| ObjectiveAcute spinal cord injury is a common injury in surgery, the consequence of which is severe and difficult to restore. Spinal cord injury includes primary damage and secondary damage. The secondary damage is more harmful than the primary damage, which is an important pathological process that influences the repair of spinal cord. The course of spinal cord injury is very complicated, as it includes a series of biochemical reactions like inflammation, free radical accumulation, apoptosis accelerating etc. Changes in internal and external environment of cells will lead to the activation of various enzymes, which can cause further damage to the organization. The nerve cell death after acute spinal cord injury includes necrosis and apoptosis. The purpose of the experiment is to study the expression of Caspase-3, AIF, Cathepsin B and to explore the significance of their expression initially by immunohistochemical study after acute spinal cord injury in rats.MethodsHealthy adult SD rats (78), regardless of male or female, weighing 220-250 g, provided by the Experimental Animal Center of China Medical University. The rats were randomized into 3 groups. Injury group:detected after spinal cord injury at six time points of 2 h,8 h,1 d,3 d,5 d,7 d, and 6 rats at each time point; sham operated group:detected after simple laminectomy at six time points of 2 h,8 h,1 d,3 d,5 d,7 d, and 6 rats at each time point; control group:detected with no treatment at six time points of 2 h,8 h,1 d,3 d,5 d,7 d, and 6 rats at each time point. HE methods were used to detect the pathologic change of spinal cord. The expression of Caspase-3, AIF and Cathepsin B were detected by immunohistochemical study. Besides, using the terminal deoxynucleotidyl transferase mediated DUTP nick and labeling (TUNEL) methods to detect the level of the apoptosis of neural cells. Statistical analysis was done by using SPSS 11.5. Data are expressed as mean±SEM. Cathepsin B, Caspase-3 positive cells and apoptotic cells was used to compare each group by ANOV. With P <0.05 as there are differences, P<0.01 for significant differences.ResultsObserved under light microscopy, normal and sham-operated group presented the rule morphology neurons and dense white matter.2 h after injury, the gray matter of damage segment appeared apparent hemorrhage.8 h after injury, compared to 2 h, hemorrhage was reduced while inflammatory cells began to appear. Spinal cord was damaged severely with loose irregular white matter from 1 to 3 d. The damage region expanded at 3 d with a large number of inflammatory cell infiltration within the damage zone. The damage region did not change significantly from 5 d to 7 d, which still with a large number of inflammatory cell infiltration. There were few expressions of Caspase-3, AIF, Cathepsin B and TUNEL positive cells in normal and sham operated spinal cord as detected by immunohistochemical study. The number of positive cells of Cathepsin B was increased clearly at 3 d, which reached a peak at 5 d and was constant at 7 d after injury. The number of positive cells of AIF was increased clearly at 1 d and then reached a peak, but it was rarely seen at 5 d after injury. The number of Caspase-3 was increased obviously at 8 h, got to a peak at 3 d and was lowest at 7 d. And the number of positive cells of TUNEL was also increased obviously at 8 h, got to a peak at 3 d, and was lowest at 7 d. There were significance differences of morphological and position of positive cells between Cathepsin B and Caspase-3 or TUNEL.ConclusionsThere is neural apoptosis after acute spinal cord injury, and AIF, Caspase-3 may involve in the regulation of the apoptosis of neural cells after acute spinal cord injury. After acute spinal cord injury, Cathepsin B may involve in the secondary injury by inflammatory cells, which is different from the role in brain injury. The experimental provide some basis to further understand the mechanism of the spinal cord injury.
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