Human umbilical cord blood-derived mesenchymal stem cells promote regeneration of crush-injured rat sciatic nerves

Several studies have demonstrated that human umbilical cord blood-derived mesenchymal stem cells can promote neural regeneration following brain injury. However, the therapeutic effects of human umbilical cord blood-derived mesenchymal stem cells in guiding peripheral nerve regeneration remain poorly understood. This study was designed to investigate the effects of human umbilical cord blood-derived mesenchymal stem cells on neural regeneration using a rat sciatic nerve crush injury model. Human umbilical cord blood-derived mesenchymal stem cells (1 × 10 6 ) or a PBS control were injected into the crush-injured segment of the sciatic nerve. Four weeks after cell injection, brain-derived neurotrophic factor and tyrosine kinase receptor B mRNA expression at the lesion site was increased in comparison to control. Furthermore, sciatic function index, Fluoro Gold-labeled neuron counts and axon density were also significantly increased when compared with control. Our results indicate that human umbilical cord blood-derived mesenchymal stem cells promote the functional recovery of crush-injured sciatic nerves.     

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.     

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。     

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).     

Time course analysis of sensory axon regeneration in vivo by directly tracing regenerating axons

Most current studies quantify axon regeneration by immunostaining regeneration-associated proteins,representing indirect measurement of axon lengths from both sensory neurons in the dorsal root ganglia and motor neurons in the spinal cord.Our recently developed method of in vivo electroporation of plasmid DNA encoding for enhanced green fluorescent protein into adult sensory neurons in the dorsal root ganglia provides a way to directly and specifically measure regenerating sensory axon lengths in whole-mount nerves.A mouse model of sciatic nerve compression was established by squeezing the sciatic nerve with tweezers.Plasmid DNA carrying enhanced green fluorescent protein was transfected by ipsilateral dorsal root ganglion electroporation 2 or 3 days before injury.Fluorescence distribution of dorsal root or sciatic nerve was observed by confocal microscopy.At 12 and 18 hours,and 1,2,3,4,5,and 6 days of injury,lengths of regenerated axons after sciatic nerve compression were measured using green fluorescence images.Apoptosis-related protein caspase-3 expression in dorsal root ganglia was determined by western blot assay.We found that in vivo electroporation did not affect caspase-3 expression in dorsal root ganglia.Dorsal root ganglia and sciatic nerves were successfully removed and subjected to a rapid tissue clearing technique.Neuronal soma in dorsal root ganglia expressing enhanced green fluorescent protein or fluorescent dye-labeled microRNAs were imaged after tissue clearing.The results facilitate direct time course analysis of peripheral nerve axon regeneration.This study was approved by the Institutional Animal Care and Use Committee of Guilin Medical University,China(approval No.GLMC201503010)on March 7,2014.     

无细胞神经支架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：作者已经成功制备了无细胞神经移植物,并且复合骨髓间充质干细胞构建组织工程人工神经桥接大鼠坐骨神经缺损。 目的：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后运动功能的恢复。 方法：成年雄性SD大鼠构建大鼠坐骨神经15 mm缺损模型,分别应用组织工程人工神经、组织工程神经支架或自行神经桥接坐骨神经缺损。桥接后20周再生神经电生理学测定,手术侧胫骨前肌湿质量、腓肠肌组织学及透视电镜分析。 结果与结论：桥接20周后,组织工程人工神经与自体神经移植组胫骨前肌湿质量比较,差异无显著性意义(P > 0.05),神经干传导速度为(30.56±2.15)m/s。结果提示,无细胞神经移植物复合骨髓间充质干细胞构建的组织工程人工神经桥接大鼠坐骨神经缺损后,可以促进再生神经运动功能的恢复。     

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.     

Schwann cell‐derived exosomes enhance axonal regeneration in the peripheral nervous system

Axonal regeneration in the peripheral nervous system is greatly supported by Schwann cells (SCs). After nerve injury, SCs dedifferentiate to a progenitor‐like state and efficiently guide axons to their original target tissues. Contact and soluble factors participate in the crosstalk between SCs and axons during axonal regeneration. Here we show that dedifferentiated SCs secrete nano‐vesicles known as exosomes which are specifically internalized by axons. Surprisingly, SC‐derived exosomes markedly increase axonal regeneration in vitro and enhance regeneration after sciatic nerve injury in vivo. Exosomes shift the growth cone morphology to a pro‐regenerating phenotype and decrease the activity of the GTPase RhoA, involved in growth cone collapse and axon retraction. Altogether, our work identifies a novel mechanism by which SCs communicate with neighboring axons during regenerative processes. We propose that SC exosomes represent an important mechanism by which these cells locally support axonal maintenance and regeneration after nerve damage. GLIA 2013;61:1795–1806     

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.     

Transplantation of olfactory ensheathing cells for promoting regeneration following spinal cord injury

OBJECTIVE: To investigate the status of olfactory ensheathing cells (OECs) transplantation in facilitating the regeneration of spinal cord injury. DATA SOURCES: Articles about OECs transplantation in treating spinal cord injury were searched in Pubmed database published in English from January 1981 to December 2005 by using the keywords of "olfactory ensheathing cells, transplantation, spinal cord injury". STUDY SELECTION: The data were checked primarily, literatures related to OECs transplantation and the regeneration of spinal cord injury were selected, whereas the repetitive studies and reviews were excluded. DATA EXTRACTION: Totally 43 articles about OECs transplantation and the regeneration and repair of spinal cord injury were collected, and the repetitive ones were excluded. DATA SYNTHESIS: There were 35 articles accorded with the criteria. OECs are the olfactory ensheathing glias isolated from olfactory bulb and olfactory nerve tissue. OECs have the characters of both Schwann cells in central nervous system and peripheral astrocytes. The transplanted OECs can migrate in the damaged spinal cord of host, can induce and support the regeneration, growth and extension of damaged neuritis. Besides, transgenic technique can enable it to carry some exogenous genes that promote neuronal regeneration, and express some molecules that can facilitate neural regeneration, so as to ameliorate the internal environment of nerve injury, induce the regeneration of damaged spinal cord neurons, which can stimulate the regeneration potential of the damaged spinal cord to reach the purpose of spinal cord regeneration and functional recovery. CONCLUSION: OECs are the glial cells with the energy for growth at mature phase, they can myelinize axons, secrete various biological nutrition factors, and then protect and support neurons, also facilitate neural regeneration. OECs have been successfully isolated from nasal olfactory mucosa and olfactory nerve. Therefore, autologous transplantation of OECs and objective genes modified OECs carrying various neurotrophic factors may become an effective method to treat spinal cord injury in the future.     

Transplantation of olfactory ensheathing cells for promoting regeneration following spinal cord injury

OBJECTIVE： To investigate the status of olfactory ensheathing cells （OECs） transplantation in facilitating the regeneration of spinal cord injury. DATA SOURCES： Articles about OECs transplantation in treating spinal cord injury were searched in Pubmed database published in English from January 1981 to December 2005 by using the keywords of ＂olfactory ensheathing cells, transplantation, spinal cord injury＂. STUDY SELECTION： The data were checked primarily, literatures related to OECs transplantation and the regeneration of spinal cord injury were selected, whereas the repetitive studies and reviews were excluded. DATA EXTRACTION： Totally 43 articles about OECs transplantation and the regeneration and repair of spinal cord injury were collected, and the repetitive ones were excluded. DATA SYNTHESIS： There were 35 articles accorded with the criteria. OECs are the olfactory ensheathing glias isolated from olfactory bulb and olfactory nerve tissue. OECs have the characters of both Schwann cells in central nervous system and peripheral astrocytes. The transplanted OECs can migrate in the damaged spinal cord of host, can induce and support the regeneration, growth and extension of damaged neuritis. Besides, transgenic technique can enable it to carry some exogenous genes that promote neuronal regeneration, and express some molecules that can facilitate neural regeneration, so as to ameliorate the internal environment of nerve injury, induce the regeneration of damaged spinal cord neurons, which can stimulate the regeneration potential of the damaged spinal cord to reach the purpose of spinal cord regeneration and functional recovery. CONCLUSION： OECs are the glial cells with the energy for growth at mature phase, they can myelinize axons, secrete various biological nutrition factors, and then protect and support neurons, also facilitate neural regeneration. OECs have been successfully isolated from nasal olfactory mucosa and olfactory nerve. Therefore, autologous transplantation of OECs and objective genes modified OECs carrying various neurotrophic factors may become an effective method to treat spinal cord injury in the future.     

Biological conduits combining bone marrow mesenchymal stem cells and extracellular matrix to treat long-segment sciatic nerve defects

The transplantation of polylactic glycolic acid conduits combining bone marrow mesenchymal stem cells and extracellular matrix gel for the repair of sciatic nerve injury is effective in some respects, but few data comparing the biomechanical factors related to the sciatic nerve are available. In the present study, rabbit models of 10-mm sciatic nerve defects were prepared. The rabbit models were repaired with autologous nerve, a polylactic glycolic acid conduit + bone marrow mesenchymal stem cells, or a polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel. After 24 weeks, mechanical testing was performed to determine the stress relaxation and creep parameters. Following sciatic nerve injury, the magnitudes of the stress decrease and strain increase at 7,200 seconds were largest in the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel group, followed by the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells group, and then the autologous nerve group. Hematoxylin-eosin staining demonstrated that compared with the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells group and the autologous nerve group, a more complete sciatic nerve regeneration was found, including good myelination, regularly arranged nerve fibers, and a completely degraded and resorbed conduit, in the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel group. These results indicate that bridging 10-mm sciatic nerve defects with a polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel construct increases the stress relaxation under a constant strain, reducing anastomotic tension. Large elongations under a constant physiological load can limit the anastomotic opening and shift, which is beneficial for the regeneration and functional reconstruction of sciatic nerve. Better regeneration was found with the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel grafts than with the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells grafts and the autologous nerve grafts.     

Ascorbic acid accelerates Wallerian degeneration after peripheral nerve injury

Wallerian degeneration occurs after peripheral nerve injury and provides a beneficial microenvironment for nerve regeneration. Our previous study demonstrated that ascorbic acid promotes peripheral nerve regeneration, possibly through promoting Schwann cell proliferation and phagocytosis and enhancing macrophage proliferation, migration, and phagocytosis. Because Schwann cells and macrophages are the main cells involved in Wallerian degeneration, we speculated that ascorbic acid may accelerate this degenerative process. To test this hypothesis, 400 mg/kg ascorbic acid was administered intragastrically immediately after sciatic nerve transection, and 200 mg/kg ascorbic acid was then administered intragastrically every day. In addition, rat sciatic nerve explants were treated with 200 μM ascorbic acid. Ascorbic acid significantly accelerated the degradation of myelin basic protein-positive myelin and neurofilament 200-positive axons in both the transected nerves and nerve explants. Furthermore, ascorbic acid inhibited myelin-associated glycoprotein expression, increased c-Jun expression in Schwann cells, and increased both the number of macrophages and the amount of myelin fragments in the macrophages. These findings suggest that ascorbic acid accelerates Wallerian degeneration by accelerating the degeneration of axons and myelin in the injured nerve, promoting the dedifferentiation of Schwann cells, and enhancing macrophage recruitment and phagocytosis. The study was approved by the Southern Medical University Animal Care and Use Committee(approval No. SMU-L2015081) on October 15, 2015.     

Acellular allogeneic nerve grafting combined with bone marrow mesenchymal stem cell transplantation for the repair of long-segment sciatic nerve defects:biomechanics and validation of mathematical models

We hypothesized that a chemically extracted acellular allogeneic nerve graft used in combination with bone marrow mesenchymal stem cell transplantation would be an effective treatment for long-segment sciatic nerve defects. To test this, we established rabbit models of 30 mm sciatic nerve defects, and treated them using either an autograft or a chemically decellularized allogeneic nerve graft with or without simultaneous transplantation of bone marrow mesenchymal stem cells. We compared the tensile properties, electrophysiological function and morphology of the damaged nerve in each group. Sciatic nerves repaired by the allogeneic nerve graft combined with stem cell trans-plantation showed better recovery than those repaired by the acellular allogeneic nerve graft alone, and produced similar results to those observed with the autograft. These ifndings conifrm that a chemically extracted acellular allogeneic nerve graft combined with transplanta-tion of bone marrow mesenchymal stem cells is an effective method of repairing long-segment sciatic nerve defects.     

Craniocerebral injury promotes the repair of peripheral nerve injury

The increase in neurotrophic factors after craniocerebral injury has been shown to promote fracture healing. Moreover, neurotrophic factors play a key role in the regeneration and repair of peripheral nerve. However, whether craniocerebral injury alters the repair of peripheral nerve injuries remains poorly understood. Rat injury models were established by transecting the left sciatic nerve and using a free-fall device to induce craniocerebral injury. Compared with sciatic nerve injury alone after 6–12 weeks, rats with combined sciatic and craniocerebral injuries showed decreased sciatic functional index, increased recovery of gastrocnemius muscle wet weight, recovery of sciatic nerve ganglia and corresponding spinal cord segment neuron morphologies, and increased numbers of horseradish peroxidase-labeled cells. These results indicate that craniocerebral injury promotes the repair of peripheral nerve injury.     

Dynamic expression of Slit1–3 and Robo1–2 in the mouse peripheral nervous system after injury

The Slit family of axon guidance cues act as repulsive molecules for precise axon pathfinding and neuronal migration during nervous system development through interactions with specific Robo receptors.Although we previously reported that Slit1–3 and their receptors Robo1 and Robo2 are highly expressed in the adult mouse peripheral nervous system,how this expression changes after injury has not been well studied.Herein,we constructed a peripheral nerve injury mouse model by transecting the right sciatic nerve.At 14 days after injury,quantitative real-time polymerase chain reaction was used to detect mRNA expression of Slit1–3 and Robo1–2 in L4–5 spinal cord and dorsal root ganglia,as well as the sciatic nerve.Immunohistochemical analysis was performed to examine Slit1–3,Robo1–2,neurofilament heavy chain,F4/80,and vimentin in L4–5 spinal cord,L4 dorsal root ganglia,and the sciatic nerve.Co-expression of Slit1–3 and Robo1–2 in L4 dorsal root ganglia was detected by in situ hybridization.In addition,Slit1–3 and Robo1–2 protein expression in L4–5 spinal cord,L4 dorsal root ganglia,and sciatic nerve were detected by western blot assay.The results showed no significant changes of Slit1–3 or Robo1–2 mRNA expression in the spinal cord within 14 days after injury.In the dorsal root ganglion,Slit1–3 and Robo1–2 mRNA expression were initially downregulated within 4 days after injury;however,Robo1–2 mRNA expression returned to the control level,while Slit1–3 mRNA expression remained upregulated during regeneration from 4–14 days after injury.In the sciatic nerve,Slit1–3 and their receptors Robo1–2 were all expressed in the proximal nerve stump;however,Slit1,Slit2,and Robo2 were barely detectable in the nerve bridge and distal nerve stump within 14 days after injury.Slit3 was highly ex-pressed in macrophages surrounding the nerve bridge and slightly downregulated in the distal nerve stump within 14 days after injury.Robo1 was upregulated in vimentin-positive cells and migrating Schwann cells inside the nerve bridge.Robo1 was also upregulated in Schwann cells of the distal nerve stump within 14 days after injury.Our findings indicate that Slit3 is the major ligand expressed in the nerve bridge and distal nerve stump during peripheral nerve regeneration,and Slit3/Robo signaling could play a key role in peripheral nerve repair after injury.This study was approved by Plymouth University Animal Welfare Ethical Review Board (approval No.30/3203) on April 12,2014.     

Robust temporal changes of cellular senescence and proliferation after sciatic nerve injury

Cellular senescence and proliferation are essential for wound healing and tissue remodeling.However,senescence-proliferation cell fate after peripheral nerve injury has not been clearly revealed.Here,post-injury gene expression patterns in rat sciatic nerve stumps(SRP113121)and L4–5 dorsal root ganglia(SRP200823)obtained from the National Center for Biotechnology Information were analyzed to decipher cellular senescence and proliferation-associated genetic changes.We first constructed a rat sciatic nerve crush model.Then,β-galactosidase activities were determined to indicate the existence of cellular senescence in the injured sciatic nerve.Ki67 and EdU immunostaining was performed to indicate cellular proliferation in the injured sciatic nerve.Both cellular senescence and proliferation were less vigorous in the dorsal root ganglia than in sciatic nerve stumps.These results reveal the dynamic changes of injury-induced cellular senescence and proliferation from both genetic and morphological aspects,and thus extend our understanding of the biological processes following peripheral nerve injury.The study was approved by the Animal Ethics Committee of Nantong University,China(approval No.20190226-001)on February 26,2019.