| 其他摘要 | 全球有超过三千五百万人患阿尔茨海默病（Alzheimer's disease, AD）。AD是一种神经退行性疾病，临床上主要表现为认知和记忆能力损伤逐渐加重的渐进性痴呆，病理上主要表现为：Aβ肽在胞外沉积形成老年斑（senile plaques, SPs）、过度磷酸化的Tau蛋白在胞内聚集形成神经纤维缠结（neurofibrillary tangles, NFTs）、大量神经元死亡和脑内炎症反应。其中老年斑和神经纤维缠结是AD的两大经典的病理学特征。然而，至今AD的病因和发病机理仍不清楚。研究发现人体内甲醛（Formaldehyde, FA）的水平随着年龄的增长而升高，AD病人的尿液、血液、脑脊液以及脑组织中甲醛含量明显高于同龄对照组，因此，近年来甲醛被认为和AD病理的发生密切相关。此外，AD病人尿液中的甲醛含量和MMSE（Mini-Mental State Examinations）得分呈负相关，并且体外研究表明，甲醛还能促使Aβ肽和Tau蛋白的聚集。本研究通过向实验猕猴（5-8岁，不携带AD相关致病突变）侧脑室注射为期12个月低浓度的甲醛（甲醛溶解于生理盐水）或生理盐水，来探索甲醛在AD发生中所发挥的作用。该研究主要检测以下指标：（1）老年斑、神经纤维缠结与配对螺旋纤维（PHF），Aβ肽、磷酸化Tau与总Tau的量；（2）甲醛对工作记忆、长时记忆与运动能力的影响；（3）反应性星形胶质细胞增生。结果显示，长期注射甲醛的猕猴出现了工作记忆、长时记忆的损伤，但没有出现明显的运动能力的损伤。并且，实验猕猴脑内与记忆相关（也是AD相关）的脑区（海马、内嗅皮层与前额叶）出现了Aβ阳性的老年斑、神经纤维缠结、PHF、Tau蛋白磷酸化和Aβ42脑内水平的升高，及反应性星形胶质细胞增生。在发现侧脑室注射甲醛诱发实验猕猴表现AD的主要行为和病理改变后，为了进一步揭示内源性甲醛在AD病理中的扮演的角色，即升高的内源性甲醛是否会发挥与长期侧脑室注射甲醛相类似的作用，我们向猕猴的侧脑室注射为期12个月的甲醇（甲醛的前体）。此外，为了探索甲醇能否在灵长类脑内被代谢成甲醛，我们向另外3只猕猴侧脑室内单次注射相同浓度和计量的甲醇并观察30小时内脑脊液中甲醛含量的动态变化情况。结果发现，实验猕猴脑脊液中甲醛的水平不仅在单次注射甲醇后出现了升高，而且在长期给药的猕猴侧脑室内出现了累积升高的现象。并且我们在长期注射甲醇的猕猴脑内发现了类似于甲醛实验的结果：记忆相关的脑区出现了Aβ阳性的老年斑、神经纤维缠结、PHF、Tau蛋白磷酸化和Aβ42脑内水平的升高，及反应性星形胶质细胞增生，并且也同时出现了空间工作记忆与长时记忆的损伤，但并未出现运动能力的损伤。综上所述，升高的颅内甲醛（源于直接注射的甲醛或由注射的甲醇在脑内代谢产生）诱发了非人灵长类实验动物表现出所有主要的AD行为和病理改变。该研究发现表明，甲醛作为一种与年龄和环境均相关的因素，在AD，尤其在散发型AD的发生和发展中扮演着重要的角色。; Alzheimer’s disease (AD) afflicts more than 35 million people worldwide. It is characterized clinically as a progressive dementia that manifests with debilitating cognitive and memory impairments. Pathologically it is characterized by neurodegeneration identified with extracellular deposits of amyloid-β (Aβ), intracellular formation of neurofibrillary tangles (NFTs), neuronal loss and inflammatory reactions. The accumulation of Aβ peptides in the form of senile plaques and hyperphosphorylated tau (a microtubule associated protein) in the form of NFTs are the two characteristic histological hallmarks and pathological features of the disease. However, the etiology and pathogenesis of the disease remain unclear. Formaldehyde (FA) has recently been implicated in AD pathology, because FA has been found to undergo age dependent increases, and to be elevated in urine, blood, cerebrospinal fluid (CSF) and the postmortem hippocampus of AD patients. FA levels in the urine of AD patients have also been inversely correlated with Mini-Mental State Examinations scores. FA is also known to aggregate Aβ and tau protein. In present study, FA and vehicle (physiological saline) were injected intracerebroventricularly into rhesus macaques (5-8 yrs, without AD related genetic mutations) over 12 months to evaluate the role of FA in AD development. Monkeys were then evaluated for the following aspects: (1) the development of senile plaques, NFTs, paired helical filament (PHF), Aβ levels, tau protein phosphorylation at residues T181 and total tau protein levels in different brain regions post-mortem; (2) effects of FA on working memory, long-term memory and motor function; and (3) the presence of reactive astrogliosis. Monkeys injected with FA were found to have spatial working memory and long-term memory impairments with no gross motor impairments. Moreover, the presence of Aβ positive senile plaques, NFTs, PHF, increased tau protein phosphorylation and Aβ42, and reactive astrogliosis were found in memory (and AD) related brain areas: the hippocampus, entorhinal cortex and prefrontal cortex. After FA was found to induce all major AD behavioral and pathological markers and changes in rhesus monkeys independent of genetic predispositions, intracerebroventricular (i.c.v.) injections of methanol, as a FA precursor, were given to rhesus macaques (5-8 yrs, without AD related genetic mutations) over a 12-month period to elucidate the role of elevated endogenous FA in AD development, that is, to determine whether elevated endogenous FA plays similar role in AD progression as exogenous FA does. Moreover, three other rhesus macaques were given single i.c.v. injection of methanol to investigate whether a metabolic process of methanol to FA occurs in nonhuman primate brain. FA levels in CSF were found to be elevated after chronic methanol i.c.v. injections as well as the single injection. Furthermore, the presence of Aβ positive senile plaques, NFTs, PHF, increased tau protein phosphorylation and reactive astrogliosis were also found in memory (and AD) related brain areas as witnessed in the monkeys received chronic FA i.c.v. injection. These changes corresponded with spatial working and long-term memory loss in the monkeys, with no gross motor impairments. All told, elevated FA levels that resulted from either direct FA i.c.v. injection or endogenous metabolism of methanol in brain were found to trigger all major AD pathological markers in nonhuman primates, suggesting that FA, as an age-related factor and environmental factor, plays a significant role in the initiation and progression of AD, especially in sporadic forms of AD that are independent of genetic predisposition. |
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