脑衰蛋白反应调节蛋白-2参与低氧预适应减轻小鼠脑缺血损伤  

CRMP-2 is involved in hypoxic preconditioning-induced neuroprotection against cerebral ischemic injuries of mice

作　　者：张彩艳[1] 封素娟[1] 刘旭[1] 卜祥宁[1] 张楠[1] 郑雅欣[1] 袁晓文[1] 李晓光[1] 李俊发[1] 

机构地区：[1]首都医科大学神经生物学系北京神经科学研究所,北京100069

出　　处：《基础医学与临床》2009年第11期1133-1138,共6页Basic and Clinical Medicine

基　　金：国家自然科学基金(30670782;30871219);国家"973"计划(2006CB5041);北京市自然科学基金(5072008);北京市教育委员会科技计划重点项目(KZ200810025012);北京市属高等学校人才强教计划项目(PHR200906116);北京市新世纪百千万人才工程培养经费资助(08-016)

摘　　要：目的研究参与低氧预适应(HPC)的经典型蛋白激酶CβⅡ(cPKCβⅡ)相互作用的脑衰蛋白反应调节蛋白-2(CRMP-2)对缺血脑是否有保护作用。方法成年雄性BALB/c小鼠随机分为常氧(Nor)、HPC、常氧假手术(Nor+sham)、HPC假手术(HPC+sham)、常氧缺血(Nor+I)和HPC缺血(HPC+I)等6组(每组n=6)。应用小鼠整体HPC和脑中动脉梗死(MCAO)致脑局部缺血模型,结合免疫沉淀、双向凝胶电泳和质谱等技术,分离和鉴定与cPKCβⅡ相互作用蛋白;利用聚丙烯酰胺凝胶电泳(SDS-PAGE)和蛋白印迹(Western blot)技术,分析CRMP-2磷酸化和蛋白降解水平在脑HPC和缺血中的变化。结果与Nor组比,HPC鼠脑皮层组织内有10种与cPKCβⅡ相互作用蛋白的表达发生了明显变化,其中CRMP-2在膜相关蛋白组分中的表达量升高,而在胞质蛋白组分中的表达量降低。在脑缺血模型中,与Nor+Sham组相比,Nor+I组小鼠脑皮层缺血核心区(Ic)CRMP-2磷酸化水平明显降低(P<0.05,n=6);与Nor+I组相比,HPC+I组小鼠脑皮层Ic区内CRMP-2磷酸化水平明显增高(P<0.05,n=6)。脑缺血可导致CRMP-2发生水解并伴随着大量55ku水解片段(BDP)的出现,但与Nor+I组相比,HPC+I组小鼠脑皮层缺血半影区(P)内CRMP-2水解片段减少,水解率明显降低(P<0.05,n=6)。结论CRMP-2参与了HPC缓解小鼠脑缺血Ic区内CRMP-2磷酸化水平的降低和减少P区内CRMP-2水解片段从而减轻缺血脑组织的损伤。Objective To investigate whether conventional protein kinase C （cPKC） βⅡ -interacting collapsin response mediator protein-2 （ CRMP-2 ） provides neuroproteetion against cerebral ischemic （ I ） injuries. Methods Male BALB/c mice were randomly divided into normoxic control （Nor） , HPC, Nor ＋ Sham, HPC ＋ Sham, Nor ＋ I and HPC ＋ I groups （ n = 6 per group）. Using our HPC and MCAO mouse models, we applied immunoprecipitation, two-dimensional electrophoresis and mass spectrometry to characterize cPKCβⅡ-interacting proteins and combined with SDS-PAGE and Western blot to quantitatively analyze CRMP-2 phosphorylation and degradation levels in the brain of mice after HPC and MCAO. Results The expression level of 10 cPKCβⅡ-interacting proteins changed obviously in cerebral cortex of HPC mice when compared with Nor group. One of these proteins, CRMP-2 protein level increased in particulate fraction and decreased in cytosolic fraction of cerebral cortex of HPC mice. CRMP-2 phosphorylation level in ischemic core （Ic） of cerebral cortex decreased significantly （ P 〈 0.05, n = 6） as com- pared with that of Nor ＋ sham group, but CRMP-2 phosphorylation level in HPC ＋ I group increased significantly as compared with that of Nor ＋ I group （P 〈 0. 05, n = 6）. In ischemic cortex, CRMP-2 degradation （proteolysis） was observed as the appearance of 55 ku breakdown products （BDP）. However, the CRMP-2 degradation level, BDPs products decreased significantly in penumbra （P） of ischemic cortex from HPC ＋ I group when we compared with that of Nor ＋ I group （P 〈 0.05, n = 6 ）. Conclusion CRMP-2 is involved in attenuating the decrease of CRMP-2 phosphorylation in ischemic core and in inhibiting its degradation in penumbra of cerebral cortex of mice thereby to lessen the ischemic injuries.

关 键 词：蛋白激酶CβⅡ 脑衰蛋白反应调节蛋白-2(CRMP-2) 蛋白质组学 低氧预适应 大脑中动脉阻塞 

分 类 号：R741[医药卫生—神经病学与精神病学]