人脑挫裂伤后神经细胞凋亡及其与caspase-3基因表达的关系

Apoptosis of neural cells and its relationship with expression of caspase-3 gene after acute traumatic brain injury

目的观察人脑急性创伤性脑损伤后神经细胞凋亡及其与caspase-3基因表达的关系。方法术中采集12例重型颅脑损伤患者挫裂伤脑组织和脑叶切除组织标本15个,分别应用凋亡细胞原位末端标记法(TUNEL)、DNA琼脂糖凝胶电泳、透射电镜等检测神经细胞凋亡的变化,采用免疫组织化学技术检测caspase-3蛋白的表达,并借助caspase-3荧光分析试剂盒检测caspase-3蛋白酶活性的变化。结果12个挫裂伤脑组织标本中10个透射电镜发现有散在分布、数量不等的凋亡神经细胞；TUNEL染色11个出现阳性凋亡神经细胞；5个DNA出现凋亡特异性的“梯状”电泳带；9个出现caspase-3蛋白超表达；11个caspase-3蛋白酶活性明显增高；凋亡出现时间从伤后4h直到216h,高峰在23～72h。结论急性创伤性脑损伤患者存在神经细胞凋亡,并且具有时间和空问依赖性。Caspase-3的超表达与激活参与了人脑创伤性脑损伤后的细胞凋亡过程。

Objective To observe the apoptosis of neural cells and its relationship with expression of caspase-3 gene in brain tissues of patients following acute severe traumatic brain injury （TBI）. Methods Twelve contusion brain tissue samples （CBTS） and three lobectomic brain tissue samples were extracted from 12 patients with severe TBI during emergency craniotomy for detecting apoptotic neural cells by TUNEL, DNA gel electrophoresis and transmission electron microscopy （TEM）, observing apoptosis-related gene caspase-3 possible expression by immunohistochemistry and detecting caspase-3 activity by caspase-3 fluorescent assay kit. Results TEM examination showed diffuse distribution of apoptotic neural cells including neurons, astrocytes and oligodendrocytes in 10 CBTS extracted from 12 patients. In 11 CBTS, TUNEL-positive cells were present in both the grey and white matter of the contusion. Gel electropboresis of DNA extracted from five CBTS presented typical ＂ladder＂ break. Caspase-3-positive immunostaining were found in nine CBTS. Caspase -3 activity was detected in 11CBTS with high value. Apoptotic neural cells presented at 4-216 hours post injury and peaked at 23-72 hours. Conclusion Apoptosis exists in patients with TBI, with time- and space-dependent fashion, Over expression and activation of Caspase-3 contribute to cell apoptosis after TBI.