Complete Spinal Cord Transection Research Articles

Modified acellular nerve-delivering PMSCs improve functional recovery in rats after complete spinal cord transection.

Due to the poor regeneration capacity of neurons and the inhibitory microenvironment, spontaneous regeneration in spinal cord injury (SCI) remains challenging. Tissue engineering is considered a promising approach for enhancing the regeneration of SCI by reconstructing the inherent structure and improving the microenvironment. In this study, the possibility of engineering a nerve complex, which is constructed by acellular nerve delivering placenta mesenchymal stem cells (PMSCs), was assessed for the recovery of a transected spinal cord. Modified acellular nerve grafts were developed, and PMSCs labeled with green fluorescent protein (GFP) were seeded on the graft to construct the engineered nerve complex. Then, the engineered nerve complex was implanted into a 2 mm-length transected gap of the spinal cord. Four weeks after the transplantation, numerous surviving PMSCs were observed in the lesion cavity by immunofluorescence staining. Moreover, co-localization between GFP and neurofilament-200 (NF200) and Neuronal Class III β-Tubulin (Tuj1) was observed at the bridge interface. The PMSCs-graft group exhibited significant function improvement as evaluated by the Basso, Beattie and Bresnahan (BBB) locomotion score and footprint analysis. Eight weeks after surgery, the evoked response was restored in the PMSCs-graft group and numerous thick myelin sheathes were observed compared to that in the control groups. Collectively, our findings suggest that the nerve complex prepared by acellular nerve delivering PMSCs enhanced the structure and function regeneration of the spinal cord after SCI.

The role of RhoA in retrograde neuronal death and axon regeneration after spinal cord injury

Paralysis following spinal cord injury (SCI) is due to interruption of axons and their failure to regenerate. It has been suggested that the small GTPase RhoA may be an intracellular signaling convergence point for several types of growth-inhibiting extracellular molecules. Even if this is true in vitro, it is not clear from studies in mammalian SCI, whether the effects of RhoA manipulations on axon growth in vivo are due to a RhoA-mediated inhibition of true regeneration or only of collateral sprouting from spared axons, since work on SCI generally is performed with partial injury models. RhoA also has been implicated in local neuronal apoptosis after SCI, but whether this reflects an effect on axotomy-induced cell death or an effect on other pathological mechanisms is not known. In order to resolve these ambiguities, we studied the effects of RhoA knockdown in the sea lamprey central nervous system (CNS), where after complete spinal cord transection (TX), robust but incomplete regeneration of large axons belonging to individually identified reticulospinal (RS) neurons occurs, and where some RS neurons show unambiguous delayed retrograde apoptosis after axotomy. RhoA protein was detected in neurons and axons of the lamprey brain and spinal cord, and its expression was increased post-TX. Knockdown of RhoA in vivo by retrogradely-delivered morpholino antisense oligonucleotides (MOs) to the RS neurons significantly reduced retrograde apoptosis signaling in identified RS neurons post-SCI, as indicated by Fluorochrome Labeled Inhibitor of Caspases (FLICA) in brain wholemounts. In individual RS neurons, the reduction of caspase activation by RhoA knockdown began at 2weeks post-TX and was still seen at 8weeks. RhoA knockdown slowed axon retraction and possibly increased early axon regeneration in the proximal stump. The number of axons regenerating beyond the lesion more than 5mm at 10weeks post-TX also was increased. Thus RhoA knockdown both enhanced true axon regeneration and inhibited retrograde apoptosis signaling after SCI.

The spinal generator of ejaculation: Functional consequences of chronic spinalization and effect of substance P in anesthetized rats

The inability to ejaculate, i.e. anejaculation, affects the vast majority of men after spinal cord injury (SCI). Ejaculation can however be obtained in approximately half of SCI men by applying extraphysiological vibratory stimulation to the penis suggesting that a spinal neural organization commanding ejaculation exists than can be activated despite disruption of cerebral connections. In the rat, a spinal generator of ejaculation (SGE) has been identified which is notably characterized by the presence of substance P (SP) receptor (neurokinin-1 receptor) onto the constituting neurons. The aim of this study was to evaluate the consequence of chronic spinal cord section and the effect of SP on the function of the rat SGE. Electrical stimulations of varying intensity were applied to SGE in anesthetized rats 4weeks after complete transection of the thoracic spinal cord (T9) and ejaculation occurrence as well as peripheral responses, i.e. bulbospongiosus electromyogram and pressure within the seminal vesicle, were monitored. Then the effect of SP locally delivered was assessed in this experimental setting. Occurrence of ejaculation elicited by SGE stimulation, SGE excitatory threshold, and amplitude of peripheral responses were unchanged in spinalized as compared to spinally intact rats. In spinalized rats, SP triggered ejaculation upon intraspinal delivery into the SGE area and decreased the SGE stimulation intensity provoking ejaculation after intrathecal administration indicating a decrease in SGE excitatory threshold. The pro-ejaculatory inducing and facilitating effects of SP were reversed by the selective neurokinin-1 receptor antagonist RP67580. It was concluded that chronic spinalization has no significant impact on SGE functioning and SP exerts a pro-ejaculatory role at the SGE level, opening new avenues for the treatment of anejaculation in SCI men.

Comparison of spinal cord contusion and transection: functional and histological changes in the rat urinary bladder

To compare the effect of complete transection (tSCI) and contusion spinal cord injury (cSCI) on bladder function and bladder wall structure in rats. A total of 30 female Sprague-Dawley rats were randomly divided into three equal groups: an uninjured control, a cSCI and a tSCI group. The cSCI group underwent spinal cord contusion, while the tSCI group underwent complete spinal cord transection. At 6 weeks post-injury, 24-h metabolic cage measurement and conscious cystometry were performed. Conscious cystometry analysis showed that the cSCI and tSCI groups had significantly larger bladder capacities than the control group. The cSCI group had significantly more non-voiding detrusor contractions than the tSCI group. Both injury groups had more non-voiding contractions compared with the control group. The mean threshold pressure was significantly higher in the tSCI group than in the control and cSCI groups. The number of voids in the tSCI group was lower compared with the control group. Metabolic cage analysis showed that the tSCI group had larger maximum voiding volume as compared with the control and cSCI groups. Vesicular acetylcholine transporter/smooth muscle immunoreactivity was higher in the control than in the cSCI or tSCI rats. The area of calcitonin gene-related peptide staining was smaller in the tSCI group than in the control or cSCI groups. Spinal cord transection and contusion produce different bladder phenotypes in rat models of SCI. Functional data suggest that the tSCI group has an obstructive high-pressure voiding pattern, while the cSCI group has more uninhibited detrusor contractions.

Mice with sclerostin gene deletion are resistant to the severe sublesional bone loss induced by spinal cord injury.

The bone loss secondary to spinal cord injury (SCI) is associated with several unique pathological features, including the permanent immobilization, neurological dysfunction, and systemic hormonal alternations. It remains unclear how these complex pathophysiological changes are linked to molecular alterations that influence bone metabolism in SCI. Sclerostin is a key negative regulator of bone formation and bone mass. We hypothesized that sclerostin could function as a major mediator of bone loss following SCI. To test this hypothesis, 10-week-old female sclerostin knockout (SOST KO) and wild type (WT) mice underwent complete spinal cord transection or laminectomy (Sham). At 8weeks after SCI, substantial loss of bone mineral density was observed at the distal femur and proximal tibia in WT mice but not in SOST KO mice. By μCT, trabecular bone volume of the distal femur was markedly decreased by 64% in WT mice after SCI. In striking contrast, there was no significant reduction of bone volume in SOST KO/SCI mice compared with SOST KO/sham. Histomorphometric analysis of trabecular bone revealed that the significant reduction in bone formation rate following SCI was observed in WT mice but not in SOST KO mice. Moreover, SCI did not alter osteoblastogenesis of marrow stromal cells in SOST KO mice. Our findings demonstrate that SOST KO mice were protected from the major sublesional bone loss that invariably follows SCI. The evidence indicates that sclerostin is an important mediator of the marked sublesional bone loss after SCI, and that pharmacological inhibition of sclerostin may represent a promising novel approach to this challenging clinical problem.

Impacts of Ultra-early Hyperbaric Oxygen Therapy on Bone Mass of Rats With Complete Spinal Cord Transection.

Seventy-five SD rats were randomly assigned to four groups: sham-operated group (Sham, n = 15), complete spinal cord transection (CSCT, n = 20) group, hyperbaric oxygenation group 1 (HBO1, n = 20), and hyperbaric oxygenation group 2 (HBO2, n = 20). The aim of this study was to analyze the impacts of ultra-early hyperbaric oxygen therapy on bone mass of rats with CSCT. Treatment of patients with complete SCI is still an unresolved medical issue and needs to be further investigated. Studies on changes in bone mass as well as osteoporosis prevention after SCI have important clinical significance. Rats in the sham group only underwent T10 laminectomy, without damaging the spinal cord. Rats in CSCT, HBO1, and HBO2 groups underwent T10 laminectomy and spinal cord transection at T10 level. Rats in HBO1 and HBO2 groups received three courses of hyperbaric oxygen therapy with 10 days per course starting at 3 and 12 hours after spinal cord injury, respectively. The femoral biomechanical characteristics, the bone calcium, and the bone hydroxyproline (B-HYP) contents were determined. Morphology of the femur bone trabecula and the bone collagen were observed by HE staining and by masson triad color staining, respectively. After 6 weeks of treatment, rats in the CSCT group showed significant decreases in femur structural and material mechanics parameters, calcium and B-HYP contents, (P < 0.01), as well as sparse, fractured, malaligned trabecular bone and collagen compared with rats in the sham group. After treatments, compared with rats in the CSCT and HBO2 groups, rats in HBO1 group showed enhancement in femur structural and material mechanics parameters, calcium and B-HYP contents, (P < 0.05), as well as trabecular bone and collagen with better continuity and neater arrangement. Ultra-early HBO therapy can significantly improve bone mass in CSCT rats. N/A.

Abstracts from The 34(th) Annual National Neurotrauma Symposium June 26-29, 2016 Lexington, Kentucky.

Abstracts from The 34(th) Annual National Neurotrauma Symposium June 26-29, 2016 Lexington, Kentucky.

Olfactory Ensheathing Cell Transplantation after a Complete Spinal Cord Transection Mediates Neuroprotective and Immunomodulatory Mechanisms to Facilitate Regeneration.

Multiple neural and peripheral cell types rapidly respond to tissue damage after spinal cord injury to form a structurally and chemically inhibitory scar that limits axon regeneration. Astrocytes form an astroglial scar and produce chondroitin sulfate proteoglycans (CSPGs), activate microglia, and recruit blood-derived immune cells to the lesion for debris removal. One beneficial therapy, olfactory ensheathing cell (OEC) transplantation, results in functional improvements and promotes axon regeneration after spinal cord injury. The lack of an OEC-specific marker, however, has limited the investigation of mechanisms underlying their proregenerative effects. We compared the effects of enhanced green fluorescent protein-labeled fibroblast (FB) and OEC transplants acutely after a complete low-thoracic spinal cord transection in adult rats. We assessed the preservation of neurons and serotonergic axons, the levels of inhibitory CSPGs and myelin debris, and the extent of immune cell activation between 1 and 8 weeks postinjury. Our findings indicate that OECs survive longer than FBs post-transplantation, preserve axons and neurons, and reduce inhibitory molecules in the lesion core. Additionally, we show that OECs limit immune-cell activation and infiltration, whereas FBs alter astroglial scar formation and increase immune-cell infiltration and concomitant secondary tissue damage. Administration of cyclosporine-A to enhance graft survival demonstrated that immune suppression can augment OEC contact-mediated protection of axons and neurons during the first 2 weeks postinjury. Collectively, these data suggest that OECs have neuroprotective and immunomodulatory mechanisms that create a supportive environment for neuronal survival and axon regeneration after spinal cord injury. Spinal cord injury creates physical and chemical barriers to axon regeneration. We used a complete spinal cord transection model and olfactory ensheathing cell (OEC) or fibroblast (FB; control) transplantation as a repair strategy. OECs, but not FBs, intermingled with astrocytes, facilitated astroglial scar border formation and sequestered invading peripheral cells. OECs attenuated immune cell infiltration, reduced secondary tissue damage, protected neurons and axons in the lesion core, and helped clear myelin debris. Immunosuppression enhanced survival of OECs and FBs, but only OEC transplantation promoted scaffold formation in the lesion site that facilitated axon regeneration and neuron preservation.

Dual Cortical Plasticity After Spinal Cord Injury.

During cortical development, plasticity reflects the dynamic equilibrium between increasing and decreasing functional connectivity subserved by synaptic sprouting and pruning. After adult cortical deafferentation, plasticity seems to be dominated by increased functional connectivity, leading to the classical expansive reorganization from the intact to the deafferented cortex. In contrast, here we show a striking "decrease" in the fast cortical responses to high-intensity forepaw stimulation 1-3 months after complete thoracic spinal cord transection, as evident in both local field potentials and intracellular in vivo recordings. Importantly, this decrease in fast cortical responses co-exists with an "increase" in cortical activation over slower post-stimulus timescales, as measured by an increased forepaw-to-hindpaw propagation of stimulus-triggered cortical up-states, as well as by the enhanced slow sustained depolarization evoked by high-frequency forepaw stimuli in the deafferented hindpaw cortex. This coincidence of diminished fast cortical responses and enhanced slow cortical activation offers a dual perspective of adult cortical plasticity after spinal cord injury.

Desenvolvimento da locomoção após injúria medular em pequenos animais – Revisão da literatura

http://dx.doi.org/10.5007/2175-7925.2016v29n2p1A injúria da medula espinhal ocorre comumente em cães e gatos e resulta em déficits neurológicos graves, podendo causar incapacidade sensitiva e motora permanente. Em pacientes com prognóstico reservado a ruim, faz-se uso dos recursos e tratamentos complementares na tentativa de desenvolver deambulação involuntária e incoordenada, que se aproxima à marcha normal, chamada comumente de andar medular ou caminhar reflexo. Sob condições experimentais, é relatado na literatura o retorno de um caminhar rítmico após completa transecção da medula espinhal. Em medicina veterinária, o papel do desenvolvimento da marcha reflexa é de extrema importância em relação à qualidade de vida dos pacientes afetados, sua independência e tranquilidade dos proprietários. O presente trabalho objetiva revisão de literatura a respeito do desenvolvimento do andar medular após injúria medular em pequenos animais.

Abnormal feeding behaviour in spinalised rats is mediated by hypothalamus: Restorative effect of exposure to extremely low frequency magnetic field.

Experimental study. To investigate the role of hypothalamus in abnormal feeding behaviour after spinal cord injury (SCI) and the effect of exposure to extremely low frequency magnetic field (ELF-MF) on it. India. Male Wistar rats (n=44) were divided into Sham (laminectomy), SCI (complete transection of T13 spinal cord), SCI+MF (ELF-MF exposure to SCI rats), VMHL (lesion of ventromedial hypothalamus; VMH), SCI+VMHL (VMHL after SCI) and SCI+VMHL+MF (ELF-MF exposure to SCI+VMHL rats) groups. Food intake (FI), water intake (WI), calorie intake (CI), body weight (BWT), taste preference and sucrose-induced biphasic (SIB) response to noxious stimulus were studied pre and post surgery. Neuronal activity at VMH was assessed by c-Fos immunohistochemistry. The extent of neuronal degeneration and regeneration in spinal cord was assessed microscopically. Data revealed post-SCI decrease in FI, WI, CI and BWT, preference for sodium chloride and citric acid, prolonged analgesic phase of SIB and increased c-Fos immunoreactivity in VMH of SCI rats vs Sham rats. VMH lesion increased FI, WI, CI, BW, preference for sweet tastants and abolished SIB, whereas in SCI+VMHL rats it abolished the effects of SCI on these parameters indicating probable involvement of VMH in SCI-induced alteration in feeding behaviour. Exposure to MF improved the study parameters in SCI rats and reduced the c-Fos immunoreactivity in VMH besides reduction in lesion volume, greater myelination and neuronal regeneration at SCI site. SCI influences VMH, leading to alteration in feeding behaviour, which is improved by exposure to ELF-MF.

Experimental Strategies to Bridge Large Tissue Gaps in the Injured Spinal Cord after Acute and Chronic Lesion.

After a spinal cord injury (SCI) a scar forms in the lesion core which hinders axonal regeneration. Bridging the site of injury after an insult to the spinal cord, tumor resections, or tissue defects resulting from traumatic accidents can aid in facilitating general tissue repair as well as regenerative growth of nerve fibers into and beyond the affected area. Two experimental treatment strategies are presented: (1) implantation of a novel microconnector device into an acutely and completely transected thoracic rat spinal cord to readapt severed spinal cord tissue stumps, and (2) polyethylene glycol filling of the SCI site in chronically lesioned rats after scar resection. The chronic spinal cord lesion in this model is a complete spinal cord transection which was inflicted 5 weeks before treatment. Both methods have recently achieved very promising outcomes and promoted axonal regrowth, beneficial cellular invasion and functional improvements in rodent models of spinal cord injury. The mechanical microconnector system (mMS) is a multi-channel system composed of polymethylmethacrylate (PMMA) with an outlet tubing system to apply negative pressure to the mMS lumen thus pulling the spinal cord stumps into the honeycomb-structured holes. After its implantation into the 1 mm tissue gap the tissue is sucked into the device. Furthermore, the inner walls of the mMS are microstructured for better tissue adhesion. In the case of the chronic spinal cord injury approach, spinal cord tissue - including the scar-filled lesion area - is resected over an area of 4 mm in length. After the microsurgical scar resection the resulting cavity is filled with polyethylene glycol (PEG 600) which was found to provide an excellent substratum for cellular invasion, revascularization, axonal regeneration and even compact remyelination in vivo.

The Sympathetic Nervous System Mediates Functional and Robust Respiratory Recovery following Acute Cervical Spinal Cord Injury

Introduction The organization of the respiratory network has traditionally been described through descending medullary tracks. The diaphragm is innervated by the phrenic nucleus (C4-C6). However, the intercostal muscles are innervated by the intercostal motor nucleus (T1-T11) located at the thoracic spinal cord segments. Therefore, cervical spinal cord injury (SCI) can typically cause respiratory motor system dysfunction. From the literature, ventilation failure within the human lower cervical SCI population typically occurs 4.5 ± 1.2 days after injury, affecting 26.6% of patients. Also, the risk of apnea following low cervical SCI is increased during sleep. However, while intercostal muscles contribute to respiratory recovery post injury the pathways mediating this effect have never been fully elucidated. Methods Following complete C7 spinal cord transection in rats, we demonstrated the presence of intercostal-EMG activity synchronized to the inspiratory bursts of the diaphragm. We explored the anatomical, and latent, pathway that enables synchronized activation of the intercostal muscles after complete cord transection. Experimental work, in the form of bilateral dorsal/ventral rhizotomies, unilateral transection of the phrenic nerve and sympathetic chain ganglia and systemic application of salubutamol, suggested that intercostal activities is mediated through the sympathetic chain ganglion by means of connection to the ipsilateral phrenic nerve. Results In anesitised and unventilated rats, rhythmic inspiratory activity was present in the intercostal muscles following acute C7 transection and bilateral dorsal/ventral rhizotomies. This activity persisted up to five days following the initial trauma. Unilateral transection of the phrenic nerve and sympathetic chain ganglia eliminated ipsilateral intercostal motor activity. Systemic application of salubutamol (β2-agonist), not only increase intercostal activity but also restored it in transected rats which had not retained this activity following transection. Conclusions These data provide the first evidence of sympathetic nerve activity mediating synchronized and robust intercostal inspiratory muscle activity following severe, lower cervical transection. This activity occurs up to 5 days following trauma, an effect which correlates with the induction of ventilation failure and respiratory dysfunction in human patients with SCI. The clinical implications of this finding are provocative. 60% of patients undergo some degree of respiratory distress following acute C5–8 injuries. Using this pathway, it may be possible to modestly stimulate or activate sympathetic activity to further aid intercostal muscle activity and respiratory function following acute SCI. This thus represents a novel target for the treatment of respiratory dysfunction following spinal trauma. [Figure: see text]

Sparing of Descending Axons Rescues Interneuron Plasticity in the Lumbar Cord to Allow Adaptive Learning After Thoracic Spinal Cord Injury.

This study evaluated the role of spared axons on structural and behavioral neuroplasticity in the lumbar enlargement after a thoracic spinal cord injury (SCI). Previous work has demonstrated that recovery in the presence of spared axons after an incomplete lesion increases behavioral output after a subsequent complete spinal cord transection (TX). This suggests that spared axons direct adaptive changes in below-level neuronal networks of the lumbar cord. In response to spared fibers, we postulate that lumbar neuron networks support behavioral gains by preventing aberrant plasticity. As such, the present study measured histological and functional changes in the isolated lumbar cord after complete TX or incomplete contusion (SCI). To measure functional plasticity in the lumbar cord, we used an established instrumental learning paradigm (ILP). In this paradigm, neural circuits within isolated lumbar segments demonstrate learning by an increase in flexion duration that reduces exposure to a noxious leg shock. We employed this model using a proof-of-principle design to evaluate the role of sparing on lumbar learning and plasticity early (7 days) or late (42 days) after midthoracic SCI in a rodent model. Early after SCI or TX at 7 days, spinal learning was unattainable regardless of whether the animal recovered with or without axonal substrate. Failed learning occurred alongside measures of cell soma atrophy and aberrant dendritic spine expression within interneuron populations responsible for sensorimotor integration and learning. Alternatively, exposure of the lumbar cord to a small amount of spared axons for 6 weeks produced near-normal learning late after SCI. This coincided with greater cell soma volume and fewer aberrant dendritic spines on interneurons. Thus, an opportunity to influence activity-based learning in locomotor networks depends on spared axons limiting maladaptive plasticity. Together, this work identifies a time dependent interaction between spared axonal systems and adaptive plasticity in locomotor networks and highlights a critical window for activity-based rehabilitation.

Targeted Inhibition of Leucine-Rich Repeat and Immunoglobulin Domain-Containing Protein 1 in Transplanted Neural Stem Cells Promotes Neuronal Differentiation and Functional Recovery in Rats Subjected to Spinal Cord Injury.

Leucine-rich repeat and immunoglobulin domain-containing protein (LINGO)-1 is expressed in neural stem cells, and its neutralization results in sustained neuronal immaturity. Thus, targeted inhibition of LINGO-1 via RNA interference may enhance transplanted neural stem cell survival and neuronal differentiation in vivo. Furthermore, LINGO-1 RNA interference in neural stem cells represents a potential therapeutic strategy for spinal cord injury. Department of Spine Surgery, First Affiliated Hospital of Sun Yat-sen University. Translational Medicine Center Research Laboratory, First Affiliated Hospital of Sun Yat-sen University. Female Sprague-Dawley rats. The animals were divided into three groups that underwent laminectomy and complete spinal cord transection accompanied by transplantation of control-RNA interference-treated or LINGO-1-RNA interference-treated neural stem cells at the injured site in vivo. In vitro, neural stem cells were divided into four groups for the following treatments: control, control RNA interference lentivirus, LINGO-1 RNA interference lentivirus and LINGO-1 complementary DNA lentivirusand the Key Projects of the Natural Science Foundation of Guangdong Province (No. S2013020012818). Neural stem cells in each treatment group were examined for cell survival and neuronal differentiation in vitro and in vivo via immunofluorescence and Western blot analysis. Axonal regeneration and tissue repair were assessed via retrograde tracing using Fluorogold, electron microscopy, hematoxylin-eosin staining and MRI. Rats were also examined for functional recovery based on the measurement of spinal cord-evoked potentials and the Basso-Beattie-Bresnahan score. LINGO-1-RNA interference-treated neural stem cell transplantation increased tissue repair and functional recovery of the injured spinal cord in rats. Similarly, LINGO-1 RNA interference increased neural stem cell survival and neuronal differentiation in vitro. The mechanism underlying the effect of LINGO-1 RNA interference on the injured rat spinal cord may be that the significant inhibition of LINGO-1 expression in neural stem cells inactivated the RhoA and Notch signaling pathways, which act downstream of LINGO-1. Our findings indicate that transplantation of LINGO-1-RNA interference-treated neural stem cells facilitates functional recovery after spinal cord injury and represents a promising potential strategy for the repair of spinal cord injury.

Gait recovery following spinal cord injury in mice: Limited effect of treadmill training

Background: Several studies in rodents with complete spinal cord transections have demonstrated that treadmill training improves stepping movements. However, results from studies in incomplete spinal cord injured animals have been conflicting and questions regarding the training dosage after injury remain unresolved.Objectives: To assess the effects of treadmill-training regimen (20 minutes daily, 5 days a week) for 3, 6 or 9 weeks on the recovery of locomotion in hemisected SCI mice.Methods: A randomized and blinded controlled experimental trial used a mouse model of incomplete spinal cord injury (SCI). After a left hemisection at T10, adult male mice were randomized to trained or untrained groups. The trained group commenced treadmill training one week after surgery and continued for 3, 6 or 9 weeks. Quantitative kinematic gait analysis was used to assess the spatiotemporal characteristics of the left hindlimb prior to injury and at 1, 4, 7 and 10 weeks post-injury.Results: One week after injury there was no movement of the left hindlimb and some animals dragged their foot. Treadmill training led to significant improvements in step duration, but had limited effect on the hindlimb movement pattern. Locomotor improvements in trained animals were most evident at the hip and knee joints whereas recovery of ankle movement was limited, even after 9 weeks of treadmill training.Conclusion: These results demonstrate that treadmill training may lead to only modest improvement in recovery of hindlimb movement after incomplete spinal cord injury in mice.

Rigid and remodelled: cerebrovascular structure and function after experimental high-thoracic spinal cord transection.

High-thoracic or cervical spinal cord injury (SCI) is associated with several critical clinical conditions related to impaired cerebrovascular health, including: 300-400% increased risk of stroke, cognitive decline and diminished cerebral blood flow regulation. The purpose of this study was to examine the influence of high-thoracic (T3 spinal segment) SCI on cerebrovascular structure and function, as well as molecular markers of profibrosis. Seven weeks after complete T3 spinal cord transection (T3-SCI, n = 15) or sham injury (Sham, n = 10), rats were sacrificed for either middle cerebral artery (MCA) structure and function assessments via ex vivo pressure myography, or immunohistochemical analyses. Myogenic tone was unchanged, but over a range of transmural pressures, inward remodelling occurred after T3-SCI with a 40% reduction in distensibility (both P < 0.05), and a 33% reduction in vasoconstrictive reactivity to 5-HT trending toward significance (P = 0.09). After T3-SCI, the MCA had more collagen I (42%), collagen III (24%), transforming growth factor β (47%) and angiotensin II receptor type 2 (132%), 27% less elastin as well as concurrent increased wall thickness and reduced lumen diameter (all P < 0.05). Sympathetic innervation (tyrosine hydroxylase-positive axon density) and endothelium-dependent dilatation (carbachol) of the MCA were not different between groups. This study demonstrates profibrosis and hypertrophic inward remodelling within the largest cerebral artery after high-thoracic SCI, leading to increased stiffness and possibly impaired reactivity. These deleterious adaptations would substantially undermine the capacity for regulation of cerebral blood flow and probably underlie several cerebrovascular clinical conditions in the SCI population.

Characterization of dendritic morphology and neurotransmitter phenotype of thoracic descending propriospinal neurons after complete spinal cord transection and GDNF treatment

After spinal cord injury (SCI), poor regeneration of damaged axons of the central nervous system (CNS) causes limited functional recovery. This limited spontaneous functional recovery has been attributed, to a large extent, to the plasticity of propriospinal neurons, especially the descending propriospinal neurons (dPSNs). Compared with the supraspinal counterparts, dPSNs have displayed significantly greater regenerative capacity, which can be further enhanced by glial cell line-derived neurotrophic factor (GDNF). In the present study, we applied a G-mutated rabies virus (G-Rabies) co-expressing green fluorescence protein (GFP) to reveal Golgi-like dendritic morphology of dPSNs. We also investigated the neurotransmitters expressed by dPSNs after labeling with a retrograde tracer Fluoro-Gold (FG). dPSNs were examined in animals with sham injuries or complete spinal transections with or without GDNF treatment. Bilateral injections of G-Rabies and FG were made into the 2nd lumbar (L2) spinal cord at 3days prior to a spinal cord transection performed at the 11th thoracic level (T11). The lesion gap was filled with Gelfoam containing either saline or GDNF in the injury groups. Four days post-injury, the rats were sacrificed for analysis. For those animals receiving G-rabies injection, the GFP signal in the T7-9 spinal cord was visualized via 2-photon microscopy. Dendritic morphology from stack images was traced and analyzed using a Neurolucida software. We found that dPSNs in sham injured animals had a predominantly dorsal-ventral distribution of dendrites. Transection injury resulted in alterations in the dendritic distribution with dorsal-ventral retraction and lateral-medial extension. Treatment with GDNF significantly increased the terminal dendritic length of dPSNs. The density of spine-like structures was increased after injury, and treatment with GDNF enhanced this effect. For the group receiving FG injections, immunohistochemistry for glutamate, choline acetyltransferase (ChAT), glycine, and GABA was performed in the T7-9 spinal cord. We show that the majority of FG retrogradely-labeled dPSNs were located in the Rexed Lamina VII. Over 90% of FG-labeled neurons were glutamatergic, with the other three neurotransmitters contributing less than 10% of the total. To our knowledge this is the first report describing the morphologic characteristics of dPSNs and their neurotransmitter expressions, as well as the dendritic response of dPSNs after transection injury and GDNF treatment.

Functionalized collagen scaffold implantation and cAMP administration collectively facilitate spinal cord regeneration

Previous studies have demonstrated that several mechanisms, including numerous inhibitory molecules, weak neurotrophic stimulation and deficient intrinsic regenerative responses, collectively contribute to the failure of mature spinal cord axon regeneration. Thus, combinatorial therapies targeting multiple mechanisms have attracted much attention. In the present study, a porous collagen scaffold was used to support neuronal attachment and bridge axonal regeneration. The scaffold was specifically functionalized using neutralizing proteins (CBD-EphA4LBD, CBD-PlexinB1LBD and NEP1-40) and collagen-binding neurotrophic factors (CBD-BDNF and CBD-NT3) to simultaneously antagonize myelin inhibitory molecules (ephrinB3, Sema4D and Nogo) and exert neurotrophic protection and stimulation. Cerebellar granular neurons cultured on the functionalized collagen scaffold promoted neurite outgrowth in the presence of myelin. Furthermore, a full combinatorial treatment comprising functionalized scaffold implantation and cAMP administration was developed to evaluate the synergistic repair ability in a rat T10 complete removal spinal cord injury model. The results showed that full combinatorial therapy exhibited the greatest advantage in reducing the volume of cavitation, facilitating axonal regeneration, and promoting neuronal generation. The newborn neurons generated in the lesion area could form the neuronal relay and enhance the locomotion recovery after severe spinal cord injury. Statement of SignificanceA porous collagen scaffold was specifically functionalized with neutralizing proteins and neurotrophic factors to antagonize the myelin inhibitory molecules and exert neurotrophic protection and stimulation for spinal cord regeneration. Cerebellar granular neurons seeded on the functionalized collagen scaffold showed enhanced neurite outgrowth ability in vitro. The functionalized scaffold implantation combined with cAMP administration exhibited synergistic repair ability for rat T10 complete spinal cord transection injury.

Inosine Improves Neurogenic Detrusor Overactivity following Spinal Cord Injury.

Neurogenic detrusor overactivity and the associated loss of bladder control are among the most challenging complications of spinal cord injury (SCI). Anticholinergic agents are the mainstay for medical treatment of detrusor overactivity. However, their use is limited by significant side effects such that a search for new treatments is warranted. Inosine is a naturally occurring purine nucleoside with neuroprotective, neurotrophic and antioxidant effects that is known to improve motor function in preclinical models of SCI. However, its effect on lower urinary tract function has not been determined. The objectives of this study were to determine the effect of systemic administration of inosine on voiding function following SCI and to delineate potential mechanisms of action. Sprague−Dawley rats underwent complete spinal cord transection, or cord compression by application of an aneurysm clip at T8 for 30 sec. Inosine (225 mg/kg) or vehicle was administered daily via intraperitoneal injection either immediately after injury or after a delay of 8 wk. At the end of treatment, voiding behavior was assessed by cystometry. Levels of synaptophysin (SYP), neurofilament 200 (NF200) and TRPV1 in bladder tissues were measured by immunofluorescence imaging. Inosine administration decreased overactivity in both SCI models, with a significant decrease in the frequency of spontaneous non−voiding contractions during filling, compared to vehicle−treated SCI rats (p<0.05), including under conditions of delayed treatment. Immunofluorescence staining demonstrated increased levels of the pan-neuronal marker SYP and the Adelta fiber marker NF200, but decreased staining for the C-fiber marker, TRPV1 in bladder tissues from inosine-treated rats compared to those from vehicle-treated animals, including after delayed treatment. These findings demonstrate that inosine prevents the development of detrusor overactivity and attenuates existing overactivity following SCI, and may achieve its effects through modulation of sensory neurotransmission.