Traumatic injuries to the spinal cord (SCI) and peripheral nerve (PNI) affect several thousand people worldwide every year. At present, there is no effective treatment for SCI and despite continuous improvements in microsurgical reconstructive techniques for PNI, many patients are still left with permanent, devastating neurological dysfunction. This thesis investigates the effects of mesenchymal stem cells (MSC) derived from adipose (ASC) and dental (DSC) tissue and chitosan/microRNA-124 polyplex particles on regeneration after spinal cord and peripheral nerve injury in adult rats. Dental stem cells were obtained from apical papilla, dental pulp, and periodontal ligament. ASC and DSC expressed MSC surface markers (CD73, CD90, CD105 and CD146) and various neurotrophic molecules including BDNF, GDNF, NGF, VEGF-A and angiopoietin-1. Growth factor stimulation of the stem cells resulted in increased secretion of these proteins. Both ASC and DSC supported in vitro neurite outgrowth and in contrast to Schwann cells, ASC did not induce activation of astrocytes. Stimulated ASC also showed an enhanced ability to induce capillary-like tube formation in an in vitro angiogenesis assay. In a peripheral nerve injury model, ASC and DSC were seeded into a fibrin conduit, which was used to bridge a 10 mm rat sciatic nerve gap. After 2 weeks, both ASC and DSC promoted axonal regeneration in the conduit and reduced caspase-3 expression in the dorsal root ganglion (DRG). ASC also enhanced GAP-43 and ATF-3 expression in the spinal cord, reduced c-jun expression in the DRG and increased the vascularity of the implant. After transplantation into injured C3-C4 cervical spinal cord, ASC continued to express neurotrophic factors and laminin and stimulated extensive ingrowth of 5HT-positive raphaespinal axons into the trauma zone. In addition, ASC induced sprouting of raphaespinal terminals in C2 contralateral ventral horn and C6 ventral horn on both sides. Transplanted cells also changed the structure and the density of the astroglial scar. Although the transplanted cells had no effect on the density of capillaries around the lesion site, the reactivity of OX42-positive microglial cells was markedly reduced. However, ASC did not enhance recovery of forelimb function. In order to reduce activation of microglia/macrophages and the secondary tissue damage after SCI, the role of microRNA-124 was investigated. In vitro transfection of chitosan/microRNA-124 polyplex particles into rat microglia resulted in the reduction of reactive oxygen species and TNF-α levels and lowered expression of MHC-II. Upon microinjection into uninjured rat spinal cords, particles formed with Cy3-labeled control sequence RNA, were specifically internalized by OX42 positive macrophages and microglia. Alternatively, particles injected in the peritoneum were transported by macrophages to the site of spinal cord injury. Microinjections of chitosan/microRNA-124 particles significantly reduced the number of ED-1 positive macrophages after SCI. In summary, these results show that human MSC produce functional neurotrophic and angiogenic factors, creating a more desirable microenvironment for neural regeneration after spinal cord and peripheral nerve injury. The data also suggests that chitosan/microRNA-124 particles could be potential treatment technique to reduce neuroinflammation.

Bakgrund Idag används olika medicinsktekniska produkter för helkroppsimmobilisering vid misstanke om spinal skada prehospitalt. Valet av immobiliseringsutrustning som skall användas görs av ansvarig sjuksköterska på plats. Den vanligaste utrustningen är ryggbräda, vaccummadrass, Kendric Extration Device (KED-väst) och nackkrage. Då utbildningen Pre Hospital Trauma Life Support (PHTLS) varit en del av specialistutbildningen för ambulanssjuksköterskor under en längre period har det funnits lite anledning till att ifrågasätta helkroppsimmobilisering och dess utrustning. All immobiliseringsutrustning klassificeras under klass 1 av Läkemedelsverket, vilket kan innebära att få vetenskapliga studier har gjorts för att utvärdera immobilieringsutrustning. Patientsäkerhetslagen ska efterlevas vad gäller vetenskaplig och beprövad metod i omvårdnaden av patienter med misstänkt spinal skada. Syfte Att genom en litteraturgranskning undersöka rekommendationer och eventuella risker med helkroppsimmobilisering vid misstanke av spinal skada prehospitalt utifrån patientsäkerhet och ett historiskt perspektiv. Metod Studien genomförs som litteraturgranskning med artikelgranskning och ett retrospektivt, deskriptivt perspektiv. Resultat Forskning rekommenderade helkroppsimmobilisering på traumapatienter där det fanns misstanke om spinal skada. Behandlingsriklinjerna följde nuvarande forskningsrekommendationer. Forskningskvaliten hade blivit bättre över tid och resultaten hade ökat i trovärdighet. Flera forskare påvisade risker med helkroppsimmobilisering. Riskerna var relaterade till dagens immobiliseringsutrustning. Ett antal alternativ till dagens immobiliseringsutrusning fanns beskrivna, men användes inte och var inte omnämnda i behandlingsriktlinjer. Slutsats Forskning rekommenderar helkroppsimmobilisering på traumapatienter där det finns misstanke om spinal skada. Immobiliseringutrustning har bevisligen flera risker för patienter, vilka orsakar allvarliga konsekvenser för hälsan. En lågt sittande ryggskada kanske inte behöver helkroppsimmobiliseras. Det kanske räcker med halvkroppsimmobilisering för vissa patienter. / Background Today, there are various medical devices for full body immobilisation in cases of suspected spinal injury prehospital. The choice of immobilisation devices to be used is the charge nurse’s on site. The most common equipment is back board, vaccummadrass, Kendric extraction Device (KED - West) and cervical collar. Because When training Pre Hospital Trauma Life Support (PHTLS) has been part of the specialist training of ambulance nurses over a longer period , there has been little reason to question full body immobilisation and its equipment . All immobilisation devices classified under Class 1 of the Medical Product Agency (MPA) , which may mean that few scientific studies have been done to evaluate immobilisation devices. The Patient Safety Act must be complied with in terms of scientific and proven method in the care of patients with suspected spinal injury. Aim Through a literature review examining recommendations and possible risks of full body immobilisation on suspicion of spinal injury pre-hospital based on patient safety and a historical perspective. Method The study is conducted as a literature review with the article review and a retrospective, descriptive perspective. Result Research recommended full body immobilisation on trauma patients where there was suspicion of spinal injury. Treatment guidelines followed the current research recommendations. The research quality had improved over time and results in increased credibility. Several researchers demonstrated the risks of full body immobilisation. The risks were related to today's immobilisation devices. A number of alternatives to current immobilisation devices was described, but was not used and was not mentioned in the treatment guidelines. Conclusion Research recommends full body immobilisation of trauma patients where there is a suspected spinal injury. Immobilisation devices has proven more risks for patients , which cause serious health consequences. A low sitting back injury may not need full body immobilisation. It might suffice with a half body immobilisation for some patients.

Peroxynitrite (PN, ONOO-), formed by nitric oxide radical (•NO) and superoxide radical (O2•-), plays an important role in post-traumatic oxidative damage. In the early work, we determined the temporal characteristics of PN-derived oxidative damage in a rat spinal cord injury (SCI) model. Our results showed 3-nitrotyrosine (3-NT), a specific marker for PN, rapidly accumulated at early time points (1 hr, 3 hrs), after when it plateaued and the high level was sustained to 1 week post injury. The co-localization of 3-NT and lipid peroxidation derived-4-HNE observed in immunohistochemistry indicates PN is involved in lipid peroxidative as well as protein nitrative damage. PN-oxidative damage exacerbates intracellular Ca2+ overload, which activates Ca2+ dependent calpain-mediated cytoskeletal protein (α-spectrin) degradation. The 145 kD fragments of α-spectrin (SBDP 145), which are specifically generated by calpain, increased dramatically as early as 1 hr after injury although the peak increase did not occur until 72 hrs post injury. The high level waned back toward sham level at one week post injury. We then carried out experiments to evaluate the beneficial effects of tempol, a scavenger of PN-derived radicals, following SCI. Three pathological events including PN-induced oxidative damage, mitochondrial dysfunction and cytoskeletal degradation were investigated. Immunoblotting and immunohistochemical studies indicated PN-mediated oxidative damage including protein nitration, protein oxidation and lipid peroxidation, were all reduced by a single dose of tempol (300mg/kg, i.p) after SCI. Spinal cord (SC) mitochondrial dysfunction in terms of the respiratory control ratio (RCR) significantly improved by both 150 mg/kg and 300 mg/kg tempol treatments. Moreover, calpain-mediated proteolysis was significantly decreased by tempol, with greater effects on calpain-specific SBDP 145 observed. Direct PN-scavenging effect of tempol was confirmed in vitro. Exposure of healthy SC mitochondria to SIN-1, a PN donor in vitro, impaired mitochondrial respiration in a dose-dependent manner. Tempol was able to protect mitochondria against SIN-1-induced damage by improving mitochondrial function and decreasing mitochondrial 3-NT formation. These findings strongly support the concept that PN is a crucial player in the secondary damage following SCI. And tempol, by scavenging PN-induced free radicals, provides a promising pharmocotherapeutic strategy for treating acute SCI.

In the hours and days following acute CNS injury, a secondary wave of events is initiated that exacerbate spinal tissue damage and neuronal cell death. A potential mechanism driving these secondary events is opening of the mitochondrial permeability transition pore (mPTP) and subsequent release of several cell death proteins. Previous studies have shown that inhibition of cyclophilin D(CypD), the key regulating component in mPTP opening, was protective against insults that induce necrotic cell death. We therefore hypothesized that CypD-null mice would show improved functional and pathological outcomes following spinal cord injury (SCI) and traumatic brain injury (TBI). Moderate and severe spinal contusion was produced in wild-type (WT) and CypD-null mice at the T-10 level using the Infinite Horizon impactor. Changes in locomotor function were evaluated using the Basso Mouse Scale (BMS) at 3 days post-injury followed by weekly testing for 4 weeks. Histological assessment of tissue sparing and lesion volume was performed 4 weeks post SCI. Calpain activity, measured by calpain-mediated spectrin degradation, was assessed in moderate injury only by western blot 24 hours post SCI. Results showed that following moderate SCI, CypD-null mice had no significant improvement in locomotor recovery or tissue sparing compared to wild-type mice. Following severe SCI, CypD-null mice showed significantly lower locomotor recovery and decreased tissue sparing compared to WT mice. Calpain-mediated spectrin degradation was not significantly reduced in CypD-null mice compared to WT mice 24h post moderate SCI. The lack of protective effects in CypD-null mice suggests that more dominant mechanisms are involved in the pathology of SCI. In addition, CypD may have a pro survival role that is dependent on the severity of the spinal cord injury.

Knowledge mobilization has been described as putting research in the hands of research users. Few studies have examined knowledge mobilization within community-based organizations (CBOs). To address this research gap, this dissertation examines knowledge mobilization within a CBO that supports people with spinal cord injury (SCI). Research suggests that communication channels, or the means by which a message is delivered, may affect knowledge mobilization. This dissertation presents four manuscripts examining how communication channels were used by the CBO to disseminate physical activity guidelines and intervention strategies to people with SCI. Manuscript 1 investigated reach and effectiveness of an event-based knowledge mobilization initiative delivered by the CBO using interpersonal communication channels to disseminate the guidelines to people with SCI. Results indicated that the event-based knowledge mobilization initiative was effective for initial dissemination of the guidelines. However, further long-term efforts are needed. Manuscript 2 describes how the event-based knowledge mobilization initiative was implemented by the CBO. Recognizing that support personnel are important messengers, the reach and effectiveness of the initiative for persuading support personnel to disseminate the guidelines was examined. Results indicated that an event-based knowledge mobilization initiative can be implemented by a CBO and may be an effective strategy for CBOs to disseminate information to support personnel. Manuscript 3 further examined the role of communication channels in the process of knowledge mobilization within the CBO using network analysis. Results indicated that CBO staff and volunteers’ integration within interpersonal communication channels was associated with greater knowledge of evidence-based physical activity resources and engagement in physical activity promotion behaviours. Manuscript 4 examined the feasibility of training peers with SCI to learn an evidence-based approach to physical activity promotion called Brief Action Planning. Findings indicated that Brief Action Planning is a tool that can be feasibly learned and potentially used by peers to promote physical activity to their mentees through interpersonal communication channels. Overall this dissertation contributes to a small but emerging body of literature examining knowledge mobilization in CBOs. Findings indicate that through a community-university multidisciplinary partnership and appropriate communication channels, a CBO can effectively and feasibly disseminate evidence-based physical activity information. / Thesis (Ph.D, Kinesiology & Health Studies) -- Queen's University, 2013-08-28 17:27:32.841

Research has begun to explore the presence and role of health-related injustice perceptions in samples of individuals who experience chronic pain associated with traumatic injury. Existing studies indicate that higher level of injustice perception is associated with poorer physical and psychosocial outcomes. However, to date, few clinical populations have been addressed. The aim of the current study was to explore injustice perceptions in a sample of individuals who have sustained a spinal cord injury (SCI), as research suggests that such individuals are likely to experience cognitive elements characteristic of injustice perception (e.g., perceptions of irreparable loss, blame, and unfairness). The study explored the relationship between participants’ level of perceived injustice and several variables associated with outcomes following SCI (depression, pain, and disability) at initial admission to a rehabilitation unit and at three months following discharge. The Injustice Experience Questionnaire was used to measure injustice perceptions. IEQ was found to significantly contribute to depression and anger at baseline. IEQ significantly contributed to depression, present pain intensity, and anger at follow-up. The implication of these preliminary findings may be beneficial for development of future interventions, as many individuals in the United States experience the lifelong physical and psychological consequences of SCI at a high personal and public cost.

Traumatic injuries to the spinal cord and brachial plexus induce a significant inflammatory response in the nervous tissue with progressive degeneration of neurons and glial cells, and cause considerable physical and mental suffering in affected patients. This thesis investigates the effects of the antioxidants N-acetyl-cysteine (NAC) and acetyl-L- carnitine (ALC) on the survival of motoneurons in the brainstem and spinal cord, the expression of pro-apoptotic and pro-inflammatory cell markers, axonal sprouting and glial cell reactions after spinal hemisection in adult rats. In addition, a novel MRI protocol has been developed to analyse the extent of neuronal degeneration in the spinal cord. Rubrospinal neurons and tibial motoneurons were pre-labelled with the fluorescent tracer Fast Blue one week before cervical C3 or lumbar L5 spinal cord hemisection. The intrathecal treatment with the antioxidants NAC (2.4mg/day) or ALC (0.9 mg/day) was initiated immediately after injury using Alzet2002 osmotic mini pumps. Spinal cord injury increased the expression of apoptotic cell markers BAX and caspase 3, induced significant degeneration of rubrospinal neurons and spinal motoneurons with associated decrease in immunoreactivity for microtubule-associated protein-2 (MAP2) in dendritic branches, synaptophysin in presynaptic boutons and neurofilaments in nerve fibers. Immunostaining for the astroglial marker glial fibrillary acidic protein and microglial markers OX42 and ED1 was markedly increased. Treatment with NAC and ALC attenuated levels of BAX, caspase 3, OX42 and ED1 expression after 2 weeks postoperatively. After 4-8 weeks of continuous intratheca ltreatment, NAC and ALC rescued approximately half of the rubrospinal neurons and spinal motoneurons destined to die, promoted axonal sprouting, restored the density of MAP2 and synaptophysin immunoreactivity and reduced the microglial reaction. However, antioxidant therapy did not affect the reactive astrocytes in the trauma zone. The inflammation modulating properties of ALC were also studied using cultures of human microglial cells. ALC increased the microglial production of interleukin IL-6 and BDNF, thereby possibly mediating the anti-inflammatory and pro-regenerative effects shown in vivo. To study degeneration in the spinal cord following pre-ganglionic and post-ganglionic brachial plexus injuries, adult rat models of ventral root avulsion and peripheral nerve injury were used. A novel MRI protocol was employed and the images were compared to morphological changes found in histological preparations. Ventral root avulsion caused degeneration of dendritic branches and axonal terminals in the spinal cord, followed by significant shrinkage of the ventral horn. Extensive astroglial and microglial reactions were detected in the histological preparations. Peripheral nerve injury reduced the density of dendritic branches but did not cause shrinkage of the ventral horn. Quantitative analysis of MRI images demonstrated changes in the ventral horn following ventral root avulsion only, thus validating the developed MRI technique as a possible tool for the differentiation of pre-ganglionic and post-ganglionic nerve injuries.

Peripheral nerve stimulation (PNS) is commonly used to promote use-dependent cortical plasticity for rehabilitation of motor function in spinal cord injury. Pairing transcranial magnetic stimulation (TMS) with PNS has been shown to increase motor evoked potentials most when the two stimuli are timed to arrive in the cortex simultaneously. This suggests that a mechanism of timing-dependent plasticity (TDP) may be a more effective method of promoting motor rehabilitation. The following thesis is the result of applying a brain-computer interface to apply PNS in closed-loop simultaneously to movement intention onset as measured by EEG of the sensorimotor cortex to test whether TDP can be induced in incomplete spinal cord injured individuals with upper limb motor impairment. 4 motor incomplete SCI subjects have completed 12 sessions of closed-loop PNS delivered over 4-6 weeks. Benefit was observed for every subject although not consistently across metrics. 3 out of 4 subjects exhibited increased maximum voluntary contraction force (MVCF) between first and last interventions for one or both hands. TMS-measured motor map volume increased for both hemispheres in one subject, and TMS center of gravity shifted in 3 subjects consistent with studies in which motor function improved or was restored. These observations suggest that rehabilitation using similar designs for responsive stimulation could improve motor impairment in SCI.

Traumatic spinal cord injury (SCI) is a physically debilitating, emotionally devastating, financially costly, and life-changing condition that afflicts more than 1,000,000 people in the United States alone. Owing to the characteristic neuropathology and low regenerative capacity of the central nervous system, many victims of SCI are left permanently paralyzed. Though the tissue damage caused by the initial insult almost certainly cannot be reversed, intensive research in recent years to elucidate the cellular and molecular events that follows has provided new grounds for optimism. Accordingly, in this dissertation, we present a number of potential treatment strategies aimed to address some of these pathological sequelae seen post-SCI so as to facilitate the regeneration of axons and the recovery of physiological functions. After the initial traumatic insult, a prominent and lasting injury-induced proliferative response occur and results in the development of a gliotic scar that isolates the lesion from the surrounding viable tissue. Although this process aids to prevent the spread of uncontrolled tissue damage, the scar nevertheless acts as a physical barrier to axonal regeneration. Furthermore, cells within the scar are a major source of axon growth-inhibitory molecules such as chondroitin sulfate proteoglycans (CSPG) and thus the scar acts concomitantly as a biochemical barrier. Concurrent to all this, inflammatory cells infiltrate the lesion and promote cell death through immunologic activation. Neuronal survival is also threatened from the lack of neurotrophic support caused by axonal severance. Finally, the pathology culminates in the formation of a fluid-filled cyst, which represents a gap that further hinders axonal regrowth. Since regeneration cannot physically occur in the presence of a cavity, we, by employing electrospinning techniques, generated a biocompatible matrix implant that can bridge and direct axonal elongation across the fluid-filled cyst. Given the complex array and scope of pathological sequelae post-SCI, it is generally recognized that a multifaceted approach is required to successfully treat SCI. In view of this, we presented novel approaches by which successful tried and true therapeutic strategies are combined to generate an enhanced matrix. An enzyme as well as a growth factor was incorporated into our matrix implants in order to respectively neutralize CSPGs and provide neurotrophic support. Using in vitro assays, we were able to demonstrate excellent protein bioactivity after incorporation. In vivo experimentation of these enhanced matrices is now ongoing. To address the injury-induced proliferative response, which represents an on-ramp off-ramp obstacle that prevents axonal regeneration onto our matrix implant, we showed how X-irradiation can be utilized to moderate this response by killing dividing cells so as to facilitate a more efficient penetration of regrowing axons into and beyond the gliotic scar. Finally, we demonstrate how a novel pharmacologic agent FTY720 can be used to attenuate the inflammatory response by preventing lymphocytic egress from lymphoid tissues. Collectively, these ideas and experimental results represent novel therapeutic strategies that can be combined in order to bring about meaningful functional recovery after SCI.

Spinal cord injury (SCI) has presented itself as a multifaceted pathology that is largely inhibitory to regeneration, and therefore to functional recovery, even though spinal cord neurons have been found to be innately regenerative. Thus, having identified the key players in the inhibition of this innate regeneration, SCI researchers have focused on two major types of approaches: (1) blocking inhibitory cues and (2) promoting innate regeneration. Schwann cells (SCs) have long been shown to promote and enhance functional recovery after SCI through providing supplemental myelination and trophic and tropic factors to regenerating axons, though singular approaches rarely address the complex SCI pathology. Guidance channels and scaffolds have been shown to provide physical support and directional cues to regeneration axons. Therefore, a combinatorial approach in which SCs migrate into and throughout a guidance scaffold would be an ideal research focus for treating SCI. However, cell migration into guidance scaffolds has been shown to be problematic. This study attempts to assess and improve two- and three-dimensional SC migration on electrospun scaffolds. Additionally, we evaluate the ability of SCs, seeded on Matrigel-coated electrospun scaffolds, to improve functional recovery in rats with completely transected spinal cords.