急性马尾神经压迫后脊髓前角运动神经元细胞凋亡相关蛋白的表达规律

Expression of apoptosis-related protein in motor neurons of anterior horn of the spinal cord after acute cauda equina compression

背景:马尾综合征往往遗留有会阴区皮肤感觉减退,尿便控制功能降低,甚至出现男性功能受损。实验研究发现外周神经纤维损伤后,神经元胞体会发生凋亡,这与损伤的性质、神经元种类、动物种属、年龄、与神经元距离等因素有关。目的:探讨急性马尾神经压迫后脊髓前角运动神经元细胞凋亡及相关蛋白的表达规律。方法:27只家犬随机分为3组,压迫组及对照组制备马尾神经压迫模型,正常组不造模。压迫组又分为水囊压迫持续4,8,12,24,48,72,168 h组,每组3只;对照组只置入水囊,不注水。采用TUNEL标记法检测脊髓前角运动神经元凋亡情况,免疫组化SABC法检测Bax、Bcl-2、Caspase-3蛋白的表达,用Qwin550Cw型图像采集与分析系统检测Bax、Bcl-2、Caspase-3蛋白表达阳性细胞灰度值。结果与结论:急性马尾神经压迫后脊髓前角运动神经元细胞发生凋亡,压迫12 h即有阳性细胞检出,72 h神经元凋亡达到高峰。Bax、Bcl-2蛋白在正常组即有少量表达,Caspase-3蛋白在正常组、对照组无表达。Bax、Bcl-2蛋白表达于压迫持续8 h开始显著增加,72 h达高峰,168 h降至正常;Bax蛋白表达增幅较Bcl-2大。Caspase-3蛋白于压迫持续12 h开始表达,72 h达高峰,168 h仍有少量表达;Bax、Caspase-3蛋白表达在72 h达高峰,Bcl-2蛋白表达则无明显上升。提示急性马尾神经压迫后脊髓前角运动神经元发生凋亡;Bax、Bcl-2蛋白表达呈相互拮抗作用,在Bax/Bcl-2复合体中,Bax蛋白占优势时促进凋亡,诱发Caspase-3蛋白表达,导致神经元凋亡发生。

BACKGROUND： Cauda equina syndrome often induces skin hypoesthesia in the perineal area, poor urine-stool control, and impairs male function. After peripheral nerve fiber injury, apoptosis of neurons appeared. This is associated with the nature of the injury, the types of neurons, the species of animals, the age, and the distance between neurons. OBJECTIVE： To explore the motor neuron apoptosis and expression of apoptosis-associated protein in the anterior horn of the spinal cord after acute cauda equina compression. METHODS： A total of 27 canines were randomly divided into three groups. In the compression and control groups, models of cauda equina compression were established. In the normal group, no models were established. Compression group received water sac compression for 4, 8, 12, 24, 48, 72 and 168 hours, with three models in each group. In the control group, only water sac was implanted, but water was not injected. Terminal deoxynucleotidyl transferase Td T-mediated biotin d UTP nick end-labeling assay was used to detect the apoptosis of neurons in the anterior horn of the spinal cord. Bcl-2, Bax and Caspase-3 protein expressions were measured by immunohistochemical staining（strept avidin-biotin complex）. Gray values of positive cells of Bax, Bcl-2 and Caspase-3 protein expressions were detected using Qwin550 Cw image collection and analysis system. RESULTS AND CONCLUSION： The apoptosis of motor neuron occurred in the compression groups. At 12 hours of compression, positive cells were detected, and the number of positive cells reached a peak at 72 hours. Bax and Bcl-2 protein expression was small in the normal group. Caspase-3 protein expression was not detected in the normal and control groups. Bax and Bcl-2 protein expression was significantly increased at 8 hours, peaked at 72 hours and reduced to a normal level at 168 hours. The increased range of Bax protein expression was bigger than that of Bcl-2. Caspase-3 protein began to express at 12 hours, peaked at 72 hours and reduced to a low level at 168 hours. Bax and Caspase-3 protein expression peaked at 72 hours, and Bcl-2 protein expression was not obviously increased. These findings verified that after acute cauda equina compression, the apoptosis of neurons occurred in the anterior horn of the spinal cord. Bax and Bcl-2 protein expression showed an antagonistic action. In the Bax/Bcl-2 complex, Bax protein in a high expression promoted apoptosis, induced Caspase-3 protein expression, and neuronal apoptosis.