建立阿尔茨海默症人源肠道菌群动物模型

目的 使用粪菌移植方法 经口途径获得阿尔茨海默症人源肠道菌群(human flora-associated,HFA)模型小鼠,并通过生物信息学、组织病理学等方法对模型进行评价.方法 无菌雌性C57BL/6J小鼠28只,随机分为阿尔茨海默症模型(AD)组和对照(CON)组,分别经口途径接种阿尔茨海默症患者及健康志愿者的新...     

早老素与阿尔茨海默症的研究进展

早老素（Presenilin,PS）是早发性阿尔茨海默症的风险基因,在外周组织和脑内均有表达。早老素行使多种生物学功能,除了是r-分泌酶的活性基团,还参与信号通路,调控细胞分化与胚胎发育。本综述旨在对早老素的多种生物学功能及其在阿尔茨海默症中的作用进行概述。     

双拷贝APP/BACE/DPsn转基因果蝇模型的建立及基因功能的研究

阿尔茨海默症 (Alzheimer′s disease, AD), 是一种以脑中β-淀粉样蛋白 (β-amyloid peptide, Aβ)沉积为主要病理改变的神经退行性疾病。在果蝇Drosophila模型中建立淀粉样蛋白前体蛋白 (amyloid precursor protein, APP)的剪切通路模拟Aβ的产生过程, 有望建立一种快速筛选治疗AD药物的动物模型。我们利用经典的Gal4/UAS系统, 将现有的APP/BACE/DPsn果蝇品系连续杂交, 通过同源重组的方法构建表达两个拷贝的APP/BACE/DPsn稳定可遗传的转基因果蝇新品系。进一步的实验结果表明： 与不表达APP/BACE/DPsn的对照果蝇w/y; APP/Cyo; BACE-DPsn/TM6BTb相比, 表达两拷贝APP/BACE/DPsn的 w/y; elav-APP; BACE-DPsn果蝇的最长寿命为52 d, 比对照组(69 d)缩短了17 d, 为对照组果蝇的75%; 中位生存时间为39 d, 比对照组(49 d)缩短了10 d, 为对照组的80%; 平均寿命为37 d, 比对照组(47 d)缩短了10 d, 为对照组的79%。同时, 表达两个拷贝APP/BACE/DPsn的果蝇所产卵的羽化时间比对照果蝇延长了3 d; 其羽化成虫的理论值为1∶9 (11%), 而实际羽化率仅为5.2%。结果提示, 由elav-Gal驱动在果蝇泛神经元内过表达APP/BACE/DPsn, 可以缩短果蝇寿命、 干扰果蝇胚胎正常发育。该果蝇有可能作为初步筛选AD治疗药物的动物模型, 为AD治疗新药的发现提供工具。     

阿尔茨海默症基因研究态势分析

阿尔茨海默症是一种起病隐匿的进行性发展的神经系统退行性疾病。随着全球老龄化日趋严重,阿尔茨海默症严重威胁人民的生命健康与生活质量。阿尔茨海默症成因复杂,治疗阿尔茨海默症仍然是重要的医学难题。通过对Web of Science核心合集中收录的阿尔茨海默症基因研究相关的文献,利用文献计量的方法,进行研究趋势、研究人员、研究机构、国家和研究热点等方面进行态势分析,为相关领域的研究人员提供重要的参考。     

阿尔茨海默症基因研究态势分析

阿尔茨海默症是一种起病隐匿的进行性发展的神经系统退行性疾病。随着全球老龄化日趋严重,阿尔茨海默症严重威胁人民的生命健康与生活质量。阿尔茨海默症成因复杂,治疗阿尔茨海默症仍然是重要的医学难题。通过对Web of Science核心合集中收录的阿尔茨海默症基因研究相关的文献,利用文献计量的方法,进行研究趋势、研究人员、研究机构、国家和研究热点等方面进行态势分析,为相关领域的研究人员提供重要的参考。     

脂筏在阿尔茨海默症中的作用

脂筏(lipid raft)是细胞膜中富含胆固醇的功能性微区,在信号转导、物质运输等方面发挥着重要作用。大量证据显示脂筏与阿尔茨海默症(Alzheimer’s disease,AD)的致病机理密切相关。β-淀粉样肽(amyloidβ-peptide,Aβ)的异常代谢和聚集可能是AD的致病主因,而脂筏不但是Aβ产生的主要场所,还能调节Aβ的聚集行为及神经毒性,因而在AD的病理过程中扮演着关键角色。     

壳寡糖可降低阿尔茨海默症模型细胞内活性氧水平

用荧光显微成像测量了两种阿尔茨海默症（Alzheimer's disease,AD）模型细胞即swe型N2a细胞和Δ9/swe型N2a细胞中活性氧的水平。培养液中适当浓度的壳寡糖降低了两种模型细胞中的活性氧水平,这一抑制作用随壳寡糖浓度的变化而变化。壳寡糖对两种模型细胞作用存在差别,提示壳寡糖抑制AD模型细胞内活性氧可能有不同的途径。     

自噬在阿尔茨海默症治疗中的研究进展

自噬(autophagy)是生物体活细胞用来维持自身平衡稳定的一个保守进程。已有研究表明,自噬与阿尔茨海默症(Alzheimer’s disease,AD)密切相关,并可能在AD的病程中起着关键作用。然而,迄今为止,其完整的作用机制仍未能得到深入的研究,自噬在AD中起到的作用仍然存在许多争议。近几年关于自噬在AD中作用和治疗方面有了一系列研究进展,陆续发现了一些能够通过调节自噬而改善AD的治疗手段,为相关的研究领域提供了一个新的认识和展望的平台。现就自噬在AD中的作用机制和相关治疗方法作一简要综述。     

阿尔茨海默症的诊断与治疗研究进展

阿尔茨海默症(AD)即老年痴呆症,是以老年斑和神经元纤维缠结为主要病理特征的神经退行性疾病.AD的发病机制是多种发病素、多种通路和分子机制的相互参与,例如信号异常、炎症和免疫系统、脂质转运、细胞内吞作用、细胞凋亡、氧化损伤和应激反应、tau 病理学、神经元和突触的损失、能量代谢等.目前没有一种 AD 治疗方法能从根本上停止其病理的退行性改变,但仍有多种治疗策略.我们从生物标志物、遗传、神经影像、药物治疗、β淀粉样蛋白免疫治疗方面,对阿尔茨海默症的治疗研究进展进行了简要综述.     

视网膜辅助阿尔茨海默症的早期诊断

阿尔茨海默症(Alzheimer's disease, AD)是一种神经系统退行性疾病,常见于老年患者,现已成为世界范围内最常见的老年痴呆疾病,迄今仍未发现效果显著的治疗方法。AD患者的认知障碍往往在脑中发生第一次退行性改变数年后才出现,同时,它的诊断方法多具侵入性、非常规、价格高昂。AD是对全人类健康的挑战,也是严峻的公共卫生问题。因此,急需确定一种能够在患病早期检测到阿尔茨海默症的新型生物标记。由于阿尔茨海默症患者通常表现出明确的视觉缺陷,且这种缺陷有时在患者认知障碍的初次表现之前就已经出现,故近年来,以视网膜辅助AD早期诊断的新概念引起了极大关注。这种概念有望为AD的早期诊断提供新方法。     

APP695^K595N／M596L突变转基因小鼠的建立和病理表型动态分析

目的建立APP695^K595N／M596L（Swedish突变）转基因小鼠和评价痴呆表型的发生和发展过程。方法将APP695^K595N／M596L突变基因插入到小鼠朊蛋白（mouse prion protein）启动子下游,构建转基因表达载体,通过显微注射法建立APP695^K595N／M596L突变转基因C57BL／6J小鼠。PCR鉴定转基因小鼠的基因表型,Western blotting检测APP突变基因表达。Thioflavin-S染色检测不同年龄转基因小鼠大脑病理改变。Morris水迷宫动态观察小鼠行为学改变。结果建立了人APP695^K595N／M596L转基因小鼠,Thioflavin-S染色显示转基因小鼠9月龄时在脑海马区可检测到老年斑形成,并且在11、12月龄时明显增多。Morris水迷宫结果发现与同月龄野生型小鼠相比,该转基因小鼠5月龄开始出现学习记忆能力缺陷,7、9、11月行为学结果证实转基因小鼠的学习记忆能力缺陷随年龄增加而日趋严重（P＜0．05）。结论建立了人APP695^K595N／M596L转基因小鼠,并能再现人类阿尔茨海默症的行为学及神经病理学特征,为阿尔茨海默病发病机制研究和药物研发提供了有价值的动物模型。     

BAG-1M is up-regulated in hippocampus of Alzheimer's disease patients and associates with tau and APP proteins

The accumulation of tau and amyloid beta proteins is the major molecular pathology of Alzheimer's disease (AD). The mechanisms leading to the accumulation of these proteins are not completely clear. Hsc-70/Hsp-70, a chaperone protein, has been shown to bind both these proteins and regulate their degradation. We have previously shown that the co-chaperone protein BAG-1 can inhibit the degradation of tau by forming a complex with Hsc-70 and tau. In this current work, we show that there is an increase in the BAG-1M isoform in the hippocampus of AD patients. In addition, BAG-1 binds to both tau and amyloid precursor protein physically, and is found highly expressed in the same neurons that contain intracellular tau or amyloid in hippocampal sections from AD patients. Over-expression of BAG-1M in cell culture also induced an increase in both tau and amyloid precursor protein levels. In conclusion, we report a specific increase of BAG-1M in human AD patients, which is both physically and functionally associated to the two major molecular markers of AD.     

Impact of cholesterol level upon APP and BACE proximity and APP cleavage

Cleavage of APP by BACE is the first proteolytic step in the production of Amyloid β (Aβ, which accumulates in senile plaques in Alzheimer’s disease. BACE-cleavage of APP is thought to happen in endosomes. However, there are controversial data whether APP and BACE can already interact on the cell surface dependent on the cholesterol level. To examine whether APP and BACE come into close proximity on the cell surface in living cells, we employed a novel technique by combining time-resolved Förster resonance energy transfer (FRET) measurements with total internal reflection microscopy (TIRET microscopy). Our data indicate that BACE and APP come into close proximity within the cell, but probably not on the cell surface. To analyze the impact of alterations in cholesterol level upon BACE-cleavage, we measured sAPP secretion. Alteration of APP processing and BACE proximity by cholesterol might be explained by alterations in cell membrane fluidity.     

Acetylcholinesterase Is Increased in the Brains of Transgenic Mice Expressing the C-Terminal Fragment (CT100) of the β-Amyloid Protein Precursor of Alzheimer's Disease

Abstract: Acetylcholinesterase (AChE) expression is markedly affected in Alzheimer's disease (AD). AChE activity is lower in most regions of the AD brain, but it is increased within and around amyloid plaques. We have previously shown that AChE expression in P19 cells is increased by the amyloid β protein (Aβ). The aim of this study was to investigate AChE expression using a transgenic mouse model of Aβ overproduction. The β-actin promoter was used to drive expression of a transgene encoding the 100-amino acid C-terminal fragment of the human amyloid precursor protein (APP CT100). Analysis of extracts from transgenic mice revealed that the human sequences of full-length human APP CT100 and Aβ were overexpressed in the brain. Levels of salt-extractable AChE isoforms were increased in the brains of APP CT100 mice. There was also an increase in amphiphilic monomeric form (G^A₁) of AChE in the APP CT100 mice, whereas other isoforms were not changed. An increase in the proportion of G^A₁ AChE was also detected in samples of frontal cortex from AD patients. Analysis of AChE by lectin binding revealed differences in the glycosylation pattern in APP CT100 mice similar to those observed in frontal cortex samples from AD. The results are consistent with the possibility that changes in AChE isoform levels and glycosylation patterns in the AD brain may be a direct consequence of altered APP metabolism.     

Purification and Solubilization of Paired Helical Filaments from Alzheimer Brains

The purpose of the present study was to develop a purification and solubilization method, compatible with current amino acid sequencing techniques, for paired helical filaments (PHFs) derived from patients with Alzheimer's disease. We have developed a mild procedure that subjects conventionally isolated PHFs to Tris/borate/sodium dodecyl sulfate/2-mercaptoethanol electrophoresis and results in the separation of the relatively insoluble PHF structures from both copurifying contaminating proteins and solubilized PHF-associated proteins. At the end of 4.5 h of electrophoresis, the purified insoluble fraction had an amino acid composition that was invariant during subsequent electrophoresis. Electron microscopy revealed an intact PHF structure before and after electrophoresis but no evidence of any other structures in the insoluble fraction, a result consistent with the removal of PHF-associated proteins from the filament structure. Isolated insoluble filament structures displayed an enhanced immunoreactivity with antibodies raised against purified PHFs in other laboratories, when compared with the fraction not subjected to electrophoresis in enzyme-linked immunosorbent assays. Solubilization of the relatively insoluble PHFs was accomplished by extending the time of electrophoresis beyond the 4.5 h required for purification. Additional electrophoresis for 34.5 h solubilized 88% of the purified, relatively insoluble PHFs. This resulted in the identification of four major protein bands between Mr values of approximately 50,000 and 70,000 on sodium dodecyl sulfate-polyacrylamide electrophoresis gel analysis, with a predominant band with an Mr of approximately 66,000. A slow fragmentation of the PHF ultrastructure occurred during this time, as judged by electron microscopy. This purification technique will permit the isolation of consistently reproducible protein fragments from solubilized PHFs, which may be used for subsequent sequence analysis.     

Chemically Regulated Expression Systems and their Applications in Transgenic Plants

In the past 20 years, several systems have been developed to control transgene expression in plants using chemicals. The components used to construct these systems are derived from regulatory sequences mostly from non-plant organisms such as bacteria, fungi, insects and mammals. These constructs allowed transgene expression to be controlled temporally, spatially and quantitatively with the help of exogenous chemicals, without disturbing endogenous plant gene expression. Various chemically regulated transgene expression systems, their advantages/disadvantages and their potential for large-scale field application are reviewed.     

Ribonuclease Activities and Distribution in Alzheimer''s and Control Brains

Levels of free and total alkaline ribonuclease, and levels of acidic ribonuclease, were measured postmortem in control brains and in the brains of patients with Alzheimer's disease. In each brain region assayed, whether control or Alzheimer's, there was a statistically significant difference between the levels of free and total alkaline ribonuclease. Between 59 and 90% of the enzyme activity was associated with alkaline ribonuclease inhibitor in an inactive complex. Levels of free and total alkaline ribonuclease varied widely among different brains and brain regions, and were always lower in cerebellum than in temporal cortex and occipital pole. There was no significant difference in the levels of total alkaline ribonuclease, free alkaline ribonuclease, or acidic ribonucleases between corresponding regions of Alzheimer's and control brains. There was also no qualitative difference in the subcellular distribution of the alkaline and acidic ribonucleases between Alzheimer's and control brain. No significant relationships were found between ribonuclease levels and age, neuritic plaque density, postmortem interval, or storage time.     

Insulin-Like Growth Factor I Receptor Binding in Brains of Alzheimer's and Alcoholic Patients

Patients with chronic alcoholism and/or Alzheimer's disease show degenerative changes in the cerebral cortex and hippocampus. To investigate possible changes in insulin-like growth factor I receptor binding sites in brain tissue of patients with these pathological conditions, the number of 125I-insulin-like growth factor I binding sites was determined in tissues obtained from control patients and those with Alzheimer's and/or with a history of alcoholism. The four experimental groups examined consisted of patients from similar age groups. Postmortem histology and a clinical history were used for the diagnosis of Alzheimer's disease and alcoholism, respectively. Careful clinical records were kept concerning other variables such as immediate cause of death and medications administered before death. Specific binding of 125I-insulin-like growth factor I to homogenates prepared from cerebral cortex of Alzheimer's, alcoholic, alcoholic Alzheimer's, and age-matched control patients was similar, although Alzheimer's patients tended to have slightly higher binding values. No significant differences in insulin-like growth factor I binding in cerebral cortex were found with regard to age of patients, the interval between death and autopsy, and CNS-active medications. No statistical differences in 125I-insulin-like growth factor I binding were noted in hippocampal tissue from the four patient groups. Thus, human insulin-like growth factor I binding sites in cerebral cortex and hippocampus appear unaffected by several variables.     

Changes of Activity and Isozyme Pattern of Phosphofructokinase in the Brains of Patients with Alzheimer's Disease

Abstract: A severe reduction of the in vivo cerebral glucose consumption rate is generally found in patients with Alzheimer's disease. In postmortem studies changes in the activities of key regulatory glycolytic enzymes, including 6-phosphofructokinase (PFK), have been reported in Alzheimer's disease brains, but the results obtained so far are inconsistent and controversial. We reevaluated the activity of PFK in brain tissue from clinically and neuropathologically confirmed cases of Alzheimer's disease using optimized tissue disintegration and assay methods and determined the PFK isozyme pattern. PFK activity in brains from patients with Alzheimer's disease was significantly increased in frontal and temporal cortex and unchanged in the other brain areas studied when compared with control brains. All three PFK isozymes were detected in each of the brain areas studied. In brains of Alzheimer's disease patients the level of the C-type PFK was slightly reduced at the expense of the M- and L-type subunits. The data presented do not support the results of other groups, which reported up to a 90% reduction of PFK activity in Alzheimer's disease. In contrast, the data presented clearly rule out the suggestion that changes of PFK activity might be one of the causes for the reduced glucose consumption in Alzheimer's disease brains.     

Molecular Characteristics of Transgenic Wheat and the Effect on Transgene Expression

A population of R0 transgenic wheat plants, generated by particle bombardment, was analyzed to define molecular, genetic and phenotypic properties resulting from transformation with a cointegrate vector, or cotransformation with two separate plasmids. By evaluating the progeny of 70 independently-derived transgenic plants, we also identified rare events such as chimerism and transgene elimination, which provide valuable information concerning the development of transgenic cereal plants following bombardment experiments. The frequency of chimerism in our transgenic wheat plants was very low. Furthermore, while transgene elimination did occur, this was also a very rare event. We determined the copy numbers of integrated transgenes and the levels of transgene expression. Comparisons to transgenic rice plants generated in the same manner demonstrated some similarities, but also important differences in transgene behavior. Whereas in rice there is no evidence for any direct relationship between transgene copy number and transgene expression or stability, multicopy populations in wheat demonstrated a bias towards higher levels of expression for the two genes and the maize ubiquitin promoter evaluated in the present study.