目的 构建新生儿缺氧缺血性脑病的经典动物模型,并分析在缺氧缺血条件下,促红细胞生成素(EPO)治疗前后胶质细胞谷氨酸运载蛋白2(EAAT2)表达情况。方法 108只出生72 h的SD乳鼠随机分为六组:空白对照组、单纯缺氧组、单纯缺血组、缺氧缺血组、促红细胞生成素治疗组和生理盐水治疗组。缺氧缺血性脑病模型通过双侧结扎颈动脉并且缺氧处理2 h建立。观察并记录不同实验组中乳鼠行为的变化。利用蛋白质印迹技术从蛋白质水平上分别检测不同实验组中EAAT2蛋白的表达情况。利用实时荧光定量聚合酶链反应(PCR)技术从mRNA水平上分别检测不同实验组中新生乳鼠的海马区、大脑皮层EAAT1、EAAT2的基因表达情况。结果 模型呈现典型的缺氧缺血脑病特征。缺氧、缺血、缺氧缺血都可以导致海马组织、大脑皮层中EAAT1的mRNA水平及EAAT2表达量的降低。经过EPO治疗后,EAAT1及EAAT2的基因表达量明显上升。并且,大脑皮层中基因表达水平要高于海马组织中基因表达水平。结论 EAAT2表达量的下降可能是HIE发病的一个原因,EPO的疗效可能是通过上调EAAT2的表达量来实现。
Abstract
Objective To assess the expression levels of excitatory amino acid transporter 2 (EAAT2) in classical rat model of hypoxic ischemic encephalopathy (HIE),and the expression level of that after erythropoietin (EPO) treatment. Methods Totally,108 postnatal 72 h SD rats were randomly assigned into six groups,including control,hypoxia,ischemia,hypoxia-ischemia,EPO treatment,and saline treatment.A neonatal rat model of HIE by bilateral ligation of carotid artery and subsequent anaerobic treatment for 2 hrs was constructed.The behavior of rats was recorded.The expression level of EAAT2 was assessed by Western blotting.Moreover,mRNA levels of EAAT1 and EAAT2 was measured using real time polymerase chain reaction. Results Typical HIE characteristics were observed on the models.The mRNA level of EAAT1 and expression level of EAAT2 was obviously reduced after hypoxia,ischemia,and hypoxia-ischemia.After treatment by EPO,the expression levels of EAAT1 and EAAT2 increased.Moreover,the gene expression levels in cortex were higher than that in hippocampe. Conclusion Reduced EAAT2 expression levels may be one reason for the development of HIE,which might also contribute to the curative effect of EPO.
关键词
促红细胞生成素 /
缺氧缺血性脑病 /
EAAT1 /
EAAT2
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基金
深圳市基础研究计划项目(JCYJ20150402095641634)
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