ADAM8 is selectively up‐regulated in endothelial cells and is associated with angiogenesis after spinal cord injury in adult mice

Endothelial cell (EC) loss and subsequent angiogenesis occur over the first week after spinal cord injury (SCI). To identify molecular mechanisms that could be targeted with intravenous (i.v.) treatments, we determined whether transmembrane “a disintegrin and metalloprotease” (ADAM) proteins are expressed in ECs of the injured spinal cord. ADAMs bind to integrins, which are important for EC survival and angiogenesis. Female adult C57Bl/6 mice with a spinal cord contusion had progressively more ADAM8 (CD156) immunostaining in blood vessels and individual ECs between 1 and 28 days following injury. Uninjured spinal cords had little ADAM8 staining. The increase in ADAM8 mRNA and protein was confirmed in spinal cord lysates, and ADAM8 mRNA was present in FACS‐enriched ECs. ADAM8 colocalized extensively and exclusively with the EC marker PECAM and also with i.v.‐injected lectins. Intravenous isolectin B4 (IB4) labels a subpopulation of blood vessels at and within the injury epicenter 3–7 days after injury, coincident with angiogenesis. Both ADAM8 and the proliferation marker Ki‐67 were present in IB4‐positive microvessels. ADAM8‐positive proliferating cells were seen at the leading end of IB4‐positive blood vessels. Angiogenesis was confirmed by BrdU incorporation, binding of i.v.‐injected nucleolin antibodies, and MT1‐MMP immunostaining in a subset of blood vessels. These data suggest that ADAM8 is vascular selective and plays a role in proliferation and/or migration of ECs during angiogenesis following SCI. J. Comp. Neurol. 512:243–255, 2009. © 2008 Wiley‐Liss, Inc.     

Neutralization of the chemokine CXCL10 enhances tissue sparing and angiogenesis following spinal cord injury

After spinal cord injury, there is a chemoattractant-mediated inflammatory response that is associated with secondary degeneration. The chemoattractant CXCL10 recruits CD4 Th1 cells via the CXCR3A receptor and inhibits growth and chemotaxis of endothelial cells via the CXCR3B receptor. To test the hypothesis that CXCL10 inhibits angiogenesis following spinal cord injury, we assayed the brainstems and spinal cords of spinal cord-injured mice treated with anti-CXCL10 antibodies for expression of angiogenesis-associated genes and quantified blood vessels within their spinal cords. Brainstem microarray analysis indicated eight angiogenesis-associated genes that had significantly higher expression levels in the treated mice than in the untreated mice. Ribonuclease protection assays of the spinal cords showed a significant increase in eight angiogenesis-associated genes in treated animals compared with untreated animals. Histological analysis of the spinal cords of treated and untreated mice showed a significant increase in the number of blood vessels in treated animals. We conclude that CXCL10 plays a critical role in vasculature remodeling following spinal cord injury and that angiogenesis is enhanced following anti-CXCL10 treatment of spinal cord injuries. Improved blood flow and oxygen supply to the injury site may contribute to the functional improvement associated with this treatment.     

Induction of endogenous neural precursors in mouse models of spinal cord injury and disease

Adult neural precursor cells (NPCs) in the mammalian central nervous system (CNS) have been demonstrated to be responsive to conditions of injury and disease. Here we investigated the response of NPCs in mouse models of spinal cord disease [motor neuron disease (MND)] with and without sciatic nerve axotomy, and spinal cord injury (SCI). We found that neither axotomy, nor MND alone brought about a response by Nestin-positive NPCs. However, the combination of the two resulted in mobilization of NPCs in the spinal cord. We also found that there was an increase in the number of NPCs following SCI which was further enhanced by systemic administration of the neuregulatory cytokine, leukaemia inhibitory factor (LIF). NPCs were demonstrated to differentiate into astrocytes in axotomized MND mice. However, significant differentiation into the various neural cell phenotypes was not demonstrated at 1 or 2 weeks following SCI. These data suggest that factors inherent to injury mechanisms are required for induction of an NPC response in the mammalian spinal cord.     

Hypoxia-specific VEGF-expressing neural stem cells in spinal cord injury model

We established three stable neural stem cell (NSC) lines to explore the possibility of using hypoxia-specific vascular endothelial growth factor (VEGF) expressing NSC lines (EpoSV-VEGF NSCs) to treat spinal cord injury. The application of EpoSV-VEGF NSCs into the injured spinal cord after clip compression injury not only showed therapeutic effects such as extended survival and angiogenesis, but also displayed its safety profile as it did not cause unwanted cell proliferation or angiogenesis in normal spinal cord tissue, as EpoSV-VEGF NSCs consistently showed hypoxia-specific VEGF expression patterns. This suggests that our EpoSV-VEGF NSCs are both safe and therapeutically efficacious for the treatment of spinal cord injury. Furthermore, this hypoxia-inducible gene expression system may represent a safe tool suitable for gene therapy.     

Endothelial progenitor cell-conditioned medium promotes angiogenesis and is neuroprotective after spinal cord injury

Endothelial progenitor cells secrete a variety of growth factors that inhibit inflammation, promote angiogenesis and exert neuroprotective effects. Therefore, in this study, we investigated whether endothelial progenitor cell-conditioned medium might have therapeutic effectiveness for the treatment of spinal cord injury using both in vitro and in vivo experiments. After primary culture of bone marrow-derived macrophages, lipopolysaccharide stimulation was used to classically activate macrophages to their proinflammatory phenotype. These cells were then treated with endothelial progenitor cell-conditioned medium or control medium. Polymerase chain reaction was used to determine mR NA expression levels of related inflammatory factors. Afterwards, primary cultures of rat spinal cord neuronal cells were prepared and treated with H_2O_2 and either endothelial progenitor cell-conditioned medium or control medium. Hoechst 33258 and propidium iodide staining were used to calculate the proportion of neurons undergoing apoptosis. Aortic ring assay was performed to assess the effect of endothelial progenitor cell-conditioned medium on angiogenesis. Compared with control medium, endothelial progenitor cell-conditioned medium mitigated the macrophage inflammatory response at the spinal cord injury site, suppressed apoptosis, and promoted angiogenesis. Next, we used a rat model of spinal cord injury to examine the effects of the endothelial progenitor cell-conditioned medium in vivo. The rats were randomly administered intraperitoneal injection of PBS, control medium or endothelial progenitor cell-conditioned medium, once a day, for 6 consecutive weeks. Immunohistochemistry was used to observe neuronal morphology. Terminal deoxynucleotidyl transferase-mediated d UTP nick-end labeling assay was performed to detect the proportion of apoptotic neurons in the gray matter. The Basso, Beattie and Bresnahan Locomotor Rating Scale was used to evaluate the recovery of motor function of the bilateral hind limbs after spinal cord injury. Compared with the other two groups, the number of axons was increased, cavities in the spinal cord were decreased, the proportion of apoptotic neurons in the gray matter was reduced, and the Basso, Beattie and Bresnahan score was higher in the endothelial progenitor cell-conditioned medium group. Taken together, the in vivo and in vitro results suggest that endothelial progenitor cell-conditioned medium suppresses inflammation, promotes angiogenesis, provides neuroprotection, and promotes functional recovery after spinal cord injury.     

Griffonia simplicifolia isolectin B4 identifies a specific subpopulation of angiogenic blood vessels following contusive spinal cord injury in the adult mouse

After traumatic spinal cord injury (SCI), disruption and plasticity of the microvasculature within injured spinal tissue contribute to the pathological cascades associated with the evolution of both primary and secondary injury. Conversely, preserved vascular function most likely results in tissue sparing and subsequent functional recovery. It has been difficult to identify subclasses of damaged or regenerating blood vessels at the cellular level. Here, adult mice received a single intravenous injection of the Griffonia simplicifolia isolectin B4 (IB4) at 1-28 days following a moderate thoracic (T9) contusion. Vascular binding of IB4 was maximally observed 7 days following injury, a time associated with multiple pathologic aspects of the intrinsic adaptive angiogenesis, with numbers of IB4 vascular profiles decreasing by 21 days postinjury. Quantitative assessment of IB4 binding shows that it occurs within the evolving lesion epicenter, with affected vessels expressing a temporally specific dysfunctional tight junctional phenotype as assessed by occludin, claudin-5, and ZO-1 immunoreactivities. Taken together, these results demonstrate that intravascular lectin delivery following SCI is a useful approach not only for observing the functional status of neovascular formation but also for definitively identifying specific subpopulations of reactive spinal microvascular elements.     

CD47 knockout mice exhibit improved recovery from spinal cord injury

Recent data have implicated thrombospondin-1 (TSP-1) signaling in the acute neuropathological events that occur in microvascular endothelial cells (ECs) following spinal cord injury (SCI) (Benton et al., 2008b). We hypothesized that deletion of TSP-1 or its receptor CD47 would reduce these pathological events following SCI. CD47 is expressed in a variety of tissues, including vascular ECs and neutrophils. CD47 binds to TSP-1 and inhibits angiogenesis. CD47 also binds to the signal regulatory protein (SIRP)α and facilitates neutrophil diapedesis across ECs to sites of injury. After contusive SCI, TSP-1(-/-) mice did not show functional improvement compared to wildtype (WT) mice. CD47(-/-) mice, however, exhibited functional locomotor improvements and greater white matter sparing. Whereas targeted deletion of either CD47 or TSP-1 improved acute epicenter vascularity in contused mice, only CD47 deletion reduced neutrophil diapedesis and increased microvascular perfusion. An ex vivo model of the CNS microvasculature revealed that CD47(-/-)-derived microvessels (MVs) prominently exhibit adherent WT or CD47(-/-) neutrophils on the endothelial lumen, whereas WT-derived MVs do not. This implicates a defect in diapedesis mediated by the loss of CD47 expression on ECs. In vitro transmigration assays confirmed the role of SIRPα in neutrophil diapedesis through EC monolayers. We conclude that CD47 deletion modestly, but significantly, improves functional recovery from SCI via an increase in vascular patency and a reduction of SIRPα-mediated neutrophil diapedesis, rather than the abrogation of TSP-1-mediated anti-angiogenic signaling.     

脊髓全横断大鼠椎管内睫状神经营养因子抗体封闭模型的建立

【摘要】　目的 建立一种特异性中和内源性CNTF表达的模型,用于研究CNTF在脊髓全横断损伤后的作用机制。方法 健康成年SD雌性大鼠45只,随机分为3组：假手术组15只、对照组15只、抗体封闭组15只。对照组和抗体封闭组大鼠行T11 脊髓全横断术和椎管内置管术,对照组大鼠经导管推注人工脑脊液20μl/只,抗体封闭组大鼠推注CNTF抗体（0.1g/ml）20μl/只。假手术组仅行T9椎板切除术。观察动物术后存活情况及24h时BBB评分,应用免疫组织化学和Western-blot技术检测术后24h、48h、72h 脊髓颈、胸、腰段CNTF的分布及表达变化。 结果　术后动物存活率100％,对照组和抗体封闭组BBB评分显著低于假手术组（Ρ<0.01）;对照组术后各时间点大量CNTF阳性细胞分布于灰质前角及白质前索、后索和外侧索;抗体封闭组T9、L2节段术后24h时未见CNTF阳性细胞,术后48h、72h出现阳性细胞,C5节段术后3个时间点均可见阳性细胞分布;Western-blot显示对照组脊髓各节段术后3个时间点CNTF表达量均较假手术组增加（Ρ<0.05）,抗体封闭组T10、L1脊髓术后24h时CNTF表达较对照组减少（Ρ<0.05）,而于48h、72h回升（Ρ<0.01）,C6节段术后各时点CNTF的变化无差异（Ρ>0.05）。 结论 该模型在24h内能够有效封闭内源性CNTF表达,为研究脊髓损伤后CNTF的作用和机制奠定了基础,并能推广应用于其它细胞因子的相关研究。 【关键词】　睫状神经营养因子(CNTF),抗体封闭,脊髓全横断,模型     

Characterization of the microvascular glycocalyx in normal and injured spinal cord in the rat

The glycocalyx of microvasculature in normal and injured spinal cord was characterized by using cationized ferritin to define anionic sites and the lectins concanavalin agglutinin (Con A) and Ricinus communis agglutinin I (RCA) to delineate carbohydrate moities. Binding of cationized ferritin was evaluated at the ultrastructural level in control animals and at 3 hours after spinal cord injury. Horseradish peroxidase (HRP) was administered intravenously before euthanasia. In control spinal cord, there was continuous even binding of cationized ferritin along the luminal front of microvasculature and no evidence of barrier permeability to HRP. After spinal cord injury, there was a reduction in binding of cationized ferritin in those regions of spinal cord that exhibited barrier breakdown to HRP. Lectin binding in the spinal cord was evaluated at 3 hours and 3 days postinjury. At the light microscopic level, there appeared to be increased binding of Con A and RCA in microvessels by 3 days postinjury as compared with the control spinal cord. At the ultrastructural level, a significant increase in RCA binding was noted along luminal fronts in the injured spinal cord. This increased binding coincided with a significant elaboration of the endothelial glycocalyx. These findings demonstrate that the charge, structure, and carbohydrate composition of the endothelial glycocalyx in microvessels in the spinal cord may be dramatically altered after spinal cord injury. Furthermore, there is an association between the loss of charge and disruption of the barrier, suggesting that anionic sites may contribute to maintenance of the blood-spinal cord barrier. © 1996 Wiley-Liss, Inc.     

Necessity for re-vascularization after spinal cord injury and the search for potential therapeutic options

Disruption of the blood-spinal cord barrier (BSCB) and microvascular changes leading to reduction of blood supply represent hallmarks of spinal cord secondary injury causing further deterioration of the traumatized patient. Injury to the blood vessels starts with prominent hemorrhage and generation of inflammation. Furthermore, spinal cord ischemia and extravasation of blood components contribute to edema formation resulting in death of neural cells. Endogenous attempts of re-vascularization have been observed although these newly formed vessels display morphological and functional abnormalities. The unfavorable regulation of angiogenic and counterregulatory anti-angiogenic factors during the complicated course of vessel remodeling after SCI is suspected to participate in the failure of re-vascularization and vessel stabilization. Repression of the expression of angiogenic factors such as vascular endothelial growth factor-A (VEGF-A), placental growth factor (PlGF), angiopoietin-1 (Ang1), and platelet-derived growth factor-BB (PDGF-BB) contributes to vessel regression. Therefore, therapeutic applications of angiogenic factors following SCI are promising strategies to restore blood flow in the lesion.     

Electrical fields and the promotion of endogenous angiogenesis in a rat spinal cord injury model

BACKGROUND:Previous studies have shown that direct current electrical fields affect development and growth of human microvascular endothelial cells,but the role of electrical fields on promoting angiogenesis in tissues following spinal cord injury remains poorly understood.OBJECTIVE:To determine the effects of electrical fields on angiogenesis and spinal cord repair following traumatic spinal cord injury in rats.DESIGN,TIME AND SETTING:A randomized,controlled,animal experiment was performed at the Chongqing Key Laboratory of Neurology,Affiliated Hospital of Chongqing Medical University,China from September 2007 to August 2008.MATERIALS:Hydrogen blood flow detector(Soochow University Medical Instrument,China),Power Lab System(AD Instruments,Colorado Springs,CO,USA)and mouse anti-vascular endothelial growth factor(VEGF)monoclonal antibody(Sigma-Aldrich,St.Louis,MO,USA)were used in this study.METHODS:A total of 60 healthy,adult,Sprague Dawley rats were equally and randomly assigned to sham-surgery,model,and electrical field groups.The Allen's weight-drop method was used to induce complete spinal cord injury in the model and electrical field groups.Rats in the electrical field group were implanted with silver needles and electrical fields(350 V/m)were applied following traumatic injury.MAIN OUTCOME MEASURES:Latency of somatosensory-evoked potential was detected and spinal cord blood flow was measured by hydrogen blood flow detector.Microvascular density was determined by histological analysis.VEGF expression in the spinal cord was observed by immunohistochemical staining.RESULTS:Recovery of spinal cord blood flow was significantly increased in the electrical field group(at 1,2,4,8,and 24 days after injury)compared with the model group(P < 0.05 or P <0.01).Latency of P1 waves in somatosensory-evoked potential of electrical field group(at 1,2,4,8,and 24 days after injury)was significantly shorter than the model group(P < 0.05 or P < 0.01).Microvascular density and VEGF expression were greater in the electrical field group compared with the model group at 24 days after injury(P< 0.01).CONCLUSION:Electrical fields(350 V/m)promoted angiogenesis within injured rat tissue following spinal cord injury and improved spinal cord function.Electrical fields could help to ameliorate spinal cord injury.The mechanisms of action could be related to increased VEGF expression.     

Elevated intraspinal pressure in traumatic spinal cord injury is a promising therapeutic target

The currently recommended management for acute traumatic spinal cord injury aims to reduce the incidence of secondary injury and promote functional recovery.Elevated intraspinal pressure(ISP)likely plays an important role in the processes involved in secondary spinal cord injury,and should not be overlooked.However,the factors and detailed time course contributing to elevated ISP and its impact on pathophysiology after traumatic spinal cord injury have not been reviewed in the literature.Here,we review the etiology and progression of elevated ISP,as well as potential therapeutic measures that target elevated ISP.Elevated ISP is a time-dependent process that is mainly caused by hemorrhage,edema,and blood-spinal cord barrier destruction and peaks at 3 days after traumatic spinal cord injury.Duraplasty and hypertonic saline may be promising treatments for reducing ISP within this time window.Other potential treatments such as decompression,spinal cord incision,hemostasis,and methylprednisolone treatment require further validation.     

Expression of angiogenic factors in rabbit spinal cord after transient ischaemia

It is known that angiogenic factors are induced in brain by ischaemia, and new vessel formation is correlated with better prognosis in patients of stroke. However, the role of angiogenesis and expression of angiogenic factors in spinal cord ischaemia is uncertain. We here investigated expression of three highly potent angiogenic peptides, i.e. basic fibroblast growth factor (bFGF), vascular endothelial growth factor (VEGF), and hepatocyte growth factor (HGF) in the rabbit spinal cord after transient ischaemia, by Western blot and immunohistochemical analysis. Western blot analysis revealed that bFGF was induced at 8 h after transient ischaemia and decreased thereafter. Immunoreactive VEGF was also induced at 8 h, and it disappeared thereafter. HGF was not detected in the spinal cord with sham-operation or ischaemic injury. By immunohistochemical analysis, bFGF was weakly expressed in only a few small interneurons in sham-operated spinal cords. However, it was induced to a marked degree in motor neurons and interneurons of the anterior horn at 8 h after reperfusion. It was also induced in small neurons of the posterior horn. The expression in the anterior horn decayed thereafter though it lasted until 7 d in the posterior horn. VEGF was not expressed in sham-operated spinal cords, but the expression was induced in large motor neurons and interneurons at 8 h with marked reduction at 1 d. In contrast, HGF was not expressed in the spinal cord with sham-operation or ischaemic injury. These factors are known to play pivotal roles in angiogenesis, regulation of blood flow, and protection of endothelial cells. Through induction of these angiogenic peptides, protection of vascular endothelial cells and improvement of regional blood flow might be occurring in the spinal cord after ischaemia.     

Nestin-positive cells in the spinal cord: a potential source of neural stem cells

Some literatures have reported neural precursor cells (NPCs) exist in spinal cord of adult mammal, however, the NPCs distribution feature in spinal cord of adult mice so far is not described in detail. In order to observe and compare the distribution feature of NPCs in various spinal cord regions of adult mice, to research a potential source of neural stem cells (NSCs), we obtained NPCs distribution feature by analyzing the distribution of the nestin-containing cells (NCCs) in spinal cord of adult nestin second-intron enhancer controlled LacZ reporter transgenic mice (pNes-Tg) with LacZ staining and positive cell quantification. The results showed that: NCCs were observed in various regions of spinal cord of adult mice, but amount of NCCs was different in distinct region, the rank order of NCCs amount in various spinal cord regions was dorsal horn region greater than central canal greater than the ventral and lateral horn. NCCs in dorsal horn region mainly distributed in substantia gelatinosa, NCCs in central canal mainly distributed in ependymal zone, on the contrary, NCCs in ventral, lateral horn, medullae, nucleus regions of spinal cord were comparatively less. The rank order of NCCs amount in various spinal cord segments was cervical segment greater than lumbar sacral segment greater than thoracic segment. There was no significantly difference between NCCs amount in the left and right sides, and within cervical 1–7, thoracic 1–12, lumbar 1–5, sacral segment of spinal cord in adult mice. These data collectively indicate that NPCs extensively distribute in various regions of spinal cord of adult mice, especially in substantia gelatinosa and ependymal zone. NPCs in cervical segment are abundant, NPCs in thoracic segment are the least while compared the different spinal cord segment, the NPCs in various regions of spinal cord of adult mice are a potential source of NSCs.     

Neural precursor cells can be delivered into the injured cervical spinal cord by intrathecal injection at the lumbar cord

Neural precursor cells (NPCs) are promising grafts for treatment of traumatic CNS injury and neurodegenerative disorders because of their potential to differentiate into neurons and glial cells. When designing clinical protocols for NPC transplantation, it is important to develop alternatives to direct parenchymal injection, particularly at the injury site. We reasoned that since it is minimally invasive, intrathecal delivery of NPCs at lumbar spinal cord (lumbar puncture) represents an important and clinically applicable strategy. We tested this proposition by examining whether NPCs can be delivered to the injured cervical spinal cord via lumbar puncture using a mixed population of neuronal-restricted precursors (NRPs) and glial-restricted precursors (GRPs). For reliable tracking, the NPCs were derived from the embryonic spinal cord of transgenic donor rats that express the marker gene, human placental alkaline phosphatase, under the control of the ubiquitous Rosa 26 promoter. We found that mixed NRP/GRP grafts can be efficiently delivered to a cervical hemisection injury site by intrathecal delivery at the lumbar cord. Similar to direct parenchymal injections, transplanted NRP/GRP cells survive at the injury cavity for at least 5 weeks post-engraftment, migrate into intact spinal cord along white matter tracts and differentiate into all three mature CNS cell types, neurons, astrocytes, and oligodendrocytes. Furthermore, very few graft-derived cells localize to areas outside the injury site, including intact spinal cord and brain. These results demonstrate the potential of delivering lineage-restricted NPCs using the minimally invasive lumbar puncture method for the treatment of spinal cord injury.     

Vasogenic edema in the injured spinal cord: A method of evaluating the extent of blood-brain barrier alteration to horseradish peroxidase

A quantitatable exogenous enzyme, horseradish peroxidase, was used as an intravascular marker to determine the extent of blood-brain barrier alteration subsequent to spinal cord compression injury in cats. Blood-brain barrier alterations to horseradish peroxidase were evaluated in the compressed region of the cord and in adjacent segments up to 9 cm at the following time intervals: during the first 6 hr, 3 days and 14 days after injury. The degree of blood-brain barrier dysfunction correlated with the severity of injury. Depending on the time interval following injury and the location of the spinal cord segment, vascular extravasation and/or abnormal endothelial uptake of horseradish peroxidase was two to six times greater in an injury sufficient to cause irreversible paraplegia, as compared to an injury resulting in a transitory paraplegia. In both types of injuries, vascular extravasation and/or abnormal endothelial uptake was significantly greater in the injured region as compared to distal segments of cord. In both injuries and at all time intervals evaluated, the degree of vascular extravasation and/or abnormal endothelial uptake exhibited and inverse relationship to the distance from the injured site. This method appears to be suitable for measuring degrees of blood-brain barrier dysfunction to horseradish peroxidase and offers a means of evaluating therapy designed to reduce vasogenic edema.     

Neurally induced umbilical cord blood cells modestly repair injured spinal cords

Umbilical cord blood (UCB) is known to have stem/progenitor cells. We earlier showed that novel progenitors could be isolated from cryopreserved human UCB with high efficiency. The multipotent progenitor cells were induced to differentiate into neural-lineage cells under the appropriate condition. In this study, we confirmed these neurally induced progenitor cells (NPCs), containing higher quantities of nerve growth factor, promoted functional recovery in rats with spinal cord injury (SCI). Sprague-Dawley rats with SCI achieved a modest improvement in locomotor rating scale until 10 weeks after transplantation of the NPCs. SCI rats treated with NPCs also showed somatosensory-evoked potentials were recovered, and grafted cells especially exhibited oligodendrocytic phenotype around the necrotic cavity. These findings suggest that UCB-NPCs might be a therapeutic resource to repair damaged spinal cords.     

Aquaporin-4 and spinal cord injury

Edema formation is a major problem following traumatic spinal cord injury (SCI) that acts to exacerbate secondary damage. Severity of edema correlates with reduced neurological outcome in human patients. To date, there are no effective treatments to directly resolve edema within the spinal cord. The aquaporin-4 (AQP4) water channel is found on membranes of astrocytic endfeet in direct contact with blood vessels, the glia limitans in contact with the cerebrospinal fluid and ependyma around the central canal. Being so locally expressed at the interface between fluid and tissue allow AQP4 channels to play an important role in the bidirectional regulation of water homeostasis under normal conditions and following trauma. With the need to better understand the pathophysiology underlying the devastating cellular events in SCI, animal models have become an integral part of exploration. Inevitably, several injury models have been developed (contusion, compression, transection) resulting in difficult interpretation between studies with conflicting results. This is true in the case of understanding the role of AQP4 in the progression and resolution of edema following SCI, whose role is still not completely understood and is highly dependent on the type of edema present (vasogenic vs cytotoxic). Here, we discuss regulation of AQP4 in varying injury models and the effects of potential therapeutic interventions on expression, edema formation and functional recovery. Better understanding of the precise role of AQP4 following a wide range of injuries will help to understand optimal treatment timing following human SCI for prime therapeutic benefit and enhanced neurological outcome.     

Blood-spinal cord barrier after spinal cord injury: relation to revascularization and wound healing

Spinal cord injury produces prominent disruption of the blood-spinal cord barrier. We have defined the blood-spinal cord barrier breakdown to the protein luciferase (61 kDa) in the acutely injured murine spinal cord and during revascularization. We show that newly formed and regenerating blood vessels that have abnormal permeability exhibit differential expression of the glucose-1 transporter (Glut-1), and that its expression is dependent on astrocytes. There was overt extravasation of luciferase within the first hour after injury, a period that coincided with marked tissue disruption within the epicenter of the lesion. Although there was a significant reduction in the number of blood vessels relative to controls by 24 hr after injury, abnormal barrier permeability remained significantly elevated. A second peak of abnormal barrier permeability at 3-7 days postinjury coincided with prominent revascularization of the epicenter. The barrier to luciferase was restored by 21 days postinjury and vascularity was similar to that of controls. During wound-healing process, the cord was reorganized into distinct domains. Between 14 and 21 days postinjury, each domain consisted primarily of nonneuronal cells, including macrophages. Astrocytes were limited characteristically to the perimeter of each domain. Only blood vessels affiliated closely with astrocytes in the perimeter expressed Glut-1, whereas blood vessels within each domain of the repairing cord did not express it. Together, these data demonstrate that both injured and regenerating vessels exhibit abnormal permeability and suggest that Glut-1 expression during revascularization is dependent on the presence of astrocytes.     

Numb rats walk - a behavioural and fMRI comparison of mild and moderate spinal cord injury

Assessment of sensory function serves as a sensitive measure for predicting the functional outcome following spinal cord injury in patients. However, little is known about loss and recovery of sensory function in rodent spinal cord injury models as most tests of sensory functions rely on behaviour and thus motor function. We used functional magnetic resonance imaging (fMRI) to investigate cortical and thalamic BOLD-signal changes in response to limb stimulation following mild or moderate thoracic spinal cord weight drop injury in Sprague-Dawley rats. While there was recovery of close to normal hindlimb motor function as determined by open field locomotor testing following both degrees of injury, recovery of hindlimb sensory function as determined by fMRI and hot plate testing was only seen following mild injury and not following moderate injury. Thus, moderate injury can lead to near normal hindlimb motor function in animals with major sensory deficits. Recovered fMRI signals following mild injury had a partly altered cortical distribution engaging also ipsilateral somatosensory cortex and the cingulate gyrus. Importantly, thoracic spinal cord injury also affected sensory representation of the upper nonaffected limbs. Thus, cortical and thalamic activation in response to forelimb stimulation was significantly increased 16 weeks after spinal cord injury compared to control animals. We conclude that both forelimb and hindlimb cortical sensory representation is altered following thoracic spinal cord injury. Furthermore tests of sensory function that are independent of motor behaviour are needed in rodent spinal cord injury research.