Electroacupuncture promotes the proliferation of endogenous neural stem cells and oligodendrocytes in the injured spinal cord of adult rats

A contusive model of spinal cord injury at spinal segment T8-9 was established in rats. Huantiao (GB30) and Huatuojiaji (Ex-B05) were punctured with needles, and endogenous neural stem cells were labeled with 5-bromo-2’-deoxyuridine (BrdU) and NG2. Double immunofluorescence staining showed that electroacupuncture markedly increased the numbers of BrdU+/NG2+ cells at spinal cord tissue 15 mm away from the injury center in the rostral and caudal directions. The results suggest that electroacupuncture promotes the proliferation of endogenous neural stem cells and oligodendrocytes in rats with spinal cord injury.     

Differential generation of oligodendrocytes from human and rodent embryonic spinal cord neural precursors

Human neural precursors are considered to have widespread therapeutic possibilities on account of their ability to provide large numbers of cells whilst retaining multipotentiality. Application to human demyelinating diseases requires improved understanding of the signalling requirements underlying the generation of human oligodendrocytes from immature cell populations. In this study, we compare and contrast the capacity of neural precursors derived from the developing human and rodent spinal cord to generate oligodendrocytes. We show that the developing human spinal cord (6-12 weeks of gestation) displays a comparable ventrodorsal gradient of oligodendrocyte differentiation potential to the embryonic rodent spinal cord. In contrast, fibroblast growth factor 2 (FGF-2) expanded human neural precursors derived from both isolated ventral or dorsal cultures show a reduced capacity to generate oligodendrocytes, whereas comparable rodent cultures demonstrate a marked increase in oligodendrocyte formation following FGF-2 treatment. In addition, we provide evidence that candidate growth factors suggested from rodent studies, including FGF-2 and platelet-derived growth factor (PDGF) do not stimulate proliferation of human oligodendrocyte lineage cells. Finally, we show that the in vivo environment of the acutely demyelinating adult rat spinal cord is insufficient to stimulate the differentiation of immature human spinal cord cells to oligodendrocytes. These results provide further evidence for inter-species difference in the capacity of neural precursors to generate oligodendrocytes.     

The fate and function of oligodendrocyte progenitor cells after traumatic spinal cord injury

Oligodendrocyte progenitor cells (OPCs) are the most proliferative and dispersed population of progenitor cells in the adult central nervous system, which allows these cells to rapidly respond to damage. Oligodendrocytes and myelin are lost after traumatic spinal cord injury (SCI), compromising efficient conduction and, potentially, the long-term health of axons. In response, OPCs proliferate and then differentiate into new oligodendrocytes and Schwann cells to remyelinate axons. This culminates in highly efficient remyelination following experimental SCI in which nearly all intact demyelinated axons are remyelinated in rodent models. However, myelin regeneration comprises only one role of OPCs following SCI. OPCs contribute to scar formation after SCI and restrict the regeneration of injured axons. Moreover, OPCs alter their gene expression following demyelination, express cytokines and perpetuate the immune response. Here, we review the functional contribution of myelin regeneration and other recently uncovered roles of OPCs and their progeny to repair following SCI.     

Human embryonic stem cells differentiate into oligodendrocytes in high purity and myelinate after spinal cord transplantation

Human embryonic stem cells (hESCs) demonstrate remarkable proliferative and developmental capacity. Clinical interest arises from their ability to provide an apparently unlimited cell supply for transplantation, and from the hope that they can be directed to desirable phenotypes in high purity. Here we present for the first time a method for obtaining oligodendrocytes and their progenitors in high yield from hESCs. We expanded hESCs, promoted their differentiation into oligodendroglial progenitors, amplified those progenitors, and then promoted oligodendroglial differentiation using positive selection and mechanical enrichment. Transplantation into the shiverer model of dysmyelination resulted in integration, differentiation into oligodendrocytes, and compact myelin formation, demonstrating that these cells display a functional phenotype. This differentiation protocol provides a means of generating human oligodendroglial lineage cells in high purity, for use in studies of lineage development, screening assays of oligodendroglial-specific compounds, and treating neurodegenerative diseases and traumatic injuries to the adult CNS.     

Integration and differentiation of neural stem cells after transplantation into the dysmyelinated central nervous system of adult mice

Mutant mice deficient in the myelin-associated glycoprotein (MAG) and the nonreceptor-type tyrosine kinase Fyn are characterized by a severely hypomyelinated central nervous system (CNS) and morphologically abnormal myelin sheaths. Despite this pronounced phenotype, MAG/Fyn-deficient mice have a normal longevity. In the present study, we took advantage of the normal life expectancy of this myelin mutant and grafted neural stem cells (NSCs) into the CNS of MAG/Fyn-deficient mice to study in short- and long-term experiments the fate of NSCs in adult dysmyelinated brains. Neural stem cells were isolated from spinal cords of transgenic mouse embryos ubiquitously expressing enhanced green fluorescent protein. Cells were expanded in vitro in the presence of mitogens for up to 5 weeks before they were grafted into the lateral ventricles or injected into white matter tracts. Analysis of mutant brains 3-15 weeks after intracerebroventricular transplantation of NSCs revealed only limited integration of donor cells into the host brains. However, injection of NSCs directly into white matter tracts resulted in widespread distribution of donor cells within the host tissue. Donor cells survived for at least 15 weeks in adult host brains. The majority of grafted cells populated white matter tracts and differentiated into oligodendrocytes that myelinated host axons. Results suggest that intraparenchymal transplantation of NSCs might be a strategy to reconstruct myelin in dysmyelinated adult brains.     

Targeted expression of anti-apoptotic protein p35 in oligodendrocytes reduces delayed demyelination and functional impairment after spinal cord injury

Functional impairment after spinal cord injury (SCI) is attributed to neuronal cell necrosis death and axonotmesis, with further worsening caused by the accompanying apoptosis of myelin-forming oligodendrocytes (OLGs). However, it is unclear as to how much OLG apoptosis contributes to functional impairment. To address this issue, we used transgenic mice characterized by the targeted expression of p35, a broad-spectrum caspase inhibitor, in OLGs using the cre/loxP system (referred to as cre/p35 transgenic mice). In this study, we examined the motor function and histopathologic changes after a contusive thoracic spinal cord injury in the cre/p35 transgenic mice. A larger number of OLGs and a lesser extent of demyelination were observed after SCI in the cre/p35 transgenic mice than in the control cre mice, which did not carry the p35 transgene. Furthermore, the motor function of the hindlimbs recovered to a significantly better degree in the cre/p35 transgenic mice than in the control cre mice. Thus, the inhibition of OLG apoptosis decreased the extent of functional impairment after SCI. These findings suggest that the inhibition of OLG apoptosis may be a potential treatment for SCI.     

他克莫司促进神经干细胞移植大鼠脊髓损伤的再生与修复

背景：研究证实,他克莫司不仅抑制T细胞的增殖、活化,还能抑制小胶质细胞、巨噬细胞等炎症细胞在损伤局部聚集、活化及相关炎症因子的释放,减轻继发性炎症反应对原发损伤周围正常组织的破坏,从而对损伤局部的神经组织起保护作用。 目的：观察他克莫司对神经干细胞移植大鼠脊髓损伤后再生修复的影响。 方法：分离培养孕13d SD大鼠神经干细胞。显微镜下动脉瘤夹夹闭SD大鼠T8脊髓,建立压迫型脊髓损伤动物模型。损伤后7 d随机数字表分为3组：对照组,于损伤中心定向注射生理盐水;细胞移植组,于损伤中心定向注射神经干细胞;他克莫司组,于损伤中心定向注射神经干细胞同时给予免疫抑制剂他克莫司1 mg/(kg•d)腹腔注射连续7 d。1,2,4,8周后,通过BDA顺行示踪、苏木精-伊红与免疫组化染色及电镜检测,观察移植后脊髓组织再生和神经元的变化。 结果与结论：对照组在损伤中心端远侧无神经纤维通过。细胞移植组与他克莫司组在治疗1周后有部分神经纤维通过,8周均有部分BDA阳性标记的皮质脊髓束再生通过脊髓损伤部位,特别是他克莫司组可延续至距损伤中心1.7 cm 。苏木精-伊红染色显示,细胞移植组与他克莫司组2周时坏死灶开始缩小,泡沫细胞减少。电镜结果显示,他克莫司组1周时即出现较正常的微丝和微管结构,8周时星形细胞、许旺细胞、髓鞘典型多见,神经轴突的终末有较多的兴奋性递质和不典型的轴树连接,出现较多的结构正常的髓鞘。说明损伤大鼠移植神经干细胞后联合应用他克莫司后可减轻早期的急性炎症反应,保证神经细胞的存活,具有神经保护和神经营养作用,可加快神经功能的恢复。     

Rolipram联合少突胶质前体细胞移植治疗大鼠脊髓损伤

背景：脊髓损伤轴突难以再生涉及到多方面原因,综合运用多种治疗方法应该更有效。 目的：应用Rolipram联合少突胶质前体细胞移植治疗大鼠脊髓损伤,观察轴突再生和再髓鞘化的情况。 方法：Wistar大鼠利用ALLEN法打击形成脊髓损伤模型。Rolipram治疗组和联合治疗组背部皮下埋置ALZET渗透性微泵内装Rolipram 200 μL,每小时释放0.5 μL,2 mg/(kg•d),可持续14 d,1周后局部注入0.012 5 μmol 的双丁酸环腺苷酸;细胞移植组和联合治疗组移植少突胶质前体细胞,脊髓损伤组注入同等量的生理盐水,术后每周测定BBB运动评分,4周取材,冰冻切片,苏木精-伊红染色,免疫荧光染色。 结果与结论：伤后4周Rolipram治疗组和联合治疗组BBB运动评分高于脊髓损伤组和细胞移植组;免疫荧光显示Rolipram治疗组和联合治疗组损伤局部NF200表达旺盛;细胞移植组和联合治疗组少突胶质前体细胞存活并表达髓鞘碱性蛋白,后者存活数量更多,并且有髓纤维增多。说明应用Rolipram提高环磷腺苷水平促进了大鼠脊髓损伤功能的恢复,联合少突胶质前体细胞移植可产生协同促进作用。     

Quantitative analysis of vascularization and cytochrome oxidase following fetal transplantation in the contused rat spinal cord

In the normal adult central nervous system, a coupling between energy consumption and vascular density is well established. Likewise, the survival of fetal neural tissue grafts is highly dependent on the establishment of functional vascular integration with the host. However, to what degree graft vascularization and tissue metabolism influence the normal host response to traumatic injury has not been extensively studied. In the present report, embryonic day 14 fetal spinal cord suspension grafts were made into the lesion epicenter of subchronic (10 days) contusion-injured rats. Three months later, intraspinal transplants were analyzed using correlative cytochrome oxidase histochemistry and vascular morphometric analysis. The same approaches were applied to the host spinal cord and injured, non-transplanted animals in order to determine the ability of a graft to alter the level of post-injury vascularization and/or metabolism. In general, graft vascular density was increased over that measured in normal or injured gray matter. Vascular density in gray matter near the host/graft interface was markedly increased when compared to either gray matter of the same spinal level in injured non-grafted animals or normal control spinal gray matter. Vascular changes were not noted in gray matter 3 mm distal to the lesion epicenter (rostral or caudal) in all groups analyzed. Cytochrome oxidase was up-regulated at this time in the graft and gray matter at the host/graft interfaces when compared to either gray matter of the same spinal level in injured, non-grafted animals or that of uninjured controls. These data indicate that an intraspinal transplant placed into the contused adult rat spinal cord reaches a metabolic capacity that is likely to be associated with high levels of oxidative metabolism in the well-vascularized graft neuropil. In addition, transplantation chronically alters vascularization and metabolic patterns of adjacent spinal gray matter following contusion injury. © 1996 Wiley-Liss, Inc.     

Differentiation of endogenous neural precursors following spinal cord injury in adult rats☆

BACKGROUND： Studies have shown that cell death can activate proliferation of endogenous neural stem cells and promote newly generated cells to migrate to a lesion site.
OBJECTIVE： To observe regeneration and differentiation of neural cells following spinal cord injury in adult rats and to quantitatively analyze the newly differentiated cells.
DESIGN, TIME AND SETTING： A cell biology experiment was performed at the Institute of Orthopedics and Medical Experimental Center, Lanzhou University, between August 2005 and October 2007.
MATERIALS： Fifty adult, Wistar rats of both sexes; 5-bromodeoxyuridine （BrdU, Sigma, USA）; antibodies against neuron-specific enolase, glial fibrillary acidic protein, and myelin basic protein （Chemicon, USA）.
METHODS： Twenty-five rats were assigned to the spinal cord injury group and received a spinal cord contusion injury. Materials were obtained at day 1, 3, 7, 15, and 29 after injury, with 5 rats for each time point. Twenty-five rats were sham-treated by removing the lamina of the vertebral arch without performing a contusion.
MAIN OUTCOME MEASURES： The phenotype of BrdU-labeled cells, i.e., expression and distribution of surface markers for neurons （neuron-specific enolase）, astrocytes （glial fibrillary acidic protein）, and oligodendrocytes （myelin basic protein）, were identified with immunofluorescence double-labeling. Confocal microscopy was used to detect double-labeled cells by immunofluorescence. Quantitative analysis of newly generated cells was performed with stereological counting methods.
RESULTS： There was significant cell production and differentiation after adult rat spinal cord injury. The quantity of newly-generated BrdU-labeled cells in the spinal cord lesion was 75-fold greater than in the corresponding area of control animals. Endogenous neural precursor cells differentiated into astrocytes and oligodendrocytes, however spontaneous neuronal differentiation was not detected. Between 7 and 29 d after spinal cord injury, newl     

Differentiation of endogenous neural precursors following spinal cord injury in adult rats

BACKGROUND:Studies have shown that cell death can activate proliferation of endogenous neural stem cells and promote newly generated cells to migrate to a lesion site.OBJECTIVE:To observe regeneration and differentiation of neural cells following spinal cord injury in adult rats and to quantitatively analyze the newly differentiated cells.DESIGN,TIME AND SETTING:A cell biology experiment was performed at the Institute of Orthopedics and Medical Experimental Center,Lanzhou University.between August 2005 and October 2007.MATERIALS:Fifty adult,Wistar rats of both sexes;5-bromodeoxyuridine(BrdU,Sigma,USA);antibodies against neuron-specific enolase,glial fibrillary acidic protein,and myelin basic protein(Chemicon,USA).METHODS:Twenty-five rats were assigned to the spinal cord injury group and received a spinal cord contusion injury.Materials were obtained at day 1,3,7,15,and 29 after injury,with 5 rats for each time point.Twenty-five rats were sham-treated by removing the lamina of the vertebral arch without performing a contusion.MAIN OUTCOME MEASURES:The phenotype of BrdU-labeled cells,i.e.,expression and distribution of surface markers for neurons(neuron-specific enolase),astrocytes(glial fibrillary acidic protein),and oligodendrocytes(myelin basic protein),were identified with immunofluorescence double-labeling.Confocal microscopy was used to detect double-labeled cells by immunofluorescence.Quantitative analysis of newly generated cells was performed with stereological counting methods.RESULTS:There was significant cell production and differentiation after adult rat spinal cord injury.The quantity of newly-generated BrdU-labeled cells in the spinal cord lesion was 75-fold greater than in the corresponding area of control animals.Endogenous neural precursor cells differentiated into astrocytes and oligodendrocytes,however spontaneous neuronal difierentiation was not detected.Between 7 and 29 d after spinal cord injury,newly generated cells expressed increasingly more mature oligodendrocyte and astrocyte markers.CONCLUSION:Spinal cord injury is a direct inducer of regeneration and differentiation of neural cells.Endogenous neural precursor cells Can difierentiate into astrocytes and oligodendrocytes following adult rat spinal cord injury.     

Migration and differentiation of nuclear fluorescence‐labeled bone marrow stromal cells after transplantation into cerebral infarct and spinal cord injury in mice

There is increasing evidence that bone marrow stromal cells (BMSC) have the potential to migrate into the injured neural tissue and to differentiate into the CNS cells, indicating the possibility of autograft transplantation therapy. The present study was aimed to clarify whether the mouse BMSC can migrate into the lesion and differentiate into the CNS cells when transplanted into the mice subjected to focal cerebral infarct or spinal cord injury. The BMSC were harvested from mice and characterized by flow cytometry. Then, the BMSC were labeled by bis‐benzimide, a nuclear fluorescence dye, over 24 h, and were stereotactically transplanted into the brain or spinal cord of the mice. The cultured BMSC expressed low levels of CD45 and high levels of CD90 and Sca‐1 on flow cytometry. A large number of grafted cells survived in the normal brain 4 weeks after transplantation, many of which were located close to the transplanted sites. They expressed the neuronal marker including NeuN, MAP2, and doublecortin on fluorescent immunohistochemistry. However, when the BMSC were transplanted into the ipsilateral striatum of the mice subjected to middle cerebral artery occlusion, many of the grafted cells migrated into the corpus callosum and injured cortex, and also expressed the neuronal markers 4 weeks after transplantation. In particular, NeuN was very useful to validate the differentiation of the grafted cells, because the marker was expressed in the nuclei and was overlapped with bis‐benzimide. Similar results were obtained in the mice subjected to spinal cord injury. However, many of the transplanted BMSC expressed GFAP, an astrocytic protein, in injured spinal cord. The present results indicate that the mouse BMSC can migrate into the CNS lesion and differentiate into the neurons or astrocytes, and that bis‐benzimide is a simple and useful marker to label the donor cells and to evaluate their migration and differentiation in the host neural tissues over a long period.     

Effect of glial cells on remyelination after spinal cord injury

Remyelination plays a key role in functional recovery of axons after spinal cord injury. Glial cells are the most abundant cells in the central nervous system. When spinal cord injury occurs, many glial cells at the lesion site are immediately activated, and different cells differentially affect inflammatory reactions after injury. In this review, we aim to discuss the core role of oligodendrocyte precursor cells and crosstalk with the rest of glia and their subcategories in the remyelination process. Activated astrocytes influence prolif-eration, differentiation, and maturation of oligodendrocyte precursor cells, while activated microglia alter remyelination by regulating the inflammatory reaction after spinal cord injury. Understanding the interac-tion between oligodendrocyte precursor cells and the rest of glia is necessary when designing a therapeutic plan of remyelination after spinal cord injury.     

Combined NgR vaccination and neural stem cell transplantation promote functional recovery after spinal cord injury in adult rats

C‐J. Xu, L. Xu, L‐D. Huang, Y. Li, P‐P. Yu, Q. Hang, X‐M. Xu and P‐H. Lu (2011) Neuropathology and Applied Neurobiology 37, 135–155
Combined NgR vaccination and neural stem cell transplantation promote functional recovery after spinal cord injury in adult rats Aims: After spinal cord injury (SCI), there are many adverse factors at the lesion site such as glial scar, myelin‐derived inhibitors, cell loss and deficiency of neurotrophins that impair axonal regeneration. Therefore, combination therapeutic strategies might be more effective than a single strategy for promoting functional recovery after SCI. In the present study, we investigated whether a Nogo66 receptor (NgR) vaccine, combined with neural stem cell (NSC) transplantation, could promote better functional recovery than when NgR vaccine or NSCs were used alone. Methods: Adult rats were immunized with NgR vaccine at 1 week after a contusive SCI at the thoracic level, and the NSCs, obtained from green fluorescent protein transgenic rats, were transplanted into the injury site at 8 weeks post injury. The functional recovery of the animals under various treatments was evaluated by three independent behavioural tests, that is, Basso, Beattie and Bresnahan locomotor rating scale, footprint analysis and grid walking. Results: The combined therapy with NgR vaccination and NSC transplantation protected more ventral horn motor neurones in the injured spinal cord and greater functional recovery than when they were used alone. Furthermore, NgR vaccination promoted migration of engrafted NSCs along the rostral‐caudal axis of the injured spinal cords, and induced their differentiation into neurones and oligodendrocytes in vivo. Conclusions: The combination therapy of NgR vaccine and NSC transplantation exhibited significant advantages over any single therapy alone in this study. It may represent a potential new therapy for SCI.     

Number of oligodendrocyte progenitors recruited to the lesioned spinal cord is modulated by the levels of the cell cycle regulatory protein p27Kip-1

Remyelination is a critical step for recovery of function after demyelination and defines the ability to generate new myelin. This repair process is dependent on the presence of resident oligodendrocyte progenitors (OLPs) that have been shown to remyelinate axons after demyelination. We have previously shown that the levels of the cell cycle inhibitor p27Kip-1 modulate the number of neonatal cortical OLPs. We now asked whether this cell cycle molecule plays also a role in regulating the number of adult OLP in the spinal cord after demyelination induced by lysolecithin injection. The proliferative response of OLP in the spinal cord of injected wild-type (wt) and p27Kip-1 null mice was evaluated 3 days after lesion. In vivo labeling with bromodeoxyuridine (BrdU) was used to identify cells in S phase. Double immunofluorescence for the OLP marker NG2, and for BrdU was used to count the number of proliferating progenitors. Consistent with a role of p27Kip-1 in regulating the number of adult OLP in the injured spinal cord, a larger number of proliferating OLPs was observed in p27Kip-1null mice compared with wild-type controls. These cells were able to differentiate as assessed by the presence of MBP+ cells in the spinal cord 14 days after injury. We conclude that the cellular levels of the cell cycle inhibitor p27Kip-1 modulate the repair response of OLPs to injury in the adult spinal cord.     

Phenotypic characterization of neural stem cells from human fetal spinal cord: synergistic effect of LIF and BMP4 to generate astrocytes

If cell based therapy for spinal cord injury is to become a reality, greater insights into the biology of human derived spinal cord stem cells are a prerequisite. Significant species differences and regional specification of stem cells necessitates determining the effects of growth factors on human spinal cord stem cells. Fetal spinal cords were dissociated and expanded as neurospheres in medium with bone morphogenetic protein 4 (BMP4), leukemia inhibitory factor (LIF) or BMP4 and LIF. First-generation neurospheres comprised a heterogeneous population of neural cell types and after plating emergent cells included neurons, oligodendrocytes and GFAP(+) cells which coexpressed stem cells markers and those of the neuronal lineage and were thus identified as GFAP(+) neural precursor cells (NPC). When plated, neurospheres maintained in BMP4 demonstrated a reduced proportion of emergent oligodendrocytes from 13 to 4%, whereas LIF had no statistically significant effect on cell type distribution. Combining BMP4 and LIF reduced the proportion of oligodendrocytes to 3% and that of neurons from 37 to 16% while increasing the proportion of GFAP(+) NPC from 45 to 79%. After 10 passages in control media aggregates gave rise to multiple neural phenotypes and only continued passage of neurospheres in the presence of BMP4 and LIF resulted in unipotent aggregates giving rise to only astrocytes. These results provide a means of obtaining pure populations of human spinal-cord derived astrocytes, which could be utilized for further studies of cell replacement strategies or in vitro evaluation of therapeutics.     

Distribution and differentiation of A2B5+ glial precursors in the developing rat spinal cord

In many regions of the rat central nervous system, oligodendrocytes develop from migratory A2B5⁺ precursor cells. In the rat spinal cord, during early embryonic development the capacity for oligodendrogenesis appears to be restricted to ventral regions of the spinal cord, while cultures of postnatal rat spinal cord contain a distinct population of A2B5⁺ astrocyte precursors. To determine if, as in other regions of the CNS, spinal cord A2B5⁺ cells give rise directly to oligodendrocytes and astrocytes, the initial distribution, and subsequent dispersion, proliferation, and differentiation of spinal cord A2B5⁺ cells have been examined in both explant and dissociated cell cultures. Spinal cord oligodendrocytes develop from A2B5⁺ cells. At E14, A2B5⁺ cells are restricted to ventral regions of the spinal cord and as development proceeds they become more uniformly distributed throughout the spinal cord. In explant cultures, greater than 95% of the explants that contain oligodendrocytes also contain A2B5⁺ cells and a proportion of mature oligodendrocytes retain detectable A2B5 immunoreactivity briefly on their surface. The maturation of spinal cord oligodendrocyte precursors occurs in a number of distinct stages characterized by the expression of O4 immunoreactivity, which first appears at E16, and GC immunoreactivity, which first appears at E18. As spinal cord oligodendrocyte precursors acquire O4 immunoreactivity they appear to lose the ability to proliferate in response to PDGF but retain the ability to proliferate in response to bFGF, suggesting that the control of proliferation of oligodendrocyte precursors is, in part, dependent on their maturational state. In the presence of high serum, spinal cord A2B5⁺ cells fail to develop in isolated E14 dorsal spinal cord cultures, while in ventral cultures they subsequently differentiate into A2B5⁺ astrocytes suggesting that A2B5⁺ astrocyte precursors are also initially ventrally located. Unlike oligodendrocyte differentiation, however, the differentiation of spinal cord A2B5⁺ cells into astrocytes is delayed in early embryonic-derived cultures compared to those from older animals. These observations suggest that local influences may regulate the timing of spinal cord A2B5⁺ astrocyte development, but not spinal cord oligodendrocyte development. © 1994 Wiley-Liss, Inc.     

Passive exercise and fetal spinal cord transplant both help to restore motoneuronal properties after spinal cord transection in rats

Spinal cord transection influences the properties of motoneurons and muscles below the lesion, but the effects of interventions that conserve muscle mass of the paralyzed limbs on these motoneuronal changes are unknown. We examined the electrophysiological properties of rat lumbar motoneurons following spinal cord transection, and the effects of two interventions shown previously to significantly attenuate the associated hindlimb muscle atrophy. Adult rats receiving a complete thoracic spinal cord transection (T-10) were divided into three groups receiving: (1) no further treatment; (2) passive cycling exercise for 5 days/week; or (3) acute transplantation of fetal spinal cord tissue. Intracellular recording of motoneurons was carried out 4-5 weeks following transection. Transection led to a significant change in the rhythmic firing patterns of motoneurons in response to injected currents, as well as a decrease in the resting membrane potential and spike trigger level. Transplants of fetal tissue and cycling exercise each attenuated these changes, the latter having a stronger effect on maintenance of motoneuron properties, coinciding with the reported maintenance of structural and biochemical features of hindlimb muscles. The mechanisms by which these distinct treatments affect motoneuron properties remain to be uncovered, but these changes in motoneuron excitability are consistent with influences on ion conductances at or near the initial segment. The results may support a therapeutic role for passive limb manipulation and transplant of stem cells in slowing the deleterious responses of motoneurons to spinal cord injury, such that they remain more viable for subsequent alternative strategies.     

Intraspinal transplantation of embryonic spinal cord tissue in neonatal and adult rats

Fetal rat spinal cord tissue was obtained on gestational day 14 (E14) and transplanted into 2-4-mm-long intraspinal cavities produced by partial spinal cord lesions in adult and neonatal rats. At regular post-transplantation intervals, light and electron microscopy, autoradiographic demonstration of tritiated thymidine labelling, and immunocytochemical localization of glial fibrillary acidic protein (GFAP) were used to identify surviving donor tissues and to study their differentiation and extent of fusion with recipient spinal cords. In some experiments, wheat germ agglutinin-conjugated horseradish peroxidase (WGA-HRP) was also employed to examine whether neurons within the grafts projected axons into the host spinal cord and vice versa. Lastly, immunocytochemistry was used to determine whether any supraspinal serotoninergic (5-HT) axons from the host extended into the transplants. Over 80% of the grafts survived in lesions of both the neonatal and adult rat spinal cord for periods of 1-16 months (duration of experiment), and considerable maturation of donor tissue was evidenced, which even included the appearance of some topographical features of the normal spinal cord. Many of the transplants extended the entire length of the lesion, and were often closely apposed to the injured surfaces of the recipient spinal cords without an intervening dense glial scar. At post-transplantation intervals of 2-4 months, injection of WGA-HRP into the host spinal cord (5 mm from the transplant in adult animals or as much as 20 mm in neonatal recipients) demonstrated retrogradely labelled neurons and anterogradely labelled axons in the grafts. Likewise, injecting WGA-HRP into transplants in adult recipients resulted in labelling of neurons in adjacent segments of the host spinal cord; some labelled axons, derived from donor neurons, were also present in neighboring spinal gray matter. Finally, immunocytochemistry revealed 5-HT-like immunoreactive fibers in transplants that had been prelabelled with tritiated thymidine. These observations demonstrate the potential of embryonic spinal cord transplants to replace damaged intraspinal neuronal populations and to restore some degree of anatomical continuity between the isolated rostral and caudal stumps of the injured mammalian spinal cord.     

Effect of methylprednisolone on motor function and spinal cord blood flow after spinal cord compression in rats

The effect of methylprednisolone (MP) on neurologic recovery and spinal cord blood flow (SCBF) was investigated up to 4 days after a spinal cord compression injury in rats. The injury was produced at midthoracic level by applying a load of 35 g on a 2.2 x 5.0 mm compression plate for 5 min, which resulted in transient paraparesis. MP was given as a bolus dose of 30 mg/kg i.v. 60 min after injury (n = 20) and controls were given saline (n = 10). The motor performance was assessed daily as the capacity angle on the inclined plane and SCBF was measured by 14C-iodoantipyrine autoradiography on Days 1 or 4. On Day 1 the capacity angle was reduced from about 63 degrees preoperatively to 33 +/- 2 degrees (mean +/- SEM) in the control group and to 50 +/- 1 degrees in the group treated with MP (p less than 0.05). Thereafter there was a slight improvement in both groups, but the difference persisted throughout the observation period. On Day 4 both gray and white matter SCBF was better preserved in MP-treated animals than in the control group (59 +/- 4 versus 49 +/- 3 ml/min/100 g tissue for gray matter and 13.6 +/- 0.6 versus 10.7 +/- 0.8 ml/min/100 g tissue for white matter). Posttraumatic treatment with MP, thus, improved both the neurologic recovery during the first 4 days and SCBF as measured on Day 4. It is speculated that the effect of MP is at least partly exerted on the vascular bed.