Myelination of two axons by a single Schwann cell

Summary A Schwann cell can form only one internode of myelin around an axon. However, we observed the formation by a single Schwann cell of myelin around two axons of different diameters in the sural nerve of a 45-year-old man with mononeuritis multiplex. Schwann cell processes spiraled in the same direction around each axon, forming mesaxons. The findings in this case appear to be an undescribed type of aberrant myelination.     

Connatal polyneuropathy — A case with proliferated microfilaments in schwann cells

Summary A case of connatal polyneuropathy is described in a boy who died of pneumonia at the age of 2 years, and from whom sural nerve biopsies had been taken when he was 4 and 16 months old. Clinically, his disease was characterized by motor weakness and muscular flaccidity in the presence of normal intellectual development. The evolution of the connatal peripheral nerve lesion could be followed from the age of 4 months to death: The first biopsy evidenced the most serious pathologic changes. The findings were reminiscent of those encountered in a fetal nerve at 18 weeks of gestation. Furthermore, it showed numerous filamentous inclusions in Schwann cells. The second biopsy showed a sparsely myelinated nerve with bands of basement membrane apparently unrelated to cells arranged around the nerve's fibers. A few Schwann cells containing filamentous inclusions were still present. At autopsy, the findings were identical to those of the second biopsy. The possibility that this patient was transitionally exposed to a neurotoxic agent during pregnancy and that the biopsy findings represent a lesion that is still florid in the first and in a residual state in the second biopsy is considered.Dedicated to Prof. A. Bischoff, Berne, on his 60th birthday     

Cultured neonatal schwann cells contain and secrete neuregulins

Neuregulins have become the focus of intense research due to their putative roles in the etiology of certain cancers and to their evolving roles in PNS and CNS development. Evidence has been presented that neuregulins are present in neuronal cells where they act as mediators of neuron-glial signaling. Consistent with this view, we now report that there is a dramatic differential response of Schwann cells to the two major isoforms of neu differentiation factor (NDFα and NDFβ). NDFβ is a potent mitogen, whereas NDFα is nonmitogenic for Schwann cells. In addition, we report that Schwann cells contain endogenous NDF as well as the mRNA for both isoforms of NDF. Evidence is also presented that several isoforms of NDF are released from cultured Schwann cells. Our data suggest that in addition to functioning as a neuron-glial mediator, endogenous neuregulins may function in an autocrine/paracrine loop or in a juxtacrine- (cell-to-cell) mediated signal between individual Schwann cells or between Schwann cells and neurons. © 1996 Wiley-Liss, Inc.     

Cation-binding sites in trigeminal ganglia and maxillary nerve: unusual reactivity of perikarya, stem axons and satellite cells

We have used the cupric/ferrocyanide reaction to study cation-binding in trigeminal ganglia and maxillary nerve of adult rats. Unmyelinated axons did not react, whereas myelinated axons were stained at nodal, paranodal or cleft sites. At 'nodal' sites, metallic deposits were found in the axoplasm, along the axolemma, and at the extracellular interfaces of the paranodal myelin. At 'paranodal' sites, particles were concentrated in the paranodal axoplasm and in the intracellular spaces of the myelin loops. Most maxillary axons examined at successive sites had all nodal or all paranodal staining, but 13 of 51 had a mixture. In trigeminal ganglia there was no staining of perineurial sheath, endoneurial cells or mast cells. Satellite cells and their basal laminae were prominently stained, with those around small neurons more reactive than those of large neurons. Patches of neuronal membrane on cell bodies were stained, more often for small than large neurons. The axon hillock and proximal stem axon were not stained in some cases, but approximately half the neurons had staining of perikaryal cytoplasm at the axon hillock or a dense asymmetric band in the proximal stem axon. Strong intraaxonal staining was found at the junction between unmyelinated proximal and myelinated distal stem axon. In distal stem axons, staining was found at the first myelin segment and at each successively thicker myelin segment; staining was mostly weak and paranodal, with intensity proportional to myelin thickness. The T-junction between stem and main myelinated axon had nodal or paranodal patterns; unmyelinated T-junctions were not stained. The varied cation-binding patterns in trigeminal ganglia show unusual properties of satellite cells and important differences between stem and main axons. The results that the cell membrane of axon hillock and proximal stem regions of many sensory large and small neurons may have numerous sodium channels and could affect signal propagation.     

GFAP expression of human Schwann cells in tissue culture

We have studied the expression of the intermediate filament (IF) proteins, vimentin and glial fibrillary acidic protein (GFAP), in cultured human Schwann cells (SC) from patients with different neuropathies and normal control cases. SC cultures from sural nerve biopsies of 8 subjects with axonal neuropathies, 8 with demyelinating neuropathies and 3 normal controls were included in this study and processed with double immunofluorescence technique, using anti-vimentin and anti-GFAP antibodies, during the 2nd, 4th and 6th week of culture. Five cultures incubated with anti-GFAP antibodies were also processed for immunoelectron microscopy. Specificity tests of the used antibodies were performed. We have found that: (1) cultured human SC constantly express vimentin; (2) SC from normal controls are GFAP-negative in the first period of culture; (3) SC from pathologic nerves can contain GFAP-immunoreactive IF and the percentage of GFAP-positive SC is higher in axonal than in demyelinating neuropathies; (4) during the permanence in culture human SC from both normal and pathologic cases acquire the ability to synthesize GFAP. The obtained data suggest that the removal from axonal contact and the resulting loss of myelinating function induce a cytoskeletal cellular response in human SC characterized by the cytoplasmic accumulation of GFAP-immunoreactive IF.     

Glial fibrillary acidic protein and intermediate filaments in human glioma cells

Summary Cultured human glioma cells were studied by double indirect immunofluorescence technique using antisera against intermediate filaments and glial fibrillary acidic protein. With both antisera cytoplasmic fibrillar fluorescence was seen. Perinuclear bundles of intermediate-sized filaments, induced by vinblastine treatment, were strongly stained with both antisera. The degree of codistribution of the two types of antigenic determinants varied considerably from cell to cell. These results suggest that two types of filament-related antigenic determinants can be present in the same cell, and also that glial fibrillary acidic protein-related filaments may possess functional similarities to the intermediate filaments found in other cells. Glial fibrillary acidic protein remains as a useful and specific antigenic marker for the study of glial cells in vitro.     

The expression of nerve growth factor receptor on Schwann cells and the effect of these cells on the regeneration of axons in traumatically injured human spinal cord

To investigate the effects of Schwann cells and nerve growth factor receptor (NGFR) on the regeneration of axons, autopsy specimens of spinal cord from 21 patients with a survival time of 2 h to 54 years after spinal cord trauma were studied using immunohistochemistry and electron microscopy. Regenerating sprouts of axons could be observed as early as 4 days after trauma. At 4.5 months after trauma, many regenerating nests of axons appeared in the injured spinal cord. The regeneration nests contained directionally arranged axons and Schwann cells. Some axons were myelinated. In injured levels of the spinal cord, the Schwann cells exhibited an increased expression of NGFR within spinal roots. These results show that an active regeneration process occurs in traumatically injured human spinal cord. The NGFR expressed on Schwann cells could mediate NGF to support and induce the axon regeneration in the central nervous system. Received: 20 June 1995 / Revised, accepted: 18 September 1995     

肌营养不良蛋白聚糖与许旺细胞的关系

肌营养不良蛋白聚糖(Dystroglycan,DG)是一种非整合素家族的粘附分子,也是一种重要的细胞膜表面成分,在骨骼肌中,它与dystrophin,carcoglycan(α,β,γ和δ),sarcospan,syntrophins(α,β1 和β2)和α-dystrobrevin共同组成肌营养不良糖蛋白复合物(DGC)[1].DG以其两个亚基α-DG和β-DG组成的二聚体形式存在.最初在研究肌营养不良疾病时被发现,后被发现广泛表达于人体骨骼肌及许多组织中.     

Transplantation of olfactory ensheathing cells or Schwann cells restores rapid and secure conduction across the transected spinal cord

Olfactory ensheathing cells (OECs) or Schwann cells were transplanted into the transected dorsal columns of the rat spinal cord to induce axonal regeneration. Electrophysiological recordings were obtained in an isolated spinal cord preparation. Without transplantation of cells, no impulse conduction was observed across the transection site; but following cell transplantation, impulse conduction was observed for over a centimeter beyond the lesion. Cell labelling indicated that the regenerated axons were derived from the appropriate neuronal source, and that donor cells migrated into the denervated host tract. As reported in previous studies, the number of regenerated axons was limited. Conduction velocity measurements and morphology indicated that the regenerated axons were myelinated, but conducted faster and had larger axon areas than normal axons. These results indicate that the regenerated spinal cord axons induced by cell transplantation provide a quantitatively limited but rapidly conducting new pathway across the transection site.     

Proliferation of schwann cells in tellurium-induced demyelination in young rats

Summary This is a study of DNA synthesis of Schwann cells during the demyelination and the remyelination of peripheral nerves secondary to the intoxication of young rats with tellurium (Te). The³H-thymidine uptake of Schwann cells begins on day 4, reaches a zenith on day 7, and ends before day 20 on the Te diet despite continuation of the diet.The chronology of pathologic events is that myelin breakdown leading to segmental demyelination occurs first, followed within 24–48 h by the appearance of paralysis and by the beginning of DNA synthesis by the Schwann cells. A quantitative study on isolated nerve fiber preparations showed that more Schwann cells are produced than necessary to cope with the remyelination and that only one of four to six Schwann cells present in the demyelinated area at day 12 will participate in the remyelinating process.This study was supported by funds from NIH grant NS-12092 and from the Faculté de Médecine Paris-Sud     

Syngeneic grafting of adult rat DRG-derived schwann cells to the injured spinal cord

A subdural inflatable microballoon was used to induce closed traumatic contusion to adult rat spinal cord. This spinal cord injury model was associated with reproducible and graded neurological deficits and histopathological alterations. At various delays after injury, transplantations of syngeneic adult cultured dorsal root ganglion-derived Schwann cells were performed into the spinal cord lesion. The transplants were well integrated and reduced the microcystic posttraumatic cavitation as well as the gliosis. Schwann cells transplants were invaded by numerous regenerating neurites most of which, based upon their neurotransmitter contents, seem to originate from the dorsal root ganglion.     

Adipose-derived stem cells differentiate into a Schwann cell phenotype and promote neurite outgrowth in vitro

Experimentally, peripheral nerve repair can be enhanced by Schwann cell transplantation but the clinical application is limited by donor site morbidity and the inability to generate a sufficient number of cells quickly. We have investigated whether adult stem cells, isolated from adipose tissue, can be differentiated into functional Schwann cells. Rat visceral fat was enzymatically digested to yield rapidly proliferating fibroblast-like cells, a proportion of which expressed the mesenchymal stem cell marker, stro-1, and nestin, a neural progenitor protein. Cells treated with a mixture of glial growth factors (GGF-2, bFGF, PDGF and forskolin) adopted a spindle-like morphology similar to Schwann cells. Immunocytochemical staining and western blotting indicated that the treated cells expressed the glial markers, GFAP, S100 and p75, indicative of differentiation. When co-cultured with NG108-15 motor neuron-like cells, the differentiated stem cells enhanced the number of NG108-15 cells expressing neurites, the number of neurites per cell and the mean length of the longest neurite extended. Schwann cells evoked a similar response whilst undifferentiated stem cells had no effect. These results indicate adipose tissue contains a pool of regenerative stem cells which can be differentiated to a Schwann cell phenotype and may be of benefit for treatment of peripheral nerve injuries.     

微囊化兔许旺细胞移植脊髓损伤大鼠：碱性成纤维细胞生长因子表达和后肢运动功能的变化

背景：研究表明微囊化兔许旺细胞移植于大鼠损伤脊髓后,能减轻炎症反应,促进脊髓再生,但具体作用途径尚不完全清楚。 目的：观察微囊化兔许旺细胞移植于损伤大鼠脊髓后,碱性成纤维细胞生长因子的表达以及后肢运动功能的恢复。 方法：取家兔坐骨神经以双酶消化法制成许旺细胞悬液后,再用气体喷入法制成海藻酸钡-许旺细胞微囊。同法制备不包被许旺细胞的空囊。SD大鼠随机分为4组：细胞组、空囊组、微囊组建立脊髓半横断损伤模型,分别于损伤处植入明胶海绵吸附的10 μL许旺细胞悬液、10 μL空囊、10 μL海藻酸钡-许旺细胞微囊;正常组不做任何干预。采用BBB评分观察大鼠后肢运动功能恢复情况,制作脊髓标本切片通过苏木精-伊红染色和尼氏染色行病理学检查,免疫组织化学染色观察碱性成纤维细胞生长因子的表达变化。 结果与结论：造模后即刻大鼠右后肢出现瘫痪;材料移植后7,14,28 d,微囊组大鼠后肢运动功能明显恢复,BBB评分明显优于细胞组、空囊组(P < 0.05或P < 0.01)。碱性成纤维细胞生长因子阳性细胞主要见于神经元细胞的胞浆及胶质细胞的胞核内。材料移植后第1,3,7天,胶质细胞主要在脊髓损伤附近处表达,其中第3天表达量最大;第14天在神经元细胞的胞浆内可见碱性成纤维细胞生长因子表达,微囊组表达程度明显高于细胞组、空囊组;之后各组表达程度均明显下降。提示微囊化兔许旺细胞移植于损伤大鼠脊髓后,可以抑制异种移植后免疫排斥,减轻炎症反应,增加碱性成纤维细胞生长因子表达,促进后肢功能恢复和脊髓再生。 关键词：脊髓损伤;移植;碱性成纤维细胞生长因子;运动功能;微囊;许旺细胞;生物材料 doi:10.3969/j.issn.1673-8225.2010.08.011     

A case of giant axonal neuropathy showing focal aggregation and hypophosphorylation of intermediate filaments

We report here the clinicopathological features of a typical case of giant axonal neuropathy (GAN). Scanning electron microscopy of the hair of this case revealed an extraordinarily irregular cuticle. Focal accumulation of intermediate filaments in axons, Schwann cells, muscle fibers and skin fibroblasts were also found under an electron microscopy. When examined immunocytochemically, muscle fibers exhibited local disruption of the filamentous network in the subsarcolemmal space and in the central cytoplasm accompanied by focal accumulation of desmin. The intracellular network of vimentin was also disrupted, exhibiting global accumulation in some of the cultured skin fibroblasts. Decreased interneurofilament spacing was found in enlarged axons, suggesting the presence of hypophosphorylation of neurofilaments in this patient. These findings suggest general disorganization, abnormal distribution and possible defective phosphorylation of intermediate filaments in GAN.     

Induction of adhesion molecules on human schwann cells by proinflammatory cytokines, an immunofluorescence study

The presence of cytokines in the peripheral nerve was positively correlated to the induction and progression of inflammation during experimental allergic neuritis (EAN) and Guillain Barré syndrome (GBS). We investigated the induction of adhesion molecules such as L-selectin, E-selectin, ICAM-1, VCAM-1 and Mac-1 on Schwann cells by proinflammatory cytokines. Cultured human Schwann cells from normal adult, fetal and diabetic nerves were studied by immunofluorescence at basal condition and after stimulation with cytokines for 6, 24, 48 and 96 h. Incubation of human Schwann cells with TNF, IFNγ and IL-1β induces the expression of ICAM-1 starting at 6 h and reaching a peak at 24 h on more than 90% of cells. VCAM-1 expression was induced after 6 h of treatment with TNF and IL-1β on almost 100% of Schwann cells. Surprisingly, stimulation with TNF, IFNγ and IL-1β also induced the expression of L-selectin on fetal and diabetic Schwann cells, but not on normal adult cells. E-selectin, an adhesion molecule classically upregulated during inflammation, as well as Mac-1, a ligand for ICAM-1, were not expressed on human Schwann cells at basal condition or after treatment with cytokines. No ICAM-1, VCAM-1 and L-selectin expression was found on unstimulated Schwann cells. Our results suggest that upregulation of adhesion molecules on Schwann cells may have a role in the pathogenesis of inflammation in the peripheral nerve.     

Purification of mouse schwann cells using neurite-induced proliferation in serum-free monolayer culture

We have recently reported that neonatal mouse dorsal root ganglionic Schwann cells will (i) survive and assume characteristic morphologies in a serum-free, fully defined culture medium (N1 medium), (ii) proliferate extensively in the same N1 medium if neurons are also present and maintained by nerve growth factor, and (iii) display a strong proliferative response to serum even in the absence of neuronal elements, while also undergoing marked changes in their morphology and their associative behavior toward neurites. In this report, we present a detailed procedure, based upon these earlier observations, which yields purified cultures of either neurons plus associated Schwann cells or Schwann cells in the absence of neurons. The procedure utilizes the neuritic mitogen for selective expansion of Schwann cell numbers in serum-free primary cultures, and a secondary culture step involving neuronal removal and additional Schwann cell expansion using the serum mitogen. The procedure requires 9 days for the generation of3−4 × 10⁶ Schwann cells from 12 newborn mice (with a Schwann cell to neuron ratio of 10) and an additional 6–7 days for the generation of a neuron-free secondary population of40 × 10⁶ Schwann cells with less than 3% contamination by identifiable ganglionic fibroblasts.     

Schwann cells differentiated from spheroid-forming cells of rat subcutaneous fat tissue myelinate axons in the spinal cord injury

In the present study, we found that nestin-expressing spheroid cells derived from multipotent adipose stem cells of subcutaneous fat tissue could efficiently differentiate into Schwann cells (SCs) in vitro based on expression of SC markers such as A2B5, GFAP, O4, p75, S100, Sox10, Krox-20, and L1. The induced SC is engrafted to spinal cord injury lesions and formed a peripheral nervous system (PNS)-type myelin sheath on central nervous system (CNS) axons. PNS-type myelin sheath formation in repaired tissue was confirmed by transplantation of both induced PKH26-labeled SC and induced EGFP-expressing SC generated from EGFP transgenic rats. In addition to direct participation as myelin sheath-forming SC in repaired tissue, the induced SC also expressed several neurotrophic factors, as did native SC, which may suggest an additional role for induced SC in stimulation of endogenous healing responses. Thus, spheroid-forming cells from subcutaneous fat tissue demonstrated rapid and efficient induction into SC, and such cells show therapeutic promise for repair of damage to the CNS and PNS.     

Isolation of cDNA clones encoding rat glial fibrillary acidic protein: expression in astrocytes and in Schwann cells.

Glial fibrillary acidic protein (GFAP) expressed by astrocytes in the central nervous system (CNS) has been extensively characterized but the molecular identity of related molecules in the peripheral nervous system (PNS) remains unclear. To examine possible structural differences between CNS and PNS GFAP, we have isolated cDNA clones for rat GFAP from both cultured astrocyte and Schwann cell libraries. Nucleotide sequence analysis indicated that the PNS and CNS GFAP clones contained identical coding regions, with a predicted protein product of 430 amino acids. However, the 5'-untranslated region of clone rGFA15, isolated from the Schwann cell library, was longer than that predicted for brain-derived GFAP mRNA. Primer extension analysis of RNA isolated from the RT4-D6 Schwann cell line indicated that the start site for PNS GFAP mRNA lies 169 bases upstream from that used in the CNS. In addition, tryptic peptide mapping of GFAP prepared from cultured astrocytes and Schwann cells revealed one major peptide fragment present in CNS GFAP but absent from PNS GFAP. These results suggest structural differences between GFAP in these two cell types, at both the nucleic acid and protein level, and are consistent with previous observations of immunochemical differences existing between CNS and PNS GFAP.     

Frequency of accumulation of filaments in adaxonal cytoplasm of Schwann cells of myelinated fibers of rat spinal roots

Summary The frequency of accumulation of 6-nm filaments in the adaxonal cytoplasm of Schwann cells in the 6th lumbar dorsal and ventral roots was evaluated in 4-, 8-, 26- and 45-week-old Sprague-Dawley rats. The frequency was higher in 4- and 8-week-old (growing) rats than in 26- and 45-week old (mature) rats, and also higher in ventral than in dorsal roots in 4-, 8- and 26-week old rats. There were no clusters on certain groups of myelinated fibers according to the size of transverse axonal area, in both the ventral and dorsal roots. Therefore, this accumulation may reflect certain functions of the adaxonal cytoplasm of Schwann cell during natural growth and maturation of the axon and myelin sheath.