新生鼠缺氧缺血性脑损伤时三磷酸肌醇激酶信号通路与缺氧诱导因子1α的调节  

PHOSPHOINOSITID 3-KINASE/Akt PATHWAY INVOLVED IN REGULATION OF HYPOXIA INDUCIBLE FACTOR 1α IN HYPOXIA ISCHEMIA BRAIN DAMAGE OF NEONATAL RATS

作　　者：李丽华[1] 屈艺[1] 毛萌[1] 母得志[1] 

机构地区：[1]四川大学华西第二医院儿科

出　　处：《中国修复重建外科杂志》2008年第9期1102-1107,共6页Chinese Journal of Reparative and Reconstructive Surgery

基　　金：国家自然科学基金资助项目(30570623,30770748);四川省科技厅应用基础资助项目(07JY029-067);教育部资金资助项目(2006331-11-7,20070610092);四川大学华西医学中心CMB人力资源资助项目(00-722)~~

摘　　要：目的探讨在体神经元缺氧缺血条件下缺氧诱导因子1α(hypoxia inducible factor1α,HIF-1α)蛋白的表达及三磷酸肌醇激酶(phosphoinositid 3-kinase/Akt,PI3K/Akt)信号通路的激活,推测PI3K/Akt信号通路与神经元HIF-1α表达调节的关系,以期更确切地阐明PI3K/Akt在发育期脑缺氧缺血性损伤(hypoxia ischemia brain damage,HIBD)神经元HIF-1α表达中的作用。方法10d龄SD大鼠56只,分为正常对照组(n=12)、假手术组(n=12)、实验组(n=24)、wortmannin干预组(n=4)和溶剂干预组(n=4)。实验组在乙醚麻醉下行右侧颈总动脉结扎,氮氧混合气(92%N2、8%O2)缺氧2.5h,即为HIBD模型;假手术组不结扎颈总动脉,不作缺氧处理;正常对照组不作任何处理。wortmannin干预组和溶剂干预组分别在侧脑室内注射wortmannin和DMSO、PBS3μL,30min后制备HIBD模型。分别于建模后4、8及24h处死动物取脑组织,应用免疫组织化学法检测HIF-1α、Akt蛋白的分布和表达,Western blot检测HIF-1α、Akt及p-Akt蛋白含量。结果实验组HIF-1α蛋白表达在术后4h明显升高,8h达高峰,24h后下降;p-Akt蛋白于术后4h明显升高,8h后下降。正常对照组各时间点HIF-1α和p-Akt均有极少量弱阳性表达。实验组各时间点HIF-1α蛋白表达明显高于正常对照组,差异有统计学意义(P<0.01);实验组p-Akt于4h和8h明显高于正常对照组,差异有统计学意义(P<0.05)。Akt蛋白表达随术后时间的延长无明显变化,与正常对照组比较差异无统计学意义(P>0.05)。术后4h wortmannin干预组的HIF-1α蛋白的表达明显下降,与溶剂干预组及实验组比较差异有统计学意义(P<0.01)。结论发育期HIBD时,可激活PI3K/Akt信号通路,并诱导HIF-1α表达增加,可针对PI3K/Akt信号通路和HIF-1α寻找新生儿HIBD新的治疗靶点。Objective To investigate the expression of hypoxia inducible factor let （HIF-1α） protein and the activation of phosphoinositid 3-kinase/Akt （PI3K/Akt） signaling pathway in neurons under hypoxia ischemia condition, and to elucidate the role of PI3K/Akt on HIF-1α regulation in the developing neurons after hypoxia ischemia brain damage （HIBD）. Methods Fifty-six SD rats aged 10 days were randomly divided into normal control group （n=12）, sham operation group （n=12）, experimental group （n=24）, wortmannin treated group （n=4） and DMSO/PBS treated group （n=4）. In the experimental group, the rats were anesthetized with ethylether. The right common carotid artery was exposed and ligated. Then, they were exposed to hypoxia in a normobaric chamber filled with 8% oxygen and 92% nitrogen for 2.5 hours. In the sham control group, the right common carotid artery was exposed but was not ligated or exposed hypoxia. In the normal control group, the rats recevied no further processing. For wortmannin treated group and DMSO/PBS treated group, the rats received intraventricular injection of wortmannin or DMSO/PBS 30 minutes before hypoxia ischemia. The brain tissues were harvested from the rats in the normal control, sham operation and experimental groups at 4, 8 and 24 hours after hypoxia ischemia, but in the wortmannin and DMSO/PBS treated groups only at 4 hours. The HIF-1α protein expression and Akt protein expression were detected with immunohistochemistry method. HIF-1α, Akt and p-Akt protein expression were measured by Western blot analysis. Results In the experimental group, the HIF-1α expression was significantly increased at 4 hours after operation, reached the peak level at 8 hours, and began to decrease at 24 hours. The p-Akt protein was significantly increased at 4 hours, and began to decrease at 8 hours. However, the expression levels of HIF- 1α and p-Akt protein in the normal control group were extremely low at each time point. So, the expression levels of HIF-1α in the experimental

关 键 词：脑 缺氧缺血 三磷酸肌醇激酶 缺氧诱导因子1Α 新生大鼠 

分 类 号：R722.1[医药卫生—儿科]