Effects of hypoxia on ischemic delayd neuronal death

1.沙土鼠暴露于舱内10%氧气中3周。在氟烷麻醉下阻断双侧颈总动脉5min，在常氧条件下进行再循环。分别于手术前、术后2、4、7天处死动物。用碱性成纤维细胞生长因子和血管内皮生长因子抗体进行免疫组织化学研究。低氧组大鼠海马区神经元死亡较重。缺氧组夹闭前在扣带回皮质、穹窿体部及内囊均有明显表达，而常氧组夹闭前未见表达。观察两组大鼠脑缺血再灌流后2、4天脑组织中bFGF和VEGF的表达。缺氧组的表达明显高于常氧组。两组在7天后均未观察到这些表达。采用差异显示技术，观察了正常氧沙土鼠脑缺血5分钟后，海马区mRNAs表达的变化。沙土鼠分为3组(对照组、6h组、2d组)。MRNAs可分为3种类型：逐渐消失、减少和恢复、新出现。对对照组大鼠海马区表达的一个在脑缺血后6h表达减少、2d消失的基因进行亚克隆和部分测序。该基因与神经元性五角蛋白有同源性。原位杂交和Northern印迹分析显示与差异显示技术显示的表达变化相同。Western印迹和免疫组织化学分析均未显示3组大鼠神经元五角蛋白水平的差异。这一观察结果表明，神经元五肽在延迟性神经元死亡中起重要作用。

1. Gerbils were exposed to 10% oxygen in a chamber for 3 weeks. The bilateral carotid arteries at the neck were occluded for 5 minutes under halothane anaesthesia, recirculated and fed in a normoxia. Animals were sacrificed before and 2,4, and 7 days after surgery. Immunohistochemical study was also done with antibodies against basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF). The neuronal death in the hippocampus was severer in the hypoxic group. Expression of both bFGF and VEGF was obvious in the cingulate cortex, corpus callosum and internal capsule before clipping in the hypoxic group, but not observed in the normoxic group before clipping. We observed the expression of both bFGF and VEGF widely in the brain at 2 and 4 days sfter recirculation in both groups. The expression in the hypoxic group was much more prominent than that in the normoxic group. These expressions were not observed at 7 days in both groups.2. Using a differential display technique, we examined the changes in expression of mRNAs in the hippocampus following 5-minute cerebral ischemia in normoxic Mongolian gerbils. Gerbils were divided into 3 groups (control, 6 hours, 2 days). mRNAs were classified into 3 patterns ; gradual disappearance, decrease and recovery, and new appearance. An mRNA species expressed in the hippocampus in the control group but which decreased at 6 hours after cerebral ischemia and disappeared at 2 days was subcloned and sequenced partly. This mRNA showed homology to neuronal pentraxin. In situ hybridization and Northern blot analyzes showed the same changes in expression as shown by the differential display technique. Neither Western blot nor immunohistochemical analyzes showed differences in neuronal pentraxin protein level among the 3 groups. This observation suggested that neuronal pentraxin plays an important role in delayd neuronal death.