GDNF 缓释微球及NogoA,ChABC 缓释微球联合促大鼠损伤脊髓再生修复的实验研究

The purpose of this study was to evaluate the effect of poly(lactide-co-glycolic acid) delayed-release microspheres,which were prepared using glial cell line-derived neurotrophic factor(GDNF),on the delayed-release,controllability,and protection of GDNF activity.The present study is the first to combine chondroitinase ABC,GDNF,and Nogo A antibody delayed-release microspheres for the treatment of spinal cord injury.Results show that the combined therapy of chondroitinase ABC,GDNF,and Nogo A antibody microspheres can increase the immunoreaction of neurofilament 200 in the injured spinal cord,and this therapeutic effect was better than chondroitinase ABC,GDNF,or Nogo A antibody microspheres administered singularly.     

Enteric glia mediate neuronal outgrowth through release of neurotrophic factors

Previous studies have shown that transplanted enteric glia enhance axonal regeneration, reduce tissue damage, and promote functional recovery following spinal cord injury. However, the mechanisms by which enteric glia mediate these beneficial effects are unknown. Neurotrophic factors can promote neuronal differentiation, survival and neurite extension. We hypothesized that enteric glia may exert their protective effects against spinal cord injury partially through the secretion of neurotrophic factors. In the present study, we demonstrated that primary enteric glia cells release nerve growth factor, brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor over time with their concentrations reaching approximately 250, 100 and 50 pg/mL of culture medium respectively after 48 hours. The biological relevance of this secretion was assessed by incubating dissociated dorsal root ganglion neuronal cultures in enteric glia-conditioned medium with and/or without neutralizing antibodies to each of these proteins and evaluating the differences in neurite growth. We discovered that conditioned medium enhances neurite outgrowth in dorsal root ganglion neurons. Even though there was no detectable amount of neurotrophin-3 secretion using ELISA analysis, the neurite outgrowth effect can be attenuated by the antibody-mediated neutralization of each of the aforementioned neurotrophic factors. Therefore, enteric glia secrete nerve growth factor, brain-derived neurotrophic factor, glial cell line-derived neurotrophic factor and neurotrophin-3 into their surrounding environment in concentrations that can cause a biological effect.     

Secretion of nerve growth factor,brain-derived neurotrophic factor,and glial cell-line derived neurotrophic factor in co-culture of four cell types in cerebrospinal fluid-containing medium

The present study co-cultured human embryonic olfactory ensheathing cells, human Schwann cells, human amniotic epithelial cells and human vascular endothelial cells in complete culture medium- containing cerebrospinal fluid. Enzyme linked immunosorbent assay was used to detect nerve growth factor, brain-derived neurotrophic factor, and glial cell line-derived neurotrophic factor secretion in the supernatant of co-cultured cells. Results showed that the number of all cell types reached a peak at 7-10 days, and the expression of nerve growth factor, brain-derived neurotrophic factor, and glial cell line-derived neurotrophic factor peaked at 9 days. Levels of secreted nerve growth factor were four-fold higher than brain-derived neurotrophic factor, which was three-fold higher than glial cell line-derived neurotrophic factor. Increasing concentrations of cerebrospinal fluid (10%, 20% and 30%) in the growth medium caused a decrease of neurotrophic factor secretion Results indicated co-culture of human embryonic olfactory ensheathing cells, human Schwann cells human amniotic epithelial cells and human vascular endothelial cells improved the expression of nerve growth factor, brain-derived neurotrophic factor, and glial cell line-derived neurotrophic factor. The reduction of cerebrospinal fluid extravasation at the transplant site after spinal cord injury is beneficial for the survival and secretion of neurotrophic factors from transplanted cells.     

Combination of epidural electrical stimulation with ex vivo triple gene therapy for spinal cord injury:a proof of principle study

Despite emerging contemporary biotechnological methods such as gene-and stem cell-based therapy,there are no clinically established therapeutic strategies for neural regeneration after spinal cord injury.Our previous studies have demonstrated that transplantation of genetically engineered human umbilical cord blood mononuclear cells producing three recombinant therapeutic molecules,including vascular endothelial growth factor(VEGF),glial cell-line derived neurotrophic factor(GDNF),and neural cell adhesion molecule(NCAM) can improve morpho-functional recovery of injured spinal cord in rats and mini-pigs.To investigate the efficacy of human umbilical cord blood mononuclear cells-mediated triple-gene therapy combined with epidural electrical stimulation in the treatment of spinal cord injury,in this study,rats with moderate spinal cord contusion injury were intrathecally infused with human umbilical cord blood mononuclear cells expressing recombinant genes VEGF165,GDNF,NCAM1 at 4 hours after spinal cord injury.Three days after injury,epidural stimulations were given simultaneously above the lesion site at C5(to stimulate the cervical network related to forelimb functions) and below the lesion site at L2(to activate the central pattern generators) every other day for 4 weeks.Rats subjected to the combined treatment showed a limited functional improvement of the knee joint,high preservation of muscle fiber area in tibialis anterior muscle and increased H/M ratio in gastrocnemius muscle 30 days after spinal cord injury.However,beneficial cellular outcomes such as reduced apoptosis and increased sparing of the gray and white matters,and enhanced expression of heat shock and synaptic proteins were found in rats with spinal cord injury subjected to the combined epidural electrical stimulation with gene therapy.This study presents the first proof of principle study of combination of the multisite epidural electrical stimulation with ex vivo triple gene therapy(VEGF,GDNF and NCAM) for treatment of spinal cord injury in rat models.The animal protocols were approved by the Kazan State Medical University Animal Care and Use Committee(approval No.2.20.02.18) on February 20,2018.     

NT-3 promotes growth of lesioned adult rat sensory axons ascending in the dorsal columns of the spinal cord

The regeneration capacity of spinal cord axons is severely limited. Recently, much attention has focused on promoting regeneration of descending spinal cord pathways, but little is known about the regenerative capacity of ascending axons. Here we have assessed the ability of neurotrophic factors to promote regeneration of sensory neurons whose central axons ascend in the dorsal columns. The dorsal columns of adult rats were crushed and either brain-derived neurotrophic factor (BDNF), glial cell line-derived neurotrophic factor (GDNF), neurotrophin-3 (NT-3) or a vehicle solution was delivered continuously to the lesion site for 4 weeks. Transganglionic labelling with cholera toxin beta subunit (CTB) was used to selectively label large myelinated Abeta fibres. In lesioned rats treated with vehicle, CTB-labelled fibres were observed ascending in the gracile fasciculus, but these stopped abruptly at the lesion site, with no evidence of sprouting or growth into lesioned tissue. No CTB-labelled terminals were observed in the gracile nucleus, indicating that the lesion successfully severed all ascending dorsal column axons. Treatment with BDNF did not promote axonal regeneration. In GDNF-treated rats fibres grew around cavities in caudal degenerated tissue but did not approach the lesion epicentre. NT-3, in contrast, had a striking effect on promoting growth of lesioned dorsal column axons with an abundance of fibre sprouting apparent at the lesion site, and many fibres extending into and beyond the lesion epicentre. Quantification of fibre growth confirmed that only in NT-3-treated rats did fibres grow into the crush site and beyond. No evidence of terminal staining in the gracile nucleus was apparent following any treatment. Thus, although NT-3 promotes extensive growth of lesioned axons, other factors may be required for complete regeneration of these long ascending projections back to the dorsal column nuclei. The intrathecal delivery of NT-3 or other neurotrophic molecules has obvious advantages in clinical applications, as we show for the first time that dorsal column axonal regeneration can be achieved without the use of graft implantation or nerve lesions.     

Transplantation of bone marrow mesenchymal stem cells reduces lesion volume and induces axonal regrowth of injured spinal cord

It has been demonstrated that transplantation of bone marrow mesenchymal stem cells (BMSCs) improves recovery of injured spinal cord in animal models. However, the mechanism of how BMSCs promote repair of injured spinal cord remains under investigation. The present study investigated the neural differentiation of BMSCs, the lesion volume and axonal regrowth of injured spinal cord after transplantation. Seven days after spinal cord injury, 3 × 10⁵ BMSCs or PBS (control) was delivered into the injury epicenter of the spinal cord. At 8 weeks after spinal cord injury, transplantation of BMSCs reduced the volume of cavity and increased spared white matter as compared to the control. BMSCs did not express the cell marker of neurons, astrocytes and oligodendrocytes in injured spinal cord. Transmission electron microscopic examination displayed an increase in the number of axons in BMSC rats. The effect of BMSCs on growth of neuronal process was further investigated by using a coculture system. The length and the number of neurites from spinal neurons significantly increased when they cocultured with BMSCs. PCR and immunochemical analysis showed that BMSCs expressed brain‐derived neurotrophic factor (BDNF) and glia cell line‐derived neurotrophic factor (GDNF). These findings demonstrate that transplantation of BMSCs reduces lesion volume and promotes axonal regrowth of injured spinal cord.     

Gene transfer of glial cell line-derived neurotrophic factor promotes functional recovery following spinal cord contusion

Neuronal cell death and the failure of axonal regeneration cause a permanent functional deficit following spinal cord injury (SCI). Administration of recombinant glial cell line-derived neurotrophic factor (GDNF) has previously been reported to rescue neurons following severe SCI, resulting in improved hindlimb locomotion in rats. In this study, thus, GDNF gene therapy using an adenoviral vector (rAd-GDNF) was examined in rats following SCI induced by dropping the NYU weight-drop impactor from a height of 25 mm onto spinal segment T9-T10. To evaluate the efficacy of intraspinal injection of recombinant adenovirus into the injured spinal cord, we observed green fluorescent protein (GFP) gene transfer in the contused spinal cord. GFP was effectively expressed in the injured spinal cord, and the most prominently transduced cells were astrocytes. The expression of GDNF was detected only in rats receiving rAd-GDNF, not the controls, and remained detectable around the injured site for at least 8 days. Open-field locomotion analysis revealed that rats receiving rAd-GDNF exhibited improved locomotor function and hindlimb weight support compared to the control groups. Immunohistochemical examination for the neuronal marker, calcitonin gene-related peptide (CGRP), showed an increase in CGRP+ neuronal fibers in the injured spinal cord in rats receiving rAd-GDNF treatment. Collectively, the results suggest that adenoviral gene transfer of GDNF can preserve neuronal fibers and promote hindlimb locomotor recovery from spinal cord contusion. This research should provide information for developing a clinical strategy for GDNF gene therapy.     

BDNF-GFP转染神经干细胞修复大鼠脊髓损伤：发挥干细胞与神经因子的双重效应？

摘要 背景：神经干细胞移植入大鼠脊髓损伤模型可以促进功能恢复,基因治疗已被广泛用于治疗脊髓损伤。 目的：确定BDNF-GFP转染后神经干细胞移植对大鼠脊髓损伤的修复效果。 设计,时间和背景：本实验是在中国医科大学基础医学院发育生物学实验室与2009年5月至2010年1月完成。 材料：10只新生Wistar大鼠和88只2-3个月大,雌雄不限的Wistar大鼠。 方法：以携带BDNF-GFP基因的腺病毒转染神经干细胞。88只Wistar大鼠中假手术组8只, 80只大鼠制成T9左侧横断模型,并随机分成四组：BDNF和GFP修饰的神经干细胞移植组,GFP修饰的神经干细胞移植组;单纯神经干细胞移植组和模型组。在各神经干细胞移植组,脊髓损伤后向横断处显微注射等体积细胞,模型组在相同的部位注射等体积的PBS。 主要观察指标： BBB评分检测脊髓损伤模型运动功能恢复情况;制备脊髓损伤模型2周后取材,免疫组化评估BDNF-GFP转染的神经干细胞移植后的细胞学特点;制备脊髓损伤模型2、4、6、8周Real-time PCR检测脊髓横断处BDNF表达情况。 结果： BDNF-GFP转染后神经干细胞在脊髓半切模型中存活并表达BDNF和GFP,移植该细胞后的大鼠体内高表达具有生物活性的BDNF,且脊髓损伤动物运动功能较对照组明显恢复。 结论：移植BDNF-GFP转染后神经干细胞可能是一种修复脊髓损伤的有效的方法。 关键词：神经干细胞,脑源性神经营养因子;绿色荧光蛋白;脊髓损伤;移植。     

骨髓间充质干细胞立体定向移植治疗大鼠脊髓损伤*

摘要 背景：传统观念认为,神经组织损伤后几乎不能再生,以往对SCI的治疗缺乏有效手段,致使本病致残率高,疗效差。干细胞治疗关键在于移植具有再生能力的干细胞,通过多种作用机制,可以重建中枢神经系统的结构和功能,近年来引起了广泛的关注。 目的：探讨立体定向移植骨髓间充质干细胞（MSCs）对大鼠脊髓损伤修复的影响并探讨其机制 设计、时间及地点：随机对照动物实验,于2007-10/2008-6在天津市环湖医院完成。 材料：1月龄SD大鼠20只,用于制备骨髓间充质干细胞;健康成年Wistar大鼠45只,雌性、同系,体质量280±20 g。将动物随机分为对照组、假手术组与移植组,每组各15只。 方法：密度梯度离心法结合贴壁筛选法分离骨髓间充质干细胞,经流式细胞仪鉴定为MSCs。以动脉瘤夹夹闭法制备大鼠脊髓损伤（SCI）模型,在SCI大鼠致伤后第7天,通过立体定向途径移植MSCs到移植组大鼠脊髓损伤中心,移植等量生理盐水至假手术组大鼠脊髓损伤中心,对照组大鼠不做处理。 主要观察指标：SCI大鼠损伤前及损伤后第7天、14天、30天、60天、90天的BBB评分;损伤后第90天处死大鼠,观察其脊髓组织中有无BrdU阳性细胞、Brdu+NSE、Brdu+GFAP、Brdu+bFGF、Brdu+BDNF免疫组化双染阳性细胞并观察NSE、GFAP、bFGF、BDNF单染阳性细胞。 结果： ①BBB评分发现,MSCs移植组大鼠BBB后肢功能评分恢复优于对照组（p<0.05）;假手术组BBB评分在损伤后30天内恢复速度慢于对照组（p<0.05）,至第90天与对照组比较无显著差异（P>0.05）;②免疫组织化学染色发现,移植组大鼠脊髓内在损伤中心及头、尾端距离脊髓损伤中心1cm处均可见BrdU染色阳性细胞及Brdu+NSE、Brdu+GFAP、Brdu+bFGF、Brdu+BDNF免疫组化双染阳性细胞。移植组NSE、GFAP、bFGF、BDNF单染阳性细胞数明显高于对照组和假手术组（p<0.05）。 结论： MSCs移植可以促进SCI大鼠的神经功能的恢复,其机制可能与移植细胞分化为神经元样和神经胶质细胞样细胞,并分泌或促进宿主分泌神经营养因子有关。 关键词 脊髓损伤 骨髓间充质干细胞 立体定向 细胞移植     

Combined transplantation of GDAsBMP and hr-decorin in spinal cord contusion repair****○

Following spinal cord injury, astrocyte proliferation and scar formation are the main factors inhibiting the regeneration and growth of spinal cord axons. Recombinant decorin suppresses inflammatory reactions, inhibits glial scar formation, and promotes axonal growth. Rat models of T8 spinal cord contusion were created with the NYU impactor and these models were subjected to combined transplantation of bone morphogenetic protein-4-induced glial-restricted precursor-derived astrocytes and human recombinant decorin transplantation. At 28 days after spinal cord contusion, double-immunofluorescent histochemistry revealed that combined transplantation inhibited the early inflammatory response in injured rats. Furthermore, brain-derived neurotrophic factor, which was secreted by transplanted cells, protected injured axons. The combined transplantation promoted axonal regeneration and growth of injured motor and sensory neurons by inhibiting astrocyte proliferation and glial scar formation, with astrocytes forming a linear arrangement in the contused spinal cord, thus providing axonal regeneration channels.     

Transfection of the glial cell line-derived neurotrophic factor gene promotes neuronal differentiation简

Glial cell line-derived neurotrophic factor recombinant adenovirus vector-transfected bone marrow mesenchymal stem cells were induced to differentiate into neuron-like cells using inductive medium containing retinoic acid and epidermal growth factor. Cell viability, micro- tubule-associated protein 2-positive cell ratio, and the expression levels of glial cell line-derived neurotrophic factor, nerve growth factor and growth-associated protein-43 protein in the su- pernatant were significantly higher in glial cell line-derived neurotrophic factor/bone marrow mesenchymal stem cells compared with empty virus plasmid-transfected bone marrow mes- enchymal stem cells. Furthermore, microtubule-associated protein 2, glial cell line-derived neurotrophic factor, nerve growth factor and growth-associated protein743 mRNA levels in cell pellets were statistically higher in glial cell line-derived neurotrophic factor/bone marrow mesen- chymal stem cells compared with empty virus plasmid-transfected bone marrow mesenchymal stem cells. These results suggest that glial cell line-derived neurotrophic factor/bone marrow mesenchymal stem cells have a higher rate of induction into neuron-like cells, and this enhanced differentiation into neuron-like cells may be associated with up-regulated expression of glial cell line-derived neurotrophic factor, nerve growth factor and growth-associated protein-43.     

Neuroprotection of glial cell line-derived neurotrophic factor in damaged spinal cords following contusive injury

Glial cell line-derived neurotrophic factor (GDNF) acts as a potent survival factor for many neuronal populations, including spinal motoneurons, indicating the therapeutic promise of GDNF for neurological disorders. Injury to spinal cord (SCI) triggers processes destructive to ascending sensory and descending motor conduction and extends tissue loss, thereby leading to permanent behavioral dysfunction. In this study, we attempted to examine whether GDNF protects neurons from SCI and subsequently lessens locomotor deficit in SCI rats. We utilized the NYU weight-drop device developed at New York University to induce spinal cord contusion at the T9-10 spinal segment. After SCI, GDNF was administrated into the cord 1-2 mm rostral and caudal to the epicenter. Animals receiving GDNF treatment showed significant improvement over phosphate-buffered saline (PBS)-treated controls on the Basso Beattie Bresnahan (BBB) locomotor rating scale (P < 0.01-0.001). GDNF treatment increased the remaining neuronal fibers with calcitonin gene-related peptide, neurofilament, and growth-associated protein 43 immunoreactivity in injured spinal tissues compared with PBS-treated controls. Moreover, treatment with GDNF caused approximately 50% cell survival in the contused spinal cord tissues. Examination of signal transduction triggered by GDNF indicated that GDNF injection transiently induced activation of the mitogen-activated protein (MAP) kinase pathway in the spinal cord. Additionally, an up-regulation of anti-apoptotic Bcl-2 levels in the contusive center of the damaged spinal cord was observed 24 hr post-GDNF injection. Together our results show that GDNF exerts behavioral and anatomic neuroprotection following SCI. Additionally, GDNF-activated MAP kinase and Bcl-2 signaling may contribute to neuronal survival after spinal cord contusion.     

骨髓间充质干细胞尾静脉移植脊髓损伤大鼠脑源性神经营养因子及神经生长因子的表达

背景：骨髓间充质干细胞移植对脊髓损伤有治疗作用,但其机制尚不完全清楚。 目的：应用免疫组织化学方法观察骨髓间充质干细胞静脉移植损伤脊髓局部脑源性神经营养因子及神经生长因子的表达,分析骨髓间充质干细胞移植治疗大鼠脊髓损伤的作用途径。 方法：运用改良Allen法制备T10脊髓外伤性截瘫大鼠模型,假手术组6只,脊髓损伤组24只随机分为对照组和骨髓间充质干细胞移植组。骨髓间充质干细胞移植组、假手术组接受骨髓间充质干细胞单细胞悬液1 mL(1×106 cells)自大鼠尾静脉缓慢注射移植,对照组静脉注射PBS 1 mL。 结果与结论：脊髓损伤后损伤局部的脑源性神经营养因子、神经生长因子表达增加,骨髓间充质干细胞静脉注射移植后能促进脊髓损伤局部脑源性神经营养因子、神经生长因子更进一步的表达,这可能是促进神经结构及神经功能恢复的因素之一。     

神经干细胞移植修复鼠脊髓损伤的实验研究

目的 :观察神经干细胞移植治疗对鼠脊髓损伤后神经结构修复和功能恢复的作用并探讨其作用机制。方法 :制备鼠T10 脊髓损伤模型 ,体外培养、诱导鼠神经干细胞 ,定量评价神经干细胞移植对脊髓损伤后神经结构修复和功能恢复的影响。结果 :与对照组相比 ,神经干细胞移植组明显的增强了GAP 43mRNA的表达 ,促进了脊髓ChAT阳性的运动神经元的再生、结构的修复和下肢运动功能的恢复。结论 :神经干细胞移植促进了脊髓损伤后神经结构的修复和功能的恢复 ,是急性脊髓损伤一种有效的治疗方案     

Rescue and sprouting of motoneurons following ventral root avulsion and reimplantation combined with intraspinal adeno-associated viral vector-mediated expression of glial cell line-derived neurotrophic factor or brain-derived neurotrophic factor

Following avulsion of a spinal ventral root, motoneurons that project through the avulsed root are axotomized. Avulsion between, for example, L2 and L6 leads to denervation of hind limb muscles. Reimplantation of an avulsed root directed to the motoneuron pool resulted in re-ingrowth of some motor axons. However, most motoneurons display retrograde atrophy and subsequently die. Two neurotrophic factors, glial cell line-derived neurotrophic factor (GDNF) and brain-derived neurotrophic factor (BDNF), promote the survival of motoneurons after injury. The long-term delivery of these neurotrophic factors to the motoneurons in the ventral horn of the spinal cord is problematic. One strategy to improve the outcome of the neurosurgical reinsertion of the ventral root following avulsion would involve gene transfer with adeno-associated viral (AAV) vectors encoding these neurotrophic factors near the denervated motoneuron pool. Here, we show that AAV-mediated overexpression of GDNF and BDNF in the spinal cord persisted for at least 16 weeks. At both 1 and 4 months post-lesion AAV-BDNF- and -GDNF-treated animals showed an increased survival of motoneurons, the effect being more prominent at 1 month. AAV vector-mediated overexpression of neurotrophins also promoted the formation of a network of motoneuron fibers in the ventral horn at the avulsed side, but motoneurons failed to extent axons into the reinserted L4 root towards the sciatic nerve nor to improve functional recovery of the hind limbs. This suggests that high levels of neurotrophic factors in the ventral horn promote sprouting, but prevent directional growth of axons of a higher number of surviving motoneurons into the implanted root.     

Cellular GDNF delivery promotes growth of motor and dorsal column sensory axons after partial and complete spinal cord transections and induces remyelination

Glial cell line-derived neurotrophic factor (GDNF) is the prototypical member of a growth factor family that signals via the cognate receptors ret and GDNF-receptor alpha-1. The latter receptors are expressed on a variety of neurons that project into the spinal cord, including supraspinal neurons, dorsal root ganglia, and local neurons. Although effects of GDNF on neuronal survival in the brain have previously been reported, GDNF effects on injured axons of the adult spinal cord have not been investigated. Using an ex vivo gene delivery approach that provides both trophic support and a cellular substrate for axonal growth, we implanted primary fibroblasts genetically modified to secrete GDNF into complete and partial mid-thoracic spinal cord transection sites. Compared to recipients of control grafts expressing a reporter gene, GDNF-expressing grafts promoted significant regeneration of several spinal systems, including dorsal column sensory, regionally projecting propriospinal, and local motor axons. Local GDNF expression also induced Schwann cell migration to the lesion site, leading to remyelination of regenerating axons. Thus, GDNF exerts tropic effects on adult spinal axons and Schwann cells that contribute to axon growth after injury.     

Role of endogenous Schwann cells in tissue repair after spinal cord injury

Schwann cells are glial cells of peripheral nervous system, responsible for axonal myelination and ensheathing, as well as tissue repair following a peripheral nervous system injury. They are one of several cell types that are widely studied and most commonly used for cell transplantation to treat spinal cord injury, due to their intrinsic characteristics including the ability to secrete a variety of neurotrophic factors. This mini review summarizes the recent findings of endogenous Schwann cells after spinal cord injury and discusses their role in tissue repair and axonal regeneration. After spinal cord injury, numerous endogenous Schwann cells migrate into the lesion site from the nerve roots, involving in the construction of newly formed repaired tissue and axonal myelination. These invading Schwann cells also can move a long distance away from the injury site both rostrally and caudally. In addition, Schwann cells can be induced to migrate by minimal insults (such as scar ablation) within the spinal cord and integrate with astrocytes under certain circumstances. More importantly, the host Schwann cells can be induced to migrate into spinal cord by transplantation of different cell types, such as exogenous Schwann cells, olfactory ensheathing cells, and bone marrow-derived stromal stem cells. Migration of endogenous Schwann cells following spinal cord injury is a common natural phenomenon found both in animal and human, and the myelination by Schwann cells has been examined effective in signal conduction electrophysiologically. Therefore, if the inherent properties of endogenous Schwann cells could be developed and utilized, it would offer a new avenue for the restoration of injured spinal cord.     

甲基强的松龙和神经干细胞移植联合治疗大鼠脊髓损伤

目的：观察甲基强的松龙和神经干细胞移植对大鼠脊髓损伤后神经结构修复和功能恢复的治疗作用并探讨其作用机制。方法：制备大鼠胸10脊髓损伤模型，体外培养、诱导分化大鼠神经干细胞，定量评价甲基强的松龙和神经干细胞移植对脊髓损伤后神经结构修复和功能恢复的影响。结果：与对照组相比，移植组明显地增强了生长相关蛋白（GAP－43）mRNA的表达，促进了乙酰胆碱转移酶（ChAT)阳性脊髓运动神经元的再生、神经结构的修复和下肢运动功能的恢复（P＜0．05）。结论：甲基强的松龙和神经干细胞移植通过增强GAP－43 mRNA的表达、运动神经元的再生而促进了脊髓损伤后神经结构的修复和功能的恢复，是急性脊髓损伤的一种有效的治疗方案。     

Neurotrophic factors expressed in both cortex and spinal cord induce axonal plasticity after spinal cord injury

We reported recently that overexpression of neurotrophin-3 (NT-3) by motoneurons in the spinal cord of rats will induce sprouting of corticospinal tract (CST) axons (Zhou et al. [2003] J. Neurosci. 23:1424-1431). We now report that overexpression of brain-derived neurotrophic factor (BDNF) or glial cell-derived neurotrophic factor (GDNF) in the rat sensorimotor cortex near the CST neuronal cell bodies together with overexpression of NT-3 in the lumbar spinal cord significantly increases axonal sprouting compared to that induced by NT-3 alone. Two weeks after unilaterally lesioning the CST at the level of the pyramids, we injected rats with saline or adenoviral vectors (Adv) carrying genes coding for BDNF (Adv.BDNF), GDNF (Adv.GDNF) or enhanced green fluorescent protein (Adv.EGFP) at six sites in the sensorimotor cortex, while delivering Adv.NT3 to motoneurons in each of these four groups on the lesioned side of the spinal cord by retrograde transport from the sciatic nerve. Four days later, biotinylated dextran amine (BDA) was injected into the sensorimotor cortex on the unlesioned side to mark CST axons in the spinal cord. Morphometric analysis of axonal sprouting 3 weeks after BDA injection showed that the number of CST axons crossing the midline in rats treated with Adv.BDNF or Adv.GDNF were 46% and 52% greater, respectively, than in rats treated with Adv.EGFP or PBS (P < 0.05). These data demonstrate that sustained local expression of neurotrophic factors in the sensorimotor cortex and spinal cord will promote increased axonal sprouting after spinal cord injury, providing a basis for continued development of neurotrophic factor therapy for central nervous system damage.