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Spinal Cord Injury (SCI) is a debilitating disease associated with a significant economic burden owing to its high level of disability; however, current treatment options have only limited efficacy. Past research has shown that iron-dependent programmed cell death, also known as ferroptosis, plays a critical role in the pathogenesis of SCI. The sigma-1 receptor (Sig-1R) is widely distributed in the central nervous system, and has been implicated in the pathophysiology of several neurological and psychiatric disorders. Several in vivo and ex vivo studies have shown that Sig-1R activation exerts unique neuroprotective effects. However, the underlying mechanisms remain unclear. To date, no study has yet demonstrated the association between Sig-1R activation and ferroptosis in patients with SCI. However, the present study found that Sig-1R activation effectively promoted the recovery of motor function in mice after spinal cord injury, attenuated neuronal apoptosis, reduced the production of pro-inflammatory cytokines and iron accumulation, and inhibited ferroptosis in spinal cord tissues following SCI in mice. Ferroptosis and IRE1α were significantly upregulated after spinal cord injury, while sigma-1 receptor agonists were able to facilitate this result through the elimination of inositol-requiring enzyme-1 alpha (IRE1α)-mediated neuronal ferroptosis. Therefore, sigma-1 receptor activation could attenuate ferroptosis after SCI by reducing IRE1α and improving functional recovery after SCI, potentially representing a new therapeutic strategy for treating SCI., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Spinal cord injury (SCI) is a serious traumatic disease with no effective treatment. This study aimed to explore the therapeutic effect of syringaresinol on SCI. First, the potential targets and associated signaling pathways of syringaresinol were predicted by bioinformatics analysis and molecular docking. Second, MTT was employed to evaluate cell proliferation rate, Western blot was performed to detect protein expression, RT-qPCR was conducted to detect mRNA expression levels, flow cytometry and 5-ethynyl-2'-deoxyuridine (EDU) staining were used to determine cell apoptosis, and immunofluorescence and immunohistochemistry were used to estimate the expression of RNA binding fox-1 homolog 3 and clipped caspase 3. Basso-Beattie-Bresnahan scores and inclined plate tests were conducted to analyze hindlimb locomotor function. Results showed that syringaresinol could inhibit the apoptosis of glutamate-treated SHSY5Y cells by upregulating the expression of ubiquitination factor E4B (UBE4B) and activating the AKT serine/threonine kinase (AKT) signaling pathway. This effect can be rescued by UBE4B knockdown or AKT pathway inhibition. Syringaresinol remarkably improved locomotor function and increased neuronal survival in SCI rats. Our results suggested that syringaresinol could promote locomotor functional recovery by reducing neuronal apoptosis by activating the UBE4B/AKT signaling pathway., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

The specific molecular mechanism of neuroprotective effects of wnt-3a on spinal cord injury (SCI) has not been elucidated. In our study, we evaluated the recovery of motor function after SCI by BBB, observed neuronal apoptosis by western blot and TUNEL, observed the changes of neuronal inflammation by western blot and immunofluorescence staining, and observed the changes of motoneurons and spinal cord area in the anterior horn of the spinal cord via Nissl and HE staining. We found that wnt-3a could significantly promote the recovery of motor function, reduce the loss of motor neurons in the anterior horn of the spinal cord, promote the recovery of injured spinal cord tissue, inhibit neuronal apoptosis and inflammatory response, and ultimately promote neuronal function after SCI. However, when XAV939 inhibits the wnt/β-catenin signaling pathway, the neuroprotective effects of wnt-3a are also significantly inhibited. The above results together indicated that wnt-3a exerts its neuroprotective effect on after SCI via activating the wnt/β-catenin signaling pathway., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Extracellular matrix (ECM) is a complex and dynamic network of proteoglycans, proteins, and other macromolecules that surrounds cells in tissues. The ECM provides structural support to cells and plays a critical role in regulating various cellular functions. ECM remodeling is a dynamic process involving the breakdown and reconstruction of the ECM. This process occurs naturally during tissue growth, wound healing, and tissue repair. However, in the context of central nervous system (CNS) injuries, dysregulated ECM remodeling can lead to the formation of fibrotic and glial scars. CNS injuries encompass various traumatic events, including concussions and fractures. Following CNS trauma, the formation of glial and fibrotic scars becomes prominent. Glial scars primarily consist of reactive astrocytes, while fibrotic scars are characterized by an abundance of ECM proteins. ECM remodeling plays a pivotal and tightly regulated role in the development of these scars after spinal cord and brain injuries. Various factors like ECM components, ECM remodeling enzymes, cell surface receptors of ECM molecules, and downstream pathways of ECM molecules are responsible for the remodeling of the ECM. The aim of this review article is to explore the changes in ECM during normal physiological conditions and following CNS injuries. Additionally, we discuss various approaches that target various factors responsible for ECM remodeling, with a focus on promoting axon regeneration and functional recovery after CNS injuries. By targeting ECM remodeling, it may be possible to enhance axonal regeneration and facilitate functional recovery after CNS injuries., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Spinal cord injury (SCI) can trigger multiple forms of neuronal cell death. Among these, ferroptosis stands out as a particularly important style of cell death due to its iron overload-dependent lipid peroxidative regulatory mechanism. The guanine-rich RNA sequence binding factor 1 (GRSF1) is an RNA-binding protein that has been implicated in cellular senescence, mitochondrial function, oxidative stress, erythropoiesis, and embryonic brain development. However, the function of GRSF1 in neuronal ferroptosis after SCI remains unclear. Here, we established a SCI rat model in vivo and evaluated the function of GRSF1 on neuronal ferroptosis by inhibiting and overexpressing GRSF1. We firstly verified the protein expression of GRSF1 and GPX4 at different time points after SCI. According of changes in expression, we chose 3 d post SCI to assess the effect of GRSF1 on ferroptosis. We found that GRSF1 expression decreased after SCI. In addition, GRSF1 was mainly localized in the cytoplasm of neurons. The results also showed that overexpression of GRSF1 promoted recovery of neurological functional after SCI. Further investigation revealed that GRSF1 might attenuate neuronal ferroptosis by regulating the GPX4 protein expression levels. In summary, our findings indicate that GRSF1 attenuates injury in SCI and reduces neuron ferroptosis and promotes functional recovery via GPX4., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

Neuroinflammation plays an important role in secondary injury after spinal cord injury (SCI) and may aggravate neurological dysfunction. Several studies have indicated that sodium houttuyfonate (SH) can significantly inhibit macrophage- mediated inflammation; however, its effects on SCI still needs to be elucidated. We found that SH improved Basso, Beattie, and Bresnahan scores and performance in the inclined plane test of SCI model rats. The injured spinal cord exhibited less neuronal loss, cell apoptosis, and M1 microglial polarization after SH treatment. In vitro, SH reduced TLR4/NF-κB expression in cultured primary microglia and decreased M1 microglial polarization and cell apoptosis in a lipopolysaccharide (LPS)-pretreated microglia and neuron coculture system. These results indicated that SH may exert a neuroprotective effect by inhibiting M1 microglial polarization after SCI via the TLR4/NF-κB signalling pathway., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Purpose: Activation of muscarinic receptors located in bladder sensory pathways is generally considered to be the primary contributor for driving the pathogenesis of neurogenic detrusor overactivity following spinal cord injury. The present study is undertaken to examine whether moxibustion improves neurogenic detrusor overactivity via modulating the abnormal muscarinic receptor pathway., Materials and Methods: Female Sprague-Dawley rats were subjected to spinal cord injury with T9-10 spinal cord transection. Fourteen days later, animals were received moxibustion treatment for one week. Urodynamic parameters and pelvic afferents discharge were measured. Adenosine triphosphate (ATP) content in the voided cystometry fluid was determined. Expressions of M2, M3, and P2X3 receptors in the bladder mucosa were evaluated., Results: Moxibustion treatment prevented the development of detrusor overactivity in spinal cord injury rats, with an increase in the intercontraction interval and micturition pressure threshold and a decrease in afferent activity during filling. The expression of M2 was markedly suppressed by moxibustion, accompanied by a reduction in the levels of ATP and P2X3. M2 receptor antagonist methoctramine hemihydrate had similar effects to moxibustion on bladder function and afferent activity, while the M2-preferential agonist oxotremorine methiodide abolished the beneficial effects of moxibustion., Conclusion: Moxibustion is a potential candidate for treating neurogenic bladder overactivity in a rat model of spinal cord injury, possibly through inhibiting the M2/ATP/P2X3 pathway., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Glutamate excitotoxicity plays a role in spinal cord injury (SCI). This study aimed to explore whether electroacupuncture (EA) improved the functional recovery of spinal cord anterior horn neurons of rats with acute SCI by regulating the GluR1 AMPA subunit in the SCI area. Eighty Sprague-Dawley rats were randomly divided into 5 groups: sham operation, model, AMPA antagonist (DNQX), EA and DNQX + EA group (n = 16/group). The models were obtained by using the modified Allen's impact method. DNQX was given by intrathecal injection 0.5 h after modeling. EA was performed at the "Dazhui" and "Mingmen" acupoints for 30 min at 0.5, 12, and 24 h. The BBB scores were evaluated before modeling and at 6, 24, and 48 h after modeling. Histopathological changes were evaluated. GluR1 expression was evaluated through immunofluorescence and western blot. Compared to the sham group, the BBB scores at 6, 24, and 48 h in the model group were all lower. The BBB scores and histopathological changes in the EA, DNQX and DNQX + EA group were between that of the sham and model group. GluR1 expression in the model group was higher than the sham group. Compared with the model group, the expression of GluR1 protein in the EA, DNQX, and DNQX + EA group was decreased, but similar among the three treatment groups, supporting the histopathological observations. In conclusion, these findings indicated that EA treatment might inhibit GluR1 expression, thus contributing to prevention of secondary nerve injury after primary acute SCI., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Spinal cord injury (SCI) is a complex central traumatic disease. STAT3 signal transduction pathway plays an important role in SCI. Wogonin has been reported to exhibit neuroprotection. However, the molecular mechanism of its potential therapeutic effect after SCI remains unclear. In this study, rats were divided into the following groups: Sham; SCI; SCI + wogonin; and SCI + wogonin + colivelin (Colivelin is an effective activator of the STAT3 pathway). Motor function was evaluated by Basso Beattie Bresnahan (BBB) score. Histomorphological changes in the spinal cords were observed by Hematoxylin-eosin (HE) staining and Nissl staining. Western blot, Transferase-mediated dUTP-biotin nick end labeling (TUNEL) staining, and immunofluorescence were used to detect changes in the neuronal inflammation, apoptosis, and STAT3 signal pathway. Western blot and immunofluorescence techniques were also performed to detect the regulatory effect and the underlying mechanism of wogonin on the inflammation and apoptosis of PC12 cells. Experimental results in vivo and in vitro showed that wogonin could promote the recovery of motor function, improve the histopathological morphology, inhibit the activation of the STAT3 signal pathway, and reduce the neuronal inflammation and apoptosis in the rats with SCI. Activation of the STAT3 signal pathway by colivelin reversed the therapeutic effect of wogonin. Therefore, wogonin could inhibit inflammation and apoptosis by inhibiting the STAT3 signal pathway and promote the functional recovery of rats with SCI., (Copyright © 2022 Elsevier B.V. All rights reserved.)

The blood-spinal cord barrier (BSCB) is a physiological barrier between the blood and spinal cord parenchyma. This study aims to determine whether Y-27632, a Rho-associated protein kinase (ROCK) inhibitor, can protect the BSCB using in vivo models. The Evans blue fluorescence assay was used to detect leakage of the BSCB. Western blotting was used to define alterations in ROCK-related and tight junction (TJ) protein expression. Immunofluorescence triple-staining was used to evaluate histologic alterations in TJs. Locomotor function was evaluated using the open-field test, the Basso-Beattie-Bresnahan score, and footprint analysis. Two peaks of BSCB leakage after spinal cord injury (SCI) occurred at 24 h and 5 days. The ROCK inhibitor reduced the BSCB leakage at the second peak after SCI. Moreover, the ROCK inhibitor ameliorated the integrity of the BSCB and improved motor function recovery after SCI by regulating the phosphorylation of myosin phosphatase subunit-1 (MYPT1) and cofilin. ROCK inhibitors might protect the BSCB, which provides a new strategy for transitioning SCI treatment from the bench to bedside., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Objectives: The aim of this study was to determine the role of microencapsulated olfactory ensheathing cells (MC-OECs) transplantation in rats with sciatic nerve injury-induced neuropathic pain, and its relationship with P2X4 receptor expression in the L4-5 spinal cord segment., Methods: Olfactory bulb tissues of healthy Sprague Dawley (SD) rats were collected to culture olfactory ensheathing cells (OECs) using differential attachment methods. Seventy-two healthy SD rats were randomly assigned to four groups: the sham, chronic constriction injury (CCI), OECs, and MC-OECs groups. Mechanical paw withdrawal thresholds were measured on days 7, 14 and 21 after surgery. The expression of P2X4 receptor genes in the L4-5 spinal cord segment was detected by in situ hybridization, western blotting and RT-PCR., Results: On post-surgical days 7, 14 and 21. The mechanical paw withdrawal thresholds of rats from low to high were the CCI, OECs, MC-OECs and sham groups. The expression level of P2X4 receptor mRNA and protein from low to high were the sham, MC-OECs, OECs and CCI groups. Compared with the OECs group, the mechanical paw withdrawal thresholds of rats were increased, and the expression level of P2X4 receptor mRNA and protein were decreased in the MC-OECs group. In addition, the mechanical paw withdrawal thresholds of rats were higher and the expression level of P2X4 receptor were lower in the MC-OECs group on post-surgical days 21 than that on days 7 and 14. All differences between groups were statistically significant (P value < 0.05)., Conclusions: OECs and MC-OECs transplantation can reduce the expression level of P2X4 receptor and relieve pain. The therapeutic efficacy was better in the MC-OECs group than in the OECs group. MC-OECs transplantation have the better effects of treatment with the increase of MC-OECs transplantation time., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Transplantation with mesenchymal stem cells (MSCs) has shown beneficial effects in treating spinal cord injury. Autophagy is an evolutionarily conserved process of degradation and recycling of cellular components that plays an important role in tissue homeostasis and cellular survival. Whether regulating autophagy in MSCs may affect their therapeutic potential in spinal cord injury repair has not yet been determined. In this study, autophagy was inhibited in MSCs with lentiviruses expressing short hairpin RNA (shRNA) to knock down Becn-1 expression, and autophagy was upregulated in MSCs under nutrient starvation. These MSCs were then labelled with Hoechst and applied to spinal cord-injured rats to evaluate their therapeutic effects. After transplanting MSCs into rats with spinal cord injuries, functional recovery, immunohistochemistry, and remyelination analyses were performed. After inducing autophagy, the MSCs exhibited an accumulation of LC3-positive autophagosomes in the cytoplasm. The expression levels of neurotrophic factors, including vascular endothelial growth factor and brain derived neurotrophic factor, were significantly higher in autophagic MSCs than normal MSCs. The in vivo study showed that more labelled MSCs migrated to the lesion site after induction of autophagy. Inducing autophagy in MSCs promoted functional recovery after spinal cord injury, whereas functional recovery was weak after inhibiting autophagy in MSCs. In contrast to the autophagy inhibition group, transplanting autophagic MSCs exhibited a greater positive impact on axon regeneration, growth of serotonergic fibers, blood vessel regeneration, and myelination, indicating a multifactorial contribution to spinal cord injury repair. These results suggest that autophagy plays important roles in MSCs during spinal cord injury repair. Regulation of autophagy in MSCs before in vivo transplantation may be a potential therapeutic interventional strategy for spinal cord injury., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Following the transection of peripherally located sympathetic preganglionic axons of the cervical sympathetic trunk (CST), transient retrograde neuronal and glial responses occur in the intermediolateral cell column (IML) of the spinal cord, the location of the parent neuronal cell bodies. The role of microglia in this central response to peripheral axon injury was examined in mice fed the PLX5622 diet containing colony-stimulating factor-1 receptor (CSF-1R) inhibitor for 28 days, which eliminated approximately 90% of spinal cord microglia. Microglia elimination did not impact baseline neurotransmitter expression in the IML neurons, and the typical neuronal plasticity observed following CST transection was unaffected. Oligodendrocyte precursor cells (OPCs) were significantly increased at one week post injury in the IML of mice fed the control diet, with no change in mature oligodendrocytes (OLs). Following microglia elimination, the baseline population of OPCs in the IML was increased, suggesting increased OPC proliferation. Injury in the microglia depleted mice resulted in no additional increase in OPCs. Though baseline astrocyte activation and GFAP protein expression were unaffected, microglia elimination led to increased activation and GFAP protein post injury when compared with mice fed the control diet. These results reveal that microglia regulate the baseline OPC population in the uninjured spinal cord and that activated microglia influence the activities of OL lineage cells as well as astrocytes. The regulatory roles of microglia observed in this study likely contribute to the long term survival of the IML neurons observed following the distal axon injury., (Copyright © 2019 Elsevier B.V. All rights reserved.)

The molecular mechanisms underlying cervical spondylotic myelopathy (CSM) are poorly understood. To assess the correlation between HIF-1α, MMP-9 and functional recovery following chronic cervical spinal cord compression (CSCI). Rats in the sham group underwent C5 semi-laminectomy, while a water-absorbable polyurethane polymer was implanted into the C6 epidural space in the chronic CSCI group. Basso, Beattie and Bresnahan score and somatosensory evoked potentials were used to evaluate neurological function. Hematoxylin and eosin staining was performed to assess pathological changes in the spinal cord, while immunohistochemical analysis was used to examine HIF-1α and MMP-9 expression on days 7, 28, 42 and 70 post-surgery. Normal rats were only used for HE staining. The BBB score was significantly reduced on day 28 following CSCI, while SEPs exhibited decreased amplitude and increased latency. In chronic CSCI group, the BBB score and SEPs significantly improved on day 70 compared with day 28. HE staining revealed different level of spinal cord edema after chronic CSCI. Compared with the sham group, immunohistochemical analyses revealed that HIF-1α- and MMP-9-positive cells were increased on day 7 and peaked on day 28. HIF-1α and MMP-9 expression were demonstrated to be significantly positively correlated, whereas HIF-1α expression and BBB score were significantly negatively correlated, as well MMP-9 expression and BBB score. HIF-1α and MMP-9 expression are increased following chronic spinal cord compression and are positively correlated with one another. Decreased expression of HIF-1α and MMP-9 may contribute to functional recovery following CSCI. This expression pattern of HIF-1α and MMP-9 may give a new perspective on the molecular mechanisms of CSM., (Copyright © 2019 Elsevier B.V. All rights reserved.)

The spinal locomotor central pattern generator (CPG) in neonatal mice exhibits diverse output patterns, ranging from sub-rhythmic to multi-rhythmic to fictive locomotion, depending on its general level of excitation and neuromodulatory status. We have recently reported that the locomotor CPG in neonatal mice rapidly recovers the ability to produce neurochemically induced fictive locomotion following an upper lumbar spinal cord compression injury. Here we address the question of recovery of multi-rhythmic activity and the serotonin-sensitivity of the CPG. In isolated spinal cords from control and 3 days post-injury mice, application of dopamine and NMDA elicited multi-rhythmic activity with slow and fast components. The slow component comprised 10-20 s episodes of activity that were synchronous in ipsilateral or all lumbar ventral roots, and the fast components involved bursts within these episodes that displayed coordinated patterns of alternation between ipsilateral roots. Rhythm strength was the same in control and injured spinal cords. However, power spectral analysis of signal within episodes showed a reduced peak frequency after recovery. In control spinal cords, serotonin triggered fictive locomotion only when applied at high concentration (30 µM, constant NMDA). By contrast, in about 50% of injured preparations fictive locomotion was evoked by 2-3 times lower serotonin concentrations (10-15 µM). This increased serotonin sensitivity was correlated with post-injury changes in the expression of specific serotonin receptor transcripts, but not of dopamine receptor transcripts., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Cell death is a key issue in spinal cord secondary injury. Ferroptosis is recently discovered as an iron-dependent type of cell death that is distinct from other forms of cell death pathways such as apoptosis and necrosis. This research is aimed to investigate the role of ferroptosis in spinal cord injury (SCI) pathophysiology, and to explore the effectiveness of ferroptosis inhibitor on SCI. We examined the ferroptosis markers and the factors in a rat contusion SCI model. Seen from transmission electron microscopy (TEM) following SCI, mitochondria showed ferroptotic characteristic changes. Treatment with a ferroptosis inhibitor SRS 16-86 enhanced functional recovery after SCI through the upregulation of anti-ferroptosis factor GPX4, GSH and xCT, and the downregulation of the lipid peroxidation marker 4HNE. SRS 16-86 treatment alleviated astrogliosis and enhanced neuronal survival after SCI. The inflammatory cytokine levels (IL-1β, TNF-α and ICAM-1) were decreased significantly post SRS 16-86 treatment after SCI. These findings suggest strong correlation between ferroptosis and the secondary injury of SCI. The effectiveness of ferroptosis inhibitor SRS-16-86 on SCI repair leads to the identification of a novel therapeutic target for SCI., (Copyright © 2018. Published by Elsevier B.V.)

Chondroitin sulfate proteoglycans produced in glial scar tissue are a major inhibitory factor for axonal regeneration after central nervous system injury in mammals. The inhibition is largely due to chondroitin sulfates, whose effects differ according to the sulfation pattern. In contrast to mammals, fish nerves spontaneously regenerate beyond the scar tissue after spinal cord injury, although the mechanisms that allow for axons to pass through the scar are unclear. Here, we used immunohistochemistry to examine the expression of two chondroitin sulfates with different sulfation variants at the lesion site in goldfish spinal cord. The intact spinal cord was immunoreactive for both chondroitin sulfate-A (CS-A) and chondroitin sulfate-C (CS-C), and CS-A immunoreactivity overlapped extensively with glial processes positive for glial fibrillary acidic protein. At 1week after inducing the spinal lesion, CS-A immunoreactivity was observed in the cell bodies and extracellular matrix, as well as in glial processes surrounding the lesion center. At 2weeks after the spinal lesion, regenerating axons entering the lesion center overtook the CS-A abundant area. In contrast, at 1week after lesion induction, CS-C immunoreactivity was significantly decreased, and at 2weeks after lesion induction, CS-C immunoreactivity was observed along the regenerating axons entering the lesion center. The present findings suggest that after spinal cord injury in goldfish, chondroitin sulfate proteoglycans are deposited in the extracellular matrix at the lesion site but do not form an impenetrable barrier to the growth of regenerating axons., (Copyright © 2017 Elsevier B.V. All rights reserved.)

The essential role of GAPDH/Siah1 signaling pathway in the pathogenesis of various injurious conditions such as traumatic spinal cord injury (SCI) has been gradually recognized. However, the drugs targeting this signaling pathway are still lacking. The endocannabinoid system, including its receptors (CB1 and CB2), act as neuroprotective and immunomodulatory modulators in SCI. WIN55212-2, an agonist for CB1 and CB2 receptors, has been demonstrated with anti-inflammatory and anti-apoptotic effects in multiple neurological diseases. Therefore, the present study aimed to investigate whether WIN55212-2 could promote functional recovery after traumatic SCI via inhibition of the GAPDH/Siah1 signaling. The traumatic SCI was induced by dropping a 10-g impactor from 25mm on the dorsal surface of T9 and T10. Our results showed that WIN55212-2 alleviated the activation of GAPDH/Siah1 signaling pathway after SCI, as indicated by the reduction in GAPDH nuclear expression, GAPDH-Siah1 complex formation and iNOS protein expression. Furthermore, WIN55212-2 reduced apoptosis, production of IL-1β and TNF-α and activation of NF-κB signaling in the spinal cord after SCI. The behavioral tests showed that WIN55212-2 improved the functional recovery after traumatic SCI as indicated by sustained increase in the locomotor scores. However, these neuroprotective effects of WIN55212-2 were blocked in the presence of the combined treatment of AM630 (an antagonist of CB2) rather than AM251 (an antagonist of CB1). In conclusion, our study indicates that, WIN55212-2 improves the functional recovery after SCI via inhibition of GAPDH/Siah1 cascades in a CB2 receptor dependent manner, indicative of its therapeutic potential for traumatic SCI or other traumatic conditions., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Aquaporin-4 (AQP4) is a water channel protein in spinal cord and plays a critical role in the pathophysiological process of spinal cord injury (SCI). However, little is known about the molecular mechanism of AQP4 involved in SCI. The present study was performed to investigate the possible molecules regulated by AQP4 after SCI by use of lentivirus-mediated RNA interference (RNAi). First, the motor function was evaluated by Basso, Beattie, Bresnahan (BBB) scale and the expression of AQP4 was measured by western blot, immunohistochemical staining and immunofluorescence in rats after contusion spinal cord injury (cSCI). After cSCI, the rats exhibited a gradual motor function recovery from 3dpo to 28dpo. And the expression and localization of AQP4 changed with different post-injury stages. At 3d after SCI, AQP4 located mainly in vascular endothelial cells (VECs) in the anterior horn of spinal cord, which was similar to the sham rats. At 7d and 28d after SCI, AQP4 was expressed both in VECs and the position of neuron membranes. The protein level of AQP4 increased significantly at 12h, 1d and 3d after SCI, and decreased slightly at 7d after SCI. Then lentivirus-mediated AQP4 RNA interference (AQP4-RNAi) was constructed and used to inhibit AQP4 expression in cSCI rats. The results from real-time quantitative polymerase chain reactions (RT-qPCR) and immunohistochemical staining suggested that the expression of nerve growth factor (NGF) was up-regulated by lentivirus-mediated AQP4 inhibition in cSCI rats, while the motor function recovery was accelerated. The results suggested that the acceleration of motor function recovery by AQP4 inhibition was associated with NGF up-regulation. This is the first report on the relationship between AQP4 and NGF in SCI, which may shed light on illustrating the role of AQP4 in SCI. These findings may also provide strategies of the clinical treatment for SCI in the future., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Salvianolic acid A (Sal A), a bioactive compound isolated from the Chinese medicinal herb Danshen, is used for the prevention and treatment of cardiovascular diseases. However, the protective function of Sal A on preserving the role of blood-spinal cord barrier (BSCB) after spinal cord injury (SCI) is unclear. The present study investigated the effects and mechanisms of Sal A (2.5, 5, 10mg/kg, i.p.) on BSCB permeability at different time-points after compressive SCI in rats. Compared to the SCI group, treatment with Sal A decreased the content of the Evans blue in the spinal cord tissue at 24h post-SCI. The expression levels of tight junction proteins and HO-1 were remarkably increased, and that of p-caveolin-1 protein was greatly decreased after SCI Sal A. The effect of Sal A on the expression level of ZO-1, occluding, and p-caveolin-1 after SCI was blocked by the HO-1 inhibitor, zinc protoporphyrin IX (ZnPP). Also, Sal A inhibited the level of apoptosis-related proteins and improved the motor function until 21days after SCI. In addition, Sal A significantly increased the expression of microRNA-101 (miR-101) in the RBMECs under hypoxia. AntagomiR-101 markedly increased the RBMECs permeability and the expression of the Cul3 protein by targeting with 3'-UTR of its mRNA. The expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and HO-1 was significantly increased after agomiR-101 treatment. Therefore, Sal A could improve the recovery of neurological function after SCI, which could be correlated with the repair of BSCB integrity by the miR-101/Cul3/Nrf2/HO-1 signaling pathway., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Spinal cord injury (SCI) leads to glial scar formation by astrocytes, which severely hinders neural regeneration. Curcumin (cur) can inhibit glial scar formation, but the underlying mechanism is not fully understood. Using both in vivo and in vitro experiments, the current study investigated the phenotypic transformation of astrocytes following cur and siRNA intervention during the processes of inflammation and fibrosis and determined details of the relationship between cur treatment and the glial scar components GFAP and CSPG. We found that cur and NF-κb p65 siRNA could inhibit astrocyte activation through suppressing NF-κb signaling pathway, which led to down-regulate the expression of chemokines MCP-1, RANTES and CXCL10 released by astrocytes and decreased macrophage and T-cell infiltration, thus reducing the inflammation in the glial scar. In addition, silencing SOX-9 may reduce the deposition of extracellular matrix CSPG; whereas its over-expression could increase the CSPG expression. Cur suppressedSOX-9-inducedCSPG deposition, reduced α-SMA (an important symbol of fibrosis) expression in astrocytes, altered astrocyte phenotype, and inhibited glial scar formation by regulating fibrosis. This study confirmed that cur could regulate both the NF-κb and SOX9 signaling pathways and reduce the expression of intracellular and extracellular glial scar components through dual-target regulating both inflammation and fibrosis after SCI in the rat. This study provides an important hypothesis centered on the dual inhibition of intracellular and extracellular glial scar components as a treatment strategy for SCI., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Previous studies have shown that inhibition of prolyl hydroxylase(PHD) stabilizes Hypoxia-inducible factor 1, alpha subunit(HIF-1α), increases tolerance to hypoxia, and improves the prognosis of many diseases. However, the role of PHD inhibitor (PHDI) in the recovery of spinal cord injury remains controversial. In this study, we investigated the protective role of a novel PHDI FG-4592 both in vivo and in vitro. FG-4592 treatment stabilized HIF1α expression both in PC12 cells and in spinal cord. FG-4592 treatment significantly inhibited tert-Butyl hydroperoxide(TBHP)-induced apoptosis and increases the survival of neuronal PC-12 cells. FG-4592 administration also improved recovery and increased the survival of neurons in spinal cord lesions in the mice model. Combination therapy including the specific HIF-1α blocker YC-1 down-regulated the HIF-1α expression and partially abolished the protective effect of FG-4592. Taken together, our results revealed that the role of FG-4592 in SCI recovery is related to the stabilization of HIF-1α and inhibition of apoptosis. Overall, our study suggests that PHDIs may be feasible candidates for therapeutic intervention after SCI and central nervous system disorders in humans., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Extensive changes occur at transcriptional level after traumatic spinal cord injury (SCI). In this study, we performed a large scale screening of expression changes of long (>200 nt) RNA transcripts including both coding and non-coding RNA species in a rat contusion SCI model. We validated significant down-regulation of one long non-coding RNA (lncSCIR1) at 1, 4, and 7 days postinjury. lncSCIR1 knockdown promoted astrocyte proliferation and migration in vitro. We further validated the strong association between lncSCIR1 knock down and the expression changes of four mRNAs after injury. Our data indicated that lncSCIR1 down-regulation might play a detrimental role in the pathophysiology of traumatic SCI and thereby provided new insights into the studies of potential therapeutic targets for traumatic central nervous system (CNS) injuries., (Copyright © 2015 Elsevier B.V. All rights reserved.)

A variety of neurotrophic factors have been used in attempts to improve morphological and behavioural outcomes after experimental spinal cord injury (SCI). Here we review many of these factors, their cellular targets, and their therapeutic impact on spinal cord repair in different, primarily rodent, models of SCI. A majority of studies report favourable outcomes but results are by no means consistent, thus a major aim of this review is to consider how best to apply neurotrophic factors after SCI to optimize their therapeutic potential. In addition to which factors are chosen, many variables need be considered when delivering trophic support, including where and when to apply a given factor or factors, how such factors are administered, at what dose, and for how long. Overall, the majority of studies have applied neurotrophic support in or close to the spinal cord lesion site, in the acute or sub-acute phase (0-14 days post-injury). Far fewer chronic SCI studies have been undertaken. In addition, comparatively fewer studies have administered neurotrophic factors directly to the cell bodies of injured neurons; yet in other instructive rodent models of CNS injury, for example optic nerve crush or transection, therapies are targeted directly at the injured neurons themselves, the retinal ganglion cells. The mode of delivery of neurotrophic factors is also an important variable, whether delivered by acute injection of recombinant proteins, sub-acute or chronic delivery using osmotic minipumps, cell-mediated delivery, delivery using polymer release vehicles or supporting bridges of some sort, or the use of gene therapy to modify neurons, glial cells or precursor/stem cells. Neurotrophic factors are often used in combination with cell or tissue grafts and/or other pharmacotherapeutic agents. Finally, the dose and time-course of delivery of trophic support should ideally be tailored to suit specific biological requirements, whether they relate to neuronal survival, axonal sparing/sprouting, or the long-distance regeneration of axons ending in a different mode of growth associated with terminal arborization and renewed synaptogenesis. This article is part of a Special Issue entitled SI: Spinal cord injury., (Copyright © 2014 Elsevier B.V. All rights reserved.)

The immunoreactive responses are a two-edged sword after spinal cord injury (SCI). Macrophages are the predominant inflammatory cells responsible for this response. However, the mechanism underlying the effects of HBOT on the immunomodulation following SCI is unclear now. The present study was performed to examine the effects of hyperbaric oxygen therapy (HBOT) on macrophage polarization after the rat compressive injury of the spinal cord. HBOT was associated with significant increases in IL-4 and IL-13 levels, and reductions in TNF-α and IFN-ɣ levels. This was associated simultaneously with the levels of alternatively activated macrophages (M2 phenotype: arginase-1- or CD206-positive), and decreased levels of classically activated macrophages (M1 phenotype: iNOS- or CD16/32-positive). These changes were associated with functional recovery in the HBOT-transplanted group, which correlated with preserved axons and increased myelin sparing. Our results suggested that HBOT after SCI modified the inflammatory environment by shifting the macrophage phenotype from M1 to M2, which may further promote the axonal extension and functional recovery., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Neural stem cell (NSC) transplantation has been reported to be a leading strategy to stimulate neuroplasticity, repair neuronal loss and promote the morphologic and functional recovery of spinal cord injury (SCI). However, massive death of transplanted NSCs is still a problem, which is considered to be related to a series of pro-inflammatory cytokines that induce apoptosis, extensive demyelination and axonal destruction. Tumor necrosis factor alpha (TNF-α), as one of the major inflammation initiators, contributes to secondary neural cell death. We previously found that the administration of the TNF-α antagonist etanercept during the acute phase of SCI can reduce the apoptosis of neurons and oligodendrocytes. To investigate whether etanercept can suppress transplanted NSC apoptosis and promote NSC survival, axon myelination and functional recovery, we tested the combination strategy of the early administration of etanercept and NSC transplantation. First we observed that etanercept suppressed the TNF-α expression and apoptosis of transplanted NSCs by Western blot, TUNEL and immunofluorescence staining. The Basso, Beattle and Bresnahan scale and motor-evoked potential were used to evaluate functional recovery. The results suggest significantly better recovery after combination therapy. Further, histopathological alterations were evaluated by hematoxylin and eosin staining and Nissl staining. These procedures showed that the early administration of etanercept improved survival of neurons in the ventral horn, restored neural morphology and produced a smaller cavity area. We observed most abundant NF-positive fibers after the combination treatment, indicating that combination therapy retained and promoted neural regeneration. Finally, the early suppression of TNF-α reduced the occurrence of demyelination, and the combination therapy led to more myelinated axons, as shown by electron microscopy. These data suggest that this strategy significantly protected transplanted NSCs via the anti-inflammation and anti-apoptosis effects of etanercept, promoting re-myelination, neural regeneration and locomotor function., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Spinal-cord injury is characterized by primary damage as a direct consequence of mechanical insult, and secondary damage that is partly due to the acute inflammatory response. The extent of any hemorrhage within the injured cord is also known to be associated with the formation of intraparenchymal cavities and has been anecdotally linked to secondary damage. This study was designed to examine the contribution of blood components to the outcome of spinal-cord injury. We stereotaxically microinjected collagenase, which causes localized bleeding, into the spinal cord to model the hemorrhage associated with spinal cord injury in the absence of significant mechanical trauma. Tissue damage was observed at the collagenase injection site over time, and was associated with localized disruption of the blood-spinal-cord barrier, neuronal cell death, and the recruitment of leukocytes. The magnitude of the bleed was related to neutrophil mobilization. Interestingly, the collagenase-induced injury also provoked extended axonal damage. With this model, the down-stream effects of hemorrhage are easily discernible, and the impact of treatment strategies for spinal-cord injury on hemorrhage-related injury can be evaluated., (Copyright © 2014. Published by Elsevier B.V.)

The glycinamide ribonucleotide transformylase (GART) gene, a trifunctional polypeptide, has phosphoribosylglycinamide formyltransferase, phosphoribosylglycinamide synthetase, and phosphoribosylaminoimidazole synthetase activity, and is required for de novo purine biosynthesis. GART is highly conserved in vertebrates. Alternative splicing of GART results in two transcript variants encoding different isoforms. However, the expression and function of GART in the central nervous system lesion are still unclear. In this study, we used a traumatic spinal cord injury (SCI) model in adult Sprague-Dawley rats and investigated the dynamic changes of GART protein expression in the spinal cord. Western blot analysis revealed that GART was present in sham-operated spinal cord. It gradually increased, reached a peak at day 3 after SCI, and then declined during the following days. Double immunofluorescence staining revealed a widespread of GART, and the majority of GARTs are detected in astrocytes. After injury, GART expression was increased predominantly in astrocytes, positively correlated with the highly expressed proliferating cell nuclear antigen (PCNA). Knockdown of GART expression in cultured primary astrocytes by siRNA revealed that expression of GART in astrocytes plays a role in the LPS-induced release of pro-inflammatory factors, such as TNF-α and IL-6. These results showed that GART may participate in the pathophysiology of SCI, and more research is needed to have a good understanding of its function and mechanism., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Amyloid-β (Aβ) is produced through the enzymatic cleavage of amyloid precursor protein (APP) by β (Bace1) and γ-secretases. The accumulation and aggregation of Aβ as amyloid plaques is the hallmark pathology of Alzheimer׳s disease and has been found in other neurological disorders, such as traumatic brain injury and multiple sclerosis. Although the role of Aβ after injury is not well understood, several studies have reported a negative correlation between Aβ formation and functional outcome. In this study we show that levels of APP, the enzymes cleaving APP (Bace1 and γ-secretase), and Aβ are significantly increased from 1 to 3 days after impact spinal cord injury (SCI) in mice. To determine the role of Aβ after SCI, we reduced or inhibited Aβ in vivo through pharmacological (using DAPT) or genetic (Bace1 knockout mice) approaches. We found that these interventions significantly impaired functional recovery as evaluated by white matter sparing and behavioral testing. These data are consistent with a beneficial role for Aβ after SCI., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Cyclic AMP (cAMP) signalling pathways are involved in axonal growth and regeneration. The calcium-calmodulin- stimulated adenylate cyclase 1 (AC1), a regulator of cAMP levels, is strongly expressed in the corticospinal motor neurons (CSMN) in cerebral cortex layer V during development, but its role in the development of the corticospinal tract (CST) is unknown. Here, we analyse the organization of the CST pathway using anterograde and retrograde tracers in the barrelless (brl) mouse that carries an inactivating mutation of the AC1 gene. We show that in brl mice the general organization of the CST is normal but there is an increase in the number of axons in the ipsilateral contingent in the dorsal and ventral medial funiculi of the cervical spinal cord. The density of CSMN in layer V of the motor cortex is increased in brl compared to wild-type mice. Thus, lack of AC1 likely perturbs late phases of CSMN and CST development. Next, we examine the motor recovery after a spinal cord injury (SCI). We find that brl mice show enhanced locomotor functions as assessed by the BMS (Basso mouse scale) as early as 6h and up to 6 weeks after SCI, indicating a smaller responsiveness of brl mice to SCI. It is therefore possible that developmental effects on motor systems might decrease the locomotor effects consecutive to a SCI. This point is particularly important with regards to the use of transgenic animals for testing SCI recovery., (Copyright © 2014 Elsevier B.V. All rights reserved.)

In previous studies with animal models of spinal cord injury (SCI) pharmacological activation of peroxisome proliferator activated receptors (PPAR) and liver X receptors (LXR) were used to reduce tissue damage and promote behavioral recovery in animal models. We have studied the endogenous expression of the transcription factors PPARα and LXRβ in the chronic stage after SCI in rats. The immunohistochemical investigation revealed a long lasting increase in the level of PPARα in white matter in the vicinity of the lesion site. The source of this signal was identified in a subpopulation of astrocytes outside of the glial scar area. Intrathecal injections of oleic acid/albumin reduced the lesion-induced PPARα immunoreactivity. In addition, ependymal cells displayed a prominent PPARα signal in the non-injured spinal cord, and continued to express the receptor as they proliferated and migrated within the damaged tissue. The nuclear receptor LXRβ was detected at similar levels after SCI as in sham operated animals. We found high levels of immunoreactivity in the gray matter, while in the white matter it was present in subpopulations of astrocytes and oligodendrocytes. Macrophages that had accumulated within the center of the lesion contained LXRβ in their cell nuclei. Possible endogenous functions of PPARα and LXRβ after SCI are discussed, specifically the control of fatty acid and cholesterol metabolism and the regulation of inflammatory reactions., (© 2013 Elsevier B.V. All rights reserved.)

The sex hormone progesterone has been shown to improve outcomes in animal models of a number of neurologic diseases, including traumatic brain injury, ischemia, spinal cord injury, peripheral nerve injury, demyelinating disease, neuromuscular disorders, and seizures. Evidence suggests it exerts its neuroprotective effects through several pathways, including reducing edema, improving neuronal survival, and modulating inflammation and apoptosis. In this review, we summarize the functional outcomes and pathophysiologic mechanisms attributed to progesterone treatment in neurologic disease. We then comment on the breadth of evidence for the use of progesterone in each neurologic disease family. Finally, we provide support for further human studies using progesterone to treat several neurologic diseases., (© 2013 Elsevier B.V. All rights reserved.)

Recent studies indicated that epigallocatechin gallate (EGCG) had neuroprotective effects on spinal cord injury (SCI).The current study was performed to determine the anti-edema effect of EGCG after SCI in rats. EGCG (100 mg/kg, i.p.) was administered to rats immediately following SCI. It was found that EGCG (100 mg/kg) could significantly reduce spinal cord water content. In addition, EGCG (100mg/kg) significantly reduced the expression of aquaporin-4(AQP4) and glial fibrillary acidic protein (GFAP) level at 24, 48 and 72h after injury, but it did not have this effect at 12 h after injury. The changes of AQP4 and GFAP protein induced by EGCG (100 mg/kg) treatment were accompanied by a reduction of spinal cord edema. Our results indicated that EGCG (100 mg/kg) could reduce spinal cord edema after SCI, which could be correlated with the down-regulation the expression of AQP4 and GFAP protein level after SCI., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Our previous studies showed that ligands to type 2 imidazoline receptors (I₂R), including 2-(2-Benzofuranyl)-2-imidazoline (2-BFI) and Idazoxan, were effective in reducing spinal cord inflammation caused by experimental autoimmune encephalomyelitis (EAE). In the present study, we determined the effective therapeutic time window of 2-BFI and found that administration of 2-BFI in mice before the appearance of ascending flaccid paralysis (1-10 days post immunization), but not during the period when neurological deficits occurred (11-20 days post immunization), significantly ameliorated EAE-induced neurobehavioral deficits, reduced the infiltration of inflammatory cells into the spinal cord, and reduced the level of demyelination. More interestingly, giving 2-BFI during 1-10 days post immunization selectively suppressed IL-17 levels in the peripheral blood, which strongly suggests that IL-17 may be a good early marker to indicate EAE progression and that 2-BFI may target CD4⁺ T lymphocytes, especially Th17 cells to reduce IL-17 expression. Collectively, these studies led us to envisage that 2-BFI can be a useful drug to treat multiple sclerosis (MS) when used in combination with an early indicator of MS progression, such as IL-17., (Crown Copyright © 2012. Published by Elsevier B.V. All rights reserved.)

Inflammation has long been implicated in secondary tissue damage after spinal cord injury (SCI). Our previous studies of inflammatory gene expression in rats after SCI revealed two temporally correlated clusters: the first was expressed early after injury and the second was up-regulated later, with peak expression at 1-2 weeks and persistent up-regulation through 6 months. To further address the role of inflammation after SCI, we examined inflammatory genes in a second species, mice, through 28 days after SCI. Using anchor gene clustering analysis, we found similar expression patterns for both the acute and chronic gene clusters previously identified after rat SCI. The acute group returned to normal expression levels by 7 days post injury. The chronic group, which included C1qB, p22(phox) and galectin-3, showed peak expression at 7 days and remained up-regulated through 28 days. Immunohistochemistry and western blot analysis showed that the protein expression of these genes was consistent with the mRNA expression. Further exploration of the role of one of these genes, galectin-3, suggests that galectin-3 may contribute to secondary injury. In summary, our findings extend our prior gene profiling data by demonstrating the chronic expression of a cluster of microglial associated inflammatory genes after SCI in mice. Moreover, by demonstrating that inhibition of one such factor improves recovery, the findings suggest that such chronic up-regulation of inflammatory processes may contribute to secondary tissue damage after SCI, and that there may be a broader therapeutic window for neuroprotection than generally accepted., (Published by Elsevier B.V.)

We examined gene expression in the lumbar spinal cord and the specific response of motoneurons, intermediate gray and proprioceptive sensory neurons after spinal cord injury and exercise of hindlimbs to identify potential molecular processes involved in activity dependent plasticity. Adult female rats received a low thoracic transection and passive cycling exercise for 1 or 4weeks. Gene expression analysis focused on the neurotrophic factors: brain-derived neurotrophic factor (BDNF), glial cell line-derived neurotrophic factor (GDNF), neurotrophin-3 (NT-3), neurotrophin-4 (NT-4), and their receptors because of their potential roles in neural plasticity. We also examined expression of genes involved in the cellular response to injury: heat shock proteins (HSP) -27 and -70, glial fibrillary acidic protein (GFAP) and caspases -3, -7, and -9. In lumbar cord samples, injury increased the expression of mRNA for TrkB, all three caspases and the HSPs. Acute and prolonged exercise increased expression of mRNA for the neurotrophic factors BDNF and GDNF, but not their receptors. It also increased HSP expression and decreased caspase-7 expression, with changes in protein levels complimentary to these changes in mRNA expression. Motoneurons and intermediate gray displayed little change in mRNA expression following injury, but acute and prolonged exercise increased levels of mRNA for BDNF, GDNF and NT-4. In large DRG neurons, mRNA for neurotrophic factors and their receptors were largely unaffected by either injury or exercise. However, caspase mRNA expression was increased by injury and decreased by exercise. Our results demonstrate that exercise affects expression of genes involved in plasticity and apoptosis in a cell specific manner and that these change with increased post-injury intervals and/or prolonged periods of exercise., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Glutamate is a major excitatory transmitter in the central nervous system that may produce cellular injury when its concentration is abnormally increased in the synaptic cleft. Glial glutamate transporters GLAST and GLT-1, which are responsible for clearing synaptic glutamate into glial cells, play an important role in the regulation of the glutamate concentration in the synaptic cleft. However, there has been no report on long-term changes in the levels of glutamate transporters following spinal cord injury. Spinal cord injury (SCI) was induced at T12 by a New York University (NYU) impactor. Segments of the spinal cord at T9-10, L1-2, L4-5 and at the epicenter were removed after SCI, and Western blots for GLAST, GLT-1 and EAAC1 were performed. GLAST and GLT-1 were significantly decreased in the epicenter from 1day up to 8weeks after SCI. GLT-1 was significantly decreased in the spinal segments rostral to the injury site, and GLAST expression was significantly increased in the L4-5 region of the spinal cord for 8weeks. Because strategies to modulate the regulation of glutamate transporters may be applied, the present data serve as a reference for further research, although the long-term roles of glutamate transporters in pathological processes caused by SCI are not clear., (Copyright © 2011 Elsevier B.V. All rights reserved.)

The study investigated the mechanism of the up-regulation of aquaporin-4 (AQP4) and aquaporin-1 (AQP1) expression induced by spinal cord injury (SCI). Using adult rat spinal cord injury model, it was found that up-regulation of hypoxia inducible factor-1α (HIF-1α), vascular endothelial growth factor (VEGF), AQP4, and AQP1 in response to spinal cord injury was greatly antagonized by 2-methoxyestradiol (2ME2), which can post-transcriptionally inhibit the expression of HIF-1α. VEGF alone significantly increased the extravasation of Evans blue and up-regulated the levels of AQP4 protein expression in the injured spinal cord issue, but the levels of AQP1 expression were not significantly changed. Taken together, our results suggest that expression of AQP4 and AQP1 is correlated with up-regulation of HIF-1α after SCI through the mechanisms that were dependent and independent of the VEGF signaling pathway, respectively. And the inhibitor of HIF-1α is a novel promising therapeutic agent for human SCI-induced edema in the future., (Copyright © 2010. Published by Elsevier B.V.)

Spontaneous and/or treatment-evoked re-modeling of the CNS following spinal cord injury is a prerequisite for functional recovery. While there has been considerable interest in the role of endogenous neurotrophins in spontaneous plasticity of several populations of spinal axons, the same cannot be said for morphological changes to dendrites. Here, we examined the responses of dendrites in the mouse lateral spinal nucleus (LSN, a site of sensory integration in the dorsolateral white matter) to exogenous and endogenous neurotrophins. We performed a septuple dorsal rhizotomy, which permanently eliminates sensory input to the spinal cord, and stimulates sprouting of spinal axons. While dendrites showed no change in density following injury alone, they sprouted vigorously (a two-fold increase in density) upon addition of exogenous brain-derived neurotrophic factor (BDNF). On the other hand, endogenous nerve growth factor (NGF) severely restricted dendritic sprouting, as TrkA-Fc treatment also roughly doubled the density of dendritic processes in the LSN. Spontaneous, BDNF- and TrkA-Fc mediated sprouting was unaffected by the absence of p75(NTR). Importantly, TrkA-Fc treatment markedly reduced expression of the truncated BDNF receptor TrkBT1 in both p75(+/+) and p75(-/-) mice, which was robustly-upregulated by deafferentation in both genotypes. We propose that the upregulation of TrkBT1 by NGF results in a reduced availability of endogenous BDNF to dendrites. Accordingly, sprouting of serotonergic axons, a BDNF-dependent consequence of dorsal root injury, was significantly enhanced in TrkA-Fc-treated animals. These results suggest that NGF and BDNF signaling differentially regulates dendritic plasticity in the deafferented spinal cord., (Copyright 2010 Elsevier B.V. All rights reserved.)

It is well known that spinal motoneurons below a spinal transection become supersensitive to a systemic administration of serotonin (5-HT) precursors, such as 5-hydroxytryptophan. This supersensitivity has been implicated in both the process of functional recovery following chronic lesions, and also in the development of symptoms such as hyperreflexia and spasticity. However, the mechanisms of this denervation supersensitivity are still largely unknown. In this study we have investigated the changes in 5-HT2A receptor immunoreactivity following chronic spinal transections at the level of the sacrocaudal spinal cord. The results show that in the spinalized rats the immunoreactivity of 5-HT2A receptors below the lesion is dramatically increased in the motoneuron soma and its proximal dendritic territory, most likely also in their distal dendritic territory, to a level 3-5-fold higher than that of sham-operated rats. We also found a small number of intraspinal 5-HT neurons and clusters of 5-HT fibers and their varicosities in the spinal cord caudal to the lesion, which may provide an intrinsic source of 5-HT to act upon the upregulated 5-HT2A receptors. These results indicate that the upregulation of 5-HT2A receptors at least partly underlies the 5-HT denervation supersensitivity of spinal motoneurons after a complete spinal transection., (Copyright 2010 Elsevier B.V. All rights reserved.)

Recent studies revealed the neuroprotective effects of epigallocatechin gallate (EGCG) on a variety of neural injury .The purpose of this study was to determine the effects of EGCG on the tissue protection and behavioral improvement after spinal cord injury (SCI). Rats were randomly divided into four groups of 18 rats each as follows: sham-operated group, trauma group, and EGCG treatment groups (50 mg/kg, i.p., immediately and 1 hour after SCI). Spinal cord samples were taken 24 hours after injury and studied for determination of malodialdehyde (MDA) levels, immunohistochemistry of Bax and Bcl-2, and TUNEL reaction. Behavioral testing was performed weekly up to 6 weeks post-injury. Then, the rats were euthanized for histopathological assessment. The results showed that MDA levels were significantly decreased in EGCG treatment groups. Greater Bcl-2 and attenuated Bax expression could be detected in the EGCG-treated rats. EGCG significantly reduced TUNEL-positive rate. Also, EGCG significantly reduced the percentage of lesion area and improved behavioral function than the trauma group. On the basis of these findings, we propose that EGCG may be effective in protecting rat spinal cord from secondary injury.

GABA signaling plays an important role in the spinal cord response to injury and subsequent motor training. Since benzodiazepines are commonly used to treat muscle spasticity in spinal cord injured subjects and the gamma2 subunit of the GABA(A) receptor is necessary for benzodiazepine binding, this subunit may be an important factor modulating sensorimotor function after an injury. Changes in gamma2 levels in muscle-specific motoneurons and surrounding astrocytes were determined approximately 3 months after a complete mid-thoracic spinal cord transection at P5 in non-trained and in step-trained spinal rats. Soleus (ankle extensor) and tibialis anterior (TA, ankle flexor) motor pools were identified using retrograde labeling via intramuscular injections of Fast Blue or Fluoro Gold, respectively. Lumbar spinal cord sections showed gamma2 immunostaining in both soleus and TA motoneurons and astrocytes. gamma2 immunoreactivity on the soma of soleus and TA motoneurons in spinal rats was differentially modulated. Compared to intact rats, spinal rats had higher levels of gamma2 in TA, and lower levels in soleus motoneurons. Step training restored GABA(A) gamma2 levels towards control values in motoneuronal pools of both muscles. In contrast, the gamma2 levels were elevated in surrounding astrocytes of both motor pools in spinal rats, and step training had no further effect. Thus, motor training had a specific effect on those neurons that were directly involved with the motor task. Since the gamma2 subunit is involved with GABA(A) receptor trafficking and synaptic clustering, it appears that this subunit could be an important component of the activity-dependent response of the spinal cord after a spinal injury.

Spinal cord stimulation (SCS) is an alternative approach for treatment of neuropathic pain when conservative management is ineffective. Previously we showed both 4 Hz and 60 Hz SCS reduces hyperalgesia in an animal model of neuropathic pain. However, the mechanisms underlying the pain reduction by SCS and how different frequencies of SCS produce the analgesic effect are unclear. To elucidate potential sites modulated by SCS we examined distribution of c-fos in Sprague-Dawley rats with spared nerve injury (SNI) and those without injury in response to SCS. SCS was delivered at one of 3 different frequencies (4 Hz, 60 Hz, and 100 Hz) for 30 min 2 weeks after SNI or in animals without SNI. Animals were perfused either 5 min or 2 h after SCS and c-fos protein examined immunohistochemically. The number of c-fos positive cells significantly increased 5 min (35 min after SCS began) after 4 Hz SCS in the NRM, but not PAG in animals with nerve injury. The number of c-fos positive cells was significantly increased bilaterally 2 h after either 4 Hz or 60 Hz SCS in the spinal cord dorsal horn in the cervical enlargement and under the electrode, but not in the lumbar enlargement in animals with nerve injury. In uninjured animals 4 Hz SCS increased c-fos expression at the electrode site and lumbar enlargement when compared to animals implanted with the electrode who did not receive SCS. 100 Hz SCS had no effect on c-fos expression. Thus, at the time points examined in this model, low frequency SCS likely activates supraspinal and spinal mechanisms to produce analgesia, while higher frequencies activate spinal mechanisms.

The study investigated whether bradykinin (BK) preconditioning could regulate the expression of aquaporin-4 (AQP4) using an in vivo transient spinal cord ischemia model in rats. BK was infused continuously via the left femoral artery with infusion pump for 15 min (10 microg/kg/min) then we induced ischemia for 20 min and reperfusion for 24 and 72 h respectively. The results demonstrated that the central part of the white matter exhibited loss of perivascular AQP4 and showed a partial recovery toward 72 h of reperfusion. The border zone of white matter was different from the central part of the white matter by showing no loss of perivascular AQP4 at 24 h of reperfusion but rather a slight increase. BK significantly reduced the expression level of AQP4 protein in the white matter, but it had none of this effect in the gray matter region at 72 h post-reperfusion. There was no difference in AQP4 protein levels between BK group and control group at the two above-mentioned spinal cord regions at 24 h after reperfusion. In addition, the changes in AQP4 protein induced by BK preconditioning were obvious at 72 h after reperfusion, which were accompanied by a reduction of spinal cord edema. Our results demonstrated that the expression of AQP4 protein after spinal cord ischemia/reperfusion was region-specific, time-dependent and also indicated that the attenuation of AQP4 expression induced by BK could be one of the important molecular mechanisms in physiopathology of spinal cord ischemic edema.

Several proteins have been identified as inhibitors of axonal regeneration following injury of the adult vertebrate central nervous system. The repulsive guidance molecule (RGMa) is considered a potent myelin-derived neurite outgrowth inhibitor. In rats, RGMa inhibition enhances the growth of injured axons and promotes functional recovery after spinal cord injury (SCI). Here, we demonstrate that RGMa inhibition induces synaptic rearrangements of spared axonal projections after SCI. Intrathecal administration of a function-blocking antibody to RGMa enhances anatomical synapse formation of the corticospinal tract in the cervical region of rats with thoracic spinal cord hemisection. These findings suggest that the suppression of synaptic rearrangements as well as axon growth inhibition in the adult spinal cord may contribute to the limitation of functional recovery after SCI.

Since hepatocyte growth factor (HGF) plays a pivotal role in the development of the central nervous system and pathological conditions, we examined the long-term changes in the mRNA and protein expression of HGF and its receptor c-Met after spinal cord injury (SCI) in rats. HGF mRNA was significantly increased from 7 days after SCI in the injured segment, and the peak was at 7 days after SCI as assessed by real-time RT-PCR. Importantly, c-met mRNA expression was up-regulated from 1 day after SCI, and reached a peak at 14 days after SCI. Although up-regulation of HGF and c-met mRNA expression in the injured segment gradually decreased, the increased expression level persisted until 56 days after SCI. Consistent with HGF mRNA expression, HGF protein level was significantly increased mainly in the injured region, which persisted until 56 days after SCI. Immunohistochemistry showed that most of GFAP-positive reactive astrocytes expressed HGF and c-Met both on 14 days and 56 days after SCI. Staining with the mitotic indicator, bromodeoxyuridine (BrdU), revealed that a small number of BrdU-incorporated cells were co-localized with HGF/GFAP-positive or c-Met/GFAP-positive cells both on 14 and 56 days. These data suggest that HGF and c-Met were up-regulated mainly in the reactive astrocytes around the injured region in the subacute to chronic stage of spinal cord injury. Since HGF plays a critical role in neurotrophic activity, activation of the HGF/c-Met signaling system might be involved in the process of post-traumatic regeneration.

NgR is a common receptor for three myelin-associated inhibitors and mediates their inhibitory activities on neurite outgrowth. In the present study, we investigated whether a DNA vaccine targeting NgR could play a beneficial role in improving recovery from spinal cord injury (SCI). We demonstrated that a DNA vaccine against NgR was successfully constructed and expressed efficiently in vitro and in vivo. After immunization with anti-NgR DNA vaccine, a low level of antibody response and a T cell-mediated immune response were induced in the vaccinated rats. And the antisera taken from the anti-NgR DNA vaccinated rats could partly reverse the inhibition of MAG on neurite outgrowth. When the rats were subjected to a contusive SCI, the vaccinated rats showed much better functional recovery than the controls. In those vaccinated rats that induced a T cell response and generated antibodies against NgR, functional improvements were even better. Histological assessments by three-dimensional reconstruction further demonstrated that the total lesion volume in the vaccinated rats was reduced by 30.8% compared to the controls. These results collectively suggest that DNA vaccine against NgR can significantly improve functional recovery in rats that received contusive SCI and that the vaccination approach may provide a promising strategy for promoting SCI repair.

The aim of this study was to examine the expression profile of the vesicular acetylcholine transporter (VAChT), which is a cholinergic pre-synaptic marker, in the lower neural tract following spinal cord injury (SCI) and its effect on coordination of micturition. In adult female Sprague-Dawley rats, SCI was induced by complete transection of the spinal cord at T9. At various time points, 3, 7, 14 and 28 days, after SCI, cystometry was performed on conscious rats. Bladder areflexia was observed during the first week. Twenty-eight days after SCI the rats showed reflex contractions and voiding. The expression of VAChT was examined with immunohistochemistry. The number of VAChT-positive nerve terminals, which were surrounding neuronal soma, was transiently decreased in pelvic ganglion and spinal cord (L1, L2, L6 and S1). In particular VAChT terminals surrounding motor neurons in the ventral horn and autonomic pre-ganglion cells were dramatically decreased from 3 to 14 days after SCI. Similarly, and the number of VAChT-positive fibers in the bladder wall was also decreased. The intensity of VAChT terminals recovered in all above regions in conjunction with recovery of bladder function. These observations indicate that the transient decrease of the VAChT-positive nerve might cause a failure of cholinergic neuronal transmission along the urinary bladder tract after SCI. As the cholinergic system was recovered at least in rat, the functional recovery of neurogenic bladder syndrome in SCI patients may become possible by further understanding the mechanism underlying the recovery of cholinergic system in rat.

Loss and damage to blood vessels are thought to contribute to secondary tissue loss after spinal cord injury. Integrins might be therapeutic targets to protect the vasculature and/or promote angiogenesis, as their activation can promote tubule formation and survival of endothelial cells in vitro. Here, we show that immunostaining with an antibody against the alpha1beta1 integrin heterodimer is present only in blood vessels from postnatal day 1 (P1) through adulthood in Sprague-Dawley rats. After a spinal cord contusion at T9 in adults, the area of alpha1beta1 integrin positive blood vessels increases within 11 mm from the injury site at 3 days post-injury and remains prominent within the injured core only at 7 days. Staining for the alpha6beta1 integrin heterodimer increases in blood vessels between P10 and adulthood and is present in preganglionic neurons of the intermediolateral cell column (IML) at all ages. The alpha6beta1 integrin is also expressed by motor neurons postnatally, and oligodendrocyte precursors (OPCs), as previously reported. After the contusion, the area of alpha6beta1-stained blood vessels is increased at 3 days and most prominently, 1 mm from the injury site, followed by a significant reduction at 7 days, when alpha6beta1 integrin staining is most prominent around the injured core. Staining is also present in a subset of microglia and/or macrophages. These results raise the possibility that alpha1beta1 and alpha6beta1 integrins in blood vessels might be targeted to reduce blood vessel loss and promote angiogenesis, which may promote tissue sparing after spinal cord injury.

Xenopus laevis tadpoles functionally recover from spinal cord transection. Because this recovery requires the tadpole to metamorphose, it may result from compensatory changes initiated by de novo growth of axons involved in limb dominant locomotion rather than from regeneration of cut axons. To determine whether axonal regrowth contributes to functional recovery, sequential retrograde double labeling with two fluorescent dextran amines was used to identify neurons with regenerated axons. Rhodamine dextran amine was applied to hemisected spinal cords of prometamorphic tadpoles between the 4th and 5th vertebrae. After metamorphosis, in animals that had recovered movement, fluorescein dextran amine was applied to the lumbar spinal cord. Two weeks later, the CNS of these animals was examined for the presence of double-labeled neurons, i.e., those whose axons had regenerated. Double-labeled neurons were found in the reticular, raphe, and solitary tract nuclei, and in the interstitial nucleus of the medial longitudinal fasciculus. Because Xenopus expresses all the known mammalian molecular inhibitors of CNS axon regeneration, the determination that these phylogenetically conserved populations of neurons are indeed capable of axon regeneration should facilitate molecular studies of successful recovery from spinal cord trauma.