| Spinal cord injury (SCI) has a high incidence and a bad prognosis. It causes serious physical and psychological hurt on the patients and brings a heavy burden on their family members and the society. Proper therapeutic strategies are urgently needed to improve the life quality of the victims of SCI and to relieve their family and the society of the burden. As we know, primary SCI is an un-reversible process, and secondary SCI is the main factor causing the expansion of pathological changes in myeloid tissue, aggravating cellula nervosa depletion and finally affecting function prognosis. So how to inhibit or reverse the secondary injury has become a focus in the SCI reasearch field. Previous studies showed that secondary SCI mainly involves the demolition of blood-spinal cord barrier (BSCB), the development of inflammatory reaction and apoptosis. Ginkgo biliba extract (EGb761), the international reference standard extractive of ginkgo leaf, which consists of terpenoid(6%) and flavonoid(24%), has been proven to possess stronger antioxidative activity and neuroprotective effect. It has been widely used in the therapy of ischemic brain injury and degenerative disease of CNS. But there are few relevant reports on its effect of SCI. In the present study, the EGb761 was used as an intervention in the acute spinal cord injury in a rat model and its protective effect on nervous tissue injury was observed. Additionally, the permeability of BSCB, the express of inflammatory factor and the cavitation in the damaged region at the advanced stage were also observed with the purpose of revealing the possible mechanism of its protective effect and identifying its therapeutical effect on SCI.Objective: (1)To observe the protective effect of the extract of Ginkgo biloba (EGb761) on the blood-spinal cord barrier after spinal cord injury in rats and study its mechanism.(2)To observe the characteristics of pathological change and apoptotic in myeloid tissue in different stages after ASCI in rat model, to survey the effect of EGb761 on apoptosis of cellula nervosa in earlier period after ASCI, and to survey the effect of EGb761 on the cavitation in the damaged region and the motor function recovery at the advanced stage, in conclusion to explore the possible mechanism of its protective effects after SCI in rats.Methods: In the present study, the experiment has two steps. In the step one, 120 SD female rats were randomly divided into three groups, including group A (sham-operated group), group B (Allen impact SCI group) and group C (EGb761 treated group). Each group had 40 rats for test. Rats in group A underwent laminectomy alone. The modified Allen method was adopted at the T9 level in groups B and C to induce the acute spinal cord injury.For group C, EGb761 was delivered 100mg/kg/d intraperitoneally half an hour after injury for the first time and per day until killed. At the same time, vehicle-treated animals received physiological saline in the same volumes as in the EGb761-treated group. The rats were sacrificed at 3, 6, 24 and 72h post-injury and T9 spinal cord specimen was extracted. The blood-spinal cord barrier (BSCB) of cords was determined by Evans blue content detection. The level of interleukin-1β(IL-1β) in the T9 spinal cord was detected by ELISA method while the protein level of intercellular cell adhesion molecule-1 (ICAM-1) in T9 spinal cord were detected by immunohistochemistry assay.(2)Another sixty SD female rats were randomly divided into three groups, including group A (sham-operated group), group B (Allen impact SCI group) and group C (EGb761 treated group). Each group had 20 rats for test. Before sacrificed, BBB assessment and footprint analysis were performed on each rat in different group at 1, 3, 7 and 14d post-injury and then T9 spinal cord specimen was extracted. Then sections were made for HE staining and histological examination to analysis and determine the quantity of the neuron around the injury area and the extent of cavitation in the damaged region. TUNEL positive cell expression and caspase-3 protein in spinal cord were detected by immunohistochemistry assay.Results: (1)In group A, there was no obvious leakage of Evans blue around the injuried site and the concentrations of IL-1βand ICAM-1 were maintained at a low level. Compared to group A, the leakage of Evans blue and the concentrations of IL-1βand ICAM-1 were significantly higher in group B. After application of EGb761, the leakage of Evans blue, as well as the concentrations of IL-1βand ICAM-1, were decreased significantly at all time points except 3 hours post-injury in group C , when compared to group B. (2) In group A, there was no obvious difference compared with normal rats. For the comparsion between group B and C, there is no significant difference in ethology assessment on 1, 3 and7 d. However, on the 14th day, the BBB score in the Group C was significantly higher (15.40±0.89 vs 13.60±0.55, P <0.05) than that in the group B. For the footprint analysis on the 14th day after surgery, the step stride length in the C group was significantly longer (10.44±0.26 vs 9.81±0.33cm, P <0.05) than that in the group B. The step stride width in the C Group was also significantly smaller (4.52±0.14 vs 5.58±0.11cm, P <0.05) than that in the group B. Histological observation showed that the cavitation area of the damaged zone in the group C was significantly less than that in the group B(2.92±0.18 vs 3.63±0.27 mmP2P, P <0.05).(3)Accroding to the histological analysis, neuronal apoptosis was induced by the acute spinal cord injury, and the quantity of apoptotic cells in gray matter reached the peak amplitude at 24h after after injury while those in white matter reached its peak on the 7th day after surgery. What's more, the majority of TUNEL positive cells were glial cells. And on the 7th day after surgery, the quantity of neuron in the Group C was significantly higher (14.04±1.80 vs 7.44±1.15, P <0.05) than that in the group B, and its apoptosis cells counting was obviously less than that in the group B(58.12±2.96 vs 69.52±4.74, P <0.05), which is coincidence with the tendency of Caspase-3 positive cell's disposition and quantity diversity.Conclusion: (1)In the Allen's acute SCI rat model, EGb761 could decrease the local concentration of IL-1β, suppress the abnormal expression of ICAM-1 protein and attenuate disruption of BSCB permeability.(2)Neuronal apoptosis definitely existed in the spinal cord injury. The apoptosis cells were mainly glial cells and have there particular characteristics of spatial and temporal distribution as well as the pathological change in myeloid tissue. (3) EGb761 has the protective effects on nervous tissues injury after ASCI. And the possible mechanisms of its protective effects were lessen myeloid tissue pathological change in the early stage after SCI, restrain neurocyte apoptosis at the region near the traumatic site, decrease the area of necrosis at the injured portion, finally improve motor function of hind limb in rats. |
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