Remyelination induced by a DNA aptamer in a mouse model of multiple sclerosis

Multiple sclerosis (MS) is a debilitating inflammatory disease of the central nervous system (CNS) characterized by local destruction of the insulating myelin surrounding neuronal axons. With more than 200 million MS patients worldwide, the absence of treatments that prevent progression or induce repair poses a major challenge. Anti-inflammatory therapies have met with limited success only in preventing relapses. Previous screening of human serum samples revealed natural IgM antibodies that bind oligodendrocytes and promote both cell signaling and remyelination of CNS lesions in an MS model involving chronic infection of susceptible mice by Theiler's encephalomyelitis virus and in the lysolecithin model of focal demyelination. This intriguing result raises the possibility that molecules with binding specificity for oligodendrocytes or myelin components may promote therapeutic remyelination in MS. Because of the size and complexity of IgM antibodies, it is of interest to identify smaller myelin-specific molecules with the ability to promote remyelination in vivo. Here we show that a 40-nucleotide single-stranded DNA aptamer selected for affinity to murine myelin shows this property. This aptamer binds multiple myelin components in vitro. Peritoneal injection of this aptamer results in distribution to CNS tissues and promotes remyelination of CNS lesions in mice infected by Theiler's virus. Interestingly, the selected DNA aptamer contains guanosine-rich sequences predicted to induce folding involving guanosine quartet structures. Relative to monoclonal antibodies, DNA aptamers are small, stable, and non-immunogenic, suggesting new possibilities for MS treatment.     

Mediators of oligodendrocyte differentiation during remyelination

Myelin, a dielectric sheath that wraps large axons in the central and peripheral nervous systems, is essential for proper conductance of axon potentials. In multiple sclerosis (MS), autoimmune-mediated damage to myelin within the central nervous system (CNS) leads to progressive disability primarily due to limited endogenous repair of demyelination with associated axonal pathology. While treatments are available to limit demyelination, no treatments are available to promote myelin repair. Studies examining the molecular mechanisms that promote remyelination are therefore essential for identifying therapeutic targets to promote myelin repair and thereby limit disability in MS. Here, we present our current understanding of the critical extracellular and intracellular pathways that regulate the remyelinating capabilities of oligodendrocyte precursor cells (OPCs) within the adult CNS.     

Cells of the oligodendroglial lineage, myelination, and remyelination

Myelin is critical in maintaining electrical impulse conduction in the central nervous system. The oligodendrocyte is the cell type responsible for myelin production within this compartment. The mutual supply of trophic support between oligodendrocytes and the underlying axons may indicate why demyelinated axons undergo degeneration more readily; the latter contributes to the neural decline in multiple sclerosis (MS). Myelin repair, termed remyelination, occurs in acute inflammatory lesions in MS and is associated with functional recovery and clinical remittances. Animal models have demonstrated that remyelination is mediated by oligodendrocyte progenitor cells (OPCs) which have responded to chemotactic cues, migrated into the lesion, proliferated, differentiated into mature oligodendrocytes, and ensheathed demyelinated axons. The limited remyelination observed in more chronic MS lesions may reflect intrinsic properties of neural cells or extrinsic deterrents. Therapeutic strategies currently under development include transplantation of exogenous OPCs and promotion of remyelination by endogenous OPCs. All currently approved MS therapies are aimed at dampening the immune response and are not directly targeting neural processes.     

Macrophages in CNS Remyelination: Friend or Foe?

Hematogenous macrophages and resident brain microglia are agents of demyelination in multiple sclerosis (MS) and paradoxically may also participate in remyelination. In vitro studies have shown that macrophage enrichment of aggregate brain cultures promotes myelination per se and enhances the capacity to remyelinate following a demyelinating episode. It has been hypothesized that remyelination in MS is implemented by surviving dedifferentiated oligodendrocytes or by newly recruited progenitors that migrate, proliferate and synthesize myelin in response to signalling molecules in the local environment. We postulate that macrophage-derived cytokines or growth factors may directly or indirectly promote oligodendroglial proliferation and differentiation, contributing to myelin repair in inflammatory demyelinating disease.     

Vitamins E and D3 Attenuate Demyelination and Potentiate Remyelination Processes of Hippocampal Formation of Rats Following Local Injection of Ethidium Bromide

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 μg/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.     

Multiple sclerosis - remyelination failure as a cause of disease progression

Multiple sclerosis (MS) is the most frequent demyelinating disease of the central nervous system (CNS) that affects worldwide about 2.5 million people. The morphological correlates of the disease are multiple lesions in brain and spinal cord which are characterized by demyelination, inflammation, gliosis and axonal damage. The underlying cause for the permanent neurological deficits in MS patients is axonal loss. Demyelinated axons are prone to damage due to the lack of trophic support by myelin sheaths and oligodendrocytes, as well as the increased vulnerability to immune mediated attacks. Remyelination occurs, but especially in chronic lesions is frequently limited to a small rim at the lesion border. Current treatment strategies are based on anti-inflammatory or immunomodulatory drugs and have the potential to reduce the numbers of newly evolving lesions, although as yet no treatment strategy exists to influence or prevent the progressive disease phase. Therefore, the development of neuroprotective treatment options, such as the promotion of endogenous remyelination is an attractive strategy. A prerequisite for the development of such new treatments is the understanding of the mechanisms leading to remyelination and the reasons for insufficient endogenous repair in chronic MS. This review will therefore provide an overview of the current concepts regarding remyelination in the rodent and human CNS. We will also summarize a selected number of inhibitory pathways and non-disease related factors which may contribute to remyelination failure in chronic MS.     

Promotion of Remyelination by Sulfasalazine in a Transgenic Zebrafish Model of Demyelination

Most of the axons in the vertebrate nervous system are surrounded by a lipid-rich membrane called myelin, which promotes rapid conduction of nerve impulses and protects the axon from being damaged. Multiple sclerosis (MS) is a chronic demyelinating disease of the CNS characterized by infiltration of immune cells and progressive damage to myelin and axons. One potential way to treat MS is to enhance the endogenous remyelination process, but at present there are no available treatments to promote remyelination in patients with demyelinating diseases.Sulfasalazine is an anti-inflammatory and immune-modulating drug that is used in rheumatology and inflammatory bowel disease. Its anti-inflammatory and immunomodulatory properties prompted us to test the ability of sulfasalazine to promote remyelination. In this study, we found that sulfasalazine promotes remyelination in the CNS of a transgenic zebrafish model of NTR/MTZ-induced demyelination. We also found that sulfasalazine treatment reduced the number of macrophages/microglia in the CNS of demyelinated zebrafish larvae, suggesting that the acceleration of remyelination is mediated by the immunomodulatory function of sulfasalazine. Our data suggest that temporal modulation of the immune response by sulfasalazine can be used to overcome MS by enhancing myelin repair and remyelination in the CNS.     

The remyelination Philosopher's Stone: stem and progenitor cell therapies for multiple sclerosis

Multiple sclerosis (MS) is an autoimmune disease that leads to oligodendrocyte loss and subsequent demyelination of the adult central nervous system (CNS). The pathology is characterized by transient phases of recovery during which remyelination can occur as a result of resident oligodendroglial precursor and stem/progenitor cell activation. However, myelin repair efficiency remains low urging the development of new therapeutical approaches that promote remyelination activities. Current MS treatments target primarily the immune system in order to reduce the relapse rate and the formation of inflammatory lesions, whereas no therapies exist in order to regenerate damaged myelin sheaths. During the last few years, several transplantation studies have been conducted with adult neural stem/progenitor cells and glial precursor cells to evaluate their potential to generate mature oligodendrocytes that can remyelinate axons. In parallel, modulation of the endogenous progenitor niche by neural and mesenchymal stem cell transplantation with the aim of promoting CNS progenitor differentiation and myelination has been studied. Here, we summarize these findings and discuss the properties and consequences of the various molecular and cell-mediated remyelination approaches. Moreover, we address age-associated intrinsic cellular changes that might influence the regenerative outcome. We also evaluate the extent to which these experimental treatments might increase the regeneration capacity of the demyelinated human CNS and hence be turned into future therapies.     

Mesenchymal Stem Cell Conditioning Promotes Rat Oligodendroglial Cell Maturation

Oligodendroglial progenitor/precursor cells (OPCs) represent the main cellular source for the generation of new myelinating oligodendrocytes in the adult central nervous system (CNS). In demyelinating diseases such as multiple sclerosis (MS) myelin repair activities based on recruitment, activation and differentiation of resident OPCs can be observed. However, the overall degree of successful remyelination is limited and the existence of an MS-derived anti-oligodendrogenic milieu prevents OPCs from contributing to myelin repair. It is therefore of considerable interest to understand oligodendroglial homeostasis and maturation processes in order to enable the development of remyelination therapies. Mesenchymal stem cells (MSC) have been shown to exert positive immunomodulatory effects, reduce demyelination, increase neuroprotection and to promote adult neural stem cell differentiation towards the oligodendroglial lineage. We here addressed whether MSC secreted factors can boost the OPC’s oligodendrogenic capacity in a myelin non-permissive environment. To this end, we analyzed cellular morphologies, expression and regulation of key factors involved in oligodendroglial fate and maturation of primary rat cells upon incubation with MSC-conditioned medium. This demonstrated that MSC-derived soluble factors promote and accelerate oligodendroglial differentiation, even under astrocytic endorsing conditions. Accelerated maturation resulted in elevated levels of myelin expression, reduced glial fibrillary acidic protein expression and was accompanied by downregulation of prominent inhibitory differentiation factors such as Id2 and Id4. We thus conclude that apart from their suggested application as potential anti-inflammatory and immunomodulatory MS treatment, these cells might also be exploited to support endogenous myelin repair activities.     

Myelin repair: the role of stem and precursor cells in multiple sclerosis

Multiple sclerosis is the most common potential cause of neurological disability in young adults. The disease has two distinct clinical phases, each reflecting a dominant role for separate pathological processes: inflammation drives activity during the relapsing-remitting stage and axon degeneration represents the principal substrate of progressive disability. Recent advances in disease-modifying treatments target only the inflammatory process. They are ineffective in the progressive stage, leaving the science of disease progression unsolved. Here, the requirement is for strategies that promote remyelination and prevent axonal loss. Pathological and experimental studies suggest that these processes are tightly linked, and that remyelination or myelin repair will both restore structure and protect axons. This review considers the basic and clinical biology of remyelination and the potential contribution of stem and precursor cells to enhance and supplement spontaneous remyelination.     

IGF binding protein alterations on periplaque oligodendrocytes in multiple sclerosis: implications for remyelination

Why myelin repair greatly fails in multiple sclerosis (MS) is unclear. The insulin-like growth factor (IGF) system plays vital roles in oligodendrocyte development, survival, and myelin synthesis. We used immunohistochemistry to study IGF-I, IGF-I receptors and IGF binding proteins (IGFBPs) 1-6 on oligodendrocytes at the edges of chronic demyelinated plaques and normal appearing white matter of MS, and in cerebral white matter of controls without neurological disease. Oligodendrocytes in all conditions were immunoreactive for IGF-I, IGF-I receptors and IGFBPs-1-5. Oligodendrocytes at the edges of demyelinated plaques displayed enhanced immunoreactivity for IGF-I, IGF-I receptors, IGFBPs-1 and -6. Because increased expression of IGFBPs-1 and -6 has been associated with impaired synthesis of myelin proteins in oligodendrocyte lineage cells, pharmacological approaches to reduce their expression might be useful for promoting remyelination of axons in MS lesions.     

A novel microtubule-associated protein-2 expressed in oligodendrocytes in multiple sclerosis lesions

Elucidation of the mechanisms involved in the regeneration of oligodendrocytes and remyelination is a central issue in multiple sclerosis (MS) research. We recently identified a novel alternatively spliced, developmentally regulated oligodendrocyte-specific protein designated microtubule-associated protein-2+13 [microtubule-associated protein-2 expressing exon 13 (MAP-2+13)]. MAP-2+13 is expressed in human fetal oligodendrocytes during process extension and myelination but is minimally expressed in normal mature CNS. To test the hypothesis that MAP-2+13 is reexpressed in regenerating oligodendrocytes in MS lesions, we examined the brains of MS patients for the expression of this protein. By immunocytochemistry using a series of monoclonal antibodies specific for MAP-2+13, we determined that MAP-2+13 expression was up-regulated in all 31 lesions from 10 different MS brains. MAP-2+13 was expressed in regenerating oligodendrocytes associated with demyelinated lesions, with the highest counts found in regions of extensive remyelination. By electron microscopy, MAP-2+13 was localized to oligodendrocytes engaged in remyelination, evident by their process extension and association with thinly myelinated (remyelinated) and demyelinated axons. These results suggest a hitherto unsuspected role for this microtubule-associated protein in oligodendrocyte function during development and myelin repair.     

The Na+/Ca2+ exchangers in demyelinating diseases

Intracellular [Na⁺]_i and [Ca²⁺]_i imbalance significantly contribute to neuro-axonal dysfunctions and maladaptive myelin repair or remyelination failure in chronic inflammatory demyelinating diseases such as multiple sclerosis. Progress in recent years has led to significant advances in understanding how [Ca²⁺]_i signaling network drive degeneration or remyelination of demyelinated axons.The Na⁺/Ca²⁺ exchangers (NCXs), a transmembrane protein family including three members encoded by ncx1, ncx2, and ncx3 genes, are emerging important regulators of [Na⁺]_i and [Ca²⁺]_i both in neurons and glial cells. Here we review recent advance highlighting the role of NCX exchangers in axons and myelin-forming cells, i.e. oligodendrocytes, which represent the major targets of the aberrant inflammatory attack in multiple sclerosis. The contribution of NCX subtypes to axonal pathology and myelin synthesis will be discussed. Although a definitive understanding of mechanisms regulating axonal pathology and remyelination failure in chronic demyelinating diseases is still lacking and requires further investigation, current knowledge suggest that NCX activity plays a crucial role in these processes. Defining the relative contributions of each NCX transporter in axon pathology and myelinating glia will constitute not only a major advance in understanding in detail the intricate mechanism of neurodegeneration and remyelination failure in demyelinating diseases but also will help to identify neuroprotective or remyelinating strategies targeting selective NCX exchangers as a means of treating MS.     

An animal model for late onset chronic demyelination disease caused by failed terminal differentiation of oligodendrocytes

Various animal models are available for studying human multiple sclerosis (MS). Most of them model the initial phase of MS,including the immune-triggered attack of the myelin membrane and/or oligodendrocytes and, occasionally, demonstrate there mission and relapsing phases. However, few mimic the late chronic demyelinating phase. Overexpression of the proteolipid protein gene (Plp) causes a unique demyelinating disorder in mice in which normal-appearing myelin forms early in life and chronic demyelination occurs later. We found that remyelination is severely affected in this late demyelinating phase, but is not caused by deprivation of oligodendrocyte progenitors expressing PDGF receptor alpha (PDGFRa) and Olig2, which are present at an even higher number in the demyelinated white matter of the mutants than in wild-type controls. Furthermore, mature oligodendrocytes containing PLP were observed, but failed to remyelinate. The ability of oligodendrocytes from older transgenic animals to produce a myelin membrane-like structure was not impaired when cultured in vitro, which indicates that the lack of remyelination is not simply caused by changes in the intrinsic properties of the oligodendrocytes. Glial activation also occurred much earlier than active demyelination in mutant mice. Thus, in addition to intrinsic mechanisms, extrinsic mechanisms might also have an important role in defects of remyelination. These features are also observed in patients at a late stage of MS, leading to chronic demyelinating lesions. Thus, this mouse model partly mimics the late stage of MS and can be used to study the cause of inhibition of remyelination.     

Thyroid hormone alleviates demyelination induced by cuprizone through its role in remyelination during the remission period

Multiple sclerosis (MS) is a disease induced by demyelination in the central nervous system, and the remission period of MS is crucial for remyelination. In addition, abnormal levels of thyroid hormone (TH) have been identified in MS. However, in the clinic, insufficient attention has been paid to the role of TH in the remission period. Indeed, TH not only functions in the development of the brain but also affects myelination. Therefore, it is necessary to observe the effect of TH on remyelination during this period. A model of demyelination induced by cuprizone (CPZ) was used to observe the function of TH in remyelination during the remission period of MS. Through weighing and behavioral tests, we found that TH improved the physical symptoms of mice impaired by CPZ. Supplementation of TH led to the repair of myelin as detected by immunohistochemistry and western blot. In addition, a sufficient TH supply resulted in an increase in myelinated axons without affecting myelin thickness and g ratio in the corpus callosum, as detected by electron microscopy. Double immunostaining with myelin basic protein and neurofilament 200 (NF200) showed that the CPZ-induced impairment of axons was alleviated by TH. Conversely, insufficient TH induced by 6-propyl-2-thiouracil resulted in the enlargement of mitochondria. Furthermore, we found that an adequate supply of TH promoted the proliferation and differentiation of oligodendrocyte lineage cells by immunofluorescence, which was beneficial to remyelination. Further, we found that TH reduced the number of astrocytes without affecting microglia. Conclusively, it was shown that TH alleviated demyelination induced by CPZ by promoting the development of oligodendrocyte lineage cells and remyelination. The critical time for remyelination is the remission period of MS. TH plays a significant role in alleviating demyelination during the remission period in the clinical treatment of MS.     

The response of NG2-expressing oligodendrocyte progenitors to demyelination in MOG-EAE and MS

Remyelination of primary demyelinated lesions is a common feature of experimental models of multiple sclerosis (MS) and is also suggested to be the normal response to demyelination during the early stages of MS itself. Many lines of evidence have shown that remyelination is preceded by the division of endogenous oligodendrocyte precursor cells (OPCs) in the lesion and its borders. It is suggested that this rapid response of OPCs to repopulate the lesion site and their subsequent differentiation into new oligodendrocytes is the key to the rapid remyelination. Antibodies to the NG2 chondroitin sulphate proteoglycan have proved exceedingly useful in following and quantitating the response of endogenous OPCs to demyelination. Here we review the literature on the response of NG2-expressing OPCs to demyelination and provide some new evidence on their response to the chronic inflammatory demyelinating environment seen in recombinant myelin oligodendrocyte glycoprotein (MOG) induced experimental allergic encephalomyelitis (EAE) in the DA rat. NG2-expressing OPCs responded to the inflammatory demyelination in this model by becoming reactive and increasing in number in a very focal manner. Evidence of NG2⁺OPCs in lesioned areas beginning to express the oligodendrocyte marker CNP was also seen. The response of OPCs appeared to occur following successive relapses but did not always lead to remyelination, with areas of chronic demyelination observed in the spinal cord. The presence of OPCs in the adult human CNS is clearly of vital importance for repair in multiple sclerosis (MS). As in rat tissue, the antibody labels an evenly distributed cell population present in both white and grey matter, distinct from HLA-DR⁺microglia. NG2⁺cells are sparsely distributed in the centre of chronic MS lesions. These cells apparently survive demyelination and exhibit a multi-processed or bipolar morphology in the very hypocellular environment of the lesion.     

Rejuvenation of regeneration in the aging central nervous system

Remyelination is a regenerative process in the central nervous system (CNS) that produces new myelin sheaths from adult stem cells. The decline in remyelination that occurs with advancing age poses a significant barrier to therapy in the CNS, particularly for long-term demyelinating diseases such as multiple sclerosis (MS). Here we show that remyelination of experimentally induced demyelination is enhanced in old mice exposed to a youthful systemic milieu through heterochronic parabiosis. Restored remyelination in old animals involves recruitment to the repairing lesions of blood-derived monocytes from the young parabiotic partner, and preventing this recruitment partially inhibits rejuvenation of remyelination. These data suggest that enhanced remyelinating activity requires both youthful monocytes and other factors, and that remyelination-enhancing therapies targeting endogenous cells can be effective throughout life.     

Glia Disease and Repair—Remyelination

The inability of the mammalian central nervous system (CNS) to undergo spontaneous regeneration has long been regarded as a central tenet of neurobiology. However, although this is largely true of the neuronal elements of the adult mammalian CNS, save for discrete populations of granular neurons, the same is not true of its glial elements. In particular, the loss of oligodendrocytes, which results in demyelination, triggers a spontaneous and often highly efficient regenerative response, remyelination, in which new oligodendrocytes are generated and myelin sheaths are restored to denuded axons. Yet, remyelination in humans is not without limitation, and a variety of demyelinating conditions are associated with sustained and disabling myelin loss. In this review, we will review the biology of remyelination, including the cells and signals involved; describe when remyelination occurs and when and why it fails and the consequences of its failure; and discuss approaches for therapeutically enhancing remyelination in demyelinating diseases of both children and adults, both by stimulating endogenous oligodendrocyte progenitor cells and by transplanting these cells into demyelinated brain.