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目的构建模拟深低温低流量过程的Akt1部分敲除(Akt1+/-)的转基因小鼠模型,并分析小鼠的表型。方法3周龄Akt1+/-转基因小鼠与野生型(WT)小鼠各为48只,分别随机均分为脑部血流量的监测(A)组、血液动力学参数指标监测(B)组、缺血再灌注后RT-PCR实验(C)组与蛋白电泳实验(D)组,每组再分为假手术组与手术组2个亚组。每个亚组6只。通过在(18.5±0.5)℃的低温下钳闭转基因与野生型小鼠颈总动脉2h,并重新开放,模拟深低温低流量的病理生理过程。检测小鼠血流动力学的变化与脑部的血流情况,统计小鼠死亡率,通过RT-PCR和Westernblot等分析小鼠表型。结果阻断颈总动脉后,激光多普勒血流仪测定脑部血流量减少86%以上;Akt1+/-小鼠再灌注24h后的Akt下游线粒体凋亡通路中的细胞色素C与Caspase-3表达水平以及死亡率较假手术对照组增加(P<0.05);转基因小鼠的Akt活性被抑制。结论这一转基因小鼠模型基本模拟了深低温低流量的临床病理生理。Akt1部分敲除后,加重了小鼠的脑损害程度,说明PI3K/Akt信号通路具有脑保护功能。
Objective To construct a murine model of Akt1 knockout (Akt1 +/-) that simulates hypobaric hypobaric flow and analyze the phenotype of mice. Methods A total of 48 Akt1 +/- transgenic mice and WT mice (3 weeks old) were randomly divided into three groups: control group (A), hemodynamic parameters (B group) After ischemia-reperfusion RT-PCR experiment (C) and protein electrophoresis experiment (D) group, each group was further divided into sham operation group and operation group 2 subgroups. 6 in each subgroup The pathophysiological process of deep hypobaric hypobaric flow was simulated by clamping the carotid artery of transgenic and wild-type mice for 2h at a low temperature of (18.5 ± 0.5) ℃ and reopening. The changes of hemodynamics and brain blood flow in mice were detected. The mortality of mice was calculated. The mouse phenotypes were analyzed by RT-PCR and Western blot. Results After the common carotid artery was blocked, the cerebral blood flow decreased by more than 86% by laser Doppler flowmetry. The expressions of cytochrome C and Caspase-3 in Akt downstream mitochondrial apoptotic pathway after Akt1 +/- mouse reperfusion for 24h The expression level and the mortality rate increased compared with the sham operation control group (P <0.05). The Akt activity of the transgenic mice was inhibited. Conclusion This transgenic mouse model basically simulates the clinicopathophysiology of deep hypothermia and low flow rate. Akt1 partially knocked out, aggravating the degree of brain damage in mice, indicating that PI3K / Akt signaling pathway with brain protection.