干细胞和脊髓全横断损伤（stem cell and spinal cordtransection）

1: Li W, Cai WQ, Li CR. Repair of spinal cord injury by neural stem cells modified with BDNF gene inrats. Neurosci Bull. 2006;22(1):34-40.School of Nursing; Department of Histology and Embryology, Third Military Medical University, Chongqing Objective To explore repair of spinal cord injury by neural stem cells (NSCs) modified with brain derived neurotrophic factor (BDNF) gene (BDNF-NSCs) in rats. Methods Neural stem cells modified with BDNF gene were transplanted into the complete transection site of spinal cord at the lumbar 4 (L4) level in rats. Motor function of rats hind limbs was observed and HE and X-gal immunocytochemical staining, in situ hybridization, and retrograde HRP tracing were also performed. Results BDNF-NSCs survived and integrated well with host spinal cord. In the transplant group, some X-gal positive, NF-200 positive, GFAP positive, BDNF positive, and BDNF mRNA positive cells, and many NF-200 positivenerve fibers were observed in the injury site. Retrograde HRP tracing through sciatic nerve showed some HRP positive cells and nerve fibers near the rostral side of the injury one month after transplant and with time, they increased in number. Examinations on rats motor function and behavior demonstrated that motor function of rats hind limbs improved better in the transplant group than the injury group. Conclusion BDNF-NSCs can survive, differentiate, and partially integrate with host spinal cord, and they significantly ameliorate rats motor function of hind limbs, indicating their promising role in repairing spinal cord injury.构建有BDNF基因的神经干细胞植入横断脊髓通过HE和X-gal（用于标记克隆产物的）在脊髓的免疫组化。结果：X-gal+，NF-200+， GFAP，BDNFmRNA含量增高。HRP从坐骨神经逆行追踪。2: Pu Y, Guo QS, Wang AM, Wu SY, Xing SX, Zhang ZR. Repair of acutely injured spinal cord through constructing tissue-engineeredneural complex in adult rats. Chin J Traumatol. 2007;10(3):171-6.Department of Orthopedics, Daping Hospital, Third Military Medical University.OBJECTIVE: To construct tissue-engineered neural complex in vitro and study its effect in repairing acutely injured spinal cord in adult rats. METHODS: Neural stem cells were harvested from the spinal cord of embryo rats and propagated in vitro. Then the neural stem cells were seeded into polyglycolic acid scaffolds and co-cultured with extract of embryonic spinal cord in vitro. Immunofluorescence histochemistry and scanning electron microscope were used to observe the microstructure of this complex. Animal model of spine semi-transection was made and tissue-engineered neural complex was implanted by surgical intervention. Six weeks after transplantation, functional evaluation and histochemistry were applied to evaluate the functional recovery and anatomic reconstruction. RESULTS: The tissue-engineered neural complex had a distinct structure, which contained neonatal neurons, oligodendrocytes and astrocytes. After tissue-engineered neural complex was implanted into the injured spinal cord, the cell components such as neurons, astrocytes and oligodendrocytes, could survive and keep on developing. The adult rats suffering from spinal cord injury got an obvious neurological recovery in motor skills. CONCLUSIONS: The tissue-engineered neural complex appears to have therapeutic effects on the functional recovery and anatomic reconstruction of the adult rats with spinal cord injury.用胎鼠脊髓匀浆的液体，共培养构建组织支架聚乙醇酸并载有神经干细胞。并用免疫荧光和电子显微镜来观察其变化。将此复合物植入半横断损伤的大鼠脊髓，6周后，运用免疫组化观察其形态结构变化，并观察其运动功能的恢复。3: Gulino R, Cataudella T, Casamenti F, Pepeu G, Stanzani S, Leanza G. Acetylcholine release from fetal tissue homotopically grafted to themotoneuron-depleted lumbar spinal cord. An in vivo microdialysis study in the awake rat. Exp Neurol. 2007;204(1):326-38. Department of Physiological Sciences, University of Catania, Viale A. Doria 6, 95125 Catania, Italy.Grafts of spinal cord (SC) tissue can survive and develop into the severed SC, but no conclusive data are available concerning the functional activity of transplanted neurons. In the present study, suspensions of prelabeled embryonic ventral SC tissue were grafted to the lumbar SC of rats with motoneuron loss induced by perinatal injection of volkensin. Eight to ten months post-grafting, acetylcholine (ACh) release was measured by microdialysis in awake rats, under either basal or stimulated conditions. In normal animals, baseline ACh output averaged 1.6 pmol/30 microl, it exhibited a 4-fold increase after KCl-induced depolarization or handling, and it was completely inhibited by tetrodotoxin administration. Moreover, ACh levels did not change following acute SC transection performed under anesthesia during ongoing dialysis, suggesting an intrinsic source for spinal ACh. Treatment with volkensin produced a severe (85%) motoneuronal loss accompanied by a similar reduction in baseline Ach release and almost completely abolished effects of depolarization or handling. In transplanted animals, many motoneuron-like labeled cells were found within and just outside the graft area, but apparently in no case were they able to extend fibers towards the denervated muscle. However, the grafts restored baseline Ach output up to near-normal levels and responded with significantly increasedrelease to depolarization, but not to handling. The present findings indicate that spinal neuroblasts can survive and develop within the motoneuron-depleted SC and release ACh in a near-normal, but apparently non-regulated, manner. This may be of importance for future studies involving intraspinal stem cell grafts.正常大鼠在清醒状态下脊髓运动神经元分泌ACH，在受到KCl刺激去极化的情况下，分泌的ACH较正常时高0.4倍，而运用河豚毒素后未测到Ach，表明ACH为神经元自分泌。建立模型使85%以上的运动神经元丢失，ACH明显降低。运用已做标记的胎鼠复角神经元植入大鼠损伤脊髓运动神经元丢失的腰髓节段。术后810周， 监测发现ACH基本恢复到正常大鼠水平。4: Burns AS, Boyce VS, Tessler A, Lemay MA. Fibrillation potentials following spinal cord injury: improvement with neurotrophins and exercise. Muscle Nerve. 2007 ;35(5):607-13.Department of Rehabilitation Medicine, Thomas Jefferson University, 132 S. 10th Street, 375 Main Building, Philadelphia, Pennsylvania 19107, USA.Fibrillation potentials and positive sharp waves (spontaneous potentials) are the electrophysiological hallmark of denervated skeletal muscle, and their detection by intramuscular electromyography (EMG) is the clinical gold standard for diagnosing denervated skeletal muscle. Surprisingly, spontaneous potentials have been described following human and experimental spinal cord injury (SCI) in muscles innervated by spinal cord segments distal to the level of direct spinal injury. To determine whether electrophysiological abnormalities are improved by two therapeutic interventions for experimental SCI, neurotrophic factors and exercise training, we studied four representative hindlimb muscles in adultdomestic short-hair cats following complete transection of the spinal cord at T11-T12. In untreated cats, electrophysiological abnormalities persisted unchanged for 12 weeks postinjury, the longest duration studied. In contrast, fibrillations and positive sharp waves largely resolved in animals that underwent weight-supported treadmill training or received grafts containing fibroblasts genetically modified to express brain-derived neurotrophic factor and neurotrophin-3. These findings suggest that neurotrophins and activity play an important role in the poorly understood phenomenon of fibrillations distal to SCI.猫T11-T12全横断后手术对照组术后12周骨骼肌电生理-，而植入神经生长因子BDNF、NT-3和术后给与运动功能锻炼恢复的两组有明显恢复。5: Vroemen M, Caioni M, Bogdahn U, Weidner N. Failure of Schwann cells as supporting cells for adult neural progenitor cell grafts in the acutely injured spinal cord. Cell Tissue Res. 2007;327(1):1-13. Epub 2006 Aug 29.Department of Neurology, University of Regensburg, Universittsstrasse 84, 93053 Regensburg, Germany.Adult neural progenitor cells (NPC) co-grafted with fibroblasts replace cystic lesion defects and promote cell-contact-mediated axonal regeneration in the acutely injured spinal cord. Fibroblasts are required as a platform to maintain NPC within the lesion; however, they are suspected to create an inhospitable milieu for regenerating central nervous system (CNS) axons. Therefore, we thought to replace fibroblasts by primary Schwann cells, which might serve as a superior scaffold to maintain NPC within the lesion and might further enhance axon regrowth and remyelination following spinal cord injury. Adult rats underwent acervical dorsal column transection immediately followed by transplant- ation of either NPC/Schwann cell or NPC/Schwann cell/fibroblast co-grafts. Animals receiving Schwann cell or fibroblast grafts alone, or Schwann cell/fibroblast co-grafts served as controls. At 3 weeks after injury/ transplantation, histological analysis revealed that only fibroblast -containing grafts were able to replace the cystic lesion defect. In both co-cultures and co-grafts, Schwann cells and NPC were segregated. Almost all NPC migrated out of the graft into the adjacent host spinal cord. As a consequence, only peripheral-type myelin, but no CNS-type myelin, was detected within co-grafts containing NPC/Schwann cells. Corticospinal axon regeneration into Schwann-cell-containing co-grafts was reduced. Taken together, Schwann cells within NPC grafts contribute toremyelination. However, Schwann cells fail as a supporting platform to maintain NPC within the graft and impair CNS axon regeneration; this makes them an unfavorable candidate to support/augment NPC grafts following spinal cord injury.联合移植成纤维细胞和神经祖细胞对周围神经损伤恢复较好，成纤维细胞可以作为一个支架，并能形成髓鞘，但在CNS却不太理想，所以考虑到了雪旺氏细胞和神经祖细胞联合移植。颈段脊髓横断后，分别移植NPC/Schwann或NPC/Schwann cell/fibroblast并单独移植Schwann，fibroblast或chwann cell/fibroblast作为对照组。3周后组织化学发现，只有fibroblast包裹NPC能发挥作用。NPC/Schwann只有边缘有髓鞘，而中心未形成。皮质脊髓束在NPC/Schwann组再生能力减低，所以说，Schwann能形成髓鞘但它对中枢神经恢复没有什么效果。6: Liang P, Jin LH, Liang T, Liu EZ, Zhao SG. Human neural stem cells promote corticospinal axons regeneration and synapse reformation in injured spinal cord of rats. Chin Med J (Engl). 2006 20;119(16):1331-8.Department of Neurosurgery, First Clinical College, Harbin Medical University.BACKGROUND: Axonal regeneration in lesioned mammalian central nervous system is abortive, and this causes permanent disabilities in individuals with spinal cord injuries. This paper studied the action of neural stem cell (NSC) in promoting corticospinal axons regeneration and synapse reformation in rats with injured spinal cord. METHODS: NSCs were isolated from the cortical tissue of spontaneous aborted human fetuses in accordance with the ethical request. The cells were discarded from the NSC culture to acquire NSC-conditioned medium. Sixty adultWistar rats were randomly divided into four groups (n = 15 in each): NSC graft, NSC medium, graft control and medium control groups. Microsurgical transection of the spinal cord was performed in all the rats at the T11. The NSC graft group received stereotaxic injections of NSCs suspension into both the spinal cord stumps immediately after transection; graft control group received DMEM injection. In NSC medium group, NSC- conditioned medium was administered into the spinal cord every week; NSC culture medium was administered to the medium control group. Hindlimb motor function was assessed using the BBB Locomotor Rating Scale.Regeneration of biotin dextran amine (BDA) labeled corticospinal tract was assessed. Differentiation of NSCs and the expression of synaptophysin at the distal end of the injured spinal cord were observed under a confocal microscope. Group comparisons of behavioral data were analyzed with ANOVA. RESULTS: NSCs transplantation resulted in extensive growth of corticospinal axons and locomotor recovery in adult rats after complete spinal cord transection, the mean BBB scores reached 12.5 in NSC graft group and 2.5 in graft control group (P 0.05). There was also significant difference in BBB score between the NSC medium (11.7) and medium control groups (3.7, P 0.05). BDA traces regenerated fibers sprouted across the lesion site and entered the caudal part of the spinal cord. Synaptophysin expression colocalized with BDA positive axons and neurons distalto the injury site. Transplanted cells were found to migrate into the lesion, but not scatter along the route of axon grows. The cells differentiated into astrocytes or oligodendrocytes, but not into the neurons after transplantation. Furthermore, NSC medium administration did not limit the degree of axon sprouting and functional recovery of the injured rats compared to the NSC graft group. CONCLUSIONS: Human embryonic neural stem cells can promote functional corticospinal axons regeneration and synapse reformation in the injured spinal cord of rats. The action is mainly through the nutritional effect of the stem cells on the spinal cord.脊髓全横断大鼠移植人的神经干细胞（术后头尾短分别注射）；并以移植NSCs培养液为对照组（并每周注射NSCs培养液），以移植DMEM培养基为另一组对照。并用BDA追踪。结果：移植NSC组皮质脊髓束再生明显，后肢功能恢复明显。NSCs培养液移植的后肢运动功能明显高于移植DMEM组。BDA追踪穿过损伤部位到达尾段。7: Chen Y, Lin G, Slack JM. Control of muscle regeneration in the Xenopus tadpole tail by Pax7.Development. 2006;133(12):2303-13. Centre for Regenerative Medicine, Department of Biology and Biochemistry,University of Bath, Bath BA2 7AY, UK.The tail of the Xenopus tadpole will regenerate completely after transection. Much of the mass of the regenerate is composed of skeletal muscle, but there has been some uncertainty about the source of the new myofibres. Here, we show that the growing tail contains many muscle satellite cells. They are active in DNA replication, whereas the myonuclei are not. As in mammals, the satellite cells express pax7. We show that a domain-swapped construct, pax7EnR, can antagonize pax7 function. Transgenic tadpoles were prepared containing pax7EnR driven by aheat-inducible promoter. When induced, this reduces the proportion of satellite cells formed in the regenerate. A second amputation of the resulting tails yielded second regenerates containing notochord and spinal cord but little or no muscle. This shows that inhibition of pax7 action does not prevent differentiation of satellite cells to myofibres, but it does prevent their maintenance as a stem cell population.8: Labombarda F, Gonzalez S, Gonzalez Deniselle MC, Garay L, Guennoun R, Schumacher M, De Nicola AF. Progesterone increases the expression of myelin basic protein and the number of cells showing NG2 immunostaining in the lesioned spinal cord. J Neurotrauma. 2006;23(2):181-92.Laboratory of Neuroendocrine Biochemistry, Institute of Experimental Biology and Medicine, and Department of Human Biochemistry, Faculty of Medicine, University of Buenos Aires, Buenos Aires, Argentina. It is now widely accepted that progesterone (PROG) brings neuroprotection in lesions of the peripheral and central nervous system. Spinal cord trauma leads to neuronal degeneration, astrogliosis, demyelination, and proliferation of oligodendrocyte-precursor cells (OPCs). In this work, we studied the effects of PROG on myelin-related parameters in rats with complete spinal cord transaction (TRX). To this end, sham-operated controls and rats with TRX at thoracic level T10 received vehicle or PROG (4 mg/kg/day) during 3 days. Three variables were measured in the lumbar L4 region below the lesion: (1) expression of myelin basic protein (MBP) at the protein and mRNA levels; (2) density of NG2-immunopositive cells as markers for OPCs; and (3) number of cells immunopositive for RIP, an antibody staining mature oligodendrocytes. TRX decreased MBP immunostaining in the corticospinal tract (CST) and dorsal ascending tract (DAT) but not the ventral funiculus (VF). NG2+ cells, which were detected in low number in controls, increased after TRX in the gray and white matter. RIP-positive cell number, however, remained unchanged. PROG treatment of rats with TRX enhanced the expression of MBP protein and mRNA in CST and DAT, but not VF and highly stimulated the number of cells showing NG2 immunostaining over untreated lesioned rats. Instead, density of RIP positive cells was similar in the PROG-treated and untreated lesioned groups. We propose that PROG effects on expression of MBP and the number of NG2 immunopositive cells may contribute to neuroprotection, as they go in parallel with previous results showing enhanced biochemical and morphological parameters of motoneurons of animals with TRX receiving PROG treatment.研究在脊髓T10全横断后黄体酮与髓鞘相关蛋白的关系，在L4监测髓鞘碱性蛋白（MBP）的蛋白及mRNA 的表达变化；用免疫组化观察NG2（少突胶质细胞的标记）的密度；结果：黄体酮促进MBP的表达和NG2阳性密度的增加。9: Hamada M, Yoshikawa H, Ueda Y, Kurokawa MS, Watanabe K, Sakakibara M, Tadokoro M, Akashi K, Aoki H, Suzuki N. Introduction of the MASH1 gene into mouse embryonic stem cells leads to differentiation of motoneuron precursors lacking Nogo receptor expression that can be applicable for transplantation to spinal cord injury. Neurobiol Dis. 2006 22(3): 509-22. Department of Immunology and Medicine, St. Marianna University School ofMedicine, 2-16-1, Sugao, Miyamae-ku, Kawasaki, Kanagawa 216-8511, Japan.ES cells transfected with the MASH1 gene yielded purified spinal motoneuron precursors expressing HB9 and Islet1. The cells lacked the expression of Nogo receptor that was of great advantage for axon growth after transplantation to an injured spinal cord. After transplantation, mice with the complete transection of spinal cord exhibited excellent improvement of the motor functions. Electrophysiological assessment confirmed the quantitative recovery of motor-evoked potential in the transplanted spinal cord. In the grafted spinal cord, gliosis was inhibited and Nogo receptor expression was scarcely detected. The transplanted cells labeled with GFP showed extensive outgrowth of axonspositive for neurofilament middle chain, connected to each other and expressed Synaptophysin, Lim1/2 and Islet1. Thus, the in vivo differentiation into mature spinal motoneurons and the reconstitution of neuronal pathways were suggested. The grafted cell population was pur