Douglas K. Anderson

Pathophysiology of spinal cord trauma

This article reviews the pathophysiology of spinal cord injury. The focus is on the role of post-traumatic membrane lipid changes, including lipid hydrolysis with enzymatic lipid peroxidation (ie, eicosanoid production) and nonenzymatic, free radical-induced lipid peroxidation in the secondary autodestruction of injured spinal cord tissue. A speculative etiopathogenesis of secondary injury is presented in an attempt to explain the importance and order of the pathophysiologic events that result in tissue death and the apparent effectiveness of diverse pharmacologic agents in the treatment of experimental spinal cord injury.

Effects of Methylprednisolone and the Combination of α-Tocopherol and Selenium on Arachidonic Acid Metabolism and Lipid Peroxidation in Traumatized Spinal Cord Tissue

Abstract: Traumatic injury of the spinal cord leads to a series of pathological events that result in tissue necrosis and paralysis. Among the earliest biochemical reactions are hydrolysis of fatty acids from membrane phospholipids, production of biologically active eicosanoids, and peroxidation of lipids. This study examines the effect of agents purported to improve recovery following spinal cord trauma, methyl‐prednisolone sodium succinate (MPSS) and the combination of α‐tocopherol and selenium (Se), on the posttraumatic alterations of membrane lipid metabolism. Pretreatment with either MPSS or a‐tocopherol and Se reduced the trauma‐induced release of total FFA including arachidonate in the injured spinal cord tissue. In addition, these agents decreased the postinjury levels of prostanoids. Pre‐treatment with either MPSS or a‐tocopherol and Se also completely prevented the trauma‐induced loss of cholesterol while inhibiting the increase of a cholesterol peroxidation product, 25‐hydroxycholesterol. These data suggest that: (a) perturbation of membrane lipid metabolism may contribute to the tissue necrosis and functional deficit of spinal cord injury and (b) MPSS or the combination of a‐tocopherol and Se may protect injured spinal cord tissue, at least in part, by limiting these posttraumatic membrane lipid changes.

Fetal cell grafts into resection and contusion/compression injuries of the rat and cat spinal cord

This article reviews recent findings concerning the feasibility, basic neurobiology, and potential functional benefits of fetal CNS tissue grafts into acute and chronic lesions of the adult spinal cord. In the rat, neuro-anatomical observations suggest that transplants into resection cavities establish neuritic projections that could functionally reunite separated rostral and caudal segments of the host spinal cord. Furthermore, some complementary electrophysiological evidence has been obtained for synaptic connectivity between host and graft neurons. In these studies, extracellular single-unit activity was evoked in fetal spinal cord (FSC) transplants by stimulating host dorsal roots that had been juxtaposed to donor tissue at the time of transplantation. In other investigations, we examined whether grafts could also establish axonal projections to appropriate areas of gray matter in the chronically injured spinal cord. For this purpose, fetal serotoninergic (5-HT) neurons were injected caudal to complete spinal cord transections that had been made 1-3 months earlier. Immunocytochemistry revealed that these cells projected their axons into gray matter regions normally innervated by bulbospinal 5-HT neurons. To investigate transplantation in a more clinically relevant lesion model, a third group of experiments involved injection of dissociated cell suspensions into acute [less than 24 h postinjury (p.i.)]), subchronic (7-10 days p.i), and chronic (greater than or equal to one month, p.i.) contusion lesions. Such grafts routinely filled areas that otherwise would have been regions of cavitation extending rostral-caudal distances of approximately 7 mm. FSC transplants in such injuries also appeared to influence some aspects of motoneuron excitability and hindlimb locomotion. More recent studies of the cat spinal cord have extended these findings in the rat by showing long-term survival (greater than 2 years) of fetal CNS allografts in recipients with either subtotal transection or compression lesions. Preliminary studies of connectivity have also shown host-graft projection patterns similar to those seen in the rat. Behavioral analyses are currently underway to examine the effects of fetal grafts in cats with chronic postcompression lesions. These observations in the rat and cat are discussed in the general context of basic biological and clinical issues relevant to the long-term objective of promoting functional improvement in the damaged spinal cord.

Feasibility and Safety of Neural Tissue Transplantation in Patients with Syringomyelia

Transplantation of fetal spinal cord (FSC) tissue has demonstrated significant potential in animal models for achieving partial anatomical and functional restoration following spinal cord injury (SCI). To determine whether this strategy can eventually be translated to humans with SCI, a pilot safety and feasibility study was initiated in patients with progressive posttraumatic syringomyelia (PPTS). A total of eight patients with PPTS have been enrolled to date, and this report presents findings for the first two patients through 18 months postoperative. The study design included detailed assessments of each subject at multiple pre- and postoperative time points. Outcome data were then compared with each subjects own baseline. The surgical protocol included detethering, cyst drainage, and implantation of 6-9-week postconception human FSC tissue. Immunosuppression with cyclosporine was initiated a few days prior to surgery and continued for 6 months postoperatively. Key outcome measures included: serial magnetic resonance imaging (MRI) exams, standardized measures of neurological impairment and functional disability, detailed pain assessment, and extensive neurophysiological testing. Through 18 months, the first two patients have been stable neurologically and the MRIs have shown evidence of solid tissue at the graft sites, without evidence of donor tissue overgrowth. Although it is still too soon to draw any firm conclusions, the findings from the initial two patients in this study suggest that intraspinal grafting of human FSC tissue is both feasible and safe.

Iron-induced lipid peroxidation in spinal cord: Protection with mannitol and methylprednisolone

The ability of the free radical scavenger, mannitol, and the synthetic glucocorticoid, methylprednisolone sodium succinate (MPSS) to reverse the effects of iron catalyzed free radical induced lipid peroxidation was assessed in the feline spinal cord. Ferrous chloride (100 mM) was infused into the gray matter of lumbar spinal cord, the region frozen in situ, removed, and homogenates of the gray matter analyzed for activity of Na+,K+-ATPase and levels of malondialdehyde (MDA). ATPase activity had declined to approximately 30% of control by 2 h after FeCl2 infusion and remained at this level through 24 h. Malondialdehyde values were elevated almost twofold at 2 h. Mannitol essentially reversed the effects of FeCl2 infusion on Na+,K+-ATPase activity and MDA production. These results may implicate the hydroxyl radical (. OH), or an oxidizing species with . OH-like reactivity, as the initiating radical species in this model of lipid peroxidation. Similarly, MPSS prevented the decline in spinal cord Na+,K+-ATPase activity and rise in MDA levels that were induced by FeCl2 infusion. This demonstrated that at the dosage levels used in this study, MPSS was an effective antioxidant. This finding provides presumptive evidence suggesting that, at least in experimental animals, the effectiveness of MPSS in preventing the tissue necrosis and paralysis that is the sequelae of spinal cord trauma may reside, in part, in the capacity of this glucocorticord to quench peroxidative reactions in the injured tissue.

Spinal cord injury and protection

Subsequent to traumatic injury of the spinal cord, a series of pathophysiological events occurs in the injured tissue that leads to tissue destruction and paraplegia. These include hemorrhagic necrosis, ischemia, edema, inflammation, neuronophagia, loss of Ca2+ from the extracellular space, and loss of K+ from the intracellular space. In addition, there is trauma-initiated lipid peroxidation and hydrolysis in cellular membranes. Both lipid peroxidation and hydrolysis can damage cells directly; hydrolysis also results in the formation of the biologically active prostaglandins and leukotrienes (eicosanoids). The time course of membrane lipid alterations seen in studies of antioxidant interventions suggests that posttraumatic ischemia, edema, inflammation, and ionic fluxes are the result of extensive membrane peroxidative reactions and lipolysis that produce vasoactive and chemotactic eicosanoids. A diverse group of compounds has been shown to be effective in ameliorating spinal cord injury in experimental animals. These include the synthetic glucocorticoid methylprednisolone sodium succinate (MPSS); the antioxidants vitamin E, selenium, and dimethyl sulfoxide (DMSO); the opiate antagonist naloxone; and thyrotropin-releasing hormone (TRH). With the exception of TRH, all of these agents have demonstrable antioxidant and/or anti-lipid-hydrolysis properties. Thus the effectiveness of these substances may lie in their ability to quench membrane peroxidative reactions or to inhibit the release of fatty acids from membrane phospholipids, or both. Whatever the mode of action, early administration appears to be a requirement for maximum effectiveness.

Fetal Neural Grafts and Repair of the Injured Spinal Cord

Solid or suspension grafts of fetal spinal cord (FSC), caudal brainstem (FBSt), neocortex (FNCx) or a combination of either FSC/FNCx or FSC/FBSt were placed into cavities produced by static loading (i.e., compression) of the spinal cord of adult cats two to 30 weeks after injury. Extensively vascularized, viable graft tissue was found in all animals with the exception of two cats which showed active rejection of their transplants. Surviving grafts showed many immature characteristics 6–9 weeks after transplantation. However, by 20–30 weeks FSC and FBSt grafts were more mature. Grafts integrated with the host gray and white matter and neuritic processes from both host and graft were seen crossing the host‐graft interface. Host calcitonin gene related peptide (CGRP)‐like immunoreactive axons could be traced into FSC and FBSt grafts. A more restricted ingrowth of host serotonin (5‐HT)‐like immunoreactive fibers was seen in FSC grafts. Our results suggest that the capacity of homotypic transplants to promote recovery of function is greater than heterotypic transplants. Additionally, it appears that the functional capacity of the graft depends upon graft survival, the time interval between injury and transplantation, and whether or not the lesion cavity was debrided prior to grafting.

Neurophysiological assessment of the feasibility and safety of neural tissue transplantation in patients with syringomyelia.

The feasibility and safety of a procedure involving fetal spinal cord tissue transplantation in patients with syringomyelia was assessed using a neurophysiological protocol designed to quantitate peripheral nerve function, spinal cord reflex excitability, and spinal cord conduction pathways essential for somatosensory evoked potentials. We report here data obtained before and for 18 months following the transplantation procedure performed on the first two patients in this study. The neurophysiological assessment protocols included measures of cortical and spinal cord evoked potentials, H-reflex excitability, and peripheral nerve conduction. Prior to the procedure, both patients had significant deficits on some of the neurophysiological measures, for example, lower extremity cortical evoked potentials. However, robust measures of intact pathways, such as upper extremity cortical evoked potentials, were also observed preoperatively in both patients. Thus, it was anticipated that conduction in these intact pathways could be at risk either from complications from the transplantation procedure and/or from continued expansion of the syrinx. Following the transplantation procedure, no negative changes were observed in any of the neurophysiological measures in either patient. In addition, patient 1 showed a decrease in the rate potentiation of tibial H-reflexes on the right side and an increase in the response probability of left tibial H-reflexes. The results of this postoperative longitudinal assessment provide a first-level demonstration of the safety of the intraspinal neural tissue transplantation procedure. However, the consideration of safety is currently limited to the grafting procedure itself, since the long-term fates of the donor tissue in these two patients remain to be shown more definitively.

Characteristics of Human Fetal Spinal Cord Grafts in the Adult Rat Spinal Cord: Influences of Lesion and Grafting Conditions ☆

The present study evaluated the growth potential and differentiation of human fetal spinal cord (FSC) tissue in the injured adult rat spinal cord under different lesion and grafting conditions. Donor tissue at 6-9 weeks of gestational age was obtained through elective abortions and transplanted either immediately into acute resection (solid grafts) or into chronic contusion (suspension and solid grafts) lesions (i.e., 14-40 days after injury) in the thoracic spinal cord. The xenografts were then examined either histologically in plastic sections or immunocytochemically 1-3 months postgrafting. Intraspinal grafts in acute lesions demonstrated an 83% survival rate and developed as well-circumscribed nodules that were predominantly composed of immature astrocytes. Solid-piece grafts in chronic contusion lesions exhibited a 92% survival rate and also developed as nodular masses. These grafts, however, contained many immature neurons 2 months postgrafting. Suspension grafts in chronic contusion lesions had an 85% survival rate and expanded in a nonrestrictive, diffuse pattern. These transplants demonstrated large neuronally rich areas of neural parenchyma. Extensive neuritic outgrowth could also be seen extending from these grafts into the surrounding host spinal cord. These findings show that human FSC tissue reliably survives and differentiates in both acute and chronic lesions. However, both the lesion environment and the grafting techniques can greatly influence the pattern of differentiation and degree of host-graft integration achieved.

Effect of glutamate and its analogs on diacylglycerol and monoacylglycerol lipase activities of neuron-enriched cultures

Neuron-enriched cultures from fetal mouse spinal cord contain diacylglycerol and monoacylglycerol lipases. The treatment of neuron-enriched cultures with glutamate or NMDA resulted in a dose- and time-dependent stimulation in diacylglycerol and monoacylglycerol lipase activities. The elevation in the activities of lipases was blocked by the NMDA receptor antagonists, dextrorphan and MK-801, suggesting that lipase stimulation is a receptor-mediated process. The treatment of neuron-enriched cultures with kainate had no effect on diacylglycerol and monoacylglycerol lipase activities. The stimulation of diacylglycerol and monoacylglycerol lipase activities by glutamate and NMDA suggests that these enzymes may play an important role in processes mediated by the NMDA type of the glutamate receptors.