Spinal Cord Of Animals Research Articles

GAP-43 Protein and Its Proteolytic Fragment in Spinal Cord Cells in Rats with Experimental Allergic Encephalomyelitis

The limited regenerative capacity of the central nervous system (CNS) is one of the key factors promoting the development of neurodegenerative diseases, including multiple sclerosis. The regenerative potential of CNS cells can be evaluated in terms of the level of GAP-43 protein, which is involved in the processes of neuritogenesis and growth cone navigation. Cleavage of GAP-43 by m-calpain to form the fragment GAP-43-3, which lacks 40 N-terminal amino acids, probably leads to loss of the initial functions of the protein, which may have adverse influences on the regenerative potential of the CNS. The present study was performed on the most appropriate model of multiple sclerosis – experimental allergic encephalomyelitis (EAE) in rats. The spinal cord of animals with EAE was shown to have inflammatory infiltrates and deformed motoneurons; limb pareses and paralysis developed. When EAE was mild, about 60% of GAP-43 in rat spinal cord cells was present in the form of GAP-43-3. In severe forms of EAE, with tetraplegia, the extent of GAP-43 proteolysis by calpain reached 85%. In this situation, GAP-43-3 was found mainly in microglial cells. We suggest that during the development of EAE, proteolytic cleavage of GAP-43 protein by m-calpain to form GAP-43-3 can significantly alter its properties, suppressing the regenerative ability of CNS cells and exacerbating the course of the pathological process.

604. Comparison of Intra-Cisterna Magna and Lumbar Puncture Intrathecal Delivery of scAAV9 GeneTherapy for Giant Axonal Neuropathy

Giant axonal neuropathy (GAN) is a rare pediatric neurodegenerative disorder characterized by progressive neuropathy that presents as early as 3 years of age and with ultimate mortality during the second or third decade of life. GAN is caused by autosomal recessive loss-of-function mutations in the GAN gene that encodes for the gigaxonin protein. Gigaxonin plays a role in the organization/degradation of intermediate filaments (IFs) and GAN patients are pathologically characterized by large axonal swellings filled with disorganized aggregates of IFs. While GAN is primarily described as a progressive peripheral neuropathy, diffuse pathology from disorganized IFs is apparent throughout the central nervous system, enteric nervous system and other organ systems. An NIH-sponsored Phase I study is underway to test the safety of intrathecal lumbar puncture (LP) administration of scAAV9/JeT-GAN to treat the most severe aspects of GAN, namely the motor and sensory neuropathy. Gigaxonin gene transfer through a single LP injection is the first proposed therapy for GAN. Intra-cisterna magna (ICM) delivery of AAV9 vectors shows high transduction of the brain and spinal cord of animals; however, this method of vector delivery has not yet been tested for the treatment of GAN. This study compared the efficacy of using ICM or LP delivery of the scAAV9/JeT-GAN vector to treat GAN KO mice. GAN KO mice were injected with scAAV9/JeT-GAN at 15 months of age using ICM or LP delivery and motor performance was tested monthly. In agreement with our previous studies, we found that GAN KO mice have impaired motor performance around 20 months of age and that this deficit is attenuated with LP delivery of scAAV9/JeT-GAN. In contrast, GAN KO mice receiving ICM-delivered scAAV9/JeT-GAN did not have significantly improved motor function as compared to vehicle treated GAN KO mice. Analyses of disease-relevant pathologies in the brain, spinal cord, and peripheral nerves of these mice are ongoing. To date, antibodies are not available to reliably detect gigaxonin protein expression via immunohistochemical (IHC) analysis, so to directly compare the transduction of the two intrathecal delivery methods, we injected wild-type mice with ~4 × 1011 vg scAAV9/GFP via an ICM or LP injection, which is a higher dose than has been reported in any publication. Mice were harvested 4-weeks post-injection and the biodistribution of scAAV9 was analyzed via semi-quantitative PCR and IHC analysis of GFP. Compared to LP-injected mice, GFP expression was notably higher and more wide-spread in the brains of ICM-injected mice, while higher amounts of GFP were detected in the sciatic nerves of LP-injected mice as compared to ICM-injected mice. IHC analysis is ongoing and we will present a detailed map of the transduction of ICM- and LP-delivered scAAV9/GFP across the central and peripheral nervous systems and different organ systems. In conclusion, we report here that ICM-delivery of scAAV9/JeT-GAN does not attenuate motor deficits in GAN KO mice. Furthermore, the vector spread varies between ICM and LP delivery, suggesting that the site of intrathecal injection may have a critical impact on the therapeutic benefit of gene vectors for a given disease.

Local versus distal transplantation of human neural stem cells following chronic spinal cord injury

Background ContextPrevious studies have demonstrated functional recovery of rats with spinal cord contusions after transplantation of neural stem cells adjacent to the site of acute injury. PurposeThe purpose of the study was to determine if the local or distal injection of neural stem cells can cause functional difference in recovery after chronic spinal cord injury. Study Design/SettingTwenty-four adult female Long-Evans hooded rats were randomized into four groups, with six animals in each group: two experimental and two control groups. Functional assessment was measured after injury and then weekly for 6 weeks using the Basso, Beattie, and Bresnahan locomotor rating score. Data were analyzed using two-sample t test and linear mixed-effects model analysis. MethodsPosterior exposure and laminectomy at the T10 level was used. Moderate spinal cord contusion was induced by the Multicenter Animal Spinal Cord Injury Study Impactor with 10-g weight dropped from a height of 25 mm. Experimental subjects received either a subdural injection of human neural stem cells (hNSCs) locally at the injury site or intrathecal injection of hNSCs through a separate distal laminotomy 4 weeks after injury. Controls received control media injection either locally or distally. ResultsA statistically significant functional improvement in subjects that received hNSCs injected distally to the site of injury was observed when compared with the control (p=.042). The difference between subjects that received hNSCs locally and the control did not reach statistical significance (p=.085). ConclusionsThe transplantation of hNSCs into the contused spinal cord of a rat led to significant functional recovery of the spinal cord when injected distally but not locally to the site of chronic spinal cord injury.

Contusive spinal cord injury up regulates mu-opioid receptor (mor) gene expression in the brain and down regulates its expression in the spinal cord: possible implications in spinal cord injury research

Traumatic spinal cord injury (SCI) is one of the dreaded neurological conditions and finding a cure for it has been a hot area of research. Naloxone – a mu-opiate receptor (mor) antagonist was considered for SCI treatment based on its positive effects under shock conditions. In contrary to animal studies based reports about the potential benefits of naloxone in treating SCI, a large scale clinical trial [National Acute Spinal Cord Injury Study II (NASCIS II)] conducted in USA failed to witness any effectiveness. The inconsistency noticed was intriguing. Therefore, the objective of the present study was to re-examine the role of naloxone in treating SCI using a highly standardised Multicenter Animal Spinal Cord Injury Study (MASCIS) animal model of contusive SCI. Results indicated that naloxone produced negligible and insignificant neuroprotection. In an attempt to understand the cause for the failure, it was found that mu-opioid receptor (mor) gene expression was upregulated in the brain but was down regulated in the spinal cord after contusive SCI. Given that the beneficial effects of naloxone are through its action on the mor, the results indicate that unlike the brain, spinal cord might not be bracing to utilise the opiate system in the repair process. This could possibly explain the failure of naloxone treatment in NASCIS II. To conclude, opiate antagonists like naloxone may be neuroprotective for treating traumatic brain injuries, but not for traumatic/contusive spinal cord injuries.

Efficacy and Safety of Riluzole in Acute Spinal Cord Injury: Rationale and Design of AOSpine Phase III Multicenter Double Blinded Randomized Controlled Trial

Introduction There is convincing evidence from the preclinical realm that the pharmacologic agent riluzole attenuates certain aspects of the secondary injury cascade leading to diminished neurological tissue destruction in animal spinal cord injury (SCI) models. The safety and pharmacokinetic profile of riluzole has been studied in a multicenter pilot study in 36 patients. Efficacy of riluzole in acute human SCI has not been established. Material and Methods This phase II/III multicenter double-blind randomized controlled trial will involve up to 35 sites. A total of 351 patients with acute C4–C8 SCI and ASIA Impairment Grade A, B, or C will be randomized 1:1 to riluzole and placebo. Primary outcome is the change in ASIA Total Motor Score between baseline and 180 days. Other measures include ASIA Upper Extremity Motor Score; ASIA Lower Extremity Motor Score; ASIA Sensory Score; ASIA Grade; Spinal Cord Independence Measure; SF-36v2; EQ-5D, and GRASSP. The statistical design utilizes two-stage sequential adaptive trial. A sample size of 316 patients (158 in each arm) will have 90% power to detect 9 points difference in the ASIA Motor Score at one-sided α = 0.025. To account for losses to follow-up of up to 10%, the study will enroll 351 patients. Results Within the phase 1 study, a matched cohort analysis was performed comparing complication rates and neurological outcomes between patients who received riluzole and non-riluzole-treated patients from a prospective SCI registry. Although the groups experienced similar rates of complications, riluzole-treated cervical SCI patients experienced an additional 15.5 points in AMS recovery at 90 days after injury as compared with non-riluzole-treated patients. Although the phase I study was underpowered to investigate the efficacy, the current phase II/III study is poised to definitively address this question. Patient enrollment for this trial began on October 1, 2013. Conclusion This is a pivotal study of riluzole in acute SCI.

Electrical stimulation and motor recovery.

In recent years, several investigators have successfully regenerated axons in animal spinal cords without locomotor recovery. One explanation is that the animals were not trained to use the regenerated connections. Intensive locomotor training improves walking recovery after spinal cord injury (SCI) in people, and >90% of people with incomplete SCI recover walking with training. Although the optimal timing, duration, intensity, and type of locomotor training are still controversial, many investigators have reported beneficial effects of training on locomotor function. The mechanisms by which training improves recovery are not clear, but an attractive theory is available. In 1949, Donald Hebb proposed a famous rule that has been paraphrased as "neurons that fire together, wire together." This rule provided a theoretical basis for a widely accepted theory that homosynaptic and heterosynaptic activity facilitate synaptic formation and consolidation. In addition, the lumbar spinal cord has a locomotor center, called the central pattern generator (CPG), which can be activated nonspecifically with electrical stimulation or neurotransmitters to produce walking. The CPG is an obvious target to reconnect after SCI. Stimulating motor cortex, spinal cord, or peripheral nerves can modulate lumbar spinal cord excitability. Motor cortex stimulation causes long-term changes in spinal reflexes and synapses, increases sprouting of the corticospinal tract, and restores skilled forelimb function in rats. Long used to treat chronic pain, motor cortex stimuli modify lumbar spinal network excitability and improve lower extremity motor scores in humans. Similarly, epidural spinal cord stimulation has long been used to treat pain and spasticity. Subthreshold epidural stimulation reduces the threshold for locomotor activity. In 2011, Harkema et al. reported lumbosacral epidural stimulation restores motor control in chronic motor complete patients. Peripheral nerve or functional electrical stimulation (FES) has long been used to activate sacral nerves to treat bladder and pelvic dysfunction and to augment motor function. In theory, FES should facilitate synaptic formation and motor recovery after regenerative therapies. Upcoming clinical trials provide unique opportunities to test the theory.

Cell Therapy Augments Functional Recovery Subsequent to Spinal Cord Injury under Experimental Conditions.

The spinal cord injury leads to enervation of normal tissue homeostasis ultimately leading to paralysis. Until now there is no proper cure for the treatment of spinal cord injury. Recently, cell therapy in animal spinal cord injury models has shown some progress of recovery. At present, clinical trials are under progress to evaluate the efficacy of cell transplantation for the treatment of spinal cord injury. Different types of cells such as pluripotent stem cells derived neural cells, mesenchymal stromal cells, neural stem cells, glial cells are being tested in various spinal cord injury models. In this review we highlight both the advances and lacuna in the field of spinal cord injury by discussing epidemiology, pathophysiology, molecular mechanism, and various cell therapy strategies employed in preclinical and clinical injury models and finally we discuss the limitations and ethical issues involved in cell therapy approach for treating spinal cord injury.

Spinal cord injury models: a review

Animal spinal cord injury (SCI) models have proved invaluable in better understanding the mechanisms involved in traumatic SCI and evaluating the effectiveness of experimental therapeutic interventions. Over the past 25 years, substantial gains have been made in developing consistent, reproducible and reliable animal SCI models. Review. The objective of this review was to consolidate current knowledge on SCI models and introduce newer paradigms that are currently being developed. SCI models are categorized based on the mechanism of injury into contusion, compression, distraction, dislocation, transection or chemical models. Contusion devices inflict a transient, acute injury to the spinal cord using a weight-drop technique, electromagnetic impactor or air pressure. Compression devices compress the cord at specific force and duration to cause SCI. Distraction SCI devices inflict graded injury by controlled stretching of the cord. Mechanical displacement of the vertebrae is utilized to produce dislocation-type SCI. Surgical transection of the cord, partial or complete, is particularly useful in regenerative medicine. Finally, chemically induced SCI replicates select components of the secondary injury cascade. Although rodents remain the most commonly used species and are best suited for preliminary SCI studies, large animal and nonhuman primate experiments better approximate human SCI. All SCI models aim to replicate SCI in humans as closely as possible. Given the recent improvements in commonly used models and development of newer paradigms, much progress is anticipated in the coming years.

Spinal cord regeneration.

Three theories of regeneration dominate neuroscience today, all purporting to explain why the adult central nervous system (CNS) cannot regenerate. One theory proposes that Nogo, a molecule expressed by myelin, prevents axonal growth. The second theory emphasizes the role of glial scars. The third theory proposes that chondroitin sulfate proteoglycans (CSPGs) prevent axon growth. Blockade of Nogo, CSPG, and their receptors indeed can stop axon growth in vitro and improve functional recovery in animal spinal cord injury (SCI) models. These therapies also increase sprouting of surviving axons and plasticity. However, many investigators have reported regenerating spinal tracts without eliminating Nogo, glial scar, or CSPG. For example, many motor and sensory axons grow spontaneously in contused spinal cords, crossing gliotic tissue and white matter surrounding the injury site. Sensory axons grow long distances in injured dorsal columns after peripheral nerve lesions. Cell transplants and treatments that increase cAMP and neurotrophins stimulate motor and sensory axons to cross glial scars and to grow long distances in white matter. Genetic studies deleting all members of the Nogo family and even the Nogo receptor do not always improve regeneration in mice. A recent study reported that suppressing the phosphatase and tensin homolog (PTEN) gene promotes prolific corticospinal tract regeneration. These findings cannot be explained by the current theories proposing that Nogo and glial scars prevent regeneration. Spinal axons clearly can and will grow through glial scars and Nogo-expressing tissue under some circumstances. The observation that deleting PTEN allows corticospinal tract regeneration indicates that the PTEN/AKT/mTOR pathway regulates axonal growth. Finally, many other factors stimulate spinal axonal growth, including conditioning lesions, cAMP, glycogen synthetase kinase inhibition, and neurotrophins. To explain these disparate regenerative phenomena, I propose that the spinal cord has evolved regenerative mechanisms that are normally suppressed by multiple extrinsic and intrinsic factors but can be activated by injury, mediated by the PTEN/AKT/mTOR, cAMP, and GSK3b pathways, to stimulate neural growth and proliferation.

The role of FGF2 in spinal cord trauma and regeneration research

The role of <scp>FGF</scp>2 in spinal cord trauma and regeneration research

Corticospinal tract sprouting in the injured rat spinal cord stimulated by Schwann cell preconditioning of the motor cortex

Peripheral nerve preconditioning lesions have been shown to consistently enhance sensory nerve regeneration in the injured spinal cord.Objective: The aim of this study was to determine if the rat motor cortex could be preconditioned through the implantation of Schwann cells (SC), thereby stimulating sprouting and regeneration of the corticospinal tract (CST).Methods: Schwann cells (cultured in vitro) were injected into the motor cortex and seven days post-surgery rats underwent a dorsal spinal hemisection injury. Eight weeks following spinal injury animals were perfused and the CST visualised by Avidin-peroxidase histochemistry for dextran-biotin.Results: Results demonstrate substantially enhanced CST collateral sprouting in both the rostral grey and white matter of the injured spinal cord in animals with SC implanted into the motor cortex compared to control animals with and without cortical vehicle injections. Corticospinal tract peri-wound regenerative sprouting was also enhanced in animals implanted with cortical SC compared to controls, however, only a small degree of CST axonal regeneration was present in the grey matter beneath the injury site. In all groups, CST peri-lesional regenerative sprouting occurred in close proximity to macrophages. Complicated and intimate relationships between CST fibres and these cells were evident.Discussion: Overall, our data demonstrates that preconditioning the motor cortex with SC prior to spinal injury results in greatly enhanced CST sprouting and that CST peri-wound sprouting takes place in juxtaposition to macrophages.

Photobiomodulation Induced by 670 nm Light Ameliorates MOG35-55 Induced EAE in Female C57BL/6 Mice: A Role for Remediation of Nitrosative Stress

BackgroundExperimental autoimmune encephalomyelitis (EAE) is the most commonly studied animal model of multiple sclerosis (MS), a chronic autoimmune demyelinating disorder of the central nervous system. Immunomodulatory and immunosuppressive therapies currently approved for the treatment of MS slow disease progression, but do not prevent it. A growing body of evidence suggests additional mechanisms contribute to disease progression. We previously demonstrated the amelioration of myelin oligodendrocyte glycoprotein (MOG)-induced EAE in C57BL/6 mice by 670 nm light-induced photobiomodulation, mediated in part by immune modulation. Numerous other studies demonstrate that near-infrared/far red light is therapeutically active through modulation of nitrosoxidative stress. As nitric oxide has been reported to play diverse roles in EAE/MS, and recent studies suggest that axonal loss and progression of disability in MS is mediated by nitrosoxidative stress, we investigated the effect of 670 nm light treatment on nitrosative stress in MOG-induced EAE.MethodologyCell culture experiments demonstrated that 670 nm light-mediated photobiomodulation attenuated antigen-specific nitric oxide production by heterogenous lymphocyte populations isolated from MOG immunized mice. Experiments in the EAE model demonstrated down-regulation of inducible nitric oxide synthase (iNOS) gene expression in the spinal cords of mice with EAE over the course of disease, compared to sham treated animals. Animals receiving 670 nm light treatment also exhibited up-regulation of the Bcl-2 anti-apoptosis gene, an increased Bcl-2:Bax ratio, and reduced apoptosis within the spinal cord of animals over the course of disease. 670 nm light therapy failed to ameliorate MOG-induced EAE in mice deficient in iNOS, confirming a role for remediation of nitrosative stress in the amelioration of MOG-induced EAE by 670 nm mediated photobiomodulation.ConclusionsThese data indicate that 670 nm light therapy protects against nitrosative stress and apoptosis within the central nervous system, contributing to the clinical effect of 670 nm light therapy previously noted in the EAE model.

SU-E-T-307: Methodology for Spinal Cord Retreat Calculations

Purpose: Often in the clinical arena, physicians rely on medical physicists to provide dose estimates. This project focused on evaluating the BED to the spinal cord for patients undergoing retreatment after conventionally fractionated RT. The BED calculation method is based on published research on human and animal spinal cord reirradiations. Methods: BED is calculated separately for the first and upcoming treatments using the planning system to estimate the dose to the spinal cord in the overlap region from each course. The cumulative BED is calculated as the sum of the BEDs of the first treatment and the upcoming treatment. The tolerance BED is calculated from the QUANTEC (2010) value for 0.2% rate of myelopathy of 50 Gy in 2 Gy fractions. If the reirradiation follows the initial irradiation by at least six months, the cumulative BED is compared to the tolerance BED multiplied by 1.25 to account for repair of RT‐induced subclinical damage. If an RT course was delivered BID, then an additional factor (Hm = 0.35) was used to account for incomplete repair of sublethal damage during the BID course. Results: BED calculations were performed using this method to assist clinical decisions for spinal cord reirradiations. Patients were treated to doses as high as 59 Gy to the spinal cord while BED calculations showed the dose to the spinal cord was effectively below the QUANTEC tolerance for 0.2% rate of myelopathy. Conclusion: Application of this BED calculation methodology for spinal reirradiations places retreatment dose in the context of current clinical research. The department is analyzing spinal cord BED calculations factoring in recovery observed in a primate model predicting 50%, 60%, and 70% recovery of the initial RT course dose at 1 year, 2 years, and 3 years post‐RT respectively given an initial treatment of up to 44 Gy.

An experimental spinal cord injury rat model using customized impact device: A cost-effective approach

Till date, NYU MASCIS (New York University, Multicenter Animal Spinal Cord Injury Study) impactor and Ohio State University electromagnetic spinal cord injury device impactor were under use for simulating an experimental spinal cord injury in rodents; functional recovery being assessed through Basso, Beattie and Bresnahan (BBB) scoring method which is an open field behavior based scoring system. Although, the cited impactors are state-of-art devices, affordability to scientists in developing and under developed countries is questionable. Since the acquisition of these impact devices are expensive, we designed a customized impact device based on the requirement, satisfying all the parameters to withstand a standard animal model for contusion type of spinal cord injury at the thoracic level without compromising the lesion reproducibility. Here, a spinal cord contusion is created using a blunt-force impactor in male Wistar rats. Our method gave consistent lesion effects as evaluated by behavior scoring methods. All the animals showed equal degree of performance in tests like narrow beam, inclined plane and horizontal ladder and in BBB scores (open field locomotor test). The aim of presenting our experience is to reinstate the fact that lack of affordability to get sophisticated instrumentation need not be a hurdle in the pursuit of science.

Autologous olfactory mucosal cell transplants in clinical spinal cord injury: a randomized double-blinded trial in a canine translational model.

This study was designed to determine whether an intervention proven effective in the laboratory to ameliorate the effects of experimental spinal cord injury could provide sufficient benefit to be of value to clinical cases. Intraspinal olfactory ensheathing cell transplantation improves locomotor outcome after spinal cord injury in ‘proof of principle’ experiments in rodents, suggesting the possibility of efficacy in human patients. However, laboratory animal spinal cord injury cannot accurately model the inherent heterogeneity of clinical patient cohorts, nor are all aspects of their spinal cord function readily amenable to objective evaluation. Here, we measured the effects of intraspinal transplantation of cells derived from olfactory mucosal cultures (containing a mean of ∼50% olfactory ensheathing cells) in a population of spinal cord–injured companion dogs that accurately model many of the potential obstacles involved in transition from laboratory to clinic. Dogs with severe chronic thoracolumbar spinal cord injuries (equivalent to ASIA grade ‘A’ human patients at ∼12 months after injury) were entered into a randomized double-blinded clinical trial in which they were allocated to receive either intraspinal autologous cells derived from olfactory mucosal cultures or injection of cell transport medium alone. Recipients of olfactory mucosal cell transplants gained significantly better fore–hind coordination than those dogs receiving cell transport medium alone. There were no significant differences in outcome between treatment groups in measures of long tract functionality. We conclude that intraspinal olfactory mucosal cell transplantation improves communication across the damaged region of the injured spinal cord, even in chronically injured individuals. However, we find no evidence for concomitant improvement in long tract function.

Phosphorylation of ezrin/radixin/moesin (ERM) protein in spinal microglia following peripheral nerve injury and lysophosphatidic acid administration

Peripheral nerve injury activates spinal glial cells, which may contribute to the development of pain behavioral hypersensitivity. There is growing evidence that activated microglia show dynamic changes in cell morphology; however, the molecular mechanisms that underlie the modification of the membrane and cytoskeleton of microglia are not known. Here, we investigated the phosphorylation of ezrin, radixin, and moesin (ERM) proteins in the spinal cord after peripheral nerve injury. ERM is known to function as membrane-cytoskeletal linkers and be localized at filopodia- and microvilli-like structures. ERM proteins must be phosphorylated at a specific C-terminal threonine residue to be in the active state. The nature of ERM proteins in the spinal cord of animals in a neuropathic pain model has not been investigated and characterized. In the present study, we observed an increase in the phosphorylated ERM in the spinal microglia following spared nerve injury. The intrathecal administration of lysophosphatidic acid induced the phosphorylation of ERM proteins in microglia along with the development of mechanical pain hypersensitivity. Intrathecal administration of ERM antisense locked nucleic acid suppressed nerve injury-induced tactile allodynia and decreased the phosphorylation of ERM, but not the Iba1 staining pattern, in spinal glial cells. These findings suggest that lysophosphatidic acid induced the phosphorylation of ERM proteins in spinal microglia and may be involved in the emergence of neuropathic pain. These findings may underlie the pathological mechanisms of nerve injury-induced neuropathic pain.

Functional assessment of the acute local and distal transplantation of human neural stem cells after spinal cord injury

Spinal cord injury can lead to severe functional impairments secondary to axonal damage, neuronal loss, and demyelination. The injured spinal cord has limited regrowth of damaged axons. Treatment remains controversial, given inconsistent functional improvement. Previous studies demonstrated functional recovery of rats with spinal cord contusion after transplantation of rat fetal neural stem cells. We hypothesized that acute transplantation of human fetal neural stem cells (hNSCs) both locally at the injury site as well as distally via intrathecal injection would lead to improved functional recovery compared with controls. Twenty-four adult female Long-Evans hooded rats were randomized into four groups with six animals in each group: two experimental and two control. Functional assessment was measured after injury and then weekly for 6 weeks using the Basso, Beattie, and Bresnahan Locomotor Rating Score. Data were analyzed using two-sample t test and linear mixed-effects model analysis. Posterior exposure and laminectomy at T10 level was used. Moderate spinal cord contusion was induced by the Multicenter Animal Spinal Cord Injury Study Impactor with 10-g weight dropped from a height of 25 mm. Experimental subjects received either a subdural injection of hNSCs locally at the injury site or intrathecal injection of hNSCs through a separate distal laminotomy. Controls received control media injection either locally or distally. Statistically significant functional improvement was observed in local or distal hNSCs subjects versus controls (p=.034 and 0.016, respectively). No significant difference was seen between local or distal hNSC subjects (p=.66). Acute local and distal transplantation of hNSCs into the contused spinal cord led to significant functional recovery in the rat model. No statistical difference was found between the two techniques.

Effects of Spinal Cord Electrical Stimulation in Patients with Vertebrospinal Pathology

Until now, no scientific neurophysiologic methods of diagnostics and treatment of vertebrospinal pathologies were developed. Previous study showed that electrical stimulation of lumbar segments of the spinal cord in animals with complete spinal cord transection induced a well-coordinated weight-bearing locomotion. The present comparative study of motor activity triggered by electrical epidural stimulation of one or two segments of the spinal cord in spinal patients showed that stimulation of lumbar (L2-L4) or sacral (S2) segments facilitated generation of motor patterns of muscle activity. Combination of electrical stimulation with locomotor training resulted in the appearance of stepping patterns characteristic of normal walking and tonic activity of the muscles needed for body balance maintenance.

Results of a phase II placebo-controlled randomized trial of minocycline in acute spinal cord injury

Preclinical studies have attributed neuroprotective properties to the antibiotic minocycline. Animal studies and early clinical trials support its use in several neurological diseases. In animal spinal cord injury models, minocycline improved neurological and histological outcomes, reduced neuronal and oligodendroglial apoptosis, decreased microglial activation and reduced inflammation. A single-centre, human, double-blind, randomized, placebo-controlled study of minocycline administration after spinal cord injury was undertaken for the purposes of dose optimization, safety assessment and to estimate outcome changes and variance. Neurological, functional, pharmacological and adverse event outcomes were compared between subjects administered 7 days of intravenous minocycline (n = 27) or placebo (n = 25) after acute traumatic spinal cord injury. The secondary outcome used to assess neurological differences between groups that may warrant further investigation was motor recovery over 1 year using the American Spinal Cord Injury Association examination. Recruitment and analyses were stratified by injury severity and injury location a priori given the expected influence of these on the sensitivity of the motor exam. Minocycline administered at higher than previously reported human doses produced steady-state concentrations of 12.7 µg/ml (95% confidence interval 11.6-13.8) in serum and 2.3 µg/ml (95% confidence interval 2.1-2.5) in cerebrospinal fluid, mimicking efficacious serum levels measured in animal studies. Transient elevation of serum liver enzymes in one patient was the only adverse event likely related to the study drug. Overall, patients treated with minocycline experienced six points greater motor recovery than those receiving placebo (95% confidence interval -3 to 14; P = 0.20, n = 44). No difference in recovery was observed for thoracic spinal cord injury (n = 16). A difference of 14 motor points that approached significance was observed in patients with cervical injury (95% confidence interval 0-28; P = 0.05, n = 25). Patients with cervical motor-incomplete injury may have experienced a larger difference (results not statistically significant, n = 9). Functional outcomes exhibited differences that lacked statistical significance but that may be suggestive of improvement in patients receiving the study drug. The minocycline regimen established in this study proved feasible, safe and was associated with a tendency towards improvement across several outcome measures. Although this study does not establish the efficacy of minocycline in spinal cord injury the findings are encouraging and warrant further investigation in a multi-centre phase III trial. ClinicalTrials.gov number NCT00559494.

Altered patterns of reflex excitability, balance, and locomotion following spinal cord injury and locomotor training.

Spasticity is an important problem that complicates daily living in many individuals with spinal cord injury (SCI). While previous studies in human and animals revealed significant improvements in locomotor ability with treadmill locomotor training, it is not known to what extent locomotor training influences spasticity. In addition, it would be of considerable practical interest to know how the more ergonomically feasible cycle training compares with treadmill training as therapy to manage SCI-induced spasticity and to improve locomotor function. Thus the main objective of our present studies was to evaluate the influence of different types of locomotor training on measures of limb spasticity, gait, and reflex components that contribute to locomotion. For these studies, 30 animals received midthoracic SCI using the standard Multicenter Animal Spinal cord Injury Studies (MASCIS) protocol (10 g 2.5 cm weight drop). They were divided randomly into three equal groups: control (contused untrained), contused treadmill trained, and contused cycle trained. Treadmill and cycle training were started on post-injury day 8. Velocity-dependent ankle torque was tested across a wide range of velocities (612–49°/s) to permit quantitation of tonic (low velocity) and dynamic (high velocity) contributions to lower limb spasticity. By post-injury weeks 4 and 6, the untrained group revealed significant velocity-dependent ankle extensor spasticity, compared to pre-surgical control values. At these post-injury time points, spasticity was not observed in either of the two training groups. Instead, a significantly milder form of velocity-dependent spasticity was detected at postcontusion weeks 8–12 in both treadmill and bicycle training groups at the four fastest ankle rotation velocities (350–612°/s). Locomotor training using treadmill or bicycle also produced significant increase in the rate of recovery of limb placement measures (limb axis, base of support, and open field locomotor ability) and reflex rate-depression, a quantitative assessment of neurophysiological processes that regulate segmental reflex excitability, compared with those of untrained injured controls. Light microscopic qualitative studies of spared tissue revealed better preservation of myelin, axons, and collagen morphology in both locomotor trained animals. Both locomotor trained groups revealed decreased lesion volume (rostro-caudal extension) and more spared tissue at the lesion site. These improvements were accompanied by marked upregulation of BDNF, GABA/GABAb, and monoamines (e.g., norepinephrine and serotonin) which might account for these improved functions. These data are the first to indicate that the therapeutic efficacy of ergonomically practical cycle training is equal to that of the more labor-intensive treadmill training in reducing spasticity and improving locomotion following SCI in an animal model.