Neuregulin-1在外周神经再生中的作用

人类外周神经再生通常导致永久性的功能丧失。在外周神经再生过程中,雪旺细胞发挥重要的作用,它可以包绕神经轴突形成髓鞘,并分泌各种神经营养因子。在这个过程中,NRG-1起着重要的作用。NRG-1通过信号传导,使雪旺细胞去分化,增殖。而且NRG-1调节神经轴突的髓鞘化,并决定髓鞘的厚度。最后,在神经肌肉接头形成的过程中,NRG-1 也起着重要的作用。总之,NRG-1促进了外周神经损伤后的再生。     

rhNGF-B腺病毒的构建及转染多种细胞的效率

研究背景：神经组织工程发展，神经断段缺损的修复是外周神经损伤后修复的核心条件。然而生物神经生长因子(NGF)在外周神经断段缺损的修复治疗中发挥了至关重要因素。 目的：建立多机能化重组腺病毒hNGF-βcDNA和调节rhNGF-β基因转染促进外周神经再生的体外实验 材料：人胎儿脑 cDNA (Human fetal brain quick clone cDNA; Clontech, USA)和pAdenoVator-CMV5-IRES-GFP(Qbiogene, USA)用于重建rhNGF-β腺病毒。原代培养雪旺细胞，血旺细胞株(ISC, ATCC, CRH-2764))，非神经细胞胚胎肾细胞(HEK293 cells)，成肌细胞CRH(ATCC, CRL-1447), 成纤维细胞fibroblast NIH3T3 cell(ATCC, CRL-1658))同样被用于该实验。 方法和主要结果测量：准备rhNGF-β腺病毒，首先，在cDNA 克隆 hNGF-β cDNA，联合E1/E3，pAdenoVator-CMV5-IRES-GFP一同转入BJ5183细胞。所以，rhNGF-β腺病毒携带全长片断的hNGF-β cDNA结合到大肠杆菌中。同时，rhNGF-β腺病毒放大于肾HEK293细胞。RT-PCR, ELISA，免疫主治化学等用于测量转化效率，表达等。用于该实验的细胞株：HEK293细胞，原代培养雪旺细胞，雪旺细胞株，NIH3T3 细胞and CRH细胞。 结果：重组的携带hNGF-β腺病毒表达核酸内切酶和DNA序列，被rhNGF-β感染的腺病毒细胞GFP表达90%。mRNA表达带明显。ELISA，感染14天后的腺病毒雪旺细胞NGF的ELISA达到18865.4±310.9pg/ml。PC-12细胞在含有rhNGF-β腺病毒感染的雪旺细胞培养后表达的神经炎高信号。 结论：重组体腺病毒载体hNGF-β在非神经细胞和神经细胞的表达成功组建为外周神经再生提供了广阔的前景。     

周围神经损伤后修复的有关影响因素

周围神经损伤后修复的有关影响因素冯凯林世和赵节绪薛志刚周围神经损伤后，由于显微外科技术的应用，使得目前已能够十分精确地将中断的神经对合，但功能完全恢复者十分罕见。近几十年来，人们已从改进神经修复的纤维外科技术转向了探索神经再生过程中的细胞、分子和基因...     

神经损伤自发放电编码的神经再生研究进展

神经损伤与修复是一道长期摆在基础与临床科研工作面前的难题．每年因中枢、外周神经损伤导致的功能瘫痪．给患、患家庭和社会都带来了巨大的痛苦和负担。解决神经损伤修复的一个最基本条件就是搞清神经再生的理论问题。由于其再生机制不清导致目前所有已发现的神经修复方法虽都有不同程度的效果，但均不能从根本上解决完全修复问题。无疑，进一步研究神经再生机制是目前解决神经修复最迫切的问题。人们早已发现外周感觉神经在受到损伤性刺激后会表现出兴奋性增强、静息膜电位不稳定、出现自发放电现象．在中枢结构重塑活跃区域会有节律性放电反应。[第一段]     

许旺细胞在周围神经损伤修复中的作用及其可能分子机制**★

背景：周围神经损伤后神经功能的恢复一直是人们关注的焦点。许旺细胞在促神经功能恢复方面有着不可替代的作用。 目的：对国内外有关许旺细胞在促神经功能恢复方面的作用及其可能的分子机制作一综述。 方法：应用计算机检索CNKI、VIP和OVID数据库中1993-01/2010-01关于周围神经损伤的文章，在标题和摘要中以“许旺细胞，周围神经，神经再生，综述”或“Schwann Cells，Peripheral Nerve，Nerve Regeneration, Review”为检索词进行检索。选择文章内容与许旺细胞对周围神经损伤的作用有关者，同一领域文献则选择近期发表或发表在权威杂志上的文章。初检得到256篇文献，根据纳入标准选择关于许旺细胞作用机制的24篇文献进行综述。 结果与结论：许旺细胞通过多种途径作用于受损的周围神经，进而促进损伤神经功能的恢复。如何促使损伤局部许旺细胞更多的增殖将成为更好地促进损伤神经功能恢复的一大突破点。综合各方面因素，电针治疗周围神经损伤将有更好的前景。     

臂丛神经损伤后神经功能的重建：标准技术vs.新技术

臂丛神经受损可导致手臂、手失神经支配和难以忍受的神经性疼痛。最常用的臂丛神经损伤修复技术是自体感觉神经移植修复神经缺损,该技术对于神经功能的恢复的神经性疼痛的减轻作用并不可靠。我们评估了目前最好的臂丛神经修复技术和用填充了自体富血小板纤维蛋白的胶原管修复臂丛神经缺损,其能更好的促进神经功能恢复,减轻或使神经性疼痛消失;这些修复效果是传统神经移植无法达到的。     

神经导管生物材料修复周围神经损伤的动物实验

目的：通过动物体内神经导管生物材料移植实验，观察、测定再生神经功能恢复的程度和神经再生的数目。 方法：应用计算机检索中国期刊全文数据库1996/2010文献，检索词为“神经导管，神经损伤，导管材料，生物材料”。纳入有关神经导管材料在周围神经损伤修复中应用的动物实验。 结果：现有的任何材料都不能制备出理想的神经导管。在生物可降解前提下，往往天然材料具有更好的生物相容性，而合成高聚物可以通过调节组分的比例和相对分子质量及相对分子质量分布等条件，从而调节降解时间和材料的机械性能以及物理性能。外周神经修复理想的导管首先需要选择合适的生物材料和组装技术，制成具有良好物理特性(通透性、柔韧性、降解性等)的导管，尤其是对于较大的神经缺损通透性和柔韧性更为重要。 结论：在单腔中空导管中填充不同材料能促进神经的再生，联合应用几种不同的填充物质可能更加有利于神经修复。 关键词：神经导管；神经损伤；导管材料；修复；生物材料 doi:10.3969/j.issn.1673-8225.2010.29.030     

专题论坛：卒中转化医学

编者按卒中不仅仅损伤神经元,更重要的是对神经血管单元和与此相关的微环境造成破坏,损伤细胞功能和细胞之间的信号传递。以往相关的科研只注重单一的神经元保护研究,现在越来越重视神经血管单元功能的修复和重塑以恢复神经功能的研究。目前,实验性卒中干预方法较多,如应用药物小分子、基因、干细胞以及基于基因的干细胞治疗等,其主要目的是通过促进卒中后神经再生和血管新生以修复     

内皮祖细胞在缺血性脑血管疾病治疗中的研究进展

近年来,成人外周血中鉴定出一类具有前体内皮细胞特性的细胞,称之为内皮祖细胞(EPC)。对于这种细胞的研究表明其具有干细胞特性,参与了神经再生和再血管化,在脑血管疾病中发挥正性作用,能够帮助缺血性卒中后的神经功能恢复。目前临床治疗缺血性脑血管疾病手段单一且疗效不确切,内皮祖细胞的修复潜力越来越受到关注。本文讨论了目前关于内皮祖细胞特性的研究进展,以及它对于缺血性脑血管疾病的治疗作用及其临床应用前景。     

骨髓间充质干细胞移植及生长相关蛋白43表达在脑修复中的作用

背景：研究证实骨髓间充质干细胞移植能够促进脑功能恢复,并对大鼠脑皮质及海马结构损伤具有修复作用,可能与细胞的自身代偿以及神经生长递质的参与有关,也可能是由于神经应激性损伤刺激靶组织细胞分泌各种神经因子的表达有关。 目的：从细胞生物学的角度,观察大鼠脑缺血损伤后骨髓间充质干细胞移植对神经再生及脑的修复作用。 方法：参考改良Nagasawa法建立大脑中动脉闭塞再灌注模型后,实施骨髓间充质干细胞移植,并分别进行跑台运动训练和水迷宫康复训练,进行神经功能评分及学习记忆评分。采用TUNEL法检测脑皮质区及海马区凋亡神经元的表达以及免疫组化技术检测生长相关蛋白43蛋白在两区的表达变化。 结果与结论：移植组16 h移植骨髓间充质干细胞在皮质区及海马CA1区表达明显增加;7 d细胞表达达高峰,分化细胞明显增加。移植后运动训练7,19,21 d移植组mNSS评分低于模型组(P均< 0.01);移植组大鼠水迷宫试验平台潜伏期的时间较模型组明显缩短(P < 0.05);移植组大鼠穿越平台次数较模型组增多(P < 0.05);缺血再灌注24 h凋亡细胞达高峰,3 d梗死体积测量为最大值;再灌注19 d生长相关蛋白43达高峰。提示大鼠脑缺血损伤介导了神经功能缺损,骨髓间充质干细胞移植促进了神经再生,生长相关蛋白43表达上调抑制神经元凋亡,进一步促进了脑梗死灶的修复。     

Numbers of regenerating axons in parent and tributary peripheral nerves in the rat

This study is concerned with numerical parameters of axonal regeneration in peripheral nerves. Our first finding is that the number of axons that regenerate into the distal stump of a somatic nerve at a particular time after transection is partially dependent on the type of lesion used to interrupt the axons. The second question concerns the proportion of axons that regenerate into the distal stump of a parent nerve compared to the proportions that regenerate into tributary nerves that arise from the parent. The proportions of regenerated myelinated axons in the nerve to the medial gastrocnemius muscle and myelinated and unmyelinated axons in the sural nerve are the same as the proportions of myelinated and unmyelinated axons that regenerate into the distal stump of the sciatic nerve for the crush, 0 and 4 mm gap transections. Proportionally fewer axons regenerate into the tributary nerves following the 8 mm gap transection, however. This implies that the length of the gap has an influence on whether or not axons in tributary nerves regenerate in concert with axons in the distal stump of the parent nerve. The unmyelinated fibers in the nerve to the medial gastrocnemius muscle are different because they do not regenerate in proportion to those in the distal stump of the sciatic nerve. We also provide evidence to indicate that myelinated axons branch whereas unmyelinated fibers end blindly when they enter the distal stump after crossing a sciatic nerve transection. Finally the normal arrangement of perineurial cells seems to be disrupted after the sciatic nerve regenerates across a gap.     

Alterations to neuronal polarity following permanent axotomy: a quantitative analysis of changes to MAP2a/b and GAP-43 distributions in axotomized motoneurons in the adult cat

Following axotomy, morphologically unusual, distal processes (UDPs) emerge from motoneuron dendrites. These processes contain an axonal protein, growth‐associated protein 43 (GAP‐43) but lack immunostaining for the dendritic protein microtubule‐associated protein 2a/b (MAP2a/b). Thus, it appears that neuronal polarity alters following axotomy. Our goal was to describe this change in neuronal polarity on a more detailed and quantitative level. We asked two questions: Following axotomy, where in the entire neuron does the immunoreactivity for MAP2a/b and GAP‐43 change and do these changes reflect a transformation of dendrite to axon or growth from terminal dendrites? Using intracellular labeling and immunocytochemistry, changes in MAP2a/b and GAP‐43 immunoreactivity were also found in processes with a morphology typical of terminal branches of intact motoneurons (called simple distal processes [SDPs]), as well as UDPs. Trajectories (the path from the soma to a single terminus) with UDPs and SDPs were longer than trajectories without these processes, and trajectories with UDPs were the longest. Trajectories without UDPs or SDPs were similar in length to trajectories from intact motoneurons. The distance from the soma to the point where MAP2a/b immunoreactivity became absent in trajectories with UDPs or SDPs was similar to the length of trajectories from intact motoneurons. Thus, following axotomy, two morphologically distinct types of axon‐like processes emerge from dendrites. The formation of these processes does not involve a transformation of the original dendrite, but rather growth at the ends of dendrites. J. Comp. Neurol. 450:318–333, 2002. © 2002 Wiley‐Liss, Inc.     

Mitogen activated protein (MAP) kinase activity is increased in adult mouse superior cervical ganglia during culturing

We have used adult mouse superior cervical ganglia (SCG) to study the role of mitogen activated protein (MAP) kinase activity during axonal outgrowth in vitro. An initial peak in activity within the first hours of culture was followed by a substantially higher activity after 1 to 2 days, a time when axons were actively growing. The latter peak is probably a result of both higher levels of protein and increased activity. The addition of nerve growth factor stimulated both outgrowth and kinase activity, whereas treating the cultures with the kinase inhibitor PD98059 had an opposite effect. Taken together, the results suggest that activation of the MAP kinase pathway could be involved in the initiation as well as regulation of axonal outgrowth from adult SCG. J. Neurosci. Res. 52:453–457, 1998. © 1998 Wiley-Liss, Inc.     

Differential gene expression in motor and sensory Schwann cells in the rat femoral nerve

Phenotypic differences in Schwann cells (SCs) may help to guide axonal regeneration down motor or sensory specific pathways following peripheral nerve injury. The goal of this study was to identify phenotypic markers for SCs harvested from the cutaneous (sensory) and quadriceps (motor) branches of the rat femoral nerve and to study the effects of expansion culture on the expression patterns of these motor or sensory phenotypic markers. RNA was extracted from SCs harvested from the motor and sensory branches of the rat femoral nerve and analyzed using Affymetrix Gene Chips (Rat Genome 230 v2.0 Array A). Genes that were upregulated in motor SCs compared with the sensory SCs or vice versa were identified, and the results were verified for a subset of genes using quantitative real-time polymerase chain reaction (qRT-PCR). The expression levels of the "phenotype-specific" genes were then evaluated in SC expansion cultures at various time points over 30 days by qRT-PCR to determine the effect of expansion on SC phenotype. Expression levels of the phenotype-specific genes were significantly altered after expansion culture for both the motor and the sensory markers compared with fresh nerve tissue. These results indicate that both motor and sensory SC gene expression patterns are disrupted during expansion in vitro and may affect the ability of SCs to express phenotype-specific genes after transplantation.     

Therapeutic strategies in promoting peripheral nerve regeneration

Currently, regeneration chambers, adrenocorticotropic hormone (ACTH) and related peptides, and gangliosides appear to be the most promising therapies in the promotion of peripheral nerve regeneration, growth, and repair. Regeneration chambers enhance rat sciatic nerve regeneration in vivo after transection by providing a structurally organized and protected preformed space within which nerve fibers are exposed to macromolecular compounds which direct and enhance nerve growth. ACTH and related peptides, independent of their corticotropic activities, increase the availability of structural proteins to the axon terminal in rats subjected to nerve crush injuries and demonstrate inotropic effects in adrenalectomized and/or hypophysectomized rats. Exogenously administered gangliosides promote neuronal sprouting, regeneration, and reinnervation in experimental situations and have undergone clinical testing in acute and chronic peripheral nerve disorders. At the current dosage levels and schedules, the clinical results of ganglioside therapy have been mixed. The success of the experimental studies supports further clinical testing of these therapies in peripheral nerve disorders.     

Omental graft improves functional recovery of transected peripheral nerve

The omentum has several properties that are advantageous for neuronal sprouting and direction. We have therefore analyzed functional recovery following transection of rat sciatic nerve using omental graft to bridge the nerve defect. In group 1, a 25-30-mm nerve defect was produced and bridged with omental graft, whereas in group 2, an end-to-end repair was performed. The sciatic function index (SFI) was assessed at 2-week intervals until 8 weeks after surgery. Functional recovery was faster in group 1 than in group 2. After 8 weeks, SFI was improved significantly from -100% to -45% (+/- -4%) in group 1 (P < 0.001) compared to -72% +/- -2% in group 2 (n = 10). Histologically, the omental graft contained more newly developed nerve fibers and less scar tissue than the end-to-end repair. Thus, omental graft appears to improve directional growth of regenerating axon sprouts and may be a means of treating peripheral nerve injury.     

An in vivo model to quantify motor and sensory peripheral nerve regeneration using bioresorbable nerve guide tubes

An in vivo preparation is presented to study the rate and time course of motor and sensory axonal regeneration. The cut ends of a transected sciatic nerve were inserted into each end of a 5-6 mm non-toxic and bioresorbable nerve guide tube to create a 4 mm nerve gap in adult mice. Subsequently, cell bodies in the ventral spinal cord and L3-L5 dorsal root ganglia that had regenerated axons across the gap were retrogradely labeled with horseradish peroxidase (HRP). The HRP was applied 3 mm distal to the nerve guide and was accessible only to axons that had regenerated through the nerve guide. Labeled cells were counted in 40 micron serial sections at 2, 4 and 6 weeks after initial nerve transection. The results indicate a significant increase in the number of labeled motor and sensory cell bodies over time. By 6 weeks after transection, approximately two thirds as many ventral horn motor cells and one third as many dorsal root ganglion sensory cells were labeled as in control non-transected animals. These data serve as a baseline to compare differential effects of additives to the nerve guide lumen in terms of sensory and motor neuron response.     

Effects of 660‐nm gallium–aluminum–arsenide low‐energy laser on nerve regeneration after acellular nerve allograft in rats

Purpose : The purpose of this study was to explore and discuss the effects of 660‐nm gallium–aluminum–arsenide low‐energy laser (GaAlAs LEL) irradiation on neural regeneration after acellular nerve allograft repair of the sciatic nerve gap in rats. Methods : Eight male and female Sprague–Dawley rats were used as nerve donors, and 32 healthy Wistar rats were randomly divided into four groups: normal control group, acellular rat sciatic nerve (ARSN) group, laser group, and autograft group. Twelve weeks after surgery, nerve conduction velocity, restoration rate of tibialis anterior wet muscle weight, myelinated nerve number, and calcitonin gene‐related peptide (CGRP) protein and mRNA expression of the spinal cord and muscle at the injury site were quantified and statistically analyzed. Results : Compared with the ARSN group, laser therapy significantly increased nerve conduction velocity, restoration rate of tibialis anterior wet muscle weight, myelinated nerve number, and CGRP protein and mRNA expression of the L₄ spinal cord at the injury site. Conclusions : These findings demonstrate that 660‐nm GaAlAs LEL therapy upregulates CGRP protein and mRNA expression of the L₄ spinal cord at the injury site and increases the rate of regeneration and target reinnervation after acellular nerve allograft repair of the sciatic nerve gap in rats. Low‐energy laser irradiation may be a useful, noninvasive adjunct for promoting nerve regeneration in surgically induced defects repaired with ARSN. Synapse 64:152–160, 2010. © 2009 Wiley‐Liss, Inc.     

Structure and function of fetal cortex implanted into degenerating peripheral nerve of adult rat

The growth, differentiation and function of E11 fetal cortex was studied over 4 months after implantation into the degenerating nerve to the biceps femoris muscle of 70 rats. At 7 days postimplantation (DPI) the implant formed a neuroepithelium which contained mainly undifferentiated cells within a reconstituted perineurium. At 14-21 DPI the neurons contained differentiated, mature nuclei, poorly developed Nissl bodies and normal cytoplasmic organelles. Neuroglia were also observed with typical nuclei and a dense cytoplasmic ground. At 30 days the implants reached their greatest degree of differentiation. Neurons had mature nuclei and cytoplasmic organelles. There were few axosomatic synapses or neuronal cell processes. Neuroglia were prominent with many cell processes. The neuropil contained myelinated and unmyelinated nerve fibers, dendrites and many axodendritic synapses. At 40-50 DPI the neurons and neuronal cell processes were degenerating. At 60-120 DPI the implant contained only neuroglia. Stimulation of only the implant resulted in biceps femoris muscle contraction in 5 animals at 30 DPI and one animal at 40, 45 and 50 days. Injection of 10 microliter of WGA-HRP into the biceps femoris muscle resulted in at least 20 neurons in 3 30-day implants that contained HRP reaction product. These data show that fetal cortical neurons can assume functions different from normal, functionally reinnervate adult muscle and can differentiate and form an ectopic nervous system for at least 30-days after implantation into a degenerating adult host peripheral nerve.     

Simultaneous gradual lengthening of both proximal and distal nerve stumps for repair of peripheral nerve defect in rats

We investigated nerve regeneration following the repair of a segmental nerve defect induced by direct end-to-end neurorrhaphy after simultaneous gradual lengthening of both proximal and distal nerve stumps in rats. A 15-mm-long nerve segment was resected from the sciatic nerve of each rat. The proximal and distal nerve stumps, respectively, were directly lengthened at a rate of 1 mm/day using a custom-made external nerve-lengthening device. After being lengthened for 14 days, both nerve stumps were refreshed, and direct end-to-end neurorrhaphy was performed. For a control, 15-mm nerve grafting was performed immediately after nerve resection. Nerve regeneration was evaluated by motor nerve conduction velocity, muscle contraction force, and histological studies at 6, 8, and 14 weeks after initial nerve resection in both groups. As a result, at 8 and 14 weeks, the motor nerve conduction velocity was significantly higher in the nerve-lengthening group than in the autografting group. In addition, at 14 weeks, the tetanic force and wet weight of the gastrocnemius muscle were significantly higher in the nerve-lengthening group than in the autografting group. Histologically, the mean axonal diameter of myelinated nerve fibers and the total number of myelinated nerve fibers were also significantly higher in the nerve-lengthening group than in the autografting group for each evaluation period. It appears that the simultaneous gradual lengthening of both proximal and distal nerve stumps might have potential application in the repair of peripheral nerve defects.