases; Diterpenes; DNA; Enzyme Activation; Fetus; Fibroblast Growth Factor 2; In Vitro Techniques; Mitosis; Myelin Basic Protein; Myelin Sheath; Oligodendroglia; Platelet-Derived Growth Factor; Protein Kinase C; Proto-Oncogene Proteins c-sis; Rats; Terpenes

The 4e transgenic mouse is characterized by overexpression of the PLP gene. Heterozygous littermates containing three PLP gene copies develop and myelinate normally. However, a progressive CNS demyelination begins at 3-4 months of age. Despite focal demyelination, these animals survive for one year with hind limb paralysis. We used this CNS demyelination model to determine if grafts of CG4 oligodendrocyte progenitors would survive and myelinate the adult CNS. Either CG4 cells, or co-grafts of CG4/B 104 cells 11:1 ratio respectively) were performed. Grafted cells survived and migrated in the normal and transgenic brain. Non-treated transgenic animals revealed extensive lack of myelin. Three months post-transplant hosts with CG4 or co-transplants displayed a near normal myelin pattern. Double immunofluorescence for neurofilament and myelin basic protein revealed the presence of many naked axons in non-grafted transgenic animals. Those grafted with progenitor CG4 cells or cografts displayed a clear increase in remyelination. This data provides a new direction for the development of cell replacement therapies in demyelinating diseases.

Topics: Animals; Brain Diseases; Cell Line; Cell Movement; Cell Survival; Demyelinating Diseases; Fluorescent Antibody Technique, Direct; Immunohistochemistry; Mice; Mice, Transgenic; Myelin Basic Protein; Myelin Sheath; Oligodendroglia; RNA, Messenger; Stem Cell Transplantation; Stem Cells; Transferrin

An 80-year-old male without abnormal past medical history presented with coma, general seizures, and fever subsequent to abnormal behavior. The pressure of the cerebrospinal fluid(CSF) elevated(13.5-20.5 cm H2O), and CSF examination revealed pleocytosis with predominant mononuclear cells(80-879/mm3) and elevated protein level(32-130 mg/dl). DNAs of herpes simplex virus(HSV) type 1 and 2 in CSF were not confirmed by polymerase chain reaction method in the acute phase. The HSV(type 1) antibody(HSV-1 Ab) ratio of serum to CSF(= [serum HSV-1 Ab]/[CSF HSV-1 Ab]) was 0.98 and HSV-1 Ab index(= [CSF HSV-1 Ab]/[serum HSV-1 Ab] divided by [CSF albumin]/[serum albumin]) was 62.4. Initial fluid attenuated inversion recovery(FLAIR) (TR/TE/TI = 6,882/110/1,700 msec) axial magnetic resonance(MR) imaging showed hyperintensity in the subfrontal area, inferomedial portions of the temporal lobes, cingulate gyri, and insular cortices bilaterally. Meningoencephalitis caused by HSV-1 was diagnosed based on the values of HSV-1 Ab ratio of serum to CSF(less than 20), of HSV-1 Ab index(larger than 1.91), and the findings of MR imaging. Diffuse white matter lesions manifesting hyperintensity on FLAIR imaging in the bilateral frontal and temporal lobes close to the affected cortices developed approximately six weeks after the onset despite administration of antiviral agent and steroid. The lesion extensively involved the white matter of the bilateral frontal and temporal lobes finally. The initial value of myelin basic protein(MBP) in CSF was 0.9 ng/ml (normal value: less than 4 ng/ml). Subsequent measurement of MBP in CSF about two, six weeks, two, three, and six months after the onset showed a marked increase of 233.9 ng/ml followed by a gradual decrease of 25.4 ng/ml, 18.4 ng/ml, 7.4 ng/ml and 4.3 ng/ml, respectively. Therefore, demyelination of the lesion in the cerebral white matter was suggested by the chronological change in FLAIR imaging and MBP in CSF.

Topics: Aged; Aged, 80 and over; Cerebral Cortex; Demyelinating Diseases; Encephalitis, Herpes Simplex; Humans; Male; Myelin Basic Protein; Simplexvirus

We had demonstrated that platelet-derived growth factor (PDGF) enhanced the reconstruction of myelin-like membranes after their disruption by lysophosphatidylcholine (LPC) in vitro. To investigate its role in vivo, a demyelinating lesion of the corpus callosum was induced in adult Wistar rats by a stereotaxic microinjection of 1 microl LPC, then 63 pairs of rats received either 1 microg PDGF, or its vehicle buffer which were injected above LPC. The effects of PDGF were significant after 2 weeks: the number of oligodendrocytes (OL) expressing 2',3'-cyclic nucleotide 3'-phosphodiesterase in the lesion increased by 49%, mature OL labelled by in situ hybridization for myelin basic protein-mRNA increased by 27% (P<10(-2)), and the total volume of demyelination decreased by 60% compared to controls. The proliferation of cells of the OL lineage was also enhanced up to 67% by PDGF compared to LPC controls (P<2.5 x 10(-2)). Ultrastructural studies confirmed this dramatic improvement, and the ratio of remyelinated to demyelinated axons, determined at the maximal demyelination site, in the centre of the lesion, increased by 10-fold (P<2.5 x 10(-3)) in animals treated with PDGF. Remyelination was complete after 3 months for both treatments. Neither exacerbation of gliosis nor glial tumoural transformation were observed. Mechanisms through which PDGF improves remyelination could involve proliferation of OL progenitors, and/or of already differentiated surviving OLs, and a chemotactic effect, which had been identified in vitro.

Topics: Animals; Axons; Cell Division; Corpus Callosum; Demyelinating Diseases; Drug Administration Schedule; Immunohistochemistry; Lysophosphatidylcholines; Male; Microscopy, Electron; Myelin Basic Protein; Myelin Sheath; Nerve Regeneration; Oligodendroglia; Platelet-Derived Growth Factor; Rats; Rats, Wistar; Recovery of Function; RNA, Messenger; Stem Cells

The immune cytokine interferon-gamma (IFN-gamma) is believed to be a key agent in the pathogenesis of immune-mediated demyelinating disorders. We have examined the possibility that one effect of this cytokine involves overloading the endoplasmic reticulum (ER) of oligodendrocytes through the induction of major histocompatibility complex (MHC) class I heavy chain (HC) gene expression. For these studies, we have utilized several genetic mouse models that yield different subcellular localizations of HC in oligodendrocytes. We show that transgenic mice that ectopically express HC in oligodendrocytes (MBP/MHC class I mice) fail to transport HC past the ER. These mice are hypomyelinated and have a tremoring phenotype. When oligodendrocytes deficient in beta-2 microglobulin (beta2m), which is required for MHC class I assembly and transport, were treated with IFN-gamma in vitro, HC was transported past the ER to the trans-Golgi network but not onto the cell surface. When an asymptomatic line of mice that expresses MHC class I in the CNS due to transgene-derived IFN-gamma (MBP/IFN-gamma mice) was crossed onto the beta2m-/- background, the resulting mice were asymptomatic. In contrast, increasing the amount of MHC class I protein transported through the ER in MBP/MHC class I transgenic mice, by crossing them to the asymptomatic MBP/IFN-gamma mice, exacerbated their phenotype. Taken together, these data indicate that the ER is a sensitive site in oligodendrocytes for accumulation of MHC class I HC and suggest a molecular mechanism for IFN-gamma's deleterious effects on these cells.

Topics: Animals; Biological Transport; Blotting, Northern; Corpus Callosum; Demyelinating Diseases; Endoplasmic Reticulum; Gene Expression; Histocompatibility Antigens Class I; Interferon-gamma; Mice; Mice, Inbred C57BL; Mice, Transgenic; Myelin Basic Protein; Myelin Sheath; Oligodendroglia; Phenotype; RNA, Messenger

Theiler's murine encephalomyelitis virus (TMEV) infection produces a chronic inflammatory disease of the spinal cord white matter, with striking similarities to both experimental allergic encephalomyelitis (EAE) and human multiple sclerosis (MS). The first phase of demyelination in this model appears to be dependent on a delayed-type hypersensitivity (DTH) response to viral antigens, driven by CD4+, Th1 lymphocytes. Macrophages, recruited in the infected CNS, would be responsible for most of the myelin damage. Recently, new populations of CD4+ lymphocytes were demonstrated in infected mice, this time with specificity for myelin antigens, particularly PLP. This suggests that, in the chronic phase of the disease, an autoimmune mechanism of demyelination, similar to EAE, may participate in the process of myelin destruction. The present study represents a first step in exploring the functional activity of these anti-myelin lymphocytes that emerge during the chronic phase of the disease. Lymphocytes were removed from chronically infected animals, they were stimulated with the major PLP encephalitogenic epitope for SJL/J mice, and they were added to organotypic myelinated spinal cord cultures for different lengths of time. Results show that lymphocytes stimulated with the major PLP epitope have a powerful capacity for demyelinating these cultures, while MBP stimulated lymphocytes and lymphocytes from control animals do not. This study, suggests that the anti-myelin response that emerges during the chronic phase of the infection is functionally active. A similar phenomenon of epitope spreading from virus to organ specific antigens may take place in humans and be involved in a number of immune-mediated diseases, including MS.

Topics: Animals; Cardiovirus Infections; Cells, Cultured; Chronic Disease; Demyelinating Diseases; Encephalitis; Epitopes; Immunization; Lymphocytes; Mice; Mice, Inbred Strains; Myelin Basic Protein; Myelin Proteolipid Protein; Myelin Sheath; Organ Culture Techniques; Ovalbumin; Theilovirus

We show that normal peripheral nerve myelination depends on strict dosage of the most abundantly expressed myelin gene, myelin protein zero (Mpz). Transgenic mice containing extra copies of Mpz manifested a dose-dependent, dysmyelinating neuropathy, ranging from transient perinatal hypomyelination to arrested myelination and impaired sorting of axons by Schwann cells. Myelination was restored by breeding the transgene into the Mpz-null background, demonstrating that dysmyelination does not result from a structural alteration or Schwann cell-extrinsic effect of the transgenic P(0) glycoprotein. Mpz mRNA overexpression ranged from 30-700%, whereas an increased level of P(0) protein was detected only in nerves of low copy-number animals. Breeding experiments placed the threshold for dysmyelination between 30 and 80% Mpz overexpression. These data reveal new points in nerve development at which Schwann cells are susceptible to increased gene dosage, and suggest a novel basis for hereditary neuropathy.

Topics: Animals; Blotting, Western; Demyelinating Diseases; Gene Dosage; Gene Expression Regulation; Mice; Mice, Inbred BALB C; Mice, Transgenic; Myelin Basic Protein; Myelin P0 Protein; Myelin Proteins; Organ Specificity; Peripheral Nervous System Diseases; Protein Isoforms; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Schwann Cells; Sciatic Nerve

Myelinogenesis in rat brain aggregate cultures is associated with a pattern of growth factor mRNA expression comparable to that of the developing brain. The rate of increase in platelet-derived growth factor-AA (PDGF-AA) expression was greatest just before the detection of myelin basic protein (MBP) mRNA in the cultures and remained high thereafter, consistent with in vivo observations. Levels of fibroblast growth factor-2 (FGF-2) and of ciliary neurotrophic factor (CNTF) mRNA increased continuously over the period of MBP accumulation. High rates of transforming growth factor beta1 (TGF-beta1), insulin-like growth factor-I (IGF-I), and neurotrophin-3 (NT-3) expression at early time points during the culture gradually decreased over time, indicative of a key regulatory role during oligodendrocyte development. The addition of demyelinative anti-myelin oligodendrocyte glycoprotein (anti-MOG) antibody resulted in a significant increase in MBP peptide fragments with a C-terminus at phenylalanine 89 indicating proteolytic breakdown of MBP after myelin phagocytosis. Immediately after antibody treatment the expression of CNTF mRNA was significantly increased, compared with controls, while that of FGF-2 and IGF-I, and of PDGF-AA peaked during the early and later stages of recovery respectively. Thus, specific growth factors combine to regulate myelination and remyelination in the aggregates; these data have implications for demyelinating disease in which protective growth factor secretion may be central to regeneration.

Topics: Animals; Antibodies; Brain; Cells, Cultured; Ciliary Neurotrophic Factor; Demyelinating Diseases; Fibroblast Growth Factor 2; Growth Substances; Insulin-Like Growth Factor I; Myelin Basic Protein; Myelin Sheath; Platelet-Derived Growth Factor; Rats; RNA, Messenger

Demyelination contributes to the loss of function consequent to central nervous system (CNS) injury. Enhanced remyelination through transplantation of myelin-producing cells may offer a pragmatic approach to restoring meaningful neurological function. An unlimited source of cells suitable for such transplantation therapy can be derived from embryonic stem (ES) cells, which are both pluripotent and genetically flexible. In this paper we show that oligodendrocyte cultures can be reliably produced from retinoic acid-induced ES cells and that these oligodendrocytes can myelinate axons in vitro. Methods were further developed for generating highly enriched cultures of oligodendrocytes through an additional culturing step, producing an intermediate "oligosphere" stage. To test whether ES cells can survive, migrate, and differentiate into mature myelin-producing cells in areas of demyelination in the adult CNS, ES cells were transplanted into the dorsal columns of adult rat spinal cord 3 days after chemical demyelination. In the demyelination site, large numbers of ES cells survived and differentiated primarily into mature oligodendrocytes that were capable of myelinating axons. Furthermore, when oligosphere cells were transplanted into the spinal cords of myelin-deficient shiverer (shi/shi) mutant mice, the ES cell-derived oligodendrocytes migrated into the host tissue, produced myelin and myelinated host axons. These studies demonstrate the ability of ES cell-derived oligodendrocytes to myelinate axons in culture and to replace lost myelin in the injured adult CNS. Transplantation of ES cells may be a practical approach to treatment of primary and secondary demyelinating diseases in the adult CNS.

Topics: Animals; Antigens, Differentiation; Biomarkers; Cell Differentiation; Cell Lineage; Cells, Cultured; Demyelinating Diseases; Female; Fetal Tissue Transplantation; Mice; Mice, Neurologic Mutants; Myelin Basic Protein; Myelin Sheath; Nerve Tissue Proteins; Oligodendroglia; Rats; Spinal Cord; Spinal Cord Injuries; Stem Cell Transplantation; Stem Cells

Quakingviable (qk(v)) is a well known dysmyelination mutation. Recently, the genetic lesion of qk(v) has been defined as a deletion 5' to the qkI gene, which results in the severe reduction of the qkI-encoded QKI RNA-binding proteins in myelin-producing cells. However, no comprehensive model has been proposed regarding how the lack of QKI leads to dysmyelination. We hypothesized that QKI binds to myelin protein mRNAs, and the lack of QKI causes posttranscriptional misregulation, which in turn leads to the loss of the corresponding myelin proteins. To test this hypothesis, we developed an RNase protection assay to directly measure the mRNA isoforms encoding the myelin basic proteins (MBPs) in the brain. Our result suggested that isoform-preferential destabilization of MBP mRNAs in the cytoplasm was responsible for the reduced MBPs in the qk(v)/qk(v) brain during early myelination. In addition, we detected markedly reduced MBP mRNAs in the qk(v)/qk(v) myelin fraction with concomitant accumulation of MBP mRNAs associated with membrane-free polyribosomes. Presumably, the impaired localization of MBP mRNAs to the myelin membrane may cause insufficient incorporation of the newly synthesized MBPs into the myelin sheath. Finally, we observed interactions between QKI and MBP mRNAs, and removing MBP 3'UTR significantly reduced QKI-binding. Taken together, these observations suggest that misregulation at multiple posttranscriptional steps is responsible for the severe reduction of MBPs in qk(v) dysmyelination, presumably because of the lack of interactions between MBP mRNAs and the QKI RNA-binding proteins.

Topics: Animals; Crosses, Genetic; Demyelinating Diseases; Female; Gene Expression Regulation, Developmental; Male; Mice; Mice, Quaking; Myelin Basic Protein; Myelin Sheath; Polyribosomes; Protein Isoforms; RNA-Binding Proteins; RNA, Messenger

Double (plp-/-mag-/-) and triple (plp-/-mbp-/-mag-/-) null-allelic mouse lines deficient in proteolipid protein (PLP), myelin-associated glycoprotein (MAG), and myelin basic protein (MBP) were generated and characterized genetically, biochemically, and morphologically including their behavioral capacities. The plp-/-mag-/- mutant develops a rapidly progressing axon degeneration in CNS with severe cognitive and motor coordinative deficits but has a normal longevity. CNS axons of the plp-/-mbp-/-mag-/- mouse are hypomyelinated and ensheathed by "pseudomyelin" with disturbed protein and complex lipid composition. The shiverer trait in the plp-/-mbp-/-mag-/- similar to the plp-/-mbp-/- mutant is significantly ameliorated, and its lifespan is considerably prolonged. The longevity of these dysmyelinosis mouse mutants recommends them as suitable models for the long-term evaluation of stem cell therapeutic strategies.

Topics: Animals; Axons; Behavior, Animal; Central Nervous System; Demyelinating Diseases; Gene Expression Regulation; Genotype; Membrane Lipids; Mice; Mice, Neurologic Mutants; Myelin Basic Protein; Myelin Proteolipid Protein; Myelin Sheath; Myelin-Associated Glycoprotein; Neural Conduction; Peripheral Nerves; Phenotype; Transcription, Genetic

Previous reports have suggested that elevated levels of phenylalanine inhibit cholesterol synthesis. The goals of this study were to investigate if perturbations in cholesterol synthesis exist in the PAH(enu2) genetic mouse model for phenylketonuria (PKU), and if so, initiate studies determining if they might underlie the white matter pathology that exists in PKU forebrain. Gross sections and electron microscopy showed that select tracts were hypomyelinated in adult PKU mouse forebrain but not hindbrain. The activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR), the rate controlling enzyme in the cholesterol biosynthetic pathway, was examined in isolated microsomes from forebrain, hindbrain, and liver to assess if perturbations in cholesterol biosynthesis were occurring. HMGR activity was normal in unaffected PKU hindbrain and was increased 2-4-fold in PKU liver compared to control. HMGR activity in the forebrain, however, was decreased by 30%. Because normal numbers of MBP-expressing glia (oligodendrocytes) were present, but the number of glia expressing HMGR was reduced by 40% in the hypomyelinated tracts, the decreased HMGR activity seemed to result from a down-regulation of HMGR expression in affected oligodendrocytes. Exposure of an oligodendrocyte-like glioma cell line to physiologically relevant elevated levels of Phe resulted in a 30% decrease in cholesterol synthesis, a 28% decrease in microsomal HMGR activity, and a 28% decrease in HMGR protein levels. Measurement of HMGR activity after addition of exogenous Phe to control brain microsomes revealed that Phe is a noncompetitive inhibitor of HMGR; physiologically relevant elevated levels of exogenous Phe inhibited HMGR activity by 30%. Taken together, these data suggest that HMGR is moderately inhibited in the PKU mouse. Unlike other cell types in the body, a subset of oligodendrocytes in the forebrain seems to be unable to overcome this inhibition. We speculate that this may be the cause of the observed pathology in PKU brain.

Topics: Alkyl and Aryl Transferases; Animals; Brain Chemistry; Cell Count; Cell Line; Cholesterol; Demyelinating Diseases; Disease Models, Animal; Farnesyltranstransferase; Hydroxymethylglutaryl CoA Reductases; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Liver; Mice; Mice, Neurologic Mutants; Mice, Transgenic; Microsomes; Myelin Basic Protein; Oligodendroglia; Phenylalanine; Phenylalanine Hydroxylase; Phenylketonurias; Prosencephalon; Rhombencephalon

The 18q- deletion syndrome (18qDS) is frequently associated with cardiac anomalies. Patients with this syndrome may also have epilepsy, which presents certain diagnostic difficulties. This case report aims to illustrate these diagnostic problems, document the usefulness of heart rate-based seizure detection algorithms in this setting, and define the epilepsy syndrome associated with 18qDS.. Closed-circuit video electroencephalogram (EEG) monitoring using a heart rate-based seizure detection software was used to identify the event in question and to establish the diagnosis of epilepsy. Chromosomal analysis and magnetic resonance imaging (MRI) were used to further define the epilepsy syndrome.. We report on a patient with an atrial septal defect, enlargement of the right heart, and incomplete right bundle branch block, who developed episodes of tachycardia, loss of consciousness, and pallor, for which he was amnesic. Chromosomal analysis demonstrated karyotype 46,XY,del(18)(q21.3). ish del(18)(wcp18+,D18Z1+) with a loss of the gene for myelin basic protein. MRI revealed multifocal dysmyelination. Video-EEG monitoring using an electrocardiogram (ECG)-triggered seizure detection software proved to be indispensable in detecting an autonomic seizure and establishing the correct diagnosis; the procedure also allowed for the definition of the epilepsy syndrome. The patient was treated with carbamazepine and remained seizure-free.. Video-EEG monitoring using a heart rate-based seizure detection software can be helpful in diagnostically differentiating autonomic seizures from syncope. Dysmyelination due to loss of the myelin basic protein gene on 18q and cortical dysgenesis may be of pathogenic relevance.

Topics: Adult; Autonomic Nervous System Diseases; Chromosome Deletion; Chromosome Mapping; Chromosomes, Human, Pair 18; Demyelinating Diseases; Diagnosis, Differential; Electrocardiography; Electroencephalography; Epilepsy; Heart Defects, Congenital; Humans; Magnetic Resonance Imaging; Male; Monitoring, Physiologic; Myelin Basic Protein; Syncope; Syndrome; Videotape Recording

Since myelination and remyelination both involve investing an axon with a myelin sheath, a plausible hypothesis is that the two processes involve the expression of similar transcription factors. In this study we have addressed this hypothesis by comparing the expression of messenger RNA of Gtx, a homeodomain transcription factor expressed within oligodendrocytes during myelination, with the expression of messenger RNAs of the major myelin proteins, myelin basic protein and proteolipid protein during remyelination of experimentally induced demyelination in the adult rat brain. We have found a close temporal and spatial association between the expression patterns of the three messenger RNA species during remyelination. By comparing the expression patterns in rapidly remyelinating lesions in young adult rats with slowly remyelinating lesions in old adult rats, we have shown that Gtx messenger RNA expression follows the reappearance of myelin basic protein and proteolipid protein messenger RNAs regardless of the rate of remyelination. This observation demonstrates a clear association between the expression of Gtx messenger RNA and myelin repair. We have also shown that there is a decrease in constitutive levels of expression of myelin basic protein, proteolipid protein and Gtx messenger RNA in old adults compared with young adults. Taken together, our results indicate that Gtx, which has multiple binding sites in the promoter regions of both myelin basic protein and proteolipid protein genes, may have a similar role in the regulation of myelin protein gene expression during remyelination as has been proposed in myelination.

Topics: Aging; Animals; Brain; Demyelinating Diseases; Female; Homeodomain Proteins; In Situ Hybridization; Myelin Basic Protein; Myelin Proteolipid Protein; Nerve Tissue Proteins; Rats; Rats, Sprague-Dawley; RNA, Messenger; Transcription Factors

Focal demyelination models provide powerful tools to study demyelination and remyelination in the central nervous system. In this report, we present a novel technique, which selectively targets oligodendrocytes within the spinal cord of transgenic mice to produce focal demyelination. Transgenic mice expressing the E. coli LacZ (beta-galactosidase) gene from the myelin basic protein promotor allowed for oligodendrocyte-specific cleavage of topically applied fluorescein-di-beta-galactopyranoside liberating photoactivatable fluorescein. Subsequent fluorescence illumination generated oxygen radicals that oxidized a second exogenous substrate, 3-amino-9-ethyl carbazole, to form a toxic precipitate within oligodendrocytes. Histochemical staining of the spinal cord dorsal columns 8 days following phototargeting revealed that the treated region no longer contained beta-galactosidase-positive cells. Focal demyelination of the dorsal columns was observed to a depth of 150 microm in transverse semithin plastic sections. Numerous bundles of naked axons interspersed with myelin, debris-laden macrophages, and reactive astrocytes were evident by electron microscopy. Remyelination of axons by both oligodendrocytes and invading Schwann cells was observed within the treated region 14 days after phototargeting. Newly generated oligodendrocytes were identified within the demyelinated region by their incorporation of bromodeoxyuridine. Thus, this novel focal demyelination protocol provides: (1) a method for selective targeted ablation of oligodendrocytes in vivo, (2) control over the extent of the demyelinated region, with (3) an environment that maintains its remyelination capacity. Phototargeted ablation of oligodendrocytes may therefore be a useful model for studying axon-glia interactions, axon regeneration within a demyelinated zone, and remyelination of axons.

Topics: Animals; beta-Galactosidase; Bromodeoxyuridine; Cell Count; Demyelinating Diseases; Disease Models, Animal; Fluoresceins; Galactosides; Light; Mice; Mice, Transgenic; Myelin Basic Protein; Oligodendroglia; Promoter Regions, Genetic; Spinal Cord

Oligodendrocytes are glial cells devoted to the production of myelin sheaths. Myelination of the CNS occurs essentially after birth. To delineate both the times of oligodendrocyte proliferation and myelination, as well as to study the consequence of dysmyelination in vivo, a model of inducible dysmyelination was developed. To achieve oligodendrocyte ablation, transgenic animals were generated that express the herpes virus 1 thymidine kinase (HSV1-TK) gene under the control of the myelin basic protein (MBP) gene promoter. The expression of the MBP-TK transgene in oligodendrocytes is not toxic on its own; however, toxicity can be selectively induced by the systemic injection of animals with nucleoside analogs, such as FIAU [1-(2-deoxy-2-fluoro-beta-delta-arabinofuranosyl)-5-iodouracil]. This system allows us to control the precise duration of the toxic insult and the degree of ablation of oligodendrocytes in vivo. We show that chronic treatment of MBP-TK mice with FIAU during the first 3 postnatal weeks triggers almost a total depletion of oligodendrocytes in the CNS. These effects are accompanied by a behavioral phenotype characterized by tremors, seizures, retarded growth, and premature animal death. We identify the period of highest oligodendrocytes division in the first 9 postnatal days. Delaying the beginning of FIAU treatments results in different degrees of dysmyelination. Dysmyelination in MBP-TK mice is always accompanied by astrocytosis. Thus, this transgenic line provides a model to study the events occurring during dysmyelination of various intensities. It also represents an invaluable tool to investigate remyelination in vivo.

Topics: Animals; Antigens, Differentiation; Arabinofuranosyluracil; Blotting, Northern; Brain; Demyelinating Diseases; Disease Models, Animal; Female; Glial Fibrillary Acidic Protein; Gliosis; Herpesvirus 1, Human; In Situ Hybridization; Male; Mice; Mice, Transgenic; Myelin Basic Protein; Myelin Sheath; Oligodendroglia; Optic Nerve; Promoter Regions, Genetic; Recombinant Fusion Proteins; RNA, Messenger; Sciatic Nerve; Thymidine Kinase

Definition of the immune process that causes demyelination in multiple sclerosis is essential to determine the feasibility of Ag-directed immunotherapy. Using the nonhuman primate, Callithrix jacchus jacchus (common marmoset), we show that immunization with myelin basic protein and proteolipid protein determinants results in clinical disease with significant demyelination. Demyelination was associated with spreading to myelin oligodendrocyte glycoprotein (MOG) determinants that generated anti-MOG serum Abs and Ig deposition in central nervous system white matter lesions. These data associate intermolecular "determinant spreading" with clinical autoimmune disease in primates and raise important issues for the pathogenesis and treatment of multiple sclerosis.

Topics: Adjuvants, Immunologic; Animals; Autoantibodies; Callithrix; Demyelinating Diseases; Disease Models, Animal; Epitopes; Injections, Intradermal; Longitudinal Studies; Magnetic Resonance Imaging; Multiple Sclerosis; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Oligodendroglia; Recombinant Fusion Proteins

Experimental autoimmune encephalomyelitis (EAE) induction in TNF gene-targeted mice has resulted in conflicting reports in part due to the strong association of TNF with the MHC locus. To define the participation of TNF in EAE development, we back-crossed TNF-deficient mice (H-2b) into the SJL/J strain and directly compared them to H-2b congenic SJL or inbred SJL/J mice. Induction of EAE with myelin basic protein (MBP) revealed that H-2b congenic SJL mice are fully susceptible, indicating that the H-2b haplotype does not affect disease susceptibility. Using H-2b congenic SJL mice we show here that TNF deficiency modifies the normal course of EAE by considerably delaying the onset for approximately 5 days, suggesting that TNF is required for the normal initiation of MBP-induced EAE. However, TNF-deficient mice eventually developed severe EAE with perivascular inflammation and primary demyelination similar to wild-type controls, indicating that TNF is not essential during these processes. Taken together, these results indicate that although TNF is not required for the progression of MBP-induced EAE, it contributes positively by advancing the onset of disease.

Topics: Animals; Central Nervous System; Demyelinating Diseases; Disease Susceptibility; Encephalomyelitis, Autoimmune, Experimental; Female; H-2 Antigens; Incidence; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Myelin Basic Protein; Tumor Necrosis Factor-alpha

Active immunisation with rubella vaccine has not been commonly associated with neurological complications. We report the case of a 23-year-old woman who developed a mild, distal demyelinating neuropathy after immunisation with the live attenuated RA 27/3 rubella strain. Post-immunisation immunologic studies carried over 24 months showed the presence of antibodies to the RV proteins, particularly to the capsid antigen, and to the myelin basic protein (MBP). A similarity between a C antigen motif and a sequence of the MBP was found by computer analysis. The cross-reactivity was confirmed by immunising mice with a synthetic peptide derived from the MBP, which developed a strong humoral response to RV and MBP. This finding raises the possibility that a virus-induced immune response could lead to an autoaggressive reaction responsible for demyelination.

Topics: Adult; Animals; Antibodies; Antibodies, Viral; Antibody Specificity; Antigens, Viral; Blotting, Western; Cross Reactions; Demyelinating Diseases; Enzyme-Linked Immunosorbent Assay; Female; Humans; Mice; Mice, Inbred BALB C; Myelin Basic Protein; Peptides; Rubella Vaccine; Rubella virus

An augmentation of experimental allergic encephalomyelitis (EAE) was observed when monoclonal antibody (mAb) to intercellular adhesion molecule 1 (ICAM-1) was administered after adoptive transfer. Clinical disease was more severe in the ICAM-1 specific mAb-treated EAE mice and included prominent ataxia compared to the PBS-treated controls or Theiler's murine encephalomyelitis virus (TMEV) infected mice treated with ICAM-1 specific mAb. Neuropathologic evaluation demonstrated a distinctly different distribution of lesions in the anti-ICAM-1-treated EAE mice which featured prominent demyelination and inflammation in the cerebellum, brainstem and cerebrum. These structures were minimally involved in the control mice and mAb treatment did not alter the neuropathology in TMEV-infected mice. These results indicate that anti-ICAM-1 can alter trafficking of lymphocytes and mononuclear cells in EAE but not TMEV-induced demyelinating disease.

Topics: Animals; Antibodies, Monoclonal; Antigens, Viral; Brain; Cardiovirus Infections; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Intercellular Adhesion Molecule-1; Lymphocyte Function-Associated Antigen-1; Mice; Myelin Basic Protein; Recurrence; Theilovirus

To gain insights into the mechanisms of myelin repair in the CNS and to establish the extent to which this process resembles myelination in development we have examined the patterns of expression of transcripts of the major myelin proteins, myelin basic protein (MBP) and proteolipid protein (PLP) during remyelination of lysolecithin-induced demyelination in the adult rat spinal cord. Injection of 1 microliter 1% lysolecithin into the dorsal funiculus caused a dramatic decrease in levels of MBP exon 1 and MBP exon 2-containing transcripts and PLP/DM20 transcripts. Between 10 and 21 days post-lesion induction there was a gradual increase in levels of expression of all transcripts, which had returned to levels associated with normally myelinated spinal cord white matter at 21 days. These increases in levels of expression corresponded to the appearance of remyelinated axons, detected on toluidine blue-stained resin sections. Foci of high levels of expression occurred in regions of the lesion in which new myelin sheath formation was occurring, although the level of expression throughout the lesion never exceeded levels associated with myelin sheath maintenance in normal white matter due to the asynchronous pattern of remyelination. The changes in levels of expression of MBP exon 2 closely followed those of MBP exon 1. Our results indicate that (i) myelin protein gene expression associated with myelinogenesis during remyelination follows a similar pattern to that of myelinogenesis during development and that (ii) in rat models of demyelination changes of expression of MBP exon 1 and exon 2-containing transcripts are of equal value, an observation relevant to quantifying the effects of putative remyelination-enhancing strategies using the lysolecithin model.

Topics: Animals; Autoradiography; Demyelinating Diseases; Exons; Female; In Situ Hybridization; Lysophosphatidylcholines; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin Sheath; Oligonucleotide Probes; Rats; Rats, Inbred Strains; RNA, Messenger; Spinal Cord

Identifying a source of cells with the capacity to generate oligodendrocytes in the adult CNS would help in the development of strategies to promote remyelination. In the present study, we examined the ability of the precursor cells of the adult mouse subventricular zone (SVZ) to differentiate into remyelinating oligodendrocytes. After lysolecithin-induced demyelination of the corpus callosum, progenitors of the rostral SVZ (SVZa) and the rostral migratory pathway (RMS), expressing the embryonic polysialylated form of the neural cell adhesion molecule (PSA-NCAM), increased progressively with a maximal expansion occurring after 2 weeks. This observation correlated with an increase in the proliferation activity of the neural progenitors located in the SVZa and RMS. Moreover, polysialic acid (PSA)-NCAM-immunoreactive cells arizing from the SVZa were detected in the lesioned corpus callosum and within the lesion. Tracing of the constitutively cycling cells of the adult SVZ and RMS with 3H-thymidine labelling showed their migration toward the lesion and their differentiation into oligodendrocytes and astrocytes but not neurons. These data indicate that, in addition to the resident population of quiescent oligodendrocyte progenitors of the adult CNS, neural precursors from the adult SVZ constitute a source of oligodendrocytes for myelin repair.

Topics: Animals; Astrocytes; Cell Count; Cell Differentiation; Cell Division; Cell Movement; Corpus Callosum; Demyelinating Diseases; Glial Fibrillary Acidic Protein; Immunohistochemistry; Lateral Ventricles; Lysophosphatidylcholines; Mice; Mice, Inbred Strains; Myelin Basic Protein; Myelin Sheath; Neural Cell Adhesion Molecule L1; Neural Cell Adhesion Molecules; Oligodendroglia; Sialic Acids; Stem Cells; Thymidine

The function of neurons residing in a particular brain area is often assessed by injecting glutamatergic excitotoxins into that area and determining the consequences for the behavior of interest. However, injections of excitotoxins into the central nervous system not only kill local neurons but also demyelinate fibers of passage. Previous studies suggest that the myelin damage is triggered by a delayed inflammatory response to cell death mediated by monocytes of peripheral origin. If so, demyelination should commence only after recruitment of monocytes, their passage through the blood-brain barrier, and their metamorphosis into macrophages. This process is estimated to require at least 48 h. Using a hematoxylin (Weil) stain and immunohistochemistry for myelin basic protein, we looked for signs of demyelination at various times after injections of N-methyl-D-aspartate into the lateral hypothalamus. Demyelination was seen within 24 h after the lesion, sooner than predicted by the monocytic infiltration hypothesis. This finding has implications for interpreting effects of excitotoxic lesions and for developing means of improving their specificity.

Topics: Animals; Demyelinating Diseases; Hypothalamus; Immunohistochemistry; Male; Microinjections; Myelin Basic Protein; N-Methylaspartate; Rats; Rats, Long-Evans; Time Factors

Previous studies, both in vitro and in vivo, suggest that small portions of the mouse myelin basic protein (MBP) promoter are sufficient to activate regulated expression of MBP. To confirm our previous in vitro studies, we prepared transgenic mice with short regions of the human MBP promoter fused to the lacZ reporter gene. We found that 750 nucleotides of the proximal human MBP promoter is sufficient to activate oligodendrocyte-specific, developmentally regulated expression of lacZ in three independent lines. This promoter, however, does not activate expression of lacZ in Schwann cells in peripheral nerve or in adult mouse brain. The relative levels of beta-galactosidase specific activity, mRNA, and transcription parallel those of MBP mRNA during myelinogenesis. Thus, we exploited this transgene as a quantitative tool to evaluate the response to stimuli known to affect myelination. Transgene expression is reduced 75 % after optic enucleation, as previously reported for levels of MBP mRNA, indicating that axons signal to this portion of the proximal MBP promoter to fully activate MBP expression during myelinogenesis. Instead, in adult shiverer mice, another setting in which MBP transcription is modulated, transgene expression is not increased, in contrast to the increased transcriptional activation of MBP previously reported in these mice. These data suggest that the regulatory region that mediates transcriptional activation of the MBP gene is modular, since discrete subregions are required for activation in Schwann cells, during myelinogenesis in oligodendrocytes, during maintenance myelination in adult brain, and in the dysmyelinating mutant shiverer mouse.

Topics: Animals; Axons; Demyelinating Diseases; Eye Enucleation; Gene Expression Regulation, Developmental; Humans; Lac Operon; Mice; Mice, Neurologic Mutants; Mice, Transgenic; Myelin Basic Protein; Oligodendroglia; Promoter Regions, Genetic; Schwann Cells

Topics: Brain; Brain Diseases; Chromosome Deletion; Chromosomes, Human, Pair 18; Demyelinating Diseases; Humans; Magnetic Resonance Imaging; Myelin Basic Protein; Syndrome

Topics: Brain; Brain Diseases; Chromosome Deletion; Chromosomes, Human, Pair 18; Demyelinating Diseases; Humans; Magnetic Resonance Imaging; Myelin Basic Protein; Syndrome

Data from several studies indicate that free radicals have a pathogenic role in experimental allergic encephalomyelitis (EAE). Iron can contribute to free radical damage by catalyzing the formation of hydroxyl radical, inducing secondary initiation of lipid peroxidation and by promoting the oxidation of proteins. The iron chelator, desferrioxamine, can limit these oxidative reactions and it can scavenge peroxynitrite independent of iron chelation. Two previous studies have examined the therapeutic value of desferrioxamine in EAE. One study observed an effect when disease was induced by spinal cord homogenates (J. Exp. Med. 160, p. 1532, 1984), but a second study found no therapeutic value of desferrioxamine for myelin basic protein (MBP)-induced EAE (J. Neuroimmunol. 17, p. 127, 1988). In the second study, the drug was only administered during the preclinical stages of disease. Since desferrioxamine scavenges free radicals and prevents their formation, we hypothesized that the drug should be given during the active stage of disease to have therapeutic value. We first demonstrated that the drug enters the CNS around inflammatory cells in EAE animals. In animals treated during the active stage of MBP-induced EAE, the clinical signs were significantly reduced compared to vehicle-treated animals. The iron-bound form of this drug, ferrioxamine, was without therapeutic value. A derivative of desferrioxamine, hydroxylethyl starch (HES)-desferrioxamine, has a greater plasma half-life than desferrioxamine and it was also tested. Although there was a suggestion of improvement in these animals, the effects were less than that observed for desferrioxamine which may be related to the greater molecular size of HES-desferrioxamine. In summary, these data suggest that chelation of iron is an effective therapeutic target for EAE.

Topics: Animals; Deferoxamine; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Ferric Compounds; Free Radicals; Hydroxyethyl Starch Derivatives; Immunohistochemistry; Iron; Iron Chelating Agents; Male; Mice; Mice, Inbred Strains; Myelin Basic Protein; Myelin Sheath; Tissue Distribution

Multiple episodes of blood-brain barrier disruption were induced by sequential intraspinal injections of ethidium bromide. In addition to the barrier disruption, there was toxic demyelination and exposure of myelin components to the immune system. Twenty-seven 3-month-old Wistar rats received 2, 3 or 4 injections of 1 microliter of either 0.1% ethidium bromide in normal saline (19 rats) or 0.9% saline (8 rats) at different levels of the spinal cord. The time intervals between the injections ranged from 28 to 42 days. Ten days after the last injection, all rats were perfused with 2.5% glutaraldehyde. The spinal sections were evaluated macroscopically and by light and transmission electron microscopy. All the lesions demonstrated a mononuclear phagocytic infiltrate apparently removing myelin. Lymphocytes were not conspicuous and were found in only 34% of the lesions. No perivascular cuffings were detected. In older lesions (38 days and older) they were found only within Virchow-Robin spaces. This result suggests that multiple blood-brain barrier disruptions with demyelination and exposure of myelin components to the immune system were not sufficient to induce an immune-mediated reaction in the central nervous system.

Topics: Animals; Blood-Brain Barrier; Central Nervous System; Demyelinating Diseases; Ethidium; Female; Injections, Spinal; Lymphocytes; Male; Microscopy, Electron; Multiple Sclerosis; Myelin Basic Protein; Nicotinic Antagonists; Rats; Rats, Wistar; Spinal Cord

Although myelin basic protein (MBP)-recognizing T cells are not readily obtained after immunization of BALB/c mice with MBP (reflecting the BALB/c resistance to actively induced experimental autoimmune encephalomyelitis (EAE)), they can be expanded and cloned after several rounds of in vitro culture. The majority of BALB/c-derived clones recognize an epitope defined by MBP peptide 59-76. When transferred to naive BALB/c recipients, these clones cause classical EAE, with characteristic inflammation and demyelination of the central nervous system (CNS). We previously showed that two related clones recognizing a minor epitope, defined by MBP peptide 151-168, cause inflammation and demyelination preferentially of the peripheral nervous system (PNS). Because MBP has alternatively spliced isoforms, residues 151-168 are not present contiguously in all MBP isoforms. In order to determine whether induction of PNS disease is idiosyncratic to these sister clones, or related to their properties of epitope recognition, an independent T-cell line with similar recognition properties was studied. Clone 116F, derived from a BALB/c shiverer mouse, expresses a different T-cell receptor (TCR), with distinct TCR contact residues, but like the previously described T cells, this clone requires residues from both exons 6 and 7 for optimal stimulation. When adoptively transferred to BALB/c recipients, this clone preferentially induces disease of the PNS. A control BALB/c shiverer-derived MBP 59-76-recognizing clone, in contrast, induces CNS disease. These data strongly suggest that the site of disease initiation may correlate with epitope recognition, particularly when alternative isoforms are involved.

Topics: Alternative Splicing; Amino Acid Substitution; Animals; Antibody Specificity; Clone Cells; Cloning, Molecular; Demyelinating Diseases; Epitopes; Exons; Female; Mice; Mice, Inbred BALB C; Mice, Neurologic Mutants; Molecular Sequence Data; Myelin Basic Protein; Neuritis; Peptide Fragments; Peripheral Nervous System; Receptors, Antigen, T-Cell; T-Lymphocytes

A rabbit antiserum (anti-EP), induced against a synthetic peptide corresponding to residues 68 to 86 of guinea pig myelin basic protein, powerfully immunostained abnormal-appearing oligodendrocytic processes and cell bodies in demyelinating areas associated with multiple system atrophy (MSA). However, as we reported previously, the antiserum, which is highly specific for the sequence QDENPVV corresponding to human myelin basic protein residues 82 to 88, failed to recognize any structures in normal human brain. QD-9, a mouse monoclonal antibody raised against human myelin basic protein residues 69 to 88, which also recognizes specifically the epitope QDENPVV, gave the same results as did anti-EP. The unusual epitope recognized by anti-EP/QD-9 antibodies appears to be accessible in areas of myelin degeneration, and the antibodies have been shown to detect such areas in multiple sclerosis and infarcted brains. These antibodies detect myelin degeneration more widely than previous conventional methods. The present study emphasizes the importance of myelin degeneration in the pathogenesis of multiple system atrophy.

Topics: Aged; Aged, 80 and over; Animals; Antibodies, Monoclonal; Blotting, Western; Cerebellum; Demyelinating Diseases; Epitopes; Female; Guinea Pigs; Humans; Image Processing, Computer-Assisted; Immunoenzyme Techniques; Inclusion Bodies; Male; Mice; Mice, Inbred BALB C; Middle Aged; Multiple System Atrophy; Myelin Basic Protein; Myelin Sheath; Nerve Degeneration; Oligodendroglia; Peptide Fragments; Rabbits; Tumor Cells, Cultured

Immunization of (PL/J x SJL/J)F1 mice with myelin basic protein (MBP) induces relapsing experimental autoimmune encephalomyelitis (EAE). Relapses occur 7 to 10 days after recovery from the initial paralysis. Staphylococcal enterotoxins (SE) A or B, administered after recovery from the initial paralysis, induce immediate relapses. IL-12 is involved in the induction of EAE. Here, we show that SEA and SEB induce IL-12 in splenocytes from (PL/J x SJL/J)F1 mice in vitro and increase the level of IL-12 in the sera of mice treated with these superantigens. IL-12 administration mimics SE in inducing spontaneous relapses and in enhancing the severity and frequency of spontaneous relapses. IL-12 neutralization blocks SE-induced and subsequent relapses of EAE, and, when instituted after recovery from the initial attack, prevents spontaneous relapse. This is the first report of prevention of relapses of EAE with anti-IL-12 Ab, an approach which may prove useful in the prevention of exacerbations in multiple sclerosis.

Topics: Animals; Antibodies, Monoclonal; Antigens, Bacterial; Autoimmune Diseases; Brain; Cytokines; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Enterotoxins; Female; Guinea Pigs; Immunization; Inflammation; Interleukin-12; Male; Mice; Molecular Mimicry; Myelin Basic Protein; Rats; Recurrence; Severity of Illness Index; Spinal Cord; Staphylococcus aureus; Superantigens; Th1 Cells

When C57BL/6J mice, 8 weeks of age, received 0.2% Cuprizone in their diet, extensive demyelination in corpus callosum was detectable after 3 weeks, and there was massive demyelination by 4 weeks. As expected, the accumulation of phagocytically active microglia/macrophages correlated closely with demyelination. When Cuprizone was removed from the diet, remyelination was soon initiated; after 6 weeks of recovery, myelin levels were near-normal and phagocytic cells were no longer prominent. Steady-state levels of mRNA for myelin-associated glycoprotein, myelin basic protein, and ceramide galactosyltransferase were already profoundly depressed after 1 week of Cuprizone exposure and were only 10-20% of control values after 2 weeks. Unexpectedly, upregulation of mRNA for these myelin genes did not correlate with initiation of remyelination but rather with accumulation of microglia/macrophages. After 6 weeks of exposure to Cuprizone, mRNA levels were at control levels or higher-in the face of massive demyelination. This suggests that in addition to effecting myelin removal, microglia/macrophages may simultaneously push surviving oligodendroglia or their progenitors toward myelination.

Topics: Animals; Brain; Corpus Callosum; Cuprizone; Demyelinating Diseases; Diet; Galactosyltransferases; Gene Expression Regulation; Macrophages; Mice; Mice, Inbred C57BL; Microglia; Myelin Basic Protein; Myelin Proteins; Myelin Sheath; Myelin-Associated Glycoprotein; N-Acylsphingosine Galactosyltransferase; Phagocytosis; RNA, Messenger; Transcription, Genetic

Acute disseminated encephalomyelitis (ADEM), a postinfectious illness of the central nervous system (CNS), is thought to be an autoimmune disease. Here, we characterized the cytokines secreted by myelin-reactive T cells generated from patients with ADEM. The frequency of MBP-reactive T cell lines was ten-fold higher in patients with ADEM compared to patients with encephalitis and normal subjects. Whereas there was no significant IFN-gamma secretion, the predominant cytokine secreted by MBP-reactive T cell lines was IL-4 in patients with ADEM. In contrast, IL-4 secretion was only rarely detected in the controls. The presence of high frequencies of MBP-reactive IL-4 secreting T cells in subjects with ADEM during their recovery phase may be similar to myelin reactive IL-4 secreting T cells observed during the spontaneous recovery of animals with EAE.

Topics: Antibodies, Viral; Autoimmunity; Child; Child, Preschool; Demyelinating Diseases; Encephalomyelitis, Acute Disseminated; Encephalomyelitis, Autoimmune, Experimental; Epitopes; Female; Herpes Zoster; Herpesvirus 3, Human; Humans; Infant; Interferon-gamma; Interleukin-2; Interleukin-4; Male; Myelin Basic Protein; T-Lymphocytes

The Long Evans shaker (les) rat is a recently identified CNS myelin mutant with an autosomal recessive mode of inheritance. Although scattered myelin sheaths are present in some areas of the CNS, most notably the ventral spinal cord in the young neonatal rat, this myelin is gradually lost, and 8-12 weeks little myelin is present throughout the CNS. Despite this severe myelin deficiency, some mutants may live beyond one year of age. Rare, thin myelin sheaths that are present early in development lack myelin basic protein (MBP) and on ultrastructural examination are poorly compacted and lack a major dense line. Many oligodendrocytes develop an accumulation of vesicles and membranous bodies, but no abnormal cell death is observed. In the optic nerve, cell kinetic studies show an increase in proliferation at early time points in les, while total glial cell counts are also increased in les from 2 months of age. In situ hybridization studies demonstrate that the numbers of mature oligodendrocytes are similar to controls early in life and increase with time compared to controls. There is both a progressive astrocyte hypertrophy and microgliosis. While les has a mutation in the myelin basic protein (mbp) gene, it is dissimilar in both genotype and phenotype to the previously described mbp mouse mutants, shiverer (shi) and shiverer(mld). Unlike shi and its allele, where myelin increases with time and oligodendrocytes become ultrastructurally normal, les oligodendrocytes are permanently disabled, continue to demonstrate cytoplasmic abnormalities, and fail to produce myelin beyond the first weeks of life.

Topics: Animals; Brain; Cell Size; Demyelinating Diseases; Immunohistochemistry; In Situ Hybridization; Microscopy, Electron; Myelin Basic Protein; Myelin Sheath; Oligodendroglia; Rats; Rats, Long-Evans; Rats, Mutant Strains; Spinal Cord

We reported previously that accumulation of myelin basic protein (MBP) in foetal brain aggregate cultures is enhanced by supplementation with peritoneal macrophages. The present study demonstrates that the rate of MBP accumulation in macrophage-enriched cultures continues to increase over time unaccompanied by a matching increase in the oligodendrocyte marker cyclic nucleotide phosphodiesterase, while that of control cultures reaches a plateau. These observations are supported by electron microscopic evidence of cumulative numbers of myelinated axons in the aggregates over time and by enhanced expression of myelin protein genes in macrophage-enriched relative to control cultures. Aggregates demyelinate following short-term exposure to cytokines and antimyelin oligodendrocyte glycoprotein antibody, and MBP synthesis resumes following removal of demyelinating agents. Supplementation of cultures with macrophages influences the degree of myelin breakdown and remyelination, drawing attention to the role that macrophage-derived growth factors may play in myelinogenesis and myelin repair in inflammatory demyelinating disease.

Topics: Animals; Blotting, Northern; Brain; Cells, Cultured; Cytokines; Demyelinating Diseases; Gene Expression; Kinetics; Macrophages, Peritoneal; Myelin Basic Protein; Myelin Proteins; Myelin Sheath; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Rats; Rats, Sprague-Dawley

We assessed human myelin basic protein (MBP) binding IgM levels in CSF. MBP is the most studied putative antigen in multiple sclerosis (MS) and immune responses against it may be involved in the demyelination process. We also correlated these levels with EDSS score and other parameters of disease progression and prognosis, both at the time of CSF analysis and during follow-up. CSF IgM anti-MBP levels were assayed by measuring total IgM levels with solid-phase ELISA in CSF samples from 66 patients with relapsing-remitting MS, 11 subjects without neurological diseases, 20 patients with non-inflammatory neurological diseases and 7 patients with lymphocytic meningitis, before and after immunoabsorption with human MBP. Confirmation of IgM binding specificity was performed by immunoblotting of positive CSF samples onto MBP coated-nitrocellulose sheets. Clinical evaluation (disability score, number and time of attacks) was performed during a mean follow-up of 2.7 +/- 1.1 years. 23 of the 66 relapsing-remitting MS patients (33.8%) had elevated IgM anti-MBP levels. In this patient subgroup, IgM anti-MBP levels correlated with the IgM index (r = 0.71; P = 0.0001), but not with CSF/serum albumin (r = 0.08; P = 0.72). In the first year of follow-up, patients with low IgM anti-MBP suffered from more numerous attacks than those with elevated levels (0.86 +/- 0.63 versus 0.43 +/- 0.58; P = 0.017). Patients with high IgM binding to MBP had a first attack during follow-up in a significantly higher time than those with low binding (28.87 +/- 4.7 versus 17 +/- 2.6 months, respectively; P = 0.005) and reached a decrease of 0.5 EDSS point significantly faster than those with low IgM (16.17 +/- 1.2 versus 29.7 +/- 2.6 months, respectively; P = 0.0002). A similar significant finding was observed when the time to reach low disability score (EDSS < or = 2.0) was analyzed (10.7 +/- 2.57 +/- 3.3 months, respectively; P = 0.014). These findings demonstrate that in a subgroup of MS patients, elevated CSF levels of IgM anti-MBP are associated with early favorable course and therefore suggest that IgM binding to MBP could be a possible prognostic marker in relapsing-remitting MS to select early MS patients for future trials.

Topics: Adolescent; Adult; Autoantibodies; Demyelinating Diseases; Disease Progression; Female; Follow-Up Studies; Humans; Immunoblotting; Immunoglobulin M; Male; Middle Aged; Multiple Sclerosis; Myelin Basic Protein; Prognosis; Survival Analysis

Hemispheres, spinal cords, and sciatic nerves were taken from taiep, carrier, and control rats at ages ranging from 1 day to 16 months. Absolute myelin yields from CNS taiep tissues peaked at approximately 2 months and then decreased until they reached a low but stable level. Myelin yield from the affected hemispheres expressed as a percentage of age-matched controls decreased continuously from 2 weeks until it reached a stable level of approximately 10-15%. The same was true for the spinal cords, but here the myelin yield reached a plateau at a slightly higher percentage of 20-25%. In comparison with control rats, isolated CNS myelin fractions from the affected rats had a greater content of high molecular weight proteins. Western blot analyses of CNS homogenates revealed that myelin basic protein (MBP), proteolipid protein, and 2',3'-cyclic nucleotide 3'-phosphodiesterase were all present but decreased to levels generally consistent with the deficiencies of myelin. However myelin-associated glycoprotein (MAG) levels always were reduced much more than those of the other three myelin proteins, and at younger ages the apparent molecular weight for MAG was increased in the mutants. Western blot analyses of sciatic nerve homogenates showed that the levels of MBP, MAG, and P0 were not significantly different in control and mutant animals. These results suggested an early hypomyelination of the CNS, with peak levels of myelin at 2 months, followed by a prolonged period of myelin loss, until a very low but stable myelin level was reached. The consistently greater loss of MAG, in comparison with other CNS myelin proteins, is different from most other hypomyelinating mutants in which MAG is relatively preserved in comparison with the proteins of compact myelin. This might be due to microtubular abnormalities in the taiep mutant interfering with transport of myelin proteins and having the greatest effect on MAG because of its most distal location in the periaxonal oligodendroglial membranes.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Blotting, Western; Brain Chemistry; Demyelinating Diseases; Electrophoresis, Polyacrylamide Gel; Molecular Weight; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin Sheath; Myelin-Associated Glycoprotein; Rats; Rats, Mutant Strains; Sciatic Nerve; Spinal Cord

The adoptive transfer of autoreactive S100 beta-specific T cells induces experimental autoimmune panencephalomyelitis and uveoretinitis in the Lewis rat, mimicking the distribution of lesions seen in a subset of patients with multiple sclerosis. We studied the frequency and functional properties of the human T-cell response to S100 beta in eight patients (two relapsing-remitting multiple sclerosis, one chronic-progressive multiple sclerosis, two with multiple sclerosis and uveitis, two neuromyelitis optica, one panuveitis) and in seven healthy individuals, using bovine S100 beta for T-cell stimulation. Both in patients and controls, the frequency of S100 beta-specific T-cell responses was half of that obtained for myelin basic protein (MBP), and only 10% of that obtained using purified protein derivative (PPD). The stimulation indices obtained in response to S100 beta were also less than half those obtained with either MBP or PPD. However, four long-term S100 beta-specific T-cell lines were established and studied in more detail. The four T-cell lines all exhibited a CD4+, CD8-, T-cell receptor alpha beta + surface phenotype and secreted tumour necrosis factor-alpha, interferon-gamma, interleukin-10 and interleukin-4 upon antigenic stimulation, but they were heterogenous with respect to T-cell receptor usage; two T-cell lines expressed V beta 2, one V beta 6.7 and one V beta 13. Antigen-specificity was confirmed using bovine S100 beta beta and alpha beta-isoforms, as well as a recombinant rat S100 beta preparation. The response to S100 beta was shown to the HLA-(human leukocyte antigen-) DR-restricted for two of the S100 beta-specific T-cell lines. Human S100 beta-specific T-cell lines were cytotoxic, although to a lesser extent than MBP-specific T-cell lines derived from the same donors. The phenotypic and functional properties of human S100 beta-specific T-cell lines raise the possibility that these T cells are pathogenic, as they are in the rat. The low frequency and proliferative index of S100 beta-specific, as opposed to MBP-specific T-cell responses suggests that the T-cell response to this widely expressed calcium-binding protein is under more efficient regulatory control.

Topics: Adult; Animals; Antibody Specificity; Autoantigens; Calcium-Binding Proteins; Cattle; Cell Line; Cytokines; Cytotoxicity Tests, Immunologic; Demyelinating Diseases; Disease Models, Animal; Encephalomyelitis, Autoimmune, Experimental; Female; Histocompatibility Antigens; Histocompatibility Testing; Humans; Immunophenotyping; Immunotherapy, Adoptive; Male; Middle Aged; Multiple Sclerosis; Myelin Basic Protein; Nerve Growth Factors; Rats; Rats, Inbred Lew; Receptors, Antigen, T-Cell; S100 Calcium Binding Protein beta Subunit; S100 Proteins; T-Lymphocytes; Tuberculin; Uveitis

The molecular mechanisms necessary for remyelination by oligodendrocytes remain unexplored. We previously characterized a myelin basic protein promoter-lacZ (MBP-lacZ) transgene whose expression is regulated uniquely during development, and also in pathological situations, suggesting that it may be a useful reporter of molecular mechanisms during remyelination. As a first step toward creating a transgenic mouse model of remyelination, we cultured oligodendrocytes from these transgenic mice and showed that expression of MBP-lacZ appeared in parallel with a marker of oligodendrocyte maturation, galactocerebroside (GC). In addition, basic fibroblast growth factor blocked the expression of both MBP-lacZ and GC in these cells. Therefore, expression of MBP-lacZ reflects not only the developmental stage of oligodendrocytes, but also extrinsic influences on oligodendrocytes. These data suggest that MBP-lacZ may be a useful marker in transgenic mouse models of remyelination.

Topics: Animals; beta-Galactosidase; Brain; Cells, Cultured; Coculture Techniques; Demyelinating Diseases; Disease Models, Animal; Mice; Mice, Transgenic; Myelin Basic Protein; Myelin Sheath; Nerve Regeneration; Neuroglia; Oligodendroglia; Platelet-Derived Growth Factor; Recombinant Fusion Proteins

To find whether CSF analysis may differentiate between relapsing-remitting and secondary progressive multiple sclerosis.. In 17 patients with relapsing-remitting and 16 patients with secondary progressive multiple sclerosis, all without current or recent relapses, albumin CSF: peripheral blood ratio, mononuclear cell number, CD4+, CD8+, and B1+ subsets, CD4+:CD8+ ratio, IgG, IgG index, IgM, IgM index, complement components C3 and C4, and C3 and C4 indices, myelin basic protein, neuron specific enolase, S100, and lactate were determined. For each measure the statistical distance measure D2 was calculated. For computation of a discriminant score variables with a P value< or =0.15 were included (two sided univariate t test). These were albumin CSF: peripheral blood ratio, mononuclear cell number, IgM, IgM index, C3, C4, neuron specific enolase, S100, and lactate. Simultaneous distributions of the variables were compared between both groups (multivariate t test) and a discriminant score was computed (linear discriminant analysis).. The discriminant score allocated all 14 relapsing-remitting patients to the relapsing-remitting group (positive score) and 12 of 13 secondary progressive patients to the secondary progressive group (negative score). One secondary progressive patient was allocated to the relapsing-remitting group.. Patients with relapsing-remitting or secondary progressive multiple sclerosis differ in CSF profile and CSF analysis may help to differentiate between relapsing-remitting and secondary progressive multiple sclerosis.

Topics: Adult; Albumins; Antigens, CD; Blood-Brain Barrier; Complement System Proteins; Demyelinating Diseases; Disease Progression; Female; Humans; Immunoglobulin G; Immunoglobulin M; Lactates; Male; Multiple Sclerosis; Myelin Basic Protein; Phosphopyruvate Hydratase; Recurrence; Remission, Spontaneous

It was recently demonstrated that selective phosphodiesterase type 4 (PDE4) inhibition suppresses the clinical manifestations of acute experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (MS), and inhibits the production of tumor necrosis factor-alpha (TNF-alpha), a pathogenetically central cytokine. Since the most common presentation of MS in humans is a relapsing-remitting course, we investigated the therapeutic potential of PDE4 inhibition in the relapsing-remitting EAE model of the SJL mouse. Administration of rolipram, the prototypic PDE4 inhibitor, reduced the clinical signs of EAE during both the initial episode of disease and subsequent relapses. In parallel, there was marked reduction of demyelination and also less inflammation throughout the central nervous system (CNS) of rolipram-treated animals. Gene expression of proinflammatory cytokines in the CNS was reduced in most of the rolipram-treated animals. Additional experiments demonstrated that PDE4 inhibition acted principally by inhibiting the secretion of Th1 cytokines, however, the encephalitogenic potential of myelin basic protein-specific T cells was not impaired. Our findings suggest that PDE4 inhibitors are a promising cytokine-directed therapy in chronic demyelinating disease.

Topics: 3',5'-Cyclic-AMP Phosphodiesterases; Animals; Autoimmune Diseases; Cell Count; Cells, Cultured; Cyclic Nucleotide Phosphodiesterases, Type 4; Demyelinating Diseases; Female; Lymph Nodes; Mice; Mice, Inbred Strains; Myelin Basic Protein; Phosphodiesterase Inhibitors; Pyrrolidinones; Rolipram; T-Lymphocytes

Many patients with AIDS have a myelopathy characterized by vacuolization of spinal cord white matter. The biochemical and molecular changes underlying this myelin disturbance have not yet been characterized. Myelin basic protein (MBP) is potentially important because it is a key structural protein of myelin with roles in compaction and stabilization. In the present study, we describe the steady-state protein concentration of MBP in 46 patients with AIDS and 12 control subjects at autopsy. Patients with myelopathy exhibited no change in the abundance of the predominant 18.5- and 17.2-kd isoforms, but a 14-kd MBP-immunoreactive degradation fragment was increased significantly. MBP degradation correlated significantly with the severity of histopathologic changes, including neutral lipid deposition, the density of vacuolated fibers, and the number of ferritin-stained activated microglia. Alkaline gel electrophoresis of isolated MBP showed preferential loss of the least cationic isomer (C-8). The concentration of MBP RNA in slot blots was normal in cords exhibiting myelopathy, and the ratio of mRNA corresponding to the 18.5- and 17.2-kd MBP isoforms, measured using reverse transcriptase-PCR, was not altered. This study suggests that mononuclear phagocyte-mediated degradation of MBP may play a role in AIDS myelopathy, and the preferential loss of the C-8 component of MBP may have mechanistic implications.

Topics: Acquired Immunodeficiency Syndrome; Adult; Blotting, Western; Demyelinating Diseases; Humans; Hydrolysis; Immunohistochemistry; Isomerism; Male; Middle Aged; Myelin Basic Protein; RNA, Messenger; Spinal Cord

Transgenic mice containing different numbers of transgenes (2-70) of the myelin proteolipid protein DM20 were phenotypically normal up to 3 mo of age, after which the mice containing 70 copies of the transgene spontaneously demyelinated and died at 10-12 mo. Since we demonstrated that demyelination in multiple sclerosis involved specific chemical changes in myelin basic protein (MBP), we investigated the MBP in our transgenic line for similar changes. Both the total amount of MBP in brain and the MBP mRNA levels were unaffected at the different ages. All the isoforms (14-21 kD) of MBP were present, but the microheterogeneity (a posttranslational event) was changed resulting in a higher proportion of the less cationic components reminiscent of the changes in MBP found in multiple sclerosis. An increased amount of the citrullinated form of MBP was found by Western blot analysis. Immunogold labeling of cryosections of brain revealed a greater density of particles with the anticitrulline antibody at 10 mo and that the levels of peptidylarginine deiminase (which deiminates protein-bound arginine to citrulline) were increased. This stable transgenic line represents a useful animal model for the human disease multiple sclerosis.

Topics: Animals; Citrulline; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Gene Dosage; Hydrolases; Isoelectric Point; Mice; Mice, Mutant Strains; Mice, Transgenic; Multiple Sclerosis; Myelin Basic Protein; Myelin Proteolipid Protein; Protein-Arginine Deiminase Type 4; Protein-Arginine Deiminases

We tested the hypothesis that immunoglobulins directed against a CNS autoantigen, myelin basic protein, may promote remyelination in the course of a chronic, immune-mediated demyelinating disease. SJL/J mice infected chronically with Daniel's strain of Theiler's virus served as an experimental model of MS. The spinal cords of these mice exhibit extensive primary demyelination and inflammation with minimal spontaneous remyelination. Treatment with whole antiserum or affinity-purified mouse immunoglobulins directed against rat or rabbit myelin basic protein increased new myelin synthesis as measured by quantitative morphometry. Electron microscopy revealed numerous oligodendrocytes in remyelinated CNS lesions and a relative lack of inflammatory cells. Viral antigen persisted in the spinal cord despite enhanced CNS-type remyelination. These findings indicate that immunoglobulins reactive with myelin autoantigens have the potential to promote myelin repair.

Topics: Animals; Blotting, Western; Cerebellum; Demyelinating Diseases; Immunoglobulins; Kidney; Mice; Mice, Inbred Strains; Myelin Basic Protein; Myelin Sheath; Poliomyelitis; Rabbits; Rats; Spinal Cord; Theilovirus

We have studied a case of acute, fulminating multiple sclerosis (MS) (Marburg type) at the pathological and biochemical levels. Postmortem examination of the brain revealed extensive areas of gross rarefaction in the hemispheric white matter. Histologically, well-demarcated areas of demyelination with a large influx of macrophages and a subtle perivascular infiltration of lymphocytes were seen with relative preservation of the axis cylinders. Myelin basic protein (MBP) was isolated and purified [correction of purifed] from noninvolved white matter. It was slightly larger in molecular weight than MBP from normal brain or from chronic MS brain. The increase in mass was accounted for, in part, by the deimination of 18 of 19 arginyl residues to citrulline, making the patient's MBP much less cationic than MBP from normal white matter. When expressed as the ratio of least cationic form of MBP to the most cationic (C-8/C-1), the normal ratio was 0.82, chronic MS 2.5, and the patient in this study 6.7. Because the ratio of 6.7 was similar to 7.5 found for a 15-month-old infant, MBP was considered to be of the immature form. The data are consistent with a genetic factor influencing the charge microheterogeneity of MBP. The resulting less cationic MBP cannot carry out its normal function of compacting multilayers.

Topics: Adult; Antibodies, Monoclonal; Arginine; Blotting, Western; Brain; Brain Chemistry; Chronic Disease; Citrulline; Demyelinating Diseases; Fatal Outcome; Female; Humans; Lymphocytes; Macrophages; Magnetic Resonance Imaging; Multiple Sclerosis; Myelin Basic Protein

Topics: Chromosomes, Human, Pair 18; Demyelinating Diseases; Humans; Multiple Sclerosis; Myelin Basic Protein; Phosphorylation

Increased expression of MHC Class I occurs in the central nervous system in association with demyelinating diseases such as multiple sclerosis and experimental allergic encephalomyelitis. To determine if MHC Class I expression by oligodendrocytes induces white matter pathology, the MHC Class I gene was expressed in transgenic mice under the control of the myelin basic protein (MBP) promoter. These mice display a neurological phenotype at 21 days-of-age. We examined these mice at 1,3, and 12 weeks-of-age. MHC Class I was detected in the brains and spinal cords of transgenic mice but not in control mice. Class I was located in oligodendrocyte perikarya but not in myelin sheaths. The central nervous system of these transgenic mice was hypomyelinated and contained hypertrophic microglia and astrocytes. These observations establish that Class I expression by oligodendrocytes delays normal myelination but does not cause inflammatory demyelination. This hypomyelinating animal model is of potential use in studying the interactions between immunologically active molecules and remyelination in disorders of myelin.

Topics: Aging; Animals; Brain; Demyelinating Diseases; Genes, MHC Class I; Histocompatibility Antigens Class I; Humans; Mice; Mice, Transgenic; Myelin Basic Protein; Oligodendroglia; Promoter Regions, Genetic; Spinal Cord

Demyelination in the transgenic mice depended on the dosage of the cDNA for DM20, in which low copy numbers (two to four and 17 copies of the minigene) showed no signs of demyelination. However when transgenic mice with 17 copies were made homozygous with 34 copies of the DM20 minigene (ND3A hm.) demyelination was observed at around 12 to 16 months compared with ND4 mice having 70 copies of the transgene which had an earlier onset of demyelinating symptoms at 3 months, demonstrating a transgene dosage effect. The process by which demyelination was initiated was associated with changes in myelin basic protein. An increased abundance of less cationic MBP (C-8) isomers occurred prior to demyelination. This increase was also associated with increased activity of peptidylarginine deiminase, the enzyme which converts arginine to citrulline in proteins, thereby providing a mechanism for generating less cationic forms of MBP. These data support a dosage effect of the DM20 transgene.

Topics: Aging; Animals; Axons; Demyelinating Diseases; Female; Heterozygote; Homozygote; Humans; Hydrolases; Male; Mice; Mice, Transgenic; Myelin Basic Protein; Myelin Proteolipid Protein; Nerve Tissue Proteins; Optic Nerve; Protein-Arginine Deiminase Type 4; Protein-Arginine Deiminases; Recombination, Genetic

The microheterogeneity of myelin basic protein, expressed as the ratio between the least cationic (C-8) charge isomer and the most cationic (C-1), was examined in experimental allergic encephalomyelitis (EAE) cases. These included acute EAE of 2 months' duration induced with bovine proteolipid protein in complete Freund's adjuvant (CFA), chronic EAE induced with mouse spinal cord homogenate in varying doses from 0.5 to 2.0 mg in CFA, and chronic relapsing EAE of 12 months' duration induced with synthetic peptide 139-151 of the proteolipid protein sequence. The C-8/C-1 ratio was within the normal range for all groups of animals. However, the C-8/C-1 ratio was six- to sevenfold increased in a spontaneously demyelinating transgenic model, ND4, which contains 70 copies of the cDNA for DM20 (Mastronardi et al.: 1996). Since an increase in the C-8/C-1 ratio was also observed in victims of multiple sclerosis but not other neurological diseases, the ND4 model may address primary changes prior to demyelination, while the EAE model addresses the autoimmune aspects of the disease.

Topics: Animals; Cattle; Cerebellum; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Humans; Mice; Mice, Inbred BALB C; Mice, Inbred Strains; Mice, Transgenic; Mycobacterium tuberculosis; Myelin Basic Protein; Myelin Proteolipid Protein; Nerve Tissue Proteins; Protein Processing, Post-Translational; Spinal Cord; Time Factors

Human JC virus (JCV) is a neurotropic human polyomavirus that was found in the plaques and oligodendroglial cells of the brains of patients with the fatal demyelinating disease, progressive multifocal leukoencephalopathy (PML). Transgenic mice expressing JCV large tumor (T)-antigen from integrated DNA showed dysmyelination in the central nervous system. However, the role of T-antigen from episomal DNA in the demyelination in PML remains unclear. In this report, we examined the effect of episomally expressed JCV T-antigen on the expression of myelin basic protein (MBP) in U-87 MG human glioblastoma cells to study the mechanism of demyelination. Expression assays of the MBP promoter in U-87 MG detected a 2.5-fold reduction in cells expressing intact T-antigen. Next, U-87 MG expressing T-antigen were examined by RNase protection assays for mRNA accumulation from the endogenous MBP promoter. Also, the expression of the MBP promoter plasmid was determined using in vitro transcription assays with extracts from T-antigen expressing cells. Both assays found a similar down-regulation of the MBP promoter by T-antigen, confirming that negative regulation occurred at the transcriptional level for the endogenous and exogenous MBP promoters. Furthermore, in situ immunofluorescence assays and quantitative Western blot analysis provided convincing evidence of a similar reduction in the level of MBP produced from the functional endogenous gene in U-87 MG glioblastoma cells expressing T-antigen. Thus, we provide evidence for the role of T-antigen in a transcriptional control mechanism for the demyelination that is caused by JCV in PML patients.

Topics: Antigens, Polyomavirus Transforming; Demyelinating Diseases; Down-Regulation; Gene Expression Regulation, Viral; Glioblastoma; Humans; JC Virus; Myelin Basic Protein; Promoter Regions, Genetic; RNA, Messenger; Transcription, Genetic; Tumor Cells, Cultured

Proteolipid protein (PLP) is a major structural component of central nervous system (CNS) myelin. Evidence exists that PLP or the related splice variant DM-20 protein may also play a role in early development of oligodendrocytes (OLs), the cells that form CNS myelin. There are several naturally occurring mutations of the PLP gene that have been used to study the roles of PLP both in myelination and in OL differentiation. The PLP mutation in the jimpy (jp) mouse has been extensively characterized. These mutants produce no detectable PLP and exhibit an almost total lack of CNS myelin. Additionally, most OLs in affected animals die prematurely, before producing myelin sheaths. We have studied cultures of jp CNS in order to understand whether OL survival and myelin formation require production of normal PLP. When grown in primary cultures, jp OLs mimic the relatively undifferentiated phenotype of jp OLs in vivo. They produce little myelin basic protein (MBP), never immunostain for PLP, and rarely elaborate myelin-like membranes. We report here that jp OLs grown in medium conditioned by normal astrocytes synthesize MBP and incorporate it into membrane expansions. Some jp OLs grown in this way stain with PLP antibodies, including an antibody to a peptide sequence specific for the mutant jp PLP. This study shows that: (1) an absence of PLP does not necessarily lead to dysmyelination or OL death; (2) OLs are capable of translating at least a portion of the predicted jp PLP; (3) the abnormal PLP made in the cultured jp cells is not toxic to OLs. These results also highlight the importance of environmental factors in controlling OL phenotype.

Topics: Animals; Cell Death; Cells, Cultured; Demyelinating Diseases; Female; Glycolipids; Immunohistochemistry; Lipoproteins; Male; Mice; Mice, Jimpy; Mutation; Myelin Basic Protein; Myelin Proteins; Oligodendroglia; Phenotype; Protein Biosynthesis; Rats; Up-Regulation

Caprine beta-mannosidosis is an inherited lysosomal storage disease that leads to a deficiency of oligodendrocytes and hypomyelination. Our previous results demonstrated that low levels of myelin-associated glycoprotein (MAG), 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNP) and proteolipid protein (PLP) found in CNS samples correlated with decreased yields of myelin. However, there was a relative preservation of myelin basic protein (MBP) in the spinal cord samples of affected goats. This report shows that the amounts of myelin protein mRNAs in the spinal cords of affected goats relative to control goats are also decreased. The levels of mRNA for MAG, MBP and PLP in affected goat spinal cords compared with those of controls were equally decreased to approximately 50% for the three myelin proteins. This suggests that the relative preservation of MBP protein in the spinal cords is not due to a higher MBP mRNA level, but might be due to a difference in post-transcriptional processes.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; alpha-Mannosidosis; Animals; Blotting, Northern; Central Nervous System; Demyelinating Diseases; Goat Diseases; Goats; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin-Associated Glycoprotein; RNA, Messenger; Spinal Cord

Murine chronic relapsing experimental autoimmune encephalomyelitis (CR-EAE) is a model of inflammatory demyelinating disease of the central nervous system (CNS) with similarity to multiple sclerosis (MS) in humans. Mice with confirmed neurologic deficits from CR-EAE were treated by oral administration of whole bovine myelin to investigate the effect of long-term oral delivery of myelin antigens on clinical disease and on the inflammatory response in the CNS. EAE-positive mice were fed doses of 1 mg, 10 mg, or 20 mg of bovine myelin every other day for 6 months. We found that prolonged oral delivery of neuroantigen suppressed inflammatory and demyelination foci in the CNS of myelin-treated mice with no exacerbation of clinical disease status compared with the control group. Analysis of histologic sections of brain and spinal cords with hematoxylin-eosin (H&E) and Luxol fast blue (LFB) staining showed a decrease in the inflammatory cell infiltration and active centers of demyelination, respectively. Furthermore, after 6 months of treatment, there was no increased sensitization to myelin antigens seen, as measured by antimyelin basic protein (MBP) or anti-proteolipid apoprotein (PLP) antibodies. These results demonstrate that prolonged oral administration of myelin antigens in diseased animals has an ameliorating effect on the pathologic process and supports its potential long-term use in humans with MS.

Topics: Administration, Oral; Animals; Antibody Formation; Cattle; Chronic Disease; Demyelinating Diseases; Disease Progression; Encephalomyelitis, Autoimmune, Experimental; Female; Mice; Mice, Inbred Strains; Myelin Basic Protein; Myelin Proteolipid Protein; Recurrence; Time Factors

Chronic progression of autoimmune disease is accompanied by the acquisition of autoreactivity to new self-determinants. Recent evidence indicates that this process, commonly referred to as determinant spreading, may be pathogenic for chronicity. Our studies on experimental autoimmune encephalomyelitis (EAE), a murine model widely used in multiple sclerosis (MS) studies, have shown that determinant spreading develops as a predictable sequential cascade of neo-autoimmunity during progression to chronic disease. By 7-8 weeks after immunization of (SWR x SJL)F1 mice with the immunodominant myelin proteolipid protein determinant (PLP 139-151), splenocytes consistently respond to the immunodominant myelin basic protein determinant (MBP 87-99). In the present study, we directly address the pathogenicity of neo-autoimmunity resulting from endogenous self-priming during the course of disease. Our results indicate that T cells responding to the spreading MBP 87-99 determinant produce a proinflammatory cytokine profile consistent with type 1 helper T cells (Th1) cells. In addition, splenocytes activated to the spreading MBP 87-99 determinant consistently transfer acute EAE in naive recipients even when T cells reactive to the priming PLP 139-151 immunogen are eliminated by bromodeoxyuridine (BUdR)-mediated photolysis. Our data indicate that endogenous neo-autoantigen priming during chronic autoimmune disease generates type 1 helper T cells (Th1) cells that are autonomously pathogenic. These results provide further evidence supporting the view that determinant spreading is a pathogenic process that leads to chronic progression of autoimmune disease.

Topics: Adoptive Transfer; Animals; Antigen Presentation; Antimetabolites; Autoantigens; Bisbenzimidazole; Bromodeoxyuridine; Central Nervous System Diseases; Cytokines; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Epitopes; Female; Fluorescent Dyes; Immunization; Mice; Mice, Inbred Strains; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Peptide Fragments; Photochemistry; Spleen; T-Lymphocytes, Helper-Inducer

Theiler's murine encephalomyelitis virus (TMEV) produces a chronic inflammatory demyelinating disease in its natural host, the mouse. A delayed-type hypersensitivity (DTH) response to viral antigens generally correlates with susceptibility to the disease and is thought to play an important role in the pathogenesis of demyelination in this model of human multiple sclerosis (MS). The hallmark of DTH responses is the recruitment by activated Th-1 cells of lymphoid cells and especially macrophages in infected areas. It is believed that soluble factors released by these cells would produce tissue damage, particularly myelin breakdown. In the present study, we compared TMEV-infected macrophages and microglia, isolated from both susceptible SJL/J and resistant C57BL/6 mice, for their ability to secrete proteolytic enzymes capable of degrading myelin basic protein. In addition, we studied whether supernatants from infected microglia/macrophages were also capable of killing oligodendrocytes in the same in vitro system. As detected by SDS-PAGE, MBP-degrading proteolytic activity was found only in supernatants from infected SJL/J microglia and macrophages, but not in supernatants collected from infected C57BL/6 microglia and macrophages, or in supernatants from mock-infected SJL/J and C57BL/6 cells. Similarly, incubation of E20.1 cells, an immortalized line of oligodendrocytes, with infected SJL/J, but not C57BL/6 supernatants, resulted in cytotoxic activity. When cells from resistant C57BL/6 mice were treated with LPS, they became susceptible to infection and also secreted proteolytic enzymes. The proteolytic activity released from infected microglia and macrophages was found to be dose-dependent, was inactivated by heat, and was inhibited by phenylmethylsulphonyl fluoride (PMSF). These results indicate that a serine protease is released from infected microglia and macrophages and suggest a role for proteases in TMEV-induced myelin injury.

Topics: Animals; Cells, Cultured; Cytotoxicity Tests, Immunologic; Demyelinating Diseases; Disease Susceptibility; Endopeptidases; Immunity, Innate; Macrophages; Mice; Mice, Inbred C57BL; Microglia; Myelin Basic Protein; Oligodendroglia; Theilovirus

Studies were conducted in experimental allergic neuritis (EAN) to evaluate the possible interaction of cellular and humoral immune mechanisms in the demyelinating process. EAN was induced in Lewis rats by passive transfer of T cells reactive to P2 myelin protein or by active immunisation with whole myelin. Animals were then given systemic antimyelin antibody or control serum and assessed clinically, electrophysiologically and with semiquantitative histological studies. Animals given intraperitoneal (i.p.) P2-reactive T cells and systemic antimyelin antibody developed much more severe disease than those given i.p. T cells alone (P < 0.001). In actively immunised animals, the addition of systemic antimyelin antibody did not significantly alter disease severity. We believe the more severe disease in animals receiving T cells and antimyelin antibody reflects synergy between cellular and humoral immune mechanisms whereby neural antigen-specific T cells breach the blood-nerve barrier, allowing demyelinating antibody access to the endoneurium. In EAN induced by active immunisation with whole myelin it is likely that both B and T cell activation occurs and that the more severe demyelination characteristic of this disease reflects the involvement of both humoral and cellular immunity.

Topics: Animals; Cattle; Demyelinating Diseases; Female; Immune Sera; Immunotherapy, Adoptive; Myelin Basic Protein; Myelin P2 Protein; Myelin Sheath; Neuritis, Autoimmune, Experimental; Rabbits; Rats; Rats, Inbred Lew; T-Lymphocytes

To compare effects of insulin-like growth factor I (IGF-I) and placebo treatment on lesions that resemble those seen during active demyelination in multiple sclerosis, we induced experimental autoimmune encephalomyelitis in Lewis rats with an emulsion containing guinea pig spinal cord and Freund's adjuvant. On day 12-13, pairs of rats with the same degree of weakness were given either IGF-I or placebo intravenously twice daily for 8 days. After 8 days of placebo or IGF-I (200 micrograms/day or 1 mg/day) treatment, the spinal cord lesions were studied by in situ hybridization and with immunocytochemical and morphological methods. IGF-I produced significant reductions in numbers and areas of demyelinating lesions. These lesions contained axons surrounded by regenerating myelin segments instead of demyelinated axons seen in the placebo-treated rats. Relative mRNA levels for myelin basic protein, proteolipid protein (PLP), and 2',3'-cyclic nucleotide 3'-phosphodiesterase in lesions of IGF-I-treated rats were significantly higher than they were in placebo-treated rats. PLP mRNA-containing oligodendroglia also were more numerous and relative PLP mRNA levels per oligodendrocyte were higher in lesions of IGF-I-treated rats. Finally, a significantly higher proportion of proliferating cells were oligodendroglia-like cells in lesions of IGF-I-treated rats. We think that IGF-I effects on oligodendrocytes, myelin protein synthesis, and myelin regeneration reduced lesion severity and promoted clinical recovery in this experimental autoimmune encephalomyelitis model. These IGF-I actions may also benefit patients with multiple sclerosis.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Cell Division; Demyelinating Diseases; Dose-Response Relationship, Drug; Encephalomyelitis, Autoimmune, Experimental; Gene Expression Regulation; Immunohistochemistry; In Situ Hybridization; Insulin-Like Growth Factor I; Male; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin Sheath; Oligodendroglia; Rats; Rats, Inbred Lew; RNA, Messenger; Spinal Cord; Transferrin

In vivo adoptive transfer of CD4+ T helper cell type 1 clones reactive with autologous myelin basic protein (MBP) may initiate an inflammatory demyelinating disease of the central nervous system called experimental autoimmune encephalomyelitis. Although MBP is also a component of peripheral nervous system (PNS) myelin, previous studies have failed to demonstrate inflammation in the PNS induced by MBP-reactive T cells. Here, we report on two MBP-specific T cell clones that preferentially initiate inflammatory and demyelinating peripheral neuritis when adoptively transferred to syngeneic recipients. The MBP epitope recognized by these clones spans the junction of exons 6 and 7 and, therefore, is present in the 21- and 18.5-kD but not the 14- and 17-kD MBP isoforms, in which exon 5 is spliced to exon 7. The data suggest that MBP may be processed and presented differently in the central nervous system and PNS, and they provide evidence for MBP as a potential target for autoimmune reactions in the PNS.

Topics: Amino Acid Sequence; Animals; Demyelinating Diseases; Female; Immunization, Passive; Mice; Mice, Inbred BALB C; Molecular Sequence Data; Myelin Basic Protein; Neuritis, Autoimmune, Experimental; Peptides; T-Lymphocytes

Experimental autoimmune encephalomyelitis (EAE) was induced in the Lewis rat by the passive transfer of a cytotoxic CD4+ T cell clone specific for the 72-89 peptide of guinea-pig myelin basic protein (MBP). Histological studies on rats with neurological signs showed that inflammation was present in the proximal peripheral nervous system (PNS), namely the spinal roots, as well as in the central nervous system (CNS). The main sites of demyelination were the spinal roots in the PNS, and the spinal cord root entry and exit zones in the CNS. The major involvement of the proximal PNS in autoimmune disease directed at MBP is in marked contrast to EAE induced by immunisation with myelin proteolipid protein, where the inflammation and demyelination are restricted to the CNS. These findings may have implications for the human inflammatory demyelinating diseases including multiple sclerosis, in which MBP is a putative target antigen.

Topics: Animals; CD4 Antigens; Clone Cells; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Guinea Pigs; Microscopy, Electron; Myelin Basic Protein; Peripheral Nervous System; Rats; Rats, Inbred Lew; Spinal Cord; Spinal Nerve Roots; T-Lymphocytes

Experimental autoimmune encephalomyelitis (EAE) is a model for multiple sclerosis (MS). However, MS is a chronic, relapsing and demyelinating disease, whereas EAE in rats is typically a brief and monophasic disorder showing little demyelination. We demonstrate here that DA rats develop severe, protracted and relapsing EAE (SPR-EAE) after a subcutaneous immunization at the tail base with syngeneic spinal cord and incomplete Freund's adjuvant (IFA). The neurological deficits were accompanied by demyelinating inflammatory lesions in the spinal cord, with infiltrating T lymphocytes and perivascular deposition of immunoglobulins and complement. The induction of SPR-EAE was associated with humoral autoreactivity to myelin oligodendrocyte glycoprotein (MOG) and cellular autoreactivity to the rat myelin basic protein (MBP) peptides 69-87 and 87-101. These two peptides, as well as whole rat MBP, were encephalitogenic. In conclusion, we believe that the presently described demyelinating SPR-EAE represents a useful model for MS.

Topics: Amino Acid Sequence; Animals; Autoantibodies; Demyelinating Diseases; Disease Models, Animal; Encephalomyelitis, Autoimmune, Experimental; Freund's Adjuvant; Guinea Pigs; Immunization; Immunohistochemistry; Injections, Subcutaneous; Molecular Sequence Data; Myelin Basic Protein; Myelin Proteins; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Peptides; Phenotype; Rats; Rats, Inbred Strains; Recurrence; Reproducibility of Results; Spinal Cord; Time Factors

Calpain secreted by lymphoid (MOLT-3, M.R.) or monocytic (U-937, THP-1) cell lines activated with PMA and A23187 degraded myelin antigens. The degradative effect of enzymes released in the extracellular medium was tested on purified myelin basic protein and rat central nervous system myelin in vitro. The extent of protein degradation was determined by SDS-PAGE and densitometric analysis. Various proteinase inhibitors were used to determine to what extent protein degradation was mediated by calpain and/or other enzymes. Lysosomal and serine proteinase inhibitors inhibited 20-40% of the myelin-degradative activity found in the incubation media of cell lines, whereas the calcium chelator (EGTA), the calpain-specific inhibitor (calpastatin), and a monoclonal antibody to m calpain blocked myelin degradation by 60-80%. Since breakdown products of MBP generated by calpain may include fragments with antigenic epitopes, this enzyme may play an important role in the initiation of immune-mediated demyelination.

Topics: Animals; Antibodies, Monoclonal; Calcimycin; Calcium; Calcium-Binding Proteins; Calpain; Chelating Agents; Culture Media, Conditioned; Demyelinating Diseases; Egtazic Acid; Humans; Leukemia-Lymphoma, Adult T-Cell; Lymphoma, Large B-Cell, Diffuse; Monocytes; Myelin Basic Protein; Myelin Sheath; Neoplasm Proteins; Protease Inhibitors; Rabbits; Rats; T-Lymphocytes; Tetradecanoylphorbol Acetate; Tumor Cells, Cultured

Matrix metalloproteinases (MMPs) are a group of enzymes responsible for the degradation of interstitial connective tissue and basement membrane. The coding sequences for five of the human MMPs, viz. interstitial collagenase, 72 kDa gelatinase, stromelysin-1, matrilysin and 92 kDa gelatinase, were cloned and expressed in Chinese hamster ovary cells, and the proteins purified. The enzymes were compared for their ability to digest myelin basic protein, the major extrinsic membrane protein of central nervous system myelin. The most active on this substrate was 72 kDa gelatinase, followed by stromelysin-1; interstitial collagenase, matrilysin and 92 kDa gelatinase were of comparable but lesser activity. Production of these enzymes by glia or infiltrating inflammatory cells could therefore contribute to demyelination in neuroinflammatory disease.

Topics: Animals; CHO Cells; Cricetinae; Demyelinating Diseases; Humans; Metalloendopeptidases; Myelin Basic Protein

We studied the shiverer mouse as a model for correcting hearing disorders resulting from genetic abnormalities of the central nervous system (CNS). Shiverer mice are homozygous for an autosomal recessive mutation (deletion) in the gene for myelin basic protein (MBP), a major protein component of the myelin sheath in the CNS. Under electron microscopic observation of the cochlear nerve, the CNS portion in shiverer mice showed hypomyelination, but the peripheral portion, including spiral ganglion cells, was normal. We produced MBP-transgenic mice by microinjection of an MBP cosmid clone into the pronucleus of fertilized eggs from shiverer mice. The transgenic mice were found to recover MBP levels up to 25% of normal. A greater number of axons in the transgenic mice were myelinated than in the shiverer mice, but the myelin sheath was not as thick as in normal controls. Every interpeak latency of brain stem auditory-evoked potentials was prolonged in the shiverer mice and improved in the transgenic mice. This study provides an example of gene therapy for hearing disorders caused by a CNS abnormality. We discuss some strategies for researching genetic hearing impairment or deafness in both animals and humans.

Topics: Animals; Auditory Threshold; Central Nervous System; Cochlear Nerve; Demyelinating Diseases; Evoked Potentials, Auditory, Brain Stem; Gene Transfer Techniques; Hearing Loss, Sensorineural; Mice; Mice, Neurologic Mutants; Mice, Transgenic; Myelin Basic Protein; Reaction Time; Temporal Bone

In patients with inflammatory demyelinating neuropathy, which is possibly mediated by autoreactive, myelin-specific T lymphocytes, most studies focusing on immune responses to the major neuritogenic myelin protein P2 have been performed with bovine P2. However, the primary structure of bovine P2 differs from the human protein by nine amino acid residues that may profoundly influence the antigen recognition by T lymphocytes. We purified bovine and human P2 from peripheral nervous tissue and established a total of 19 T cell lines (TCL) reactive with bovine P2 from blood of two patients with acute Guillain-Barré syndrome (n = 5 TCL) and from six healthy individuals. Only four of these TCL, all raised from the blood of the GBS patients, transiently cross-recognized human P2 protein. Our results suggest that the use of human autoantigen may be crucial for the characterization of T cellular immune responses against P2 protein both in patients with inflammatory demyelinating neuropathy and in healthy controls.

Topics: Animals; Cattle; Cell Line; Chromatography, High Pressure Liquid; Demyelinating Diseases; Humans; Myelin Basic Protein; Myelin P2 Protein; Rats; Rats, Inbred Lew; T-Lymphocytes

The demyelinative potential of the cytokines interleukin-1 alpha (IL-1 alpha), interferon-gamma (IFN-gamma), and tumor necrosis factor-alpha (TNF-alpha) has been investigated in myelinating aggregate brain cell cultures. Treatment of myelinated cultures with these cytokines resulted in a reduction in myelin basic protein (MBP) content. This effect was additively increased by anti-myelin/oligodendrocyte glycoprotein (alpha-MOG) in the presence of complement. Qualitative immunocytochemistry demonstrated that peritoneal macrophages, added to the fetal telencephalon cell suspensions at the start of the culture period, successfully integrated into aggregate cultures. Supplementing the macrophage component of the cultures in this fashion resulted in increased accumulation of MBP. The effect of IFN-gamma on MBP content of cultures was not affected by the presence of macrophages in increased numbers.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Antibodies, Monoclonal; Astrocytes; Biomarkers; Blood Physiological Phenomena; Cell Aggregation; Cells, Cultured; Complement System Proteins; Culture Media, Serum-Free; Cytokines; Demyelinating Diseases; Glutamate-Ammonia Ligase; Guinea Pigs; Interferon-gamma; Interleukin-1; Macrophages, Peritoneal; Membrane Glycoproteins; Mice; Myelin Basic Protein; Myelin Proteins; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Nerve Tissue Proteins; Oligodendroglia; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Telencephalon; Tumor Necrosis Factor-alpha

Low density lipoprotein (LDL), the major carrier of plasma cholesterol, may enter the parenchyma of early multiple sclerosis (MS) lesions as a result of blood-brain barrier damage. We have used antibodies against LDL and epitopes found in LDL oxidized by two peroxidative end-products, malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE), to immunocytochemically stain MS plaques at different stages of pathology. Native LDL, epitopes of MDA-LDL, peptides of myelin basic protein and neutral lipid oil red O (ORO) staining were found to be co-localized within foamy macrophages in early and actively demyelinating MS plaques. Thus cholesterol esters, which are seen as Maltese crosses under polarized light in a proportion of foamy macrophages, appear to be derived from both LDL and myelin. ORO-negative astrocytes were strongly stained with the antibodies against 4-HNE-LDL and MDA-LDL, suggesting uptake of oxidatively modified protein products alone. Our findings suggest that a large proportion of the plasma LDL which enters the parenchyma of MS plaques is oxidatively modified in the lesion. Lipid peroxidation and oxidized LDL uptake by activated microglia and infiltrating macrophages in the early stages of MS plaque development may play important roles in demyelination.

Topics: Adult; Aged; Aldehydes; Astrocytes; Demyelinating Diseases; Humans; Immunohistochemistry; Lipoproteins, LDL; Macrophages; Malondialdehyde; Middle Aged; Multiple Sclerosis; Myelin Basic Protein; Oxidation-Reduction

A Japanese boy with the typical manifestations of 18q-syndrome and delayed myelination on magnetic resonance imaging is described. Cytogenetic investigation revealed a deletion at 18q21.3. Three serial magnetic resonance images demonstrated that myelination in the central nervous system was delayed except for the corpus callosum and brainstem. This pattern of delayed myelination appears to be peculiar to the 18q- syndrome. Because the gene for myelin basic protein has been localized to the distal end of the long arm of chromosome 18, we speculate that the abnormal myelination in our patient was partly due to the failure of expression of the myelin basic protein gene.

Topics: Brain; Brain Stem; Child, Preschool; Chromosome Aberrations; Chromosome Deletion; Chromosome Disorders; Chromosomes, Human, Pair 18; Corpus Callosum; Demyelinating Diseases; Developmental Disabilities; Glucuronidase; Humans; Magnetic Resonance Imaging; Male; Muscle Hypotonia; Myelin Basic Protein; Neurologic Examination

The twitcher (twi/twi) is an authentic murine model of human globoid cell leukodystrophy (GLD), caused by a deficiency of galactosylceramidase. Similar to human GLD, the twitcher shows progressive deterioration of neurological function and its neuropathology is characterized by a collection of periodic acid-Schift stain (PAS)-positive macrophages in the areas of demyelination. However, there are some differences in the clinico-pathological aspects between human and murine GLD. We investigated the spacio-temporal progression of neuropathology in the twitcher from postnatal day (PND) 10 to 45. No clinical symptoms or neuropathological changes were apparent in twi/twi until PND 15. Generally, infiltration of macrophages, concomitant with myelin degeneration, was recognized in the cerebellar white matter and the brain stem after PND 20, then in cerebral white matter after PND 25, and in cerebral and cerebellar gray matter after PND 30. The demyelination was very severe in the radix of the 8th and the 5th cranial nerves. The neurological symptoms such as tremor, spasticity and cranial nerve dysfunction were well correlated with the progression of pathological changes. Demyelination progressed in an orderly fashion such that myelin degeneration began 10 to 20 days after the commencement of myelination in any of the given nerve fiber tracts. This suggests that there are no significant differences in the metabolism of galactocerebroside in the myelin and myelin-forming cells in individual nerve fiber tracts throughout the murine brain. Over-expression of glial fibrillary acidic protein was already present before the initiation of obvious demyelination.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Aging; Animals; Animals, Newborn; Brain; Demyelinating Diseases; Leukodystrophy, Globoid Cell; Mice; Mice, Neurologic Mutants; Myelin Basic Protein; Periodic Acid-Schiff Reaction; Spinal Cord; Time Factors

To assess the distribution of insulin-like growth-factor-related proteins during autoimmune CNS demyelination and remyelination, experimental autoimmune encephalomyelitis was produced by injecting Lewis rats with an emulsion containing guinea pig spinal cord and complete Freund's adjuvant. Tail weakness appeared at 10-12 days and was followed by hind and forelimb weakness. Paraplegia and incontinence were observed in some animals. From 8-40 days postinoculation (dpi), spinal cord sections were used to correlate lesion location and severity with mRNA distributions of insulin-like growth factor I (IGF-I), IGF-binding protein 2 (IGFBP-2), IGF-I-receptor (IGFR-I), glial fibrillary acidic protein (GFAP), and myelin basic protein (MBP). These were determined semiquantitatively by in situ hybridization. Fourteen dpi, there were inflammatory infiltrates and demyelination in both white matter (WM) and grey matter (GM). IGF-I and GFAP mRNAs were increased in these lesions and transcripts encoding myelin basic protein (MBP) were greatly reduced. Large lesions with extensive demyelination were evident in both WM and GM when mRNA levels of GFAP and IGF-I peaked 26 dpi. MBP mRNA levels began increasing 21 dpi and peaked 26 dpi, when a few thin regenerating myelin sheaths were found morphologically. Astrocytes, identified by their morphology and GFAP immunoreactivity, expressed very low levels of IGFBP-2 mRNA and peptide in normal controls; their levels were significantly higher 14 dpi, peaked 26 dpi, and then gradually decreased. Some neurons, as well as oligodendroglia in areas undergoing remyelination, expressed IGFR-I. Although levels of IGF-I, IGFBP-2, and GFAP mRNAs were highest in lesion areas, levels were also elevated around lesions and in some normal-appearing areas of WM and GM 14-40 dpi. The gene expression of both IGF-I and IGFBP-2 by hypertrophic GFAP-positive astrocytes was demonstrated 14-40 dpi by combined in situ hybridization and immunocytochemistry as well as by double immunostaining. Coexpression of IGF-I and IGFBP-2 in the same astrocyte was a frequent finding. Relative increases in both IGF-I, GFAP, IGFBP-2, IGFR-I, and MBP mRNAs peaked at about the same time. This suggests that during lesion progression and recovery, astrocytic expression of IGF-I-related peptides may reduce immune-mediated myelin injury. We also suggest that astrocytic IGFBP-2 in lesions may help target IGF-I to IGFR-I-expressing oligodendrocytes and promote remyelination of dem

Topics: Animals; Astrocytes; Base Sequence; Carrier Proteins; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Glial Fibrillary Acidic Protein; Guinea Pigs; Immunohistochemistry; In Situ Hybridization; Insulin-Like Growth Factor Binding Protein 2; Insulin-Like Growth Factor I; Molecular Probes; Molecular Sequence Data; Myelin Basic Protein; Rats; Rats, Inbred Lew; Receptors, Somatomedin; RNA, Messenger

Degradation of purified myelin basic protein (MBP) was studied by SDS gel electrophoresis after addition of CSF samples obtained from HIV-1-infected patients. An increase in MBP degradation was detected in patients with neurological complications, such as AIDS dementia complex (ADC) or progressive multifocal leukoencephalopathy (PML), when compared with patients with no neurological symptoms (NA) or with other neurological opportunistic infections (OI). In the ADC and PML patients, in addition to CSF proteolytic activity, an increase in CSF-MBP levels and presence of white matter lesions were also observed by neuroimaging (MRI). In other opportunistic infections of the brain, MBP levels but not anti-MBP proteolytic activity increased. Results suggest the involvement of proteases in the virus-induced demyelination.

Topics: Adult; AIDS Dementia Complex; AIDS-Related Opportunistic Infections; Cerebrospinal Fluid; Demyelinating Diseases; HIV Infections; HIV-1; Humans; Leukoencephalopathy, Progressive Multifocal; Myelin Basic Protein; Nervous System Diseases; Peptide Hydrolases

A transgenic mouse containing 70 copies (ND4) of the transgene encoding DM20, a myelin proteolipid protein, appeared clinically normal up to 3 months of age. By 8-10 months, it showed tremors, unsteady gait, and died shortly thereafter. We concluded that the clinical symptoms correlated with demyelination based on the following criteria: 1) at 10 months of age only 17% of the amount of myelin obtained from normal mice was isolated from the ND4 mice; 2) astrogliosis, a prominent feature of demyelinating disease was minimal at 3 months of age but prominent by 10 months; 3) at the electron microscopic level disrupted myelin was seen at 8 months of age in the ND4 mice and ingested myelin debris was found in astrocytes; 4) lymphocytic infiltration in association with endothelial cells was observed routinely in the ND4 mice; 5) sections through optic nerves showed denuded and thinly myelinated axons in the 8 month old ND4 mice. Although the mechanism by which demyelination takes place is not fully understood, measurements of the amounts of PLP suggest it is down-regulated by the large amount of DM20. Since DM20 is a major proteolipid in the young but a minor one in the adult, the persistence of high levels in the adult results in improperly assembled myelin which is prone to disruption. Therefore demyelination in the ND4 mouse appears to result from the persistence of immature myelin into the adult.

Topics: Animals; Astrocytes; Axons; Demyelinating Diseases; Disease Models, Animal; Glial Fibrillary Acidic Protein; Immunohistochemistry; Mice; Mice, Transgenic; Microscopy, Electron; Multiple Sclerosis; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin Sheath; Nerve Tissue Proteins; Optic Nerve; Proteolipids

We report an autopsied case of paraneoplastic necrotizing myelopathy. The patient had bilateral blindness, quadriplegia, and dyspnea of acute onset and died without remission 7 weeks later. The severe tissue necrosis and demyelination were found in the optic chiasm and from the medulla oblongata throughout the whole length of spinal cord. A papillary carcinoma was found in the thyroid gland at autopsy. In the present case IgG, myelin basic protein and activated helper T cells were increased in the CSF at onset, suggesting a mechanism of autoimmune demyelination for the condition.

Topics: Autoimmune Diseases; Autopsy; Carcinoma, Papillary; Demyelinating Diseases; Female; Humans; Immunoglobulin G; Medulla Oblongata; Middle Aged; Myelin Basic Protein; Necrosis; Optic Chiasm; Paraneoplastic Syndromes; Spinal Cord; Thyroid Neoplasms

Mast cells are known for their participation in immediate and, more recently, delayed hypersensitivity reactions. They have been found in the meninges and certain brain areas where they are strictly perivascular, in close apposition to neurons, and they are activated by direct nerve stimulation or by neuropeptides. Intracranial mast cells contain many vasoactive substances which can increase the permeability of the blood-brain barrier, proteolytic enzymes which can degrade myelin in vitro, as well as chemotactic molecules which can attract inflammatory molecules in vivo. Connective tissue mast cells, with which intracranial mast cells share many characteristics, contain cytokines which can cause inflammation directly. Multiple sclerosis is a human demyelinating disease of unknown etiology, with a high prevalence in women which results in penetration of blood-borne immune cells within the brain parenchyma and subsequent destruction of myelin. Here, we report that 17 beta-estradiol and myelin basic protein, a major suspected immunogen in multiple sclerosis, had a synergistic action on inducing mast cell secretion. This effect was more pronounced in Lewis rats, which are susceptible to the development of experimental allergic encephalomyelitis, an animal model for multiple sclerosis, than in Sprague-Dawley rats, which are fairly resistant. Moreover, 18 h incubation of purified peritoneal mast cells with homogeneic slices of brain white matter in the presence of 17 beta-estradiol and myelin basic protein resulted in myelin changes resembling early stages of brain demyelination, which were also more evident in Lewis rats than in Sprague-Dawley rats. These results support the notion that mast cells could participate in the pathophysiology of demyelinating diseases.

Topics: Animals; Brain; Demyelinating Diseases; Drug Synergism; Estradiol; Image Processing, Computer-Assisted; In Vitro Techniques; Male; Mast Cells; Microscopy, Electron; Myelin Basic Protein; Myelin Sheath; Rats; Rats, Inbred Lew; Rats, Sprague-Dawley

Myelin basic protein (MBP), a major protein of myelin, is thought to play an important role in myelination, which occurs postnatally in mouse. Here we report that the MBP gene is expressed from the 12th embryonic day in mouse brain and that most of the predominant embryonic isoforms are not those reported previously. These isoforms have a deletion of a sequence encoded by exon 5 from the well-known isoforms. These isoforms show a unique developmental profile, i.e., they peak in the embryonic stage and decrease thereafter. In jimpy, a dysmyelinating mutant, the level of these isoforms remains high even in the older ages. These results suggest that MBPs have heretofore unknown functions unrelated to myelination before myelinogenesis begins. The possible presence of 18 isoforms of MBP mRNA, which are classified into at least three groups with different developmental profiles, is also reported here.

Topics: Animals; Base Sequence; Brain; Cloning, Molecular; Demyelinating Diseases; Exons; Galactosylceramides; Gene Expression; Male; Mice; Mice, Inbred ICR; Mice, Neurologic Mutants; Molecular Sequence Data; Mutation; Myelin Basic Protein; Nucleic Acid Hybridization; Oligodendroglia; Polymerase Chain Reaction; Promoter Regions, Genetic; RNA, Messenger

To study the effect of SV40 large T-antigen expression in myelin-forming cells of both the central and peripheral nervous system, a series of transgenic mice were generated expressing the SV40 large T-antigen under control of the myelin basic protein (MBP) promoter. Two neurologic phenotypes, designated A and B, appeared among individual transgenic founders and their progeny. The A mice developed a severe action tremor at about 10 days of age that progressed into periods of convulsions and early death by three to four weeks of age. In contrast, the B mice exhibited a progressive hindlimb ataxia and had a more normal lifespan. The A mice displayed hypomyelinating lesions in the central nervous system (CNS), whereas the B mice had lesions in either the peripheral nervous system (PNS) alone or in both the PNS and CNS. Immunohistochemical staining of spinal cord sections of a type A mouse showed a substantial depletion in MBP. Moreover, T-antigen-positive cells appeared predominantly in white matter tracts as randomly distributed single cells. Double labeling immunocytochemistry demonstrated that some of these T-antigen-positive cells were positive for oligodendrocyte differentiation markers MBP and O4. Thus, T-antigen expression appeared to coincide with a terminal stage of oligodendrocyte differentiation.

Topics: Animals; Antigens, Polyomavirus Transforming; Ataxia; Central Nervous System; Cloning, Molecular; Demyelinating Diseases; DNA; Female; Immunohistochemistry; Male; Mice; Mice, Inbred C57BL; Mice, Inbred DBA; Mice, Transgenic; Microscopy, Electron; Myelin Basic Protein; Myelin Sheath; Oligodendroglia; Peripheral Nerves; Phenotype; Promoter Regions, Genetic; Spinal Cord; Tremor

Topics: AIDS-Related Opportunistic Infections; Demyelinating Diseases; Humans; Myelin Basic Protein; Toxoplasmosis, Cerebral

Populations of phagocytic cells in multiple sclerosis (MS) plaques were examined by quantitative immunocytochemical analysis of macrophage markers and myelin degradation products in serial cryostat sections from 10 cases of MS. Around lesions with ongoing demyelination expression of the Class II antigen HLA-DQ appeared to be a marker of microglial activation. Alpha 1-antichymotrypsin+ monocytes and myelin-laden macrophages expressing the later differentiation markers Ber-MAC3 and RFD7 were predominantly perivascular in location. On the basis of the distribution of oil red O (ORO)+ phagocytes and myelin loss, plaques were divided into groups representing different stages in lesion development. In early lesions (group 1), there was no apparent myelin loss around ORO+ macrophages although these cells contained material stained with antibodies against myelin basic protein (MBP) epitopes and neoepitopes. However, patchy myelin loss was detectable around the phagocytic macrophages uniformly distributed throughout group 2 plaques. ORO+ macrophages containing MBP peptides were confined to the hypercellular border of group 4 lesions, in which the demyelinating process may be recurrent.

Topics: Adult; Biomarkers; Demyelinating Diseases; HLA-DQ Antigens; Humans; Immunohistochemistry; Macrophages; Middle Aged; Monocytes; Multiple Sclerosis; Myelin Basic Protein; Phagocytosis

There is suggestive but inconclusive evidence for a contribution of T cells and antimyelin antibodies to the pathogenesis of the Guillain-Barré polyneuropathy. We have studied the potential synergism of cellular and humoral immunity in the adoptive transfer model of EAN. EAN was induced in Lewis rats by injecting varying doses of P2 peptide (SP26)-sensitized T lymphocytes. Disease severity was dose-dependent. The addition of intravenous GC-AB to a subclinical dose of SP26-sensitized T cells resulted in overt clinical disease and markedly enhanced demyelination. Intravenous injection of antibody alone had no effect. We conclude that activated neuritogenic T cells, while entering into peripheral nerves, alter the blood-nerve barrier, which gives circulating demyelinating antibodies access to the endoneurium. The observations support the concept of a synergistic role of T-cell autoimmunity and humoral responses in the inflammatory demyelination of Lewis rat EAN.

Topics: Animals; Antibodies; Axons; Demyelinating Diseases; Edema; Female; Galactosylceramides; Ganglia, Spinal; Immunization, Passive; Lymphocyte Activation; Macrophages; Male; Motor Neurons; Myelin Basic Protein; Myelin P2 Protein; Nerve Degeneration; Neurilemma; Neuritis, Autoimmune, Experimental; Neurons, Afferent; Rabbits; Rats; Rats, Inbred Lew; Sciatic Nerve; Spinal Cord; Spinal Nerve Roots; T-Lymphocytes

The goals of this study were to quantify myelin-associated changes in the brain following single doses of radiation and to determine their relationship to the dose limits that this tissue can tolerate. Mice developed a transient loss of balance 1 month after 60 Gy doses 250 kVp X-rays to the brain and 3-4 months after 30-45 Gy radiation, but not after lower doses. The symptoms were transient and lasted approximately 1 month. The ED50/300 for radiation-induced brain death, which occurred large between 200 and 240 days, was 32.4 Gy (29.1, 35.5 Gy, 95% confidence limit of mean). At the time that animals developed neurological symptoms, 3-4 months after irradiation with doses of 30-45 Gy, biochemical assays of myelin-associated proteins showed decreases in 2',3'-cyclic nucleotide phosphohydrolase (CNPase) and myelin basic protein (MBP) levels that were not seen with lower radiation doses. By 120-180 days, further dose-dependent decreases in both CNPase and MBP levels were found after 20-45 Gy irradiation that preceded and correlated with death. The correlation of the decrease in CNPase and MBP levels with the incidence of transient neurological malfunction and animal death, together with histological evidence, suggests that demyelination is responsible for these phenomena.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Brain; Demyelinating Diseases; Male; Mice; Myelin Basic Protein; Radiation Dosage; Specific Pathogen-Free Organisms; Survival Rate

Rumpshaker (rsh) is an X-linked mutation causing hypomyelination of the CNS of mice and has recently been identified as an allele of jimpy (jp). The mutation (known as jprsh) differs in several respects from other X-linked myelin mutants, including jp, in that mice have normal longevity, oligodendrocyte numbers are not decreased, and cell death is not a feature. Myelin sheaths are deficient in immunostainable PLP protein. The present study examines the developmental expression of the major myelin protein genes and translatability of PLP and MBP mRNA. Differences between the spinal cord and brain of mutants are evident in that mRNA levels are more markedly decreased in the brain. Protein levels are severely reduced in both locations and to a proportionately greater extent than the mRNA, particularly in the spinal cord where PLP RNA and protein are approximately 80% and 10-20%, respectively, of age-matched wild type mice. DM-20 protein, the other major product of the PLP gene, is disproportionately expressed in rumpshaker as is a 10 kDa proteolipid. In vitro translation studies indicate a marked decrease in PLP translation products from mutant RNA. There is no deficiency in the number of PLP mRNA-expressing oligodendrocytes although the abundance per cell is reduced. The data suggest that the phenotypic effects of the mutation may be associated with reduced translation of major myelin proteins, in particular PLP and its incorporation into compact myelin. However, the mutation is compatible with survival of oligodendrocytes and their differentiation to the stage of expressing PLP/DM-20 mRNA.

Topics: Aging; Alleles; Animals; Blotting, Northern; Brain; Demyelinating Diseases; Gene Expression Regulation; Immunoblotting; Immunohistochemistry; In Situ Hybridization; Mice; Mice, Mutant Strains; Myelin Basic Protein; Myelin Proteins; Protein Biosynthesis; Proteolipids; RNA; RNA, Messenger; Spinal Cord; X Chromosome

It has previously been shown in the adult rat optic nerve that cells with many features of oligodendrocytes are capable of surviving for extended periods of time in the absence of axons. This is in contrast to the situation in the developing nervous system, where removal of axons leads to the failure of differentiation and to the death of oligodendrocytes. In the adult, these surviving oligodendrocytes were not typical in their appearance, and could only be identified with certainty using cell specific markers. In the present experiments, the functional capacity of these long-term quiescent cells to regenerate and myelinate was tested using the Shiverer mouse, a mutant lacking the gene for myelin basic protein (MBP), as a host animal. Fragments of optic nerve from adult rats which had been enucleated up to 2 years previously, were implanted into neonatal Shiverer mice. Four weeks later, the brains were removed and the formation of myelin investigated with antibodies to MBP, to ensure that this was of donor origin. Axons were demonstrated to have grown into the implants, and may have provided the stimulus for the production of MBP by the oligodendrocytes, which were stained positively within the implant. Myelin was demonstrated both within and adjacent to the implant. This study indicates that in the adult central nervous system, cells can survive for extended periods of time in the absence of axons, albeit in an inactive state, and are then capable of functional regeneration when placed in contact with unmyelinated axons. The origin of these cells, either from surviving oligodendrocytes which had previously myelinated the axons, or from progenitors lying within the adult nerve is unclear. The implications of these results are of importance in the further investigation of the potential for central nervous system regeneration.

Topics: Animals; Axons; Brain; Brain Tissue Transplantation; Demyelinating Diseases; Immunohistochemistry; Male; Mice; Mice, Neurologic Mutants; Microscopy, Immunoelectron; Myelin Basic Protein; Myelin Sheath; Oligodendroglia; Optic Nerve; Rats; Transplantation, Heterologous; Wallerian Degeneration

Myelin basic protein (MBP) was measured in cerebrospinal fluid (CSF) of patients with acquired immunodeficiency syndrome (AIDS) dementia complex (ADC) in order to investigate the degree of white matter destruction. Results show that increased CSF levels of MBP were detected in all patients with severe ADC (10/10) and, less often, in subjects with mild (2/7) or moderate dementia (7/16). No evidence of MBP-elevated concentration was observed in 14 human immunodeficiency virus (HIV)-seropositive subjects without neurological disorders and in nine HIV-seronegative controls. Our findings suggest that the measurement of CSF MBP concentration may represent a predictive marker of myelin injury and neurologic damage during the course of ADC.

Topics: Adult; Aged; AIDS Dementia Complex; Biomarkers; Demyelinating Diseases; Female; Humans; Male; Middle Aged; Myelin Basic Protein

To investigate insulin-like growth factor I (IGF-I) and IGF-I receptor gene expression during experimental demyelination and myelin regeneration, young mice were fed cuprizone (( bis(cyclohexanone) oxaldihydrazone )). This copper-chelating agent produces demyelination in the corpus callosum and superior cerebellar peduncles, and when treatment is stopped, there is rapid remyelination. At intervals during cuprizone treatment and recovery, brain sections were hybridized with specific probes and immunostained with antibodies to determine the localization and relative amounts of IGF-I and IGF-I receptor mRNAs and peptides. In untreated littermates, IGF-I and IGF-I receptor mRNAs and peptides were not detected in white matter. In cuprizone-treated mice, high levels of both IGF-I mRNA and peptide were expressed by astrocytes in areas of myelin breakdown. Astrocyte IGF-I expression decreased rapidly during recovery and oligodendroglial expression of myelin-related genes increased. In severely demyelinated areas, immature oligodendroglia exhibited a transient increase in IGF-I receptor mRNA and peptide immunoreactivity during early recovery. This highly specific pattern of IGF-I induction in astrocytes during demyelination and the expression of the IGF-I receptor in regenerating oligodendrocytes during recovery suggest that IGF-I functions in the regulation of oligodendrocyte and myelin metabolism in vivo.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Astrocytes; Cuprizone; Demyelinating Diseases; Gene Expression; Glial Fibrillary Acidic Protein; Immunoenzyme Techniques; Insulin-Like Growth Factor I; Male; Mice; Myelin Basic Protein; Nerve Tissue Proteins; Nucleic Acid Hybridization; Proteolipids; Receptors, Cell Surface; Receptors, Somatomedin; RNA, Messenger

We examined the development of the inflammatory cellular response and demyelination in P2 protein-induced experimental allergic neuritis (EAN). Collections of inflammatory cells were first identified in nerve roots 14 days after immunization. Ia+ cells predominated in the evolving lesions and T-helper cells were the dominant T-cell type with T-suppressor/cytotoxic cells appearing later in the course of the disease. Vesiculation, the earliest change seen in the myelin sheath, appeared before the wave of cellular infiltration. These results indicate that myelin injury precedes inflammation in P2 protein-induced EAN, and provide further evidence that this disorder is indistinguishable from EAN induced with whole peripheral nerve myelin.

Topics: Animals; Demyelinating Diseases; Inflammation; Male; Myelin Basic Protein; Myelin P2 Protein; Myelin Sheath; Neuritis, Autoimmune, Experimental; Rats; Rats, Inbred Lew; Spinal Nerve Roots

The expression of myelin-specific protein genes, i.e. myelin proteolipid (PLP), basic (BP), and myelin associated glycoproteins (MAG) was studied in normal and severely undernourished 20-day-old rats. The undernutrition paradigm resulted in reductions of approximately 50, 25 and 65% in body weight, brain weight and brain myelin yield, respectively. The amount of total brain RNA was not significantly altered, although the amount of cyclophilin (CYC) mRNA was increased. In contrast, the steady-state levels of myelin specific mRNAs were significantly decreased by approximately 40, 20 and 40% for PLP, BP and MAG, respectively. In addition, polyadenylation of the PLP transcript was altered, producing an abnormal ratio of the 1.6 kb to the 3.2 kb PLP mRNAs. The results indicate that down-regulation of myelin-specific gene expression is involved in the mechanisms of hypomyelination in hunger disease, although the individual genes are differently altered. Furthermore, undernutrition may have additional effects on the posttranscriptional processing of the transcripts as indicated by the abnormal size distribution of PLP messages.

Topics: Animals; Blotting, Northern; Body Weight; Brain; Demyelinating Diseases; DNA Probes; Down-Regulation; Female; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin Sheath; Myelin-Associated Glycoprotein; Nucleic Acid Hybridization; Nutrition Disorders; Organ Size; Pregnancy; Rats; RNA, Messenger

The X-linked mutation rumpshaker (rsh), which is probably an allele of jimpy (jp), causes hypomyelination in the CNS of mice. This study examines the developmental expression of the morphology, glial cells, and immunostaining of myelin proteins in the optic nerve and spinal cord. The optic nerve contains varying numbers of amyelinated and myelinated fibres. The majority of such sheaths are of normal thickness whereas in the spinal cord most axons are associated with a disproportionately thin sheath which changes little in thickness during development. In the optic nerve glial cell numbers are elevated in mutants during early and peak myelination but then fall slightly below normal in adults. In contrast, the number of glial cells is consistently elevated after 16 days of age in the spinal cord. The majority of the alterations to total glial cells are due to corresponding changes in the oligodendrocyte population. Immunostaining intensity is somewhat reduced for myelin basic protein (MBP) and the C-terminal common to proteolipid protein (PLP) and DM-20 and profoundly decreased for the PLP-specific peptide. Glial fibrillary acidic protein (GFAP) is increased in rsh. It is probable that some of the variation in myelination between optic nerve and cord in rsh is related to the difference in axon diameter in the two locations, as there are adequate numbers of oligodendrocytes at the time of myelination. However, the effect of the mutation on cell development in the brain and the spinal cord may be different. The immunostaining indicates a marked deficiency in PLP in myelin but suggests that DM-20 levels may be relatively normal. rsh shows several major differences from jp and other X-linked myelin mutants, particularly in relation to oligodendrocyte numbers, and will be useful to elucidate the role of the PLP gene in influencing oligodendrocyte differentiation and survival.

Topics: Animals; Demyelinating Diseases; Female; Glial Fibrillary Acidic Protein; Immunohistochemistry; Lipoproteins; Mice; Mice, Neurologic Mutants; Mutation; Myelin Basic Protein; Neuroglia; Oligodendroglia; Optic Nerve; Spinal Cord; X Chromosome

The mAb R73 directed to the TCR-alpha/beta of rat lymphocytes was tested for its therapeutic potential during the effector phase of experimental autoimmune neuritis (EAN) in Lewis rats. EAN can be actively induced by immunization with bovine peripheral nerve myelin, bovine P2 protein, or a peptide containing its neuritogenic epitope and serves as a model of the human Guilain-Barré syndrome. Adoptive transfer of activated P2-specific T lymphocytes also produces the monophasic disease (AT-EAN) characterized by inflammation and demyelination of peripheral nerves and highlights the central role of T lymphocytes in the pathogenesis of EAN. A single administration of the mAb R73 immediately after injection of activated P2-specific T line cells completely prevented the development of clinical and electrophysiologic signs of EAN in most animals and greatly alleviated the disease in the others. In further experiments mAb R73 was applied after the appearance of first clinical signs of EAN actively induced by immunization with a neuritogenic peptide or bovine peripheral nerve myelin. In both cases the anti-TCR-alpha/beta mAb reversed clinical signs of EAN and prevented the development of peripheral nerve dysfunction. In vivo and in vitro data suggest that impairment of Ag recognition and T cell function by occupancy of the TCR and R73-induced TCR-modulation rather than depletion of TCR-alpha/beta-bearing lymphocytes is the decisive mechanism underlying suppression of EAN that is apparent already within 48 h of the first R73 injection.

Topics: Animals; Antibodies, Monoclonal; Demyelinating Diseases; Endocytosis; Female; Immunosuppression Therapy; Immunotherapy; Myelin Basic Protein; Myelin P2 Protein; Neuritis, Autoimmune, Experimental; Rats; Rats, Inbred Lew; Receptors, Antigen, T-Cell, alpha-beta

We report evoked potential findings in a patient with 18q-syndrome (18q22.3----qter). The deletion included the locus for myelin basic protein (MBP). Clinical manifestations were mild intellectual deficit, involuntary movements and ataxia. MRI of the brain showed diffusely abnormal white matter. Visual evoked responses were normal. Central conduction was prolonged on median somatosensory evoked potentials and no central response was seen with posterior tibial somatosensory potentials. Putative congenital deficiency of MBP does not necessarily cause abnormal visual evoked responses.

Topics: Adult; Cerebral Cortex; Chromosome Deletion; Chromosome Mapping; Chromosomes, Human, Pair 18; Demyelinating Diseases; Electroencephalography; Evoked Potentials, Somatosensory; Evoked Potentials, Visual; Female; Humans; Myelin Basic Protein; Signal Processing, Computer-Assisted

A chronic relapsing model of demyelinating experimental allergic encephalomyelitis (EAE) was induced in Lewis rats by the repeated co-transfer of encephalitogenic, myelin basic protein (MBP)-specific T cells in combination with a demyelinating monoclonal antibody (mAb) specific for the myelin oligodendrocyte glycoprotein (MOG). In controls, repeated injections of 5 x 10(5) MBP-specific T cells at intervals of 18-21 days resulted in an increasing resistance to the induction of further episodes of EAE. However, intravenous injection of the mAb 4 days after each T cell transfer overcame this 'vaccination' effect and induced severe clinical relapses associated with an increasing and persistent neurological deficit. Histological examination revealed that four cycles of treatment with T cells and mAb were sufficient to result in the formation of large plaques of demyelination in the spinal cord that failed to undergo significant remyelination within 60 days of the final injection of mAb. These lesions consisted of a matrix of astrocytic scar tissue traversed by numerous naked axons. These observations demonstrate that the formation of large, persistently, demyelinated lesions in a T cell-mediated model of EAE in the Lewis rat is dependent on the presence of an appropriate anti-myelin autoantibody response.

Topics: Animals; Antibodies, Monoclonal; Cell Line; Central Nervous System Diseases; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Female; Membrane Glycoproteins; Myelin Basic Protein; Myelin Proteins; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Nervous System; Rats; Rats, Inbred Lew; T-Lymphocytes

Remyelination can be studied in aggregating rat brain cell cultures after limited demyelination. Demyelination was induced using a monoclonal antibody against myelin/oligodendrocyte glycoprotein (MOG mAb), in the presence of complement. De- and remyelination were assessed by measuring myelin basic protein (MBP). Two days after removing the MOG mAb, MBP levels reached 50% of controls and after 7 days 93%. During this period, cell proliferation determined by [14C]thymidine incorporation was similar in remyelinating and control cultures. Hormones and growth factors were tested for possible stimulatory effect on remyelinating cultures. Bovine growth hormone (bGH), triiodothyronine (T3), basic fibroblast growth factor (bFGF) and platelet-derived growth factor (PDGF) did not improve remyelination. Only epidermal growth factor (EGF) increased the level of remyelination. PDGF increased the rate of cell proliferation in both control and remyelinating cultures. A significant proportion of oligodendrocytes entered the cell division cycle and were not available for remyelination. The results obtained with PDGF and FGF (inhibition) support the idea that a pool of progenitor cells was still present and able to proliferate and differentiate into myelinating oligodendrocytes. The levels of myelin protein mRNAs were investigated during de- and remyelination. During demyelination, myelin protein mRNA levels decreased to approximately 50% of control cultures and returned to normal during remyelination. These preliminary results indicate that normal levels of gene transcription are sufficient to meet the increased need for newly synthesized myelin proteins during remyelination.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Brain; Cattle; Cell Aggregation; Cells, Cultured; Demyelinating Diseases; Growth Hormone; Growth Substances; Myelin Basic Protein; Myelin Sheath; Rats; RNA, Messenger; Triiodothyronine

Cerebrospinal fluid samples were obtained at lumbar puncture from 53 patients with a wide variety of neurological disorders. Cerebrospinal fluid samples were tested for the presence of P2 protein, a constituent of myelin, with an enzyme linked immunosorbent assay technique using a specific polyclonal antibody. High concentrations of P2 in the cerebrospinal fluid paralleled a raised IgG index (clearance ratio), the presence of oligoclonal bands, as well as raised white cell counts or depressed albumin:IgG ratios. Twenty one patients had been diagnosed as having definite or probable multiple sclerosis and the remaining 32 had other conditions. Of the 13 patients with high positive P2, 12 (92%) were in the multiple sclerosis category; of the 40 patients with low (12) or undetectable (28) P2 concentrations, only nine (23%) were diagnosed as having multiple sclerosis. In this patient population the presence of high immunoreactive P2 concentrations in cerebrospinal fluid was closely associated with evidence of intrathecal immunoglobulin synthesis and with the clinical diagnosis of multiple sclerosis. On this basis it is suggested that immunoassay of P2 concentration in the cerebrospinal fluid may be of potential value in the investigation of patients with demyelinating disorders.

Topics: Adolescent; Adult; Aged; Child; Child, Preschool; Demyelinating Diseases; Enzyme-Linked Immunosorbent Assay; Female; Humans; Immunoglobulin G; Infant; Male; Middle Aged; Multiple Sclerosis; Myelin Basic Protein; Myelin P2 Protein; Sensitivity and Specificity

The influence of experimental autoimmune encephalomyelitis (EAE) on the course and outcome of pregnancy, and the effect of pregnancy on EAE development, was investigated in rabbits and rats. Animals were immunized with encephalitogenic antigen in complete Freund's adjuvant (CFA) either before or during pregnancy. Abortion or fetal resorption was observed in most of the rabbits immunized before or during pregnancy, but not in pregnant rabbits injected with CFA or saline alone. Fetal loss was higher in those rabbits that developed clinical EAE. In rats, fetal loss occurred only when immunization was carried out during the first half of pregnancy. The appearance of EAE in pregnant rabbits, but not in rats, was delayed until after abortion or termination of pregnancy. The incidence of EAE in rabbits was lower, with milder severity and longer duration. Serum antibody levels to myelin basic protein, the autoantigen of EAE, was lower in pregnant rabbits, but not in rats, as compared to non pregnant animals. These results indicate that in species where pregnancy has a suppressive influence on the development of experimental autoimmune demyelinating disease, immunization with the neuroantigen induces a high rate of fetal loss.

Topics: Animals; Autoantibodies; Autoantigens; Autoimmune Diseases; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Female; Fetal Death; Fetal Resorption; Myelin Basic Protein; Pregnancy; Pregnancy Complications; Pregnancy Outcome; Rabbits; Rats

IL2-PE40 is a chimeric protein composed of human interleukin-2 (IL2) genetically fused to a modified form of Pseudomonas exotoxin lacking the cell recognition domain. IL2-PE40 is cytotoxic for IL2 receptor-bearing lymphocytes in culture and can inhibit activation of T cells in vivo. IL2-PE40 can significantly diminish antigen-stimulated proliferation of lymphocytes sensitized to myelin basic protein. Intraperitoneal administration of IL2-PE40 not only markedly inhibits the clinical manifestations of adoptively transferred relapsing experimental allergic encephalomyelitis but also dramatically reduces both inflammation and demyelination characteristic of the disease.

Topics: Animals; Bacterial Proteins; Brain; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Exotoxins; Female; Guinea Pigs; Immunotherapy, Adoptive; Immunotoxins; Interleukin-2; Lymphocyte Activation; Mice; Myelin Basic Protein; Pseudomonas aeruginosa; Receptors, Interleukin-2; Recombinant Proteins

(SJL/J x PL/J)F1 mice immunized with myelin basic protein (MBP) develop an autoimmune demyelinating disease termed relapsing experimental allergic encephalomyelitis (rEAE). The acute state of disease is mediated by CD4+ T cells specific for MBP amino acids 1-9. To determine the immunologic bases for disease relapse, host sensitization to additional autoantigens of the central nervous system was measured. Results indicate that most animals develop T cell reactivity to endogenous myelin proteolipid protein (PLP) during rEAE. However, PLP-specific immunity does not appear to account for expression of relapse episodes of demyelination.

Topics: Animals; Autoimmunity; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Histocompatibility Antigens Class II; Mice; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Recurrence; T-Lymphocytes

Two children with acute transverse myelopathy following adenovirus and Borrelia Burgdorferi infections are presented. The diagnosis stems from the clinical presentation, the determination of specific antibodies in serum and the favorable response to penicillin treatment in the case of neuroborreliosis. Both children made a good recovery. The cerebrospinal fluid examination showed a highly increased myelin basic protein concentration, indicating demyelination.

Topics: Adenovirus Infections, Human; Borrelia burgdorferi Group; Child; Demyelinating Diseases; Humans; Lyme Disease; Male; Myelin Basic Protein; Myelitis, Transverse; Penicillins

Trophic control over the expression and membrane distribution of voltage-dependent ion channels is one of the principal organizing events underlying the maturation of excitable cells. The myelin sheath is a major structural determinant of regional ion channel topography in central axons, but the exact molecular signals that mediate local interactions between the oligodendrocyte and axolemma are not known. We have found that large caliber fibre pathways in the brain of the mutant mouse shiverer (shi, gene on chromosome 18), whose developmental fate of myelination is averted by deletion of five exons in the myelin basic protein gene, have a striking excess of sodium channels. As cytoplasmic membranes of shiverer oligodendroglia still adhere to axons, the evidence indicates that myelin basic protein or a myelin basic protein-dependent glial transmembrane signal associated with compact myelin formation, rather than a simple glial-axon contact inhibition or an intrinsic genetic program of neuronal differentiation, could be critical in downregulating sodium channel density in axons. Here we use the shiverer mutant to show that mature central nervous system projection neurons with large caliber unmyelinated fibres sustain functional excitability by increasing sodium channel density. This axon plasticity, triggered by the absence of a single glial protein, contributes to the unexpectedly mild degree of neurological impairment in the mutant brain without myelin, and may be a potentially inducible mechanism determining the recovery of function from dysmyelinating disease.

Topics: Animals; Axons; Brain; Cell Membrane; Chromosome Deletion; Demyelinating Diseases; Exons; Mice; Mice, Neurologic Mutants; Myelin Basic Protein; Myelin Sheath; Nerve Fibers; Oligodendroglia; Saxitoxin; Sodium Channels

Myelin basic protein (MBP) was isolated from guinea-pig spinal cord in a form retaining the binding to all the myelin lipids. This new, lipid-bound and native-like preparation was used to immunize Lewis rats in complete Freund's adjuvant (CFA) in order to produce experimental allergic encephalomyelitis (EAE). The clinical features were compared with those of Lewis rats immunized with lipid-free MBP (LF-MBP), myelin, LF-MBP + octyl-POE (the non-ionic detergent used for the purification of LB-MBP) and octyl-POE alone. The clinical observation indicate that LB-MBP exerts an encephalitogenic activity on Lewis rats which is more intense than LF-MBP and includes demyelinating lesions in the central nervous system (CNS). The data suggest that LB-MBP is a new encephalitogenic antigen, which may induce more intensive immunization in rats and may be relevant in humans for autoimmune demyelinating diseases of the CNS.

Topics: Animals; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Guinea Pigs; Lipids; Myelin Basic Protein; Rats

Oral administration of proteins is a long-recognized method of inducing antigen-specific peripheral immune tolerance. We previously showed that oral administration of myelin basic protein suppresses monophasic experimental autoimmune encephalomyelitis in the Lewis rat when it is given in association with immunization and prior to disease onset. As a potential therapy for human autoimmune disease, it is crucial to determine whether oral tolerance can ameliorate an ongoing immune response. We therefore asked whether oral administration of myelin antigens, after sensitization and disease expression has occurred, could affect immunological, clinical, or pathological features of experimental autoimmune encephalomyelitis. Chronic relapsing experimental autoimmune encephalomyelitis was induced in the Lewis rat and strain 13 guinea pig by immunization with whole guinea pig cord homogenate, complete Freund's adjuvant, and Mycobacterium tuberculosis. Following recovery from the first attack, animals were orally given bovine myelin, guinea pig myelin, or guinea pig myelin basic protein three times per week for up to 3 months. Animals receiving myelin products orally had decreased severity and frequency of clinical relapses, decreased delayed-type hypersensitivity responses to myelin antigens, diminished inflammation in the central nervous system (CNS), and decreased areas of CNS demyelination. In the rat, guinea pig myelin basic protein was as effective as guinea pig myelin in ameliorating the disease and also resulted in decreased serum anti-myelin basic protein antibody levels. No exacerbation of disease or worsening of pathological findings occurred in the animals given myelin products. These results demonstrate that oral administration of myelin antigens can suppress chronic relapsing experimental autoimmune encephalomyelitis and have direct relevance to therapy of human demyelinating disorders such as multiple sclerosis.

Topics: Administration, Oral; Animals; Autoantigens; Autoimmune Diseases; Cattle; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Guinea Pigs; Hypersensitivity, Delayed; Immune Tolerance; Immunotherapy; Myelin Basic Protein; Myelin Proteins; Rats; Rats, Inbred Lew

Thymidine incorporation proliferation assays to whole bovine P2 protein and its 58-81 and 14-25 synthetic peptides were performed on blood mononuclear cells from ten patients with Guillain-Barré syndrome (GBS), six patients with chronic idiopathic demyelinating polyradiculoneuropathy (CIDP), and age and sex matched normal subjects. The only patients whose cells showed any response were two out of four with very early GBS. One responded to P2 and both synthetic peptides. One responded to P2 but to neither peptide. The results support a role for cell mediated immunity to P2 protein in some patients with Guillain-Barré syndrome.

Topics: Adult; Aged; Child; Chronic Disease; Demyelinating Diseases; Female; Humans; Lymphocyte Activation; Male; Middle Aged; Myelin Basic Protein; Myelin P2 Protein; Peptides; Polyradiculoneuropathy

We report here for the first time the occurrence of apoptosis of cells in the spinal cord in experimental allergic encephalomyelitis (EAE), an autoimmune, T-cell-mediated demyelinating disease. Four different forms of EAE were studied in the Lewis rat: (i) acute EAE induced by inoculation with whole spinal cord and adjuvants; (ii) acute EAE induced by inoculation with myelin basic protein (MBP) and adjuvants; (iii) acute EAE induced by the passive transfer of MBP-sensitized spleen cells; (iv) chronic relapsing EAE induced by inoculation with whole spinal cord and adjuvants followed by treatment with low-dose cyclosporin A. Cells undergoing apoptosis were recognized at light and electron microscopy by the presence of either crescentic masses of condensed chromatin lying against the nuclear envelope or rounded masses of uniformly dense chromatin. They were found in both the white and grey matter of the spinal cord in all 4 forms of this disease. Although it was not possible to identify definitively the types of cells undergoing apoptosis, the size and location of some of the affected cells suggested that they were oligodendrocytes. As there is now a large body of evidence that T-cell-induced target cell death takes the form of apoptosis, it is attractive to hypothesize that oligodendrocyte apoptosis is occurring in EAE as a result of oligodendrocyte-directed T-cell cytotoxicity. However, other apoptotic cells were located within the myelin sheath, meninges and perivascular spaces and were clearly not oligodendrocytes but were most likely blood-derived mononuclear cells. The sparsity of their cytoplasm and the absence of phagocytosed material suggested that they were mainly lymphocytes rather than macrophages. Apoptosis has been shown to be involved in deleting autoreactive T-cells during the normal development of tolerance. Thus apoptotic deletion of myelin/oligodendrocyte-specific lymphocytes in the central nervous system in EAE might explain both the subsidence of inflammation and the acquisition of tolerance in this autoimmune disease.

Topics: Animals; Cyclosporine; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Ganglia, Spinal; Immunization, Passive; Inflammation; Meninges; Microscopy, Electron; Myelin Basic Protein; Nerve Degeneration; Rats; Rats, Inbred Strains; Spinal Cord; Spleen

A common feature of demyelinating diseases such as multiple sclerosis in humans and experimental autoimmune encephalomyelitis in rodents is the marked elevation in the expression of the major histocompatibility complex (MHC) antigens in the involved sites. By specific targeting of a syngeneic MHC class I gene to oligodendrocytes, we have generated transgenic mice which not only exhibit severe involuntary tremors and develop tonic seizures but also show extensive demyelination in both the brain and the spinal cord. The fact that demyelination in these mice occurs in the absence of immune infiltration dismisses an autoimmune involvement but suggests that the MHC class I antigens play a direct role in inducing disease. Our findings lend support to the possibility that demyelinating diseases are induced by infectious agents such as viruses which can either directly activate MHC gene expression in oligodendroglia or indirectly activate expression through the release by reactive T cells of gamma interferon in the brain.

Topics: Animals; Base Sequence; Chimera; Demyelinating Diseases; Disease Models, Animal; DNA; DNA Probes; Female; Gene Expression; Genes, MHC Class I; Male; Mice; Mice, Transgenic; Molecular Sequence Data; Myelin Basic Protein; Oligonucleotides; Optic Nerve; Organ Specificity; Polymerase Chain Reaction; Transcription, Genetic

Major histocompatibility complex (MHC) molecules are not normally expressed in the central nervous system (CNS). However, aberrant expression has been observed in multiple sclerosis lesions and could contribute to the destruction of myelin or the myelinating cells known as oligodendrocytes. The mechanism of cell damage associated with aberrant MHC molecule expression is unclear: for example, overexpression of class I and class II MHC molecules in pancreatic beta cells in transgenic mice leads to nonimmune destruction of the cells and insulin-dependent diabetes mellitus. We have generated transgenic mice that express class I H-2Kb MHC molecules, under the control of the myelin basic protein promoter, specifically in oligodendrocytes. Homozygous transgenic mice have a shivering phenotype, develop tonic seizures and die at 15-22 days. This phenotype, which we term 'wonky', is due to hypomyelination in the CNS, and not to involvement of the immune system. The primary defect appears to be a shortage of myelinating oligodendrocytes resulting from overexpression of the class I MHC molecules.

Topics: Animals; Brain; Demyelinating Diseases; Gene Expression; H-2 Antigens; Mice; Mice, Transgenic; Multiple Sclerosis; Myelin Basic Protein; Oligodendroglia; Promoter Regions, Genetic; Spinal Cord

We describe the measurement of myelin basic protein gene transcription rate, and of the accumulation of both mature mRNA and total transcripts from the myelin basic protein gene in brains from mice of wild-type and homozygous shiverer genotypes at several ages spanning postnatal development. In wild-type brains the accumulation of total transcripts as well as mature mRNA, and the transcription rate, all follow the same general pattern of rising sharply from a low level at birth to a peak at 20 days, and continuing at a somewhat reduced level into adulthood. Thus a major factor in the developmental regulation of myelin basic protein expression is the control of transcription rate. The shiverer mutation consists of a deletion of the 3' end of the myelin basic protein gene which completely prevents production of mature mRNA and protein, and results in severe dysmyelination and a trembling behavior. In shiverer brains, the transcription rates for the intact 5' end of the gene follow closely those seen in wild-type animals up to the age at which maximal myelination normally occurs. Total myelin basic protein transcripts follow a similar profile but at less than 5% the level seen in wild-type, and, as expected, no mature mRNA is detected. Thus the shiverer deletion does not remove information required for efficient, developmentally regulated transcription, and the low level of myelin basic protein gene transcripts in this mutant must be a result of their reduced stability. A higher than normal myelin basic protein gene transcription rate in older shiverer animals raises interesting questions regarding the regulatory mechanisms controlling myelinogenesis.

Topics: Animals; Brain Chemistry; Chromosome Deletion; Demyelinating Diseases; Gene Expression Regulation; Mice; Mice, Neurologic Mutants; Myelin Basic Protein; Promoter Regions, Genetic; RNA, Messenger; Transcription, Genetic

Intracerebral inoculation of Theiler's murine encephalomyelitis virus (TMEV) into susceptible mouse strains produces a chronic demyelinating disease in which mononuclear cell-rich infiltrates in the central nervous system (CNS) are prominent. Current evidence strongly supports an immune-mediated basis for myelin breakdown, with an effector role proposed for TMEV-specific, major histocompatibility complex (MHC) class II-restricted delayed-type hypersensitivity (DTH) responses in which lymphokine-activated macrophages mediate bystander demyelination. The present study examined the possibility that concomitant or later-appearing neuroantigen-specific autoimmune T cell responses, such as those demonstrated in chronic-relapsing experimental allergic encephalomyelitis (R-EAE), may contribute to the demyelinating process following TMEV infection. T cell responses against intact, purified major myelin proteins (myelin basic protein (MBP) and proteolipid protein (PLP], and against altered myelin constituents were readily demonstrable in SJL/J mice with R-EAE, but were not detectable in SJL/J mice with TMEV-induced demyelinating disease. TMEV-infected mice also did not display T cell responses against the peptide fragments of MBP(91-104) and PLP(139-151) recently shown to be encephalitogenic in SJL/J mice. In addition, induction of neuroantigen-specific tolerance to a heterogeneous mixture of CNS antigens, via the i.v. injection of syngeneic SJL/J splenocytes covalently coupled with mouse spinal cord homogenate, resulted in significant suppression of clinical and histologic signs of R-EAE and the accompanying MBP- and PLP-specific DTH responses. In contrast, neuroantigen-specific tolerance failed to alter the development of clinical and histologic signs of TMEV-induced demyelinating disease or the accompanying virus-specific DTH and humoral immune responses. These findings demonstrate that TMEV-induced demyelinating disease can occur in the apparent absence of neuroantigen-specific autoimmune responses. The relationship of the present results to the immunopathology of multiple sclerosis is discussed.

Topics: Animals; Antibody Formation; Antibody Specificity; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Enterovirus Infections; Epitopes; Female; Immune Tolerance; Maus Elberfeld virus; Mice; Mice, Inbred Strains; Myelin Basic Protein; Myelin Sheath; Nervous System; Proteolipids; Spinal Cord; T-Lymphocytes

Myelin basic proteins (MBPs) from 6-day-old, 10-day-old, 20-day-old and adult normal mouse brain were compared with those from 20-day-old jimpy (dysmyelinating mutant) mouse brain to determine the effect of reduced levels of proteolipid protein (PLP) on MBPs. Alkaline-urea-gel electrophoresis showed that 6-day-old and 10-day-old normal and jimpy MBPs lacked charge microheterogeneity, since C8 (the least cationic of the components; not be confused with complement component C8) was the only charge isomer present. In contrast, MBPs from 20-day-old and adult normal mouse brain displayed extensive charge microheterogeneity, having at least eight components. A 32 kDa MBP was the major isoform observed on immunoblots of acid-soluble protein from 6-day-old and 10-day-old normal and 20-day-old jimpy mouse brain. There were eight bands present in 20-day-old and adult normal mouse brain. Purified human MBP charge heteromers C1, C2, C3 and C4 reacted strongly with rat 14 kDa MBP antiserum, whereas the reaction with human C8 was weak. This suggested that MBPs from early-myelinating and jimpy mice did not react to MBP antisera because C8 was the major charge isomer in these animals. Purification of MBPs from normal and jimpy brain by alkaline-gel electrophoresis showed that both normal and jimpy MBPs have size heterogeneity when subjected to SDS/PAGE. However, the size isoforms in normal mouse brain (32, 21, 18.5, 17 and 14 kDa) differed from those in jimpy brain (32, 21, 20, 17, 15 and 14 kDa) in both size and relative amounts. Amino acid analyses of MBPs from jimpy brain showed an increase in glutamic acid, alanine and ornithine, and a decrease in histidine, arginine and proline. The changes in glutamic acid, ornithine and arginine are characteristic of the differences observed in human C8 when compared with C1.

Topics: Aging; Amino Acids; Animals; Brain; Cations; Demyelinating Diseases; Electrochemistry; Electrophoresis; Electrophoresis, Polyacrylamide Gel; Immunoblotting; Mice; Mice, Jimpy; Molecular Weight; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein

An immunohistochemical study was performed to compare glial reactions in recent and old lesions of central pontine myelinolysis (CPM). Regions of demyelination and destruction of oligodendrocytes, showed reduced immunoreactivity of myelin basic protein (MBP), myelin-associated glycoprotein (MAG), transferrin, and carbonic anhydrase C (CA C). In addition, labeling of glial fibrillary acidic protein (GFAP) and S-100 protein revealed distinct dystrophic alterations of the astroglia. Remarkably, immunolabeling of GFAP was drastically reduced in astrocytic cytoplasm within freshly demyelinated lesions. Immunostaining of vimentin revealed a differential intracytoplasmic decoration of hypertrophic and dystrophic astrocytes in recent and old CPM lesions. Immunolabeling of desmin failed to stain glial cells. Monoclonal antibodies against HNK-1 exhibited greatly increased immunoreactivity both of persisting oligodendrocytes and of reactive fibrillary astrocytes in old CPM foci. In freshly demyelinated lesions, enhanced immunoreactivity of the X-hapten (3-fucosyl-N-acetyllactosamine) was prominent in astroglia and oligodendrocytes. Simultaneously, reactive astrocytes revealed intracytoplasmic labeling of laminin. Quantitation of GFAP+ astroglia in fresh CPM and control cases revealed an increase in the number of astrocytes within the demyelinated foci and in the surrounding non-demyelinated pontine tissue of CPM cases. The occurrence of astroglial alterations in the demyelinated foci of CPM could be interpreted as "astroglial dystrophy" which may represent a pathogenic factor in CPM. Furthermore, it is possible that changes of the glial microenvironment may influence the astroglia to revert transiently back to an immature phenotype as indicated by the enhanced expression of the X-hapten and HNK-1, and the de novo synthesis of vimentin and laminin.

Topics: Adult; Cell Count; Demyelinating Diseases; Female; Humans; Immunohistochemistry; Intermediate Filament Proteins; Male; Middle Aged; Myelin Basic Protein; Myelin Proteins; Myelin-Associated Glycoprotein; Oligodendroglia; Pons

To study the possible in vivo activity of experimental allergic neuritis (EAN) serum, we injected serum from rats immunized with whole nerve, P2 protein or adjuvant alone into the sciatic nerve of normal Lewis rats. Serum from whole nerve and P2-immunized animals produced demyelination 24 h after injection. Only high-titer anti-P2 serum was active and no control serum had this effect. Anti-P2 antibodies or other serum factors may contribute to the pathogenesis of whole nerve and P2-induced EAN.

Topics: Animals; Autoantibodies; Demyelinating Diseases; Immunization, Passive; Myelin Basic Protein; Myelin P2 Protein; Neuritis, Autoimmune, Experimental; Rats; Rats, Inbred Lew; Sciatic Nerve

A monoclonal antibody (8-18C5) directed against myelin/oligodendrocyte glycoprotein (MOG) induced demyelination in aggregating brain cell cultures. With increasing doses of anti-MOG antibody in the presence of complement, myelin basic protein (MBP) concentration decreased in a dose-related manner. A similar, albeit less pronounced, effect was observed on specific activity of 2',3'-cyclic nucleotide 3'-phosphohydrolase. In the absence of complement, anti-MOG antibody did not induce detectable demyelination. In contrast to the effect of anti-MOG antibody and as expected, anti-MBP antibody did not demyelinate aggregating brain cell cultures in the presence of complement. These results provide additional support to the suggestion that MOG, a quantitatively minor myelin component located on the external side of the myelin membrane, is a good target antigen for antibody-induced demyelination. Indeed, they show that a purified anti-MOG antibody directed against a single epitope on the glycoprotein can produce demyelination, not only in vivo as previously shown, but also in cultures. Such an observation has not been made with polyclonal antisera raised against purified myelin proteins like MBP and proteolipid protein, the major protein components of the myelin membrane, or myelin-associated glycoprotein. These observations may have important implications regarding the possible role of anti-MOG antibodies in demyelinating diseases.

Topics: 2',3'-Cyclic Nucleotide 3'-Phosphodiesterase; 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Antibodies, Monoclonal; Antigens; Brain; Cells, Cultured; Complement System Proteins; Demyelinating Diseases; Epitopes; Immunoglobulin G; Membrane Glycoproteins; Myelin Basic Protein; Myelin Proteins; Myelin Sheath; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Phosphoric Diester Hydrolases; Rats

This is the first description of acute demyelinating experimental autoimmune encephalomyelitis (EAE) induced in rats by myelin basic protein (BP)-specific T lymphocytes without the administration of demyelinating antibodies. BP-specific T cell lines were selected from inbred Buffalo-strain rats (Rt-1b) following techniques used to develop similar lines from Lewis rats (Rt-1l). Unlike those of Lewis rats, the spinal cords of Buffalo rats with T cell line-mediated EAE had prominent perivascular demyelination associated with mononuclear inflammation. Like Lewis rat lines. Buffalo rat BP-specific T cell lines transferred acute, non-relapsing EAE into syngeneic recipients, demonstrating that demyelination in passive acute EAE can occur without subsequent clinical relapses.

Topics: Animals; Autoimmune Diseases; Cell Line; Demyelinating Diseases; Encephalomyelitis; Female; Guinea Pigs; Hypersensitivity, Delayed; Immunotherapy, Adoptive; Myelin Basic Protein; Rats; Rats, Inbred BUF; Rats, Inbred Lew; Spinal Cord; T-Lymphocytes

A previous study from this laboratory on adoptively transferred experimental allergic encephalomyelitis (EAE) induced by myelin basic protein-responsive (MBP+) 11C-labeled lymphocytes showed that MBP+ cells entered the central nervous system (CNS) before signs, migrated through the endothelium, and remained within the perivascular space. The majority of cells effecting CNS damage were nonradiolabeled and appeared to be host-derived and non-CNS antigen specific. The present study defined the immunocytochemical and initial structural events occurring between lymphocytes and endothelial cells (EC) on CNS blood vessels during EAE induced with MBP+ lymph node cells or T-cell lines. Monoclonal antibodies against lymphocyte function-associated molecule LFA-1 and its ligand, intercellular adhesion molecule-1 (ICAM-1), and the addressin MECA-325, a marker of mouse lymph node high endothelial venules, were tested on frozen sections in combination with the avidin-biotin-complex technique. The attachment and infiltration of lymphocytes correlated with the onset of signs and the appearance in the CNS of MECA-325 and ICAM-1 on vessels with cellular infiltrates and sometimes with plump EC. The cellular infiltrates were composed largely of LFA-1+ lymphocytes. Ultrastructurally, pseudopodia from lymphocytes were seen to attach to and penetrate EC in the CNS and form small gap junction-like contacts. On the EC surface, some processes from lymphocytes made larger synapse-like contacts while others were associated with coated pits suggestive of receptor mediation. The results are in accord with specific homing and attachment of lymphocytes to the CNS vasculature being early features of the disease process in EAE and with some CNS vessels acquiring properties of lymph node elements. Understanding of the mechanisms underlying these lymphocyte/EC interactions has therapeutic import for multiple sclerosis, for which EAE is the prime model.

Topics: Animals; Autoimmune Diseases; Brain; Cell Adhesion; Cell Adhesion Molecules, Neuronal; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Endothelium, Vascular; Female; Mice; Microscopy, Electron; Myelin Basic Protein; T-Lymphocytes

To identify and assess the consequences of the mutation in myelin-deficient (md) rats, the myelin proteolipid protein (PLP) gene and its expression were studied in md rats. Southern blots of the PLP gene demonstrated that no major deletions or insertions have occurred in this gene. In addition, the mutation in this gene does not result in a splicing defect in the RNAs, since all exons are represented in md PLP RNAs. These data are consistent with results in another laboratory indicating that a point mutation in the PLP gene in md rats results in a single amino acid alteration in the protein. To elucidate the molecular mechanisms producing reduced levels of PLP, myelin basic protein (MBP) and glycerol phosphate dehydrogenase (GPDH) mRNAs, and their corresponding proteins in md rats, in vitro transcription assays were performed. Transcription of the PLP gene in nuclei isolated from 23-day-old md rat brains was dramatically reduced relative to normal tissue. Thus, the single amino acid alteration in this protein alters the regulation of transcription of this gene. In contrast, the transcriptional activities of the MBP and GPDH genes in md rats were indistinguishable from normal animals. Thus, the lower level of MBP and GPDH mRNA and protein in md rats relative to normal results from a posttranscriptional event.

Topics: Animals; Blotting, Southern; Cell Nucleus; Demyelinating Diseases; Exons; Glycerolphosphate Dehydrogenase; Male; Mice; Mice, Neurologic Mutants; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin Sheath; RNA, Messenger; Single-Strand Specific DNA and RNA Endonucleases; Transcription, Genetic

Batch purification of the myelin basic protein (MBP) in the lipid-bound form was obtained from bovine brain white matter by using the slightly polydisperse nonionic detergent, n-octyl-pentaoxyethylene (octyl-POE) and hydroxyapatite. This large-scale procedure can also be carried out in laboratories without chromatographic equipment, and is applicable to small amounts of myelin. More interestingly, removal and inhibition of the proteolytic activity associated with myelin allowed us to obtain more stable and intact forms of the protein when compared with MBP isolated in the lipid-bound form by our previous method. Since it retains binding to all myelin lipids, this purified MBP may be considered as being in a native-like form. In this article, we suggest why this more intact form of MBP could be used to advantage as an alternative to lipid-free, water-soluble MBP in the study, detection, and treatment of myelin damage in pathology.

Topics: Animals; Brain; Cattle; Chromatography, Thin Layer; Demyelinating Diseases; Electrophoresis, Polyacrylamide Gel; Indicators and Reagents; Lipid Metabolism; Molecular Weight; Myelin Basic Protein

In order to determine if free (F) and bound (B) levels of autoantibodies to myelin basic protein (anti-MBP) are present from the onset of multiple sclerosis (MS), 201 patients referred to our clinic were clinically divided into a group diagnosed as having an initial MS relapse and a group of non-MS controls. Ninety-four of 106 patients thought to have an initial MS relapse had increased CSF anti-MBP, while only 14 of 95 controls had elevated antibody levels; 9 of these 14 positive controls were subsequently shown to have MS by magnetic resonance imaging and/or clinical follow-up. CSF anti-MBP was more frequently abnormal than 3 estimates of intrathecal IgG synthesis in the group with suspected MS. Kinetics of F and B CSF anti-MBP were determined in a group of 29 patients with clinically definite MS during an acute relapse and 97.4 +/- 54 days later in the subsequent convalescent phase when in clinical remission. F and B anti-MBP levels were highly dependent on the timing of the CSF sampling; generally, as patients entered into clinical remission F anti-MBP declined, B antibody levels rose and F/B anti-MBP ratios initially above unity gradually declined towards zero. These data suggest that anti-MBP may be involved in the mechanism of MS.

Topics: Acute Disease; Autoantibodies; Convalescence; Demyelinating Diseases; Humans; Kinetics; Multiple Sclerosis; Myelin Basic Protein; Time Factors

We evaluated 130 consecutive HIV-infected neurologically asymptomatic individuals for intrathecal IgG production and myelin basic protein (MBP) levels. Although 56.7% of immunologically normal and 68.8% of immunocompromised patients had some CSF abnormality, no patient had an abnormal MBP level. The lack of MBP elevation in the CSF of these patients suggests that the production of intrathecal IgG is not related to active demyelination.

Topics: Acquired Immunodeficiency Syndrome; Acute Disease; Adult; Dementia; Demyelinating Diseases; Follow-Up Studies; Humans; Immunoglobulin G; Male; Myelin Basic Protein; Neurologic Examination

Three patterns of EAE with different morbidity and mortality rates were induced in the guinea pigs inoculated with various doses of tryptophane peptide (TP) and complete Freund's adjuvant. TP-sensitized animals manifested the delayed type hypersensitivity (DTH) reactions to TP and circulating anti-TP and -BPF antibodies were not found, polypeptide fraction of myelin basic protein (BPF while a A correlation was revealed between the DTH-reactions and EAE development. Intracutaneous TP and BPF injections at the early period before the EAE onset resulted in reduction of morbidity rate from 90 to 50 per cent. For the first time with the help of modified Marchi method demyelination has been shown to be highly marked in CNS tissue process in CNS can be caused by cell-mediated immune of animals given TP. It is supposed that the demyelinating reaction to encephalitogenic fragment of BP molecule. The data indicate a possibility of EAE inhibition by means of TP and BPF injections in saline solution.

Topics: Animals; Brain; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Guinea Pigs; Hypersensitivity, Delayed; Myelin Basic Protein; Peptides; Spinal Cord; Tryptophan

C57BI/6N mice develop a CNS demyelinating disease when inoculated intracranially at 4 weeks of age with the A59 strain of mouse hepatitis virus (MHV-A59). In order to explore the virus-host interactions, the histological features of the demyelinating disease were correlated with the spatial and temporal distribution of viral transcripts and the expression of oligodendrocyte-specific genes (myelin basic protein, proteolipid protein, myelin-associated glycoprotein, and 2',3' cyclic nucleotide 3'-phosphohydrolase) in the spinal cord of diseased mice. Three distinct phases in the disease were identified. In the first phase, 1 week postinfection (1 WPI), virus replication was widespread in both gray and white matter but was preferentially occurring in glial cells. In the ventral and dorsal root zones where viral transcripts were most abundant, all myelin gene transcripts were decreased before demyelination was seen. During the second phase of the disease (2-3 WPI), viral transcripts decreased in abundance and became restricted to the white matter. Numerous demyelinating lesions were observed and were characterized by inflammatory cells, paucity of oligodendrocytes, and a profound decrease of all myelin gene transcripts. In the third phase of the disease (4-6 WPI) no viral transcripts were detected, and remyelination began. In the lesions and the tissue surrounding them, transcripts of all myelin genes increased to levels above normal. The increased expression of myelin gene transcripts occurred in a synchronized manner and with a cellular distribution reminiscent of that seen in developmental myelination. These molecular events correlated with efficient remyelination and clinical recovery in this murine demyelinating disease.

Topics: Animals; Coronaviridae Infections; Demyelinating Diseases; Gene Expression Regulation; Gene Expression Regulation, Viral; Mice; Mice, Inbred C57BL; Myelin Basic Protein; RNA, Messenger; Spinal Cord

To test the hypothesis that T lymphocytes sensitized to central nervous system (CNS) antigens may quantitatively induce more demyelination in neural tissue than T cells sensitized to non-CNS antigens, we established T cell lines specific for myelin basic protein (MBP) or the purified protein derivative (PPD) of M. tuberculosis. The potential of T cells to cause myelin pathology was determined by measuring the activity of the myelin-associated enzyme 2'-3'-cyclic nucleotide 3'-phosphodiesterase (CNPase) in organotypic cultures of syngeneic spinal cord after incubation with the T cell lines. The activity of CNPase in neural tissue has been shown to correlate positively with the amount and integrity of CNS myelin. Although both MBP- and PPD-specific T cells caused decreases in CNPase activity, the MBP line caused significantly greater and consistent changes. This finding indicates that T cell-mediated CNS demyelination may be comprised of CNS antigen-specific and CNS non-specific components, the former causing more pathology.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Antigens, Surface; Cells, Cultured; Demyelinating Diseases; Mice; Mice, Inbred Strains; Myelin Basic Protein; Organ Culture Techniques; Spinal Cord; T-Lymphocytes; Tuberculin

We report the development of a simple and reliable method for the study of demyelination in vitro based on the measurement of 2':3'-cyclic nucleotide 3'-phosphodiesterase in isolated myelin. Using only small quantities of myelin (equivalent to 100 micrograms of myelin protein) the system was tested under conditions that are believed to approximate those found at the site of an inflammatory demyelinating lesion. Treatment with a combination of trypsin, phospholipase A2, and lysophosphatidylcholine was used to evaluate the method. This microsystem has the potential not only for testing the myelinotoxicity of soluble factors but also for investigating the involvement of inflammatory cells in the demyelinating process. Myelin degradation by elicited peritoneal macrophages could be demonstrated at relatively high densities of these cells. Nylon wool purified lymph node T cells from myelin basic protein-primed SJL/J mice, after selective expansion with antigen and interleukin 2, failed to induce any significant myelin breakdown unless a limited number of syngeneic activated macrophages were also present. T cells from mice that had been inoculated with keyhole limpet haemocyanin failed to show any effect. The advantages of this technique over other in vitro systems are that it enables the study of demyelination using syngeneic sources of myelin and defined cell populations.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Animals; Antigens; Demyelinating Diseases; Female; Hemocyanins; In Vitro Techniques; Lysophosphatidylcholines; Macrophages; Male; Mice; Mice, Inbred BALB C; Myelin Basic Protein; Myelin Sheath; Phagocytosis; Phospholipases A; Phospholipases A2; Sulfoglycosphingolipids; T-Lymphocytes; Trypsin

Topics: Animals; Brain; Demyelinating Diseases; Humans; Myelin Basic Protein; Peptide Fragments; Peptide Hydrolases; Rats

The new synthesis of myelin and the proliferation of oligodendrocytes was stimulated by serum from syngeneic mice immunized with homogenized spinal cord (SCH). Treatment with this antiserum produced a 10-fold increase in the area of remyelination in spinal cords that had become demyelinated previously as a result of infection by Theiler's murine encephalomyelitis virus. Inflammation was decreased in regions of white matter that showed remyelination. Oligodendrocytes exposed to anti-SCH in vitro incorporated three to five times more [3H]thymidine than resting cells did and expressed more myelin basic protein in their cytoplasm, suggesting stimulation of myelinogenesis. Thus, there is a factor present in anti-SCH antiserum that stimulates central nervous system-type remyelination. This finding may provide clues for the therapy of patients with demyelinating disorders such as multiple sclerosis.

Topics: Animals; Cell Division; Demyelinating Diseases; Female; Fluorescent Antibody Technique; Immunization, Passive; Maus Elberfeld virus; Mice; Multiple Sclerosis; Myelin Basic Protein; Myelin Proteins; Myelin Sheath; Neuroglia; Oligodendroglia; Spinal Cord; Thymidine

Beagle dogs were experimentally infected with the Cornell A75-17 strain of canine distemper virus. At three time points post-infection (PI), immunoreactive myelin basic protein (MBP) was measured in cerebrospinal fluid (CSF). Levels were correlated with neuropathological findings, interferon in CSF and virus isolation from the brain. CSF from animals inoculated with Cornell A75-17 strain often showed detectable immunoreactive MBP late in the disease course. As anticipated from earlier morphological studies, CSF drawn around day 20 PI lacked MBP while subsequent samples were positive. Dogs with severe demyelination had elevated values of immunoreactive MBP while dogs with only mild inflammation had little or none. Release of MBP or MBP peptides into CSF of dogs with canine distemper may be a valuable laboratory test in studies of the natural history of this disease and in assessing the response to treatment. Whether an immune response to MBP plays an immunopathogenic role in the chronic, demyelinating phase of canine distemper encephalitis remains to be determined.

Topics: Animals; Brain; Demyelinating Diseases; Distemper; Distemper Virus, Canine; Dogs; Encephalomyelitis; Myelin Basic Protein

The premise that acute non-fatal viral infections of the central nervous system (CNS) predispose to the subsequent development of chronic immune-mediated neurologic disease was investigated. Adult C57Bl/6 mice inoculated peripherally with 10(4) PFU of the A774 strain of Semliki Forest virus (SFV) develop a transient encephalomyelitis and sporadic (less than 20%) mild symptoms of paralysis with demyelination in the cerebellum from which they recover. Such recovered mice were found to develop signs characteristic of experimental allergic encephalomyelitis (EAE) 2 to 8 wk after either immunization with myelin basic protein (MBP) or receipt of 1 to 2 X 10(7) lymph node cells from MBP-primed syngeneic donors. These two methods of disease induction were unsuccessful when applied to normal B6 mice or those previously inoculated with noninfectious SFV. These findings suggest the possibility that virus-induced damage to CNS tissue may facilitate subsequent priming or clonal expansion of pre-existing myelin-reactive lymphoid cells.

Topics: Acute Disease; Animals; Demyelinating Diseases; Disease Susceptibility; Encephalomyelitis; Encephalomyelitis, Autoimmune, Experimental; Female; Immunization, Passive; Mice; Mice, Inbred C57BL; Myelin Basic Protein; Semliki forest virus; Togaviridae Infections

Following intracerebral inoculation of Theiler's murine encephalomyelitis virus (TMEV), susceptible mouse strains develop a chronic demyelinating disease characterized by mononuclear cell-rich infiltrates in the central nervous system. Current evidence strongly supports an immune-mediated basis for myelin breakdown, with an effector role proposed for TMEV-specific, major histocompatibility class II-restricted delayed-type hypersensitivity, which temporally correlates with disease onset and remains chronically elevated in susceptible mice. This study examined the fine specificity of class II-restricted T cell responses in TMEV-infected mice to better define the relevant virus-encoded T cell determinant(s) responsible for triggering the demyelinating process, and to determine if class II-restricted neuroantigen-specific autoimmune responses could be detected in mice with TMEV-induced demyelination. The data clearly show that T cell responses in TMEV-infected mice are directed against determinants shared by closely related TMEV strains and are cross-reactive with related picornaviruses, such as encephalomyocarditis virus. In contrast, class II-restricted autoimmune responses against syngeneic mouse spinal cord homogenate and the two major protein components of myelin, myelin basic protein and proteolipid protein, are not demonstrable in susceptible SJL/J mice undergoing chronic TMEV-induced demyelinating disease, but are readily seen in SJL/J mice displaying chronic, relapsing experimental allergic encephalomyelitis. Cross-reactivity (or lack thereof), as determined by functional T cell analyses, was found to correlate with the extent of exact amino acid homology between the TMEV capsid proteins, the two neuroantigens, and related picornaviruses. The data thus do not support a major role for autoimmune responses against myelin proteins in TMEV-induced demyelinating disease, but are consistent with our previously proposed hypothesis that TMEV-specific T cell responses constitute a major effector mechanism of myelin breakdown.

Topics: Animals; Cross Reactions; Demyelinating Diseases; Enterovirus; Histocompatibility Antigens Class II; Hypersensitivity, Delayed; Immunity, Cellular; Male; Maus Elberfeld virus; Mice; Mice, Inbred Strains; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Nerve Tissue Proteins; Picornaviridae; T-Lymphocytes

The mld mutation on chromosome 18 in the mouse is a putative allele of the shiverer (shi) mutation. We have analyzed the structure of myelin basic protein (MBP) gene sequences in mld DNA by restriction mapping of genomic DNA. The results indicate that the mld chromosome carries two copies of the MBP structural gene, one of which is intact and one of which is interrupted. Genetic analysis indicates that the interrupted gene is close to the intact MBP structural gene and cosegregates with the mld mutation. We have also analyzed the levels of MBP polypeptides and MBP-specific mRNA in wild-type, homozygous and heterozygous shiverer and mld mice and in mice carrying both mutations. The results indicate that both shi and mld are cis-acting codominant mutations that cause severely reduced steady state levels of MBP-specific mRNA and MBP polypeptides in the brain. We have analyzed the total number of oligodendrocytes and the number of MBP-positive oligodendrocytes in mld and shi brain primary cultures. In shi cultures, none of the oligodendrocytes expresses MBP. However, in mld cultures, approximately 5% of the oligodendrocytes express MBP. The nature of the "revertant" mld oligodendrocytes is not known.

Topics: Animals; Chromosome Mapping; Demyelinating Diseases; DNA Restriction Enzymes; Gene Expression Regulation; Genes; Mice; Mice, Neurologic Mutants; Mutation; Myelin Basic Protein; Phenotype

Both immunohistochemical and biochemical evidence is presented to show for the first time that carbonic anhydrase II (CA II) activity falls in the brain of mice in cuprizone (bis(cyclohexanone)oxalyldihydrazone) induced demyelination well before demyelination develops. This fall began during the first week, whereas the first signs of myelin degeneration induced by cuprizone did not appear until 3 weeks and demyelination in the superior cerebellar peduncle in the mouse took 6-8 weeks to develop. The findings suggest that oligodendrocyte CA II activity is essential either for the survival of oligodendrocytes or for the maintenance of central myelin.

Topics: Animals; Brain; Carbonic Anhydrases; Cuprizone; Demyelinating Diseases; Glial Fibrillary Acidic Protein; Male; Mice; Myelin Basic Protein; Myelin Sheath; Nerve Degeneration; Neuroglia; Oligodendroglia; Time Factors

Using two methods of immunization (A and B), 5/10 (A) and 27/45 (B), BALB/c by J mice immunized with human myelin proteolipid protein developed a demyelinating disease with a spectrum of chronic progressive to relapsing-remitting courses. Demyelinative lesions were seen histopathologically in all clinically affected animals that were examined. Many of the clinically unaffected animals also had histopathologic evidence of demyelination. Some of the animals had evidence of multiple ages of foci of activity. Evidence is presented that contamination with myelin basic protein could not account for the disease. This is a new model for multiple sclerosis in mice.

Topics: Animals; Brain; Chronic Disease; Demyelinating Diseases; Disease Models, Animal; Female; Glial Fibrillary Acidic Protein; Humans; Hypersensitivity, Delayed; Immunization; Mice; Mice, Inbred BALB C; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Neurons; Spinal Cord

Chronic relapsing experimental autoimmune encephalomyelitis is commonly seen in a number of species after a single injection of whole white matter in adjuvant but not after inoculation with myelin basic protein, the major encephalitogen of central myelin. In the present report on large groups of SJL mice, we describe a form of chronic relapsing experimental autoimmune encephalomyelitis with destructive lesions after a single inoculation of myelin basic protein in complete Freund's adjuvant. This condition was studied for up to 19 months postinoculation and was characterized by a relapsing-remitting or a chronic progressive course, usually with a prolonged latent period. Higher doses of 400 and 800 micrograms of myelin basic protein were more effective in inducing this condition than were lower doses of 100 and 200 micrograms. Large lesions were apparent in the white matter. These comprised widespread destruction and Wallerian degeneration with some demyelination towards the margins. Demyelination was an initial, albeit transient, event which was subsequently masked by nerve fiber destruction. Polymorphonuclear leukocytes were early and prominent components of the inflammatory infiltrate and together with macrophages appeared to be involved in the lysis of myelin and axons. Thus, despite the clinical similarities, these features contrast the model with the more purely demyelinative lesions of chronic relapsing experimental autoimmune encephalomyelitis in other species and multiple sclerosis in man.

Topics: Adjuvants, Immunologic; Animals; Demyelinating Diseases; Disease Models, Animal; Encephalomyelitis, Autoimmune, Experimental; Inflammation; Macrophages; Mice; Mice, Inbred Strains; Multiple Sclerosis; Myelin Basic Protein; Neutrophils; Wallerian Degeneration

Expression of the myelin proteolipid protein (PLP) was examined in the nuclei and polysomes of 12-27-day-old quaking, jimpy, and shiverer mouse brains and in 2-27-day-old normal brains and compared with expression of the myelin basic proteins (MBPs). Northern blots showed the presence of multiple mouse PLP RNAs, the developmental expression of which coincided with myelination. Two major mouse PLP RNAs, 3.5 and 2.6 kilobases in length, were observed in both cytoplasmic polyribosomes and nuclei, and, in addition, a larger 4.6-kilobase PLP RNA was observed in nuclei. Quantitative measurements with slot blot analyses showed that the levels of PLP and MBP RNAs peaked simultaneously at 18 days in nuclei but that maximal levels of PLP RNA lagged behind MBP RNA by several days in the polysomes. The developmental expression of both major classes of myelin protein mRNAs was affected in all three mutants. In shiverer brains, the levels of PLP mRNA in polysomes and nuclei were only 30-55% of control levels after 15 days. Thus, the deletion of a portion of the MBP gene appeared to have a major effect on the expression of the PLP gene in this mutant. In jimpy mice, where the mutation has been shown to involve the PLP gene, expression of MBP mRNA was also severely reduced, to less than 25% of control values. In quaking brains, the expression of each gene followed its own developmental course, different from each other and different from the normal mouse. The extent to which the expression of PLP and MBP was affected by the quaking mutation depended on the age at which it was examined.

Topics: Animals; Brain; Cell Nucleus; Demyelinating Diseases; DNA; DNA, Recombinant; Mice; Mice, Neurologic Mutants; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Nucleic Acid Hybridization; Polyribosomes; RNA, Messenger

The pathology of demyelination in rabbits with experimental allergic neuritis (EAN) or galactocerebroside-induced neuritis was compared to that in rabbits inoculated with either an emulsion of lipid haptens (gangliosides, lecithin and cholesterol) and Freund's complete adjuvant or Freund's complete adjuvant (FCA) alone. In rabbits inoculated with bovine peripheral myelin in FCA, perivenular demyelination associated with infiltrates of lymphocytes and macrophages occurred after 30 days, while those animals inoculated with galactocerebroside (GC) in Freund's adjuvant did not develop lesions until 60-90 days. GC rabbits had demyelination and severe nerve edema without cellular infiltrates. In rabbits inoculated with FCA alone, demyelination was restricted to ganglia and proximal nerve roots. Myelin basic protein (MBP) and GC antibodies from EAN, GC and lipid hapten-inoculated rabbits were detected by ELISA in sera at all post-inoculation time points. Appreciable P0 and P2 antibody titers were detected only in EAN animals. The results indicate that Freund's complete adjuvant alone or in combination with lipid haptens is capable of producing neuropathic effects in the rabbit independent of those produced by EAN or galactocerebroside neuritis.

Topics: Animals; Antibodies; Demyelinating Diseases; Female; Freund's Adjuvant; Galactosylceramides; Haptens; Lipids; Myelin Basic Protein; Myelin Sheath; Neuritis; Peripheral Nerves; Rabbits

Lewis and Brown Norway rats were infected at different ages with the neurotropic murine coronavirus strain, JHM and the resultant central nervous system diseases were studied. Suckling rats of both strains came down with a fatal, acute encephalomyelitis. Weanling Lewis rats developed a subacute demyelinating encephalomyelitis which neuropathologically revealed changes of an immunopathologic reaction. In contrast, Brown Norway rats developed a clinically silent subacute demyelinating encephalomyelitis with a persistent JHM virus infection which was less severe and quite different from the subacute demyelinating encephalomyelitis in Lewis rats with respect to size, distribution, and localization of the demyelinating plaques as well as the type of infiltrating cells. In addition, infected Lewis rats showed a pronounced lymphocyte proliferation to myelin basic protein and JHM virus whereas lymphocytes from infected Brown Norway rats did not react to these two antigens. These observations demonstrate the pathogenetic importance of host factors in the development of virus-induced demyelination.

Topics: Aging; Animals; Animals, Newborn; Antigens, Viral; Brain; Coronaviridae; Coronaviridae Infections; Demyelinating Diseases; Encephalomyelitis; Immunity, Cellular; Mice; Microscopy, Electron; Myelin Basic Protein; Rats; Rats, Inbred BN; Rats, Inbred Lew; Species Specificity; Spinal Cord

Antibody responses to the myelin/oligodendrocyte glycoprotein (MOG) and myelin basic protein (MBP) were determined in the sera of Hartley guinea pigs with chronic relapsing experimental allergic encephalomyelitis (CREAE) using an enzyme-linked immunoassay. The sera were also tested for in vivo demyelinating activity by infusion into the subarachnoid space of normal rats. In contrast to the MBP titres, the anti-MOG antibody titres showed good correlation with the in vivo demyelinating activity of the sera (r = 0.91, P less than 0.001). This result suggests that antibodies directed against MOG may be involved in the pathogenesis of demyelination in CREAE.

Topics: Animals; Autoantibodies; Chronic Disease; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Glycoproteins; Guinea Pigs; Humans; Myelin Basic Protein; Myelin Proteins; Neuroglia; Oligodendroglia

The levels of myelin basic protein (MBP) in the cerebrospinal fluid (CSF) have been evaluated as a method of detecting central nervous system (CNS) demyelination. MBP levels were measured in the CSF of 129 patients with neurological illnesses of whom 26 had a demyelinating illness. High levels of MBP were found in 81% of patients with recent episodes of demyelination but were not seen in patients with multiple sclerosis that was clinically stable or in remission. Patients with CNS trauma also had high levels of MBP in their CSF. The detection of a high level of MBP in CSF may be useful in the diagnosis of acute CNS demyelination but the finding is not specific for multiple sclerosis.

Topics: Demyelinating Diseases; Diagnosis, Differential; Humans; Multiple Sclerosis; Myelin Basic Protein; Nervous System Diseases

Platelet aggregation (PA) stimulated by encephalitogenic peptide (EP) and PA induced by ADP were measured in 83 multiple sclerosis (MS) patients and 70 control subjects with other neurological diseases (OND). EP-stimulated PA was significantly increased in MS patients as compared with the controls. There was no significant difference in ADP-induced PA between patients with MS and OND. The results are discussed in terms of EP-stimulated platelets playing a role in the pathogenesis of MS by affecting the venular permeability of the brain.

Topics: Adenosine Diphosphate; Adolescent; Adult; Aged; Capillary Permeability; Demyelinating Diseases; Female; Humans; Male; Middle Aged; Multiple Sclerosis; Myelin Basic Protein; Nervous System Diseases; Platelet Aggregation; Polyradiculopathy

Radioimmunoassay-determined myelin basic protein (MBP) shed to CSF during active demyelination, has been found to be a useful but non-specific test for MS. CSF creatin kinase BB (CK-BB), as measured by radioimmunoassay, is increased in a variety of neurological diseases, and has been considered a useful indication of brain damage but not of demyelinating diseases. Taking into account that the mean concentration of CSF CK-BB should not be increased in patients during the acute phase of MS, we suggest that the CSF MBP/CK-BB ratio could be a more specific index to demyelination than CSF-MBP alone. We also defined a laboratory demyelination pattern (CSF MBP greater than mean control MBP + 2 S.D. and CK-BB less than MBP). CSF levels of MBP and CSF levels of CK-BB were determined by radioimmunoassay in 232 patients with several neurological disorders and 33 control subjects. Patients diagnosed as having MS with clinical exacerbation had significantly higher values of CSF-MBP/CSF-BB ratio than control subjects. Our study showed a significant presence of demyelination pattern in CSF of patients with MS. We conclude that the CSF MBP/CK-BB ratio and the CSF demyelination pattern may be new and reliable tests for the diagnosis of MS.

Topics: Creatine Kinase; Demyelinating Diseases; Humans; Isoenzymes; Multiple Sclerosis; Myelin Basic Protein; Radioimmunoassay

A chronic form of experimental allergic encephalomyelitis can be produced by sensitization of rabbits with bovine myelin proteolipid apoprotein (PLP). To investigate the humoral immune response in this model, serum PLP antibodies were determined by enzyme-linked immunosorbent and dot immunobinding assays. In an initial experiment, 3 PLP-sensitized rabbits with severe chronic experimental allergic encephalomyelitis had a positive antibody response whereas 3 with mild disease, or with no visible clinical disease, had no detectable antibodies against PLP. In a second experiment, 3 rabbits were preimmunized with PLP in incomplete Freund's adjuvant, followed by a single immunization with PLP in complete Freund's adjuvant. These animals developed chronic experimental allergic encephalomyelitis with different progression rates, although all eventually became severely paralyzed. In both experiments the anti-PLP response was maximal before or immediately after disease onset and tended to decline during disease progression. The degree of the anti-PLP response correlated with clinical and histologic disease severity. These data suggest a possible role for humoral factors in the modulation of the chronic EAE induced in PLP-immunized rabbits.

Topics: Animals; Antibodies; Antibody Formation; Apoproteins; Cattle; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Enzyme-Linked Immunosorbent Assay; Immunization; Myelin Basic Protein; Myelin Proteins; Proteolipids; Rabbits

Guinea pigs received a suboptimal transfer of lymphocytes sensitized to myelin basic protein (BP) and were then immunized with guinea pig BP, BP plus chicken brain or chicken myelin, or chicken brain alone. Sera from these animals were tested for the presence of myelinotoxic antibodies, as detected by the myelination inhibition assay. Myelination inhibition activity correlated with the histologic severity of demyelination.

Topics: Animals; Antibodies; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Guinea Pigs; Lymphocyte Transfusion; Lymphocytes; Myelin Basic Protein

Of 66 patients (31 female and 35 male) with demyelinating inflammatory polyradiculoneuropathy (DIP), 12% (8/66) had a chronic relapsing and/or progressive course (CR-DIP) and 88% (58/66) had an acute monophasic illness (acute Guillain-Barré syndrome or GBS). Ten (15%) of the 66 had one or more associated putative autoimmune diseases; of these ten, five had CR-DIP and five had GBS. Cell-mediated immune responsiveness (CMI) of 30 cases with DIP was tested in vitro by lymphocyte transformation. Peripheral nervous system neuritogenic protein (NP) and central nervous system encephalitogenic myelin basic protein were the challenge antigens. Eighteen (60%) of the 30 patients had CMI to NP of human peripheral nervous system origin when a stimulation index (SI) of 2 or more was evaluated as positive; eight 27% (1) had CMI to NP when a positive SI was 3 or more. Of the 44 control patients with other neuropathies, only two (4.6%) demonstrated CMI to NP (SI, greater than or equal to 2). The in vitro response of patients with DIP to myelin basic protein (7/30) was not significantly different from that of the control population (16/44). The high incidence of DIP associated with autoimmune diseases and the CMI to NP in this group suggest that DIP may be an autoimmune disease with NP as one possible major antigen.

Topics: Acute Disease; Adolescent; Adult; Aged; Autoimmune Diseases; Central Nervous System; Child; Child, Preschool; Chronic Disease; Demyelinating Diseases; Female; Humans; Immunity, Cellular; Infant; Lymphocyte Activation; Lymphocytes; Male; Middle Aged; Myelin Basic Protein; Peripheral Nerves; Polyradiculoneuropathy

Utilizing a competitive inhibition enzyme-linked immunosorbent assay (ELISA) we measured the amount of myelin basic protein (MBP) in brain, liver, and kidney of newborn mouse and compared these values with myelinated cerebellar explant cultures under various conditions. Explant cultures demyelinated with anti-myelin serum showed a 90% decrease in MBP when compared with controls. These results correlated with myelination and demyelination as observed by light microscopy. Newborn tissues including brain showed only negligible activity for MBP. This technique provides a sensitive and objective method for quantitating in-vitro demyelination.

Topics: Animals; Animals, Newborn; Cerebellum; Culture Techniques; Demyelinating Diseases; Enzyme-Linked Immunosorbent Assay; Kidney; Liver; Mice; Myelin Basic Protein; Radioimmunoassay

Demyelination is a major feature of CNS injury and disease, including multiple sclerosis. To examine the potential for myelination by transplanted oligodendrocytes, initially described by Gumpel et al., we have transplanted neonatal cortex of mice with normal myelin into a dysmyelinating mutant, the shiverer mouse. We have found that oligodendrocyte precursors mature and synthesize myelin following transplantation. Immunostaining with antibodies to myelin basic protein (MBP), neurofilament protein and glial fibrillary acidic protein, demonstrates myelination both within the graft and extending out into the host, axonal sprouting from the graft which parallels the MBP-reactivity, and minimal astrocytic proliferation in response to the transplant.

Topics: Animals; Animals, Newborn; Demyelinating Diseases; Fetus; Glial Fibrillary Acidic Protein; Intermediate Filament Proteins; Mice; Mice, Neurologic Mutants; Myelin Basic Protein; Myelin Sheath; Neurofilament Proteins; Neuroglia; Oligodendroglia

The presence of myelin basic protein (MBP)-like material in cerebrospinal fluid (CSF) usually reflects breakdown of central nervous system myelin. Immunoreactive MBP levels were measured in 70 CSF specimens from 66 patients with a variety of peripheral neuropathies. Immunoreactive MBP was present in CSF in 70% (16 of 23) of patients with biopsy-proved chronic demyelinating polyneuropathies, whereas in neuropathies producing primarily axonal damage it was present in only 22% (5 of 23). In Guillain-Barré syndrome, 45% (9 of 20) of patients had immunoreactive MBP in the CSF. We conclude that MBP-like material is present in the CSF of most patients with longstanding demyelinating polyradiculoneuropathies and probably reflects detection of peripheral nervous system P1 protein.

Topics: Demyelinating Diseases; Humans; Myelin Basic Protein; Peripheral Nervous System Diseases; Polyradiculoneuropathy

To investigate a role for T lymphocytes in primary demyelination of central nervous system (CNS) tissue, antigen-specific T cell lines sensitized to myelin-associated and myelin-unrelated antigens were developed from SJL mice and tested on myelinated organotypic cultures of syngeneic spinal cord. Demyelination was assessed morphologically by electron microscopy. Antigen responsiveness and specificity, and the phenotypes of the cell lines, were determined by thymidine uptake (3H-TdR) assays and flow cytometry (FC), respectively. Although all T cell lines caused pathologic changes in myelin, the CNS-antigen-specific line induced the most pronounced effects. 3H-TdR uptake assays and FC showed that after three cycles of incubation in the presence of interleukin-2 (IL-2) or antigen, the T cell lines had increased specificity and responsiveness to the priming antigen and were enriched for the L3T4 (helper/inducer) phenotype. This represents the first direct demonstration of T-cell-mediated demyelination, supports a role for the helper/inducer subset in CNS lesion development, and may prove relevant to the human demyelinating disease multiple sclerosis.

Topics: Animals; Antigens, Surface; Cells, Cultured; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Interleukin-2; Mice; Mice, Inbred Strains; Myelin Basic Protein; Phenotype; Spinal Cord; T-Lymphocytes; T-Lymphocytes, Helper-Inducer

The loss of myelin-associated glycoprotein (MAG) and myelin basic protein (MBP) was compared by quantitative immunocytochemistry in demyelinating lesions of measles encephalomyelitis (ME), multiple sclerosis (MS), and progressive multifocal leukoencephalopathy (PML). Serial sections from paraffin-embedded tissue were reacted with antisera for MAG and MBP, and areas of staining loss were compared morphometrically. Lesions in ME showed MAG loss equal to that of MBP, lesions of PML showed MAG loss greater than that of MBP, and MS lesions showed a mixture of patterns. These data demonstrate distinctive patterns of MAG and MBP loss in these three diseases.

Topics: Central Nervous System; Demyelinating Diseases; Encephalomyelitis; Humans; Immunoenzyme Techniques; Leukoencephalopathy, Progressive Multifocal; Measles; Multiple Sclerosis; Myelin Basic Protein; Myelin Proteins; Myelin Sheath; Myelin-Associated Glycoprotein; Nerve Fibers, Myelinated

In dissociated mouse brain cell cultures we frequently observed an association between myelin basic protein (MBP) positive oligodendrocytes and proliferating astrocytes. When MBP was added in a purified form to the culture medium, it greatly stimulated the proliferation of astrocytes, while other proteins tested did not. This finding allows us to speculate that the gliosis observed in demyelinating diseases or/and in central nervous system (CNS) injury would be due to the mitogenic effect exerted by MBP or its fragments when there is myelin breakdown.

Topics: Animals; Astrocytes; Cells, Cultured; Central Nervous System; Demyelinating Diseases; Gliosis; Humans; Mice; Mitosis; Multiple Sclerosis; Myelin Basic Protein; Stimulation, Chemical

Acute demyelinating and relapsing demyelinating lesions from spinal cords of mice infected with the WW strain of Theiler's encephalomyelitis virus (TMEV) were studied immunocytochemically with antisera to various myelin constituents. Acute lesions were studied for differences in the distribution of myelin basic protein (MBP) and myelin associated glycoprotein (MAG). Relapsing lesions, characterized by demyelination of areas previously remyelinated by Schwann cells, were studied for differences in the distribution of P0 and MAG. In both instances the earliest lesions were characterized by preferential disappearance of MBP and P0 respectively when compared to MAG. In well-developed lesions, MAG, MBP and P0 were absent in essentially equal proportion. These observations are in agreement with previous findings suggesting a primary loss of myelin rather than a direct attack on oligodendrocytes as the main pathogenetic mechanism of demyelination in this viral model.

Topics: Acute Disease; Animals; Demyelinating Diseases; Enterovirus Infections; Histocytochemistry; Male; Maus Elberfeld virus; Mice; Myelin Basic Protein; Myelin P0 Protein; Myelin Proteins; Myelin-Associated Glycoprotein; Recurrence

We examined the in vivo demyelinating activity of serum from dogs with acute canine idiopathic polyneuropathy (ACIP), a Guillain-Barré syndrome (GBS)-like illness. Sera from 6 ACIP cases demyelinated rat sciatic nerves more intensely than 11 control sera. Serum activity increased after guinea pig serum (gps) was added, although gps alone had little effect. ACIP sera did not bind more to whole nerve cross sections or Schwann cells in vitro than control sera, and did not contain elevated antigalactocerebroside titers. We do not as yet know the pathogenic importance of the demyelinating factor in ACIP and control dog serum, or understand its relationship to the demyelinating constituent in serum from humans with GBS.

Topics: Animals; Demyelinating Diseases; Dog Diseases; Dogs; Guinea Pigs; Myelin Basic Protein; Myelin Sheath; Peripheral Nerves; Peripheral Nervous System Diseases; Rats; Schwann Cells; Sciatic Nerve

An immunoblotting technique has been used to screen serum samples from patients with demyelinating disease for antibody directed against central nervous system proteins. Antibodies of the IgM, IgG and IgA class directed against one or more of the particulate fraction proteins tubulin, myelin basic protein, 69 K neurofilament protein, glial fibrillary acidic protein, myelin associated glycoprotein or Wolfgram protein were present in 94, 54 and 47%, respectively, of multiple sclerosis sera examined. IgM antibodies against tubulin and myelin basic protein predominated. A similar antibody spectrum was seen in a significant proportion of sera from patients with optic neuritis, subacute sclerosing panencephalitis and motor neurone disease, in which primary or secondary demyelination occurs. Antibodies of all three classes directed against the 169 K and 220 K neurofilament proteins and against some unidentified proteins of human peripheral nerve, kidney, liver, spleen and skeletal muscle were detected in sera from healthy subjects and patients with neurological disease.

Topics: Adolescent; Adult; Antibody Specificity; Autoantibodies; Brain; Child; Demyelinating Diseases; Electrophoresis, Polyacrylamide Gel; Glial Fibrillary Acidic Protein; Humans; Immunoenzyme Techniques; Immunoglobulin A; Immunoglobulin G; Immunoglobulin M; Intermediate Filament Proteins; Middle Aged; Myelin Basic Protein; Nerve Tissue Proteins; Neurofilament Proteins; Tubulin

A 30-year-old homosexual man presented with widespread Burkitt's lymphoma. On the basis of immunologic and viral studies, he was suspected of having the acquired immune deficiency syndrome. Following chemotherapy that included intrathecal cytosine arabinoside and methotrexate, brain stem edema, paraplegia, and an elevated cerebrospinal fluid level of myelin basic protein developed. Autopsy revealed vacuolar demyelination of spinal cord, brain stem, and cerebellum. The pathologic findings were similar to those reported to occur in myelopathy associated with intrathecal chemotherapy, but far more extensive. The contribution of the suspected acquired immune deficiency syndrome is unknown.

Topics: Acquired Immunodeficiency Syndrome; Adult; Antineoplastic Combined Chemotherapy Protocols; Brain Diseases; Brain Stem; Burkitt Lymphoma; Cerebellum; Demyelinating Diseases; Homosexuality; Humans; Injections, Spinal; Male; Myelin Basic Protein; Spinal Cord Diseases

We aimed to study the level of CNPase activity in the cerebrospinal fluid of patients with demyelinating diseases and other neurological diseases, particularly multiple sclerosis, with reference to CSF myelin basic protein content. CNPase activity was measured paper chromatographically using radioactive 2',3'-cAMP as a substrate. Myelin basic protein content was measured with a radioimmunoassay. The mean level of CNPase activity was significantly higher for multiple sclerosis than for nonneurological controls. Dividing the disease phases of multiple sclerosis into the three periods, the CNPase activity was found to be significantly elevated in the worsening period and reduced in the improving period and the inactive period. The level of CNPase activity in the cerebrospinal fluid of multiple sclerosis coincided with the clinical activity of the disease. The level of CNPase activity correlated well (r = 0.84) with the level of myelin basic protein content in cerebrospinal fluid. The ratio for CNPase activity and myelin basic protein content in cerebrospinal fluid was almost the same as that in human central nerve myelin. We concluded that CNPase activity in the cerebrospinal fluid from neurological patients is an indicator of destruction of myelin in the central nervous system, and the measurement of CNPase activity in the cerebrospinal fluid of multiple sclerosis could be useful in the clinical management.

Topics: 2',3'-Cyclic Nucleotide 3'-Phosphodiesterase; 2',3'-Cyclic-Nucleotide Phosphodiesterases; Demyelinating Diseases; Humans; Multiple Sclerosis; Myelin Basic Protein; Phosphoric Diester Hydrolases; Radioimmunoassay

Total polyribosomes were isolated from the brains of 16-20 day C57BL/6 mice, four neurological mutants (qk/qk, shi/shi, mld/mld, and jp/Y), and four heterozygote or littermate controls (qk/+, shil/+, mld, and jp littermates) and translated in a homologous, cell-free system. No differences were observed among the nine genotypes in either the yield of polysomes (32.2 +/- 0.6 A260/g brain) or in the incorporation of [35S]methionine into trichloroacetic acid-precipitable protein. However, when the four myelin basic proteins (BPs) were isolated from the translation mixtures little incorporation of [35S]methionine into the BPs was noted in those assays directed by polysomes from mld/mld or from shi/shi animals. Compared with C57BL/6 polysomes, mld littermate and shi/+ polysomes incorporated approximately half the levels of label into the four BPs while qk/+ and qk/qk incorporated normal and close-to-normal levels. Polysomes from jp littermates and jp/Y brains synthesized 66% and less than 15% of the levels of the 14K BP compared with C57BL/6 polysomes. Incorporation of label into the other three BPs was normal with jp littermate polysomes and about half the control levels with jp/Y polysomes. The data indicate that shi/shi and mld/mld mutants either produce altered BPs not recognized by our antibody or synthesize very low levels of BP. The data provide additional support for the notion that the qk/qk mutant synthesizes much higher levels of MBP than are incorporated into myelin. They also indicate that in the jimpy mutant the synthesis of the four BPs is affected to differing extents; thus, the mutant cannot be easily characterized as either an "assembly" or "synthesis" defect.

Topics: Animals; Cell-Free System; Demyelinating Diseases; Genotype; Mice; Mice, Jimpy; Mice, Neurologic Mutants; Myelin Basic Protein; Ribosomes; Species Specificity

Topics: Animals; Antigens, Viral; Coronaviridae; Coronaviridae Infections; Cross Reactions; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Immunity, Cellular; Myelin Basic Protein; Rats; Rats, Inbred Lew; Recurrence

Topics: Animals; Chimera; Demyelinating Diseases; Genotype; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Neurologic Mutants; Mosaicism; Myelin Basic Protein

Immunoblot analysis of antiserum to rat central nervous system (CNS) myelin revealed antibodies to myelin basic protein (MBP), proteolipid protein (PLP), and numerous high molecular weight proteins. In addition, anti-CNS myelin serum exclusively immunostained 4 basic proteins of rat peripheral nervous system (PNS) myelin. Similarly, anti-PNS myelin sera immunostained many high molecular weight proteins in both CNS and PNS myelin in addition to P0 and 4 basic proteins. Purified MBP and PLP were immunostained by anti-CNS myelin sera and MBP and P0 by anti-PNS myelin sera, indicating that antigenic sites are preserved during protein purification. Immunohistochemical localization with antisera was confined to the myelin sheath except that antisera to CNS myelin also stained oligodendrocytes during the active period of myelination. While anti-CNS myelin sera specifically demyelinated centrally myelinated fibers in culture, none of the anti-PNS myelin sera used here demyelinated organotypic spinal cord-dorsal root ganglion cultures.

Topics: Animals; Antibodies; Central Nervous System; Culture Techniques; Demyelinating Diseases; Immune Sera; Myelin Basic Protein; Myelin Proteins; Myelin Proteolipid Protein; Myelin Sheath; Peripheral Nerves; Rats

Topics: Animals; Astrocytes; Autoimmune Diseases; Cell Line; Cell Movement; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Genetic Predisposition to Disease; Histocompatibility Antigens Class II; Humans; Major Histocompatibility Complex; Myelin Basic Protein; Organ Specificity; Rats; Rats, Inbred Lew; T-Lymphocytes

Eleven dogs were experimentally infected with canine distemper virus and studied for periods of up to 63 days post-inoculation. The responsiveness of lymphocytes in vitro toward phytohemagglutinin, myelin basic protein and galactocerebroside was tested at regular intervals during the course of infection by means of [3H]thymidine incorporation and flow cytometry. All dogs developed a marked decrease of lymphocyte responsiveness toward phytohemagglutinin. Four dogs recovered rapidly from the immunosuppression and did not develop demyelination or had only mild lesions, while two others failed to recover at all and developed severe non-inflammatory demyelinating lesions. The remaining dogs exhibited a slow or partial immune recovery and had various degrees of inflammatory demyelination. Lymphocytes from 2 dogs with demyelination and 2 dogs without myelin lesions responded to myelin antigens. The findings indicate that the degree of immunosuppression in canine distemper virus infection may determine the type of demyelination and autoimmune reactions that occur during the inflammatory stage of demyelination may be epiphenomena.

Topics: Animals; Antigens; Brain Diseases; Demyelinating Diseases; Distemper; Dogs; Galactosylceramides; Immunosuppression Therapy; Lectins; Lymphocyte Activation; Myelin Basic Protein; Phytohemagglutinins

A temporal series of demyelinating lesions in experimental canine distemper virus (CDV) infection was examined with immunohistological techniques demonstrating myelin basic protein (MBP), myelin-associated glycoprotein (MAG), and glial fibrillary acidic protein (GFAP) on serial sections. The earliest lesions were characterized by decreased MBP and MAG and increased GFAP. During the further progression of the disease, MBP and MAG losses continued to match each other. There was no indication of MAG loss preceding the disappearance of MBP. In the more advanced lesions there was a marked decrease of GFAP positive cells. Since these findings differed considerably from similar immunohistochemical studies in progressive multifocal leukoencephalopathy (PML) where demyelination results from oligodendroglial infection, it was concluded that the oligodendroglial cell body is not the primary target of CDV. The marked astroglial changes were also considered to contribute to demyelination in CDV infection but the mechanism by which this happens remains unknown.

Topics: Animals; Astrocytes; Demyelinating Diseases; Distemper; Dogs; Glial Fibrillary Acidic Protein; Glycoproteins; Intermediate Filament Proteins; Myelin Basic Protein; Myelin Proteins; Nerve Tissue Proteins; Neuroglia

Sections of the central nervous system of the leukodystrophic mouse mutants 'jimpy' (jp) and 'myelin deficient' (mld), as well as of healthy littermates, were immunostained for glial fibrillary acidic protein (GFA), myelin basic protein (MBP) and myelin-associated glycoprotein (MAG). Adjacent sections were stained conventionally for myelin. In jp, GFA-stained astrocytes were abnormally prominent already at the age of 12 days. A considerable amount of MBP and MAG was present in the vicinity of axons, although no myelin was visible in the conventional stains for myelin. In mld, GFA-staining astrocytes were present in normal numbers. MAG could be demonstrated in its normal localization along axons, but MBP was visible only in the comparatively old animal (85 days). Here, it was demonstrated in an abnormal site--the perikarya and the proximal parts of the processes of oligodendrocytes. Thin myelin sheaths present in this animal could not be stained for MBP.

Topics: Animals; Brain Stem; Cerebellum; Demyelinating Diseases; Glial Fibrillary Acidic Protein; Immunoassay; Intermediate Filament Proteins; Mice; Mice, Mutant Strains; Microscopy, Electron; Myelin Basic Protein; Myelin Proteins; Myelin Sheath; Myelin-Associated Glycoprotein; Optic Nerve; Spinal Cord

Topics: Animals; Brain; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Lipid Metabolism; Membranes; Myelin Basic Protein; Myelin Sheath; Proteins; Spinal Cord

A general outline of the morphologic structure and biochemistry of myelin, its mode of development and assembly, and its alterations in diseased states, including diagnoses and prognoses of diseases affecting myelin, is given.

Topics: Demyelinating Diseases; Humans; Lipid Metabolism; Lipoproteins; Multiple Sclerosis; Myelin Basic Protein; Myelin Sheath

Nine dogs with canine distemper encephalitis (CDE) were examined with immunological techniques including demonstration of antibodies against canine distemper virus (CDV) in the serum and against myelin basic protein (MBP) in serum and in CSF. Mitogen stimulation tests of lymphocytes were also done. The brains were examined pathologically and immunoglobulin and C3 were demonstrated in lesions by means of immunohistological techniques. Six dogs with acute CDE had none or low antibody levels against CDV or MBP, and there was no immunoglobulin in demyelinating lesions. Some of these dogs had depressed lymphocyte mitogen responses. Two dogs with chronic CDE showed recovery of lymphocyte mitogen responses. One of these had a significant antibody response against CDV and MBP in the serum. Both dogs with chronic CDE had very high antibody titers against MBP in the CSF and demyelinating lesions contained immunoglobulin. These results suggest that acute demyelination in CDE is probably due to some direct viral activity and that the progression of demyelination in chronic CDE is associated with a local immune response.

Topics: Animals; Antibodies, Viral; Brain Chemistry; Complement C3; Demyelinating Diseases; Distemper; Distemper Virus, Canine; Dogs; Encephalitis; Female; Immunoglobulins; Lymphocyte Activation; Male; Myelin Basic Protein

Myelin basic protein (BP) is a specific constituent of the myelin sheath. This structural protein cannot be detected in the cerebrospinal fluid (CSF) unless myelin is acutely degraded. In order to detect active demyelinating diseases, BP was measured in CSF samples of radioimmunoassay. The assay is specific and sensitive to as little as 1.5 to 2.5 ng/ml BP. A moderate non-parallelism between the standard curve and various dilutions of CSF samples indicates that in CSF BP is present in an altered state. Over 1000 CSF samples have been measured in a double-blind study, in which 100 patients were selected and their clinical records evaluated. Twenty-eight patients without demyelinating disease had BP levels lower than 2.5 ng/ml. 72 patients had values higher than 2.5 ng/ml. Among them, the most frequent causes of demyelination were multiple sclerosis (19 cases), brain tumors (22 cases) and cerebral or spinal vascular accidents (12 cases). During a single acute demyelinating episode, BP levels revert to background levels within a few days. In contrast to immunological anomalies observed in the CSF, the presence of BP is concomitant with the breakdown of myelin. The size and location of the lesion influence the level of BP in the CSF. Thus, the assay is useful for the detection of active demyelination in the central nervous system and in following the course of the disease, although normal values do not rule out the presence of demyelinating lesions. For the time being, therefore, this assay should be restricted to specialized neurological centers and selected patients.

Topics: Adult; Brain Diseases; Brain Neoplasms; Central Nervous System Diseases; Child; Demyelinating Diseases; Female; Humans; Male; Meningoencephalitis; Multiple Sclerosis; Myelin Basic Protein; Radioimmunoassay

Topics: Collodion; Demyelinating Diseases; Electrophoresis, Polyacrylamide Gel; Glial Fibrillary Acidic Protein; Humans; Methods; Multiple Sclerosis; Myelin Basic Protein; Nerve Tissue Proteins

Topics: Demyelinating Diseases; Humans; Myelin Basic Protein; Radioimmunoassay

It is postulated that the pathogenesis of demyelination in multiple sclerosis (MS) might lie in the cooperative effect of a T cell response against one myelin antigen (e.g. myelin basic protein--MBP) and a B cell response against a second myelin component which may act as a hapten or a carrier for the primary antigen. The hypothesis is based upon recent experiments in guinea pigs in which the encephalitogenicity of MBP was enhanced by the myelin glycolipid, galactocerebroside. This pathogenetic mechanism might be analogous to antibody-dependent, cell-mediated demyelination. Based upon this assumption, therapeutic trials in MS should take into consideration the possibility that instead of MBP alone, MBP might be more effective in combination with a lipid hapten.

Topics: Animals; Antigens; Autoimmune Diseases; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Galactosylceramides; Guinea Pigs; Haptens; Male; Multiple Sclerosis; Myelin Basic Protein

Topics: Animals; Antibodies; Brain; Demyelinating Diseases; Dogs; Endopeptidases; Female; Injections, Spinal; Male; Microscopy, Electron; Myelin Basic Protein; Myelin Sheath

Topics: Animals; Axons; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Guinea Pigs; Microscopy, Electron; Myelin Basic Protein; Optic Nerve; Optic Neuritis; Papilledema

Levels of cerebrospinal fluid (CSF) basic protein (BP) immunoreactive material and CSF lymphocyte in vitro reactivity to BP were determined in patients with demyelinating and other inflammatory diseases of the nervous system. Elevated levels of BP and enhanced in vitro reactivity were observed, but there was no correlation between the magnitude of the in vitro response and the amount of BP-like material in CSF.

Topics: Demyelinating Diseases; Humans; Lymphocytes; Multiple Sclerosis; Myelin Basic Protein

Activated macrophages secrete a variety of neutral proteinases, including plasminogen activator. Since macrophages are implicated in primary demyelination in the peripheral nervous system (PNS) in Guillain-Barré syndrome and experimental allergic neuritis, we have investigated the ability of plasmin and of conditioned media from cultured macrophages, in the presence of plasminogen, to degrade the proteins in bovine and rat PNS myelin. The results indicate that (a) the major glycoprotein P0 and the basic P1 and Pr proteins in PNS myelin are extremely sensitive to plasmin, perhaps more so than is the basic protein in CNS myelin; (b) the initial product of degradation of P0 by plasmin has a molecular weight higher than that of the "X" protein; (c) large degradation products of P0 are relatively insensitive to further degradation; and (d) the neuritogenic P2 protein in PNS myelin is quite resistant to the action of plasmin. Results similar to those with plasmin were obtained with conditioned media from macrophages and macrophage-like cell lines together with plasminogen activator, and the degradation of the PNS myelin proteins, Po and P1, under these conditions was inhibited by p-nitrophenylguanidinobenzoate, an inhibitor of plasmin and plasminogen activator. The results suggest that the macrophage plasminogen activator could participate in inflammatory demyelination in the PNS.

Topics: Animals; Brain; Cattle; Demyelinating Diseases; Fibrinolysin; In Vitro Techniques; Macrophages; Molecular Weight; Myelin Basic Protein; Myelin P0 Protein; Myelin Proteins; Peripheral Nerves; Rats; Spinal Nerve Roots

The encephalitogenic effects of bovine galactocerbroside and total myelin lipids given in the presence or absence of a known encephalitogenic dose of bovine myelin basic protein (MBP) in complete Freund's adjuvant have been examined in Hartley strain guinea pigs. The lipid haptens and MBP were given in the ratio in which they occur in intact myelin and were compared with experimental allergic encephalomyelitis (EAE) induced by whole bovine white matter in complete Freund's adjuvant. Clinically, bovine white matter- and MBP-induced EAE were similar, but in lesions in the central nervous system. Lipid/complete Freund's adjuvant emulsions were ineffective both clinically and histologically. In combination with MBP, galactocerebroside and total myelin lipids induced an EAE as severe as MBP-induced disease except that central nervous system lesions also showed demyelination. When given separately into opposite hindfeet, the lipid haptens and MBP produced EAE, but the lesions were not demyelinative. It appears, therefore, that lipid haptens have an augmenting effect on MBP when given in the same emulsion and produce central nervous system lesions which are both inflammatory and demyelinative.

Topics: Animals; Cerebrosides; Demyelinating Diseases; Galactosylceramides; Guinea Pigs; Haptens; Lipids; Male; Micelles; Myelin Basic Protein

Myelinated cultures of mouse spinal cord have been exposed to sera raised in rabbits against whole white matter (anti-WM), myelin basic protein (anti-MBP) and galactocerebroside (anti-GC), the major glycolipid of CNS myelin, to determine which factor in central nervous system (CNS) tissue in vitro is the target of serum demyelinating and myelin swelling antibodies. The sera were tested by radioimmunoassay for activity against MBP and against GC and were also specifically absorbed with MBP, GC and control antigens. Studies were also performed with and without active complement. The findings show that demyelination and myelin swelling in vitro are caused by antibodies against GC and not against MBP. Ultrastructurally, the effects of anti-WM and anti-GC sera with and without complement were indistinguishable. This study demonstrates that GC is a major target in antibody-mediated demyelination.

Topics: Absorption; Animals; Cells, Cultured; Cerebrosides; Complement System Proteins; Demyelinating Diseases; Galactosylceramides; Guinea Pigs; Immune Sera; In Vitro Techniques; Male; Mice; Microscopy, Electron; Myelin Basic Protein; Myelin Sheath; Rabbits; Spinal Cord

Cerebrospinal fluid from 582 persons was analyzed by a double-antibody radioimmunoassay for the presence of material cross-reactive with peptide 43-88 of human myelin basic protein (BP). In a group of 104 patients with multiple sclerosis (MS), 23 of 33 individuals clinically judged to have had an exacereation within two weeks prior to the time CSF was obtained had detectable material ranging from 2 to 200 ng/ml. In the remaining 71 MS patients who either were stable or had had an exacerbation more than two weeks before, only 1 patient had a marginally elevated level of immunoreactive material. CSF from 53 persons with cerebrovascular disease was studied, and 13 of 29 with recent infarctions had values of 2 to 540 ng/ml. The degree of elevation in strokes generally paralleled the predicted volume of the lesion, but the amounts detected did not correlate quite so closely temporally with onset as they did with the periods of active disease in MS. Of the remaining 425 patients, 29 had immunoreactive material of 2 to 400 ng/ml in their CSF. Most of these patients with detectable material had acute diseases known to affect the myelin sheath. Eight of 10 persons with acute disseminated encephalomyelitis had no detectable material. The presence in CSF of material cross-reactive with BP peptide 43-88 does not have diagnostic specificity for MS but can be used as a means for determining recent myelin injury. The type of BP peptide formed and mechanisms for clearance of BP and BP peptides may be important in determining the biological consequences following release of this potentially immunogenic material from the central nervous system.

Topics: Antigen-Antibody Reactions; Central Nervous System Diseases; Demyelinating Diseases; Humans; Multiple Sclerosis; Myelin Basic Protein; Nervous System Diseases; Radioimmunoassay

Topics: Adolescent; Adult; Brain; Brain Diseases; Child; Child, Preschool; Demyelinating Diseases; Humans; Leukemia, Lymphoid; Methotrexate; Myelin Basic Protein

Topics: Animals; Brain; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Humans; Membrane Lipids; Membrane Proteins; Multiple Sclerosis; Myelin Basic Protein; Myelin Sheath; Nerve Fibers, Myelinated; Neural Conduction; Oligodendroglia; Schwann Cells

Topics: Amino Acid Sequence; Animals; Demyelinating Diseases; Female; Fibroblast Growth Factors; Fibroblasts; Growth Substances; Humans; Mice; Mice, Inbred BALB C; Mice, Inbred Strains; Mitogens; Models, Molecular; Multiple Sclerosis; Muscular Dystrophies; Myelin Basic Protein; Peptides

Topics: Animals; Central Nervous System Diseases; Demyelinating Diseases; Disease Models, Animal; Guinea Pigs; Mice; Mice, Neurologic Mutants; Myelin Basic Protein; Myelin Sheath; Schwann Cells

We have developed a radioimmunoassay to measure a specific neurological component, the basic protein of myelin, and have used this test for assessing this component in spinal fluid. The levels of basic protein in spinal fluid correlate closely with the clinical activity of multiple sclerosis; therefore the test can be used for objective evaluation of disease activity in patients with that disorder. Moreover, it is a useful adjunct in the diagnosis of multiple sclerosis and evaluation of potential therapy. In addition, the test is helpful in diagnosing other diseases in which acute breakdown of myelin occurs, especially leukoencephalopathy resulting from irradiation and chemotherapy for treatment of leukemia in children.

Topics: Demyelinating Diseases; Humans; Multiple Sclerosis; Myelin Basic Protein; Optic Neuritis; Radioimmunoassay

Material cross-reactive with myelin basic protein (BP) peptide 43-88 has been identified as a major BP-like antigen appearing in human cerebrospinal fluid during acute myelin injury. In an effort to develop a means for detecting this material in the more accessible body fluids of blood and urine, as well as to determine its metabolic fate, the manner in which rabbits handle human BP peptide 43-88 was investigated. Unlabeled peptide was administered intravenously, and its concentration in plasma was monitored by radioimmunoassay. In studies of 10 rabbits, the peptide disappeared from blood in two phases, the first showing a half-life of 6.0 +/- 1.2 minutes and the second a half-life of 51.6 +/- 5.4 minutes. Organ-exclusion experiments indicated a rapid clearance, predominantly by the kidney, for the peptide, with tubular reabsorption and cleavage into smaller peptide fragments a probable catabolic mechanism.

Topics: Animals; Demyelinating Diseases; Female; Half-Life; Humans; Kidney; Myelin Basic Protein; Nephrectomy; Peptides; Rabbits; Radioimmunoassay

In inbred Lewis rats, P2 basic protein from bovine peripheral nervous system (PNS) myelin produced experimental allergic neuritis (EAN) without involvement of the brain or spinal cord. In guinea-pigs, bovine P2 did not produce EAN but large doses produced mild experimental allergic encephalomyelitis (EAE). In rabbits, bovine P2 produced both mild EAE and EAN. Human P2 produced severe EAN in the Lewis rat, but only mild EAN with quite marked EAE in the guinea-pig. Material cross-reacting with bovine P on immunodiffusion was identified in the extracts from the nerves of all three species but only in the spinal cord of the guinea-pig and rabbit, not in the rat spinal cord. The species differences in response to immunisation with P2 cannot be simply explained by the presence or absence of P2 in their PNS or CNS, but may reflect differences in the immune response.

Topics: Animals; Antigens; Autoimmune Diseases; Cattle; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Guinea Pigs; Humans; Male; Myelin Basic Protein; Neuritis; Rabbits; Rats; Rats, Inbred Lew; Species Specificity; Spinal Nerve Roots

The light microscopical examinations were performed on the peripheral nerve (PN) lesions in rabbits induced by bovine peripheral nerve (BPN) myelin and by its components. Morphology of the PN lesions induced by P2 protein-ganglioside complex were equivalent to those induced by PN myelin (original EAN); and some of them were more remarkable in severity than the original EAN. The lesions were composed of myelin destruction, numerous macrophages containing myelin debris, completely demyelinated axons and remyelinating fibers. P2-I, acid treated P2-I and P2-II proteins induced PN lesions. Histologically they were moderate to minimal in severity in that order. However, when each protein was injected as the complex with ganglioside, PN lesions obviously became severer, respectively. These findings indicate the possibility that the P2-ganglioside complex may be involved in the chemical principle of EAN, and also the findings strongly support the theory proposed by Nagai et al. that gangliosides may play an essential role in rabbit EAN to give P2 protein a special conformation necessary for antigenic activity.

Topics: Animals; Autoimmune Diseases; Demyelinating Diseases; Gangliosides; Immunization; Male; Myelin Basic Protein; Myelin P2 Protein; Myelin Proteins; Neuritis; Peripheral Nerves; Rabbits

Groups of juvenile Strain 13 guinea pigs sensitized for chronic relapsing experimental allergic encephalomyelitis (EAE) with isogeneic central nervous system (CNS) tissue in complete Freund's adjuvant (CFA) were either left to develop late-onset chronic EAE (unsuppressed), or given a series of injections of bovine myelin basic protein (MBP) in incomplete Freund's adjuvant (IFA) to suppress the disease. All unsuppressed animals developed disease and all suppressed animals remained healthy over a 27-month period of study. some unsuppressed and suppressed animals were rechallenged with CNS tissue in CFA 12 or 26 months post-inoculation (PI). Unsuppressed animals all became sick 2-4 weeks after rechallenge, while rechallenged, suppressed animals were protected, indicating that the suppression was permanent. Pathologic findings in the CNS complemented the clinical changes. Circulating lymphocyte studies were performed on animals from all groups. Early (active, high-affinity rosetting) T cell levels in unsuppressed animals showed significant decreases during exacerbations (P less than 0.01) and normal values during remissions. After rechallenge, circulating early T cells decreased in unsuppressed animals with the development of signs. In suppressed animals, early T cells showed significant elevations during, and for a short time after, the period of suppressive injections, and normal values afterwards. These levels did not change significantly after rechallenge. Late (total, 24 hour rosetting) T cell and B cell values showed minor fluctuations only which did not correlate with disease activity. These results indicate that chronic relapsing EAE can be successfully suppressed with MBP in IFA, that this suppression is permanent and that the immunologic findings presented correlate well with the clinical and pathologic facets of the disease. the findings are presented in terms of their relevance to multiple sclerosis.

Topics: Animals; Chronic Disease; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Freund's Adjuvant; Guinea Pigs; Leukocyte Count; Lymphocytes; Male; Myelin Basic Protein; Spinal Cord

Topics: Animals; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Humans; Isoelectric Point; Methods; Multiple Sclerosis; Myelin Basic Protein; Myelin Proteins; Peptide Hydrolases; Structure-Activity Relationship

In the inflammatory demyelinating diseases, such as multiple sclerosis, Landry-Guillain-Barré syndrome and experimental allergic encephalomyelitis, demyelination occurs in the vicinity of infiltrating mononuclear cells. Although the histopathology is characteristic of each disease, the general observation that myelin destruction in inflammatory lesions begins prior to phagocytosis suggests a common mechanism for myelinolysis in these diseases. Recent studies show that stimulated macrophages secrete several neutral proteinases, including plasminogen (Plg) activator. We have tested the possibility that these proteinases could, directly or indirectly, initiate myelin destruction. Isolated brain myelin was incubated with supernatant media from cultures of stimulated mouse peritoneal macrophages in the presence and absence of Plg. Cell supernatants alone caused some degradation of basic protein (BP) in myelin. The amount degraded was considerably enhanced in the presence of Plg. The other myelin proteins remained essentially intact. While the Plg-independent proteolytic activity in the supernatants was abolished by EDTA, known to inhibit the neutral proteinases, the Plg-dependent hydrolysis was inhibited by p-nitrophenylguanidinobenzoate, an inhibitor of Plg activator and plasmin. These results suggested that the Plg activator secreted by the macrophages generated plasmin, which selectively degraded BP. This interpretation was confirmed by the observation that urokinase, a Plg activator, plus Plg was effective in degrading BP in myelin. We propose that the action of neutral proteinases released by stimulated macrophages, and its amplification by the Plg-plasmin system, may play a significant role in several inflammatory demyelinating diseases; and that the relative specificity of these reactions for myelin lies in the extreme susceptibility of BP to proteolysis.

Topics: Animals; Brain; Cats; Cattle; Demyelinating Diseases; Fibrinolysin; Inflammation; Macrophages; Molecular Weight; Myelin Basic Protein; Myelin Sheath; Peptide Hydrolases; Plasminogen; Plasminogen Activators

Topics: Animals; Autoantibodies; Binding Sites, Antibody; Cerebrosides; Culture Techniques; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Galactosylceramides; Humans; Immune Sera; Immunoenzyme Techniques; Multiple Sclerosis; Myelin Basic Protein; Myelin Proteins; Myelin Sheath; Rabbits; Spinal Cord

Bulk-isolated human and bovine oligodendroglia, practically free from myelin, have been used in attempts to elicit an autoimmune response which has been compared with acute experimental allergic encephalomyelitis (EAE). For these experiments, a total of 20 Hartley guinea pigs, 33 Lewis rats and 16 rabbits have been studied. Animals were inoculated with a range of doses of purified preparations of both human and bovine oligodendroglial cells in complete Freund's adjuvant (CFA) and compared with others challenged with whole white matter in CFA. The latter animals all developed clinical and histological signs of experimental allergic encephalomyelitis (EAE) 2-3 weeks post-inoculation. In general, oligodendroglial cells were encephalitogenically less potent than white matter. Guinea pigs were the most susceptible to inoculations of oligodendroglia. In several given human oligodendroglia 14 days earlier, a paraparesis indistinguishable from conventional EAE was seen. Animals receiving bovine cells showed no clinical signs. Histologically, the CNS of afflicted guinea pigs displayed severe inflammation but, in contrast to conventional EAE in the same species, demyelination was rare in the small group of animals tested. After sensitization with oligodendroglia, rats displayed no clinical disease. Histologically, some given human cells had positive evidence of disease while bovine cells in others gave a mild response. Rabbits showed no clinical and very little histological disease. Although more extensive studies are needed to confirm the findings, from the animals studied it appears that (1) variation in response to inocula containing oligodendroglia exists among the species tested, (2) that human oligodendroglia are more potent immunologically than bovine cells, (3) that CNS lesions produced by these cells in guinea pigs, lack a strong demyelinative component and (4) a specific antigen might exist in oligodendrocytes which is distinct from myelin basic protein. The possible reasons underlying our findings are discussed.

Topics: Animals; Cattle; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Guinea Pigs; Humans; Male; Meninges; Myelin Basic Protein; Neuroglia; Oligodendroglia; Rabbits; Rats; Rats, Inbred Lew; Spinal Cord; Spinal Nerve Roots

Topics: Animals; Autolysis; Brain; Brain Edema; Central Nervous System; Demyelinating Diseases; Edema; Female; Lipid Metabolism; Myelin Basic Protein; Myelin Proteins; Myelin Sheath; Rats; Spinal Cord; Spinal Cord Diseases; Time Factors; Triethyltin Compounds

Neuropathological evidence of demyelination was found in the brain and sciatic nerve of diabetic patients at autopsy. The activity of acid proteinase was somewhat increased in the white matter but decreased in the gray matter of diabetic patients. No increase was observed in the activity of neutral proteinase in diabetic white and gray matter. The activities of beta-glucuronidase and 2',3'-cyclic nucleotide-3'-phosphohydrolase (CNP) were of the same level as those of the controls. The activities of all 4 enzymes appeared to be increased in the diabetic nerve, with the possible exception of CNP which was measured from only 1 nerve. Furthermore, the amount of total protein was markedly decreased in diabetic peripheral myelin. The encephalitogenic basic protein of diabetic brain myelin was normal in the disc gel electrophoretic patterns of brain myelin proteins. However, the basic proteins of peripheral myelin were reduced in a number of diabetic patients. The present biochemical findings for diabetic white and gray matter were largely normal. Instead, the increased activities of at least the proteinases and beta-glucuronidase in diabetic peripheral nerve, together with the loss of basic proteins, indicate extensive biochemical damage of the peripheral nervous system in diabetes. They suggest that demyelination and other phenomena observed in diabetic peripheral nerve are not caused only by angiopathy and impaired circulation.

Topics: 2',3'-Cyclic-Nucleotide Phosphodiesterases; Adult; Aged; Brain; Demyelinating Diseases; Diabetic Neuropathies; Female; Glucuronidase; Humans; Male; Middle Aged; Myelin Basic Protein; Myelin Proteins; Nervous System; Peptide Hydrolases; Peripheral Nerves

Topics: Animals; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Epitopes; Freund's Adjuvant; Mice; Mutation; Myelin Basic Protein; Myelin Sheath

Complement-dependent demyelinating activity of whole brain homogenate (WBH)-induced experimental allergic encephalomyelitis (EAE) sera was tested on long term tissue cultures of in vitro myelinated fetal guinea pig cerebellum. Complement-fixing (CF) auto-antibodies were shown to be the responsible agents, as demonstrated in experiments where all reagents belonged to the same species: guinea pigs of outbred (Hartley) and even of inbred (S2 or S13) strains. These antibodies were of the IgG2 class as shown by Sephadex G-200 and DEAE cellulose fractionation experiments. The corresponding auto-antigen was present in the homogenate and myelin of the central nervous system (CNS) tissue. It was different from the encephalitogenic basic protein of CNS myelin (BP), as shown in experiments where the demyelinating auto-antibodies were induced, detected, and absorbed by WBH or by CNS myelin but not by BP. They were neither induced by nor cross-reacting with cerebroside and peripheral nervous system (PNS) tissue.

Topics: Animals; Autoantibodies; Autoimmune Diseases; Brain; Central Nervous System; Cerebrosides; Complement Fixation Tests; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Guinea Pigs; Immunoglobulin G; In Vitro Techniques; Myelin Basic Protein; Peripheral Nerves; Tissue Extracts

In order to elucidate the role of humoral antibodies in the pathogenesis of myelin lesions in experimental allergic encephalomyelitis (EAE) a combined in vivo and in vitro study was done using rabbits immunized with the purified A1 basic protein. Rabbits injected with whole white matter were used for comparison. Demyelinating activity appeared in the rabbit sera 5 days after injection, as tested in myelinated organotypic tissue cultures. In spite of this no lesions of the myelin preceded the appearance of inflammatory cells in the living animals. In the spinal cord changes in vascular permeability, as revealed by leakage of Evans blue-albumin complex, appeared at the same time as the cells. In contrast to in vitro, the mere presence of circulating antibodies in vivo does not appear to be enough to cause structural changes of the myelin. Possible reasons for this discrepancy are discussed; it is emphasized that the inflammatory changes develope first in areas where the so-called blood-brain barrier to diffusion of proteins is lacking.

Topics: Animals; Autoantibodies; Blood-Brain Barrier; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Myelin Basic Protein; Myelin Sheath; Rabbits

Topics: Animals; Antigens; Cattle; Central Nervous System; Cerebrosides; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Guinea Pigs; Inflammation; Membranes, Artificial; Myelin Basic Protein; Myelin Sheath; Time Factors

Topics: Animals; Brain; Chronic Disease; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Guinea Pigs; Lymphocytes; Macrophages; Microscopy, Electron; Myelin Basic Protein; Optic Nerve; Plasma Cells; Spinal Cord

Sensitization of guinea pigs with purified myelin basic protein induces experimental allergic encephalomyelitis (EAE) but does not induce a serum factor which inhibits myelin formation in vitro. This factor, induced by some unidentified constituent of whole central nervous system tissue, should not be characterized as a component of "EAE serum."

Topics: Animals; Antibody Formation; Antigen-Antibody Reactions; Antigens; Cerebellum; Culture Techniques; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Guinea Pigs; Immune Sera; Iodine Isotopes; Myelin Basic Protein; Myelin Sheath; Nerve Tissue Proteins; Rabbits; Spinal Cord

Topics: Animals; Brain; Cell Differentiation; Central Nervous System Diseases; Cholesterol; Chromatography, Thin Layer; Demyelinating Diseases; Diffuse Cerebral Sclerosis of Schilder; Disease Models, Animal; DNA; Electrophoresis, Polyacrylamide Gel; Male; Mice; Mice, Inbred Strains; Myelin Basic Protein; Myelin Sheath; Nerve Fibers, Myelinated; Nerve Tissue Proteins; Neuroglia; Phospholipids; RNA; Sulfoglycosphingolipids

Topics: Animals; Brain; Brain Stem; Cerebral Cortex; Demyelinating Diseases; Disease Models, Animal; Electrophoresis, Polyacrylamide Gel; Encephalomyelitis, Autoimmune, Experimental; Haplorhini; Macaca; Multiple Sclerosis; Myelin Basic Protein; Necrosis; Nerve Tissue Proteins; Peptide Hydrolases

Topics: Amino Acids; Animals; Cyclopentanes; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Fluorescent Antibody Technique; Freund's Adjuvant; Immunosuppressive Agents; Lymph Nodes; Male; Mercaptopurine; Mycobacterium; Myelin Basic Protein; Rats; Rats, Inbred Lew; Spinal Cord

Topics: Animals; Brain; Demyelinating Diseases; Encephalomyelitis, Autoimmune, Experimental; Female; Freund's Adjuvant; Guinea Pigs; Male; Myelin Basic Protein; Proteins; Spinal Cord

Topics: Acid Phosphatase; Animals; Cell Separation; Demyelinating Diseases; Electrophoresis, Polyacrylamide Gel; Encephalomyelitis, Autoimmune, Experimental; Female; Freund's Adjuvant; Guinea Pigs; Hexosaminidases; Leukocyte Count; Lymphocytes; Lysosomes; Male; Myelin Basic Protein; Nitrophenols; Phosphates; Ultrasonics