The transition zone of the central nervous system-peripheral nervous system of the adult rat; ultrastructural and immunocytochemical studies: a new function of the astroglia?]

At the transition between central nervous system (CNS) and peripheral nervous system (PNS), the CNS compartment forms cone-shaped incursions into the peripheral part of the dorsal root. The ultrastructural study of the CNS-PNS transitional zone shows that this region is particularly rich in astrocyte processes. In an attempt to investigate the possible role of the CNS-PNS interface astrocytes in myelin formation, a photonic microscopy immunocytochemical study has been done with anti-GFAP and anti-MBP sera. The CNS glial expansion shows an important GFAP immunoreactivity with intimate association between astrocyte processes and myelinated axons. This may indicate that the transitional myelin originates from astrocytes. The same region is also MBP-positive. Two explanations are considered: some astrocytes form transitional myelin sheathes and express MBP epitopes, or oligodendrocytes, with cell bodies distant from the CNS-PNS interface, send myelinating cytoplasmic expansions which are not shown by the techniques we used.     

Schwann cell expression of an oligodendrocyte-like remyelinating pattern after ethidium bromide injection in the rat spinal cord

Schwann cells are recognized by their capacity of producing single internodes of myelin around axons of the peripheral nervous system. In the ethidium bromide (EB) model of primary demyelination in the brainstem, it is observed the entry of Schwann cells into the central nervous system in order to contribute to the myelin repair performed by the oligodendrocytes that survived to the EB gliotoxic action, being able to even remyelinate more than one axon at the same time, in a pattern of repair similar to the oligodendroglial one. The present study was developed in the spinal cord to observe if Schwann cells maintained this competence of attending simultaneously different internodes. It was noted that, on the contrary of the brainstem, Schwann cells were the most important myelinogenic cells in the demyelinated site and, although rare, also presented the capacity of producing more than one internode of myelin in distinct axons.     

Behaviour of oligodendrocytes and Schwann cells in an experimental model of toxic demyelination of the central nervous system.

Oligodendrocytes and Schwann cells are engaged in myelin production, maintenance and repairing respectively in the central nervous system (CNS) and the peripheral nervous system (PNS). Whereas oligodendrocytes act only within the CNS, Schwann cells are able to invade the CNS in order to make new myelin sheaths around demyelinated axons. Both cells have some limitations in their activities, i.e. oligodendrocytes are post-mitotic cells and Schwann cells only get into the CNS in the absence of astrocytes. Ethidium bromide (EB) is a gliotoxic chemical that when injected locally within the CNS, induce demyelination. In the EB model of demyelination, glial cells are destroyed early after intoxication and Schwann cells are free to approach the naked central axons. In normal Wistar rats, regeneration of lost myelin sheaths can be achieved as early as thirteen days after intoxication; in Wistar rats immunosuppressed with cyclophosphamide the process is delayed and in rats administered cyclosporine it may be accelerated. Aiming the enlightening of those complex processes, all events concerning the myelinating cells in an experimental model are herein presented and discussed.     

In Vivo Expression of the Arf6 Guanine-Nucleotide Exchange Factor Cytohesin-1 in Mice Exhibits Enhanced Myelin Thickness in Nerves

The myelin sheath consists of a unique multiple layer structure that acts as an insulator between neuronal axons to enhance the propagation of the action potential. In neuropathies such as demyelinating or dismyelinating diseases, chronic demyelination and defective remyelination occur repeatedly, leading to more severe neuropathy. As yet, little is known about the possibility of drug target-specific medicine for such diseases. In the developing peripheral nervous system (PNS), myelin sheaths form as Schwann cells wrap individual axons. It is thought that the development of a drug promoting myelination by Schwann cells would provide effective therapy against peripheral nerve disorders: to test such treatment, genetically modified mice overexpressing the drug target molecules are needed. We previously identified an Arf6 activator, the guanine-nucleotide exchange factor cytohesin-1, as the signaling molecule controlling myelination of peripheral axons by Schwann cells; yet, the important issue of whether cytohesin-1 itself promotes myelin thickness in vivo has remained unclear. Herein, we show that, in mouse PNS nerves, Schwann cell-specific expression of wild-type cytohesin-1 exhibits enhanced myelin thickness. Downstream activation of Arf6 is also seen in these transgenic mice, revealing the involvement of the cytohesin-1 and Arf6 signaling unit in promoting myelination. These results suggest that cytohesin-1 may be a candidate for the basis of a therapy for peripheral neuropathies through its enhancement of myelin thickness.     

Neuropathology of Charcot-Marie-Tooth and related disorders

The peripheral nervous system (PNS), with all its branches and connections, is so complex that it is impossible to study all components at the light or electron microscopic level in any individual case; nevertheless, in certain diseases a simple nerve biopsy may suffice to arrive at a precise diagnosis. Structural changes of the PNS in neuropathies of the Charcot-Marie-Tooth (CMT) type and related disorders comprise various components of the PNS. These include peripheral motor, sensory, and autonomous neurons with their axons, Schwann cells, and myelin sheaths in the radicular and peripheral nerves as well as satellite cells in spinal and autonomous ganglia. Astrocytes, oligodendroglial cells, and microglial cells around motor neurons in the anterior horn and around sensory neurons in other areas of the spinal cord are also involved. In addition, connective tissue elements such as endoneurial, perineurial, and epineurial components including blood and lymph vessels play an important role. This review focuses on the cellular components and organelles involved, that is, myelin sheaths, axons with their micro-tubules and neurofilaments; nuclei, mitochondria, endoplasmic reticulum, and connective tissue including the perineurium and blood vessels. A major role is attributed to recent progress in the pathomorphology of various types of CMT1, 2,4, CMTX, and HMNSL, based on light and electron microscopic findings, morphometry, teased fiber studies, and new immunohisto-chemical results such as staining of certain periaxin domains in CMT4F.     

Spinal cord multiple sclerosis lesions in Japanese patients: Schwann cell remyelination occurs in areas that lack glial fibrillary acidic protein (GFAP)

Summary To extend earlier observations on Schwann cell remyelination in multiple sclerosis (MS) lesions (Itoyama et al. 1983) we immunostained spinal cord sections from eight Japanese MS patients with antiserum to P_o glycoprotein, a major constituent of peripheral nervous system (PNS) myelin, myelin basic protein (MBP), and glial fibrillary acidic protein (GFAP). Spinal cord sections from six of the eight Japanese MS patients contained large clusters of peripheral myelin sheaths with anti-P_o immunoreactivity. In lesions found in four of the six patients, thousands of P_o-stained PNS myelin sheaths were present. Necrosis was prominent in these lesions which included more than half of the spinal cord's transverse area. The number and density of regenerating myelin sheaths of peripheral origin were much greater than we observed in MS spinal cord lesions of white people (Itoyama et al. 1983). Anti-GFAP immunoreactivity was present in most brain and spinal cord lesions. However, the areas in lesions that contained large groups of PNS myelin sheaths lacked anti-GFAP immunoreactivity. Our data suggest that spinal MS lesions that are large, severely demyelinated, and partially necrotic may contain factors that inhibit fibrous astrogliosis. These factors, other substances in the large lesions and/or the lack of astrocytic scarring could then promote Schwann cell invasion, multiplication, and remyelination of surviving axons.     

Cytochemical demonstration of TPPase in myelinated fibers in the central and peripheral nervous system of the rat

Thiamine pyrophosphatase (TPPase) activity was demonstrated by means of cytochemistry and electron microscopy in association with myelinated fibers in the central and peripheral nervous system of the rat. The areas studied included corpus callosum, hippocampus, cerebral cortex, cervical spinal cord and sciatic nerves. In the myelin sheaths, the enzymatic activity was found in 3 locations: (1) within oligodendroglial and Schwann cytoplasmic clefts between myelin lamellae; (2) in the major dense line of myelin; and (3) within the periaxonal space. In addition to this myelin-associated TPPase, enzymatic activity was also observed in specific cytoplasmic localizations in myelinogenic cells. Oligodendrocytes displayed TPPase activity within the nuclear envelope and the endoplasmic reticulum cisternae, whereas Schwann cells displayed TPPase activity within the endoplasmic reticulum and Golgi saccules. The results are discussed in relation to the role that TPPase might play in myelinated fibers, including roles in the conduction of nerve impulses or roles in the maintenance of structural configuration of myelin sheaths.     

Studies with antisera against peripheral nervous system myelin and myelin basic proteins. I. Effects of antiserum upon living cultures of nervous tissue

We studied the effects of antiserum against rat peripheral nervous system (PNS) myelin, rat or chicken central nervous system myelin basic protein (BP), or rabbit P₂ protein from PNS myelin on myelinated cultures containing only rat dorsal root ganglion neurons and Schwann cells. While anti-PNS myelin serum consistently produced segmental PNS demyelination, anti-BP serum and anti-P₂ serum did not. The culture results suggest that the myelin PNS proteins P₁ (identical to basic protein from central nervous system myelin) and P₂ are not exposed on the extracellular surfaces of myelin-related Schwann cells in tissue culture.     

Discontinuous transition of myelin structure at the junction between central and peripheral components of the eighth cranial nerve, as disclosed by X-ray diffraction

Scanning along human acoustovestibular nerves from cross-sections closely proximal to the brain to locations distinctly peripheral thereto, by means of small-angle X-ray diffraction, has disclosed transitional junctions at which the myelin structure typical of central nervous system (CNS) axons gives way to one characteristic of peripheral (PNS) fibers. The junctions correspond to regions along the nerves, previously recognized histologically, at which the satellite cells responsible for axon myelination change character, from the oligodendrocytes of the CNS to the Schwann cells of the PNS. Thus the structural discontinuity between the CNS and PNS myelins can be ascribed to differences in the biosynthetic processes of the respective satellite cells. Junctions of this kind are to be expected in all cranial and spinal roots near the locations where they leave the CNS.     

Reinnervation, myelination and organization of iris tissue implanted into the rat midbrain--an ultrastructural study

The ultrastructure of autologous irides implanted into the midbrain of mature Sprague-Dawley rats was studied over a time period of 4-14 days. Most features of the normal iris still persisted throughout this time, including typical iris blood vessels and amelanotic melanocytes. At 5-4 days after implantation, the normal innervation of the implanted iris had degenerated, except for the presence of intact Schwann cells and some myelin debris. Reinnervation, beginning about the seventh day, proceeded rapidly. By the fourteenth day, extensive reinnervation of the implant was evidenced by the presence of numerous small (0.1-0.6 micrometer) and large (2-4 micrometer) non-myelinated axons ensheathed in Schwann cell cytoplasm. Axonal varicosities, filled either with dense core or clear core vesicles, formed junctions with axons. These junctions were characterized by an accentuation of areas of the axonal membrane and pre- or post-junctional thickenings; however, we did not observe typical synaptic complexes. Some large axons within the myelinated iris developed thick myelin sheaths of the peripheral nervous system (PNS) type; we believe this is the first reported instance in which myelination of central axons by Schwann cells within the brain parenchyma has been produced by the implantation of PNS elements.     

Influence of aging on peripheral nerve function and regeneration

Aging deeply influences several morphologic and functional features of the peripheral nervous system (PNS). Morphologic studies have reported a loss of myelinated and unmyelinated nerve fibers in elderly subjects, and several abnormalities involving myelinated fibers, such as demyelination, remyelination and myelin balloon figures. The deterioration of myelin sheaths during aging may be due to a decrease in the expression of the major myelin proteins (P0, PMP22, MBP). Axonal atrophy, frequently seen in aged nerves, may be explained by a reduction in the expression and axonal transport of cytoskeletal proteins in the peripheral nerve. Aging also affects functional and electrophysiologic properties of the PNS, including a decline in nerve conduction velocity, muscle strength, sensory discrimination, autonomic responses, and endoneurial blood flow. The age-related decline in nerve regeneration after injury may be attributed to changes in neuronal, axonal, Schwann cell and macrophage responses. After injury, Wallerian degeneration is delayed in aged animals, with myelin remnants accumulated in the macrophages being larger than in young animals. The interaction between Schwann cells and regenerative axons takes longer, and the amount of trophic and tropic factors secreted by reactive Schwann cells and target organs are lower in older subjects than they are in younger subjects. The rate of axonal regeneration becomes slower and the density of regenerating axons decrease in aged animals. Aging also determines a reduction in terminal and collateral sprouting of regenerated fibers, further limiting the capabilities for target reinnervation and functional restitution. These age-related changes are not linearly progressive with age; the capabilities for axonal regeneration and reinnervation are maintained throughout life, but tend to be delayed and less effective with aging.     

α-Synuclein immunoreactivity in normal and neoplastic Schwann cells

Alpha-synuclein is known to play an important role in several neurodegenerative diseases. Moreover, it is expressed in central nervous system neuronal tumors, and another member of the synuclein family, gamma-synuclein, is overexpressed in breast and ovarian carcinomas. However, the expression of alpha-synuclein has not been reported hitherto in the peripheral nervous system (PNS). In the present study, we investigated normal PNS tissue and schwannomas in human postmortem and biopsy samples using both immunocytochemistry and immunoelectron microscopy with antibodies against alpha-, beta- and gamma-synuclein. In normal PNS tissue, Schwann cells, but not axons or myelin, were immunopositive for alpha-synuclein. In schwannomas, almost all of the tumor cells showed diffuse cytoplasmic staining for alpha-synuclein (30 cases). Ultrastructurally, alpha-synuclein immunoreactivity was found in the cytoplasm of normal and neoplastic Schwann cells, in association with the plasma membrane, ribosomes, rough endoplasmic reticulum, small vesicles, Golgi apparatus and the nuclear outer membrane. No beta- or gamma-synuclein immunoreactivity was found in those cells. These results indicate that in the PNS, alpha-synuclein is a useful marker of Schwann cells and that it is not involved in tumorigenesis.     

SENSORY GANGLIONOPATHY ASSOCIATED WITH SJÖGREN'S SYNDROME PRESENTING WITH DYSPHAGIA

In immune-mediated demyelination of the nervous system, glial cell apoptosis has been observed recently; however, the relevance of the phenomenon and the characterization of the involved molecules are still controversial. Cytokines are secreted by many cells, including inflammatory and glial cells, and appear to play a relevant role in the peripheral nervous system (PNS) immuno-mediated demyelination, being active in promoting the damage to Schwann cells, myelin, and axons. Even though the exact role of the different cytokines is at present uncertain, they have a sequential different expression in PNS immune-mediated demyelination and could induce apoptotic death of Schwann cells in the vicinity of the inflammatory reaction via the expression of CD95 (Apo1/Fas). This study has been designed to detect in rat primary Schwann cell tissue cultures whether the administration of IL-1B and IFN-y can induce cell death. Identification of apoptotic Schwann cell was performed by morphological, immunohistochemical, and electron-microscopy analysis. Our results show that Schwann cells stimulated by proinflammatory cytokines IL-1B and IFN-y show morphological evidence of nuclear chromatin condesation at the DAPI staining and are TUNEL positive. The same features of apoptotic cell death were observed by electron microscopy. These findings provide evidence to support the hypothesis that cytokines can directly damage Schwann cells in disorders of the PNS.     

Hypomyelinated mutant mice IV: Peripheral myelin injp

This study compares peripheral myelination in a specific subdivision of the sciatic nerve ofjp^msd and unaffected littermate mice. No significant differences are found in numbers of myelinated and unmyelinated axons, diameters of axons, thickness of myelin sheaths relative to axon diameter, extent of unmyelinated axon segregation by Schwann cell processes, or in the ultrastructure of myelin and Schwann cells. By contrast,jp^msd mutant mice show severe CNS hypomyelination. This evidence, that thejp^msd mutation affects only oligodendrocytes, distinguishes mutations at this locus from others producing CNS hypomyelination in which PNS myelin is also affected.     

Remyelination of the demyelinated CNS: the case for and against transplantation of central,peripheral and olfactory glia

Although originally developed as a research tool for studying glial-glial and glial-axonal interactions, the technique of transplanting glial cell into the central nervous system has more recently been employed as a potential means for repairing persistent demyelination in clinical disease. It has now been clearly established using various experimental models that oligodendrocyte lineage cells, Schwann cells and olfactory ensheathing cells can all produce new myelin sheaths around demyelinated or amyelinated axons following transplantation. However, this property alone does not necessarily mean that transplantation of these cells into demyelinated lesions in clinical disease will be successful. This article considers some of the properties that would be required of a transplanted myelinogenic cell and assesses the advantages and disadvantages of the currently available cell types.     

Signals for proinflammatory cytokine secretion by human Schwann cells

Wallerian degeneration is a post-traumatic process of the peripheral nervous system whereby damaged axons and their surrounding myelin sheaths are phagocytosed by infiltrating leukocytes. Our studies indicate that Schwann cells could initiate the process of Wallerian degeneration by releasing proinflammatory cytokines involved in leukocyte recruitment and differentiation including IL-1beta, MCP-1, IL-8 and IL-6. A comparison of the secretory pattern between nerve explants and cultured Schwann cells showed that each cytokine was differentially regulated by growth factor deprivation or axonal membrane fragments. Since Wallerian-like degeneration occurs in a wide variety of peripheral neuropathies, Schwann cell-mediated cytokine production may play an important role in many disease processes.     

Schwann cells and oligodendrocytes read distinct signals in establishing myelin sheath thickness

Schwann cells and oligodendrocytes produce myelin sheaths of widely varying sizes. How these cells determine the size of myelin sheath for a particular axon is incompletely understood. Axonal diameter has long been suspected to be a signal in this process. We have analyzed myelin sheath thickness in L5 lumbar root and spinal cord white matter of a series of mouse mutants with diminished axonal calibers resulting from a deficiency of neurofilaments (NFs). In the PNS, average axonal diameters were reduced by 20-37% in the NF mutants. Remarkably, the average myelin sheath thickness remained unchanged from control values, and regression analysis showed sheaths abnormally thick for a given size of axon. These data show that a genetically induced reduction in axonal caliber does not cause a reduction in myelin sheath thickness in PNS and indicate that Schwann cells read some intrinsic signal on axons that can be uncoupled from axonal diameter. Interestingly, myelin sheaths in the spinal cord of these animals were not abnormally thick, arguing that axonal diameter may contribute directly to the regulation of myelination in the CNS and that oligodendrocytes and Schwann cells use different cues to set myelin sheath thickness.     

In vitro myelination of regenerating adult rat retinal ganglion cell axons by Schwann cells.

Schwann cell cultures provide a highly favorable substrate for retinal ganglion cell (RGC) survival and axon growth in vitro (B?hr and Bunge, Exp Neurol 106:27, 1989; Hopkins and Bunge, Glia 4:46, 1991). In this report we have extended former studies to obtain axon regeneration, long-term survival, and myelination of adult rat RGC axons in co-cultures of retinal explants with purified Schwann cells. By using modified co-culture conditions, we observed myelination of regenerating adult RGC axons by Schwann cells after 3-4 weeks in vitro. Myelination was associated with a one-to-one Schwann cell-axon relationship, characteristic of the formation of peripheral myelin. Under culture conditions that supported myelination, long-term survival (more than 12 weeks) of a small population of RGCs was observed. These findings highlight the remarkable ability of Schwann cells to support long-term survival of adult rat RGCs in the absence of either central nervous system (CNS) target tissue or other peripheral nervous system (PNS) components. This tissue culture system may serve as a model for the systematic study of the molecular mechanisms which are involved in axon regeneration and myelination of adult CNS neurons.     

Crucial Role for the Myelin-associated Glycoprotein in the Maintenance of Axon-Myelin Integrity

It has recently been shown that mice deficient in the gene for myelin-associated glycoprotein develop normal myelin sheaths in the peripheral nervous system. Here we report that in mutant mice older than 8 months the maintenance of axon-myelin units is disturbed, resulting in both axon and myelin degeneration. Morphological features include those typically seen in human peripheral neuropathies, where demyelination-induced Schwann cell proliferation and remyelination lead to the formation of so-called onion bulbs. Expression of tenascin-C, a molecule indicative of peripheral nerve degeneration, was up-regulated by axon-deprived Schwann cells and regenerating axons were occasionally seen. Myelin-associated glycoprotein thus appears to play a crucial role in the long-term maintenance of the integrity of both myelin and axons.