4-hydroxy-2-nonenal and Spinal-Cord-Injuries

Spinal cord injury (SCI) is a deliberating disorder with impairments in locomotor deficits and incapacitating sensory abnormalities. Harpagophytum procumbens (Hp) is a botanical widely used for treating inflammation and pain related to various inflammatory and musculoskeletal conditions. Using a modified rodent contusion model of SCI, we explored the effects of this botanical on locomotor function and responses to mechanical stimuli, and examined possible neurochemical changes associated with SCI-induced allodynia. Following spinal cord contusion at T10 level, Hp (300 mg/kg, p.o.) or vehicle (water) was administered daily starting 24 h post-surgery, and behavioral measurements made every-other day until sacrifice (Day 21). Hp treatment markedly ameliorated the contusion-induced decrease in locomotor function and increased sensitivity to mechanical stimuli. Determination of Iba1 expression in spinal cord tissues indicated microglial infiltration starting 3 days post-injury. SCI results in increased levels of 4-hydroxynonenal, an oxidative stress product and proalgesic, which was diminished at 7 days by treatment with Hp. SCI also enhanced antioxidant heme oxygenase-1 (HO-1) expression. Concurrent studies of cultured murine BV-2 microglial cells revealed that Hp suppressed oxidative/nitrosative stress and inflammatory responses, including production of nitric oxide and reactive oxygen species, phosphorylation of cytosolic phospholipases A

Topics: Aldehydes; Animals; Drug Evaluation, Preclinical; Gene Expression Regulation; Harpagophytum; Heme Oxygenase (Decyclizing); Hyperalgesia; Inflammation; Male; Mice; Motor Activity; NF-E2-Related Factor 2; Nitric Acid; Oxidative Stress; Phytotherapy; Plant Extracts; Rats; Rats, Sprague-Dawley; Reactive Nitrogen Species; Reactive Oxygen Species; Single-Blind Method; Spinal Cord Injuries; Touch

Substantial experimental evidence supports that reactive species mediate secondary damage after traumatic spinal cord injury (SCI) by inducing oxidative stress. Removal of reactive species may reduce secondary damage following SCI. This study explored the effectiveness of a catalytic antioxidant - Mn (III) tetrakis (4-benzoic acid) porphyrin (MnTBAP) - in removing reactive oxygen species (ROS), reducing oxidative stress, and improving functional recovery in vivo in a rat impact SCI model. The efficiency of MnTBAP was also compared with that of methylprednisolone - the only drug used clinically in treating acute SCI.. In vivo measurements of time courses of ROS production by microdialysis and microcannula sampling in MnTBAP, methylprednisolone, and saline (as vehicle control)-treated SCI rats showed that both agents significantly reduced the production of hydrogen peroxide, but only MnTBAP significantly reduced superoxide elevation after SCI. In vitro experiments further demonstrated that MnTBAP scavenged both of the preceding ROS, whereas methylprednisolone had no effect on either. By counting the immuno-positive neurons in the spinal cord sections immunohistochemically stained with anti-nitrotyrosine and anti-4-hydroxy-nonenal antibodies as the markers of protein nitration and membrane lipid peroxidation, we demonstrated that MnTBAP significantly reduced the numbers of 4-hydroxy-nonenal-positive and nitrotyrosine-positive neurons in the sections at 1.55 to 2.55 mm and 1.1 to 3.1 mm, respectively, rostral to the injury epicenter compared to the vehicle-treated animals. By behavioral tests (open field and inclined plane tests), we demonstrated that at 4 hours post-SCI treatment with MnTBAP and the standard methylprednisolone regimen both significantly increased test scores compared to those produced by vehicle treatment. However, the outcomes for MnTBAP-treated rats were significantly better than those for methylprednisolone-treated animals.. This study demonstrated for the first time in vivo and in vitro that MnTBAP significantly reduced the levels of SCI-elevated ROS and that MnTBAP is superior to methylprednisolone in removing ROS. Removal of ROS by MnTBAP significantly reduced protein nitration and membrane lipid peroxidation in neurons. MnTBAP more effectively reduced neurological deficits than did methylprednisolone after SCI - the first most important criterion for assessing SCI treatments. These results support the therapeutic potential of MnTBAP in treating SCI.

Topics: Aldehydes; Analysis of Variance; Animals; Blood-Brain Barrier; Cell Count; Disease Models, Animal; Hydrogen Peroxide; Locomotion; Male; Metalloporphyrins; Methylprednisolone; Microdialysis; Nervous System Diseases; Neuroprotective Agents; Oxidative Stress; Psychomotor Performance; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Recovery of Function; Spinal Cord Injuries; Time Factors; Tyrosine

The pathogenesis of cervical spondylotic myelopathy (CSM) is related to both primary mechanical and secondary biological injury. The authors of this study explored a novel, noninvasive method of promoting neuroprotection in myelopathy by using curcumin to minimize oxidative cellular injury and the capacity of omega-3 fatty acids to support membrane structure and improve neurotransmission.. An animal model of CSM was created using a nonresorbable expandable polymer placed in the thoracic epidural space, which induced delayed myelopathy. Animals that underwent placement of the expandable polymer were exposed to either a diet rich in docosahexaenoic acid and curcumin (DHA-Cur) or a standard Western diet (WD). Twenty-seven animals underwent serial gait testing, and spinal cord molecular assessments were performed after the 6-week study period.. At the conclusion of the study period, gait analysis revealed significantly worse function in the WD group than in the DHA-Cur group. Levels of brain-derived neurotrophic factor (BDNF), syntaxin-3, and 4-hydroxynonenal (4-HNE) were measured in the thoracic region affected by compression and lumbar enlargement. Results showed that BDNF levels in the DHA-Cur group were not significantly different from those in the intact animals but were significantly greater than in the WD group. Significantly higher lumbar enlargement syntaxin-3 in the DHA-Cur animals combined with a reduction in lipid peroxidation (4-HNE) indicated a possible healing effect on the plasma membrane.. Data in this study demonstrated that DHA-Cur can promote spinal cord neuroprotection and neutralize the clinical and biochemical effects of myelopathy.

Topics: Aldehydes; Animals; Brain-Derived Neurotrophic Factor; Chronic Disease; Curcuma; Diet; Disease Models, Animal; Docosahexaenoic Acids; Male; Neuroprotective Agents; Qa-SNARE Proteins; Rats; Rats, Sprague-Dawley; Spinal Cord Injuries

The reactive nitrogen species peroxynitrite (PN) has been suggested to be an important mediator of the secondary oxidative damage that occurs following acute spinal cord injury (SCI). The PN decomposition products nitrogen dioxide (*NO(2)), hydroxyl radical (*OH), and carbonate radical (*CO(3)) are highly reactive with cellular lipids and proteins. In this immunohistochemical study, we examined the temporal (3, 24, and 72 h, and 1 and 2 weeks) and spatial relationships of PN-mediated oxidative damage in the contusion-injured rat thoracic spinal cord (IH device, 200 kdyn, T10) using 3-nitrotyrosine (3-NT), a marker for protein nitration by PN-derived *NO(2) and 4-hydroxynonenal (4-HNE), an indicator of lipid peroxidation (LP) initiated by any of the PN radicals. Minimal 3-NT or 4-HNE immunostaining was seen in sham, non-injured spinal cords. In contrast, both markers showed a substantial increase at 3 h post-injury at the epicenter, that extended throughout the gray matter and into the surrounding white matter. At 24 and 72 h, the oxidative damage expanded circumferentially to involve all but a small rim of white matter tissue at the injury site, and longitudinally as much as 6-9 mm in the rostral and caudal directions. The staining was observed in neuronal soma, axons, and microvessels. At all time points except 3 h, there was no significant difference in the mean rostral or caudal extent of 3-NT and 4-HNE staining. By 1, and more so at 2 weeks, the longitudinal extent of the oxidative damage staining was greatly decreased. The spatial and temporal overlap of 3-NT and 4-HNE staining supports the concept that PN is involved in both damage produced by lipid peroxidation and protein nitration, and that antioxidant agents that target PN or PN-derived radicals should be effective neuroprotectants for acute SCI if administered during the first post-injury hours.

Topics: Aldehydes; Animals; Female; Immunohistochemistry; Lipid Peroxidation; Neurons; Oxidative Stress; Peroxynitrous Acid; Rats; Rats, Sprague-Dawley; Spinal Cord; Spinal Cord Injuries; Thoracic Vertebrae; Time Factors; Tyrosine

We hypothesized that heme oxygenase (HO)-1, the inducible form of HO, represents an important defense against early oxidative injury in the traumatized spinal cord by stabilizing the blood-spinal cord barrier and limiting the infiltration of leukocytes. To test this hypothesis, we first examined the immunoexpression of HO-1 and compared barrier permeability and leukocyte infiltration in spinal cord-injured HO-1-deficient (+/-) and wild-type (WT, +/+) mice. Heme oxygenase was expressed in both endothelial cells and glia of the injured cord. Barrier disruption to luciferase and infiltration of neutrophils were significantly greater in the HO-1+/- than WT mice at 24 h postinjury (P

Topics: Aldehydes; Animals; Blood Vessels; Blotting, Western; Brain; Enzyme Induction; Heme Oxygenase (Decyclizing); Heme Oxygenase-1; Immunohistochemistry; Inflammation; Male; Malondialdehyde; Mice; Mice, Inbred C57BL; Mice, Knockout; Myelin Basic Protein; Neutrophil Infiltration; Oxidative Stress; Spinal Cord; Spinal Cord Injuries

Peroxynitrite (PON, ONOO(-)), formed by nitric oxide synthase-generated nitric oxide radical ( NO) and superoxide radical (O(2) (-)), is a crucial player in post-traumatic oxidative damage. In the present study, we determined the spatial and temporal characteristics of PON-derived oxidative damage after a moderate contusion injury in rats. Our results showed that 3-nitrotyrosine (3-NT), a specific marker for PON, rapidly accumulated at early time points (1 and 3 h) and a significant increase compared with sham rats was sustained to 1 week after injury. Additionally, there was a coincident and maintained increase in the levels of protein oxidation-related protein carbonyl and lipid peroxidation-derived 4-hydroxynonenal (4-HNE). The peak increases of 3-NT and 4-HNE were observed at 24 h post-injury. In our immunohistochemical results, the co-localization of 3-NT and 4-HNE results indicates that PON is involved in lipid peroxidative as well as protein nitrative damage. One of the consequences of oxidative damage is an exacerbation of intracellular calcium overload, which activates the cysteine protease calpain leading to the degradation of several cellular targets including cytoskeletal protein (alpha-spectrin). Western blot analysis of alpha-spectrin breakdown products showed that the 145-kDa fragments of alpha-spectrin, which are specifically generated by calpain, were significantly increased as soon as 1 h following injury although the peak increase did not occur until 72 h post-injury. The later activation of calpain is most likely linked to PON-mediated secondary oxidative impairment of calcium homeostasis. Scavengers of PON, or its derived free radical species, may provide an improved antioxidant neuroprotective approach for the treatment of post-traumatic oxidative damage in the injured spinal cord.

Topics: Aldehydes; Animals; Biomarkers; Calcium Signaling; Calpain; Disease Progression; Female; Free Radical Scavengers; Free Radicals; Lipid Peroxidation; Nerve Degeneration; Nitric Oxide; Oxidative Stress; Peptide Fragments; Peroxynitrous Acid; Rats; Rats, Sprague-Dawley; Spectrin; Spinal Cord Injuries; Time Factors; Tyrosine; Up-Regulation

Mitochondrial dysfunction following spinal cord injury (SCI) may be critical for the development of secondary pathophysiology and neuronal cell death. Previous studies have demonstrated a loss of mitochondrial bioenergetics at 24 h following SCI. To begin to understand the evolution and study the contribution of mitochondrial dysfunction in pathophysiology of SCI, we investigated mitochondrial bioenergetics in the mid-thoracic region at 6, 12, and 24 h following contusion SCI. It is widely accepted that increased free radical generation plays a critical role in neuronal damage after SCI. Hence, to ascertain the role of free radicals in SCI-induced mitochondrial dysfunction, markers for oxidative damage, including nitrotyrosine (3-NT), lipid peroxidation byproduct (4-hydroxynonenal [HNE]), and protein oxidation (protein carbonyls) were quantified in the same samples of isolated mitochondria during the 24-h time course. The results demonstrate that a significant decline in mitochondrial function begins to occur 12 h post-injury and persists for a least 24 h following SCI. Furthermore, there was a progressive increase in mitochondrial oxidative damage that preceded the loss of mitochondrial bioenergetics, suggesting that free radical damage may be a major mitochondrial secondary injury process. Based on the present results, the temporal profile of mitochondrial dysfunction indicates that interventions targeting mitochondrial oxidative damage and dysfunction may serve as a beneficial pharmacological treatment for acute SCI.

Topics: Aldehydes; Animals; Disease Models, Animal; Disease Progression; Energy Metabolism; Female; Free Radicals; Lipid Peroxidation; Mitochondria; Oxidative Stress; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Spinal Cord; Spinal Cord Injuries; Time Factors; Tyrosine

To investigate whether phospholipase A2 (PLA2) plays a role in the pathogenesis of spinal cord injury (SCI).. Biochemical, Western blot, histological, immunohistochemical, electron microscopic, electrophysiological, and behavior assessments were performed to investigate (1) SCI-induced PLA2 activity, expression, and cellular localization after a contusive SCI; and (2) the effects of exogenous PLA2 on spinal cord neuronal death in vitro and tissue damage, inflammation, and function in vivo.. After SCI, both PLA2 activity and cytosolic PLA2 expression increased significantly, with cytosolic PLA2 expression being localized mainly in neurons and oligodendrocytes. Both PLA2 and melittin, an activator of endogenous PLA2, induced spinal neuronal death in vitro, which was substantially reversed by mepacrine, a PLA2 inhibitor. When PLA2 or melittin was microinjected into the normal spinal cord, the former induced confined demyelination and latter diffuse tissue necrosis. Both injections induced inflammation, oxidation, and tissue damage, resulting in corresponding electrophysiological and behavioral impairments. Importantly, the PLA2-induced demyelination was significantly reversed by mepacrine.. PLA2, increased significantly after SCI, may play a key role in mediating neuronal death and oligodendrocyte demyelination following SCI. Blocking PLA2 action may represent a novel repair strategy to reduce tissue damage and increase function after SCI.

Topics: Aldehydes; Animals; Apoptosis; Blotting, Western; CD11b Antigen; Cell Count; Cells, Cultured; Cytokines; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Interactions; Embryo, Mammalian; Female; Gene Expression; Glial Fibrillary Acidic Protein; Hydro-Lyases; Immunohistochemistry; Intracellular Signaling Peptides and Proteins; Melitten; Microscopy, Electron, Transmission; Motor Activity; Neurons; Oligodendroglia; Phospholipases A; Phospholipases A2; Phosphopyruvate Hydratase; Rats; Rats, Sprague-Dawley; Spinal Cord; Spinal Cord Injuries; Time Factors

Treatment with a monoclonal antibody (mAb) against the CD11d subunit of the leukocyte integrin CD11d/CD18 after spinal cord injury (SCI) decreases intraspinal inflammation and oxidative damage, improving neurological function in rats. In this study we tested whether the anti-CD11d mAb treatment reduces intraspinal free radical formation and cell death after SCI. Using clip-compression SCI in rats, reactive oxygen species (ROS) generated in injured spinal cord were detected using 2',7'-dichlorofluorescin-diacetate and hydroethidine as fluorescent probes. ROS in the injured cord increased significantly after SCI; anti-CD11d mAb treatment significantly reduced this ROS formation. Immunohistochemistry and western blotting were employed to assess the effects of anti-CD11d mAb treatment on spinal cord expression of gp91Phox (a subunit of NADPH oxidase producing superoxide) on formation of 4-hydroxynonenal (HNE, indicating lipid peroxidation) and on expression of caspase-3. We also assessed effects on cell death, determined by cell morphology. The expression of gp91Phox, formation of HNE, and cell death increased after SCI. Anti-CD11d mAb treatment clearly attenuated these responses. In conclusion, anti-CD11d mAb treatment significantly reduces intraspinal free radical formation caused by infiltrating leukocytes after SCI, thereby reducing secondary cell death. These effects likely underlie tissue preservation and improved neurological function that result from the mAb treatment.

Topics: Aldehydes; Animals; Antibodies, Monoclonal; CD11 Antigens; Cell Death; Ethidium; Female; Fluoresceins; Free Radicals; Membrane Glycoproteins; NADPH Oxidase 2; NADPH Oxidases; Rats; Rats, Wistar; Spinal Cord Compression; Spinal Cord Injuries; Superoxides

Reactive oxygen species and resultant lipid peroxidation (LPO) have been associated with central nervous system trauma. Acrolein (2-propenal) and 4-hydroxynonenal (HNE) are the most toxic byproducts of LPO, with detrimental effects in various types of cells. In this study, we used immunoblotting techniques to detect the accumulation of protein-bound acrolein and HNE. We report that protein-bound acrolein and HNE were significantly increased in guinea pig spinal cord following a controlled compression injury. The acrolein and HNE protein-adducts increased in the damaged spinal cord as early as 4 h after injury, reached a peak at 24 h after injury, and remained at a significantly high level up to 7 days after injury. Such increase of protein adducts was also observed in the adjacent segments of the injury site beginning at 24 h post injury. These results suggest that products of lipid peroxidation, especially acrolein, may play a critical role in the secondary neuronal degeneration, which follows mechanical insults.

Topics: Acrolein; Aldehydes; Animals; Blotting, Western; Densitometry; Female; Guinea Pigs; Immunohistochemistry; Lipid Peroxidation; Protein Binding; Proteins; Spinal Cord Injuries

Oxidative stress plays an important role in the pathogenesis of neurodegeneration after the acute central nervous system injury. We reported previously that increased nitric oxide (NO) production following spinal cord hemisection tends to lead to neurodegeneration in neurons of the nucleus dorsalis (ND) that normally lacks expression of neuronal NO synthase (nNOS) in opposition to those in the red nucleus (RN) that constitutively expresses nNOS. We wondered whether oxidative stress could be a mechanism underlying this NO involved neurodegeneration. In the present study, we examined oxidative damage evaluated by the presence of 4-hydroxynonenal (HNE) and iron accumulation and expression of putative antioxidant enzymes heme oxygenase-1 (HO-1) and superoxide dismutase (SOD) in neurons of the ND and RN after spinal cord hemisection. We found that HNE expression was induced in neurons of the ipsilateral ND from 1 to 14 days following spinal cord hemisection. Concomitantly, iron staining was seen from 7 to 14 days after lesion. HO-1, however, was only transiently induced in ipsilateral ND neurons between 3 and 7 days after lesion. In contrast to the ND neurons, HNE was undetectable and iron level was unaltered in the RN neurons after spinal cord hemisection. HO-1, SOD-Cu/Zn and SOD-Mn were constitutively expressed in RN neurons, and lesion to the spinal cord did not change their expression. These results suggest that oxidative stress is involved in the degeneration of the lesioned ND neurons; whereas constitutive antioxidant enzymes may protect the RN neurons from oxidative damage.

Topics: Aldehydes; Animals; Antioxidants; Cell Count; Functional Laterality; Gene Expression Regulation; Histocytochemistry; Immunohistochemistry; Iron; Male; Mediodorsal Thalamic Nucleus; Neurons; Oxidative Stress; Rats; Rats, Wistar; Red Nucleus; Spinal Cord Injuries; Staining and Labeling; Superoxide Dismutase; Time Factors

The goal of the present study was to determine in vivo whether peroxynitrite, at the concentration and duration produced by SCI, contributes to membrane lipid peroxidation (MLP) after traumatic spinal cord injury (SCI) and the capability of a broad spectrum scavenger of reactive species, Mn (III) tetrakis (4-benzoic acid) porphyrin (MnTBAP), to reduce MLP. This was accomplished by administering a peroxynitrite donor 3-morpholinosydnonimine (SIN-1) into the gray matter of an uninjured rat spinal cord through a microdialysis fiber to generate ONOO at the SCI-elevated levels. The resulting MLP was characterized by measuring the productions of extracellular malondialdehyde and of intracellular 4-hydroxynonenal. We demonstrated that extracellular SIN- 1 administration significantly increased the concentration of malondialdehyde (p < 0.001) and the numbers of hydroxynonenal-positive cells (p < 0.001) as compared to a control group in which ACSF was administered. Simultaneous administration of MnTBAP through a second microdialysis fiber significantly reduced SIN-1-induced malondialdehyde production (p < 0.001) and the numbers of HNE-positive cells (p < 0.001). There was no significant difference between MnTBAP-treated and ACSF-controls (p = 0.3). These results demonstrate in vivo that (1) SCI-produced levels of peroxynitrite sufficient to cause MLP, and therefore that peroxynitrite is an agent of secondary damage after acute SCI; (2) MnTBAP can efficiently reduce SIN-1-induced MLP.

Topics: Aldehydes; Animals; Cell Membrane; Free Radical Scavengers; Lipid Peroxidation; Male; Malondialdehyde; Metalloporphyrins; Molsidomine; Nitric Oxide Donors; Peroxynitrous Acid; Phospholipids; Rats; Rats, Sprague-Dawley; Spinal Cord Injuries

Lipid peroxidation (LPO) is considered a major factor in damage spread after spinal cord injury (SCI). Therapies that limit LPO after SCI have demonstrated some utility in clinical trials, but more effective treatments are needed. In the present study the effects of augmenting SC levels of the endogenous antioxidant glutathione (GSH) on LPO after SCI were studied in a rat contusion injury model. A significant decrease in GSH occurred 1h after SCI which was paralleled by increases of 123% in malondialdehyde (MDA) and >500% in the 4-hydroxyalkenals (4-HA's), two LPO products. SC irrigation with gamma-glutamylcysteine (GC) preserved GSH and reduced 4-HA's below naive levels but had no effect on MDA. By 24 h after SCI, MDA returned to naive levels but 4-HA's were still elevated. Once again, GC treatment reduced 4-HA's. 4-HA's are much more reactive than MDA and are considered among the most toxic LPO products. These results suggest that (1) conditions after SCI may favor particular branches of the LPO pathway leading to differential LPO product levels, (2) MDA measurement is not by itself an adequate test for the presence or magnitude of LPO after SCI, (3) binding of GSH to 4-HA's may be an important mechanism by which the GSH system confers protection against LPO after SCI, and (4) SC GSH can be augmented after trauma by local irrigation with GC. These results also suggest that GSH augmentation may be an effective strategy for curtailment of LPO-mediated damage in acute phase SCI.

Topics: Acute Disease; Aldehydes; Animals; Antioxidants; Female; Glutathione; Glutathione Disulfide; Glutathione Transferase; Infusion Pumps, Implantable; Lipid Peroxidation; Rats; Rats, Sprague-Dawley; Spinal Cord Injuries

The authors tested the hypothesis that breach of the blood-spinal cord barrier (BSCB) will produce evidence of oxidative stress and that a similar staining pattern will be seen between 4-hydroxynonenal (HNE)/protein complexes and extravasated immunoglobulin G (IgG).. Adult female Fischer 344 rats, each weighing 200 to 225 g, were subjected to a spinal cord contusion at T-10 by means of a weight-drop device. Spinal cord tissue was assessed for oxidative stress by localizing extravasated plasma contents with a monoclonal antibody for rat IgG and protein conjugation with HNE, which is an aldehyde byproduct of lipid peroxidation. The animals were killed at 1 and 6 hours, and 1, 2, and 7 days after surgery. Maximum HNE/protein staining was observed at 2 days postinjury, and HNE/protein and IgG manifested similar staining patterns. Analysis revealed a graduated but asymmetrical rostral-caudal response relative to the T-10 injury site. Both HNE/protein complex and IgG staining revealed that the caudal levels T-11 and T-12 stained significantly more intensely than the rostral levels T-9 and T-8, respectively. A higher percentage of neurons positive for HNE/protein immunostaining was observed in spinal cord levels caudal to the injury site compared with equidistant rostral regions. Protein dot-blot assays also revealed a similar asymmetrical rostral-caudal HNE/protein content. To analyze the timing of the BSCB breach, another group of animals received identical contusions, and horseradish peroxidase (HRP) was injected 10 minutes before or at various times after injury (1, 3, and 6 hours, and 1, 2, and 7 days). Maximum HRP permeability was seen immediately after injury, with a significant decrease occurring by 1 hour and a return to control levels by 2 days posttrauma.. Data from this study indicate possible compromise of neuronal, axonal, glial, and synaptic function after trauma, which may be a factor in motor deficits seen in animals after spinal cord contusion. The colocalization of the IgG stain with the HNE/protein stain is consistent with the hypothesis of a mutual cause-effect relationship between BSCB and oxidative stress in central nervous system trauma.

Topics: Aldehydes; Analysis of Variance; Animals; Antibodies, Monoclonal; Axons; Blood; Coloring Agents; Contusions; Cross-Linking Reagents; Cysteine Proteinase Inhibitors; Disease Models, Animal; Female; Follow-Up Studies; Horseradish Peroxidase; Immunoblotting; Immunoenzyme Techniques; Immunoglobulin G; Lipid Peroxidation; Neuroglia; Neurons; Oxidative Stress; Paralysis; Permeability; Protein Binding; Proteins; Rats; Rats, Inbred F344; Spinal Cord Injuries; Synapses

Traumatic injury to the spinal cord initiates a host of pathophysiological events that are secondary to the initial insult. One such event is the accumulation of free radicals that damage lipids, proteins, and nucleic acids. A major reactive product formed following lipid peroxidation is the aldehyde, 4-hydroxynonenal (HNE), which cross-links to side chain amino acids and inhibits the function of several key metabolic enzymes. In the present study, we used immunocytochemical and immunoblotting techniques to examine the accumulation of protein-bound HNE, and synaptosomal preparations to study the effects of spinal cord injury and HNE formation on glutamate uptake. Protein-bound HNE increased in content in the damaged spinal cord at early times following injury (1-24 h) and was found to accumulate in myelinated fibers distant to the site of injury. Immunoblots revealed that protein-bound HNE levels increased dramatically over the same postinjury interval. Glutamate uptake in synaptosomal preparations from injured spinal cords was decreased by 65% at 24 h following injury. Treatment of control spinal cord synaptosomes with HNE was found to decrease significantly, in a dose-dependent fashion, glutamate uptake, an effect that was mimicked by inducers of lipid peroxidation. Taken together, these findings demonstrate that the lipid peroxidation product HNE rapidly accumulates in the spinal cord following injury and that a major consequence of HNE accumulation is a decrease in glutamate uptake, which may potentiate neuronal cell dysfunction and death through excitotoxic mechanisms.

Topics: Aldehydes; Amino Acid Transport System X-AG; Animals; Antibody Specificity; ATP-Binding Cassette Transporters; Biological Transport; Blotting, Western; Cell Death; Cross-Linking Reagents; Female; Free Radicals; Glutamic Acid; Immunohistochemistry; Lipid Peroxidation; Neurons; Neurotoxins; Rats; Rats, Inbred Strains; Spinal Cord Injuries; Wounds and Injuries