[Show abstract][Hide abstract] ABSTRACT: Central nervous system remyelination by oligodendrocyte progenitor cells (OPCs) ultimately fails in the majority of multiple sclerosis (MS) lesions. Remyelination benefits from transient expression of factors that promote migration and proliferation of OPCs, which may include fibronectin (Fn). Fn is present in demyelinated lesions in two major forms; plasma Fn (pFn), deposited following blood-brain barrier disruption, and cellular Fn, synthesized by resident glial cells and containing alternatively spliced domains EIIIA and EIIIB. Here, we investigated the distinctive roles that astrocyte-derived Fn (aFn) and pFn play in remyelination. We used an inducible Cre-lox recombination strategy to selectively remove pFn, aFn or both from mice, and examined the impact on remyelination of toxin-induced demyelinated lesions of spinal cord white matter. This approach revealed that astrocytes are a major source of Fn in demyelinated lesions. Furthermore, following aFn conditional knockout, the number of OPCs recruited to the demyelinated lesion decreased significantly, whereas OPC numbers were unaltered following pFn conditional knockout. However, remyelination completed normally following conditional knockout of aFn and pFn. Both the EIIIA and EIIIB domains of aFn were expressed following demyelination, and in vitro assays demonstrated that the EIIIA domain of aFn mediates proliferation of OPCs, but not migration. Therefore, although the EIIIA domain from aFn mediates OPC proliferation, aFn is not essential for successful remyelination. Since previous findings indicated that astrocyte-derived Fn aggregates in chronic MS lesions inhibit remyelination, aFn removal may benefit therapeutic strategies to promote remyelination in MS. GLIA 2014
[Show abstract][Hide abstract] ABSTRACT: The role of remote astrocyte (AC) reaction to central or peripheral axonal insult is not clearly understood. Here we use a transgenic approach to compare the direct influence of normal with diminished AC reactivity on neuronal integrity and synapse recovery following extracranial facial nerve transection in mice. Our model allows straightforward interpretations of AC-neuron signalling by reducing confounding effects imposed by inflammatory cells. We show direct evidence that perineuronal reactive ACs play a major role in maintaining neuronal circuitry following distant axotomy. We reveal a novel function of astrocytic signal transducer and activator of transcription-3 (STAT3). STAT3 regulates perineuronal astrocytic process formation and re-expression of a synaptogenic molecule, thrombospondin-1 (TSP-1), apart from supporting neuronal integrity. We demonstrate that, through this new pathway, TSP-1 is responsible for the remote AC-mediated recovery of excitatory synapses onto axotomized motor neurons in adult mice. These data provide new targets for neuroprotective therapies via optimizing AC-driven plasticity.
[Show abstract][Hide abstract] ABSTRACT: Functional screening for compounds that promote remyelination represents a major hurdle in the development of rational therapeutics for multiple sclerosis. Screening for remyelination is problematic, as myelination requires the presence of axons. Standard methods do not resolve cell-autonomous effects and are not suited for high-throughput formats. Here we describe a binary indicant for myelination using micropillar arrays (BIMA). Engineered with conical dimensions, micropillars permit resolution of the extent and length of membrane wrapping from a single two-dimensional image. Confocal imaging acquired from the base to the tip of the pillars allows for detection of concentric wrapping observed as 'rings' of myelin. The platform is formatted in 96-well plates, amenable to semiautomated random acquisition and automated detection and quantification. Upon screening 1,000 bioactive molecules, we identified a cluster of antimuscarinic compounds that enhance oligodendrocyte differentiation and remyelination. Our findings demonstrate a new high-throughput screening platform for potential regenerative therapeutics in multiple sclerosis.
[Show abstract][Hide abstract] ABSTRACT: Genome-wide association studies have revolutionised the genetic analysis of multiple sclerosis. Through international collaborative efforts involving tens of thousands of cases and controls, more than 100 associated common variants have now been identified. These variants consistently implicate genes associated with immunological processes, overwhelmingly lie in regulatory rather than coding regions, and are frequently associated with other autoimmune diseases. The functional implications of these associated variants are mostly unknown; however, early work has shown that several variants have effects on splicing that result in meaningful changes in the balance between different isoforms in relevant tissues. Including the well established risk attributable to variants in genes encoding human leucocyte antigens, only about a quarter of reported heritability can now be accounted for, suggesting that a substantial potential for further discovery remains.
The Lancet Neurology 07/2014; · 21.82 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The vacuole formation (VF) rat is an autosomal recessive myelin mutant characterized by generalized tremor, hypomyelination, and periaxonal vacuole formation of the central nervous system (CNS). Here, we report the most likely causative gene for neurological disease in the VF rat and pursue its roles in the development and maintenance of the CNS myelin. We identified a nonsense mutation in the dopey family member 1 (Dopey1) located on rat chromosome 8. Expression level of Dopey1 mRNA was decreased and DOPEY1 protein was undetectable both in the white and gray matter of the spinal cords in the VF rats. Double immunohistochemistry demonstrated that DOPEY1 was mainly expressed in neurons and oligodendrocytes in the wild-type rats, whereas no positive cells were detected in the VF rats. We also demonstrated a marked reduction in myelin components both at mRNA and protein levels during myelinogenesis in the VF rats. In addition, proteolipid protein and myelin-associated glycoprotein accumulated in oligodendrocyte cell body, suggesting that Dopey1 is likely to be involved in the traffic of myelin components. Our results highlighted the importance of Dopey1 for the development and maintenance of the CNS myelin. GLIA 2014.
[Show abstract][Hide abstract] ABSTRACT: Oligodendrocyte progenitors (OPs) are a major proliferating cell population within the adult CNS. In response to myelin loss or increasing demand, OPs have the capacity to differentiate into mature, myelinating oligodendrocytes. The name 'oligodendrocyte progenitor' suggests restriction to the oligodendrocyte cell lineage. However, with growing evidence of the lineage plasticity of OPs both in vitro and in vivo, we discuss whether they have potential beyond that expected of dedicated progenitor cells, and hence may justify categorization as adult stem cells.
[Show abstract][Hide abstract] ABSTRACT: Embryonic stem cells (ESCs) self-renew in a state of naïve pluripotency in which they are competent to generate all somatic cells. It has been hypothesized that, before irreversibly committing, ESCs pass through at least one metastable transition state. This transition would represent a gateway for differentiation and reprogramming of somatic cells. Here, we show that during the transition, the nuclei of ESCs are auxetic: they exhibit a cross-sectional expansion when stretched and a cross-sectional contraction when compressed, and their stiffness increases under compression. We also show that the auxetic phenotype of transition ESC nuclei is driven at least in part by global chromatin decondensation. Through the regulation of molecular turnover in the differentiating nucleus by external forces, auxeticity could be a key element in mechanotransduction. Our findings highlight the importance of nuclear structure in the regulation of differentiation and reprogramming.
[Show abstract][Hide abstract] ABSTRACT: Myelination allows rapid saltatory propagation of action potentials along the axon and is an essential prerequisite for the normal functioning of the nervous system. During peripheral nervous system (PNS) development, myelin-forming Schwann cells (SCs) generate radial lamellipodia to sort and ensheath axons. This process requires controlled cytoskeletal remodeling, and we show that SC lamellipodia formation depends on the function of profilin 1 (Pfn1), an actin-binding protein involved in microfilament polymerization. Pfn1 is inhibited upon phosphorylation by ROCK, a downstream effector of the integrin linked kinase pathway. Thus, a dramatic reduction of radial lamellipodia formation is observed in SCs lacking integrin-linked kinase or treated with the Rho/ROCK activator lysophosphatidic acid. Knocking down Pfn1 expression by lentiviral-mediated shRNA delivery impairs SC lamellipodia formation in vitro, suggesting a direct role for this protein in PNS myelination. Indeed, SC-specific gene ablation of Pfn1 in mice led to profound radial sorting and myelination defects, confirming a central role for this protein in PNS development. Our data identify Pfn1 as a key effector of the integrin linked kinase/Rho/ROCK pathway. This pathway, acting in parallel with integrin β1/LCK/Rac1 and their effectors critically regulates SC lamellipodia formation, radial sorting and myelination during peripheral nervous system maturation.
[Show abstract][Hide abstract] ABSTRACT: Microglia are the resident macrophages of the central nervous system that survey the microenvironment for signals of injury or infection. The response to such signals induces an inflammatory response involving macrophages derived from both resident microglia and recruited circulating monocytes. Although implicated as contributors to autoimmune-mediated injury, microglia/ macrophages have recently been shown to be critical for the important central nervous system regenerative process of remyelination. This functional dichotomy may reflect their ability to be polarized along a continuum of activation states including the well-characterized cytotoxic M1 and regenerative M2 phenotypes. Here we review the roles of microglia, monocytes and the macrophages they give rise to in creating lesion environments favourable to remyelination, highlighting the specific roles of M1 and M2 phenotypes and how the pro-regenerative role of the innate immune system is altered by ageing. This article is protected by copyright. All rights reserved.
Journal of Neurochemistry 03/2014; · 4.24 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Amongst neurological diseases, multiple sclerosis (MS) presents an attractive target for regenerative medicine. This is because the primary pathology, the loss of myelin-forming oligodendrocytes, can be followed by a spontaneous and efficient regenerative process called remyelination. While cell transplantation approaches have been explored as a means of replacing lost oligodendrocytes, more recently therapeutic approaches that target the endogenous regenerative process have been favored. This is in large part due to our increasing understanding of (1) the cell types within the adult brain that are able to generate new oligodendrocytes, (2) the mechanisms and pathways by which this achieved, and (3) an emerging awareness of the reasons why remyelination efficiency eventually fails. Here we review some of these advances and also highlight areas where questions remain to be answered in both the biology and translational potential of this important regenerative process. GLIA 2014;
[Show abstract][Hide abstract] ABSTRACT: During the last two decades, many experiments have examined the ability of cell transplants to ameliorate the loss of function after spinal cord injuries, with the hope of developing interventions to benefit patients. Although many reports suggest positive effects, there is growing concern over the quality of the available preclinical data. It is therefore important to ask whether this worldwide investigative process is close to defining a cell transplant protocol that could be translated into human patients with a realistic chance of success. This review systematically examines the strength of the preclinical evidence and outlines mechanisms by which transplanted cells may mediate their effects in spinal cord injuries. First, we examined changes in voluntary movements in the forelimb associated with cell transplants after partial cervical lesions. Second, we examined the efficacy of transplanted cells to restore electrophysiological conduction across a complete thoracic lesion. We postulated that cell therapies found to be successful in both models could reasonably have potential to treat human patients. We conclude that although there are data to support a beneficial effect of cell transplantation, most reports provide only weak evidence because of deficits in experimental design. The mechanisms by which transplanted cells mediate their functional effects remain unclear.
[Show abstract][Hide abstract] ABSTRACT: Myelination is essential for rapid impulse conduction in the CNS, but what determines whether an individual axon becomes myelinated remains unknown. Here we show, using a myelinating coculture system, that there are two distinct modes of myelination, one that is independent of neuronal activity and glutamate release and another that depends on neuronal action potentials releasing glutamate to activate NMDA receptors on oligodendrocyte lineage cells. Neuregulin switches oligodendrocytes from the activity-independent to the activity-dependent mode of myelination by increasing NMDA receptor currents in oligodendrocyte lineage cells 6-fold. With neuregulin present myelination is accelerated and increased, and NMDA receptor block reduces myelination to far below its level without neuregulin. Thus, a neuregulin-controlled switch enhances the myelination of active axons. In vivo, we demonstrate that remyelination after white matter damage is NMDA receptor-dependent. These data resolve controversies over the signalling regulating myelination and suggest novel roles for neuregulin in schizophrenia and in remyelination after white matter damage.
[Show abstract][Hide abstract] ABSTRACT: The increasing effectiveness of new disease-modifying drugs that suppress disease activity in multiple sclerosis has opened up opportunities for regenerative medicines that enhance remyelination and potentially slow disease progression. Although several new targets for therapeutic enhancement of remyelination have emerged, few lend themselves readily to conventional drug development. Here, we used transcription profiling to identify mitogen-activated protein kinase (Mapk) signalling as an important regulator involved in the differentiation of oligodendrocyte progenitor cells (OPCs) into oligodendrocytes. We show in tissue culture that activation of Mapk signalling by elevation of intracellular levels of cyclic adenosine monophosphate (cAMP) using administration of either dibutyryl-cAMP or inhibitors of the cAMP-hydrolysing enzyme phosphodiesterase-4 (Pde4) enhances OPC differentiation. Finally, we demonstrate that systemic delivery of a Pde4 inhibitor leads to enhanced differentiation of OPCs within focal areas of toxin-induced demyelination and a consequent acceleration of remyelination. These data reveal a novel approach to therapeutic enhancement of remyelination amenable to pharmacological intervention and hence with significant potential for translation.
EMBO Molecular Medicine 12/2013; 5(12):1918-34. · 7.80 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The lack of therapies for progressive multiple sclerosis highlights the need to understand the regenerative process of remyelination that can follow CNS demyelination. This involves an innate immune response consisting of microglia and macrophages, which can be polarized to distinct functional phenotypes: pro-inflammatory (M1) and anti-inflammatory or immunoregulatory (M2). We found that a switch from an M1- to an M2-dominant response occurred in microglia and peripherally derived macrophages as remyelination started. Oligodendrocyte differentiation was enhanced in vitro with M2 cell conditioned media and impaired in vivo following intra-lesional M2 cell depletion. M2 cell densities were increased in lesions of aged mice in which remyelination was enhanced by parabiotic coupling to a younger mouse and in multiple sclerosis lesions that normally show remyelination. Blocking M2 cell-derived activin-A inhibited oligodendrocyte differentiation during remyelination in cerebellar slice cultures. Thus, our results indicate that M2 cell polarization is essential for efficient remyelination and identify activin-A as a therapeutic target for CNS regeneration.
[Show abstract][Hide abstract] ABSTRACT: The myelin sheath, generated by oligodendrocytes in the central nervous system (CNS), is crucial to neuronal function, enabling rapid propagation of nerve impulses and providing trophic support to the axon. Remyelination is the default response to myelin damage. Oligodendrocyte precursor cells, distributed throughout both the grey and white matter of the CNS, are activated in response to myelin injury, undergoing proliferation, migration to the site of damage and differentiation into mature myelinating oligodendrocytes. The end result is complete reconstruction of the area of myelin loss. However, this remarkable regenerative capacity of the CNS becomes less efficient with age and can show clinically significant failure in diseases such as multiple sclerosis. Without the myelin sheath, neuronal function and survival is compromised, leading to axonal degeneration and progressive deterioration in neurological function. Therapies to enhance remyelination could offer a means to prevent the neurological decline of chronic demyelinating disease. In order to develop such therapies, a detailed understanding of the process of remyelination, the major cellular players involved and the mechanisms of remyelination failure is needed. As the intricacies of remyelination continue to be unravelled, effective remyelination therapies are ever closer to becoming a reality.
Journal of comparative pathology 07/2013; · 1.73 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Ischaemia leads to increased proliferation of progenitors in the subependymal zone (SEZ) neurogenic niche of the adult brain and to generation and migration of newborn neurons. Here we investigated the spatiotemporal characteristics of the mitotic activity of adult neural stem and progenitor cells in the SEZ during the sub-acute and chronic post-ischemic phases. Ischaemia was induced by performing a 1h unilateral middle cerebral artery occlusion (MCAO) and tissue was collected 4/5 weeks and 1 year after the insult. Neural stem cells (NSCs) responded differently from their downstream progenitors to MCAO, with NSCs being activated only transiently while progenitors remain activated even at 1 year post-injury. Importantly, mitotic activation was observed only in the affected areas of the niche and specifically in the dorsal half of the SEZ. Analysis of the topography of mitoses, in relation to the anatomy of the lesion and to the position of ependymal cells and blood vessels, suggested an interplay between lesion-derived recruiting signals and the local signals that normally control proliferation in the chronic post-ischemic phase.
[Show abstract][Hide abstract] ABSTRACT: Peroxisome proliferator-activated receptor gamma-coactivator-1 alpha (PGC1a) is involved in energy and lipid metabolism, and its loss leads to neurodegenerative changes in the striatum. Here we performed lipidomic analysis on brain extracts from PGC1a mutant and wild-type mice. We found increased phosphatidylcholine and decreased ceramides in the brain of PGC1a-deficient mice. An analysis of lipid raft fractions revealed increased ceramide, glucocylceramides and GM1 ganglioside in the PGC1a mutants. In the cerebellum, we observed a decrease in proteins associated with myelination, but were unable to detect any morphological abnormalities in compact myelin formation in PGC1a mutants compared with wild-type mice. Although PGC1a is involved in lipid biosynthesis, we concluded that altered lipid composition in the PGC1a mutant did not directly affect central nervous system myelin morphology.
European Journal of Neuroscience 06/2013; · 3.67 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The growing burden of the rapidly ageing global population has reinvigorated interest in the science of ageing and rejuvenation. Among organ systems, rejuvenation of the central nervous system (CNS) is arguably the most complex and challenging of tasks owing, among other things, to its startling structural and functional complexity and its restricted capacity for repair. Thus, the prospect of meaningful rejuvenation of the CNS has seemed an impossible goal; however, advances in stem cell science are beginning to challenge this assumption. This Review outlines these advances with a focus on ageing and rejuvenation of key endogenous stem and progenitor cell compartments in the CNS. Insights gleaned from studies of model organisms, chiefly rodents, will be considered in parallel with human studies.
Development 06/2013; 140(12):2562-2575. · 6.27 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Traditionally, in vitro stem cell systems have used oxygen tensions that are far removed from the in vivo situation. This is particularly true for the central nervous system, where oxygen (O2) levels range from 8% at the pia to 0.5% in the midbrain, whereas cells are usually cultured in a 20% O2 environment. Cell transplantation strategies therefore typically introduce a stress challenge at the time of transplantation as the cells are switched from 20% to 3% O2 (the average in adult organs). We have modeled the oxygen stress that occurs during transplantation, demonstrating that in vitro transfer of neonatal rat cortical neural precursor cells (NPCs) from a 20% to a 3% O2 environment results in significant cell death, whereas maintenance at 3% O2 is protective. This survival benefit translates to the in vivo environment, where culture of NPCs at 3% rather than 20% O2 approximately doubles survival in the immediate post-transplantation phase. Furthermore, NPC fate is affected by culture at low, physiological O2 tensions (3%), with particularly marked effects on the oligodendrocyte lineage, both in vitro and in vivo. We propose that careful consideration of physiological oxygen environments, and particularly changes in oxygen tension, has relevance for the practical approaches to cellular therapies.
STEM CELLS TRANSLATIONAL MEDICINE 05/2013; · 3.60 Impact Factor