Motor function recovery during peripheral nerve multiple regeneration

Neuronal functional compensation and multiple regenerating axon sprouting occur during peripheral nerve regeneration. Sprouting nerve buds were quantitatively maintained and had matured when multiple injured distal nerves were anastomosed to smaller number of proximal nerve stumps; this has positive clinical significance for proximal stump damage. This study investigated whether sprouting axon buds would reinnervate the distal neuromuscular junction and maintain the function of the target organ under compensation conditions. The results showed that the sprouting axon buds maintained the numbers and morphology of motor end plates repaired by a smaller number of proximal nerve stumps, and recovered 80.0% tetanic muscle force compared with the normal side. Meanwhile, nerve conduction velocity, compound muscle action potential and diameter of muscular fibres declined 72.7%, 73.2% and 61.8%, respectively, compared with normal. This observation indicates the potential functional reserve of neurons and that it is feasible to repair nerve fibre injury through anastomosis of multiple distal nerve stumps with a smaller number of proximal nerve stumps, within the limits of compensation. Copyright © 2013 John Wiley & Sons, Ltd.     

一种新的生物材料-海藻酸盐修复长距离周围神经缺损的实验研究

目的探讨一种新的神经再生引导材料--可生物降解并吸收的海藻酸盐对兔坐骨神经缺损修复的作用.方法切除兔一侧坐骨神经的一部分,形成一35mm缺损,用两片海藻酸盐制成的海棉状材料,不经缝合地植入缺损并桥接两侧断端,术后行电生理、解剖学、组织学检查.结果术后6周起右下肢体出现电生理上的恢复,术后16周解剖学检查见原缺损处再生组织连接两断端,组织学检查证实再生组织内有大量有髓及无髓的轴突组织,未发现有海藻酸盐的残留.(而在未使用海藻酸盐修复的对照组动物,各项检查均为阴性.)结论海藻酸盐能促进长距离周围神经缺损的再生,同时也表明不缝合修复神经缺损的可能性,这为临床应用提供了新的材料和方法.     

大鼠胫神经损伤后远侧端髓鞘和轴索再生的研究

目的:探讨大鼠胫神经损伤时远侧端髓鞘和轴突的再生速度。方法:大鼠54只,建立神经原位桥接动物模型,其中36只大鼠用于荧光示踪注射,根据缝合口神经远端荧光示踪剂注射点位置不同,分为10 mm组和30 mm组各18只,分别于术后4、8、12周三个时间点进行处理,每组每个时间点6只;其他18只大鼠用于髓鞘染色,分别于术后4、8、12周三个时间点进行处理。结果:术后8、12周,10 mm和30 mm组的L3~L6脊神经节内有荧光标记细胞,30 mm组明显少于10 mm组,有显著性差异(P0.01);术后8、12周,患侧的损伤神经远侧端有大量排列有序的有髓神经纤维,10 mm组与30 mm组间差异无统计学意义(P0.05)。10 mm和30 mm组术后8、12周的胫神经运动神经传导速度高于术后4周,有显著性差异(P0.01);术后各时间点,患侧的胫神经运动神经传导速度均低于正常对照(健侧),有显著性差异(P0.01)。结论:大鼠胫神经损伤后远侧端髓鞘和轴索生长速度不一致。     

应用人工神经修复大鼠坐骨神经损伤的实验研究

目的观察应用复合碱性成纤维细胞生长因子(bFGF)-壳聚糖导管修复大鼠周围神经损伤的效果。方法成年Wistar大鼠25 只分为假手术组(n=10)、单纯损伤组(n=5)和人工神经修复组(n=10)。假手术组仅暴露坐骨神经5 mm,单纯损伤组暴露坐骨神经并切断,人工神经修复组以复合bFGF-壳聚糖导管桥接缺损。术后对实验动物的坐骨神经进行大体观察,对运动进行行为观察,以及组织化学和免疫组织化学方法评价神经再生情况和靶肌肉恢复情况。结果术后5 周,与单纯损伤组相比,人工神经修复组运动功能有一定程度恢复。人工神经修复组再生的坐骨神经已经通过bFGF-壳聚糖导管越过缺损并与远端相连接。免疫组织化学方法显示,坐骨神经再生段可观察到神经微丝(NF)阳性和S-100 阳性纤维。应用Masson 染色方法,可观察到人工神经修复组腓肠肌去纤维化程度相对于单纯损伤组有明显改善。结论应用bFGF-壳聚糖导管能有效修复周围神经损伤并使大鼠运动功能得到改善。     

周围神经术前肌电图结果与术中所见的比较研究及误诊原因分析

目的 探讨肌电图(EMG)检查对周围神经损伤的诊断意义，分析误诊的原因。方法 收集2000年1月至2003年4月行手术治疗的周围神经损伤患者63例(69条神经)，按神经损伤特点分为开放性周围神经损伤组、闭合性周围神经损害组、臂丛损伤组和神经修复后再生组，各组患者均于术前进行肌电图检测，并将检测结果与术中所见进行比较、分析。结果 开放性周围神经损伤组术前EMG对神经完全损伤的诊断符合率为73.08％，与术中所见结果比较，差异有统计学意义。闭合性周围神经损害组对受损神经的定性、定位诊断，均在术中得到证实。臂丛损伤组的大体定位正确率达96．30％，完全符合率达70．37％；对臂丛完全根性损伤的检出率为68．52％，与磁共振的检出率(55．56％)相比，差异无统计学意义。神经修复后再生组5条神经，EMG结果与术中所见3条符合，2条不符合。69条神经中，EMG检查完全符合率为71.01％，基本符合率为85．51％，完全不符率为13．04％；假阳性率为4．49％，假阴性率为22．73％。结论 EMG检查对损伤神经的定位、定性诊断及神经修复后再生状况的评价在临床诊治中具有重要的指导意义，但可出现假阳性及假阴性结果，且以运动诱发电位的假阴性为多。术前EMG与磁共振检查相结合，可提高对臂丛神经完全根性损伤的检出率。     

脑源性生长因子与神经损伤的修复及再生

背景:脑源性生长因子具有促进神经元生长存活,引导轴突延伸塑型的作用.周围神经损伤后的再生和髓鞘形成需要内源性脑源性神经生长因子.目的:归纳总结脑源性生长因子的研究现状.方法:以中文检索词"神经再生;脑源性生长因子"和英文检索词"nerve,regeneration,BDNF"检索2000-01/2009-08中国期刊全文数据库和Pubmed数据库.纳入具有原创性、论点论据可靠的试验文章,观点明确,分析全面的文章,及文献主题与此课题关系紧密的文章.排除重复性研究和综述文章.结果与结论:神经损伤后在髓鞘形成过程中脑源性神经生长因子通过高亲和力Trk受体和低亲和力受体P75~(NTR)促进髓鞘形成.与神经生长因子在周围靶组织合成不同,脑源性神经生长因子主要在中枢神经系统中合成,但当周围神经受损后其mRNA表达增多,大量的实验表明正常周围神经的许旺细胞同样有少量脑源性神经生长因子表达.现在人们通过将脑源性神经生长因子基因通过病毒介导转染干细胞后移植到神经损伤区域治疗疾病,有望成为新的治疗方法.     

Changes of pro-inflammatory and anti-inflammatory macrophages after peripheral nerve injury

Macrophages are notable immune cells that are recruited to the injury sites after peripheral nerve injury. Following peripheral nerve injury, increasing numbers of macrophages engulf debris and promote nerve regeneration. However, changes of pro-inflammatory (M1) and anti-inflammatory (M2) macrophages, two types of macrophages with dissimilar biological functions, have not been discovered. In the current study, the expression profiles of M1 and M2 macrophage marker genes in the sciatic nerve stumps and dorsal root ganglions (DRGs) after rat sciatic nerve injury were determined using RNA sequencing. Robust up-regulation of macrophage marker genes was observed in the injured sciatic nerve stumps as compared with in the DRGs. Measurement of the dynamic expression levels of M1 macrophage specific marker genes CD38 and Gpr18 as well as M2 macrophage specific marker genes Egr2 and Myc suggested that M1 macrophages were highly involved at all tested time points after peripheral nerve injury while M2 macrophage might be more involved in the later phase after nerve injury. Dynamic changes of M1 macrophage-inducing miRNAs showed that miR-18a, miR-19b, miR-21, miR-29a, and miR-29b were elevated in the injured nerve stump. These up-regulated miRNAs might mediate macrophage polarization by targeting multiple genes, such as Pten. Collectively, our study explored the unique temporal patterns of pro-inflammatory and anti-inflammatory macrophages after peripheral nerve injury for genetic aspects and provided a deeper understanding of the cellular and molecular basis of microenvironment reconstruction after peripheral nerve injury.

The temporal patterns of pro-inflammatory and anti-inflammatory macrophages after peripheral nerve injury.     

神经生长因子对挤压伤坐骨神经再生的促进作用

背景神经生长因子(nerve growth factor,NGF)不仅是维持促进中枢神经系统发育、分化、存活的基本的生长因子,而且在外周神经损伤的修复中也起着重要的作用.目的观察NGF肌注对大鼠坐骨神经挤压伤后神经再生恢复以及功能的影响.设计以实验动物为研究对象,随机对照的重复观察测量,探索性研究.单位一所军医大学的新药评价中心.材料实验于1999-07/2000-03在第二军医大学基础部新药评价中心完成.SD大鼠40只,雌雄各半,体质量200～250 g,上海西普尔-必凯实验动物有限公司提供.方法将40只大鼠随机分成NGF高、中、低剂量组,正常对照和模型对照组.距坐骨切迹远端6 mm处钳夹坐骨神经,使产生-4 mm宽的挤压伤.NGF高、中、低剂量组分别给予NGF 8,4和2μg/kg(1.6×103,8×102和4×102 IU/kg)药物,肌注1次/d,连续56 d.主要观察指标①手术后的不同时间神经传导速度(nerve conduction ve1ocities,NCV)和坐骨神经功能指数(sciatic functional index,SFI).②组织形态学评价.结果与模型对照组相比,高剂量组在35,56 d,中剂量组在35 d的NCV均显著加快(t=2.32～5.14,P＜0.05～0.01);高、中、低3个剂量组在手术14 d后的SFI与模型组相比,差异均有显著性意义(t=2.29～6.28,P＜0.05～0.01),且高剂量组恢复较明显,至56 d各剂量组SFI各组差异均无显著性意义;组织形态学显示,NGF治疗组神经髓鞘、轴索与正常对照组相比,差异无显著性意义;而模型组髓鞘出现脱失,细微结构不清晰,许旺细胞变性坏死.结论NGF对大鼠坐骨神经受损部位的神经髓鞘及轴索有明显的改善作用,能促进其神经纤维的再生及神经功能的恢复.     

Let-7 microRNAs Regenerate Peripheral Nerve Regeneration by Targeting Nerve Growth Factor

Peripheral nerve injury is a common clinical problem. Nerve growth factor (NGF) promotes peripheral nerve regeneration, but its clinical applications are limited by several constraints. In this study, we found that the time-dependent expression profiles of eight let-7 family members in the injured nerve after sciatic nerve injury were roughly similar to each other. Let-7 microRNAs (miRNAs) significantly reduced cell proliferation and migration of primary Schwann cells (SCs) by directly targeting NGF and suppressing its protein translation. Following sciatic nerve injury, the temporal change in let-7 miRNA expression was negatively correlated with that in NGF expression. Inhibition of let-7 miRNAs increased NGF secretion by primary cultured SCs and enhanced axonal outgrowth from a coculture of primary SCs and dorsal root gangalion neurons. In vivo tests indicated that let-7 inhibition promoted SCs migration and axon outgrowth within a regenerative microenvironment. In addition, the inhibitory effect of let-7 miRNAs on SCs apoptosis might serve as an early stress response to nerve injury, but this effect seemed to be not mediated through a NGF-dependent pathway. Collectively, our results provide a new insight into let-7 miRNA regulation of peripheral nerve regeneration and suggest a potential therapy for repair of peripheral nerve injury.     

Metallothionein deficiency in the injured peripheral nerves of complex regional pain syndrome as revealed by proteomics

Complex regional pain syndrome (CRPS) is characterized by persistent and severe pain after trauma or surgery; however, its molecular mechanisms in the peripheral nervous system are poorly understood. Using proteomics, we investigated whether injured peripheral nerves of CRPS patients have altered protein profiles compared with control nerves. We obtained nerve samples from 3 patients with CRPS-2 who underwent resection of part of an injured peripheral nerve. Sural nerves from fresh cadavers with no history of trauma or neuropathic pain served as controls. Proteomic analysis showed that the number and functional distribution of proteins expressed in CRPS and control nerves was similar. Interestingly, metallothionein was absent in the injured nerves of CRPS-2, although it was readily detected in control nerves. Western blotting further confirmed the absence of metallothionein in CRPS-2 nerves, and immunohistochemistry corroborated the deficiency of metallothionein expression in injured nerves from 5 of 5 CRPS patients and 2 of 2 patients with painful neuromas. In contrast, all control nerves, including 5 sural nerves from fresh cadavers and 41 nerves obtained from surgically resected tumors, expressed MT. Furthermore, expression of S100 as a marker for Schwann cells, and neurofilament M as a marker of axons was comparable in both CRPS-2 and controls. Metallothioneins are zinc-binding proteins that are probably involved in protection against injury and subsequent regeneration after CNS damage. Their absence from the injured peripheral nerves of patients with CRPS-2 suggests a potential pathogenic role in generating pain in the damaged peripheral nerves.     

Visualization of peripheral nerve degeneration and regeneration: monitoring with diffusion tensor tractography

We applied diffusion tensor tractography (DTT), a recently developed MRI technique that reveals the microstructures of tissues based on its ability to monitor the random movements of water molecules, to the visualization of peripheral nerves after injury. The rat sciatic nerve was subjected to contusive injury, and the data obtained from diffusion tensor imaging (DTI) were used to determine the tracks of nerve fibers (DTT). The DTT images obtained using the fractional anisotropy (FA) threshold value of 0.4 clearly revealed the recovery process of the contused nerves. Immediately after the injury, fiber tracking from the designated proximal site could not be continued beyond the lesion epicenter, but the intensity improved thereafter, returning to its pre-injury level by 3 weeks later. We compared the FA value, a parameter computed from the DTT data, with the results of histological and functional examinations of the injured nerves, during recovery. The FA values of the peripheral nerves were more strongly correlated with axon-related (axon density and diameter) than with myelin-related (myelin density and thickness) parameters, supporting the theories that axonal membranes play a major role in anisotropic water diffusion and that myelination can modulate the degree of anisotropy. Moreover, restoration of the FA value at the lesion epicenter was strongly correlated with parameters of motor and sensory functional recovery. These correlations of the FA values with both the histological and functional changes demonstrate the potential usefulness of DTT for evaluating clinical events associated with Wallerian degeneration and the regeneration of peripheral nerves.     

电疗法对周围神经损伤修复的研究进展

近年来,周围神经损伤的修复和再生有了若干进展。本文分别从低频电疗法与直流电疗法、中频电疗法、高频电疗法（超短波、分米波与毫米波）等角度探讨国内外学者在电疗法对周围神经损伤修复方面的研究及进展。 由于周围神经对不同程度的损伤有不同的病理过程和不同的再生方式,因此,周围神经损伤后,电疗法的选择应按照神经恢复的不同阶段要求来调整治疗方法。但其作用机制仍有待进一步深入研究。     

The dynamic changes of main cell types in the microenvironment of sciatic nerves following sciatic nerve injury and the influence of let-7 on their distribution

Schwann cells (SCs), fibroblasts and macrophages are the main cells in the peripheral nerve stumps. These three types of cell play important roles in regulating the regeneration microenvironment and improving the regeneration effect through a variety of manual interventions. However, the dynamic distribution of these cells during the different stages of peripheral nerve regeneration remain unclear. In this study, we systematically explored the number/ratio and distribution changes of SCs, fibroblasts, and macrophages at different time points after sciatic nerve injury. Moreover, considering that let-7 is an ideal molecule for regulating the regeneration environment, we further studied the entrance and influence of let-7 antagomir on these three main cell types. Collectively, our current study revealed the cell basis of the microenvironment of peripheral nerve regeneration and indicated that let-7 could modify the regenerative microenvironment by regulating the number/ratio and distribution of SCs, fibroblasts, and macrophages. Our study would help to open a new potential therapeutic window for peripheral nerve injury.

Schwann cells (SCs), fibroblasts and macrophages are the main cells in the peripheral nerve stumps.     

脊髓损伤模型大鼠鞘内注射甲基强的松龙后的蛋白质组学变化

背景：临床研究表明,早期使用甲基强的松龙可以促进损伤神经功能恢复,合理的初始剂量、剂量间隔时间和治疗持续时间是甲基强的松龙治疗急性脊髓损伤取得良好效果的关键。目的：观察急性脊髓损伤模型大鼠鞘内注射大剂量甲基强的松龙后的脊髓组织差异蛋白表达质谱。方法：取8只SD大鼠,建立急性脊髓损伤模型,随机分2组,分别于造模后0,8 h鞘内注射甲基强的松龙7.5 mg/kg,于注射24 h后取损伤段脊髓组织,采用同位素标记相对和绝对定量技术分析两组差异蛋白质及表达明显相关神经再生的差异蛋白。结果与结论：共鉴定到了87个差异表达的蛋白,与0 h组相比,8 h组上调的差异蛋白有43个,下调的差异蛋白数是44个。相关神经再生的差异蛋白18个,上调的差异蛋白有8个,下调的差异蛋白数是10个,其中OMgp是一个潜在的神经轴突生长抑制因子,OMgp与NgR/P75/TROY/Lingo-1组成受体复合体特异性结合,通过第二信使cAMP激活RhoA作用可抑制轴突生长锥的崩解。提示鉴定鞘内注射甲基强的松龙治疗大鼠急性脊髓损伤后脊髓差异蛋白和脊髓相关神经再生因子,分别进行蛋白数据库检索和功能分析,其表达可能作为急性脊髓损伤后临床上监测神经损伤再生的指标。     

A role for apolipoprotein E, apolipoprotein A-I, and low density lipoprotein receptors in cholesterol transport during regeneration and remyelination of the rat sciatic nerve.

Recent work has demonstrated that apo E secretion and accumulation increase in the regenerating peripheral nerve. The fact that apoE, in conjunction with apoA-I and LDL receptors, participates in a well-established lipid transfer system raised the possibility that apoE is also involved in lipid transport in the injured nerve. In the present study of the crushed rat sciatic nerve, a combination of techniques was used to trace the cellular associations of apoE, apoA-I, and the LDL receptor during nerve repair and to determine the distribution of lipid at each stage. After a crush injury, as axons died and Schwann cells reabsorbed myelin, resident and monocyte-derived macrophages produced large quantities of apoE distal to the injury site. As axons regenerated in the first week, their tips contained a high concentration of LDL receptors. After axon regeneration, apoE and apoA-I began to accumulate distal to the injury site and macrophages became increasingly cholesterol-loaded. As remyelination began in the second and third weeks after injury, Schwann cells exhausted their cholesterol stores, then displayed increased LDL receptors. Depletion of macrophage cholesterol stores followed over the next several weeks. During this stage of regeneration, apoE and apoA-I were present in the extracellular matrix as components of cholesterol-rich lipoproteins. Our results demonstrate that the regenerating peripheral nerve possesses the components of a cholesterol transfer mechanism, and the sequence of events suggests that this mechanism supplies the cholesterol required for rapid membrane biogenesis during axon regeneration and remyelination.     

生物粘接端端缝合法修复外周神经的实验研究

目的　研究生物粘接端端缝合修复外周神经的早期效果。方法　选用 Wister大鼠 32只 ,在双目放大镜下分别用生物粘接端端缝合法和单纯端端缝合法修复坐骨神经损伤 ,2周后进行组织形态学、电生理学、坐骨神经功能指数检测评定。结果　生物粘接端缝合组各项指标均优于对照组 ,其神经纤维的生长速度、数量明显优于单纯缝合组 ,SFI的恢复优于单纯缝合组。结论　生物粘接端端缝合法可以促进神经再生 ,加快神经再生速度与神经功能恢复。     

鼠神经生长因子不同给药方式修复周围神经损伤

背景：鼠神经生长因子对神经损伤后的修复和再生有促进作用,但目前实验研究表明不同用药方式的作用尚有争议。目的：评价鼠神经生长因子不同给药方式治疗周围神经损伤的临床疗效。方法：52例周围神经损伤的患者随机分为2组,试验组27例局部注射鼠神经生长因子;对照组25例全身注射鼠神经生长因子,1次/d,一个疗程4周,比较两组患者神经功能的修复情况及疗效。结果与结论：与对照组相比,试验组优良率85%（23例）,有效率93%（25例）;对照组的优良率72%（18例）,有效率84%（21例）,两组优良率与有效率相比试验组显著优于对照组（P〈0.05）。试验组中13例出现注射部位一过性痛,其中1例患者口服镇痛药物缓解;对照组中12例患者出现注射部位一过性疼痛,未做处理。结果提示,鼠神经生长因子治疗周围神经损伤安全有效,局部注射疗效优于全身用药。     

周围神经损伤的早期显微外科修复

目的探索周围神经损伤早期显微外科修复的临床效果及预后。方法根据神经损伤的部位及程度应用不同的显微外科方法进行修复。具体方法为神经探查、神经松解、神经外膜修补、神经外膜端端吻合、神经束膜端端吻合、神经移植、神经移位、神经植入等。结果共修复周围神经118例12条166处，获随访72例12条98处，运动功能恢复达M3以上31．82％，感觉功能恢复达S3以上67．74％，其中前臂内外侧皮神经、股外侧皮神经、腓肠神经损伤修复后感觉恢复均达到S3，28处指神经损伤修复后感觉恢复达s3以上占82．14％，尺神经、腓总神经的运动功能无一例恢复，桡神经及股神经的运动功能恢复最理想。结论应用显微外科技术早期修复周围神经损伤效果良好，但应注意修复时机及方式。     

自体肋间神经联合酸性成纤维细胞生长因子移植治疗高位脊髓损伤

背景:酸性成纤维细胞生长因子具有调节细胞增殖、移行、分化和生存的作用,也可以下调已知轴突再生的抑制因子如蛋白聚糖等,帮助轴突克服这些抑制因子,对神经纤维再生有重要作用.目的:观察酸性成纤维细胞生长因子联合周围神经移植治疗大鼠高位脊髓损伤的可行性及效果.方法:健康成年雌性SD大鼠108只随机抽签法分为自体神经组、自体神经联合生长因子组、高位脊髓横断组.咬除大鼠T_(8-10)棘突、椎板,显露硬膜囊,水平切断高位脊髓并切除3 mm,显微镜下确认无神经纤维相连.自体神经组、自体神经联合生长因子组取双侧第8～10对肋间神经各2 cm,将肋间神经交叉移植入高位脊髓缺损处(近端白质与远端灰质、远端白质与近端灰质),分别以纤维蛋白凝胶、含有酸性成纤维细胞生长因子的纤维蛋白凝胶固定植入的肋间神经,缝合硬膜.高位脊髓横断组断端间旷置.术后90 d,行体感诱发电位及运动诱发电位检测观察神经电生理恢复情况.术后76 d,生物素葡聚糖胺顺行神经示踪观察运动传导束恢复情况.术后60 d,后肢BBB运动功能评分观察肢体运动恢复情况.结果与结论:高位脊髓横断组大鼠均未引出体感及运动诱发电位波形.自体神经组、自体神经联合生长因子组均可引出体感及运动诱发电位,自体神经联合生长因子组体感诱发电位及运动诱发电位的平均潜伏期和波幅、BBB评分均明显优自体神经组(P＜0.01).自体神经组和自体神经联合生长因子组在损伤区有较多生物素葡聚糖胺标记阳性神经纤维通过,明显多于高位脊髓横断组(P＜0.01),自体神经联合生长因子组多于自体神.经组(P＜0.01).示自体周围神经移植酸性成纤维细胞生长因子能更好地恢复高位脊髓损伤后大鼠肢体运动功能.     

低强度超声促进周围神经损伤后的再生

目的：探讨低强度超声对周围神经损伤后再生的作用。方法：将64只大鼠右侧坐骨神经重度钳夹伤制作神经损伤模型，随机分为2组各32只。超声组造模侧神经损伤处以声强250mW／cm^2、频率1．0MHz的超声进行体外治疗，隔天1次；对照组相应部位予以未启动治疗系统的假治疗。术后不同时期进行电生理、坐骨神经功能指数等指标测定及组织学检查。结果：超声组损伤神经远侧Wallerian变性进程加速、雪旺细胞增殖、变性组织吸收，轴索及髓鞘再生、感觉传导速度及坐骨神经功能的恢复等与对照组比较均提前（P〈0．01或P〈0．05）。结论：低强度超声通过影响神经再生的多个环节而促进周围神经的再生和功能恢复。