Layers Of Myelin Sheath Research Articles

4D printing of stretchable nanocookie@conduit material hosting biocues and magnetoelectric stimulation for neurite sprouting

A high-frequency magnetic field (MF) generates an electric current by charging conductors that enable the induction of various biological processes, including changes in cell fate and programming. In this study, we show that electromagnetized carbon porous nanocookies (NCs) under MF treatment facilitate magnetoelectric conversion for growth factor release and cell stimulation to induce neuron cell differentiation and proliferation in vitro and in vivo. Integrating four-dimensional printing technology, the NCs are exposed on the surface, which enhances the cell adhesion and allows direct manipulation of electromagnetic stimulation of the cells. Remarkably, large amounts of growth factor encapsulated in NC@conduit resulted in excellent permeability and on-demand release, improving the in vivo layers of myelin sheaths and directing the axon orientation at 1 month postimplantation. This study offers proof of principle for MF-guided in vivo neuron regeneration as a potentially viable tissue regeneration approach for neuronal diseases.

Effect of the CSF1R inhibitor PLX3397 on remyelination of corpus callosum in a cuprizone-induced demyelination mouse model.

Multiple sclerosis (MS) is a chronic inflammatory disease affecting the central nervous system (CNS). Despite introducing multiple immunomodulatory approaches for MS, there are still major concerns about possible ways for improving remyelination in this disease. Microglia exert essential roles in regulation of myelination processes, and interaction between colony-stimulating factor 1 (CSF1) with its receptor CSF1R is considered as a key regulator of microglial differentiation and survival. The aim of this study was to investigate possible roles for a CSF1R inhibitor PLX3397 in recovery of central myelination processes. Chronic demyelination was induced in mice by addition of 0.2% cuprizone to the chow for 12 weeks. Next, animals were undergoing a diet containing 290 mg/kg PLX3397 to induce microglial ablation. The PLX3397 treatment caused a significant decrease in the rate of expression for the CSF1/CSF1R axis, and a reduction in the protein expressions for the microglial marker Iba-1 and for the oligodendrocyte marker Olig-2. Findings from Luxol fast blue (LFB) staining and transmission electron microscopy (TEM) showed an increase in the rate of myelination for the mice receiving PLX3397. The rate of destruction in the nerve fibers and the extent of the gaps formed between layers of myelin sheaths was also reduced after the treatment with PLX3397. In addition, animals experienced an improvement in recovery of motor deficit after receiving PLX3397 (for all P < 0.05). It could be concluded that PLX3397 could retain myelination in the MS model possibly through regulation of the myelin environment.

Functional regeneration of the brain: white matter matters.

Functional regeneration of the brain: white matter matters.

Direct visualization of membrane architecture of myelinating cells in transgenic mice expressing membrane-anchored EGFP.

Myelinogenesis is a complex process that involves substantial and dynamic changes in plasma membrane architecture and myelin interaction with axons. Highly ramified processes of oligodendrocytes in the central nervous system (CNS) make axonal contact and then extrapolate to wrap around axons and form multilayer compact myelin sheathes. Currently, the mechanisms governing myelin sheath assembly and axon selection by myelinating cells are not fully understood. Here, we generated a transgenic mouse line expressing the membrane-anchored green fluorescent protein (mEGFP) in myelinating cells, which allow live imaging of details of myelinogenesis and cellular behaviors in the nervous systems. mEGFP expression is driven by the promoter of 2'-3'-cyclic nucleotide 3'-phosphodiesterase (CNP) that is expressed in the myelinating cell lineage. Robust mEGFP signals appear in the membrane processes of oligodendrocytes in the CNS and Schwann cells in the peripheral nervous system (PNS), wherein mEGFP expression defines the inner layers of myelin sheaths and Schmidt-Lanterman incisures in adult sciatic nerves. In addition, mEGFP expression can be used to track the extent of remyelination after demyelinating injury in a toxin-induced demyelination animal model. Taken together, the membrane-anchored mEGFP expression in the new transgenic line would facilitate direct visualization of dynamic myelin membrane formation and assembly during development and process remodeling during remyelination after various demyelinating injuries.

Morphological Progression of Myelin Abnormalities in IgM-Monoclonal Gammopathy of Undetermined Significance Anti-Myelin-Associated Glycoprotein Neuropathy

To characterize the morphological progression of neuropathy associated with immunoglobulin M-monoclonal gammopathy of undetermined significance with anti-myelin-associated glycoprotein antibody, we assessed histopathologic features of sural nerve specimens from 15 patients, emphasizing widely spaced myelin (WSM), demyelination, and tomaculous changes. The frequency of WSM correlated with that of demyelination and tomaculous appearance in teased-fiber preparations. In longitudinal sections at nodes of Ranvier and paranodal regions, the spaces between terminal myelin loops, particularly those adjacent to the node of Ranvier, were widened, indicating an early change before demyelination, and there was concomitant swelling of terminal myelin loops. Some conspicuously swollen terminal myelin loops were detached from the paranodal axolemma, thereby widening the nodes of Ranvier. Tomacula coexisted frequently with redundant myelin loops and WSM, particularly in the outermost layer of myelin sheaths, suggesting that loosening of the outer layers contributes to their formation. By immunofluorescence microscopy, immunoglobulin M and myelin-associated glycoprotein were colocalized in paranodal regions and Schmidt-Lanterman incisures. Confocal analysis revealed colocalization of immunoglobulin M and complement product C3d corresponding to the area of WSM. Thus, morphological changes in terminal myelin loops, formation of WSM at paranodes, and subsequent dissociation from paranodal axolemma (which may be associated with activation of the complement pathway) likely contribute to demyelination in this condition. Loosening of compact myelin seems to contribute to tomacula formation.

MMP-mediated cleavage of β-dystroglycan in myelin sheath is involved in autoimmune neuritis

α-/β-Dystroglycans (DG) located at the outmost layer of myelin sheath play a critical role in its formation and stability in the peripheral nerve system. The demyelination of nerve fibers is present in autoimmune neuritis, however, it is not known about the molecular mechanisms underlying this pathological process. In an animal model of experimental autoimmune neuritis, we observed that β-DG cleavage was associated with the demyelination of peripheral nerves. The neuritis and β-DG cleavage were accompanied by matrix metalloproteinase (MMP)-2/-9 over-expressions and attenuated by captopril, a MMP inhibitor. The blockade of MMPs also improves clinical signs. Our results reveal a crucial role of MMP-mediated β-DG cleavage in autoimmune neuritis, such as Guillain–Barre’ syndrome, and bring insights into therapeutic strategies for autoimmune diseases.

Characterization of glial cell line‐derived neurotrophic factor family receptor α‐1 in peripheral nerve Schwann cells

Glial cell line-derived neurotrophic factor (GDNF) family receptor alpha-1 (GFRalpha-1) is a receptor component of GDNF that associates with and activates the tyrosine kinase receptor Ret. To further understand GDNF and its receptor system in the PNS, we first characterized the expression of GFRalpha-1 in bovine peripheral nerve in vivo. GFRalpha-1 immunoreactivity was localized adjacent to the outermost layer of myelin sheath, as well as in the endoneurium and axoplasm. In a fractionation study, GFRalpha-1 was recovered mostly in the soluble fraction, although a small amount was recovered in the membrane fraction. A substantial amount of GFRalpha-1 in the membrane fraction was extractable by detergent and alkaline conditions. To further clarify the expression of GFRalpha-1 in Schwann cells, we examined cultured rat Schwann cells and the Schwannoma cell line RT4. Schwann cells expressed GFRalpha-1 in both the soluble/cytosolic and membrane fractions, and the membrane form of GFRalpha-1 was expressed at the outer surface of the Schwann cell plasma membrane. We also confirmed the secretion of the soluble form of GFRalpha-1 from Schwannoma cells in a metabolic labeling experiment. These data contribute to our knowledge of the production, expression and functions of GFRalpha-1 in the PNS.

Anti-MOG autoantibodies in Italian multiple sclerosis patients: specificity, sensitivity and clinical association.

There is considerable evidence that multiple sclerosis (MS) is an immune-mediated disease characterized by infiltration of inflammatory cells into the CNS and demyelination. Several myelin proteins may be encephalitogenic, including myelin basic protein, proteolipid protein and myelin oligodendrocyte glycoprotein (MOG), the latter being expressed on the external layer of myelin sheaths and hence accessible to antibody attack. We investigated MOG autoreactivity in serum and cerebrospinal fluid (CSF) by ELISA, employing the recombinant extracellular domain of MOG as antigen. We tested serum samples from 262 MS patients (175 relapsing-remitting, 43 primary progressive and 44 secondary progressive), 131 patients with other neurological diseases (OND) and 307 healthy controls. No patients or controls were receiving immunomodulating treatments. We found anti-MOG antibodies in the serum of 13.7% MS patients, mainly in those with secondary progressive MS (25%), in 13.7% of OND patients and in 6.2% of controls. We found a direct correlation (R(2) = 0.6, P = 0.002) between disease severity and anti-MOG titer only in patients with primary and secondary progressive MS. Anti-MOG antibodies were present in the CSF of 11.4% MS patients and 18.9% OND patients. Intrathecal synthesis of anti-MOG antibodies was demonstrated in four (4.5%) of MS patients and no OND patients. Anti-MOG antibodies are not specific for MS; however, they may characterize a subset of MS patients and this may be revealed by serial assays in relation to changing disease phase.

EXPRESSION AND DISTRIBUTION OF TRANSCRIPTION FACTOR NF‐KB IN PERIPHERAL NEUROPATHIES OF DIFFERENT ORIGIN

Background: Nuclear Factor‐kB (NF‐kB) is a transcription factor expressed ubiquitously, that plays an important role in immune and inflammatory responses by regulating the expression of different genes. Furthermore many studies have shown its involvement in the apoptosis of neurodegenerative disorders. Objective: The aim of the study was to evaluate the expression of this factor in inflammatory demyelinating and axonal neuropathies as well as in non‐inflammatory neuropathies. Materials and Methods: We studied the expression of NF‐kB by immunocytochemistry and Western Blot of nuclear extract proteins in sural nerve biopsies of 11 patients with peripheral neuropathies of different origin: 3 with CIDP, 2 with chronic axonal inflammatory neuropathy, 2 with vasculitis, 2 with CMT1 and 2 with CMT2. Three normal nerves served as controls. Immunohistochemistry for macrophages, B, CD4 and CD8 cell subsets was also performed. Results: NF‐kB expression was found in some endoneurial vessel walls, many cell nuclei and in the outermost layer of myelin sheath of some nerve fibers in inflammatory axonal and demyelinating neuropathies. Few positive cells were found in hereditary neuropathies whereas no immunoreactivity was detected in normal controls. Western blot analysis confirmed such findings. Conclusion: Our findings underline the role of NF‐kB in the immune response of inflammatory neuropathies. NF‐kB reactivity in hereditary neuropathies may be related to the apoptotic mechanism.

Perineurium talin immunoreactivity decreases in diabetic neuropathy

We studied the immunolocalization of Dp116 (a 116 kDa protein product of the dystrophin gene), vinculin, talin, vimentin, desmin, spectrin and titin in the sural nerve biopsies of 25 patients with peripheral neuropathies of different origin. 4 patients presented with HMSN type 1, 4 with HMSN type 2, 2 with HNPP, 4 with CIDP, 5 with chronic axonal neuropathy of unknown origin, 3 with vasculitic neuropathy, 3 with diabetic neuropathy. Expression and localization of Dp116, vinculin, vimentin, desmin, spectrin and titin did not differ from normal control cases. Spectrin and titin immunoreactivities were absent and desmin was occasionally found in few epineurial vessels. A thin rim of Dp116 binding surrounded the outermost layer of myelin sheaths. Perineurium and epineurial vessels stained deeply for vinculin. Vimentin immunoreactivity was seen in all endoneurial, perineurial and epineurial cells. Immunoreactivity for talin was normally found at endoneurial and epineurial vessel walls, perineurial cells and epineurial fibroblasts in all the sural nerves except diabetic nerves. In the latter, whereas talin binding was normal in the vessel walls and epineurial fibroblasts, it was markedly reduced in the perineurium. On immunoblot, two bands at 235 and 190 kDa were found in the sural nerves with the antibody anti-talin, and both were reduced only in the patients with diabetic neuropathy. We postulate that decreased perineurium talin in diabetic polyneuropathy may be related to the known alterations of the tight junctions of the perineurial cells, which have been proposed to be a contributory factor to impaired permeability barrier properties. © 1997 Elsevier Science B.V. All rights reserved.

Characteristics of the myelin sheath membrane of the spiral ganglion cell and the cochlear nerve of guinea pig.

The membrane structures of the myelin sheath of guinea pig spiral ganglion cell and cochlear nerve were examined using freeze-fracturing, rapid freezing and freeze substitution, and deep-etching techniques. The intermediate dense lines of the myelin sheath were observed only in the outermost several layers using thin and thick (1 micron) section techniques. The spaces between the two adjacent major dense lines at any layer of myelin sheath were occupied by flocculent materials in the freeze-substituted specimen. Periods between the two adjacent major dense lines were 13.3 nm in conventional fixation and 11.7 nm in rapid freezing followed by freeze substitution, respectively. A freeze-fracturing study showed no differences in membrane structures at any layer of myelin sheath. Developed tight-junction strands were observed parallel to the long axis of the myelin sheath in the deep-etched specimen. These findings may show the characteristics of myelin sheath membrane in the spiral ganglion cell and cochlear nerve of the guinea pig.

Dystroglycan Is a Dual Receptor for Agrin and Laminin-2 in Schwann Cell Membrane

We have shown previously that alpha-dystroglycan with a molecular mass of 120 kDa is a Schwann cell receptor of laminin-2, the endoneurial isoform of laminin comprised of the alpha2, beta1, and gamma1 chains. In this paper, we show that Schwann cell alpha-dystroglycan is also a receptor of agrin, an acetylcholine receptor-aggregating molecule having partial homology to laminin alpha chains in the C terminus. Immunochemical analysis demonstrates that the peripheral nerve isoform of agrin is a 400-kDa component of the endoneurial basal lamina and is co-localized with alpha-dystroglycan surrounding the outermost layer of myelin sheath of peripheral nerve fibers. Blot overlay analysis demonstrates that both endogenous peripheral nerve agrin and laminin-2 bind to Schwann cell alpha-dystroglycan. Recombinant C-terminal fragment of the peripheral nerve isoform of agrin also binds to Schwann cell alpha-dystroglycan, confirming that the binding site for Schwann cell alpha-dystroglycan resides in the C terminus of agrin molecule. Furthermore, the binding of recombinant agrin C-terminal fragment to Schwann cell alpha-dystroglycan competes with that of laminin-2. All together, these results indicate that alpha-dystroglycan is a dual receptor for agrin and laminin-2 in the Schwann cell membrane.

Differentiation between dysmyelination and demyelination using magnetic resonance diffusional anisotropy

Using magnetic resonance (MR) diffusion-weighted method, we examined the optic and the trigeminal nerves of jimpy and twitcher mice, considered to be animal models of Pelizaeus-Merzbacher disease, hypomyelination disorder, and Krabbe disease, demyelination disorder, respectively. In jimpy mice, diffusional anisotropy of optic nerve did not show a significant difference compared to age-matched control mice, suggesting that diffusional anisotropy does exist in absence of multiple layers of myelin sheath. In twitcher mice, diffusional anisotropy was attenuated remarkably in the optic and trigeminal nerves. Loss of axonal straightness on longitudinal section confirmed by electron microscopy appeared to be the principal explanation for it. It is further suggested that this MR diffusion-weighted imaging method enables us to differentiate hypomyelination from demyelination in vivo.

Differential expression of dystrophin, utrophin and dystrophin-associated proteins in peripheral nerve

The dystrophin-glycoprotein complex is a novel laminin receptor in skeletal muscle. Dystrophin-associated proteins are comprised of an extracellular glycoprotein of 156 kDa (156DAG), transmembrane glycoproteins of 50 kDa (50DAG), 43 kDa (43DAG) and 35 kDa (35DAG), and a cytoskeletal protein of 59 kDa (59DAP). The laminin-binding 156DAG and 43DAG are encoded by a single gene and are now called α- and β-dystroglycan, respectively. In neuromuscular junctions, utrophin, an autosomal homologue of dystrophin, is associated with sarcolemmal proteins identical or immunologically homologous to the dystrophin-associated proteins. Here we demonstrate the co-localization of Dp116 (a 116 kDa protein product of the DMD gene), full-size utrophin, α- and β-dystroglycan, 59DAP and 35DAG in a thin rim surrounding the outermost layer of myelin sheath of peripheral nerve fibers. The α-dystroglycan in peripheral nerve had molecular weight of 120 kDa instead of 156 kDa, suggesting different levels of glycosylation between skeletal muscle and peripheral nerve. In sharp contrast to skeletal muscle, however, full-size dystrophin and 50DAG were undetectable in peripheral nerve. Our results demonstrate the varied expression of the components of the dystrophin/utrophin-glycoprotein complex between skeletal muscle and peripheral nerve suggesting the complex may exist in varied compositions and have varied functions in these two tissues.

The monoclonal antibodies Elec-39, HNK-1 and NC-1 recognize common structures in the nervous system and muscles of vertebrates

The IgM monoclonal antibodies, Elec-39, HNK-1 and NC-1, recognize the same subset of Torpedo electric organ acetylcholinesterase (AChE). We show that they react against a glycosphingolipid (SGPG) containing a sulfated glucuronic acid (SGA). The three antibodies appear essentially identical in their specificity but differ in their affinity for the antigens. We have examined their binding in the CNS, nerves and muscles of several vertebrate species, at the optical and in some cases at the electron microscope level. All three antibodies label the same structures: they show diffuse staining around neuromuscular endplates and label the plasma membrane of the Schwann cells, surrounding the outer layer of myelin sheaths. In the adult rat CNS, the antibodies label certain defined structures, notably extracellular material in the habenula and in the CA2 layer of the hippocampus. In the cortex and cerebellum, they label the surface of neural processes and terminals apposed to large multipolar neurons and Purkinje cells, as well as membranous material contained in inclusions dispersed in the cytoplasm of these neurons. These localizations are consistent with the suggestion that the SGA-antigens may be involved in cellular interactions.

Uncompacted myelin lamellae in dysglobulinemic neuropathy

Uncompacted lamellae, located preferentially in inner layers of myelin sheath, were observed in biopsied sural nerves of 3 cases of dysglobulinemic neuropathy, in which the main pathological findings of myelinated fibers were those of segmental demyelination and remyelination, and axonal degeneration with concurrent marked decrease of myelinated fiber density. The presence of uncompacted myelin lamellae is well explained by the irregular distribution of adaxonal, incisural and paranodal cytoplasm of the Schwann cell.

Ultrastructural study of myelinating cells and sub-pial astrocytes in developing rat spinal cord

The anterior funiculus of the spinal cervical cord of post-natal rats was examined ultrastructurally. The myelinating cells found one day after birth contained a large amount of evenly distributed ribosomes up to the outer tongue of mesaxons, representing the cytoplasmic density. These cells were separated by astrocytic processes from the pial basement membrane, even when they were located on the pial surface. Astrocytes contained glial fibrils from one day onwards and often attached their processes to the pial basement membrane. Although the cytoplasmic processes of astrocytes occasionally wrapped axons, they were never shown to form the initial layer of myelin sheaths. However, the tenuous processes of the sub-pial astrocytes were occasionally rolled in myelin lamellae, as if a part of the myelin sheaths was constructed by astrocytic processes. The interpretation for this finding is discussed in relation to function and potency of the astrocytes, and variations and anomalies of nervous ontogeny.

RESPONSE OF SINGLE RANVIER NODES TO ELECTRICAL STIMULI

A VERTEBRATE motor nerve fiber is thickly covered with a layer of myelin sheath except at narrow gaps called nodes of Ranvier. As the myelin sheath is shown to be composed of an electric insulator, investigation of the physiological properties of these nodes seems to disclose the nature of the plasma membrane of an infinitesimal dimension and consequently to enable us better to understand the activity of the surface layer of a cell in general. The present investigation is undertaken to secure a series of oscillograms which seemed to us to indicate the basic properties of the plasma membrane at the nodes. The temporal configuration of the electric responses and the latencies of the responses to rectangular current pulses are our main concern in this paper.