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目的研究大鼠脑外伤后溶酶体酶cathepsin-B和-D是否被激活及其不同时段表达变化,阐述其与凋亡执行因子caspase-3表达的关系,并探讨对脑外伤诊断及形成时间的意义。方法采用自由落体打击法建立脑外伤动物模型,并对模型及对照样本进行免疫荧光、双标和激光共聚焦检测,结果用SPSS10.0软件处理。结果脑外伤后1hcathepsin-B表达即增加,4～8d达高峰,脑外伤后32d仍处于高表达水平;cathepsin-D的表达于脑外伤后12h增加,4～8d达高峰,32d的表达仍然高于12h的表达水平。脑外伤初期,cathepsin-B和-D阳性细胞与caspase-3阳性细胞重叠较少,脑外伤后6h开始增加,32d仍然有很多阳性细胞重叠。结论脑外伤后cathepsin-B和-D被激活,其激活在脑外伤早期可能抑制细胞凋亡执行因子caspase-3的激活,之后(6h后)则与caspase-3起协同作用,共同促进细胞死亡;cathepsin-B和-D表达的时程变化对于脑外伤的法医学诊断和中晚期的时间推断有参考意义。
Objective To investigate whether the lysosomal enzymes cathepsin-B and -D are activated and expressed at different time points after traumatic brain injury in rats, and to elucidate the relationship between cathepsin-B and -D and the expression of caspase-3, an inhibitor of brain injury. Meaning. Methods The animal models of traumatic brain injury were established by free-fall strike method. Immunofluorescence, double-labeled and confocal laser scanning confocal microscope were used to detect the model and control samples. The results were processed by SPSS 10.0 software. Results The expression of cathepsin-B increased 1 h after TBI and peaked at 4 to 8 days. The expression of cathepsin-D was still high at 32 days after traumatic brain injury. The expression of cathepsin-D increased at 12 hours and reached the peak at 4 to 8 days, and remained high at 32 days The expression level at 12h. In the early stage of traumatic brain injury, cathepsin-B and -D-positive cells overlap with caspase-3-positive cells less, and they began to increase at 6h after traumatic brain injury. There were still many positive cells overlapping at 32d. Conclusions Catsps-B and -D are activated after traumatic brain injury, and their activation may inhibit the activation of caspase-3 in early stage of traumatic brain injury. Afterwards, they may cooperate with caspase-3 after 6h to promote cell death ; The time-course changes of cathepsin-B and -D expression have reference significance for the forensic diagnosis of brain trauma and the inference of middle and late stages.