| Objective: To study the effect of Cyclosporin A(CsA) on the expression of Bc1-2, Bax and AQP-4 after acute spinal cord injury (ASCI), and investigate the possible mechanism of Cyclosporin A on inhibiting nerve cell edema and necrosis after ASCI.Methods:1. One hundred and twenty six SD rats, weight 200-250g, were randomly divided into three groups: laminectomy only group (group A, n=42), ASCI group (group B, n=42), and Cyclosporin A treated group after ASCI (group C, n=42).2. Group B and C received acute spinal cord injury using modified Allen's method, and group C received intraperitoneal injection of 2.5 mg/kg Cyclosporin A immediately and every 12h for the following 3 days after the operation. Normal saline of the same dose were applied to group B (control group) and group A. Rats in each group were further randomly divided into 7 sub-group, i.e. 12h, 24h, 3d, 7d, 14d, 21d, and 28d after surgery.3. The hindlimb locomotor function of every group were evaluated by BBB score at different time points after ASCI; and histologic morphology changes were observed by hematoxylin and eosin (HE) staining.4. The expression of Bcl-2, Bax and AQP-4 were detected by immunohisto- chemistry and compared among different experimental groups.5. The gene transcription changes of Bcl-2 and Bax in different groups at different time points were detected by RT-PCR. 6. The expressions of AQP-4 protein in different groups and different time points were detected by Western Blot.Results:1. The rat SCI model is successfully established. The spinal cord injury and animal death rate met the experimental requirements.2. Twelve and one day after SCI, no significant differences of BBB scores were found between group B and C. Three days after SCI, BBB score in group C is obviously higher than that of group B, which has statistically significance. However, the BBB score of both group B and C were significantly lower than that of group A. The HE staining revealed severe edema of tissues and cells in group B which lead to scar formation at later stage,while less edema in group C and none in group A.3. The positive expression of AQP-4, Bcl-2 and Bax Protein detected by immunohistochemistry were shown in all groups at different time points after SCI. After ASCI, the expression of AQP-4, Bcl-2 and Bax Protein in the injury areas increased. The expressions of Bcl-2 and Bax were increased 1day after SCI in group B and C, reach the peak at 7d, and still significant higher than that in group A at 14d after SCI. The expression of AQP-4 in group B and C increased 12 hours after injury and reached the peak at 3d after SCI, and still much higher than that in group A at 7d. The expressions of Bcl-2 in group C were significant higher than that in group B, while the expressions of Bax and AQP-4 in Group B were significant higher than that in group C at different time points. All the differences were statistically significant.4. RT-PCR results. The mRNA levels of Bcl-2 and Bax in group A almost the same in different time points after SCI. The mRNA levels of Bcl-2 and Bax in group B and C increased at 1d after SCI which were much higher than that of group A and climb up continuously to the peak at 7d, and still higher than group A at 14d with significant difference. Bcl-2 mRNA level in group B is always lower than in group C at different time points, while the Bax mRNA level is always higher in group C than in group B with significant difference.5. The Western blot results showed an increase of AQP-4 expression level in group B and C 12 hours after operation and climbs up to the peak at 3d after SCI, and still higher than that in group A at 7d with significant difference. The AQP-4 protein expressions in group B were statistically significant higher than in group C at different time points. AQP-4 expressions in group A always the same at each time point and statistically significant lower than the other two groups.Conclusion:Cyclosporine A can inhabit the expression of AQP-4 in SD rats'spinal cord after ASCI, promote Bcl-2 to express, and inhabit Bax to express, and raise the Bcl-2/Bax ratio, so as to reduce edema, necrosis and apoptosis of injured spinal.
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