Effects of hypoxia on ischemic delayd neuronal death

1. 沙鼠在一个房间里暴露在10%的氧气中3周。双侧颈动脉在氟烷麻醉下闭塞5分钟，再循环，在常氧环境下喂养。分别于手术前、术后2、4、7天处死。同时用抗碱性成纤维细胞生长因子（bFGF）和血管内皮生长因子（VEGF）抗体进行免疫组化研究。缺氧组海马神经元死亡更严重。低氧组扣带皮层、胼胝体和内包膜中bFGF和VEGF表达明显，而常氧组扣带皮层、胼胝体和内包膜中bFGF和VEGF表达不明显。我们观察到两组脑循环后2天和4天bFGF和VEGF的广泛表达。低氧组的表达明显高于常氧组。7 d时，两组均未见上述表达。利用差异显示技术，我们检测了蒙古沙鼠在正常缺氧5分钟脑缺血后海马中mrna表达的变化。沙鼠分为3组（对照组、6 h组、2 d组）。mrna可分为3种模式；逐渐消失、减少、恢复，并重新出现。对对照组海马中表达的一种在脑缺血后6 h下降，2 d消失的mRNA进行亚克隆和部分测序。该mRNA与神经元戊烷素具有同源性。原位杂交和Northern blot分析显示，与差异显示技术显示的表达变化相同。Western blot和免疫组化分析均未发现三组间神经元戊烷素蛋白水平的差异。这一观察结果提示神经元戊烷素在延迟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.