Remodeling of the interstitial extracellular matrix in white matter multiple sclerosis lesions: Implications for remyelination (failure)

The extracellular matrix (ECM) provides protection, rigidity, and structure toward cells. It consists, among others, of a wide variety of glycoproteins and proteoglycans, which act together to produce a complex and dynamic environment, most relevant in transmembrane events. In the brain, the ECM occupies a notable proportion of its volume and maintains the homeostasis of central nervous system (CNS). In addition, remodeling of the ECM, that is transient changes in ECM proteins regulated by matrix metalloproteinases (MMPs), is an important process that modulates cell behavior upon injury, thereby facilitating recovery. Failure of ECM remodeling plays an important role in the pathogenesis of multiple sclerosis (MS), a neurodegenerative demyelinating disease of the CNS with an inflammatory response against protective myelin sheaths that surround axons. Remyelination of denuded axons improves the neuropathological conditions of MS, but this regeneration process fails over time, leading to chronic disease progression. In this review, we uncover abnormal ECM remodeling in MS lesions by discussing ECM remodeling in experimental demyelination models, that is when remyelination is successful, and compare alterations in ECM components to the ECM composition and MMP expression in the parenchyma of demyelinated MS lesions, that is when remyelination fails. Inter- and intralesional differences in ECM remodeling in the distinct white matter MS lesions are discussed in terms of consequences for oligodendrocyte behavior and remyelination (failure). Hence, the review will aid to understand how abnormal ECM remodeling contributes to remyelination failure in MS lesions and assists in developing therapeutic strategies to promote remyelination.     

Tenascin-R and C in multiple sclerosis lesions: relevance to extracellular matrix remodelling

In the present study the distribution of the inhibitory extracellular molecules tenascin-R (TN-R) and tenascin- C (TN-C) was examined by immunocytochemistry during evolution of the multiple sclerosis (MS) lesion, in which astrogliosis is a prominent feature. Sections were cut from five control cases and from 22 blocks containing lesions representing different pathological stages in 18 cases of secondary progressive MS. Widespread expression of TN-R was found in the normal human central nervous system (CNS), while that of TN-C was in general restricted to white matter. In acute MS plaques however, there was a similar striking loss of both TN-R and TN-C up to the edge of the lesion, where the macrophage density is greatest, extending into the apparently normal white matter. In subacute lesions a TN-C and/or TN-R-immunopositive reactive astrocyte subpopulation was prominent, reflecting synthesis of extracellular matrix molecules. Both tenascins were expressed throughout chronic MS plaques at levels similar to those seen in adjacent white matter. The loss of TN-R and TN-C in acute plaques is indicative of enzyme-mediated breakdown of the matrix which may be a marker of blood-brain barrier breakdown and leucocyte extravasation. Subsequent production of tenascins by reactive astrocytes may result in glial scar formation impeding remyelination and axonal repair in MS lesions.     

Adult rat oligodendrocytes grown in vitro upon an extracellular matrix have the ability to proliferate

The use of extracellular matrix (ECM) as a natural substrate for cell culture has markedly improved the growth and morphological differentiation of isolated adult rat oligodendrocytes. ECM-grown oligodendrocytes exhibited cyclic nucleotide phosphodiesterase (CNP)-activity which increased with time in culture and network formation. As much as 50-70% of the cells incorporated [3H]thymidine as visualized by the high labeling index of galactocerebroside (GalC)-positive cells. Chemical and enzymatic modifications of the ECM suggested that laminin in conjunction with other ECM constituents, plays a role in the induction of proliferation and/or differentiation responses in mature oligodendrocytes.     

Preparation of a monoclonal antibody to citrullinated epitopes: its characterization and some applications to immunohistochemistry in human brain

Using hybridoma technology, an IgM monoclonal antibody (mAb), designated as F95, was developed against a deca-citrullinated peptide (DCP) consisting of 10 citrulline residues and a carboxyl Gly-Gly-Cys through which DCP was covalently linked to an activated carrier protein, keyhole limpet hemocyanin (KLH). Clones were selected on the basis of not reacting with human unmodified and noncitrullinated myelin basic protein (MBP), MBP-C1, but reacting well with human citrullinated MBP (MBP-C8). When tested by ELISA, this mAb demonstrated minimal reactivity with human MBP-C1, varying reactivity with the C2-C5 isomers of human MBP, moderate binding with guinea pig MBP-C8, and strong reactivity with human MBP-C8. By ELISA, mAb F95 was directed predominantly against citrulline, not MBP, as revealed by its binding to DCP linked with activated KLH, bovine serum albumin (BSA), or ovalbumin (OA), but not with KLH, BSA, or OA alone. Immunohistochemistry of normal human brain demonstrated that F95 stained central nervous system myelin and a subset of astrocytes. Given the citrulline-directed features of mAb F95, this immunohistochemical pattern suggests that certain astroglial filaments expressing glial fibrillary acidic protein also contain citrulline-bearing components. These potentially implicate citrullinated proteins, notably in astroglial filaments, in a variety of normal and pathological neurobiological processes.     

Fetal rat septal cells adhere to and extend processes on basement membrane, laminin, and a synthetic peptide from the laminin A chain sequence

Responses of rat embryonic septal cells to reconstituted basement membrane, laminin, and laminin A chain-derived synthetic peptides were studied in culture. Dissociated fetal E16/17 septal cells were grown for three days on differently coated plastic substrata. Reconstituted basement membrane (Matrigel), laminin, and a 19-amino acid synthetic peptide CSRARKQAASIKVAVSADR-NH2 (PA22-2) from the laminin A chain sequence mediated cell-substratum adhesion and promoted neurite outgrowth. In contrast, cells did not attach to or form processes on uncoated plastic or on plastic substrata coated with synthetic, laminin-derived control peptides. Polyethylenimine (PEI) supported the adhesion and survival of fetal septal cells; however, when laminin was added to the medium during cell plating or 18 hr afterward, a dose-dependent increase was observed in neurite outgrowth of cells attached to this substratum. Cells grown for 6 days on PEI in the presence of laminin showed a determined increase in the number of cholinergic neurons as marked by acetylcholinesterase staining. These data suggest that the subpopulation of cholinergic septal neurons present in the septal cells studied here were also responding to laminin. The results of this in vitro study suggest potential uses for basement membrane, laminin, or synthetic peptides, such as PA22-2, in fetal septal grafts to enhance regeneration in the damaged septo-hippocampal system.     

Serum gelatinase B/MMP-9 in primary progressive multiple sclerosis patients treated with interferon-beta-1a

Background: Interferon- beta (IFN) acts by a variety of mechanisms in relapsing-remitting multiple sclerosis (MS). One of these is a cellular down-regulation of gelatinase B or matrix metalloproteinase-9 (MMP-9), which is known from biochemical, biological and immunohistochemical evidences to play a disease-promoting role in MS.Aims: a) To investigate the influence of IFN-1a (30 or 60 µg I. M./week) on serum MMP-9 levels in patients with primary progressive MS (PPMS). b) To correlate serum MMP-9 levels with clinical and magnetic resonance imaging (MRI) findings.Methods: Serial blood samples were collected every 3 months from 49 patients participating in a phase II trial of IFN-1a in PPMS. Serum MMP-9 was quantified by ELISA and correlations with clinical (EDSS) as well as MRI findings (brain and spinal cord atrophy, ventricular volume, T1 and T2 lesion load) were calculated.Results: No significant differences were found between serial serum MMP-9 levels in IFN-treated versus placebo-treated patients. MMP-9 levels did not differ between patients who progressed or did not progress during the study interval. Although mean absolute serum MMP-9 levels over the study period correlated with an increase in T2 lesion load (relative T2 change: r=0.51, p<0.001; absolute T2 change: r=0.30, p=0.038), absolute increase in brain ventricular volume (r=0.29, p=0.05) and increased brain atrophy (r=0.35, p=0.02), only the correlation with T2 lesion load was sustained throughout the study period. No correlations were found between MMP-9 levels and relative changes in ventricular volume or with relative/absolute changes in T1 lesion load and in spinal cord atrophy. None of the MRI measures correlated with MMP-9 changes between baseline levels and those on treatment.Conclusion: Although some evidence suggests a down-regulating effect of IFN on MMP-9, this was not confirmed for a once weekly intramuscular dose of IFN-1a in patients with PPMS. The sustained correlation between serum MMP-9 and changes in T2 volumes, and the lack of correlation with clinical or MRI measures of disease progression may suggest that MMP-9 is more directly related to non-specific central nervous system damage than to axonal loss.     

Individual isoforms of the amyloid βprecursor protein demonstrate differential adhesive potentials to constituents of the extracellular matrix

The amyloid β precursor protein (AβPP) can exist as a membrane-bound glycoprotein which modulates neural cell adhesion. The adhesion of clones of the AtT20 mouse pituitary cell line, transfected with cDNA coding for the 695 (AβPP₆₉₅) and 751 (AβPP₇₅₁) amino acid forms of the protein, to individual components of the extracellular matrix was determined using a centrifugal shear assay. On laminin, poly-L-lysine, fibronectin, and uncoated glass substrata, the cells transfected with AβPP₆₉₅ (6A1 cells) demonstrated a 50% increase in adhesivity over non-transfected cells, while those transfected with AβPP₇₅₁ (7A1 cells) showed a significant decrease in adhesion. There was, however, a significant increase in the adhesive strength of the 7A1 cells to collagen type IV with no change in the adhesivity of the 6A1 cells when compared with control. These changes in adhesivity could be attributed to changes in the levels of the membrane-bound protein and were not due to the interaction of soluble AβPP with elements of the extracellular matrix. These studies provide evidence for differential adhesivities of the constituent AβPP isoforms and the possible role of the Kunitz protease inhibitor (KPI) domain in influencing the adhesive properties of the protein backbone. J. Neurosci. Res. 49:154–160, 1997. © 1997 Wiley-Liss, Inc.     

Protective effect of a synapsin peptide genetically fused to the B subunit of Escherichia coli heat‐labile enterotoxin in rat autoimmune encephalomyelitis

Experimental autoimmune encephalomyelitis (EAE) is an inflammatory demyelinating disease with similarities to multiple sclerosis that requires the activation of auto reactive T cells that infiltrate the central nervous system. In previous studies we have shown that intraperitoneal administration of synaptosomal antigens could suppress EAE. Herein we examined the effect in this animal model of a fusion protein comprising the C domain of synapsin Ia and the B subunit of Escherichia coli heat‐labile enterotoxin (LTBSC). Oral administration to rats of low amounts of LTBSC induced immunological systemic tolerance to the encephalitogenic myelin basic protein. Treatment with LTBSC prior to EAE induction diminished disease incidence, DTH reaction to myelin basic protein, and central nervous system inflammation. LTBSC treatment also reduced the specific T‐cell proliferative response to myelin basic protein, decreased nitric oxide production, and augmented arginase activity by peritoneal macrophages. All animals challenged for EAE developed antibody response specific for myelin basic protein, but rats treated with LTBSC showed a lower IgG2b/IgG1 ratio, indicating a shift to a Th2‐type milieu. The data presented here suggest that well‐conserved synapsin peptides conjugated to the B subunit of enterotoxins from the cholera toxin family have a protective role and provide a potential therapeutic tool for intervention in EAE as well as in multiple sclerosis. © 2009 Wiley‐Liss, Inc.     

Microglial expression of alphavbeta3 and alphavbeta5 integrins is regulated by cytokines and the extracellular matrix: beta5 integrin null microglia show no defects in adhesion or MMP-9 expression on vitronectin

As the primary immune effector cells in the CNS, microglia play a central role in regulating inflammation. The extracellular matrix (ECM) protein vitronectin is a strong inducer of microglial activation, switching microglia from a resting into an activated potentially destructive phenotype. As the activating effect of vitronectin is mediated by alphav integrins, the aim of the current study was to evaluate the requirement of the alphavbeta5 integrin in mediating microglial adhesion and activation to vitronectin, by studying these events in beta5 integrin-null murine microglia. Surprisingly, beta5 integrin null microglia were not defective in adhesion to vitronectin. Further analysis showed that microglia express the alphavbeta3 integrin, in addition to alphavbeta5. Flow cytometry revealed that microglial alphav integrin expression is regulated by cytokines and ECM proteins. alphavbeta3 integrin expression was downregulated by IFN-gamma, TNF, LPS, and TGF-beta1. alphavbeta5 expression was also reduced by IFN-gamma, TNF, and LPS, but strongly increased by the antiactivating factors TGF-beta1 and laminin. Gel zymography revealed that beta5 integrin null microglia showed no deficiency in their expression of matrix metalloproteinase (MMP)-9 in response to vitronectin. Taken together, these data show that microglia express two different alphav integrins, alphavbeta3 and alphavbeta5, and that expression of these integrins is independently regulated by cytokines and ECM proteins. Furthermore, it reveals that the alphavbeta5 integrin is not essential for mediating microglial adhesion and MMP-9 expression in response to vitronectin.     

Once-weekly 22microg subcutaneous IFN-beta-1a in secondary progressive MS: a 3-year follow-up study on brain MRI measurements and serum MMP-9 levels

OBJECTIVE: To study the effect of weekly injected subcutaneous interferon (IFN)-beta-1a 22 microg on the extent of brain lesions on magnetic resonance imaging (MRI) and the level of serum matrix metalloproteinase (MMP)-9 in patients with secondary progressive multiple sclerosis (SPMS). SUBJECTS AND METHODS: All the 28 Finnish patients participating in the Nordic multicentre trial on the clinical efficacy of weekly IFN-beta-1a (Rebif) 22 microg in SPMS were studied neurologically and by volumetric MRI during a 3-year follow-up. The levels of MMP-9 in serum were measured over the 3-year study. RESULTS: There was no obvious effect on the number of contrast medium-enhancing lesions, the volume of T1 or T2 lesions or level of serum MMP-9, nor was any effect detected on the relapse rate and the Expanded Disability Status Scale (EDSS). Brain atrophy progression was not affected by the treatment. CONCLUSION: The lack of effect on MRI, clinical outcomes or the levels of MMP-9 indicates that subcutaneous administration of low-dose low-frequency IFN-beta-1a is insufficient in controlling either the inflammatory constitutes or the neurodegenerative changes of advanced SPMS.     

血清单核细胞趋化蛋白1、基质金属蛋白酶9及胱抑素C水平对脑梗死患者预后的评估价值

目的探究血清单核细胞趋化蛋白1(MCP-1)、基质金属蛋白酶9(MMP-9)及胱抑素C(Cys-C)水平对脑梗死患者预后的评估价值。方法回顾性分析2016年6月—2018年5月收治的173例脑梗死患者的临床资料,根据改良Rankin量表(mRS)评分结果将患者分为预后良好组(MRS≤2分,98例)和预后不良组(MRS2分,75例),比较两组患者的血清MCP-1、MMP-9、Cys-C水平及美国国立卫生研究院卒中量表(NIHSS)评分,分析MCP-1、MMP-9、Cys-C水平与患者NIHSS评分的相关性;绘制受试者工作特征(ROC)曲线,评估MCP-1、MMP-9及Cys-C对脑梗死患者预后的预测价值。结果预后不良组患者的血清MCP-1、MMP-9、Cys-C水平及NIHSS评分均高于预后良好组(P 0.05)。Pearson相关性分析结果显示,脑梗死患者的血清MCP-1、MMP-9、Cys-C水平与NIHSS评分均呈正相关(r=0.637、0.579、0.476,均P 0.05)。ROC曲线分析结果显示,MCP-1、MMP-9和Cys-C水平对脑梗死患者预后均有预测价值,其ROC曲线下面积(AUC)分别为0.846、0.764和0.688,对脑梗死患者预后的评估均具有统计学意义(P 0.001)。结论血清MCP-1、MMP-9和Cys-C水平均可对脑梗死患者的预后进行有效评估,可作为脑梗死患者预后的参考指标。[国际神经病学神经外科学杂志, 2021, 48(1):59-62]     

Intravenous synthetic peptide MBP8298 delayed disease progression in an HLA Class II-defined cohort of patients with progressive multiple sclerosis: results of a 24-month double-blind placebo-controlled clinical trial and 5 years of follow-up treatment

MBP8298 is a synthetic peptide with a sequence corresponding to amino acid residues 82–98 of human myelin basic protein (DENPVVHFFKNIVTPRT). It represents the immunodominant target for both B cells and T cells in multiple sclerosis (MS) patients with HLA haplotype DR2. Its administration in accordance with the principle of high dose tolerance results in long‐term suppression of anti‐myelin basic protein (MBP) autoantibody levels in the cerebrospinal fluid (CSF) of a large fraction of progressive MS patients. MBP8298 was evaluated in a 24‐month placebo‐controlled double‐blinded Phase II clinical trial in 32 patients with progressive MS. The objective was to assess the clinical efficacy of 500 mg of MBP8298 administered intravenously every 6 months, as measured by changes in Expanded Disability Status Scale (EDSS) scores. Contingency analysis for all patients at 24 months showed no significant difference between MBP8298 and placebo‐treatments (n = 32, P = 0.29). Contingency analysis in an HLA Class II defined subgroup showed a statistically significant benefit of MBP8298 treatment compared with placebo in patients with HLA haplotypes DR2 and/or DR4 (n = 20, P = 0.01). Long‐term follow‐up treatment and assessment of patients in this responder group showed a median time to progression of 78 months for MBP8298 treated patients compared with 18 months for placebo‐treatment (Kaplan–Meier analysis, P = 0.004; relative rate of progression = 0.23). Anti‐MBP autoantibody levels in the CSF of most MBP8298 treated patients were suppressed, but antibody suppression was not predictive of clinical benefit. Anti‐MBP autoantibodies that reappeared in the CSF of one patient at 36 months, whilst under treatment with MBP8298, were not reactive with the MBP8298 peptide in vitro. The identification of a responder subgroup (62.5% of the patients in this study) enables a more efficient design of a large confirmatory clinical trial of MBP8298. The probability that patients with other less common HLA‐DR haplotypes will respond to this treatment should not be ignored.     

Dynamic culture of a thermosensitive collagen hydrogel as an extracellular matrix improves the construction of tissue-engineered peripheral nerve

Tissue engineering technologies offer new treatment strategies for the repair of peripheral nerve injury, but cell loss between seeding and adhesion to the scaffold remains inevitable. A thermosensitive collagen hydrogel was used as an extracellular matrix in this study and combined with bone marrow mesenchymal stem cells to construct tissue-engineered peripheral nerve composites in vitro. Dynamic culture was performed at an oscillating frequency of 0.5 Hz and 35° swing angle above and below the horizontal plane. The results demonstrated that bone marrow mesenchymal stem cells formed membrane-like structures around the poly-L-lactic acid scaffolds and exhibited regular alignment on the composite surface. Collagen was used to fill in the pores, and seeded cells adhered onto the poly-L-lactic acid fibers. The DNA content of the bone marrow mesenchymal stem cells was higher in the composites constructed with a thermosensitive collagen hydrogel compared with that in collagen I scaffold controls. The cellular DNA content was also higher in the thermosensitive collagen hydrogel composites constructed with the thermosensitive collagen hydrogel in dynamic culture than that in static culture. These results indicate that tissue-engineered composites formed with thermosensitive collagen hydrogel in dynamic culture can maintain larger numbers of seeded cells by avoiding cell loss during the initial adhesion stage. Moreover, seeded cells were distributed throughout the material.     

SC1, a brain extracellular matrix glycoprotein related to SPARC and follistatin, is expressed by rat cerebellar astrocytes following injury and during development

In the nervous system, extracellular matrix components are believed to influence cell shape, proliferation and migration during development and following injury. SC1 is a secreted glycoprotein expressed during neural development and in the adult brain. The molecule shows partial sequence homology to the anti-adhesive extracellular matrix molecule SPARC/osteonectin and to follistatin. We have made a surgical lesion in the adult rat cerebellum and examined changes in SC1 expression at 1 to 14 days after injury. Dual in situ hybridization/immunohistochemistry demonstrated that SC1 mRNA was induced in astrocytes surrounding the wound, reaching maximal levels at 10 days post-lesion. Immunohistochemistry revealed changes in the deposition of SC1 protein in radial fibres of Bergmann glia. SC1 protein was also detected at the border of the lesion, suggesting an association with the glial scar. Double immunohistochemistry with the astrocytic marker GFAP demonstrated that astrocytes also express SC1 during postnatal development.     

Disruption of the retinal basal lamina during early embryonic development leads to a retraction of vitreal end feet,an increased number of ganglion cells,and aberrant axonal outgrowth

Bacterial collagenase was injected into the vitreous of the eye of chick and quail embryos. Immunocytochemical and ultrastructural studies revealed that the collagenase dissolved the retinal basal lamina of the injected eye. The basal lamina disruption was first detectable 1 hour after enzyme injection and was complete within 3 hours. With further development, the retinal basal lamina was not reestablished; newly developing neuroepithelium in the peripheral retina, however, generated an intact basal lamina. Western blot analysis showed that Clostridial collagenase degraded various collagens but spared noncollagenous proteins. Basal lamina disruption of embryonic day 3 to 6 retinae led to the retraction of the end feet of the neuroepithelial cells, caused an increase in the number of Islet-1⁺ cells (most likely ganglion cells), an increase in the thickness of the optic fiber layer, and aberrant growth of optic axons on their way toward the optic disc. None of these changes were observed when retinal basal laminae were disrupted at later stages of development. The present data demonstrate that the retinal basal lamina, by anchoring the neuroepithelial cells to the pial surface of the retina, has an important function in the development of the normal cytoarchitecture of this structure. It is proposed that the altered extracellular environment in the vitreal part of the retina, resulting in the retraction of the neuroepithelial end feet, is responsible for the increased number of Islet-1⁺ cells and the aberrant axonal navigation. J. Comp. Neurol. 397:89–104, 1998. © 1998 Wiley-Liss, Inc.     

Ex vivo and in vivo studies of CME-1, a novel polysaccharide purified from the mycelia of Cordyceps sinensis that inhibits human platelet activation by activating adenylate cyclase/cyclic AMP

Introduction

CME-1, a novel water-soluble polysaccharide, was purified from the mycelia of Cordyceps sinensis, and its chemical structure was characterized to contain mannose and galactose in a ratio of 4:6 (27.6 kDa). CME-1 was originally observed to exert a potent inhibitory effect on tumor migration and a cytoprotective effect against oxidative stress. Activation of platelets caused by arterial thrombosis is relevant to various cardiovascular diseases (CVDs). However, no data are available concerning the effects of CME-1 on platelet activation. Hence, the purpose of this study was to examine the ex vivo and in vivo antithrombotic effects of CME-1 and its possible mechanisms in platelet activation.

Methods

The aggregometry, immunoblotting, flow cytometric analysis and platelet functional analysis were used in this study.

Results

CME-1 (2.3-7.6 μM) exhibited highly potent activity in inhibiting human platelet aggregation when stimulated by collagen, thrombin, and arachidonic acid but not by U46619. CME-1 inhibited platelet activation accompanied by inhibiting Akt, mitogen-activated protein kinases (MAPKs), thromboxane B₂ (TxB₂) and hydroxyl radical (OH^●) formation. However, CME-1 interrupted neither FITC-triflavin nor FITC-collagen binding to platelets. CME-1 markedly increased cyclic AMP levels, but not cyclic GMP levels, and stimulated vasodilator-stimulated phosphoprotein (VASP) phosphorylation. SQ22536, an inhibitor of adenylate cyclase, but not ODQ, an inhibitor of guanylate cyclase, obviously reversed the CME-1-mediated effects on platelet aggregation and vasodilator-stimulated phosphoprotein (VASP), Akt, p38 MAPK phosphorylation, and TxB₂ formation. CME-1 substantially prolonged the closure time of whole blood and the occlusion time of platelet plug formation.

Conclusion

This study demonstrates for the first time that CME-1 exhibits highly potent antiplatelet activity that may initially activate adenylate cyclase/cyclic AMP and, subsequently, inhibit intracellular signals (such as Akt and MAPKs), ultimately inhibiting platelet activation. This novel role of CME-1 indicates that CME-1 exhibits high potential for application in treating and preventing CVDs.     

Myelin-, reactive glia-, and scar-derived CNS axon growth inhibitors: expression, receptor signaling, and correlation with axon regeneration

Axon regeneration is arrested in the injured central nervous system (CNS) by axon growth-inhibitory ligands expressed in oligodendrocytes/myelin, NG2-glia, and reactive astrocytes in the lesion and degenerating tracts, and by fibroblasts in scar tissue. Growth cone receptors (Rc) bind inhibitory ligands, activating a Rho-family GTPase intracellular signaling pathway that disrupts the actin cytoskeleton inducing growth cone collapse/repulsion. The known inhibitory ligands include the chondroitin sulfate proteoglycans (CSPG) Neurocan, Brevican, Phosphacan, Tenascin, and NG2, as either membrane-bound or secreted molecules; Ephrins expressed on astrocyte/fibroblast membranes; the myelin/oligodendrocyte-derived growth inhibitors Nogo, MAG, and OMgp; and membrane-bound semaphorins (Sema) produced by meningeal fibroblasts invading the scar. No definitive CSPG Rc have been identified, although intracellular signaling through the Rho family of G-proteins is probably common to all the inhibitory ligands. Ephrins bind to signalling Ephs. The ligand-binding Rc for all the myelin inhibitors is NgR and requires p75(NTR) for transmembrane signaling. The neuropilin (NP)/plexin (Plex) Rc complex binds Sema. Strategies for promoting axon growth after CNS injury are thwarted by the plethora of inhibitory ligands and the ligand promiscuity of some of their Rc. There is also paradoxical reciprocal expression of many of the inhibitory ligands/Rc in normal and damaged neurons, and NgR expression is restricted to a limited number of neuronal populations. All these factors, together with an incomplete understanding of the normal functions of many of these molecules in the intact CNS, presently confound interpretive acumen in regenerative studies.     

A role for the MAPK/ERK pathway in oligodendroglial differentiation in vitro: stage specific effects on cell branching

The mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) pathway is important for both long-term survival and timing of the progression of oligodendrocyte differentiation. Oligodendroglial cells treated with MEK inhibitor were distinguished by using stage specific markers: NG2 proteoglycan, A2B5, 2′3′nucleotide-cyclic 3′phosphodiesterase (CNPase) and myelin basic protein (MBP), and classified according to their morphology into different developmental stages. Treatment significantly increased the number of cells with more immature morphologies and decreased the number of mature cells. Furthermore, it increased the number of rounded cells that could not be classified into any of the oligodendroglial developmental stages. The strongest effects were usually observed shortly after treatment. Rounded cells were CNPase/MBP positive and they were not stained by anti-NG2 or A2B5, indicating that they were mature cells unable either to extend and/or to maintain their processes. These data showed an effect of the MAPK/ERK pathway on oligodendroglial branching, with possible consequences for the formation of the myelin sheath.