大鼠脊髓损伤后挫伤部位的X线照射治疗对运动功能恢复的作用研究

目的 探讨脊髓挫伤部位的X线照射治疗对脊髓损伤后运动功能恢复的作用.方法 将70只雌性Wistar大鼠按照纽约大学的重力冲击方法建立大鼠脊髓(T10)损伤动物模型,按照随机数字表法分为7组,每组10只,其中6组大鼠在损伤后不同时间(损伤后20 min、1 d、2 d、4 d、7 d、17 d)对挫伤部位进行X线(20 Gy)照射,第7组则不予照射(对照组).然后根据Basso、Beattie、Bresnahan(BBB)评分标准评测各组大鼠运动功能恢复情况,并进行统计学比较.采用快蓝染色对存活6周以上的大鼠进行挫伤脊髓的组织形态学观察.结果脊髓挫伤后20 min、1 d、2 d行X线照射组BBB评分明显高于对照组,差异均有统计学意义(P<0.05),且在脊髓损伤后的2～3周进展较快,后期恢复缓慢.组织形态学观察可见应用X线治疗组周边组织残存区面积大于对照组.结论 脊髓挫伤部位伤后早期行X线照射治疗可保护脊髓残存的神经组织,改善运动功能的恢复.     

慢性渐进性压迫对大鼠行为功能及脊髓运动神经递质表达的影响

目的：观察脊髓慢性受压后实验动物的行为功能与运动神经递质表达的变化情况。方法：采用大鼠后路渐进性脊髓压迫动物模型，观察联合行为评分（CBS），常规病理及免疫组化检测胆碱乙酰转移酶（ChAT)的变化。结果：免疫组织化学染色显示，在正常大鼠脊髓前角运动神经元及大小神经元ChAT均表达阳性，脊髓损伤后ChAT阳性细胞数减少，CBS升高，二者具有相关关系。结论：脊髓受压抑制大鼠脊髓神经元合成ChAT，从而影响实验动物的行为功能。     

驽药针刺对大鼠脊髓损伤后运动功能及BDNF表达的影响

目的研究驽药针刺在大鼠脊髓损伤后运动功能变化以及BDNF表达的变化。方法采用脊髓半横断损伤模型。100只SD大鼠随机分为对照组、假手术组、脊髓损伤组、单纯针刺组、驽药针刺组,每组分为3天、7天、14天、21天共4个亚组,每组5只。BBB法评定大鼠后肢运动功能变化,免疫组化法检测大鼠脊髓中BDNF的表达变化。结果 BBB评分显示驽药针刺组的各时间点评分均高于脊髓损伤组(P0.05),驽药针刺组7、14、21d的BDNF表达均高于脊髓损伤组(P0.05),且与BBB评分呈正相关(r=0.717,P0.05)。结论驽药针刺可明显改善脊髓损伤大鼠的运动功能,并可明显促进大鼠脊髓损伤后BDNF的表达。     

移植分泌神经营养素3的人胚神经干细胞在脊髓损伤大鼠体内存活及对运动功能的影响

BACKGROUND： Many methods have been attempted to repair nerves following spinal cord injury, including peripheral nerve transplantation, Schwann cell transplantation, olfactory ensheathing cell transplantation, and embryonic neural tissue transplantation. However, there is a need for improved outcomes.
OBJECTIVE： To investigate the repair feasibility for rat spinal cord injury using human neural stem cells （hNSCs） genetically modified by lentivirus to express neurotrophin-3.
DESIGN, TIME AND SETTING： In vitro cell biological experiment and in vivo randomized, controlled genetic engineering experiment were performed at the Third Military Medical University of Chinese PLA and First People＇s Hospital of Yibin, China from March 2006 to December 2007.
MATERIALS： A total of 64 adult, female, Wistar rats were used for the in vivo study. Of them, 48 rats were used to establish models of spinal cord hemisection, and were subsequently equally and randomly assigned to model, genetically modified hNSC, and normal hNSC groups. The remaining 16 rats served as normal controls.
METHODS： hNSCs were in vitro genetically modified by lentivirus to secrete both green fluorescence protein and neurotrophin-3. Neurotrophin-3 expression was measured by Western blot. Genetically modified hNSC or normal hNSC suspension （5 × 10^5） was injected into the rat spinal cord following T10 spinal cord hemisection. A total of 5μL Dulbecco＇s-modified Eagle＇s medium was infused into the rat spinal cord in the model grop. Transgene expression and survival of transplanted hNSCs were determined by immunohistochemistry. Motor function was evaluated using the Basso, Beattie, and Bresnahan （BBB） scale.
MAIN OUTCOME MEASURES： The following parameters were measured： expression of neurotrophin-3 produced by genetically modified hNSCs, transgene expression and survival of hNSCs in rats, motor function in rats.
RESULTS： hNSCs were successfully genetically modified by lentivirus to stably express neurotrophin-3. The transplanted hNSCs primarily gathered at, or around, the injection site two weeks following transplantation, and gradually migrated towards the surrounding tissue. Transplanted hNSCs were observed 7.0-8.0 mm away from the injection site. In addition, hNSCs were observed 10 weeks after transplantation. At week 4, BBB locomotor scores were significantly greater in the genetically modified hNSC and normal hNSC groups, compared with the model group （P 〈 0.05）, and scores were significantly greater in the genetically modified hNSC group compared with the normal hNSC group （P 〈 0.05）.
CONCLUSION： hNSCs were genetically modified with lentivirus to stably secrete neurotrophin-3. hNSCs improved motor function recovery in rats following spinal cord injury.     

人脐带间充质干细胞移植对大鼠脊髓损伤神经功能恢复的评价

目的 观察人脐带间充质干细胞（human umbilical cordmesenchymal stem cell，hUCMSC）移植对大鼠脊髓损伤神经功能恢复的影响。方法 SD大鼠70只，随机分为3组：脊髓半切＋hUCMSC组（n=30）、脊髓半切＋PBS组（n=30）和假手术组（n=10）。脊髓半切＋hUCMSC组和PBS组又分为头侧注射、尾侧注射和头尾两侧注射三个亚组。移植后1、7、14、21、28d观察大鼠神经功能恢复情况，应用免疫组化检测移植到脊髓的hUCMSC胶质纤维酸性蛋白（GFAP）和神经元特异性烯醇化酶（NSE）表达情况。结果 大鼠脊髓半切损害后，hUCMSC组动物较PBS组有明显的神经功能恢复。植入后28d在宿主脊髓中存活的hUCMSC细胞MABl281（mouse antiuman nuclei monoclonal antibody）染色阳性，免疫组化双标染色显示MABl28l阳性细胞亦分别有NSE或GFAP表达并向损伤部位迁移，hUCMSC来源的GFAP阳性细胞可见明显的树突生长。结论 hUCMSC移植到宿主损伤脊髓后可以存活、向损伤部位迁移，并向神经元样和星形胶质细胞分化，且可促进大鼠脊髓损伤后神经功能恢复。hUCMSC作为一种来源广泛的干细胞用于治疗脊髓损伤可能具有重要的价值。     

Delayed treatment with Rho-kinase inhibitor does not enhance axonal regeneration or functional recovery after spinal cord injury in rats

Axonal regeneration in the central nervous system is blocked by many different growth inhibitory factors. Some of these inhibitors act on neurons by activating RhoA and Rho-kinase, an effector of RhoA. Several studies have shown that Rho-kinase inhibition immediately after spinal cord injury enhances axonal sprouting and functional recovery. In this study, we ask whether delayed treatment with Rho-kinase inhibitor is effective in promoting regeneration and functional recovery. We administered Fasudil, a Rho-kinase inhibitor, locally to the injury site 4 weeks or immediately after contusion of the thoracic spinal cord in rats. Although the immediate treatment significantly stimulated axonal sprouting and recovery of hindlimb function, treatment started 4 weeks after surgery had no effect on fiber sprouting or locomotor recovery. Our findings suggest that RhoA/Rho-kinase alone may not account for the irreversible arrest of axon outgrowth in the chronic stage of injury in the central nervous system.     

骨髓间质干细胞移植对大鼠脊髓损伤神经功能恢复的影响

目的：观察成人骨髓间质干细胞（hBMSCs)移植对大鼠脊髓损伤神经功能恢复的影响.方法：Wistar大鼠90只，随机分为脊髓半切＋hBMSCs组、脊髓半切＋PBS组、单纯脊髓半切组和假手术组。脊髓半切＋hBMSCs组和PBS组又分别分为头侧注射、尾侧注射和头尾两侧注射三个亚组。移植后1、7、14、21、28d观察大鼠神经功能恢复情况，应用免疫组化和免疫荧光技术检测BrdU标记hBMSCs的胶质纤维酸性蛋白（GFAP）和神经元特异性核蛋白（NeuN)表达情况。结果：大鼠脊髓半切损害后，hBMSCs组动物较PBS组死亡率下降并有明显的神经功能恢复。移植的hBMSCs 在宿主脊髓中存活，从第7天开始即有NeuN和GFAP表达并向损伤部位及对侧迁移，第28天hBMSCs来源GFAP阳性细胞可见明显的树突生长。结论：hBMSCs可在宿主损伤脊髓中存活、向损伤部位迁移并向神经元和星形胶质细胞分化，并促进神经功能恢复，降低死亡率，成人骨髓间质干细胞作为一种独特的干细胞来源用于治疗脊髓损伤可能具有非常重要的价值。     

胚胎脊髓移植在恢复损伤脊髓传导功能中的作用

目的：探讨胚胎脊髓移植在恢复损伤脊髓传导功能中的作用。方法：将Ｅ１４胚胎脊髓植入成鼠损伤脊髓后３０、４５、６０天时，用单位放电记录技术观察了正常脊髓神经元和移植物神经元的自发放电活动，及其对刺激坐骨神经、红核和同时刺激的反应。结果：正常脊髓神经元的自发单位放电多是一个低频的单发脉冲活动。无论选择那种刺激方式，都可见兴奋、抑制和无反应三种反应。术后３０天时，胚胎神经元的自发单位放电以高频电脉冲活动为主，簇状放电所占比例较大，对刺激有反应的放电单位数也较少；随着动物存活时间的延长，这些单位放电的情况逐渐向着低频、单脉冲以及高反应率的方向发展。至术后６０天时，胚胎神经元单位放电的频率、形式以及对刺激的反应情况都变得和正常神经元的相似。结论：胚胎神经元移植后经历了一个逐渐发育分化过程，在这个过程中它们有可能逐渐和宿主神经元形成了功能性突触连接。     

A noninvasive ultrasonographic method to evaluate bladder function recovery in spinal cord injured rats

Suprasacral spinal cord injury induces changes in the mechanical and neuronal properties of the bladder resulting in bladder areflexia followed by bladder-sphincter dyssynergia and detrusor muscle hypertrophy, which lead to urinary retention and increased bladder size. These changes are most often quantified using highly skilled urodynamic techniques that involve catheterization. We investigated whether a hand-held digital ultrasound imaging system could monitor urinary retention in the bladder following spinal cord injury in adult rats. Our findings indicate that contusive spinal cord injury resulted in high residual bladder volumes that decreased and stabilized by 2 weeks post-injury but remained significantly higher than control bladder volumes up to 46 days post-injury (the longest time point examined). Post hoc analysis indicated that the degree of bladder function recovery recorded at the end of the study correlated with the degree of bladder function recovery recorded at 6 days post-injury, indicating that bladder function recovery can be predicted by analyzing bladder volume as early as 6 days post-injury. Bladder function recovery correlated with locomotor recovery as assessed using the BBB locomotor rating scale. While providing a noninvasive assessment of bladder function with no detrimental impact on locomotor function or assessment, this protocol provides researchers with a clinically relevant outcome measure for quantifying bladder function recovery after spinal cord injury or after experimental treatments for spinal cord injury.     

以NgR为靶点治疗脊髓损伤的研究进展

NgR是最近发现并克隆的一种糖基磷脂酰肌醇（GPI）锚定膜蛋白，因其在髓磷脂抑制轴突再生信号转导过程中特殊的靶分子效应，日益受到重视。以NgR为靶点促进脊髓轴突再生的尝试为中枢神经系统（CNS）损伤的治疗提供了有益的启示。     

Immunization with recombinant Nogo-66 receptor (NgR) promotes axonal regeneration and recovery of function after spinal cord injury in rats

Nogo-66 receptor (NgR), a common receptor for the three known myelin-associated inhibitors, i.e., Nogo-A, myelin-associated glycoprotein (MAG), and oligodendrocyte myelin glycoprotein (OMgp), plays a key role in the failure of axonal regeneration in the adult mammalian central nervous system (CNS). Here we report a novel vaccine approach that stimulates the production of anti-NgR antibody to overcome NgR-mediated growth inhibition after spinal cord injury (SCI). We showed that adult rats immunized with recombinant NgR produced high titers of the anti-NgR antibody and that antisera obtained from the immunized rats promoted neurite outgrowth of rat cerebellar neurons on the inhibitory MAG substrate in vitro. In a spinal cord dorsal hemisection model, NgR immunization promoted regeneration of lesioned corticospinal tract (CST) axons, anterogradely labeled with biotin dextran amine (BDA), beyond the lesion site. In a contusive SCI model, NgR immunization markedly reduced the total lesion volume and improved Basso, Beattie, and Bresnahan (BBB) locomotor rating scale and grid walking performance. Thus, the NgR vaccine approach may represent a promising repair strategy to promote structural and functional recovery following SCI.     

Methylprednisolone fails to improve functional and histological outcome following spinal cord injury in rats

Currently, methylprednisolone sodium succinate (MPSS) is the standard treatment following acute spinal cord injury (SCI) as a consequence of the results obtained from the National Acute Spinal Cord Injury Studies. However, many have questioned the efficacy of MPSS because of its marginal effects. Additionally there has been criticism of both study design and statistical interpretation. The functional consequences of experimental SCI have been assessed in many ways. The purpose of this investigation was to determine the effects of MPSS vs. saline solution (SS) following moderate T10 contusion injury in rat. Functional recovery was evaluated using the 21-point Basso, Beattie and Bresnahan (BBB) locomotor recovery scale, the inclined plane, the beam walk, footprint analysis and the horizontal ladder. To optimize the precision and accuracy of functional results we examined the locomotion on a treadmill using three-dimensional (3D) analysis. Stereology was used to estimate the amount of damaged tissue. The results of the traditional functional methods showed that administration of the NASCIS dosage of MPSS following acute spinal cord contusion did not lead to any significant differences in the functional recovery of MPSS- vs. SS-treated animals. More importantly, the results of the 3D kinematic showed that the MPSS administration did not affect the flexion/extension of the hip, knee and ankle joints during the step cycle. Finally, stereological results revealed no statistically significant differences between the two experimental groups. Altogether, our results support data previously reported by several authors, suggesting that MPSS does not lead to improved functional outcome following experimental acute SCI.     

Leuprolide acetate induces structural and functional recovery of injured spinal cord in rats

Gonadotropin-releasing hormone (GnRH) and its synthetic analog leuprolide acetate, a GnRH agonist, have neurotrophic properties. This study was designed to determine whether administration of leuprolide acetate can improve locomotor behavior, gait, micturition reflex, spinal cord morphology and the amount of microglia in the lesion epicenter after spinal cord injury in rats. Rats with spinal cord compression injury were administered leuprolide acetate or saline solution for 5 weeks. At the 5^th week, leuprolide acetate-treated rats showed locomotor activity recovery by 38%, had improvement in kinematic gait and exhibited voiding reflex recovery by 60%, as compared with the 1^st week. By contrast, saline solution-treated rats showed locomotor activity recovery only by 7%, but voiding reflex did not recover. More importantly, leuprolide acetate treatment reduced microglial immunological reaction and induced a trend towards greater area of white and gray matter in the spinal cord. Therefore, leuprolide acetate has great potential to repair spinal cord injury.