Central and peripheral nerve regeneration by transplantation of Schwann cells and transdifferentiated bone marrow stromal cells

In contrast to the peripheral nervous system (PNS), little structural and functional regeneration of the central nervous system (CNS) occurs spontaneously following injury in adult mammals. The inability of the CNS to regenerate is mainly attributed to its own inhibitorial environment such as glial scar formation and the myelin sheath of oligodendrocytes. Therefore, one of the strategies to promote axonal regeneration of the CNS is to experimentally modify the environment to be similar to that of the PNS. Schwann cells are the myelinating glial cells in the PNS, and are known to play a key role in Wallerian degeneration and subsequent regeneration. Central nervous system regeneration can be elicited by Schwann cell transplantation, which provides a suitable environment for regeneration. The underlying cellular mechanism of regeneration is based upon the cooperative interactions between axons and Schwann cells involving the production of neurotrophic factors and other related molecules. Furthermore, tight and gap junctional contact between the axon and Schwann cell also mediates the molecular interaction and linking. In this review, the role of the Schwann cell during the regeneration of the sciatic (representing the PNS) and optic (representing the CNS) nerves is explained. In addition, the possibility of optic nerve reconstruction by an artificial graft of Schwann cells is also described. Finally, the application of cells not of neuronal lineage, such as bone marrow stromal cells (MSCs), in nerve regeneration is proposed. Marrow stromal cells are known as multipotential stem cells that, under specific conditions, differentiate into several kinds of cells. The strategy to transdifferentiate MSCs into the cells with a Schwann cell phenotype and the induction of sciatic and optic nerve regeneration are described.     

In vivo proliferation,migration and phenotypic changes of Schwann cells in the presence of myelinated fibers

Following injury to a peripheral nerve, changes in the behavior of Schwann cells help to define the subsequent microenvironment for regeneration. Such changes, however, have almost exclusively been considered in the context of Wallerian degeneration distal to an injury, where loss of axonal contact or input is thought to be critical to the changes that occur. This supposition, however, may be incorrect in the proximal stumps where axons are still in contact with their cell bodies. In this work, we studied aspects of in vivo Schwann cell behavior after injury within the microenvironment of proximal stumps of transected rat sciatic nerves, where axons are preserved. In particular we studied this microenvironment proximal to the outgrowth zone, in an area containing intact myelinated fibers and a perineurial layer, by using double immunolabelling of Schwann cell markers and 5-bromo-2'-deoxyuridine (BrdU) labeling of proliferating cells.In normal sciatic nerve, Schwann cells were differentiated, in an orderly fashion, into those associated with unmyelinated fibers that labeled with glial fibrillary acidic protein (GFAP) and those associated with myelinated fibers that could be identified by individual axons and myelin sheaths. After sciatic nerve transection, there was rapid and early expansion in the population of GFAP-labeled cells in proximal stumps that was generated in part, by de novo expression of GFAP in Schwann cells of myelinated fibers. Schwann cells from this population also underwent proliferation, indicated by progressive rises in BrdU and GFAP double labeling. Finally, this Schwann cell pool also developed the property of migration, traveling to the distal outgrowth zone, but also with lateral penetration into the perineurium and epineurium, while in intimate contact with new axons.The findings suggest that other signals, in the injured proximal nerve stumps, beyond actual loss of axons, induce 'mature' Schwann cells of myelinated axons to dedifferentiate into those that up-regulated their GFAP expression, proliferate and migrate with axons.     

坐骨神经瓦勒变性大鼠许旺细胞生物学特性及分泌功能变化

背景：研究表明外周神经损伤后,许旺细胞在基底膜管内形成Bunger带,引导再生轴突延伸,但具体作用机制目前尚不清楚。 目的：观察大鼠坐骨神经损伤后瓦勒变性对许旺细胞生物学特性及分泌功能的影响。 方法：建立大鼠坐骨神经横切模型,分为坐骨神经瓦勒变性组(坐骨神经横断组)和手术对照组。采用神经段单酶消化法分离培养许旺细胞,光镜下观察细胞形态变化,S-100免疫荧光鉴定。取第1代许旺细胞,利用计数法绘制14 d内许旺细胞的生长曲线,MTT法检测14 d内许旺细胞增殖活性,酸性磷酸酶法检测许旺细胞黏附能力,ELISA法检测神经生长因子浓度。 结果与结论：坐骨神经段培养第14天,坐骨神经横断组神经段边缘可见大量许旺细胞,呈线形排列;手术对照组许旺细胞数量少,呈散在分布,两组许旺细胞S-100均呈阳性表达。许旺细胞传代培养第3天,两组许旺细胞均进入对数增长期,随时间延长,细胞数及细胞增殖吸光度值均呈上升趋势,坐骨神经横断组细胞数及增殖吸光度值明显高于手术对照组(P < 0.05);坐骨神经横断组许旺细胞黏附能力明显高于手术对照组(P < 0.05);ELISA法检测示,坐骨神经横断组神经生长因子浓度在培养第4,6,8,10,12,14天时均高于手术对照组(P < 0.05)。结果表明大鼠坐骨神经损伤后两三周,瓦勒变性对许旺细胞生物学功能具有显著影响,可诱导许旺细胞幼稚化,促使许旺细胞在短期内迅速分裂增殖,并分泌大量神经营养因子及细胞外黏附成分,为再生轴突的延伸提供适宜的神经微环境;并增加细胞黏附能力,为外周神经损伤修复提供适宜的神经微环境。 中国组织工程研究杂志出版内容重点：组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程全文链接：     

The effect of sciatic nerve transection on myelinated fibers in the L5 dorsal root and lumbar dorsal column

Summary The occurrence of Marchi-positive structures (MPS) in the L5 dorsal root and lumbar dorsal column was examined 1–18 weeks after unilateral sciatic nerve transection in rats, and compared to the occurrence of MPS during Wallerian degeneration seen after transection of L4 and L5 dorsal roots. There was an increasing number of MPS centrally to the junction between the peripheral (PNS) and central nervous system (CNS) and in the lumbar dorsal column ipsilateral to sciatic nerve transection throughout the examined time period. In the portion of the root distal to the PNS-CNS junction MPS were rare before 12 weeks postoperatively after which time small groups of MPS appeared. At all stages the incidence of MPS was just a fraction of that seen during Wallerian degeneration. From these observations it is inferred that few ganglion cells with myelinated central processes undergo complete disintegration after peripheral nerve transection. In addition, some of the myelinated central ganglion cell processes appear to be more severely affected proximal to the PNS-CNS junction than distally to it.     

Olfactory ensheathing cells: historical perspective and therapeutic potential

Olfactory ensheathing cells (OECs) are the glial cells that ensheath the axons of the first cranial nerve. They are attracting increasing attention from neuroscientists as potential therapeutic agents for use in the repair of spinal cord injury and as a source of myelinating glia for use in remyelinating axons in demyelinating diseases such as multiple sclerosis. This review mainly addresses the cell biological aspects of OECs pertinent to addressing two questions. Namely, where do OECs fit into the groupings of central nervous system (CNS)/peripheral nervous system (PNS) glial cells and should OECs be viewed as a clinically relevant alternative to Schwann cells in the treatment of spinal cord injury? The evidence indicates that OECs are indeed a clinically relevant alternative to Schwann cells. However, much more work needs to be done before we can even come close to answering the first question as to the lineage and functional relationship of OECs to the other types of CNS and PNS glial cells.     

Responses of macrophages in rat dorsal root ganglia following peripheral nerve injury

Summary Immunohistochemical studies with monoclonal antibodies to macrophage antigens were performed on sections of rat lumbar dorsal root ganglia. In confirmation of previous observations, cells with macrophage antigenicity were detected in normal ganglia. Many of these presumptive macrophages were perineuronal in contact with the neuron/satellite cell complex, a few were perivascular, and others were in interstitial position not in apparent contact with either blood vessels or neurons. The number of macrophages in lumbar dorsal root ganglia started to increase 2–4 days after sciatic nerve transection and remained elevated for four weeks. Perineuronal macrophages resembled satellite glial cells in light microscope appearance but were distinguished from glial cells by their lack of S-100 immunoreactivity. Following this sciatic nerve injury, macrophage counts were modestly increased in contralateral lumbar dorsal root ganglia but not in cervical dorsal root ganglia. Thus peripheral nerve injury induces a recruitment and/or proliferation of macrophages in the corresponding dorsal root ganglion. Although the functions of these macrophages are unclear, those in perineuronal position could contribute to the survival or regeneration of axotomized neurons.     

Ultrastructural study of anterograde transport of glial cell line-derived neurotrophic factor from dorsal root ganglion neurons of rats towards the nerve terminal

The glial cell line-derived neurotrophic factor (GDNF) is a potent neurotrophic substance in the central and peripheral nervous systems. The present immunohistochemical study clarified the ultrastructural localization of GDNF-immunoreactive substance (GDNF-IR) accumulated at transfected sciatic nerve stumps and also at normal spinal dorsal horn, and has demonstrated that GDNF-IR products appear to be located in dense-cored vesicles within the axons. Furthermore, to determine the source of proximally accumulated GDNF in the transected sciatic nerve, we attempted a transection and a double ligation maneuver involving the sciatic nerve. In the early period after the ligation (20 h), GDNF-IR fibers were observed in the proximal and distal segment of the ligations, but no immunoreactivities were detected in the middle segment. On the other hand, at a late period (8 days) after the transection, GDNF-IR fibers had almost disappeared, but weak GDNF-IR was observed in Schwann cells in the proximal and distal stumps of transected nerve. These findings suggest that most of GDNF-IR was transported from the proximal or distal side in the early period, but was locally synthesized by Schwann cells around the ligations in the late period. Spinal rhizotomy caused prominent accumulation of GDNF-IR products at the cut end of the ganglion side of the dorsal root, but not at the ventral root. These results suggested that dorsal root ganglionic (DRG) sensory neurons are one of the origins of GDNF. The fact that small- to medium-sized DRG neurons show enhanced GDNR-IR after the colchicine treatment may support the above suggestion. In conclusion, the present results strongly suggest that a subgroup of DRG sensory neurons synthesized GDNF-containing dense-cored vesicles in the neuronal somata and anterogradely transports the vesicles to peripheral or central axon terminals.     

Trophic activity derived from bone marrow mononuclear cells increases peripheral nerve regeneration by acting on both neuronal and glial cell populations

A rat model of complete sciatic nerve transection was used to evaluate the effect of bone marrow mononuclear cells (BMMC) transplanted to the injury site immediately after lesion. Rats treated with BMMC had both sensory and motor axons reaching the distal stump earlier compared to untreated animals. In addition, BMMC transplantation reduced cell death in dorsal root ganglia (DRG) compared to control animals. Transplanted BMMC remained in the lesion site for several days but there is no evidence of BMMC differentiation into Schwann cells. However, an increase in the number of Schwann cells, satellite cells and astrocytes was observed in the treated group. Moreover, neutralizing antibodies for nerve growth factor (NGF) (but not for brain-derived neurotrophic factor and ciliary-derived neurotrophic factor) added to the BMMC-conditioned medium reduced neurite growth of sensory and sympathetic neurons in vitro, suggesting that BMMC release NGF, improve regeneration of the sciatic nerve in the adult rat and stimulate Schwann and satellite cell proliferation or a combination of both.     

Delayed nerve repair increases number of caspase 3 stained Schwann cells

Caspase 3 staining in Schwann cells was investigated with immunohistochemistry, as a measure of Schwann cell apoptosis, after transection and immediate (day 0) or delayed rat sciatic nerve repair (30, 90 and 180 days post injury). Cleaved caspase 3 stained Schwann cells significantly increased at the site of lesion (SNL; median [IQR], 15.2 [7.0] %) and in the distal nerve segment (SND; 9.5 [3.6] %) 10 days after immediate repair. The number of cleaved caspase stained Schwann cells also increased significantly after delayed repair, irrespective of length of delay, at both locations (SNL: 22.0–27.1%; SND: 18.5–22.1%; p < 0.05). Some cleaved caspase 3 stained satellite cells were seen in dorsal root ganglia on the injured side, but no stained motor or sensory neurons were observed at any time-point. Delayed nerve repair is associated with more pronounced Schwann cell apoptosis which may explain impaired nerve regeneration after nerve injury and delayed repair.     

The induction of a regenerative propensity in sensory neurons following peripheral axonal injury

Summary Injury of the peripheral axons of primary sensory neurons has been previously shown to increase the probability that the corresponding central axons would grow from the injured spinal cord into a peripheral nerve graft. This phenomenon has been used to investigate the nature of extrinsic cues from injured nerves that trigger an enhanced regenerative propensity within sensory neurons. In 13 groups of rats, a segment of the right sciatic nerve was grafted to the dorsal columns of the spinal cord and the left sciatic nerve was subjected to mechanical injury, injection of colchicine or infusion of nerve growth factor. Subsequently, neurons in lumbar dorsal root ganglia with axons growing from the spinal cord into a graft were identified by retrograde perikaryal labelling and compared for the two sides. The aim was to mimic or modify the inductive effect of nerve transaction by alternative or additional manipulation of the nerve. Growth of central axons was less enhanced by peripheral axonal interruption if the length of the proximal stump was increased or if a distal stump was present to permit rapid regeneration. However, the regenerative response following nerve transection was altered little by crushing the proximal stump or injecting it with colchicine or nerve growth factor. It is suggested that sensory neurons are stimulated to regenerate by peripheral axonal injuries that reduce some normal retrograde regulatory influence of Schwann cells.     

Selective expression of L-serine synthetic enzyme 3PGDH in schwann cells, perineuronal glia, and endoneurial fibroblasts along rat sciatic nerves and its upregulation after crush injury

Non-essential amino acid L-serine functions as a highly potent, glia-derived neurotrophic factor, because it is a precursor for syntheses of proteins, other amino acids, membrane lipids, and nucleotides, and also because its biosynthetic enzyme 3-phosphoglycerate dehydrogenase (3PGDH) is preferentially expressed in particular glial cells within the brain. Here we pursued 3PGDH expression in peripheral nerves and its change after crush injury. In the pathway of rat sciatic nerves, 3PGDH was selectively expressed in non-neuronal elements: Schwann sheaths and endoneurial fibroblasts in sciatic nerves, satellite cells in dorsal root ganglia, and astrocytes and oligodendrocytes in the spinal ventral horn. In contrast, 3PGDH was immunonegative in axons, somata of spinal motoneurons and ganglion cells, and endoneurial macrophages. One week after crush injury, 3PGDH was upregulated in the distal segment of injured nerves, where 3PGDH was intensified in activated Schwann cells and fibroblasts. 3PGDH was still negative in activated macrophages, which were instead associated or surrounded by activated Schwann cells with intensified 3PGDH. These results suggest that in the peripheral nervous system, these non-neuronal cells synthesize and may supply L-serine to satisfy metabolic demands for maintenance and regeneration of peripheral nerves and for proliferation and activation of macrophages upon nerve injury.     

Transient modulation of Schwann cell antigens after peripheral nerve transection and subsequent regeneration

Summary Schwann cells within the distal portion of a transected nerve undergo a series of poorly understood events in response to injury and loss of axonal contact. These events may influence the regeneration of PNS neurons. In this study we examined the alteration of antigens located in the basal lamina, plasma membrane and cytoplasm of Schwann cells within the distal nerve stump: (a) after a complete transection of the sciatic nerve, and (b) subsequent to reestablished contact between regenerating axons and dedifferentated Schwann cells separated from contact with neurons. Visualization of laminin and heparan sulphate proteoglycan molecules at various intervals after nerve transection always revealed intact basal lamina channels. In response to loss of axonal contact, vimentin expression by Schwann cells within the distal nerve stump increased, becoming a predominant intermediate filament protein of the cytoskeleton while glial fibrillary acid protein (GFAP) expression decreased. This reversal in the prominence of intermediate filament proteins was maintained until the onset of axonal reinnervation, at which point expression of GFAP increased and vimentin decreased. Expression of the Schwann cell plasma membrane associated protein, C4, closely mimicked GFAP expression during axon degeneration and subsequent reinnervation. In the normal uninjured nerve, tissue plasminogen activator (tPA) and S-100 were localized in the region near the Schwann cell-axon interface and the outer Schwann cell plasma membrane. In response to loss of axonal contact, the S-100 and tPA immunoreactivity associated with the Schwann cell-axon interface was lost while that localized around the outer Schwann cell plasma membrane remained unchanged. The results of this study demonstrate that Schwann cells modulate a portion of their antigenic repertoire in response to a loss of axonal contact and after contact with regenerating axons.     

Peripheral nerve explants grafted into the vitreous body of the eye promote the regeneration of retinal ganglion cell axons severed in the optic nerve

Summary We have conducted experiments in the adult rat visual system to assess the relative importance of an absence of trophic factors versus the presence of putative growth inhibitory molecules for the failure of regeneration of CNS axons after injury. The experiments comprised three groups of animals in which all optic nerves were crushed intra-orbitally: an optic nerve crush group had a sham implant-operation on the eye; the other two groups had peripheral nerve tissue introduced into the vitreous body; in an acellular peripheral nerve group, a frozen/thawed teased sciatic nerve segment was grafted, and in a cellular peripheral nerve group, a predegenerate teased segment of sciatic nerve was implanted. The rats were left for 20 days and their optic nerves and retinae prepared for immunohistochemical examination of both the reaction to injury of axons and glia in the nerve and also the viability of Schwann cells in the grafts. Anterograde axon tracing with rhodamine-B provided unequivocal qualitative evidence of regeneration in each group, and retrograde HRP tracing gave a measure of the numbers of axons growing across the lesion by counting HRP filled retinal ganglion cells in retinal whole mounts after HRP injection into the optic nerve distal to the lesion. No fibres crossed the lesion in the optic nerve crush group and dense scar tissue was formed in the wound site. GAP-43-positive and rhodamine-B filled axons in the acellular peripheral nerve and cellular peripheral nerve groups traversed the lesion and grew distally. There were greater numbers of regenerating fibres in the cellular peripheral nerve compared to the acellular peripheral nerve group. In the former, 0.6–10% of the retinal ganglion cell population regenerated axons at least 3–4 mm into the distal segment. In both the acellular peripheral nerve and cellular peripheral nerve groups, no basal lamina was deposited in the wound. Thus, although astrocyte processes were stacked around the lesion edge, a glia limitans was not formed. These observations suggest that regenerating fibres may interfere with scarring. Viable Schwann cells were found in the vitreal grafts in the cellular peripheral nerve group only, supporting the proposition that Schwann cell derived trophic molecules secreted into the vitreous stimulated retinal ganglion cell axon growth in the severed optic nerve. The regenerative response of acellular peripheral nerve-transplanted animals was probably promoted by residual amounts of these molecules present in the transplants after freezing and thawing. In the optic nerves of all groups the astrocyte, microglia and macrophage reactions were similar. Moreover, oligodendrocytes and myelin debris were also uniformly distributed throughout all nerves. Our results suggest either that none of the above elements inhibit CNS regeneration after perineuronal neurotrophin delivery, or that the latter, in addition to mobilising and maintaining regeneration, also down regulates the expression of axonal growth cone-located receptors, which normally mediate growth arrest by engaging putative growth inhibitory molecules of the CNS neuropil.     

Quantification of the mononuclear phagocyte response to Wallerian degeneration of the optic nerve

Summary We investigated the numbers, origin and phenotype of mononuclear phagocytes (macrophages/microglia) responding to Wallerian degeneration of the mouse optic nerve in order to compare it with the response to Wallerian degeneration in the PNS, already described. We found macrophage/microglial numbers elevated nearly four fold in the distal segments of crushed optic nerves and their projection areas in the contralateral superior colliculus 1 week after unilateral optic nerve crush. This relative increase in mononuclear phagocyte numbers compared well with the four-to five-fold increases reported in the distal segments of transected saphenous or sciatic nerves. Moreover, maximum numbers are reached at 3, 5 and 7 days in the saphenous, sciatic and optic nerves respectively, suggesting that the very slow clearance of axonal debris and myelin in CNS undergoing Wallerian degeneration is not simply due to a slow or small mononuclear phagocyte response. The apparent delay in the response in the CNS occurs because the mononuclear phagocytes respond to the Wallerian degeneration of axons, which is slightly slower in the CNS than the PNS, rather than to events associated with the crush itself, such as the abolition of normal electrical activity in the distal segment. This was demonstrated by the protracted time course of the mononuclear phagocyte response in the distal segment following optic nerve crush in mice carrying theWld ^smutation which dramatically slows the rate at which the axons undergo Wallerian degeneration. By³H-Thymidine labelling or by blocking microglial proliferation by X-irradiation of the head prior to optic nerve crush, we showed that the majority of macrophages/microglia initiating the response to Wallerian degeneration were of local, CNS origin but these cells rapidly (from 3 days post crush) upregulate endocytic and phagocytic functional markers although they do not resemble rounded myelin-phagocytosing macrophages observed in degenerating peripheral nerves. We speculate that the poor clearance of myelin in CNS fibre tracts undergoing Wallerian degeneration compared to the PNS, in the face of a mononuclear phagocyte response which is similar in relative magnitude and time course, is because Schwann cells in degenerating peripheral nerves promptly modify their myelin sheaths such that they can be recognized and phagocytosed by macrophages, whilst in the CNS oligodendrocytes do not.     

Inverse relation between intensity of GFAP expression in the substantia gelatinosa and degree of chronic mechanical allodynia

Glial cells are known to have a large impact on neuropathic pain conditions. Within the spinal cord, microglia rapidly respond to peripheral nerve injury, resulting in central sensitization and ultimately in the onset of enhanced pain behaviour. Astroglia respond with a short delay and are thought to contribute to the early maintenance of neuropathic pain. Nevertheless, it is unknown whether the roles of these glial cell types can be influenced by the chronicity of the neuropathology. Here, the persistent responses of astroglia and microglia to peripheral nerve injury within central pain networks in the upper dorsal horn laminae were studied. At 12 weeks after complete sciatic nerve injury, upregulation of glial fibrillary acidic protein (GFAP), but not complement receptor-3, could be detected in laminae II and III. Moreover, it was found that neuropathic animals with a higher degree of mechanical allodynia had a lower intensity of GFAP expression in lamina II (substantia gelatinosa). From these data we conclude that the role of astroglial responses in mechanical allodynia after peripheral nerve injury may be less straightforward as previously thought. Although astroglia are known to play a pro-nociceptive role in early neuropathic pain states, this role may shift to anti-nociception in more chronic pain states.     

The CNS-PNS transitional zone of the rat. Morphometric studies at cranial and spinal levels.

The transitional zone is that length of rootlet containing both central and peripheral nervous tissue. The CNS-PNS interface may be defined as the basal lamina covering the intricately interwoven layer of astrocyte processes which forms the CNS surface and which is pierced by axons passing between the CNS and PNS. Study of transitional zone development defines morphologically the growth, relative movement and interaction of central and peripheral nervous tissues as they establish their mutually exclusive territories on either side of the CNS-PNS boundary, and helps to explain the wide variations in the form of the mature transitional zone. Nerve rootlets at first consist of bundles of bare axons. These become segregated by matrices of fine Schwann cell processes peripherally and of astrocyte processes centrally. The latter may prevent Schwann cell invasion of the CNS. Astrocyte processes branch profusely and come to form the principal central nervous tissue component of the transitional zone. Developmental changes in the transitional zone vary markedly between nerves, reflecting differences in its final morphology. Widespread relative movements and migration of CNS and PNS tissues take place during development, so that the central-peripheral interface changes shape and position, commonly oscillating along the proximodistal axis of the rootlet. For example, developing cervical ventral rootlets contain a transient central tissue projection, while that of lumbar ventral rootlets and to a lesser extent that of cervical dorsal rootlets alternately increase and decrease in length. In the developing cochlear nerve, a central tissue projection is present before birth, but regresses somewhat before a marked outgrowth of central nervous tissue along the nerve takes place, which reaches into the modiolus during the first week postnatum. During development, some astrocytic tissue may even break off and migrate distally into the root, giving rise to one or more glial islands within it. During the period immediately preceding birth, Schwann cells come to be present in very large numbers in that part of the rootlet immediately distal to the CNS-PNS interface, the proximal rootlet segment. Here they form prominent sleeves or clusters of closely packed cells which intertwine with and encapsulate one another on the rootlet surface. Such Schwann cell overcrowding in the proximal rootlet segment could result in part from distal overgrowth of the rapidly expanding CNS around axon bundles, which might strip the Schwann cells distally off the bundle segments so engulfed.(ABSTRACT TRUNCATED AT 400 WORDS)     

The interaction and adhesive mechanisms between axon and Schwann cell during central and peripheral nerve regeneration

It is well known that the injured mammalian PNS can successfully regenerate, while the CNS such as the optic nerve of adult mammals is incapable of regeneration. It is now generally accepted that the inability of CNS neurons to regenerate appears to be caused by the glial environment made up of astrocytes and oligodendrocytes. However, recent studies show that such CNS neurons have the intrinsic capacity to regenerate which is triggered by an experimental replacement of inhibitorial glial environment to peripheral nerve segment. Thus, the PNS environment is suitable not only for the regeneration of PNS itself, but also for the elicitation of CNS regeneration. Schwann cell is the major component of PNS, which plays a central role both in PNS and CNS regeneration by producing various kinds of functional substances. The contact of axons to Schwann cells based upon the structural and molecular linkages seems to be indispensable for stable and successful regeneration. In addition to cell adhesion molecules, Schwann cells utilize short focal tight junctions to provide morphological stabilization of the contact with the elongating axon, as well as small scale gap junctions to facilitate traffic of substances between them. Thus, nerve regeneration is not a simple phenomenon of axonal elongation on the part of the Schwann cell membrane, but is based on direct and dynamic communication between the axon and the neighboring Schwann cell, which may be partly associated with the mechanisms of neural regeneration.     

Expression of glial fibrillary acidic protein in the CNS and PNS of murine globoid cell leukodystrophy, the twitcher.

The expression of glial fibrillary acidic protein (GFAP) was found to be markedly enhanced immunohistochemically and biochemically both in the central (CNS) and peripheral (PNS) nervous systems of the twitcher mutant, an authentic murine model of human globoid cell leukodystrophy. The astrocytes in the CNS, the unmyelinated Schwann cells in the sciatic nerve, and the satellite cells in the trigeminal ganglion stained heavily with anti-GFAP antiserum. These changes in GFAP expression occurred shortly before the initiation of demyelination and coincided chronologically and topographically with infiltration of macrophages, suggesting that the same or closely related factors trigger the infiltration of macrophages and activate expression of GFAP. Cytoskeletal protein preparations showed increases in GFAP as well as in vimentin in the brainstem, spinal cord, and sciatic nerve. These results demonstrate that at least two types of peripheral glia (the unmyelinated Schwann cell and the satellite cell), in addition to the astrocyte, respond to some pathologic stimuli with an increased expression of GFAP. However, two other GFAP-positive structures, the Bergmann and radial glia, showed no significant changes in their immunostaining.     

The role of basic fibroblast growth factor in peripheral nerve regeneration

In the peripheral nervous system regeneration and gradual functional restoration occur following peripheral nerve injury. Growth of regenerating axons depends on the presence of diffusible neurotrophic factors, in addition to the substratum. Neurotrophic factors that are involved in peripheral nerve regeneration include nerve growth factor, brain-derived neurotrophic factor, ciliary neurotrophic factor, glial cell line-derived neurotrophic factor, and interleukin-6. Recent functional and expression studies of basic fibroblast growth factor and its receptors have emphasized a physiological role of these molecules in the peripheral nervous system. Basic fibroblast growth factor and its receptors are constitutively expressed in dorsal root ganglia and the peripheral nerve. These molecules display an upregulation in dorsal root ganglia and in the proximal and distal nerve stumps following peripheral nerve injury. In the ganglia these molecules show a mainly neuronal expression, whereas at the lesion site of the nerve, Schwann cells and invading macrophages represent the main cellular sources of basic fibroblast growth factor and the receptors 1–3. Exogenously applied basic fibroblast growth factor mediates rescue effects on injured sensory neurons and supports neurite outgrowth of transectioned nerves. Regarding the expression pattern and the effects after exogenous administration of basic fibroblast growth factor, this molecule seems to play a physiological role during nerve regeneration. Thus, basic fibroblast growth factor could be a promising candidate to contribute to the development of new therapeutic strategies for the treatment of peripheral nerve injuries.     

GAP-43 in the axons of mammalian CNS neurons regenerating into peripheral nerve grafts

Summary Although mature mammalian CNS neurons do not normally regenerate axons after injury, it is well established that they will regrow axons over long distances into peripheral nerve implants. We have autografted segments of sciatic nerve into the brains of adult albino rats and have used light and electron microscopic immunocytochemistry to examine the distribution of the growth associated protein GAP-43 in and around the graft in the first two weeks following implantation. GAP-43 was present, 3–14 days after grafting, in small non-myelinated axonal sprouts in the brain parenchyma around the proximal tip of the graft. At 11–14 days after implantation similar sprouts within the graft itself were GAP-43 immunoreactive. The sprouts were either naked or associated with other cell processes (chiefly of Schwann cells; to a lesser extent of astrocytes). We also show that small numbers of neuronal perikarya around the tip of the graft become GAP-43 immunoreactive 11–14 days after implantation. Thus mature mammalian CNS neurons regenerating axons into a PNS graft display a marked increase in their content of GAP-43. In addition, we report that small plaques of GAP-43 reaction product are sometimes present on the plasma membranes of Schwann cells or astrocytes adjacent to immunoreactive axons, and that narrow sheet-like or filopodial processes of astrocytes, Schwann cells and possibly other non-neuronal cell types, may contain small amounts of GAP-43.