The Effect Of Cyclosporin A On The Neuroinflammation In The Early Period Of Rat Spinal Cord Injury

Objective:To investigate the role and possible mechanism of cyclosporine A on the neuroinflammation in the early period of rat spinal cord injury.Methods:Total of 78 adult male Sprague-Dawley(SD) rats weighing 220 to 260 g were used in this experiment. The 15 rats among all the animals were randomly assigned to sham-operated rats(sham group), Cs A-treated SCI rats(Cs A group), and normal saline-treated SCI rats(NS group) for Basso, Beattie, and Bresnahan(BBB) score and H&E staining 28 d post SCI(n=5). The 20 rats among all the animals were randomly assigned to sham group and NS group for H&E staining. Sham group and SCI group were then allocated to 2 subgroups according to different time-point(6 h, 1 d), respectively(n=5, respectively). The remaining 60 rats were randomly divided to sham group, Cs A group, and NS group for q RT-PCR and immunohistochemistry, respectively. The three groups were then allocated to 2 subgroups according to different time-point(6 h, 1 d), respectively(n=5, respectively).A modified Allen's weight drop method was applied to induce a rat moderate contusion injury model. BBB score was quantified to evaluate locomotor ability of SCI rats, and the H&E staining was used to assess spinal cord histopathology by the quantification of lesioned area within spinal cord. Additionally, q RT-PCR and immunohistochemistry were used to determine pro-inflammatory mediator gene expression.The data were expressed as mean ± SD, except for data from the behavioral assessment of BBB scores, which is presented as mean ± SEM. Statistical analyses were performed using SPSS 13.0(SPSS Inc., Chicago, IL, USA). Data from the BBB scores were analyzed using repeated measures ANOVA. For other data, statistical comparisons were analyzed using a one-way ANOVA followed by a SNK test. A P-value less than 0.05 was considered to be statistically significant.Results:1. Behavioral assessment: BBB scores in sham group maintained at 21 points, indicating that hindlimb movement of sham group was normal(p>0.05). During the observation period, as illustrated by increased BBB scores, hindlimb locomotor activity gradually improved in Cs A group and NS group(p<0.01). In addition, compared with NS group, BBB scores from Cs A group significantly increased since 14 days after SCI(p<0.01,14d?21d?28d;p>0.05,1d?3d?7d). These results indicated that Cs A treatment is beneficial for the motor function recovery of SCI rats.2. Histopathological evaluation: 6 hours post-SCI, the spinal cord transverse sections from SCI group showed significant structure destroy, hemorrhage, edema formation, and inflammatory cell infiltration. Similarly, sections from SCI group 1 d post-SCI showed parallel pathologic changes. 28 days post-injury, the spinal cord transverse sections from sham group showed a compact structure, regular morphology. However, transverse sections from Cs A group and NC group revealed obvious various size cystic cavities, and glia scar formation surround cavity. More importantly, compared with NC group, the lesioned volume of epicenter from Cs A group significantly decreased(p<0.01).3. The expression of pro-inflammatory cytokines: 6 hours post-injury, the m RNA expression of IL-1?, IL-6, and TNF-? in Cs A group and NC group significantly increased when compared with sham group(p<0.01). But, the m RNA level of IL-1?, IL-6, and TNF-? was significantly reduced by Cs A treatment in comparison to NS group(p<0.01). Similarly, data from 1 d post-SCI showed parallel expression tendency. Immunohistochemistry data 6 hours post-injury showed that IL-1?, IL-6, and TNF-? were mainly localized in the cytoplasm of gray matter neurons and glial cells. The percentage of positive cells in Cs A group demonstrated a significant decrease in comparison to NS group(p<0.01). Similarly, data from 1 d post-SCI showed parallel expression tendency.Conclusions:1. Cs A significantly improved the motor function recovery of SCI rats.2. Cs A significantly alleviated histopathological progress.3. Cs A significantly reduced m RNA and protein expression of IL-1?, IL-6, and TNF-? in injured spinal cord.4. Cs A may attenuate spinal cord injury via neuroinflammation reduction.