【摘要】：阿爾茨海默病(Alzheimer’s disease, AD)是一種以老年人記憶和認知功能損傷為臨床表征的神經(jīng)退行性疾病。AD是老年癡呆疾病中最常見的形式，病理特征包括A聚集形成的老年斑(senile plaques, SP)、Tau蛋白異常磷酸化形成的神經(jīng)纖維纏結(jié)(neurofibrillary tangles, NFT)、神經(jīng)元丟失導(dǎo)致的腦萎縮、膠質(zhì)增生以及炎癥。在AD的發(fā)病過程中，Aβ扮演著重要的角色。Aβ是新陳代謝的自然產(chǎn)物，由39-42個氨基酸組成，由β-分泌酶(BACE1)和γ-分泌酶水解加工淀粉樣前體蛋白(APP)產(chǎn)生。大量的研究表明，Aβ的聚集在AD的發(fā)病機制中發(fā)揮關(guān)鍵性作用。Aβ單體聚集形成的可溶性的寡聚體是引發(fā)神經(jīng)細胞毒性的主要原因，而氧化應(yīng)激與炎癥反應(yīng)是其誘發(fā)細胞毒性的主要機制。 到目前為止，盡管不能確定AD的發(fā)生是否伴隨氧化應(yīng)激反應(yīng)，但是，氧化應(yīng)激反應(yīng)在AD的病程進展中發(fā)揮著重要的作用。氧化應(yīng)激過程中Aβ聚集物能夠誘導(dǎo)活性氧(Reactive oxygen species, ROS)的過度產(chǎn)生。產(chǎn)生的ROS可能損傷包括蛋白、脂質(zhì)以及DNA在內(nèi)的多種類型的生物大分子。雖然脂質(zhì)過氧化反應(yīng)產(chǎn)生的丙二醛(MDA)具有神經(jīng)毒性，尤其是可能導(dǎo)致蛋白質(zhì)變性，影響蛋白質(zhì)功能，但是脂質(zhì)過氧化本身可以通過直接損傷細胞膜結(jié)構(gòu)的完整性導(dǎo)致細胞凋亡甚至死亡。另一方面，據(jù)報道，在AD病人的腦中，對ROS具有解毒作用的內(nèi)源性的抗氧化物酶的水平降低。其中超氧化物歧化酶(SOD)與谷胱甘肽過氧化物酶(GSH-Px)是最主要的抗氧化物酶，能夠抵抗自由基誘導(dǎo)的氧化性損傷，發(fā)揮細胞保護作用。此外，Aβ聚集物能夠激活小膠質(zhì)細胞，促進前炎癥細胞因子諸如腫瘤壞死因子α(TNF-α)和白介素1β(IL-1β)的釋放，進而促進下游炎癥級聯(lián)反應(yīng)的發(fā)生。因此，在AD的修飾型藥物研發(fā)過程中，降低Aβ42聚集引起的神經(jīng)毒性、抑制Aβ42誘導(dǎo)產(chǎn)生的氧化應(yīng)激以及前炎癥細胞因子的釋放成為治療與預(yù)防AD的可行性計策。 在先前的研究工作中，實驗室以Aβ單體為靶分子，利用噬菌體展示技術(shù)，從環(huán)型七肽庫中經(jīng)過三輪篩選、富集、洗脫得到了能夠特異性與Aβ結(jié)合的多肽，并命名為XD3。本文在原來研究的基礎(chǔ)上，對XD3在體內(nèi)以及體外水平的藥效學(xué)進行了研究。 在細胞水平研究結(jié)果表明，多肽XD3能夠抑制Aβ42誘導(dǎo)產(chǎn)生的神經(jīng)細胞毒性；通過減少活性氧(ROS)以及氧化型谷胱甘肽(GSSG)的生成，，增加還原型谷胱甘肽GSH的含量，增強SOD、GSH-Px的活性降低Aβ42誘導(dǎo)產(chǎn)生的氧化應(yīng)激反應(yīng)；此外，XD3可以下調(diào)Aβ42誘導(dǎo)的促炎癥細胞因子TNF-α和IL-1β的釋放。 同時，應(yīng)用XD3對AD轉(zhuǎn)基因小鼠的治療結(jié)果表明，XD3可以明顯改善小鼠的認知記憶功能、通過降低腦內(nèi)的膠質(zhì)細胞增生抑制炎癥反應(yīng)、降低腦內(nèi)老年斑的沉積以及氧化應(yīng)激等病理性變化。 XD3作為小分子多肽，具有靶點明確、治療作用明顯、副作用小以及制備方便的特性，為AD的治療提供了具有良好應(yīng)用前景的免疫制劑。
[Abstract]:Alzheimer's disease (Alzheimer's disease, AD) is a neurodegenerative disease characterized by memory and cognitive impairment in the elderly. Ad is the most common form of Alzheimer's disease. The pathological features included brain atrophy, glial hyperplasia and inflammation caused by the loss of (neurofibrillary tangles, NFT), neurons in neurofibrillar tangles caused by abnormal phosphorylation of (senile plaques, SP), Tau protein in elderly plaques. A 尾 plays an important role in the pathogenesis of AD. A 尾 is a natural product of metabolism, which is composed of 39 amino acids and is produced by hydrolysis of 尾-secretase (BACE1) and gamma-secretase to process starch precursor protein (APP). A large number of studies have shown that the aggregation of A 尾 plays a key role in the pathogenesis of AD. The soluble oligomer formed by A 尾 monomer aggregation is the main cause of neurocytotoxicity. Oxidative stress and inflammation are the main mechanisms of cytotoxicity. Up to now, although it is impossible to determine whether the occurrence of AD is accompanied by oxidative stress, oxidative stress plays an important role in the course of AD. During oxidative stress, A 尾 aggregates can induce the overproduction of reactive oxygen species (Reactive oxygen species, ROS). The resulting ROS may damage many types of biological macromolecules, including proteins, lipids and DNA. Although malondialdehyde (MDA) (MDA) produced by lipid peroxide is neurotoxic, especially it may lead to protein denaturation and affect protein function. However, lipid peroxide itself can directly damage the integrity of cell membrane structure leading to apoptosis and even death. On the other hand, it is reported that the level of endogenous antioxidant enzymes that detoxify ROS is reported to be decreased in the brains of AD patients. Among them, (SOD) and GSH-Px are the most important antioxidant enzymes, which can resist oxidative damage induced by free radicals and play a role in cell protection. In addition, A 尾 aggregates can activate microglia and promote the release of proinflammatory cytokines such as tumor necrosis factor 偽 (TNF- 偽) and IL 1 尾 (IL-1 尾), thus promoting the occurrence of downstream inflammatory cascade reaction. Therefore, in the process of modified drug development of AD, reducing the neurotoxicity caused by A 尾 42 aggregation, inhibiting the oxidative stress induced by A 尾 42 and the release of proinflammatory cytokines are the feasible strategies for the treatment and prevention of AD. In the previous research work, using A 尾 monomer as the target molecule and bacteriophage display technique, the polypeptide which can specifically bind to A 尾 was obtained by three rounds of screening and eluting from the cyclic seven peptide library, and named XD3.. On the basis of the original study, the pharmacokinetics of XD3 in vivo and in vitro was studied. The results showed that polypeptide XD3 could inhibit the neurotoxicity induced by A 尾 42. By reducing the production of reactive oxygen species (ROS) and oxidized glutathione (GSSG), the content of reduced glutathione GSH was increased, and the activity of SOD,GSH-Px was enhanced to decrease the oxidative stress induced by A 尾 42. In addition, XD3 could down-regulate the release of pro-inflammatory cytokines TNF- 偽 and IL-1 尾 induced by A 尾 42. At the same time, the treatment of AD transgenic mice with XD3 showed that XD3 could significantly improve the cognitive memory function of mice and inhibit inflammatory response by reducing the proliferation of glial cells in the brain. Reduce the deposition of aging plaques in the brain and oxidative stress and other pathological changes. As a small molecular polypeptide, XD3 has the characteristics of clear target, obvious therapeutic effect, small side effects and convenient preparation, which provides an immune preparation with good application prospect for the treatment of AD.
【學(xué)位授予單位】：安徽農(nóng)業(yè)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2013
【分類號】：R749.16;R96

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1 莊瑩;陳杰;;阿爾茨海默病病因及發(fā)病機制研究進展[J];吉林醫(yī)藥學(xué)院學(xué)報;2008年02期

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