Wild-type mice and mice lacking nitric oxide synthase (NOS) of neuronal type or TNF-alpha were subjected to an extradural compression of the thoracic spinal cord. The functional outcome of the hind limbs was assessed by using a motor function score (MFS). The injury resulted in paraplegia of the hind limbs in wild-type mice at day 1 after injury. Gradual recovery was observed during the following 14 days. Injured NOS -/- animals had an improved hind limb motor function during the entire observation period compared to wild-type controls. The difference was statistically significant on day 10 (p < 0.022) and day 14 (p < 0.048) after injury. At the site of injury, there was a trend of gray matter preservation in NOS -/- mice, as measured by MAP2 staining (p < 0.077). Injured mice lacking TNF-alpha had the lowest motor score among all the groups on day 1. During the following period, they had motor scores similar to those of wild-type controls and there was no significant difference at any time point. TNF-alpha -/- animals showed a trend of decreased white matter preservation compared to wild-type animals (p < 0.097). Our study shows that after spinal cord injury, mice lacking NOS have a better functional ability of their hind limbs than controls with the same degree of injury. This would indicate that the functional outcome is influenced in a negative way in wild mice by the presence of NO. The degree of secondary damage to the spinal cord might be attenuated in NOS-deficient mice.

Traumatic spinal cord injury is followed by infiltration of leukocytes, influenced by endothelial adhesion molecules such as ICAM-1 and P-selectin. In order to evaluate the pathogenetical role of these molecules, wild-type mice and mice lacking ICAM-1 and P-selectin were subjected to an experimental spinal cord compression of two degrees of severity. Hind limb motor function decreased after injury in all animals but the groups of injured ICAM-1/P-selectin knockout animals had a better functional outcome during the entire observation period of 14 days. This difference was statistically significant on day 1. Our results indicate that adhesion molecules influence the functional outcome after spinal cord injury in a negative way and may be a target for future therapy of neurotrauma.

We have performed an immunohistochemical study on the expression of the adhesion molecules ICAM-1 and CD11b 1 h to 1 week following a compression injury to the rat spinal cord. The spinal cord of control animals showed ICAM-1 expression in some vessels and in the leptomeninges. Mechanical compression of the spinal cord induced an endothelial upregulation of ICAM-1 that was maximal in rats surviving 1-2 days after injury. This reaction was seen at the center of the lesion as well as in the perifocal zones. Apart from the endothelial upregulation, increased ICAM-1 expression also was found in leptomeningeal and ependymal cells of traumatized animals. In control animals resting microglial cells were moderately CD11b immunoreactive. Trauma induced a rapid microglial upregulation of CD11b in the white matter that was evident even at 1 h after injury. By 1 day to 1 week posttrauma conformational changes consistent with microglial activation, i.e., transformation into phagocytic microglial cells, were seen in the white matter. In the gray matter, CD11b immunohistochemistry revealed massive infiltration of phagocytic microglial cells and macrophages in animals surviving 1 day to 1 week. Intravascular and infiltrating leukocytes were intensely CD11b immunopositive. As reflected by CD11b immunohistochemistry, the maximal infiltration of polymorphonuclear leukocytes occurred at 2 days after the insult. Endothelial upregulation of ICAM-1 facilitates adhesion and extravasation of leukocytes by binding to the counterreceptor CD11b. Knowledge regarding the expression and cellular distribution of such molecules after central nervous system trauma is important since inflammatory mechanisms have been suggested to be involved in secondary neurological damage and thus constitute potential targets of therapy.

Clomethiazole (CMZ) has a neuroprotective effect in experimental focal and global forebrain ischemia. This neuroprotective effect may depend on its ability to enhance GABA receptor activity. We have studied the effect of pretreatment with CMZ on motor function recovery and nerve cell damage after spinal cord injury (SCI). Rats were randomized and 30 min before SCI they received a single intraperitoneal dose of CMZ (150 mg/kg) or saline. The spinal cord was injured with a 50 g (4.5 g/mm2) load, applied over the exposed dura, through a curved rectangular plate (2.2 x 5.0 mm) for 5 min at T8-9. The animals became paraplegic 1 day after injury. The rats were evaluated for recovery of hind limb motor function. All animals recovered to some extent over the observation period of 12 weeks. However, hind limb motor function was significantly better in the animals pretreated with CMZ. At 12 weeks the rats were killed and perfused/fixed for morphological investigations. Microtubule-associated protein 2 (MAP2) immunostaining was used to stain neurons and dendrites and Luxol-fast blue to stain myelinated tracts of the white matter. The injured segment of the spinal cord showed severe atrophy, distortion, cavitation and necrosis of grey and white matter. Compared to uninjured controls the transverse sectional area was reduced to 32.7 +/- 4% in untreated animals but only to 38.5% +/- 4.1 in CMZ-treated animals. MAP2 staining showed that, compared to uninjured controls, grey matter was reduced to 7.4 +/- 2.7% in saline-treated injured animals and to 22.7 +/- 5.4% in CMZ-treated rats. Our results thus show that in this model CMZ improves hind limb motor function and attenuates the morphological damage to the spinal cord.

Study design: Functional outcome was evaluated following experimental compression-type spinal cord injury (SCI) in wild-type mice and knockout mice, lacking the inducible nitric oxide synthase (iNOS) gene. Objectives: To evaluate the role of the nitric oxide generating enzyme iNOS in SCI. Methods: The experimental animals were subjected to an extradural compression of the thoracic spinal cord. Functional outcome was studied during the first 2 weeks post-injury using a scoring system for assessment of hind limb motor function. Results: Injury resulted in initial paraplegia followed by gradual improvement of motor function in most cases. Mice lacking the iNOS gene (iNOSÀ/À) clearly tended to have a better functional outcome than wild-type mice. The difference was significant on day 14 after injury. Conclusion: In accordance with a few earlier experimental studies, showing beneficial effects of pharmacological iNOS inhibition, the present report would indicate a destructive influence of iNOS following spinal cord trauma.

A study was made on the expression of the intercellular adhesion molecule 1 (ICAM-1) in cerebral microvessels after cortical contusion trauma of the rat brain. The trauma was produced by a free-falling weight on the exposed dura of one fronto-parietal lobe. Immunohistochemistry was done on cryostat sections using a monoclonal antibody and the reaction product was visualized using the avidin-biotin-peroxidase complex method. Control and sham-operated rats showed immunostaining of some penetrating arteries of the cerebral cortex, the epithelial cells of the choroid plexus and occasional microvessels of the brain parenchyma. The same pattern of immunostaining was seen in rats that were subjected to trauma and killed after 30 min. All rats with contusion trauma that were allowed to survive for 6-72 h showed a substantial increase in the number of immunostained capillaries throughout the site of the lesion. The ipsilateral hippocampus showed a mild to moderate increase in the number of immunostained microvascular profiles. This phenomenon was also present in the lateral thalamus of some rats. The staining was seen as an uninterrupted line at the position of the endothelial cells, indicating an upregulation of this adhesion molecule after brain trauma. Up-regulation of ICAM-1 is a well-known phenomenon in inflammatory and ischemic lesions of the brain but has not previously been described in detail in traumatic brain injury. ICAM-1 may be involved in the production of several post-traumatic events such as leukocyte adhesion, microcirculatory disturbances and edema formation.

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Adhesion molecule-mediated adhesion and extravasation of leukocytes may constitute a mechanism of secondary tissue damage following spinal cord injury (SCI). The objective of the present study was to determine to what extent genetic deficiency in the adhesion molecule ICAM-1 influences functional and histopathological measures of outcome following SCI. ICAM-1-/- (n = 11) and wild-type (n = 9) mice were subjected to a compression-type SCI. Assessment of hind-limb motor function was done on days 1, 2, 4, 7, 10, and 14 after injury, using a motor function scoring system. Injury resulted in a drastically impaired hind limb motor function at day one after injury followed by a partial recovery during the observation period. No significant functional differences were found between the experimental groups at any time-point. Fourteen days after injury the animals were sacrificed and the spinal cords were processed for histopathological and immunohistochemical evaluation. Luxol-stained, MAP2-, GFAP- and iba-1-immunostained cross-sectional areas were quantitated using a computerized image analysis system to investigate white matter damage, neuronal loss, astrocytic response and microglial activation respectively. None of these parameters differed significantly between the groups. Separate experiments revealed that the early (24 h postinjury) infiltration of polymorphonuclear leukocytes was significantly reduced in white matter but not in the grey matter of ICAM-1-/- mice, compared to injured controls. In summary, these results do not support the concept that ICAM-1 alone mediates secondary tissue damage following traumatic SCI in the mouse.