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目的探讨p-Akt、Bid在小鼠局灶性脑缺血再灌注(I/R)及缺血后处理(IP)后的启动规律,探讨p-Akt、Bid与IP保护作用的关系。方法将315只雄性昆明小鼠采用线栓法建立大脑中动脉栓塞模型后,随机分成假手术组(Sham组)、缺血再灌注组(I/R组)、缺血后处理组(IP组)、缺血后处理+PI-3K抑制剂LY294002组(IP+LY组)4组,于再灌注相应时间点取材,免疫组织化学方法检测p-Akt、Bid的表达及分布情况;免疫印迹法检测皮质p-Akt、Bid蛋白表达情况。氯化三苯基四氮唑(TTC)染色测定脑梗死体积。结果与Sham组比较,I/R组神经元胞质中可见p-Akt及Bid的表达明显上调,p-Akt表达于30min增加,1 h达高峰(P<0.05),24 h回落至基线水平,Bid表达于6 h增加,24 h达高峰(P<0.05),48～72 h有所回落;IP组各相应时间点较I/R组p-Akt表达显著增加(P<0.05),Bid表达明显降低(P<0.05)。应用PI-3K特异性阻断剂LY294002阻断PI-3K/Akt信号系统活化的同时,也抑制了IP的保护作用。p-Akt蛋白表达量与Bid免疫反应性呈负相关(r=-0.8265,P<0.05)。结论 p-Akt、Bid参与缺血后处理过程;缺血后处理可能通过PI-3K/Akt通路发挥脑保护作用;Akt可能通过死亡受体通路调控Bid的表达。
Objective To investigate the activation of p-Akt and Bid after focal cerebral ischemia-reperfusion (I / R) and post-ischemic postconditioning (IP) in mice and to explore the relationship between p-Akt, Bid and IP. Methods Thirty-five male Kunming mice were randomly divided into three groups: Sham group, I / R group, IP group, ischemia-reperfusion group ), Ischemic postconditioning + PI-3K inhibitor LY294002 group (IP + LY group) were taken at the corresponding time point after reperfusion. The expression and distribution of p-Akt and Bid were detected by immunohistochemistry. Detection of cortex p-Akt, Bid protein expression. Measurement of infarct volume by triphenyltetrazolium chloride (TTC) staining. Results Compared with Sham group, the expressions of p-Akt and Bid were significantly up-regulated in neuronal cytoplasm of I / R group. The expression of p-Akt increased at 30 min and peaked at 1 h (P <0.05) (P <0.05). The expression of Bid increased at 6 h, peaked at 24 h (P <0.05), and decreased at 48-72 h. The expression of p-Akt in Bid group increased significantly at each time point The expression was significantly decreased (P <0.05). PI-3K-specific blocker LY294002 blocked PI-3K / Akt signaling system activation, but also inhibited the protective effect of IP. The expression of p-Akt protein was negatively correlated with Bid immunoreactivity (r = -0.8265, P <0.05). Conclusions p-Akt and Bid are involved in the process of ischemic postconditioning. Post-ischemic postconditioning may play a protective role through PI-3K / Akt pathway. Akt may regulate the expression of Bid via death receptor pathway.