Cervical Spinal Cord Contusion Injury Research Articles

PTEN inhibition promotes robust growth of bulbospinal respiratory axons and partial recovery of diaphragm function in a chronic model of cervical contusion spinal cord injury

High spinal cord injury (SCI) leads to persistent and debilitating compromise in respiratory function. Cervical SCI not only causes the death of phrenic motor neurons (PhMNs) that innervate the diaphragm, but also damages descending respiratory pathways originating in the rostral ventral respiratory group (rVRG) located in the brainstem, resulting in denervation and consequent silencing of spared PhMNs located caudal to injury. It is imperative to determine whether interventions targeting rVRG axon growth and respiratory neural circuit reconnection are efficacious in chronic cervical contusion SCI, given that the vast majority of individuals are chronically-injured and most cases of SCI involve contusion-type damage to the cervical region. We therefore employed a rat model of chronic cervical hemicontusion to test therapeutic manipulations aimed at reconstructing damaged rVRG-PhMN-diaphragm circuitry to achieve recovery of respiratory function. At a chronic time point post-injury, we systemically administered: an antagonist peptide directed against phosphatase and tensin homolog (PTEN), a central inhibitor of neuron-intrinsic axon growth potential; an antagonist peptide directed against receptor-type protein tyrosine phosphatase sigma (PTPσ), another important negative regulator of axon growth capacity; or a combination of these two peptides. PTEN antagonist peptide (PAP4) promoted partial recovery of diaphragm motor activity out to nine months post-injury (though this effect depended on the anesthetic regimen used during recording), while PTPσ peptide did not impact diaphragm function after cervical SCI. Furthermore, PAP4 promoted robust growth of descending bulbospinal rVRG axons caudal to the injury within the denervated portion of the PhMN pool, while PTPσ peptide did not affect rVRG axon growth at this location that is critical to control of diaphragmatic respiratory function. In conclusion, we find that, when PTEN inhibition is targeted at a chronic time point following cervical contusion, our non-invasive PAP4 strategy can successfully promote significant regrowth of damaged respiratory neural circuitry and also partial recovery of diaphragm motor function.

Autophagy exerts a protective role in cervical spinal cord injury by microglia inhibition through the nuclear factor kappa-B pathway.

Spinal cord injury (SCI) is a serious trauma to the central nervous system. M1/M2 microglial polarization as well as the following neuroinflammatory response are crucial factors in SCI. Autophagy plays an important role in SCI, but its neuroprotective or neurodegenerative role remains controversial. Here, we majorly examined the properties of autophagy in SCI and uncovered the regulatory relationship between autophagy and microglial polarization in SCI. In our study, the Basso-Beattie-Bresnahan (BBB) score was declined in SCI. The cervical contusion SCI stimulated a sustaining neuropathic pain-linked phenotype characterized by thermal hyperalgesia as well as mechanical allodynia. It was revealed the structural damage to the spinal cord in SCI. Besides, the expression of microglia markers as well as inflammatory factor were promoted in SCI. Cervical contusion SCI induced autophagy inhibition and nuclear factor kappa-B (NF-κB) activation in mice. More importantly, enhanced autophagy induced by rapamycin suppressed the NF-κB pathway and alleviated cervical contusion SCI-induced neurological function damage in mice. Additionally, rapamycin promoted microglia M2 polarization and improved microglia-mediated inflammatory response. In conclusion, our study demonstrated that autophagy played a protective role in cervical SCI by promoting microglia polarization toward M2 through the NF-κB pathway. Our study may provide a novel sight for SCI treatment.

The biomechanical implications of neck position in cervical contusion animal models of SCI.

Large animal contusion models of spinal cord injury are an essential precursor to developing and evaluating treatment options for human spinal cord injury. Reducing variability in these experiments has been a recent focus as it increases the sensitivity with which treatment effects can be detected while simultaneously decreasing the number of animals required in a study. Here, we conducted a detailed review to explore if head and neck positioning in a cervical contusion model of spinal cord injury could be a factor impacting the biomechanics of a spinal cord injury, and thus, the resulting outcomes. By reviewing existing literature, we found evidence that animal head/neck positioning affects the exposed level of the spinal cord, morphology of the spinal cord, tissue mechanics and as a result the biomechanics of a cervical spinal cord injury. We posited that neck position could be a hidden factor contributing to variability. Our results indicate that neck positioning is an important factor in studying biomechanics, and that reporting these values can improve inter-study consistency and comparability and that further work needs to be done to standardize positioning for cervical spinal cord contusion injury models.

Metformin promotes Schwann cell remyelination, preserves neural tissue and improves functional recovery after spinal cord injury

Patients with a spinal cord injury (SCI) usually suffer lifelong disability as a result. Considering this, SCI treatment and pathology study are urgently needed. Metformin, a widely used hypoglycemic drug, has been indicated for its important role in central nervous system diseases. This study aimed to investigate the potential effect of metformin on remyelination after SCI. In the present study, we established a cervical contusion SCI model and metformin treatment was applied after SCI. Biomechanical parameters and behavioral assessment were used to evaluate the severity of injury and the improvement of functional recovery after SCI, respectively. The immunofluorescence and western blot were performed at the terminal time point. Our results showed that treating with metformin after SCI improved functional recovery by reducing the white matter loss and promoting Schwann cell remyelination, and the Nrg1/ErbB signaling pathway may be involved in promoting remyelination mediated by oligodendrocytes and Schwann cells. In addition, the area of spared tissues was significantly increased in the metformin group. However, metformin had no significant effects on the glial scar and inflammation after SCI. In summary, these findings indicated that the role of metformin in Schwann cell remyelination after SCI was probably related to the regulation of the Nrg1/ErbB pathway. It is, therefore, possible to suggest that metformin may be a potential therapy for SCI.

Transplantation of Human Induced Pluripotent Stem Cell-Derived Neural Progenitor Cells Promotes Forelimb Functional Recovery after Cervical Spinal Cord Injury.

Locomotor function after spinal cord injury (SCI) is critical for assessing recovery. Currently, available means to improve locomotor function include surgery, physical therapy rehabilitation and exoskeleton. Stem cell therapy with neural progenitor cells (NPCs) transplantation is a promising reparative strategy. Along this line, patient-specific induced pluripotent stem cells (iPSCs) are a remarkable autologous cell source, which offer many advantages including: great potential to generate isografts avoiding immunosuppression; the availability of a variety of somatic cells without ethical controversy related to embryo use; and vast differentiation. In this current work, to realize the therapeutic potential of iPSC-NPCs for the treatment of SCI, we transplanted purified iPSCs-derived NPCs into a cervical contusion SCI rat model. Our results showed that the iPSC-NPCs were able to survive and differentiate into both neurons and astrocytes and, importantly, improve forelimb locomotor function as assessed by the grooming task and horizontal ladder test. Purified iPSC-NPCs represent a promising cell type that could be further tested and developed into a clinically useful cell source for targeted cell therapy for cervical SCI patients.

Acute Magnetic Resonance Imaging Predictors of Chronic Motor Function and Tissue Sparing in Rat Cervical Spinal Cord Injury.

Predicting functional outcomes from spinal cord injury (SCI) at the acute setting is important for patient management. This work investigated the relationship of early magnetic resonance imaging (MRI) biomarkers in a rat model of cervical contusion SCI with long-term functional outcome and tissue sparing. Forty rats with contusion injury at C5 at either the spinal cord midline (bilateral) or over the lateral cord (unilateral) were examined using in vivo multi-modal quantitative MRI at 1 day post-injury. The extent of T2-weighted hyperintensity reflecting edema was greater in the bilateral model compared with the unilateral injury. Diffusion tensor imaging (DTI) exhibited microscopic damage in similar regions of the cord as reductions in fractional anisotropy (FA) and mean diffusivity (MD), but DTI parameter maps were also confounded by the presence of vasogenic edema that locally increased FA and MD. In comparison, filtered diffusion-weighted imaging (fDWI) more clearly delineated the location of acute axonal damage without effects of vasogenic edema. Pairwise correlation analysis revealed that 28-day motor functional outcomes were most strongly associated with the extent of edema (R = -0.69). Principal component analysis identified close associations of motor functional score with tissue sparing, the extent of edema, lesion area, and injury type (unilateral or bilateral). Among the diffusion MRI parameters, lesion areas measured with fDWI had the strongest association with functional outcome (R = -0.41). Voxelwise correlation analysis identified a locus of white matter damage associated with function in the dorsal white matter, although this was likely driven by variance across the two injury patterns (unilateral and bilateral injury). Nonetheless, correlation with motor function within the damaged region found in the voxelwise analysis outperformed morphological lesion area measurement as a predictor of chronic function. Collectively, this study characterized anatomical and diffusion MRI signatures of acute SCI at cervical spine and their association with chronic functional outcomes and histological results.

Rehabilitative training improves skilled forelimb motor function after cervical unilateral contusion spinal cord injury in rats

Animal models of cervical spinal cord injury (SCI) have frequently utilized partial transection injuries to evaluate plasticity promoting treatments such as rehabilitation training of skilled reaching and grasping tasks. Though highly useful for studying the effects of cutting specific spinal tracts that are important for skilled forelimb motor function, cervical partial-transection SCI-models underappreciate the extensive spread of most human SCIs, thus offering poor predictability for the clinical setting. Conversely, moderate cervical contusion SCI models targeting the spinal tracts important for skilled reaching and grasping can better replicate the increased size of most human SCIs and are often considered more clinically relevant. However, it is unknown whether animals with moderate cervical contusion SCIs that damage key spinal motor tracts can train in skilled reaching and grasping tasks. In this study, we quantify the impact of injury size and distribution on recovery in a skilled motor task called the single pellet reaching, grasping and retrieval (SPRGR) task in rats with cervical unilateral contusion injuries (UCs), and compare to rats with a partial transection SCIs (i.e., dorsolateral quadrant transection; DLQ). We found that UCs damage key tracts important for performing skilled motor tasks, similar to DLQs, but UCs also produce more extensive grey matter damage and more ventral white matter damage than DLQs. We also compared forelimb functionality at 1, 3, and 5 weeks of rehabilitative motor training between trained and untrained rats and found a more severe drop in SPRGR performance than in DLQ SCIs. Nevertheless, despite more severe injuries and initially low SPRGR performance, rehabilitative training for contusion animals resulted in significant improvements in SPRGR performance and proportionally more recovery than DLQ rats. Our findings show that rehabilitative motor training can facilitate considerable amounts of motor recovery despite extensive spinal cord damage, especially grey matter damage, thus supporting the use of contusion or compression SCI models and showing that ventral grey and white matter damage are not necessarily detrimental to recovery after training.

Diffusion-prepared fast spin echo for artifact-free spinal cord imaging.

Diffusion MRI provides unique contrast important for the detection and examination of pathophysiology after acute neurologic insults, including spinal cord injury. Diffusion weighted imaging of the rodent spinal cord has typically been evaluated with axial EPI readout. However, Diffusion weighted imaging is prone to motion artifacts, whereas EPI is prone to susceptibility artifacts. In the context of acute spinal cord injury, diffusion filtering has previously been shown to improve detection of injury by minimizing the confounding effects of edema. We propose a diffusion-preparation module combined with a rapid acquisition with relaxation enhancement readout to minimize artifacts for sagittal imaging. Sprague-Dawley rats with cervical contusion spinal cord injury were scanned at 9.4 Tesla. The sequence optimization included the evaluation of motion-compensated encoding diffusion gradients, gating strategy, and different spinal cord-specific diffusion-weighting schemes. A diffusion-prepared rapid acquisition with relaxation enhancement achieved high-quality images free from susceptibility artifacts with both second-order motion-compensated encoding and gating necessary for reduction of motion artifacts. Axial diffusivity obtained from the filtered diffusion-encoding scheme had greater lesion-to-healthy tissue contrast (52%) compared to the similar metric from DTI (25%). This work demonstrated the feasibility of high-quality diffusion sagittal imaging in the rodent cervical cord with diffusion-prepared relaxation enhancement. The sequence and results are expected to improve injury detection and evaluation in acute spinal cord injury.

Utility of somatosensory and motor-evoked potentials in reflecting gross and fine motor functions after unilateral cervical spinal cord contusion injury.

Fine motor skills are thought to rely on the integrity of ascending sensory pathways in the spinal dorsal column as well as descending motor pathways that have a neocortical origin. However, the neurophysiological processes underlying communication between the somatosensory and motor pathways that regulate fine motor skills during spontaneous recovery after spinal cord contusion injury remain unclear. Here, we established a rat model of cervical hemicontusive injury using C5 laminectomy followed by contusional displacement of 1.2 mm (mild injury) or 2.0 mm (severe injury) to the C5 spinal cord. Electrophysiological recordings were performed on the brachial muscles up to 12 weeks after injury to investigate the mechanisms by which spinal cord pathways participate in motor function. After spinal cord contusion injury, the amplitudes of somatosensory and motor-evoked potentials were reduced, and the latencies were increased. The forelimb open field locomotion test, grooming test, rearing test and Montoya staircase test revealed improvement in functions. With increasing time after injury, the amplitudes of somatosensory and motor-evoked potentials in rats with mild spinal cord injury increased gradually, and the latencies gradually shortened. In comparison, the recovery times of somatosensory and motor-evoked potential amplitudes and latencies were longer, and the recovery of motor function was delayed in rats with severe spinal cord injury. Correlation analysis revealed that somatosensory-evoked potential and motor-evoked potential parameters were correlated with gross and fine motor function in rats with mild spinal cord contusion injury. In contrast, only somatosensory-evoked potential amplitude was correlated with fine motor skills in rats with severe spinal cord injury. Our results show that changes in both somatosensory and motor-evoked potentials can reflect the changes in gross and fine motor functions after mild spinal cord contusion injury, and that the change in somatosensory-evoked potential amplitude can also reflect the change in fine motor function after severe spinal cord contusion injury. This study was approved by the Animal Ethics Committee of Nanfang Hospital, Southern Medical University, China (approval No. NFYY-2017-67) on June 11, 2017.

Correlating Tissue Mechanics and Spinal Cord Injury: Patient-Specific Finite Element Models of Unilateral Cervical Contusion Spinal Cord Injury in Non-Human Primates.

Non-human primate (NHP) models are the closest approximation of human spinal cord injury (SCI) available for pre-clinical trials. The NHP models, however, include broader morphological variability that can confound experimental outcomes. We developed subject-specific finite element (FE) models to quantify the relationship between impact mechanics and SCI, including the correlations between FE outcomes and tissue damage. Subject-specific models of cervical unilateral contusion SCI were generated from pre-injury MRIs of six NHPs. Stress and strain outcomes were compared with lesion histology using logit analysis. A parallel generic model was constructed to compare the outcomes of subject-specific and generic models. The FE outcomes were correlated more strongly with gray matter damage (0.29 < R2 < 0.76) than white matter (0.18 < R2 < 0.58). Maximum/minimum principal strain, Von-Mises and Tresca stresses showed the strongest correlations (0.31 < R2 < 0.76) with tissue damage in the gray matter while minimum principal strain, Von-Mises stress, and Tresca stress best predicted white matter damage (0.23 < R2 < 0.58). Tissue damage thresholds varied for each subject. The generic FE model captured the impact biomechanics in two of the four models; however, the correlations between FE outcomes and tissue damage were weaker than the subject-specific models (gray matter [0.25 < R2 < 0.69] and white matter [R2 < 0.06] except for one subject [0.26 < R2 < 0.48]). The FE mechanical outputs correlated with tissue damage in spinal cord white and gray matters, and the subject-specific models accurately mimicked the biomechanics of NHP cervical contusion impacts.

The spatiotemporal spread of cervical spinal cord contusion injury pathology revealed by 3D in-line phase contrast synchrotron X-ray microtomography

Extensive structural changes occur within the spinal cord following traumatic injury. Acute tissue debris and necrotic tissue are broken down, proliferating local glia and infiltrating leukocytes remodel tissue biochemical and biophysical properties, and a chronic cavity surrounded by a scar forms at the injury epicentre. Serial-section 2D histology has traditionally assessed these features in experimental models of spinal cord injury (SCI) to measure the extent of tissue pathology and evaluate efficacy of novel therapies. However, this 2D snapshot approach overlooks slice intervening features, with accurate representation of tissue compromised by mechanical processing artefacts. 3D imaging avoids these caveats and allows full exploration of the injured tissue volume to characterise whole tissue pathology. Amongst 3D imaging modalities, Synchrotron Radiation X-ray microtomography (SRμCT) is advantageous for its speed, ability to cover large tissue volumes at high resolution, and need for minimal sample processing. Here we demonstrate how extended lengths of formalin-fixed, paraffin-embedded (FFPE) rat spinal cord can be completely imaged by SRμCT with micron resolution. Label-free contrast derived from X-ray phase interactions with low-density soft tissues, reveals spinal cord white matter, gray matter, tissue damage and vasculature, with tissue still viable for targeted 2D-histology after 3D imaging. We used SRμCT to quantify tissue pathology after a midline, cervical level (C6), 225 kDyne contusion injury over acute-to-chronic (24 h to 5 weeks) post injury time points. Quantification revealed acute tissue swelling prior to chronic atrophy across the whole imaged region (spanning 2 spinal segments above and below injury), along with rostro-caudal asymmetries in white and gray matter volume loss. 3D volumes revealed satellite damage in tissue far removed from the epicentre, and extensive rostro-caudal spread of damage through the base of the dorsal columns at 24 h post injury. This damage overlapped regions of vasogenic oedema, confirmed with subsequent histology. Tissue damage at later time points in border regions was most prominent in the dorsal columns, where it overlapped sites of damaged venous vasculature. Elaborating rostro-caudal and spatiotemporal asymmetries in reduced traumatic injury models centred on these regions may inform future treatments that seek to limit the spread of tissue pathology to these ‘at-risk’ regions.

Bilateral cervical contusion spinal cord injury: A mouse model to evaluate sensorimotor function

Spinal cord injury is a severe condition, resulting in specific neurological symptoms depending on the level of damage. Approximately 60% of spinal cord injuries affect the cervical spinal cord, resulting in complete or incomplete tetraplegia and higher mortality rates than injuries of the thoracic or lumbar region. Although cervical spinal cord injuries frequently occur in humans, there are few clinically relevant models of cervical spinal cord injury. Animal models are critical for examining the cellular and molecular manifestations of human cervical spinal cord injury, which is not feasible in the clinical setting, and to develop therapeutic strategies. There is a limited number of studies using cervical, bilateral contusion SCI and providing a behavioral assessment of motor and sensory functions, which is partly due to the high mortality rate and severe impairment observed in severe cervical SCI models. The goal of this study was to develop a mouse model of cervical contusion injury with moderate severity, resulting in an apparent deficit in front and hindlimb function but still allowing for self-care of the animals. In particular, we aimed to characterize a mouse cervical injury model to be able to use genetic models and a wide range of viral techniques to carry out highly mechanistic studies into the cellular and molecular mechanisms of cervical spinal cord injury. After inducing a bilateral, cervical contusion injury at level C5, we followed the recovery of injured and sham-uninjured animals for eight weeks post-surgery. Hindlimb and forelimb motor functions were significantly impaired immediately after injury, and all mice demonstrated partial improvement over time that remained well below that of uninjured control mice. Mice also displayed a significant loss in their sensory function throughout the testing period. This loss of sensory and motor function manifested as a reduced ability to perform skilled motor tasks in all of the injured mice. Here, we describe a new mouse model of moderate bilateral cervical spinal cord injury that does not lead to mortality and provides a comprehensive assessment of histological and behavioral assessments. This model will be useful in enhancing our mechanistic understanding of cervical spinal cord injury and in the development of treatments targeted at promoting neuroprotection, neuroplasticity, and functional recovery after cervical SCI.

The effects of mouse strain and age on a model of unilateral cervical contusion spinal cord injury.

There are approximately 1.2 million people currently living with spinal cord injury (SCI), with a majority of cases at the cervical level and half involving incomplete injuries. Yet, as most preclinical research has been focused on bilateral thoracic models, there remains a disconnect between bench and bedside that limits translational success. Here, we profile a clinically relevant model of unilateral cervical contusion injury in the mouse (30kD with 0, 2, 5, or 10 second dwell time). We demonstrate sustained behavioral deficits in performance on grip strength, cylinder reaching, horizontal ladderbeam and CatWalk automated gait analysis tasks. Beyond highlighting reliable parameters for injury assessment, we also explored the effect of mouse strain and age on injury outcome, including evaluation of constitutively immunodeficient mice relevant for neurotransplantation and cellular therapy testing. Comparison of C57Bl/6 and immunodeficient Rag2gamma(c)-/- as well as Agouti SCIDxRag2Gamma(c)-/- hybrid mouse strains revealed fine differences in post-injury ipsilateral grip strength as well as total number of rearings on the cylinder task. Differences in post-SCI contralateral forepaw duty cycle and regularity index as measured by CatWalk gait analysis between the two immunodeficient strains were also observed. Further, assessment of young (3–4 months old) and aging (16–17 months old) Rag2gamma(c)-/- mice identified age-related pre-injury differences in strength and rearing that were largely masked following cervical contusion injury; observations that may help interpret previous results in aged rodents as well as human clinical trials. Collectively, the work provides useful insight for experimental design and analysis of future pre-clinical studies in a translational unilateral cervical contusion injury model.

5-HT7 Receptor Inhibition Transiently Improves Respiratory Function Following Daily Acute Intermittent Hypercapnic-Hypoxia in Rats With Chronic Midcervical Spinal Cord Contusion

Background. Intermittent hypoxia can induce respiratory neuroplasticity to enhance respiratory motor outputs following hypoxic treatment. This type of respiratory neuroplasticity is primarily mediated by the activation of Gq-protein-coupled 5-HT2 receptors and constrained by Gs-protein-coupled 5-HT7 receptors. Objective. The present study hypothesized that the blockade of 5-HT7 receptors can potentiate the effect of intermittent hypercapnic-hypoxia on respiratory function after cervical spinal cord contusion injury. Methods. The ventilatory behaviors of unanesthetized rats with midcervical spinal cord contusions were measured before, during, and after daily acute intermittent hypercapnic-hypoxia (10 episodes of 5 minutes of hypoxia [10% O2, 4% CO2, 86% N2] with 5 minutes of normoxia intervals for 5 days) at 8 weeks postinjury. On a daily basis, 5 minutes before intermittent hypercapnic-hypoxia, rats received either a 5-HT7 receptor antagonist (SB269970, 4 mg/kg, intraperitoneal) or a vehicle (dimethyl sulfoxide). Results. Treatment with intermittent hypercapnic-hypoxia induced a similar increase in tidal volume between rats that received SB269970 and those that received dimethyl sulfoxide within 60 minutes post-hypoxia on the first day. However, after 2 to 3 days of daily acute intermittent hypercapnic-hypoxia, the baseline tidal volumes of rats treated with SB269970 increased significantly. Conclusions. These results suggest that inhibiting the 5-HT7 receptor can transiently improve daily intermittent hypercapnic-hypoxia–induced tidal volume increase in midcervical spinal contused animals. Therefore, combining pharmacological treatment with rehabilitative intermittent hypercapnic-hypoxia training may be an effective strategy for synergistically enhancing respiratory neuroplasticity to improve respiratory function following chronic cervical spinal cord injury.

Fecal transplant prevents gut dysbiosis and anxiety-like behaviour after spinal cord injury in rats.

Secondary manifestations of spinal cord injury beyond motor and sensory dysfunction can negatively affect a person’s quality of life. Spinal cord injury is associated with an increased incidence of depression and anxiety; however, the mechanisms of this relationship are currently not well understood. Human and animal studies suggest that changes in the composition of the intestinal microbiota (dysbiosis) are associated with mood disorders. The objective of the current study is to establish a model of anxiety following a cervical contusion spinal cord injury in rats and to determine whether the microbiota play a role in the observed behavioural changes. We found that spinal cord injury caused dysbiosis and increased symptoms of anxiety-like behaviour. Treatment with a fecal transplant prevented both spinal cord injury-induced dysbiosis as well as the development of anxiety-like behaviour. These results indicate that an incomplete unilateral cervical spinal cord injury can cause affective disorders and intestinal dysbiosis, and that both can be prevented by treatment with fecal transplant therapy.

Application of Hepatocyte Growth Factor for Amyotrophic Lateral Sclerosis

Amyotrophic lateral sclerosis (ALS) is an adult onset neurodegenerative disorder characterized by the death of upper and lower motor neurons. About 10% of all ALS cases are familial, and we have identified SOD1 and FUS mutations as the most common causes in a consecutive series of 111 familial ALS pedigrees in Japan (Nishiyama A, 2017). From studies of the TDP43, FUS, and C9orf72 genes, perturbations of RNA processing can be highly adverse in motor neurons. Hepatocyte growth factor (HGF) is one of the most potent survival-promoting factors for motor neurons. We administered human recombinant HGF (hrHGF) by continuous intrathecal delivery to transgenic rats at onset of paralysis for 4 weeks. Intrathecal administration of hrHGF attenuated motor neuron degeneration and prolonged the duration of the disease by 63% (Ishigaki A, 2007). To translate this strategy to human treatment, we induced a contusive cervical spinal cord injury in the common marmoset, a primate, and then administered hrHGF intrathecally. We conducted a first-in-human phase I trial of intrathecal hrHGF in 15 Japanese patients with ALS (Warita H, 2019). Based on the results, we are conducting a phase II trial of intrathecal hrHGF for patients with ALS.

Behavioral and histopathological studies of cervical spinal cord contusion injury in rats caused by an adapted weight-drop device

Abstract Background Models of spinal cord injury (SCI) caused by weight-drop devices to cause contusion have been used extensively, and transient behavioral deficits after thoracic injury have been demonstrated. The severity of the injury caused by the device should be mild enough to allow recovery. Objective To determine whether our adapted weight-drop device with a small tip can effectively induce mild hemicontusion at the level of the fifth cervical vertebra. Methods We divided 15 adult male Sprague Dawley rats into groups of 5 for the following treatments: sham (SH, laminectomy only), mild (MSCI) or severe SCI (SSCI). Behavioral tests and histopathology were used before (day 1) and after the treatment on days 3, 7, 14, 21, 28, and 35 to assess the injury. Results Rats with SSCI showed a significant somatosensory deficit on days 3 and 7 compared with rats in the SH group, recovering by day 14. In a horizontal-ladder test of skilled locomotion, rats with SSCI showed a significant increase in error scores and percentage of total rungs used, and a decrease in the percentage of correct paw placement compared with rats in the SH group. There was greater recovery to normal paw placement by rats with MSCI than by rats with SSCI. These behavioral deficits were consistent with histopathology using hematoxylin and eosin counterstained Luxol fast blue, indicating the degree of injury and lesion area. Conclusions Mild hemicontusion caused by the adapted device can be used to evaluate SCI and provides a model with which to test the efficacy of translational therapies for SCI.

Injectable hydrogels of optimized acellular nerve for injection in the injured spinal cord

Objective. Spinal cord injury (SCI) affects a quarter million individuals in the United States, and there is currently no clinical treatment. Both fresh and acellular peripheral nerve grafts can induce spinal axon regeneration and support functional recovery in experimental injury models. Nonetheless, a scaffold that can be injected into a spinal contusion would be far less invasive to apply. We aimed to develop the first injectable acellular nerve graft for promoting repair after contusion SCI. Approach. We report a method to enzymatically solubilize optimized acellular (OA) nerve—a decellularized peripheral nerve graft developed in our laboratory and currently used clinically—to obtain an injectable solution that undergoes thermal gelation under physiological conditions. We quantified multiple physical and compositional properties of this novel material as well as tested its efficacy at acute and chronic time points following cervical contusion SCI. Main Results. This injectable optimized acellular (iOA) nerve graft retains native chemical cues such as collagens and glycosaminoglycans. By varying hydrogel concentration, the rheological properties and compressive modulus of iOA were similar to that previous reported for rat central nervous tissue. iOA solution was compatible with rat Schwann cells in culture, and hydrogel injection into a rat cervical contusion model significantly reduced the ratio of M1:M2 macrophages after one week, favoring regenerative phenotypes (p < 0.05). Furthermore, while iOA treatment did not affect locomotor or respiratory recovery over an eight week period, the percentage of axonal coverage increased at the distal tissue interface (p < 0.05), suggesting enhanced axonal extension within this region. Significance. Our data indicate that this novel injectable form of acellular nerve grafts is amenable for use after contusion SCI and may bolster a simultaneous therapy by acutely modulating the inflammatory milieu and supporting axonal growth.

Combining Constitutively Active Rheb Expression and Chondroitinase Promotes Functional Axonal Regeneration after Cervical Spinal Cord Injury

After spinal cord injury (SCI), severed axons in the adult mammalian CNS are unable to mount a robust regenerative response. In addition, the glial scar at the lesion site further restricts the regenerative potential of axons. We hypothesized that a combinatorial approach coincidentally targeting these obstacles would promote axonal regeneration. We combined (1) transplantation of a growth-permissive peripheral nerve graft (PNG) into an incomplete, cervical lesion cavity; (2) transduction of neurons rostral to the SCI site to express constitutively active Rheb (caRheb; a Ras homolog enriched in brain), a GTPase that directly activates the growth-promoting pathway mammalian target of rapamycin (mTOR) via AAV-caRheb injection; and (3) digestion of growth-inhibitory chondroitin sulfate proteoglycans within the glial scar at the distal PNG interface using the bacterial enzyme chondroitinase ABC (ChABC). We found that expressing caRheb in neurons post-SCI results in modestly yet significantly more axons regenerating across a ChABC-treated distal graft interface into caudal spinal cord than either treatment alone. Excitingly, we found that caRheb+ChABC treatment significantly potentiates the formation of synapses in the host spinal cord and improves the animals' ability to use the affected forelimb. Thus, this combination strategy enhances functional axonal regeneration following a cervical SCI.

Motor cortex and spinal cord neuromodulation promote corticospinal tract axonal outgrowth and motor recovery after cervical contusion spinal cord injury

Cervical injuries are the most common form of SCI. In this study, we used a neuromodulatory approach to promote skilled movement recovery and repair of the corticospinal tract (CST) after a moderately severe C4 midline contusion in adult rats. We used bilateral epidural intermittent theta burst (iTBS) electrical stimulation of motor cortex to promote CST axonal sprouting and cathodal trans-spinal direct current stimulation (tsDCS) to enhance spinal cord activation to motor cortex stimulation after injury. We used Finite Element Method (FEM) modeling to direct tsDCS to the cervical enlargement. Combined iTBS-tsDCS was delivered for 30min daily for 10days. We compared the effect of stimulation on performance in the horizontal ladder and the Irvine Beattie and Bresnahan forepaw manipulation tasks and CST axonal sprouting in injury-only and injury+stimulation animals. The contusion eliminated the dorsal CST in all animals. tsDCS significantly enhanced motor cortex evoked responses after C4 injury. Using this combined spinal-M1 neuromodulatory approach, we found significant recovery of skilled locomotion and forepaw manipulation skills compared with injury-only controls. The spared CST axons caudal to the lesion in both animal groups derived mostly from lateral CST axons that populated the contralateral intermediate zone. Stimulation enhanced injury-dependent CST axonal outgrowth below and above the level of the injury. This dual neuromodulatory approach produced partial recovery of skilled motor behaviors that normally require integration of posture, upper limb sensory information, and intent for performance. We propose that the motor systems use these new CST projections to control movements better after injury.