DAPK1在弥漫性轴索损伤后大鼠脑组织中的表达及其与神经元凋亡的关系

Expression of death-associated protein kinase 1 and its relationship with neuron apoptosis following diffuse axonal injury in rats

目的探讨大鼠弥漫性轴索损伤(diffuse axonal injury,DAI)后神经元凋亡及死亡相关蛋白激酶1(DAPK1)表达变化,为临床治疗提供新的靶点及治疗时间窗。方法 40只SD成年雄性大鼠随机分为对照组(n=8)和DAI组(n=32)。DAI组选用大鼠头颅瞬间旋转装置制备DAI模型,分为6h、24h、72h、7d组共4个亚组(n=8)。各组在造模前及处死前采用改良神经功能缺损评分(mNSS)对大鼠神经功能进行评分;同时将各组大鼠一半用于Western bolt检测大鼠胼胝体、脑干组织内DAPK1表达情况;另一半用于TUNEL检测大鼠脑组织内神经元细胞凋亡情况。结果 DAI后大鼠均出现明显体质量下降、神经功能缺失;DAI后6h组大鼠胼胝体、脑干组织可见DAPK1蛋白表达明显升高,并出现明显神经元细胞凋亡,DAI后24h组DAPK1表达及神经元细胞凋亡均到达高峰值,DAI后72h、7d组表达开始下降,仍明显高于对照组。结论 DAI后出现神经功能缺失、体质量下降,可能与损伤后DAPK1表达及神经元细胞凋亡有关,二者在损伤后24h一致性达到高峰;提示DAPK1抑制剂药物治疗时间窗为伤后24h。

Objective To explore the change of death-associated protein kinase 1 （DAPK1） expression and neuron apoptosis so as to provide a therapeutic time window for diffuse axonal injury in rats. Methods Totally 40 adult male SD rats were randomly divided into control group （n = 8） and DAI groups （n = 32）. The model of DAI was produced by instant head rotation, and the model group was further divided into 4 subgroups, with 8 rats in each： 6 h, 24 h, 72 h and 7 d. Rats in each group were evaluated for the modified neurological severity score （mNSS） before injury and before sacrifice. Meanwhile half of the rats in each group were used to assay the expression of DAPK1 in the corpus callosum and the brain stem after DAI by Western blot, while the other half were used to detect the brain neuron apoptosis after DAI. Results There were obvious weight loss and neurological deficit in the rats after DAI. At 6 h after brain injury, the expression of DAPK 1 increased obviously and obvious neuron apoptosis was observed. At 24 h after injury, both the expression of DAPK1 and neuron apoptosis reached the peak. However, they began to decrease at 72 h and 7d after injury, but were still higher than those in control group. Conclusion Weight decrease and neurological deficit in the rats after DAI may be related to the expression of DAPK1 and neuron apoptosis after DAI, both of which reached the peak at 24 h after injury. It suggested that the optimum therapeutic time window of DAPK1 inhibitors was 24 h after DA[.