大鼠脊髓缺血再灌注损伤后ERK、Akt的表达与细胞凋亡关系的研究

Neuronal apoptosis and expressions of ERK、Akt in rats with spinal cord ischemia reperfusion injury

目的:观察在大鼠脊髓缺血再灌注损伤过程中ERK、Akt的表达规律及其与神经细胞凋亡的关系。方法:采用胸主动脉阻断法建立脊髓缺血(25分)/再灌注损伤模型。健康成年SD大鼠118只,用随机数字表法分为两组:1脊髓缺血再灌注组96只,每个时间点12只;2假手术对照组12只大鼠,只置入球囊而不阻断。用形态学、分子生物学等方法,分别于缺血再灌注后0,1,2,4,12,24,48和72小时检测缺血段脊髓组织中细胞凋亡以及P-ERK、P-Akt的表达水平变化情况。结果:TUNEL染色结果显示,脊髓缺血再灌注损伤后12小时凋亡细胞明显增加,48小时达高峰。损伤后4小时细胞的P-ERK表达水平达高峰,损伤后12小时表达至低谷。损伤后2小时细胞的P-Akt表达水平至高峰,损伤后12小时表达至低谷。假手术组:P-ERK、P-Akt少有表达,少见TUNEL阳性细胞。结论:P-ERK、P-Akt均参与脊髓缺血再灌注损伤后的神经细胞凋亡,ERK、Akt信号通路被调控的有效时间窗可能在再灌注后12小时内。

Objective： To observe the rules of cellular apoptotic change and ERK and AKT expression in he rat spinal cord ischemia/reperfusion injury（SCII）. Methods： one hundred eighteen adult SD rats （250- 300g） were randomly divided into 2 groups, the control group（ n = 12）and operation group （n = 96）. Spinal cord ischemia was induced in male Sprague-Dawley rats by balloon occlusion of the thoracic aorta. The operation process in the control group was the same as the operation group except the ischemia/reperfusion of the spinal cord. The segments of ischemia/reperfusion spinal cord were properly taken at an interval of 0,1,2 ,4 ,12 ,24 ,48 and 72 h after reperfusion. The ischemia/reperfusion segments of spinal cord were examined by Nissl, TUNEL staining, Western blot and image analysis. Results： TUNEL-positive cells were significantly at 12 hour after spinal cord injury, and peaked at 48 hour after injury. P-ERK positive cells were significantly reached a peak at 4 hour or a trough at 12 hour after spinal cord injury, P-Akt positive cells were significantly reached a peak at 2 hour or a trough at 12 hour after spinal cord injury. P-ERK, P-Akt, TUNEL-positive cells were rare in control group. Conclusion： P-ERK, P-Akt signaling pathway are involved in mechanism of neuronal apoptosis after spinal cord ischemia and reperfusion, the effective time window that ERK, Akt signaling pathway could been regulated may be with in 12 hours after reperfusion.