Human amnion tissue injected with human umbilical cord mesenchymal stem cells repairs damaged sciatic nerves in rats

Human umbilical cord mesenchymal stem cells,incorporated into an amnion carrier tubes,were assessed for nerve regeneration potential in a rat nerve defect model.Damaged nerves were exposed to human amnion carriers containing either human umbilical cord mesenchymal stem cell (cell transplantation group)or saline(control group).At 8,12,16 and 20 weeks after cell implantation,the sciatic functional index was higher in the cell transplantation group compared with the control group.Furthermore,electrophysiological examination showed that threshold stimulus and maximum stimulus intensity gradually decreased while compound action potential amplitude gradually increased.Hematoxylin-eosin staining showed that regenerating nerve fibers were arranged in nerve tracts in the cell transplantation group and connective tissue between nerve tracts and amnion tissue reduced over time.Gastrocnemius muscle cell diameter,wet weight and restoration ratio were increased.These data indicate that transplanted human umbilical cord mesenchymal stem cells,using the amnion tube connection method,promote restoration of damaged sciatic nerves in rats.     

Human umbilical cord blood-derived stem cells and brain-derived neurotrophic factor protect injured optic nerve：viscoelasticity characterization

The optic nerve is a viscoelastic solid-like biomaterial.Its normal stress relaxation and creep properties enable the nerve to resist constant strain and protect it from injury.We hypothesized that stress relaxation and creep properties of the optic nerve change after injury.Moreover,human brain-derived neurotrophic factor or umbilical cord blood-derived stem cells may restore these changes to normal.To validate this hypothesis,a rabbit model of optic nerve injury was established using a clamp approach.At 7 days after injury,the vitreous body received a one-time injection of 50 μg human brain-derived neurotrophic factor or 1 × 106 human umbilical cord blood-derived stem cells.At 30 days after injury,stress relaxation and creep properties of the optic nerve that received treatment had recovered greatly,with pathological changes in the injured optic nerve also noticeably improved.These results suggest that human brain-derived neurotrophic factor or umbilical cord blood-derived stem cell intervention promotes viscoelasticity recovery of injured optic nerves,and thereby contributes to nerve recovery.     

Transplantation of placenta-derived mesenchymal stem cell-induced neural stem cells to treat spinal cord injury

Because of their strong proliferative capacity and multi-potency, placenta-derived mesenchymal stem cells have gained interest as a cell source in the field of nerve damage repair. In the present study, human placenta-derived mesenchymal stem ceils were induced to differentiate into neural stem cells, which were then transplanted into the spinal cord after local spinal cord injury in rats. The motor functional recovery and pathological changes in the injured spinal cord were observed for 3 successive weeks. The results showed that human placenta-derived mesenchymal stem cells can differentiate into neuron-like cells and that induced neural stem cells contribute to the restoration of injured spinal cord without causing transplant rejection. Thus, these cells promote the recovery of motor and sensory functions in a rat model of spinal cord injury. Therefore, human placenta-derived mesenchymal stem cells may be useful as seed cells during the repair of spinal cord injury.     

Combination of olfactory ensheathing cells and human umbilical cord mesenchymal stem cell-derived exosomes promotes sciatic nerve regeneration

Olfactory ensheathing cells(OECs)are promising seed cells for nerve regeneration.However,their application is limited by the hypoxic environment usually present at the site of injury.Exosomes derived from human umbilical cord mesenchymal stem cells have the potential to regulate the pathological processes that occur in response to hypoxia.The ability of OECs to migrate is unknown,especially in hypoxic conditions,and the effect of OECs combined with exosomes on peripheral nerve repair is not clear.Better understanding of these issues will enable the potential of OECs for the treatment of nerve injury to be addressed.In this study,OECs were acquired from the olfactory bulb of Sprague Dawley rats.Human umbilical cord mesenchymal stem cell-derived exosomes(0–400μg/mL)were cultured with OECs for 12–48 hours.After culture with 400μg/mL exosomes for 24 hours,the viability and proliferation of OECs were significantly increased.We observed changes to OECs subjected to hypoxia for 24 hours and treatment with exosomes.Exosomes significantly promoted the survival and migration of OECs in hypoxic conditions,and effectively increased brain-derived neurotrophic factor gene expression,protein levels and secretion.Finally,using a 12 mm left sciatic nerve defect rat model,we confirmed that OECs and exosomes can synergistically promote motor and sensory function of the injured sciatic nerve.These findings show that application of OECs and exosomes can promote nerve regeneration and functional recovery.This study was approved by the Institutional Ethical Committee of the Air Force Medical University,China(approval No.IACUC-20181004)on October 7,2018;and collection and use of human umbilical cord specimens was approved by the Ethics Committee of the Linyi People’s Hospital,China(approval No.30054)on May 20,2019.     

Use of hybrid chitosan membranes and human mesenchymal stem cells from the Wharton jelly of umbilical cord for promoting nerve regeneration in an axonotmesis rat model

Many studies have been dedicated to the development of scaffolds for improving post-traumatic nerve regeneration. The goal of this study was to assess the effect on nerve regeneration, associating a hybrid chitosan membrane with non-differentiated human mesenchymal stem cells isolated from Wharton’s jelly of umbilical cord, in peripheral nerve reconstruction after crush injury. Chromosome analysis on human mesenchymal stem cell line from Wharton’s jelly was carried out and no structural alterations were found in metaphase. Chitosan membranes were previously tested in vitro, to assess their ability in supporting human mesenchymal stem cell survival, expansion, and differentiation. For the in vivo testing, Sasco Sprague adult rats were divided in 4 groups of 6 or 7 animals each:Group 1, sciatic axonotmesis injury without any other intervention (Group 1-Crush); Group 2, the axonotmesis lesion of 3 mm was infiltrated with a suspension of 1 250-1 500 human mesenchymal stem cells (total volume of 50 μL) (Group 2-CrushCell); Group 3, axonotmesis lesion of 3 mm was enwrapped with a chitosan type III membrane covered with a monolayer of non-differentiated human mesenchymal stem cells (Group 3-CrushChitIIICell) and Group 4, axonotmesis lesion of 3 mm was enwrapped with a chitosan type III membrane (Group 4-CrushChitIII). Motor and sensory functional recovery was evaluated throughout a healing period of 12 weeks using sciatic functional index, static sciatic index, extensor postural thrust, and withdrawal reflex latency. Stereological analysis was carried out on regenerated nerve fibers. Results showed that infiltration of human mesenchymal stem cells, or the combination of chitosan membrane enwrapment and human mesenchymal stem cell enrichment after nerve crush injury provide a slight advantage to post-traumatic nerve regeneration. Results obtained with chitosan type III membrane alone confirmed that they significantly improve post-traumatic axonal regrowth and may represent a very promising clinical tool in peripheral nerve reconstructive surgery. Yet, umbilical cord human mesenchymal stem cells, that can be expanded in culture and induced toform several different types of cells, may prove, in future experiments, to be a new source of cells for cell therapy, including targets such as peripheral nerve and muscle.     

Collagen scaffold combined with human umbilical cord-mesenchymal stem cells transplantation for acute complete spinal cord injury

Currently, there is no effective strategy to promote functional recovery after a spinal cord injury. Collagen scaffolds can not only provide support and guidance for axonal regeneration, but can also serve as a bridge for nerve regeneration at the injury site. They can additionally be used as carriers to retain mesenchymal stem cells at the injury site to enhance their effectiveness. Hence, we hypothesized that transplanting human umbilical cord-mesenchymal stem cells on collagen scaffolds would enhance healing following acute complete spinal cord injury. Here, we test this hypothesis through animal studies and a phase I clinical trial.(1) Animal experiments: Models of completely transected spinal cord injury were established in rats and canines by microsurgery. Mesenchymal stem cells derived from neonatal umbilical cord tissue were adsorbed onto collagen scaffolds and surgically implanted at the injury site in rats and canines;the animals were observed after 1 week–6 months. The transplantation resulted in increased motor scores, enhanced amplitude and shortened latency of the motor evoked potential, and reduced injury area as measured by magnetic resonance imaging.(2) Phase I clinical trial: Forty patients with acute complete cervical injuries were enrolled at the Characteristic Medical Center of Chinese People's Armed Police Force and divided into two groups. The treatment group(n = 20) received collagen scaffolds loaded with mesenchymal stem cells derived from neonatal umbilical cordtissues;the control group(n = 20) did not receive the stem-cell loaded collagen implant. All patients were followed for 12 months. In the treatment group, the American Spinal Injury Association scores and activities of daily life scores were increased, bowel and urinary functions were recovered, and residual urine volume was reduced compared with the pre-treatment baseline. Furthermore, magnetic resonance imaging showed that new nerve fiber connections were formed, and diffusion tensor imaging showed that electrophysiological activity was recovered after the treatment. No serious complication was observed during follow-up. In contrast, the neurological functions of the patients in the control group were not improved over the follow-up period. The above data preliminarily demonstrate that the transplantation of human umbilical cord-mesenchymal stem cells on a collagen scaffold can promote the recovery of neurological function after acute spinal cord injury. In the future, these results need to be confirmed in a multicenter, randomized controlled clinical trial with a larger sample size. The clinical trial was approved by the Ethics Committee of the Characteristic Medical Center of Chinese People's Armed Police Force on February 3, 2016(approval No. PJHEC-2016-A8). All animal experiments were approved by the Ethics Committee of the Characteristic Medical Center of Chinese People's Armed Police Force on May 20, 2015(approval No. PJHEC-2015-D5).     

Transplantation of mesenchymal stem cells from human umbilical cord versus human umbilical cord blood for peripheral nerve regeneration

BACKGROUND: Mesenchymal stem cells (MSCs) appear to be a good alternative to Schwann cells in the treatment of peripheral nerve injury. Fetal stem cells, like umbilical cord blood (UCB) and umbilical cord (UC) stem cells, have several advantages over adult stem cells.OBJECTIVE: To assess the effects of UC-derived MSCs (UCMSCs) and UCB-derived MSCs (UCBMSCs) in repair of sciatic nerve defects. DESIGN, TIME AND SETTING: A randomized controlled animal experiment was performed at the laboratory of Department of Oral and Maxillofacial Surgery, Seoul National University Dental Hospital, from July to December 2009. MATERIALS: UCMSCs were provided by the Research Institute of Biotechnology, Dongguk University. UCBMSCs were provided by the Laboratory of Stem Cells and Tumor Biology, College of Veterinary Medicine, Seoul National University. Dulbecco's modified Eagle's medium (DMEM) was purchased from Gibco-BRL, USA. METHODS: Seven-week-old Sprague-Dawley rats were randomly and evenly divided into three groups: DMEM, UCBMSCs, and UCMSCs. A 10-mm defect in the left sciatic nerve was constructed in all rats. DMEM (15 μL) containing 1 × 106 UCBMSCs or UCMSCs was injected into the gap between nerve stumps, with the surrounding epineurium as a natural conduit. For the DMEM group, simple DMEM was injected. MAIN OUTCOME MEASURES: At 7 weeks after sciatic nerve dissection, dorsal root ganglia neurons were labeled by fluorogold retrograde labeling. At 8 weeks, electrophysiology and histomorphometry were performed. At 2, 4, 6, and 8 weeks after surgery, sciatic nerve function was evaluated using gait analysis.RESULTS: The UCBMSCs group and the UCMSCs group exhibited similar sciatic nerve function and electrophysiological indices, which were better than the DMEM group, as measured by gait analysis (P < 0.05). Fluorogold retrograde labeling of sciatic nerve revealed that the UCBMSCs group demonstrated a higher number of labeled neurons; however, the differences were not significant. Histomorphometric indices were similar in the UCBMSCs and UCMSCs groups, and total axon counts, particularly axon density (P < 0.05), were significantly greater in the UCBMSCs and UCMSCs groups than in the DMEM group. CONCLUSION: Transplanting either UCBMSCs or UCMSCs into axotomized sciatic nerves could accelerate and promote sciatic nerve regeneration over 8 weeks. Both treatments had similar effects on nerve regeneration.     

HRP逆行示踪评价复合骨髓间充质干细胞的无细胞神经移植物

背景：作者前期将无细胞神经移植物与骨髓间充质干细胞复合培养,成功构建了组织工程人工神经。 目的：应用辣根过氧化物酶(HRP)神经逆行示踪技术对无细胞神经移植物复合骨髓间充质干细胞构建的神经移植复合体桥接大鼠坐骨神经缺损后运动神经元的保护作用进行评价。 方法：成年清洁级健康雄性SD大鼠,随机分成3组：①实验组：采用复合骨髓间充质干细胞的无细胞神经移植物桥接大鼠坐骨神经缺损。②空白对照组：采用无细胞神经移植物桥接大鼠坐骨神经缺损。③自体神经对照组：采用自体神经移植桥接大鼠坐骨神经缺损。术后12周应用辣根过氧化物酶神经逆行示踪技术对脊髓前角运动神经元的再生进行评价。 结果与结论：术后12周脊髓前角运动神经元再生评价结果显示：实验组优于无细胞神经移植物组,而与自体神经移植物组相比差异无显著性意义。证实无细胞神经移植物复合骨髓间充质干细胞构建组织工程人工神经修复大鼠坐骨神经缺损,对大鼠脊髓运动神经元具有保护作用,可能达到与自体神经移植相似的效果。 关键词：无细胞神经移植物;骨髓间充质干细胞;辣根过氧化物酶;神经移植;大鼠     

Human umbilical cord blood stem cells and brain-derived neurotrophic factor for optic nerve injury:a biomechanical evaluation

Treatment for optic nerve injury by brain-derived neurotrophic factor or the transplantation of human umbilical cord blood stem cells has gained progress, but analysis by biomechanical indicators is rare. Rabbit models of optic nerve injury were established by a clamp. At 7 days after injury, the vitreous body received a one-time injection of 50 μg brain-derived neurotrophic factor or 1 × 106 human umbilical cord blood stem cells. After 30 days, the maximum load, maximum stress, maximum strain, elastic limit load, elastic limit stress, and elastic limit strain had clearly improved in rabbit models of optical nerve injury after treatment with brain-derived neurotrophic factor or human umbilical cord blood stem cells. The damage to the ultrastructure of the optic nerve had also been reduced. These findings suggest that human umbilical cord blood stem cells and brain-derived neurotrophic factor effectively repair the injured optical nerve, improve biomechanical properties, and contribute to the recovery after injury.     

The interaction of stem cells and vascularity in peripheral nerve regeneration

The degree of nerve regeneration after peripheral nerve injury can be altered by the microenvironment at the site of injury. Stem cells and vascularity are postulated to be a part of a complex pathway that enhances peripheral nerve regeneration; however, their interaction remains unexplored. This review aims to summarize current knowledge on this interaction, including various mechanisms through which trophic factors are promoted by stem cells and angiogenesis. Angiogenesis after nerve injury is stimulated by hypoxia, mediated by vascular endothelial growth factor, resulting in the growth of preexisting vessels into new areas. Modulation of distinct signaling pathways in stem cells can promote angiogenesis by the secretion of various angiogenic factors. Simultaneously, the importance of stem cells in peripheral nerve regeneration relies on their ability to promote myelin formation and their capacity to be influenced by the microenvironment to differentiate into Schwann-like cells. Stem cells can be acquired through various sources that correlate to their differentiation potential, including embryonic stem cells, neural stem cells, and mesenchymal stem cells. Each source of stem cells serves its particular differentiation potential and properties associated with the promotion of revascularization and nerve regeneration. Exosomes are a subtype of extracellular vesicles released from cell types and play an important role in cell-to-cell communication. Exosomes hold promise for future transplantation applications, as these vesicles contain fewer membrane-bound proteins, resulting in lower immunogenicity. This review presents pre-clinical and clinical studies that focus on selecting the ideal type of stem cell and optimizing stem cell delivery methods for potential translation to clinical practice. Future studies integrating stem cell-based therapies with the promotion of angiogenesis may elucidate the synergistic pathways and ultimately enhance nerve regeneration.     

Combining acellular nerve allografis with brain- derived neurotrophic factor transfected bone marrow mesenchymal stem cells restores sciatic nerve injury better than either intervention alone

In this study, we chemically extracted acellular nerve allografts from bilateral sciatic nerves, and repaired 10-mm sciatic nerve defects in rats using these grafts and brain-derived neurotrophic factor transfected bone marrow mesenchymal stem cells. Experiments were performed in three groups： the acellular nerve allograft bridging group, acellular nerve allograft ＋ bone marrow mesenchymal stem cells group, and the acellular nerve allograft ＋ brain-derived neurotrophic factor transfected bone marrow mesenchyrnal stem cells group. Results showed that at 8 weeks after bridging, sciatic functional index, triceps wet weight recovery rate, myelin thickness, and number of myelinated nerve fibers were significantly changed in the three groups. Variations were the largest in the acellular nerve allograft ＋ brain-derived neurotrophic factor transfected bone marrow mesenchymal stem cells group compared with the other two groups. Experimental findings suggest that chemically extracted acellular nerve allograft combined nerve factor and mesenchymal stem cells can promote the restoration of sciatic nerve defects. The repair effect seen is better than the single application of acellular nerve allograft or acellular nerve allograft combined mesenchymal stem cell transplantation.     

Functional recovery and microenvironmental alterations in a rat model of spinal cord injury following human umbilical cord blood-derived mesenchymal stem cells transplantation

BACKGROUND: Transplantation of human umbilical cord blood-derived mesenchymal stem cells (MSCs) has been shown to benefit spinal cord injury (SCI) repair. However, mechanisms of microenvironmental regulation during differentiation of transplanted MSCs remain poorly understood. OBJECTIVE: To observe changes in nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and interleukin-8 (IL-8) expression following transplantation of human umbilical cord-derived MSCs, and to explore the association between microenvironment and neural functional recovery following MSCs transplantation.DESIGN, TIME AND SETTING: A randomized, controlled, animal experiment was performed at the Department of Orthopedics, First Affiliated Hospital of Soochow University from April 2005 to March 2007. MATERIALS: Human cord blood samples were provided by the Department of Gynecology and Obstetrics, First Affiliated Hospital of Soochow University. Written informed consent was obtained. METHODS: A total of 62 Wister rats were randomly assigned to control (n = 18), model (n = 22, SCI + PBS), and transplantation (n = 22, SCI + MSCs) groups. The rat SCI model was established using the weight compression method. MSCs were isolated from human umbilical cord blood and cultured in vitro for several passages. 5-bromodeoxyuridine (BrdU)-labeled MSCs (24 hours before injection) were intravascularly transplanted. MAIN OUTCOME MEASURES: The rats were evaluated using the Basso, Beattie and Bresnahan (BBB) locomotor score and inclined plane tests. Transplanted cells were analyzed following immunohistochemistry. Enzyme-linked immunosorbant assay was performed to determine NGF, BDNF, and IL-8 levels prior to and after cell transplantation.RESULTS: A large number of BrdU-positive MSCs were observed in the SCI region of the transplantation group, and MSCs were evenly distributed in injured spinal cord tissue 1 week after transplantation. BBB score and inclined plane test results revealed significant functional improvement in the transplantation group compared to the model group (P< 0.05), which was maintained for 2-3 weeks. Compared to the model group, NGF and BDNF levels were significantly increased in the injured region following MSCs transplantation at 3 weeks (P < 0.05), but IL-8 levels remained unchanged (P > 0.05).CONCLUSION: MSCs transplantation increased NGF and BDNF expression in injured spinal cord tissue. MSCs could promote neurological function recovery in SCI rats by upregulating NGF expression and improving regional microenvironments.     

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     

靶肌肉注射人间充质干细胞对坐骨神经损伤保护性作用的电生理研究

目的探讨电生理在大鼠失神经支配的骨骼肌中移植间充质干细胞对坐骨神经损伤后修复中的作用。方法体外分离培养人间充质干细胞,经鉴定标记后备用。将36只SD大鼠随机分为肌肉注射干细胞组(实验组)、肌肉注射生理盐水组(对照组),每组18只,建立大鼠坐骨神经损伤模型,术后3d将干细胞和等量生理盐水注射到损伤侧坐骨神经支配的骨骼肌中。于移植后3、7、14、21、28、60d,观察坐骨神经功能指数(SFI)、腓肠肌肌电图(EMG)、运动神经传导速度(MCV),肌肉复合动作电位幅值(CMAP)检查。结果腓肠肌EMG实验组自发电位减少和动作电位出现时间均早于对照组。坐骨神经MCV和CMAP波幅呈恢复趋势。移植后:21d开始,实验组明显快于对照组(P<0.05),60d时,传导速度两组无显著性(P>0.05),波幅有显著性差异(P<0.05)。结论肌肉注射人间充质干细胞对坐骨神经损伤具有促修复作用。     

Adipose-derived mesenchymal stem cells accelerate nerve regeneration and functional recovery in a rat model of recurrent laryngeal nerve injury

Medialization thyroplasty or injection laryngoplasty for unilateral vocal fold paralysis cannot restore mobility of the vocal fold. Recent studies have shown that transplantation of mesenchymal stem cells is effective in the repair of nerve injuries. This study investigated whether adipose-derived stem cell transplantation could repair recurrent laryngeal nerve injury. Rat models of recurrent laryngeal nerve injury were established by crushing with micro forceps. Adipose-derived mesenchymal stem cells(ADSCs; 8 × 105) or differentiated Schwann-like adipose-derived mesenchymal stem cells(d ADSCs; 8 × 105) or extracellular matrix were injected at the site of injury. At 2, 4 and 6 weeks post-surgery, a higher density of myelinated nerve fiber, thicker myelin sheath, improved vocal fold movement, better recovery of nerve conduction capacity and reduced thyroarytenoid muscle atrophy were found in ADSCs and d ADSCs groups compared with the extracellular matrix group. The effects were more pronounced in the ADSCs group than in the d ADSCs group. These experimental results indicated that ADSCs transplantation could be an early interventional strategy to promote regeneration after recurrent laryngeal nerve injury.     

Human umbilical cord Wharton’s jelly-derived oligodendrocyte precursor-like cells for axon and myelin sheath regeneration

Human umbilical mesenchymal stem cells from Wharton’s jelly of the umbilical cord were induced to differentiate into oligodendrocyte precursor-like cells in vitro. Oligodendrocyte precursor cells were transplanted into contused rat spinal cords. Immunofluorescence double staining indicated that transplanted cells survived in injured spinal cord, and differentiated into mature and immature oligodendrocyte precursor cells. Biotinylated dextran amine tracing results showed that cell transplantation promoted a higher density of the corticospinal tract in the central and caudal parts of the injured spinal cord. Luxol fast blue and toluidine blue staining showed that the volume of residual myelin was significantly increased at 1 and 2 mm rostral and caudal to the lesion epicenter after cell transplantation. Furthermore, immunofluorescence staining verified that the newly regenerated myelin sheath was derived from the central nervous system. Basso, Beattie and Bresnahan testing showed an evident behavioral recovery. These results suggest that human umbilical mesenchymal stem cell-derived oligodendrocyte precursor cells promote the regeneration of spinal axons and myelin sheaths.     

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。     

Transplantation of human umbilical cord blood mesenchymal stem cells to treat a rat model of traumatic brain injury

In the present study, human umbilical cord blood mesenchymal stem cells were injected into a rat model of traumatic brain injury via the tail vein. Results showed that 5-bromodeoxyuridine-labeled cells aggregated around the injury site, surviving up to 4 weeks post-transplantation. In addition, transplantation-related death did not occur, and neurological functions significantly improved. Histological detection revealed attenuated pathological injury in rat brain tissues following human umbilical cord blood mesenchymal stem cell transplantation. In addition, the number of apoptotic cells decreased. Immunohistochemistry and in situ hybridization showed increased expression of brain-derived neurotrophic factor, nerve growth factor, basic fibroblast growth factor, and vascular endothelial growth factor, along with increased microvessel density in surrounding areas of brain injury. Results demonstrated migration of transplanted human umbilical cord blood mesenchymal stem cells into the lesioned boundary zone of rats, as well as increased angiogenesis and expression of related neurotrophic factors in the lesioned boundary zone.     

无细胞神经移植物复合骨髓间充质干细胞构建组织工程人工神经修复坐骨神经缺损

背景：作者前期已经成功将无细胞神经移植物复合骨髓间充质干细胞构建组织工程人工神经,并证明可以促进周围神经再生。 目的：构建组织工程人工神经,观察和验证桥接大鼠坐骨神经缺损后的神经功能恢复情况。 方法：成年雄性SD大鼠60只构建大鼠坐骨神经15 mm缺损模型。随机分成3组,每组20只。桥接大鼠坐骨神经缺损,实验组采用组织工程人工神经,空白对照组采用无细胞组织工程神经支架,自体神经对照组采用自体神经移植。桥接后12周通过大体观察、胫骨前肌湿质量、组织学等方法分析坐骨神经组织学及功能恢复情况。 结果与结论：桥接术后12周：实验组大鼠肢体可以支撑着地,钳夹大鼠手术侧足底皮肤出现逃避反射,足底皮肤s-100蛋白染色呈阳性反应。实验组与自体神经移植组胫骨前肌湿质量比差异无显著性意义(P > 0.05)。实验组辣根过氧化物酶逆行示踪实验显示脊髓、后根神经节均可见数量不等的辣根过氧化物酶标记阳性细胞。实验组移植物与自体神经移植组有髓神经纤维数、髓鞘厚度、神经组织面积比较差异无显著性意义。实验结果验证了无细胞神经移植物复合骨髓间充质干细胞构建组织工程人工神经修复大鼠坐骨神经缺损,可以促进神经组织学的修复重建和功能的恢复。