Caspase-3 Expression And Its Relationships With Neuronal Apoptosis In Spinal Cord After Hemitransection Injury In Rats

Objective: Apoptosis is one of principal ways for secondary lesion of spina1 cord after injury. The objectives of this study were to establish hemitransecti on of spinal cord injury (hSCI) model of rats, to investigate the changes in e-xpression of cystein-dependent aspartate-specific protease-3 (caspase-3) in n-eurons of ventral horn following hSCI and its relationships with the apoptosis, to explore the nerve repair function acted by caspase-3,and finally to offer the data for inhibitting cell apoptosis and accelerating the recovery of spinal cord after injury.Method: Forty five adult healthy Wistar rats were randomly divided into 9groups(5 each group): sham-operated control group and other 8 groups 6 hours, 1 day, 4 days, 7 days, 14 days, 21 days, 28 days and 35 days after spinal cord injury. The operation was performed on the rats and their thoracic cord were hemitransectioned at T8.The animals were sacrificed and perfused at diffferent time points after operation. The spinal cord of each rat was taken out and two blocks were kept. One block was the rostral part 0-1 cm rostral and another was the caudal part 0-1 cm caudal to the lesioned point. The two blocks were sectioned in a cryostat. The anti-caspase-3 immunohistochemistry SP method and TUNEL staining were performed on these sections. We respectively observed positive caspase-3 immunoreactive neurons,and TUNEL positive neurons. Those two types of positive neurons were also counted in the ventral horn of spinal cord for all groups. The data were analyzed by ANOVA and t test.Result: 1.TUNEL staining: TUNEL positive neurons were firstly observedin 6h group,lasting at least 5 weeks. The TUNEL positive neurons in lessioned groups increased in number by contrast of sham-operational group. The more positive neurons in the rostral part on lesioned side were observed during day 4 to day 35, had a peak at day 4.The differences were all significant. The number of the positive cells in the caudal part on lesioned side increased significantly at hour 6, day 1, day 4, day 7 and day 14 group, with a peak at day 1.2.Anti-caspase-3 immunohistochemistry: The positive caspase-3 immunoreactive products were mainly localized in cytoplasm and neural processes of the neurons,sometimes in their nuclei. For the spinal cord injury groups, the positive neurons all increased in number in both rostral and caudal parts on lesioned side,comparing with sham-operational group. And in the rostral part on lesioned side,all increases at different time points were significant except the day 35 group. The number of the positive neurons increased significantly in the caudal part on lesioned side at all time pointsexcept the day 21 group.Conclusion: For each lesioned group after hSCI in rats,TUNEL positive neurons and positive caspase-3 immunoreactive neurons were observed in the ventral horn of the rostral and caudal parts of the spinal cord and the number of both positive neurons were increased, comparing with sham-operational group. The change of caspase-3 immunoreactivity and TUNEL positive cell number were almost consistent. It indicates caspase-3 may involve in neuronal apoptosis in spinal cord following hSCI, and suggests that 24 hours following SCI is the optimal therapeutic window.