2014-08-012024-05-17https://scholars.lib.ntu.edu.tw/handle/123456789/694733摘要：腦中風是造成全世界人口死亡的第三位，也是導致永久殘障的第一位。缺血性腦中風佔所有中風的七到八成，當腦血管急性阻塞之後會引起急性缺血系列反應導致神經細胞死亡。CPEB (cytoplasmic polyadenylation element binding protein)-類似蛋白，包括CPEB2，CPEB3 及CPEB4 是去氧核糖核酸的序列特異蛋白。其中CPEB3 及CPEB4 主要在神經細胞上表現，並且CPEB3 調控報導去氧核糖核酸轉錄的機制已經被證實與細胞NMDA 接受器訊息傳遞相關。過去的一個研究顯示，缺血性中風後CPEB4 會從細胞質轉移到核內，並且將CPEB4 表現量減少後的神經細胞較能抵抗缺氧缺糖的環境刺激。然而，究竟CPEB 相關的蛋白在缺血性中風是否扮演關鍵的角色仍舊需要更多的研究結果佐證。最近我們成功的培育出CPEB3 及CPEB4 兩種基因剔除鼠，我們及其他實驗室的研究結果也發現CPEB蛋白在不同的疾病狀況下與缺氧誘發因子及組織型纖維蛋白溶酶原的活化有關。因此，本研究預計以三年的時間從臨床與基礎的面向來探討CPEB 蛋白在缺血性中風扮演的病生理角色。在基礎研究的部分，我們首先利用小鼠急性腦中風的中大腦動脈阻塞及再灌流手術及神經細胞的缺氧缺血模式來探討神經細胞的CPEB 蛋白及相關訊息在缺血後的生理變化。之後在CPEB4 的基因剔除鼠並且CPEB 4 基因剔除的神經細胞進行急性腦中風模式並與非基因剔除的對照組比較中風範圍，神經功能，細胞的存活是否有顯著差異，並且中風後CPEB 的下游及相關訊息蛋白的變化。同樣的實驗步驟也將在CPEB3 的基因剔除鼠並且CPEB 3 基因剔除的神經細胞進行。最後我們將在CPEB4 剔除神經細胞上進一步減少CPEB2 及3 的表現量來決定CPEB 相關蛋白在缺血性腦中風是否有協同的關鍵作用。在臨床的部分，我們將針對急性腦中風病患接受開刀所切除的腦部組織進行染色來驗證神經細胞的CPEB 相關蛋白在急性缺血性中風後會從細胞質轉移入細胞核的現象。整體而言，本研究計畫預期將能建立神經細胞的CPEB 相關蛋白在急性腦中風的重要角色，並且提供未來發展腦中風治療的可能對象。<br> Abstract: Stroke is among the three leading causes of death worldwide and the most frequent causeof permanent disability. The ischemic stroke (IS) accounts for 70-80% of all strokes andocclusion of cerebral arteries induce acute ischemic cascades, leading to neuronal death.CPEB (cytoplasmic polyadenylation element binding protein)-like proteins, CPEB2, CPEB3and CPEB4, are sequence-specific RNA binding proteins. CPEB3 and CPEB4 are expressedpredominantly in neurons and the CPEB3- controlled translation of reporter RNA is regulatedby the NMDA receptor signaling. In IS, there was one study showing that CPEB4 maybecome nuclear in response to focal ischemia and CPEB4 knockdown neurons survived betterthan controls under oxygen and glucose-deprived (OGD) condition. However, whetherneuronal CPEBs play any crucial role in IS requiring stronger evidence. Recently, ourlaboratory has successfully generated CPEB3 and CPEB4 knockout (KO) mice. Results fromours and other laboratories also demonstrated the link between CPEBs and the activations ofhypoxic inducible factor and tissue plasminogen activator in various pathological conditions.Therefore, this 3 years study aims to investigate the role of CPEBs in acute IS through basicand clinical aspects.In basic part, we will perform MCA ischemic/reperfusion injury on wild type mice andOGD on wild type neurons to study the physiological changes of neuronal CPEBs andassociated signaling in response to ischemic damage. Then we will perform theaforementioned experimental stroke models on CPEB4 KO mice and neurons as well as wildtype controls to see any difference in brain infarct volume, behavior score and cell viability.Downstream pathways of CPEBs in CPEB4 KO and wild type control mice and neurons willalso be investigated. The same experimental procedures will be repeated on CPEB3 KO miceand neurons. Furthermore, we will produce double knock down of CPEB2 & 3 genes onCPEB4 KO neuron for OGD to determine the synergistic effect of neuronal CPEBs on IS.In clinical part, brain tissues from acute IS patients receiving surgery will be obtained forimmunostaining to demonstrate the phenomenon of CPEBs nuclear translocation from cytosolin IS patients. Overall speaking, our study will be anticipated to establish a pivotal role andfuture therapeutic target of neuronal CPEB in acute IS.缺血性腦中風CPEB 蛋白神經細胞ischemic strokeCPEBneuron