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Background: Among spinal cord injuries, secondary injury is considered to be a "reversible" process and seems to be a key target for the treatment of spinal cord injury. Recently, macrophage migration inhibitory factor (MIF) has been reevaluated as being one of the most important cytokines which act during wound healing, proliferation and differentiation of cells. However, the expression of MIF in the spinal cord has not been investigated yet., Purpose: The purpose of this paper is to demonstrate the MIF expression in normal rat spinal cord and to evaluate the kinetics of MIF after spinal cord injury., Materials & Methods: Female Wistar (280-320 g) rats were studied. Spinal cord injury was made by the clip compression method at the level of C7/Th1 (56 g, For 1 min.). The expression of MIF was examined by immunohistochemistry and northern blot analysis. MIF content in the cerebrospinal fluid (CSF) was measured by enzyme-linked immunosorbent assays (ELISA). Furthermore, to examine the MIF function on neuronal cell, cell proliferation assay (MTS assay) was carried out using PC12, pheochromocytoma cell line, and LN444, glioblastoma cell line, in the presence of anti-MIF monoclonal antibody., Results: MIF stain was positive in normal rat spinal cord white matter. The expression of MIF decreased between 1 hour and 6 hours after injury. It was found to have re-appeared 24 hours after injury. The kinetics of MIF mRNA expression showed reverse-correlation with those of the MIF positive stain. MIF content in CSF was found to be elevated soon after injury. MTS assay suggested that MIF had some proliferative function on neuronal cells., Conclusion: MIF exists in the rat white matter. And it's immediately released into the CSF and then re-synthesized 24-hr after injury. MIF shows a cell proliferative function on neuronal cells. These results suggest that MIF plays an important role for secondary spinal cord injury.

Glucocorticoid has been clinically used for the treatment of acute spinal cord injury (SCI) to enhance the neurological recovery, but the relevance of the use of steroid is not fully discussed. Neurotrophins play important roles in normal development of central and peripheral nervous system. It is reported that traumatic insults to the brain alter the expression of these neurotrophins. These responses are considered to trigger a cascade of cellular protection and repair. First, we investigated the temporal and spatial expression patterns of neurotrophin and IL-1 beta mRNAs in the area of spinal cord lesion. Second, we examined if methylprednisolone (MP) affects the expression of these genes in SCI. Male Sprague-Dawley rats (250-300 gms) were laminectomized at T10. Spinal cord was crushed by clipping (holding force 60 gms, 1 sec). Rats were killed at 1, 6, 12, 24 h, and 72 h after the injury and the spinal cord was rapidly removed and frozen sections were cut. Second, the other group of rats were treated with MP (165 mg/kg) just after SCI and sacrificed at 6 h. The levels of neurotrophins and IL-1 beta mRNAs were evaluated by in situ hybridization histochemistry. IL-1 beta mRNA level was elevated at 1 h and 6 h and attenuated at 12 h. The increased level of BDNF and NT3 mRNAs were first observed at 6 h and the labeling was enhanced at 24 h and 72 h. In MP treated group, the levels of IL-1 beta, BDNF and NT3 mRNAs were attenuated compared with those of MP-untreated SCI group. Steroid hormone therapy diminishes the post-traumatic inflammatory cascades which produce edema and swelling and worsen neuronal injury. However, glucocorticoid may hinder the endogenous repair mechanism. Our data show that MP depress the production of BDNF and NT3 following SCI, which might be disadvantageous to the survival of spinal neurons.

Unlabelled: Acute spinal cord trauma may initiate a cascade of hemodynamic and biochemical events characterized by direct mechanical destruction of neurons, hemorrhages and significant increases in active substances (glutamate, prostaglandins (PGs), ets.). However, there are no data which define the effect of acute spinal cord trauma on biochemical changes in the spinal cord. The aim of the present study was to examine time-dependent biochemical changes in the spinal cord by intrathecal microdialysis in the acute stage of spinal cord turama., Methods: Male Sprague-Dawley rats (300-325g) were implanted with a dialysis catheter and PE-10 catheter in the intrathecal space of the lumbar spinal cord. Three days later the animals were anesthetized with halothane and 10-min dialysis samples were collected. Compression spinal cord injury was produced by inflating an extradural 2F-Fogarty balloon catheter (L4-L5) for 60 sec. After compression, samples were collected while the animals remained anesthetized for 30 min, and additional samples were collected until 4hr post-injury. At the end of the dialysis period neurological function was briefly evaluated. Microdialysate samples were analyzed for glutamate, aspartate, and taurine by HPLC-UV and for PGE2 by RIA., Results: Compression of the spinal cord caused significant increases in the levels of all amino acids. The greatest in creases were in glutamate and aspartate at 10 min, followed by a decrease over the rest of the 4-hr period. Similarly, PGE2 showed an immediate increase after compression, with normalization between 1-2 hr, but this was followed by a secondary increase at 2-4 hr. Neurologically, all animals displayed complete paralysis with loss of sensory function that remained unchanged for the 4-hr period., Comment: The present study clearly demonstrated the time course of spinal amino acids and PGE2 release by using a chronically implanted dialysis catheter after spinal cord compression and also showed a correlation with neurological deterioration. These biochemical changes in the extracellular space may display various, multiphasic patterns reflecting progressively developing edema and corresponding ischemia.

Brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3) are neurotrophic factors involved in the neurotrophin family. However, while their biological activities on cultured neurons and their distribution have been reported, their functions in the central nervous system including the spinal cord are still unknown. We have recently developed antibodies to BDNF and to NT-3 synthetic peptides with different amino acid sequences as tools to visualize these molecules in in vivo tissue. Each antibody was specific to either BDNF or NT-3. Using these antibodies, we investigated the distribution of BDNF- or NT-3-like immunoreactivities (LI) in the spinal cord following transection in rats. In Group with an uninjured spinal cord as control, both BDNF-LI and NT-3-LI were localized in the gray matter, and particularly in the motor neurons of the ventral horn, and in some axons and glial cells of the white matter. In Group after transection of the spinal cord as a model of spinal injury, the motor neurons in the spinal cord, were strongly stained by anti-NT-3 antibodies and less strong by anti-BDNF antibodies at 3 weeks after injury. In the gray matter at 4 weeks after injury, astrocyte-like cells were markedly stained with anti-BDNF antibodies These findings indicate that in the spinal cord after injury, the biosynthesis of BDNF was upregulated in a different manner from that of NT-3 and that BDNF and NT-3 were both involved in the repair mechanisms, directing the amelioration of the spinal cord injury.

Spinal microenvironment and metabolic alterations after experimental contusional injury of the spinal cord were evaluated in the same Wistar rats. Severe spinal cord injury was made under light GOF anesthesia with a 10 g weight drop onto the exposed Th-8 spinal cord from a 10 cm height and then halothane was ceased. The author studied extracellular potassium activity ([K+]e) and DC potential for 2 hours after paraplegic spinal cord injury in conscious rats. Furthermore, at 2 hours after cord injury, local spinal cord glucose utilization (1-SCGU) was measured with quantitative autoradiographic 2-[14C] deoxy-glucose method (Sokoloff et al.). [K+]e in injured spinal cords was 59 +/- 5 (mean +/- S.E.M.) mEq at 10 min after injury and was cleared with an exponential half-life of 1 hour. At 2 hours after injury [K+]e was still high with a value of 16 +/- 1 mEq compared with 4 mEq of control animals. DC potential changes was a mirror image of that of [K+]e. DC potential changed by a mean of 10.7 mV positively from 10 min. to 2 hours after injury. 1-SCGU at the impact site was extremely low in both white and gray matters. At 6mm rostral from the impact center 1-SCGU was remarkably reduced in the gray matter, and in the lateral white matter. But at 3 mm rostral 1-SCGU was well preserved. And at 20 mm rostral there was no difference in 1-SCGU with control animals. Massive potassium efflux from the injured spinal cord to the adjacent spinal segment was clarified at this experiment.(ABSTRACT TRUNCATED AT 250 WORDS)

Thromboxane A2 (TXA2) is an eicosanoid with potent platelet aggregation and vasoconstricting activity, while prostaglandin I2 (PGI2) antagonizes its activity. But these eicosanoids are so labile that the stable degradation products, thromboxane B2 (TXB2) and 6-keto-prostaglandin F1 alpha (6-keto-PGF1 alpha), are determined in biological materials. In a rat spinal cord compression injury model, a production of TXB2 reached a peak (133.6 +/- 13.8 pmol/g cord) 5 minutes after compression injury, while that of 6-keto-PGF1 alpha slightly increased (26.2 +/- 11.7 pmol/g cord). And the magnitude of the increase in TXB2 and the extent of post-traumatic vascular damage as determined by fluorescein uptake were both dependent on the degree of spinal cord compression injury. We also studied the effect of a selective TXA2 synthetase inhibitor, OKY-046 [E)-3-[4-(1-imidazolylmethyl)phenyl]-2-propenoic acid), both on TXB2 production in the injured spinal cord and post-traumatic vascular damage. When OKY-046 was administered intravenously 10 minutes prior to compression injury at a dose of 500 micrograms/kg body weight, the increased production of TXB2 was inhibited by about 80% and uptake of sodium fluorescein was reduced by a maximum of 40%. When the compression injury was induced before OKY-046 was administered, the inhibitory effect of OKY-046 on TXB2 production decreased depending on the duration before administration. In contrast, the 6-keto-PGF1 alpha level was not affected in the presence of OKY-046.

Using a rat severe spinal cord injury model, serial determinations of local spinal cord blood flow and energy metabolism including ATP, ADP, AMP, energy charges, local tissue oxygen consumption and changes in pH values in the injured spinal cord tissue were compared topographically. Initially, following injury, energy metabolism was increased with increased ATP utilization and within 30 minutes energy metabolites were severely depleted. By 2 hours after spinal cord injury, a state of severe anaerobic metabolism was associated with the depletion of energy metabolites. This severe state of metabolic disruption and energy metabolite depletion, especially ATP, was followed by a progressive disturbance of spinal cord blood flow. By 3 hours following injury, the disturbances of spinal cord blood flow and energy metabolism were equal in severity. This study suggested that in the severely injured rat spinal cord, an initial depletion of energy metabolites creates a disruption of metabolic function which results in a progression of pathophysiological factors including.

Local tissue oxygen consumption, nicotinamide-adenine dinucleotide hydrogenase, coenzyme-Q and alpha-tocopherol were measured and the relationships between damage to the hydrogen electron transport system and free radical reactions were examined in a irreversible rat spinal cord injury model. Damage to the hydrogen electron transport system became apparent in the injured spinal cord segment earlier than expected. Oxygen consumption declined to 26% of the baseline level within five to 30 minutes after injury, by one hour, further declined by 21% and by two hours another 17% and by three to four hours by an additional 13%. This severe disturbance of oxygen metabolism was associated with a marked reduction of adenosine triphosphate. A reduction in coenzyme-Q by 50% was noted within 10 minutes after injury and might be at least partially responsible for these changes since a reduction of coenzyme-Q promotes the semiquinone (.coenzyme-Q) forming reaction and also produces the superoxide radical X O2-. While coenzyme-Q reacts with H+ ion, this superoxide radical X O2-, produce a state of scavenger wastage and hyperoxygenation of nicotinamide-adenine dinucleotide hydrogenase at two hours after injury. Lipid peroxigenation resulted from damage to the hydrogen electron transport system which created a state of energy metabolite disruption and cellular membrane damage and ultimately led to cellular autolysis.