NR2A反义寡核苷酸对短暂性脑缺血/再灌注大鼠海马神经元损伤的保护作用  

Neuroprotective effects of NR2A antisense oligodeoxynucleotides on transient brain ischemia/reperfusion-induced neuronal injure in rat hippocampus

作　　者：刘志安[1,2] 李梅[1] 张旭[3] 滕大才[1] 裴冬生[2] 徐铁军[1,2] 

机构地区：[1]徐州医学院解剖学和神经生物学教研室 [2]江苏省麻醉学重点实验室,江苏徐州221002 [3]北京大学附属首钢医院神经内科,北京100144

出　　处：《中国药理学通报》2010年第11期1424-1428,共5页Chinese Pharmacological Bulletin

基　　金：国家自然科学基金资助项目(No30800309);江苏省麻醉学重点实验室课题资助项目(NoKJS07005)

摘　　要：目的探讨NMDA受体亚单位2A(NR2A)反义寡核苷酸在短暂性脑缺血/再灌注大鼠海马神经元损伤中的保护作用,为研制和开发针对NR2A的特异性新药提供理论基础和形态学依据。方法健康♂SD大鼠随机分为正常对照组、假手术对照组和缺血/再灌注组。经生理盐水、错义寡核苷酸和反义寡核苷酸预处理后,以四血管阻断法建立短暂性全脑缺血(15min)/再灌注(1、2、3和5d)动物模型。在确定的时间点进行灌注固定、取材、石蜡包埋和组织切片(片厚8μm),然后行TUNEL反应、焦油紫染色、原位杂交染色以及免疫组织化学染色。结果短暂性脑缺血/再灌注(I/R)3d,大鼠海马CA1区出现大量的凋亡阳性细胞;I/R5d大鼠海马CA1区细胞严重受损,与对照组相比二组差异具有显著性(P<0.05)。经NR2A反义寡核苷酸预处理后,I/R3d和I/R5d海马CA1区的细胞凋亡和细胞损伤明显减轻,与对照组相比二组差异具有显著性(P<0.05)。NR2A反义寡核苷酸能抑制I/R1d NR2A mRNA表达和I/R2d蛋白质表达,与对照组相比差异具有显著性(P<0.05)。结论短暂性全脑缺血后,NR2A反义寡核苷酸能明显地减轻缺血诱导的大鼠海马CA1区细胞凋亡和细胞损伤,且这种作用与NR2A反义寡核苷酸特异性抑制NR2A及其mRNA的表达密切相关。Aim To investigate the neuroprotective effects of NMDA receptor subunit 2A（NR2A） antisense oligodeoxynucleotides on transient brain ischemia/reperfusion-induced neuronal injure in rat hippocampus,and to provide theoretical basis and morphologic accordance for developing new drugs aiming at NR2A subunit.Methods Healthy male SD rats were randomly divided into normal control group,sham operation control group and ischemia/reperfusion group.After pretreatment with saline,misssense oligonucleotide and antisense oligonucleotide,the four-vessel occlusion method was used to establish transient forebrain ischemia（15 min） and reperfusion（1,2,3 d and 5 d） animal model.At the specific time-points,the rats were killed,the hippocampus paraffin-embedded and tissue sections made （slice thickness is 8 μm）.The slices were processed by TUNEL reaction,Cresyl violet staining,in situ hybridization staining and immunohistochemistry staining.Results In transient cerebral ischemia/reperfusion 3 d,TUNEL staining showed that the number of apoptotic cells was significantly increased in CA1 subfield of rat hippocampus,and in ischemia/reperfusion 5 d,Cresyl violet staining showed that the neurons in CA1 subfield of rat hippocampus were severely damaged,compared with the control group,which both manifested significant difference （P0.05）.After pretreatment with antisense oligonucleotide,the number of apoptotic cells at I/R 3 d and the damaged neurons at I/R 5 d in CA1 subfield of rat hippocampus were decreased significantly （P0.05）.NR2A antisense oligonucleotide can significantly inhibit the expression of NR2A mRNA and NR2A protein in CA1 subfield of rat hippocampus,respectively in I/R 1 d and I/R 2 d,Which was markedly different from the two control groups （P0.05）.Conclusion After transient forebrain ischemia,NR2A antisense oligonucleotide can significantly reduce the ischemic apoptosis and the neuronal damage in CA1 subfield of rat hippocampus induced by cerebral ischemia and reperfusion,and this effect closely co

关 键 词：NR2A反义寡核苷酸 神经保护 NR2A 脑缺血 神经元损伤 大鼠 海马 

分 类 号：R-332[医药卫生] R322.81