Israel Grijalva

Cyclosporin-A inhibits lipid peroxidation after spinal cord injury in rats

Besides its immunosuppressive/anti-inflammatory activity, cyclosporin-A (CsA) may protect damaged tissues from lipid peroxidation (LP) by free radicals. To determine the effect of CsA on LP spinal cord (SC) injury, Wistar rats were treated with either vehicle or CsA (2.5 mg/kg per 12 h i.p.) 1, 2, 6 or 12 h after SC trauma by T8-T9 spinal cord contusion, analyzing LP 24 h after injury at the lesion site by the lipid fluorescent products formation method. CsA significantly diminished LP to levels below control values after contusion (P < 0.05). The greater inhibition was observed when CsA was given during the first 6 h after injury, furthermore, animals showed a significant clinical improvement. Results show that CsA may be beneficial to injured tissue by inhibiting the levels of LP.

Development of post-traumatic cysts in the spinal cord of rats-subjected to severe spinal cord contusion.

To study the development of post-traumatic spinal cord (SC) cysts, and their fine anatomic characteristics, rats were subjected to severe SC contusion. Specimens were analyzed from day 1 to 1 year post-injury. Using conventional light, and transmission and scanning electron microscopy, three stages were typified, namely: necrosis, repair, and stability. The final cell composition and thickness of the cyst walls were not uniform. Astrocytes, fibroblasts, ependymal cells, and collagen fibers were the main constituents. Chronic inflammatory cells were also observed. The neuropathologic characterization of posttraumatic SC cysts could be useful in planning strategies for SC reconstruction at different times post-injury.

Effects of cyclosporin-A on immune response, tissue protection and motor function of rats subjected to spinal cord injury

The aim of this work was to test the effect of cyclosporin-A (CsA) on some immunological, morphological and functional aspects developed after spinal cord injury. The specific cellular immune response against spinal cord constituents, the amount of spared tissue and myelination at the site of injury, and the motor function outcome were assessed in a first series of experiments. Rats were subjected to spinal cord compression and treated with cyclosporin-A before lesion and during the entire study. A specific lymphocyte response against spinal cord antigens was found in untreated spinal cord injured rats but not in cyclosporine-A treated injured rats. A significantly better myelination index was also found in injured cyclosporin-A-treated rats, as compared to untreated animals. The amount of spared spinal cord tissue at the epicenter was not significantly different comparing CsA-treated with vehicle-treated rats. Looking for a potential therapeutic use of CsA, in a second series of experiments, rats were subjected to spinal cord contusion and treated with cyclosporin-A from 1 to 72 h after lesion. Motor recovery and red nuclei neurons survival, were evaluated, and found to be significantly better in spinal cord injured rats treated with cyclosporin-A than in injured-untreated rats. This work confirms the existence of an autoimmune cellular reaction after injury that can be inhibited by cyclosporin-A treatment. Furthermore, cyclosporin-A promotes neuroprotection by diminishing both demyelination and neuronal cell death, resulting in a better motor outcome after spinal cord injury.

Lipid peroxidation inhibition in spinal cord injury: cyclosporin-A vs methylprednisolone.

To compare the effectiveness of cyclosporin-A (CsA) with methylprednisolone (MP) or a combination of both upon inhibition of lipid peroxidation (LP) after spinal cord (SC) injury, rats were treated with either CsA, MP, CSA+MP or vehicle starting 1 h after SC contusion at T9 level. LP was assessed 24 h after injury by the lipid fluorescent product formation method. The survival rate was also evaluated in other series of rats by the Kaplan–Meier curves. Lipid peroxidation was similarly inhibited in rats treated with CsA, MP, or CSA+MP (p >0.05). Animals receiving MP (alone or combined with CsA) showed the poorest surviving rate. LP was inhibited by CsA to the same extent as by MP but without the lethal effect of the latter.

Spontaneous long-term remyelination after traumatic spinal cord injury in rats.

The capability of the central nervous system to remyelinate axons after a lesion has been well documented, even though it had been described as an abortive and incomplete process. At present there are no long-term morphometric studies to assess the spinal cord (S.C.) remyelinative capability. With the purpose to understand this phenomenon better, the S.C. of seven lesionless rats and the S.C. of 21 rats subjected to a severe weight-drop contusion injury were evaluated at 1, 2, 4, 6, and 12 months after injury. The axonal diameter and the myelination index (MI = axolemmal perimeter divided by myelinated fiber perimeter) were registered in the outer rim of the cord at T9 SC level using a transmission electron microscope and a digitizing computer system. The average myelinated fiber loss was 95.1%. One month after the SC, 64% of the surviving fibers were demyelinated while 12 months later, only 30% of the fibers had no myelin sheath. The MI in the control group was 0.72 +/- 0.07 (X +/- S.D.). In the experimental groups, the greatest demyelination was observed two months after the lesion (MI = 0.90 +/- 0.03), while the greatest myelination was observed 12 months after the injury (MI = 0.83 +/- 0.02). There was a statistical difference (p < 0.02) in MI between 2 and 12 months which means that remyelination had taken place. Remyelination was mainly achieved because of Schwann cells. The proportion of small fibers (diameter = 0.5 micron or less) considered as axon collaterals, increased from 18.45% at 1 month to 27.66% a year after the contusion. Results suggest that remyelination is not an abortive phenomenon but in fact a slow process occurring parallel to other tissue plastic phenomena, such as the emission of axon collaterals.

Constitutive and inducible nitric oxide synthase activities after spinal cord contusion in rats

Nitric oxide (NO) plays a role in the secondary damage after spinal cord (SC) injury. NO is produced by the activity of two classes of enzymes: calcium-dependent constitutive nitric oxide synthase (NOS) and calcium-independent inducible NOS. To determine the time course of both NOS activities after SC injury, 50 Wistar rats were submitted to severe SC contusion. NOS activities were assayed at the site of SC injury at several times after lesion. Results showed a significant increase of 138 and 96% in the constitutive NOS activity at 4 and 8 h after the lesion, respectively, as compared to sham-operated rats. iNOS activity was increased 72 h after lesion by 103% (P<0.05). In conclusion, both isoforms of NOS increase their activity at different time periods after SC injury.

Efficacy and Safety of 4-Aminopyridine in Patients with Long-Term Spinal Cord Injury: A Randomized, Double-Blind, Placebo-Controlled Trial

Objectives. To study the efficacy and safety of 4‐aminopyridine (4‐AP), and to document sensorimotor changes after discontinuation of the drug in patients with long‐term spinal cord injury.

Neuroprotection of completely lacerated spinal cord of adult rats by homotopic and heterotopic transplantation

To evaluate the neuroprotective effect of transplants placed in the lesion zone after a complete spinal cord (SC) laceration, two independent series of experiments were carried out. In the first, allogeneic or xenogeneic fetal SC was transplanted into the gaps of the damaged lower thoracic SC of adult rats. In the transplanted rats the incidence of life-threatening complications was reduced, and the survival rate was increased compared with the control group (lesion, without implant). Histological examination showed less damage to the neighboring SC parenchyma in the transplanted rats. The measurement of this neuroprotective effect was made in a second series of experiments. Using the same model of SC injury, allogeneic fetal SC, autologous peripheral nerve and/or adipose tissue were implanted. Rats with implants of Gelfoam and damaged rats without implants were the controls. The implanted rats of all groups, including the Gelfoam group, showed a better survival rate than the nonimplanted rats. Significantly less damage to the neighboring SC parenchyma was measured in implanted rats with any of the live tissues tested compared with non-implanted rats, although no significant differences were observed between the Gelfoam group and the nonimplanted rats. Histological evidence of tissue implant survival was observed in all corresponding groups. It is concluded that the transplanted tissues tested here have a neuroprotective effect, possibly by acting as a buffer to neurotoxic substance(s) released by the stumps, and/or by exerting trophic effect(s) on the host.

Spontaneous and induced aberrant sprouting at the site of injury is irrelevant to motor function outcome in rats with spinal cord injury.

In the absence of effective regeneration following spinal cord (SC) injury, sprouting from undamaged axons has been regarded as an underlying factor for functional improvement after incomplete SC injury. The influence of spontaneous and induced axonal sprouting at the injury site on motor function was tested using rats subjected to moderate SC contusion at T9 level, using megadoses of methylprednisolone (MP) and intralesion implantation of cells from sciatic nerve (PNI). Groups using MP and PNI combined, implant vehicle, and injury with no treatment were also included. Amount of sprouting at the injury sites was significantly different depending on treatment. It was abundant in PNI-treated rats, moderate in rats treated with vehicle or nontreated, and limited in rats given MP with or without PNI (chi2, p=0.0084). This sprouting showed an aberrant course and was located in proliferating tissue at the site of injury, characterized by the presence of ependymal cells, macrophages, and myelinating and nonmyelinating Schwann cells. Functional scores and amount of spared white matter were not significantly different among groups. Correlation of the amount of sprouting vs. functional outcome or vs. amount of spared tissue was not significant, while correlation of functional outcome vs. amount of spared tissue was significant (p<0.0001). In conclusion, PNI increase aberrant sprouting at the injury site, while MP limits such sprouting, in either case without impact on motor function outcome. Missing guiding channels for sprouting axons could explain the absence of any functional improvement.

High Doses of 4-Aminopyridine Improve Functionality in Chronic Complete Spinal Cord Injury Patients with MRI Evidence of Cord Continuity

BACKGROUND AND AIMSnMany patients with complete spinal cord injury (SCI) exhibit demyelinated and poorly myelinated nerve fibers traversing the lesion site. Conventional doses of 4-aminopyridine (4-AP, 30 mg/day) have shown to provide no or minor functional improvement in these patients. We undertook this study to test the functional effect of high doses of 4-AP on patients with chronic complete SCI with cord continuity at the site of injury demonstrated by magnetic resonance imaging.nnnMETHODSnFourteen patients were included in a double-blind, randomized, placebo-controlled trial followed by an open label long-term follow-up. Initially, patients received 4-AP or placebo orally, with 4-AP being increased gradually (5 mg/week) to reach 30 mg/day. For long-term treatment, 4-AP was increased 10 mg periodically according to negative electroencephalogram and blood test abnormalities and minor adverse reactions. Pre-treatment, 12 and 24 weeks of the controlled trial, and 6 and 12 months of open trial evaluations, or with the highest doses reached were obtained.nnnRESULTSnThree of 12 patients were able to walk with the assistance of orthopedic devices, 1/12 became incomplete (AIS B), 7/12 improved their somatosensory evoked potentials, 5/12 had sensation and control of bladder and anal sphincters, and 4/9 male patients had psychogenic erection.nnnCONCLUSIONSnPositive changes were seen mainly in patients with cyst (4/5) or atrophy (3/5) of the injury site. Two patients withdrew from the study: one had seizures and one had intolerant adverse reactions. We conclude that high doses of 4-AP in the studied population produced several functional benefits not observed using lower doses.