老年大鼠脑细胞内钙的变化及β-淀粉样蛋白的影响

目的 观察老年大鼠脑细胞内钙浓度 (〔Ca2 + 〕i)的变化及βAP对〔Ca2 + 〕i的影响。方法 以Fura 2 /AM为荧光指示剂 ,采用荧光分光光度计测定急性分离的大鼠脑〔Ca2 + 〕i。结果 静息状态下老年大鼠脑〔Ca2 + 〕i及对高钾除极化的反应均明显低于青年对照组 (P <0 0 1 ) ,βAP对高钾除极化下脑〔Ca2 + 〕i的增加有明显增强作用 (P <0 0 1 ) ,但其增强作用在老年大鼠与青年大鼠间无明显差异 (P >0 0 5)。结论 老年大鼠脑〔Ca2 + 〕降低 ,βAP能使老年动物脑细胞内钙更明显超载 ,且在相同情况下老年神经元的受损更加严重。

血清中β淀粉样蛋白检测对老年性痴呆的临床意义

目的 :研究阿尔茨海默病 (AD)患者血清中 β淀粉样蛋白 (Aβ)含量及临床意义。方法 :采用放射免疫分析法 (RIA)检测 2 2例AD患者和 30例正常对照者血清中Aβ含量。结果 :AD患者血清中Aβ含量明显高于对照组 (P <0 0 5 ) ;以 72 3pg/ml作为正常人群Aβ含量的上限 ,诊断老年性痴呆的敏感性为6 3 6 % ,特异性为 83 3% ;经相关分析 ,血清Aβ含量与MMSE评分之间未发现显著相关关系 (P >0 0 5 )。结论 :AD患者血清中Aβ含量升高 ,对诊断老年性痴呆有较高的敏感性和特异性 ,有一定的临床应用价值。

糖尿病患者血浆β-淀粉样蛋白、雌二醇水平临床意义探讨

目的 :探讨糖尿病 (DM)作为老年性痴呆 (AD)危险因素的可能机理。方法 :放免法检测 DM组 5 8例 ,正常对照组33人的血浆 β-淀粉样蛋白 (Aβ)、雌二醇 (E2 )水平 ,DM组 2 6例脑 CT扫描。结果 :1 5例脑 CT皮质萎缩 ,脑室扩大。 DM组女性 E2 为 4 .4 6± 4 .31 pg/ml,对照组女性 E2 为 9.4 3± 7.87pg/ml,P<0 .0 1 ,两组有显著差异。DM组 Aβ为 5 8.37± 8.6 5 pg/m l,对照组 Aβ为 6 1 .1 5± 8.39pg/ml,两组无显著差异。结论 :DM作为 AD的危险因素的可能机理 :脑皮质萎缩 ;E2 水平显著降低 ;Aβ被晚期糖基化终末产物 (AGE)

神经节苷脂促进β-淀粉样蛋白生成及其二者协同分布

目的 :探讨神经节苷脂GM1干扰 β -淀粉样蛋白前体 (APP)正常代谢途径以及增加 β -淀粉样蛋白 (Aβ)生成的可能机制 ,以确立GM1在阿尔茨海默病 (AD)脑淀粉样变性过程中的特定作用。 方法 :选用人类 β -淀粉样蛋白前体基因 (APP6 95)转染细胞模型 ,用IP -Western印迹法观察GM1,对细胞APP代谢产物Aβ释放的影响 ;应用激光共聚焦扫描显微术 (LSCM )分别定量分析不同浓度GM1作用下细胞内Aβ40和Aβ42荧光强度的变化 ;通过双重免疫荧光标记确定GM1与Aβ4 0 和 /或Aβ4 2 的协同定位分布。结果 :GM1干扰细胞APP水解代谢和促进Aβ的释放 ;细胞内Aβ4 0 和Aβ4 2 荧光强度均随GM1浓度的增加而增强 ;内、外源性GM1分别与Aβ4 0 和Aβ4 2 在细胞内协同定位。结论 :提示GM1通过与Aβ的协同分布 。

载脂蛋白E与脑血管病

载脂蛋白E(apoE)是血液中最重要的载脂蛋白成分之一，它具有基因多态性，有六种遗传表现型。apoE与A1zheimer病(AD)的关系十分密韧，体外研究发现，apoEε3具有减少β—淀粉样蛋白(Aβ)积聚和保护tau蛋白磷酸化区，减少神经纤维缠结形成的作用，apoEε2的作用次之，apoEε4的作用最弱。apoE多态性与脑血管病的关系近几年越来越被重视。apoE为最常见的出血性卒中的危险指标；淀粉样脑血管病(CAA)相关的脑出血病病人可见ε2和ε4等位基因的过度量表达；ε4的脑出血(1CH)病人预后较差，其死亡率是无ε4的3倍；动脉硬化性脑血管病和冠心病的病变基础是相似的，所以叩oE也合影响脑血管病，动脉硬化脑梗塞(ACl)组ε3／3和ε3／4亚组Lp(a)均明显高于对照组，HDL低于对照组，ε3／4型Aβ组的LDL值高于对照组，揭示ε3／4与中青年ACI的发生有关，ε4也是中青年ACI的一种遗传易感性因子。CAA血管内沉积的Aβ与AD老年斑中的Aβ基本相似，93例尸检证实的中、重度CAA病人的ε4频率，与无ε4的病人相比增加2．9倍。13例重度CAA的ICH病人，ε4频率为62％，说明ε4在CAA的发生中起重要作用。ε4携带者的CAA病人，与无这一等位基因的病人相比，ICH的发病年龄平均提前5年以上。apoE导致脑出血的可能机制是，apoE基因的表达可导致正常分泌的Aβ在早期CAA血管壁上快速螯合和聚积。CAA分子基础的阐明有助于探索新的、有益的防治方法；在评估相对年轻(小于70岁)的可疑CAA病人时，apoE基因型的确认有助于CAA的诊断。因此，更深入地研究apoE多态性的病理学和分子生物学，将会清楚的阐明其与人类疾病的关系。

老年痴呆关联基因的研究进展

阿尔茨海默类痴呆(AD)是老年痴呆中最常见的一种,它以渐进性的神经功能退化并伴随着整体认知能力的下降为特征。早发性AD主要是由β-淀粉样前体蛋白(APP)基因和早老素基因突变引起,而与晚发性AD发病明显相关的只有载脂蛋白E-ε4(APOE-ε4)等位基因。但是APOE-ε4等位基因对AD发病既非充分又非必要,而且只能解释少于50％的AD的遗传变异。所以有必要进一步寻找与AD的关联基因。

阿尔茨海默病的药物治疗进展

对阿尔茨海默病 ( AD)药物治疗的新进展 ,如改善胆碱能神经传递 ,促进神经营养代谢 ,雌激素替代治疗 ,消炎镇痛药的应用 。

Targeted migration and differentiation of engrafted neural precursor cells in amyloid β-treated hippocampus in rats

Objective To observe the migration and differentiation of the neural precursor cells （NPCs） that derived from murine embryonic stem cells （ESCs） when they were transplanted into amyloid β （Aβ）-treated rat hippocampus. Methods MESPU35, a murine ESC cell line that express the enhanced green fluorescent protein （EGFP）, was induced differentiation into nestin-positive NPCs by modified serum-free methods. The Aβ plaques and the differentiation of the grafted cells were observed by immunofluorescent staining. Results Comparing 16 weeks with 4 weeks post-transplantation, the migration distance increased about 5 times; the rate of migratory NPCs differentiating into glial fibrillary acidic protein （GFAP）-positive cells kept rising from （30.41 ± 1.45）% to （49.25± 1.23）%, and the rate of NPCs differentiating into neurofilament 200 （NF200） positive cells increased from （16.68±0.95）% to （27.94± 1.21）%. Meanwhile, the GFAP-positive cells targeting to the ipsilateral side of Aβ plaques increased from 60.2% to 81.3 %, while the NF200-positive cells increased from 61.3% to 84.1%. The migration distance had significant positive linear correlations to the neuronal differentiation rate （r = 0.991） and to the astrocytic differentiation rate （r = 0.953）. Conclusion Engrafted NPCs migrate targetedly to the Aβ injection site and differentiate into neurons and astrocytes.

The Value of CSF Level of β- amyloid Protein in the Diagnosis of Alzheimer's Disease

Objective:To evaluate the diagnostic potential of cerebrospmal fluid (CSF) levels of β-amyloid protein (Aβ) as biochemical marker for senile dementia in clinical practice. Methods : Sensitive enzyme-linked immunosorbent assay (ELISA) was performed in our lalxrratory to delect the CSF levels of Aβt-40, Aβ1-42 in 54 patients with Alzlteimer's disease (AD), and 30 normal controls (NC). Results: The cut off value of Aβ ratio and Aβ1-42 concentration in NC group provided 54. 51%, 90. 00% sensitivity and 81. 25%, 84. 38% specificity respectively in diagnosis of AD. Conclusion : AD group had a significant decreased level of Aβ1-42 and an increased level of Aβ ratio, compared with NC group.

Alzheimer’s disease:epidemiology,genetics,and beyond

Alzheimer＇s disease （AD） is an increasing epidemic threatening public health. Both men and women are susceptible to the disease although women are at a slightly higher risk. The prevalence of AD rises exponentially in elderly people from 1% at age of 65 to approximately 40%-50% by the age of 95. While the cause of the disease has not been fully understood, genetics plays a role in the onset of the disease. Mutations in three genes （APP, PSENI, and PSEN2） have been found to cause AD and APOE4 allele increases the risk of the disease. As human genomic research progresses, more genes have been identified and linked with AD. Genetic screening tests for persons at high risk of AD are currently available and may help them as well as their families better prepare for a later life with AD.

Observation of amyloid precursor protein cleavage and Aβ generation in living cells by using multiphoton laser scanning microscopy

Objective To investigate the proteolytic mechanism of amyloid precursor protein （APP） and to explore amyloidbeta （Aβ） generation in living neurons. Methods DNA fragments were amplified by PCR or synthesized. The four fragments, CFP, 54bp, YFP and C99 were ligated into pcDNA3.0 vector to construct the recombinant plasmids pcDNA3.0-CFP-54bp- YFP and pcDNA3.0-CFP-54bp-YFP-C99. The SH-SY5Y cells were transiently transfected with pcDNA3.0-CFP-54bp-YFP or pcDNA3.0-CFP-54bp-YFP-C99. The expression of fusion gene was examined under a multiphoton laser scanning microscope. Fluorescence resonance energy transfer （FRET） was used to measure the β cleavage and γ cleavage of APE Aβ generation was confirmed by immunocytochemistry and multiphoton laser scanning microscopy. Cell viability was tested by MTT assay at different time points. Results （1） The double restriction endonuclease digestion and sequencing analysis confirmed the authenticity of the recombinant plasmids pcDNA3.0-CFP-54bp-YFP and pcDNA3.0-CFP-54bp- YFP-C99. （2） Blue and yellow fluorescences were detected in the transfected cells. （3） FRET occurred in pcDNA3.0-CFP- 54bp-YFP-transfected cells but not in pcDNA3.0-CFP-54bp-YFP-C99-transfected cells. （4） Aβ was produced in the pcDNA3.0- CFP-54bp-YFP-C99 transfected cells. （5） Aβ-deposition was widespread in the cell. （6） Cell viability decreased along with the intracellular Aβ deposition. Conclusion C99 is important for the APP β cleavage. Aβ may be generated and deposited in cells at the early stage of Alzheimer＇s disease. Intracellular Aβ accumulation brings deleterious effects on cells.

Receptor tyrosine kinases positively regulate BACE activity and Amyloid-β production through enhancing BACE internalization

Amyloid-β （Aβ） peptide, the primary constituent of senile plaques in Alzheimer＇s disease （AD）, is generated by β-secretase- and y-secretase-mediated sequential proteolysis of the amyloid precursor protein （APP）. The aspartic protease, β -site APP cleavage enzyme （BACE）, has been identified as the main β-secretase in brain but the regulation of its activity is largely unclear. Here, we demonstrate that both BACE activity and subsequent Aβ production are enhanced after stimulation of receptor tyrosine kinases （RTKs）, such as the receptors for epidermal growth factor （EGF） and nerve growth factor （NGF）, in cultured cells as well as in mouse hippocampus. Furthermore, stimulation of RTKs also induces BACE internalization into endosomes and Golgi apparatus. This enhancement of BACE activity and A β production upon RTK activation could be specifically inhibited by Src family kinase inhibitors and by depletion of endogenous c-Src with RNAi, and could be mimicked by over-expressed c-Src. Moreover, blockage of BACE internalization by a dominant negative form of Rab5 also abolished the enhancement of BACE activity and Aβ production, indicating the requirement of BACE internalization for the enhanced activity. Taken together, our study presents evidence that BACE activity and Aβ production are under the regulation of RTKs and this is achieved via RTK-stimulated BACE internalization, and suggests that an aberration of such regulation might contribute to pathogenic Aβ production.

Structural Dynamics of Amyloid β Peptide Binding to Acetylcholine Receptor and Virtual Screening for Effective Inhibitors

The interaction between Amyloid β(Aβ) peptide and acetylcholine receptor is the key for our understanding of how Aβ fragments block the ion channels within the synapses and thus induce Alzheimer’s disease.Here,molecular docking and molecular dynamics(MD)simulations were performed for the structural dynamics of the docking complex consisting of Aβ and α7-n ACh R(α7 nicotinic acetylcholine receptor),and the inter-molecular interactions between ligand and receptor were revealed.The results show that Aβ_(25-35) is bound toα7-n ACh R through hydrogen bonds and complementary shape,and the Aβ_(25-35) fragments would easily assemble in the ion channel of α7-n ACh R,then block the ion transfer process and induce neuronal apoptosis.The simulated amide-I band of Aβ_(25-35) in the complex is located at 1650.5 cm^(-1),indicating the backbone of Aβ_(25-35) tends to present random coil conformation,which is consistent with the result obtained from cluster analysis.Currently existing drugs were used as templates for virtual screening,eight new drugs were designed and semi-flexible docking was performed for their performance.The results show that,the interactions between new drugs and α7-n ACh R are strong enough to inhibit the aggregation of Aβ_(25-35) fragments in the ion channel,and also be of great potential in the treatment of Alzheimer’s disease.

Tetrahydroxy stilbene glucoside reduces the cognitive impairment and overexpression of amyloid precursor protein induced by aluminum exposure

Objective Excessive aluminum （Al） exposure impairs neurocognitive function in humans and animals. Epidemiologic studies have shown a potential linkage between chronic Al exposure and Alzheimer’s disease. The present study aims to evaluate the effects of tetrahydroxy stilbene glucoside （TSG）, the extract from herbal medicine Polygoni Multiflori, on cognitive impairment and the over-expression of hippocampal amyloid precursor protein （APP） induced by chronic exposure to Al in rats. Methods Rats were treated with 0.3% aluminum chloride （AlCl3） prepared in the drinking water for 90 d. AlCl3-treated animals were then randomly assigned to receive vehicle, TSG （4 g/kg）, or Vitamin E （VE; 40 mg/kg） treatment for 5 months. VE served as a positive control. The effect of TSG was evaluated by passive avoidance task, and APP expression was evaluated by Western blotting. Results Following exposure to AlCl3 for 90 d, animals displayed a striking decrease （〉80%） in step-through latency in the passive avoidance task and a significant increase in the expression of APP in the hippocampus. Both TSG and VE significantly ameliorated the performance impairment in the passive avoidance task, and suppressed the over-expression of APP. Moreover, the effects of TSG, but not of VE, were in a time-dependent manner. Conclusion TSG may possess therapeutic effects against Alzheimer’s disease.

Effects of free radicals and amyloid β protein on the currents of expressed rat receptors in Xenopus oocytes

Objective To investigate the effects of free radicals (FRs) and amyloid β protein 1 40 (Aβ 1 40 ) on the functions of expressed neurotransmitter receptors (NRs) in Xenopus oocytes Methods Total RNA and messenger RNA (mRNA) was prepared from 3 month old Wistar rat brain tissues with Promega kits and microinjected into maturated Xenopus oocytes (stages Ⅴ Ⅵ) with 50?nl (50?ng) for each oocyte The microinjected oocytes were incubated with modified Bath's solution at 19 0℃±1 0℃ for receptor expression and their currents were recorded with double electrode voltage clamp technique Superoxide anion free radicals (SAFRs) were produced via a reaction system (HPX/XO) with hypoxanthine (HPX, 0 05?mol/L) and xanthine oxidase (XO, 0 1?U/L) In order to observe the effects of Aβ and SAFRs on the expressed glutamate receptor, HPX/XO and Aβ 1 40 were added to incubation solution at 12?h, 24?h and 96?h before recording Results The results showed that the oocytes expressed functional NRs originating from rat brain tissues These NRs included muscarinic acetylcholine (mACh), glutamate (Glu), dopamine (DA), serotonin (5 HT) and γ aminobutyric acid (GABA) The current characteristics of expressed receptors were inward currents carried by chloride ion with their equibrilium potentials close to -22?mV The extent of effect on the current of expressed glutamate receptor from rat brain was different among different Aβ concentrations and incubation times Aβ 1 40 at a concentration of 20?nmol/L had little effect on the currents of expressed rat brain glutamate receptors up to 24?h of incubation period; but the currents of glutamate receptor were significantly decreased (25% off, P <0 01) in the treatment of 60?nmol/L Aβ 1 40 over 24?h Moreover, when 20?nmol/L Aβ 1 40 was co incubated over 12?h with SAFRs produced by the reaction system of HPX/XO, it was found that the currents of expressed rat brain glutamate receptors had been changed markedly When the oocytes were co treated with 60?nmol/L Aβ 1 40 and SAFRs over a period of 12?h, the currents of glutamate receptor significantly decreased (21% off, P <0 05), and the decreased percentage reached 52% over 24?h co treatment with 60?nmol/L Aβ 1 40 and SAFRs In addition, vitamin E had a partial effect against this inhibitory effect Conclusion The results suggest that Aβ has a kind of inhibitory effect upon the current of the glutamate receptor, similar to the effects of free radicals The effects can be antagonized by vitamin E These imply that Aβ may play a role via inhibiting receptor function in the pathophysiology of Alzheimer's disease

Vanadyl complexes work with cinnamaldehyde in promoting cell viability under the β-amyloid burden in SH-SY5Y neural cells

The Alzheimer＇s disease （AD） is one of the common cognitive disorders in the elderly. AD shares some similar pathological characters with diabetes mellitus （DM）, suggesting potential application of anti-diabetic agents, such as vanadyl complexes, in therapeutic treatment of AD. In the present work, we studied the effects of vanadyl acetylacetonate （VO（acac）2） and cinnamaldehyde （CA） on an AD model based on SH-SY5Y neural cells. The experimental results showed that VO（acac）2 at sub-micromolar concentrations could improve the viability of neural cells with or without increased β-amyloid （Aβ） burden; and the combination of VO（acac）2 and CA showed an additive cell protection effects. Further investigation revealed that for SH-SY5Y neural cells, VO（acac）2 could activate PPART-AMPK signal transduction and inhibit GSK 3β, one of the major kinases for Tau hyperphosphorylation. Meanwhile, CA could correct the abnormal mitochondrial morphology due to Aβ-induced excessive mitochondrial fission, thus restoring/enhancing the mitochondrial function. In addition, both VO（acac）2 and CA decreased intracellular reactive oxygen species （ROS） level and inhibited formation of toxic Aβ oligomers. Overall, VO（acac）2 might work with CA in improving the neural cell viability under the Aβ burden, suggesting application of vanadium metallodrugs in AD treatment.

Regulation of β cleavage of amyloid precursor protein

Alzheimer’s disease ranks the first cause for senile dementia.The amyloid cascade is proposed to contribute to the pathogenesis of this disease.In this cascade,amyloid β peptide（Aβ）is produced through a sequential cleavage of amyloid precursor protein（APP）by β and γ secretases,while its cleavage by α secretase precludes Aβ production and generates neurotrophic sAPPα.Thus,enhancing α secretase activity or suppressing β and γ cleavage may reduce Aβ formation and ameliorate the pathological process of the disease.Several regulatory mechanisms of APP cleavage have been established. The present review mainly summarizes the signaling pathways pertinent to the regulation of APP β cleavage.

Monocytes and Alzheimer's disease

Alzheimer＇s disease（AD） is a neurodegenerative disease characterized by extracellular amyloid beta（Aβ） deposition and intracellular neurofibrillary tangle formation.Monocyte is part of the innate immune system and can effectively remove dead cells and debris.It has been suggested that Aβ can recruit monocytes into brain in AD mice,resulting in restriction of cerebral amyloidosis.However,monocyte may act as a double-edged sword,either beneficial（e.g.,clearance of Aβ） or detrimental（e.g.,secretion of neurotoxic factors）.In addition,recent studies indicate that in AD patients,Aβ phagocytosis by monocytes is ineffective.The present review mainly summarized the current knowledge on monocytes and their potential roles in AD.

Presenilins as endoplasmic reticulum calcium leak channels and Alzheimer's disease pathogenesis

Alzheimer disease(AD) is the most common neurodegenerative disorder worldwide and is at present,incurable.The accumulation of toxic amyloid-beta(Aβ) peptide aggregates in AD brain is thought to trigger the extensive synaptic loss and neurodegeneration linked to cognitive decline,an idea that underlies the'amyloid hypothesis'of AD etiology in both the familal(FAD) and sporadic forms of the disease.Genetic mutations causing FAD also result in the dysregulation of neuronal calcium(Ca2+) handling and may contribute to AD pathogenesis,an idea termed the'calcium hypothesis'of AD.Mutations in presenilin proteins account for majority of FAD cases.Presenilins function as catalytic subunit ofγ-secretase involved in generation of Aβ peptide Recently,we discovered that presenilns function as low-conductance,passive ER Ca2+ leak channels,independent of γ-secretase activity.We further discovered that many FAD mutations in presenilins result in loss of ER Ca2+ leak function activity and Ca2+ overload in the ER.These results provided potential explanation for abnormal Ca2+ signaling observed in FAD cells with mutations in presenilns.Our latest work on studies of ER Ca2+ leak channel function of presenilins and implications of these findings for understanding AD pathogenesis are discussed in this article.

Influence of electroacupuncture therapy of tonifying the kidney and regulating governor vessel on Aβ related degradation enzymes in the hippocampus of a rat model of Alzheimer's disease induced by Aβ_(1-42)

Objective： To explore influence of electroacupuncture（EA） therapy of tonifying the kidney and regulating governor vessel on amyloid beta（Aβ） related degradation enzymes in the hippocampus of a rat model of Alzheimer＇s disease（AD） induced by Aβ（1-42）.Methods： Forty Wistar male rats were randomly divided into 4 groups： a normal group, a sham operation group, a model group and an EA group, 10 rats in each one. The rats in normal group were normally fed. The rats in sham operation group were bilaterally injected in the hippocampus with 5 μL of saline and they were normally fed after the injection. The rats in the model group and the EA group were bilaterally injected in the hippocampus with 5 μL of Aβ（1-42） on each side. Rats in the EA group received EA of 5 Hz continuous wave at the ＂Bǎihuì（百会 GV20）＂ and bilateral ＂Shènshū（肾俞 BL23）＂ for a duration of 15 min per time every day and continuously for 15 days. After 15 days, the hippocampal expression levels of insulin degrading enzyme（IDE）, lipoprotein（LPL）, transthyretin（TTR）, apolipoprotein E（APoE）,a2 macroglobulin（a2 M） and Aβ（1-42） of the 4 groups were tested by Western blot.Results： Compared with the sham operation group, the expression levels of IDE, LPL, TTR, APoE and a2 M in the hippocampus were significantly lower（P〈 0.05, P〈 0.01） and the expression of Aβ（1-42） was significantly higher（P〈 0.01） in the model group. Compared with the model group, the expression levels of IDE, LPL, TTR,APoE and a2 M in the hippocampus of these rats were significantly lower（P〈 0.05,P〈 0.01）, the expression of Aβ（1-42） was significantly higher（P〈 0.01） in the EA group.Conclusion： EA therapy of tonifying the kidney and regulating governor vessel can enhance the expression of IDE, LPL, TTR, APoE, and a2 M in the hippocampus of AD rats injected by Aβ（1-42）, and may consequently promote the degradation of aβ（1-42） to help improve the pathological manifestations of AD and therefore delay its progression.