Cationic liposome-mediated GDNF gene transfer after spinal cord injury

Glial cell line-derived neurotrophic factor (GDNF) has been shown to protect cranial and spinal motoneurons, which suggests potential uses of GDNF in the treatment of spinal cord injury (SCI) and motor neuron disease. We examined neuroprotective effect of cationic liposome-mediated GDNF gene transfer in vivo on axonal regeneration and locomotor function recovery after SCI in adult rats. The mixture of DC-Chol liposomes and recombinant plasmid pEGFP-GDNF cDNA was injected after SCI. RT-PCR confirmed the increased expression of GDNF mRNA in the injected areas at 7 days after injection. The expression of EGFP-GDNF was observed in the cells around the injection locus by fluorescence microscope at least 4 weeks after injection. Four weeks after GDNF gene transfer, regeneration of the corticospinal tracts was assessed using anterograde tract tracing. There are more HRP labeling of corticospinal tract axons across the lesion in GDNF group compared with control group. In GDNF group, the maximum distance these labeled axons extended varied in different animals and ranged from 5 mm to approximately 9 mm from the lesion. In control group, no HRP labeled axons extended caudal to the lesion. The locomotion function of hindlimbs of rats was evaluated using inclined plane test and BBB locomotor scores. The locomotion functional scores in GDNF group were higher than that in control group within 1-4 weeks after SCI (p < 0.05). These data demonstrate that in vivo transfer of GDNF cDNA can promote axonal regeneration and enhance locomotion functional recovery, suggesting that cationic liposome-mediated delivery of GDNF cDNA may be a practical gene transfer method for traumatic SCI treatment.     

胚胎脊髓不同移植方法对损伤后大鼠脊髓轴突再生的作用

[目的]研究胚胎脊髓不同移植方法对损伤后大鼠脊髓轴突再生的作用。[方法]将成年大鼠分为四组，A组：单纯脊髓挫伤和半切洞组；B组：脊髓挫伤和半切洞＋胚胎脊髓移植组；C组：脊髓挫伤和半切洞＋胚胎脊髓移植＋损伤区上下神经根吻合组；D组：脊髓挫伤和半切洞＋胚胎脊髓移植＋带蒂大网膜移植组。移植后6周，应用行为学检查，观察大鼠功能恢复情况，用光、电镜检查计算轴突和观察轴突病理变化，采用计算机图像分析技术，进行定量分析。[结果]作者发现脊髓损伤后6周A组轴突丢失最严重，各组轴突都有不同程度的减少，其减少程度顺序是A组〉B组〉C组〉D组。D组与其它三组比较有显著差异性。残留的轴突数目与神经功能的改善相平行。[结论]大鼠胚胎脊髓和大网膜移植后能增强损伤脊髓轴突再生并能促进大鼠后肢神经功能的恢复。     

Calcitonin gene-related peptide immunoreactivity in chronic human spinal cord injury

STUDY DESIGN: Histopathological study of the human spinal cord. SETTING: International Collaboration on Repair Discoveries, Vancouver, BC, Canada. RATIONALE: In animals, primary dorsal root afferent fibers, which are immunoreactive for calcitonin gene-related peptide (CGRP), sprout following spinal cord injury (SCI) into deeper laminas of the dorsal horn below the level of injury. It has been suggested that this aberrant sprouting plays a role in altering cardiovascular control after SCI and could be responsible for life-threatening episodes of autonomic dysreflexia (AD). OBJECTIVES: To observe the changes of CGRP distribution after SCI and compare the differences between normal and injured human spinal cord. METHODS: Upper thoracic segments from individuals with chronic cervical SCI (n=4) and individuals with intact spinal cords (n=5) were processed immunocytochemically to identify CGRP fibers and histologically to identify the severity of degeneration. RESULTS: Semiquantitative analysis showed a significant increase in CGRP immunoreactivity in the dorsal horns of individuals with chronic SCI (P<0.001). Furthermore, one of the SCI individuals in this study displaying significant CGRP sprouting had well documented episodes of AD. CONCLUSIONS: Our observations suggest that SCI in humans results in significant sprouting of CGRP fibers. This aberrant sprouting of sensory fibers could contribute to the abnormal cardiovascular control and pain commonly observed following chronic human SCI.     

胚胎脊髓不同移植方法对损伤后大鼠脊髓轴索病理的影响

目的研究胚胎脊髓不同移植方法对损伤后大鼠脊髓轴索病理的影响。方法将成年大鼠40只均分为四组,A组：单纯脊髓挫伤和半切洞组;B组：脊髓挫伤和半切洞＋胚胎脊髓移植组;C组：脊髓挫伤和半切洞＋胚胎脊髓移植＋损伤区上下神经根吻合组;D组：脊髓挫伤和半切洞＋胚胎脊髓移植＋带蒂大网膜移植组。移植后6周,应用行为学检查,观察大鼠功能恢复情况,用光、电镜检查计算轴索和观察轴索病理变化,采用计算机图像分析技术,进行定量分析。结果脊髓损伤后6周A组轴索丢失最严重,各组轴索都有不同程度的减少,其减少程度顺序是A组〉B组〉C组〉D组。D组与其它三组比较有显著差异性。残留的轴索数目与神经功能的改善相平行。结论大鼠胚胎脊髓和大网膜移植后对损伤的大鼠脊髓轴索有保护作用并能促进大鼠后肢神经功能的恢复。     

In silico modeling of axonal reconnection within a discrete fiber tract after spinal cord injury

Following spinal cord injury (SCI), descending axons that carry motor commands from the brain to the spinal cord are injured or transected, producing chronic motor dysfunction and paralysis. Reconnection of these axons is a major prerequisite for restoration of function after SCI. Thus far, only modest gains in motor function have been achieved experimentally or in the clinic after SCI, identifying the practical limitations of current treatment approaches. In this paper, we use an ordinary differential equation (ODE) to simulate the relative and synergistic contributions of several experimentally-established biological factors related to inhibition or promotion of axonal repair and restoration of function after SCI. The factors were mathematically modeled by the ODE. The results of our simulation show that in a model system, many factors influenced the achievability of axonal reconnection. Certain factors more strongly affected axonal reconnection in isolation, and some factors interacted in a synergistic fashion to produce further improvements in axonal reconnection. Our data suggest that mathematical modeling may be useful in evaluating the complex interactions of discrete therapeutic factors not possible in experimental preparations, and highlight the benefit of a combinatorial therapeutic approach focused on promoting axonal sprouting, attraction of cut ends, and removal of growth inhibition for achieving axonal reconnection. Predictions of this simulation may be of utility in guiding future experiments aimed at restoring function after SCI.     

Effect of fetal spinal cord graft with different methods on axonal pathology after spinal cord contusion

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Antibodies neutralizing Nogo-A increase pan-cadherin expression and motor recovery following spinal cord injury in rats

STUDY DESIGN: A rat model of spinal cord injury was used to test the hypothesis that Nogo-A monoclonal antibody (NEP1-40) promotes morphologic and functional recoveries of injured spinal cord. OBJECTIVE: Nogo-A is a myelin-associated neurite outgrowth inhibitory protein, which blocks elongation nerve fibers and limits neuronal regeneration after central nervous system injury. METHODS: Forty-four rats were utilized and allocated into control (vehicle) and NEP1-40-treated groups. In all animals, the spinal cord was hemi-transected at Th-10 and phosphate-buffered saline solution was immediately applied on the injured area in the control group. NEP1-40 solution was immediately applied on the hemi-transected area in the treatment group. Each group was subdivided into three subgroups according to the postsurgical day of killing (3, 8 and 21 days). The spinal cords were removed for analysis. RESULTS: Motor scores in the NEP1-40-treated groups were significantly higher than those in the vehicle groups both at 8 and 21 days post injury. Immunohistochemical staining for pan-cadherin, a marker of neuronal cell adhesion and axonal sprouting, revealed a significant increase in staining in the NEP1-40 treatment group at 8 and 21 days post injury. Transmission electron microscopical evaluation revealed degeneration of the myelin and loss of cytoarchitectural organization in the axons of controls. Better preservation and normal histologic features were observed in the NEP1-40-treated groups. CONCLUSION: We have demonstrated improved preservation of injured axons and significant pan-cadherin expression after NEP1-40 treatment after the spinal cord injury. Inhibition of Nogo-A may improve the capacity for neuronal regeneration after spinal cord injury.     

NT-3基因修饰许旺细胞与神经干细胞联合移植促进大鼠全横断脊髓受损伤神经元的存活及其轴突再生

目的探讨神经营养素-3基因修饰许旺细胞（NT-3-SCs）与神经干细胞（NSCs）联合移植对大鼠因脊髓全横断而受损伤的神经元存活及其轴突再生的作用。方法将NT-3-SCs及LacZ报告基因修饰SCs（LacZ-SCs）与NSCs联合移植到全横断性脊髓损伤处，60d后在脊髓横断处尾侧注射荧光金（FG）进行逆行标记。第67天，取材进行组织学检测大脑皮质体感区、红核及脊髓横断处头侧FG标记神经元；大脑皮质体感区、红核和脊髓背核内存活的神经元以及脊髓损伤处再生的轴突。结果大脑皮质体感区、红核及脊髓L1背核内存活的神经元由多到少依次为NT-3-SCs与NSCs联合组、LacZ—SCs与NSCs联合组和实验对照组。NT-3-SCs与NSCs联合组和LacZ—SCs与NSCs联合组的大脑皮质体感区、红核和脊髓横断处头侧有FG标记神经元；脊髓横断处及其附近组织有5-HT、CGRP和SP阳性神经纤维。结论NT-3-SCs与NSCs联合移植能够促进脊髓全横断损伤后神经元的存活以及轴突再生。     

Resection of glial scar following spinal cord injury

While many studies have focused on modulating the immune response and enhancing axonal regeneration after spinal cord injury (SCI), there is limited work being performed on evaluating the role of glial scar in SCI. We sought to evaluate the effects of glial scar resection in contusion models and dorsal hemisection models of SCI. At 1‐week postinjury, 2 mm of glial scar was excised from specimens in one of the two groups from each injury model. Functional outcome was measured weekly using the Basso, Beattie, Bresnahan (BBB) Locomotor Rating Scale along with histologic evaluation of spinal cord tracts to determine axonal regeneration. Within the dorsal hemisection model, there was no significant difference in recovery for animals that underwent glial scar excision versus animals that did not have scar excision (p = 0.61). Animals subjected to the contusion model, however, demonstrated lower BBB scores in the glial resection group during the earlier postoperative periods (<4 weeks; p < 0.05). Histological analysis revealed no axons within the glial resection contusion model, and moderate axonal growth within the nonresection contusion group and both hemisection groups (p > 0.05 for differences among the three groups). While glial scar may serve to stabilize the preserved axonal tracts and thereby permit modest recovery in a contusion model of SCI, it may be of less importance with a dorsal hemisection model. These experiments highlight that basic biologic processes following SCI may vary tremendously based on the injury mechanism and that the role of glial scar in spinal cord regeneration must be elucidated. © 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 27: 931–936, 2009     

嗅鞘细胞移植在大鼠脊髓损伤处能够迁移和促进神经轴突生长吗?

目的：探讨嗅鞘细胞（olfactory ensheathing cells,OEC）移植在大鼠损伤脊髓后是否有独特的迁移和轴突生长导向特性。方法：将表达绿色荧光蛋白基因的OEC注入到C4脊髓损伤大鼠距离损伤部位头端1mm及尾端1mm背柱白质处,注射后1、3、12、24h及3、7、28d灌注固定取材,冰冻切片和免疫组化分析。用骨髓基质细胞和成纤维细胞移植到同样的损伤部位做为对照进行同样的分析。另将OEC注射到距离损伤部位头尾侧1mm处脊髓灰质内、小剂量注射在白质内、在损伤前3d或损伤后9d注射、注射到未损伤脊髓白质中,观察细胞迁移情况。结果：OEC在细胞注射后1h内即形成由注射压力造成的从注射部位向损伤处的被动性延伸带,并且不断地从注射部位向损伤处延伸扩散,在形态学方面好象起到＂桥接＂损伤处的作用。对照组骨髓基质细胞和成纤维细胞注射后也迅速形成细胞＂桥接＂带,并扩散至损伤空腔。将OEC注射到脊髓灰质内或小剂量注射或注射到未损伤的脊髓白质内、在脊髓损伤3d前或9d后注射细胞均没有见到细胞带延伸进入损伤部位。OEC在注射部位、细胞带以及损伤部位有增殖现象。28d后,共焦免疫酶标法证实细胞带取代了脊髓原有星形胶质细胞,但是下行或上行长束轴突没有优先延伸至该细胞带,也没能起到维持皮层脊髓轴突的桥接作用。结论：OEC在植入大鼠损伤脊髓后没有独特的迁移特性,与骨髓基质细胞或成纤维细胞相比没有明显促进轴突生长的作用,也没有支持皮层脊髓轴突在脊髓损伤处形成桥接的作用。     

A histopathological analysis of the human cervical spinal cord in patients with acute traumatic central cord syndrome

STUDY DESIGN: We have applied conventional histochemical and morphometric techniques to study the changes within the human spinal 'hand' motor neuron pool after spinal cord injury in patients who presented with acute traumatic central cord syndrome (ATCCS). OBJECTIVE: To determine whether a reduction of large alpha motor neurons at the C7, C8 and T1 spinal cord levels underlies the mechanism which causes hand dysfunction seen in patients with (ATCCS). BACKGROUND: The etiology of upper extremity weakness in ATCCS is debated and injury and/or degeneration of motor neurons within the central gray matter of the cervical enlargement has been advanced as one potential etiology of hand weakness. METHODS: The spinal cords of five individuals with documented clinical evidence of ATCCS and three age-matched controls were obtained. The ATCCS spinal cords were divided into acute/sub-acute (two cases) and chronic (three cases) groups depending on the time to death after their injury; the chronic group was further subdivided according to the epicenter of injury. We counted the motor neurons using light microscopy in 10 randomly selected axial sections at the C7, C8 and T1 spinal cord levels for each group. We also analyzed the lateral and ventral corticospinal tracts (CST) in all groups for evidence of Wallerian degeneration and compared them to controls. RESULTS: A primary injury to the lateral CST was present in each case of ATCCS with evidence of Wallerian degeneration distal to the epicenter of injury. There was minimal Wallerian degeneration within the ventral corticospinal tracts. In the chronic low cervical injury group, there was a decrease in motor neurons supplying hand musculature relative to the other injury groups where the motor neurons sampled at the time of death were not reduced in number when compared to the control group. CONCLUSIONS: We hypothesize that hand dysfunction in ATCCS can be observed after spinal cord injury without any apparent loss in the number of motor neurons supplying the hand musculature as seen in our acute/sub-acute (n=2) and our chronic high injury (n=1) groups. The motor neuron loss seen in the chronic low level injury was felt to be secondary to the loss of C7, C8, and T1 neurons adjacent to the injury epicenter.     

Present situation and future aspects of spinal cord regeneration

The central nervous system (CNS) has a limited capacity for regeneration after injury. In spinal cord injury (SCI) patients, total loss of all motor and sensory function occurs below the level of injury. Advances in treatment are expected for orthopedic and spinal surgeons. Recently, evidence of axonal regeneration and functional recovery has been reported in animal spinal cord injury models. Our studies on the roles of inhibitory molecules with a comparison between neonatal and adult animals may help serve as therapeutic targets to enhance axonal regeneration for the injured spinal cord. Also, our cell replacement study indicates the possibility of transplanting neural stem cells to supply the cell source for immature oligodendrocytes, which are thought to be essential for both the myelination and trophic support of regenerating axons in the spinal cord. Administration of neurotrophic factors, prevention of inhibitory factors, and stem cell technology have clinical applications in SCI patients. However, spinal cord regeneration involves a multistep process, and several factors have to be controlled after injury. A combination of several treatments could overcome a nonpermissive environment for spinal cord regeneration. Further understanding of the mechanisms and finding optimal targets of spinal cord regeneration are necessary to obtain successful therapies for SCI patients.Presented at the 76th Annual Meeting of the Japanese Orthopaedic Association, Kanazawa, Japan, May 23, 2003     

Axonal remyelination by cord blood stem cells after spinal cord injury

Human umbilical cord blood stem cells (hUCB) hold great promise for therapeutic repair after spinal cord injury (SCI). Here, we present our preliminary investigations on axonal remyelination of injured spinal cord by transplanted hUCB. Adult male rats were subjected to moderate SCI using NYU Impactor, and hUCB were grafted into the site of injury one week after SCI. Immunohistochemical data provides evidence of differentiation of hUCB into several neural phenotypes including neurons, oligodendrocytes and astrocytes. Ultrastructural analysis of axons reveals that hUCB form morphologically normal appearing myelin sheaths around axons in the injured areas of spinal cord. Colocalization studies prove that oligodendrocytes derived from hUCB secrete neurotrophic hormones neurotrophin-3 (NT3) and brain-derived neurotrophic factor (BDNF). Cord blood stem cells aid in the synthesis of myelin basic protein (MBP) and proteolipid protein (PLP) of myelin in the injured areas, thereby facilitating the process of remyelination. Elevated levels of mRNA expression were observed for NT3, BDNF, MBP and PLP in hUCB-treated rats as revealed by fluorescent in situ hybridization (FISH) analysis. Recovery of hind limb locomotor function was also significantly enhanced in the hUCB-treated rats based on Basso-Beattie-Bresnahan (BBB) scores assessed 14 days after transplantation. These findings demonstrate that hUCB, when transplanted into the spinal cord 7 days after weight-drop injury, survive for at least 2 weeks, differentiate into oligodendrocytes and neurons, and enable improved locomotor function. Therefore, hUCB facilitate functional recovery after moderate SCI and may prove to be a useful therapeutic strategy to repair the injured spinal cord.     

Benefit of chondroitinase ABC on sensory axon regeneration in a laceration model of spinal cord injury in the rat

BACKGROUND: Chondroitin sulfate proteoglycans are up-regulated in the spinal cord after SCI, creating a molecular barrier inhibitory to axon growth. Chondroitinase ABC degrades CSPGs in vitro and in vivo. METHODS: We studied whether IT ChABC promotes axonal regeneration in a laceration model of SCI. Three groups of Sprague-Dawley rats were used: control and rats treated with low-dose and high-dose IT ChABC. Chondroitin sulfate proteoglycan breakdown products were measured by 2-B-6 expression, and intact CSPGs by CS-56 expression. Sensory axonal regeneration was traced after CTB injection into the median, ulnar, and sciatic nerves. RESULTS: CS-56 expression was down-regulated and 2-B-6 expression was increased in the groups treated with IT ChABC but not in the control. Laminin and GFAP immunoreactivity was unaltered in the ChABC groups. The number of axons growing into the scar was 3.1 times greater (P < .01) in the high-dose ChABC group and 2.1 times greater (P < .01) in the low-dose group compared with the controls. The length of axonal growth after high- and low-dose ChABC was 9.9 (P < .01) and 8.3 (P < .01) times greater, respectively, than in the control group. Axons extended across the lesion gap and into the distal spinal cord stump in 2 of 8 (low dose) and in 3 of 9 (high dose) rats compared with none in the control group. CONCLUSIONS: Intrathecal ChABC administration caused a slight decrease in CSPGs in the scar after a laceration SCI with a minimal increase in sensory axonal regeneration into and across the laceration gap.     

Influences of olfactory ensheathing cells transplantation on axonal regeneration in spinal cord of adult rats

Objective: To observe whether offactory ensheathing cells could be used to promote axonal regeneration in a slmntaneously nonregenerating system. Methods: After laminectomy at the lower thoracic level, the spinal cords of adult rats were exposed and completely transected at T10. A suspension of ensheathing cells was injected into the lesion site in 12 adult rats, and control D/F-12 (1:1 mixture of DMEM and Ham‘s F-12) was injected in 12 adult rats. Six weeks and ten weeks after cell transplantation, the rats were evaluated by climbing test and motor evoked potentials (MEPs) monitoring. The samples were procured and studied with histologiel and immounohistochemical methods. Results: At the 6th week after cell transplantation,d the rats in both the transplanted and control groups were paraplegic and the MEPs could not be recorded. At the 10th week after cell transplantation, of 7 rats in the control group, 2 rats had muscles‘ contraction of the lower extremities, 2 rats had hips and/or knees‘ active movement; and 5 rats‘ MEPs could be recorded in the hind limbs in the transplanted group ( n = 7). None of the rats in the control group had functional improvement and no MEPs recorded ( n = 7 ). Numerous regenerating axons were observed through the transplantation and continued to regenerate into the denervated host tract. Cell labelling using anti-Myelin Basic Protein (MBP) and anti-Nerve Growth Factor Receptor (anti-NGFR) indicated that the regenerated axons were derived from the appropriate neuronal source and that donor cells migrated into the denervated host tract. But axonal degeneration existed and regenerating axons were not observed within the spinalcords of the adult rats with only D/F-12 injection. Conclusions: The axonal regeneration in the transected adult rat spinal cord is possible after eusheathing cells transplantation.     

Chronic cervical spinal cord injury: DTI correlates with clinical and electrophysiological measures

Diffusion tensor imaging (DTI) is rarely applied in spinal cord injury (SCI). The aim of this study was to correlate diffusion properties after SCI with electrophysiological and neurological measures. Nineteen traumatic cervical SCI subjects and 28 age-matched healthy subjects participated in this study. DTI data of the spinal cord were acquired with a Philips Achieva 3 T MR scanner using an outer volume suppressed, reduced field of view (FOV) acquisition with oblique slice excitation and a single-shot EPI readout. Neurological and electrophysiological measures, American Spinal Injury Association (ASIA) impairment scale scores, and motor (MEP) and somatosensory evoked potentials (SSEP) were assessed in SCI subjects. Fractional anisotropy (FA) values were decreased in the SCI subjects compared to the healthy subjects. In upper cervical segments, the decrease in FA was significant for the evaluation of the entire cross-sectional area of the spinal cord, and for corticospinal and sensory tracts. A decreasing trend was also found at the thoracic level for the corticospinal tracts. The decrease of DTI values correlated with the clinical completeness of SCI, and with SSEP amplitudes. The reduced DTI values seen in the SCI subjects are likely due to demyelination and axonal degeneration of spinal tracts, which are related to clinical and electrophysiological measures. A reduction in DTI values in regions remote from the injury site suggests their involvement with wallerian axonal degeneration. DTI can be used for the quantitative evaluation of the extent of spinal cord damage, and eventually to monitor the effects of future regeneration-inducing treatments.     

cAMP诱导大鼠脊髓损伤后神经再生

目的观察在体给入环磷酸腺苷(cyclic adinosine monophosphate,cAMP)对大鼠脊髓损伤后神经再生的作用。方法56只SD大鼠制成脊髓T10背侧半切断损伤模型,随机分为六组。各组分别在脊髓损伤局部、大脑运动皮层和T6蛛网膜下腔内给入cAMP或生理盐水。动物存活6周后处死,制作脊髓组织切片,通过免疫组化染色观察损伤区局部神经丝的分布;通过皮质脊髓束和脊髓顺行追踪观察皮质脊髓束和脊髓神经纤维的再生;通过动物后肢运动BBB评分观察后肢运动功能恢复的程度,并以此来评价治疗措施的脊髓损伤修复效果。结果用三种方式给入cAMP,损伤区神经丝沿脊髓纵轴延伸分布,但未与尾端重新连接。在蛛网膜下腔给入cAMP无明显的脊髓神经再生;在损伤区局部和大脑运动皮层给入cAMP,损伤区可见大量脊髓再生纤维,包括部分皮质脊髓神经纤维再生。各组动物在术后4～5周均恢复正常行走,BBB评分超过20分。结论在脊髓损伤局部和在大脑运动皮层给入cAMP能诱导大鼠脊髓损伤后神经再生,但尚不能达到有效促进实验动物后肢运动功能的效果。     

手术减压时间对损伤脊髓轴索病理和损伤区面积的影响

目的 探讨大鼠脊髓损伤后手术减压时间对大鼠脊髓轴索病理和损伤区面积的影响。方法 将动物分为两组：大鼠脊髓挫伤2h手术减压组（A组）,大鼠脊髓挫伤8h手术减压组（B组）。手术后1、3、7、14、28d进行轴索病理变化的观察并测量脊髓损伤面积,采用计算机图像分析技术,进行定量分析。计算Tarlv评分并检测感觉诱发电位（SEP）和运动诱发电位（MEP）。结果 图象分析发现：脊髓损伤后B组轴索丢失明显多于A组。脊髓损伤面积B组亦明显大于A组;大鼠后肢功能Tarlv评分和电生理检查也有类似的变化趋势。结论 大鼠脊髓损伤后早期手术减压对损伤的大鼠脊髓轴索有保护作用,能减少脊髓损伤面积,并促进大鼠后肢功能恢复。     

完全横断性脊髓损伤后跨神经元变性的实验研究

目的：探讨大鼠完全性脊髓损伤后损伤平面以下下运动神经元轴突参与组成的周围神经的变化及其变化规律，为临床治疗、康复及预后的判断提供相关的理论基础。方法：48只成年雄性Wistar大鼠随机分为假手术组和脊髓损伤组，各组24只，每组均对应分为7d、1个月、2个月、3个月4个时间点，每组每个时间点6只大鼠。完全性脊髓损伤组大鼠制作T10水平完全性脊髓横断模型。在预定取材时间点处死相应动物，自腓总神经入肌点处向近端切取10mm该神经，分别行天青美-蓝染色光镜观察及超微结构观察。结果：各时间点假手术组腓总神经光镜下观察，神经纤维排列均匀；轴突外形正常、染色均匀。超微结构观察，髓鞘外形正常，板层清晰；轴索位置正常，轴索内微丝微管正常。完全性脊髓损伤组光镜观察发现腓总神经髓鞘和轴索出现明显退变，且退变随时间推移逐渐加重，同时出现轴突发芽现象。超微结构观察可见轴索内线粒体肿胀．轴浆内微丝、微管溶解，髓鞘板层结构破坏，雪旺氏细胞增生及空泡变性现象。退变的同时．腓总神经出现大量的新生神经纤维即轴突发芽现象。结论：完全性横断性脊髓损伤后周围神经存在退变现象．且退变程度随时间的延长逐渐加重，说明大鼠完全横断性脊髓损伤可以导致损伤水平以下周围神经发生跨神经元变性。     

The role of transcranial motor evoked potentials in predicting neurologic and histopathologic outcome after experimental spinal cord ischemia

BACKGROUND: Monitoring of myogenic motor evoked potentials to transcranial stimulation (tcMEPs) is clinically used to assess motor pathway function during aortic and spinal procedures that carry a risk of spinal cord ischemia (SCI). Although tcMEPs presumably detect SCI before irreversible neuronal deficit occurs, and prolonged reduction of tcMEP signals is thought to be associated with impending spinal cord damage, experimental evidence to support this concept has not been provided. In this study, histopathologic and neurologic outcome was examined in a porcine model of SCI after different durations of intraoperative loss of tcMEP signals. METHODS: In 15 ketamine-sufentanil-anesthetized pigs (weight, 35-45 kg) the spinal cord feeding lumbar arteries were exposed. tcMEP were recorded from the upper and lower limbs. Under normothermic conditions, animals were randomly allocated to undergo short-term tcMEP reduction (group A, < 10 min, n = 5) or prolonged tcMEP reduction (group B, 60 min, n = 10), resulting from temporary or permanent clamping of lumbar segmental arteries. Neurologic function was evaluated every 24 h, and infarction volume and the number of eosinophilic neurons and viable motoneurons in the lumbosacral spinal cord was evaluated 72 h after induction of SCI. RESULTS: In all animals except one, segmental artery clamping reduced tcMEP to below 25% of baseline. All but one animal in group A had reduced tcMEP for less than 10 min and had normal motor function and no infarction at 72 h after the initial tcMEP reduction. Seven animals in group B (70%) had reduced tcMEP signals for more than 60 min and were paraplegic with massive spinal cord infarction at 72 h. Two animals (one in both groups) had tcMEP loss for 40 min, with moderate infarction and normal function. In general, histopathologic damage and neurologic dysfunction did not occur when tcMEP amplitude recovered within 10 and 40 min after the initial decline, respectively. CONCLUSION: Prolonged reduction of intraoperative tcMEP amplitude is predictive for postoperative neurologic dysfunction, while recovery of the tcMEP signal within 10 min after the initial decline corresponds with normal histopathology and motor function in this experimental model. This finding confirms that intraoperative tcMEPs have a good prognostic value for neurologic outcome during procedures in which the spinal cord is at risk for ischemia.