Amyloid Deposition In Alzheimer's Disease Research Articles

The role of fibrinogen glycation in ATTR: evidence for chaperone activity loss in disease.

Transthyretin amyloidosis (ATTR) belongs to a class of disorders caused by protein misfolding and aggregation. ATTR is a disabling disorder of autosomal dominant trait, where transthyretin (TTR) forms amyloid deposits in different organs, causing dysfunction of the peripheral nervous system. We previously discovered that amyloid fibrils from ATTR patients are glycated by methylglyoxal. Even though no consensus has been reached about the actual role of methylglyoxal-derived advanced glycation end-products in amyloid diseases, evidence collected so far points to a role for protein glycation in conformational abnormalities, being ubiquitously found in amyloid deposits in Alzheimer's disease, dialysis-related amyloidosis and Parkinson's diseases. Human fibrinogen, an extracellular chaperone, was reported to specifically interact with a wide spectrum of stressed proteins and suppress their aggregation, being an interacting protein with TTR. Fibrinogen is differentially glycated in ATTR, leading to its chaperone activity loss. Here we show the existence of a proteostasis imbalance in ATTR linked to fibrinogen glycation by methylglyoxal.

Anti-amyloidogenic property of human gastrokine 1.

Gastrokine 1 (GKN1) is a stomach-specific protein expressed in normal gastric tissue but absent in gastric cancer. GKN1 plays a major role in maintaining gastric mucosa integrity and is characterized by the presence of a BRICHOS domain consisting of about 100 amino acids also found in several unrelated proteins associated with major human diseases like BRI2, related to familial British and Danish dementia and surfactant protein C (SP-C), associated with respiratory distress syndrome. It was reported that recombinant BRICHOS domains from BRI2 and SP-C precursor (proSP-C) prevent fibrils formation of amyloid-beta peptide (Aβ), that is the major component of extracellular amyloid deposits in Alzheimer's disease. Here we investigated on the interaction between human recombinant GKN1 (rGKN1) and Aβ peptide (1-40) that derives from the partial hydrolysis of the amyloid precursor protein (APP). GKN1 prevented amyloid aggregation and fibrils formation by inhibiting Aβ(1-40) polymerization, as evaluated by SDS-PAGE, thioflavin-T binding assay and gel filtration experiments. Mass spectrometry showed the formation of a prevailing 1:1 complex between GKN1 and Aβ(1-40). SPR analysis of GKN1/Aβ interaction led to calculate a dissociation constant (KD) of 34μM. Besides its interaction with Aβ(1-40), GKN1 showed also to interact with APP as evaluated by confocal microscopy and Ni-NTA pull-down. Data strongly suggest that GKN1 has anti-amyloidogenic properties thus functioning as a chaperone directed against unfolded segments and with the ability to recognize amyloidogenic polypeptides and prevent their aggregation.

Multifunctional nanoliposomes with curcumin–lipid derivative and brain targeting functionality with potential applications for Alzheimer disease

With the objective to formulate multifunctional nanosized liposomes to target amyloid deposits in Alzheimer Disease (AD) brains, a lipid–PEG–curcumin derivative was synthesized and characterized. Multifunctional liposomes incorporating the curcumin derivative and additionally decorated with a Blood Brain Barrier (BBB) transport mediator (anti-Transferin antibody) were prepared and characterized. The fluorescence intensity of curcumin derivative was found to increase notably when the curcumin moiety was in the form of a diisopropylethylamine (DIPEA) salt. Both curcumin-derivative liposomes and curcumin-derivative Anti-TrF liposomes showed a high affinity for the amyloid deposits, on post-mortem brains samples of AD patients. The ability of both liposomes to delay Aβ1-42 peptide aggregation was confirmed by Thioflavin assay. However, the decoration of the curcumin-derivative liposomes with the Anti-TrF improved significantly the intake by the BBB cellular model. Results verify that the attachment of an antibody on the curcumin-liposome surface does not block deposit staining or prevention of Aβ aggregation, while the presence of the curcumin–PEG–lipid conjugate does not reduce their brain-targeting capability substantially, proving the potential of such multifunctional NLs for application in Alzheimer disease treatment and diagnosis.

Age-Related Brain Expression and Regulation of the Chemokine CCL4/MIP-1β in APP/PS1 Double-Transgenic Mice

The detrimental effect of activation of the chemokine CCL4/MIP-1β on neuronal integrity in patients with HIV-associated dementia has directed attention to the potential role of CCL4 expression and regulation in Alzheimer disease. Here, we show that CCL4 mRNA and protein are overexpressed in the brains of APPswe/PS1ΔE9 (APP/PS1) double-transgenic mice, a model of cerebral amyloid deposition; expression was minimal in brains from nontransgenic littermates or single-mutant controls. Increased levels of CCL4 mRNA and protein directly correlated with the age-related progression of cerebral amyloid-β (Aβ) levels in APP/PS1 mice. We also found significantly increased expression of activating transcription factor 3 (ATF3), which was positively correlated with age-related Aβ deposition and CCL4 in the brains of APP/PS1 mice. Results from chromatin immunoprecipitation-quantitative polymerase chain reaction confirmed that ATF3 binds to the promoter region of the CCL4 gene, consistent with a potential role in regulating CCL4 transcription. Finally, elevated ATF3 mRNA expression in APP/PS1 brains was associated with hypomethylation of the ATF3 gene promoter region. These observations prompt the testable hypothesis for future study that CCL4 overexpression, regulated in part by hypomethylation of the ATF3 gene, may contribute to neuropathologic progression associated with amyloid deposition in Alzheimer disease.

LRP-1 polymorphism is associated with global and regional amyloid load in Alzheimer's disease in humans in-vivo

ObjectiveImpaired amyloid clearance has been proposed to contribute to β-amyloid deposition in sporadic late-onset Alzheimer's disease (AD). Low density lipoprotein receptor-related protein 1 (LRP-1) is involved in the active outward transport of β-amyloid across the blood–brain barrier (BBB). The C667T polymorphism (rs1799986) of the LRP-1 gene has been inconsistently associated with AD in genetic studies.We aimed to elucidate the association of this polymorphism with in-vivo brain amyloid load of AD patients using amyloid PET with [11C]PiB.Materials and methods72 patients with very mild to moderate AD were examined with amyloid PET and C667T polymorphism was obtained using TaqMan PCR assays. The association of C667T polymorphism with global and regional amyloid load was calculated using linear regression and voxel based analysis, respectively. The effect of the previously identified modulator of amyloid uptake, the apolipoprotein E genotype, on this association was also determined.ResultsThe regression analysis between amyloid load and C667T polymorphism was statistically significant (p = 0.046, β = 0.236). In an additional analysis ApoE genotype and gender were identified to explain further variability of amyloid load. Voxel based analysis revealed a significant (p < 0.05) association between C667T polymorphism and amyloid uptake in the temporo-parietal cortex bilaterally. ApoE did not interact significantly with the LRP-1 polymorphism.DiscussionIn conclusion, C667T polymorphism of LRP-1 is moderately but significantly associated with global and regional amyloid deposition in AD. The relationship appears to be independent of the ApoE genotype. This finding is compatible with the hypothesis that impaired amyloid clearance contributes to amyloid deposition in late-onset sporadic AD.

Pyroglutamate-Modified Amyloid-β Protein Demonstrates Similar Properties in an Alzheimer's Disease Familial Mutant Knock-In Mouse and Alzheimer's Disease Brain

Background: N-terminally truncated, pyroglutamate-modified amyloid-β (Aβ) peptides are major constituents of amyloid deposits in Alzheimer's disease (AD). Methods: Using a newly developed ELISA for Aβ modified at glutamate 3 with a pyroglutamate (pE3Aβ), brain pE3Aβ was characterized in human AD in an AD mouse model harboring double knock-in amyloid precursor protein (APP)-KM670/671NL and presenilin 1 (PS1)-P264L (APP/PS1-dKI) mutations, and in a second mouse model with transgenic overexpression of human APP695 with APP-KM670/671NL (Tg2576). Results: pE3Aβ increased in the AD brain versus age-matched controls, with pE3Aβ/total Aβ at 45 and 10%, respectively. Compared to controls, the AD brain demonstrated 8.5-fold increased pE3Aβ compared to non-pE3Aβ species, which increased 2.7-fold. In the APP/PS1-dKI brain, pE3Aβ/total Aβ increased from 7% at 3 months to 16 and 19% at 15 and 19 months, respectively. In Tg2576, pE3Aβ/total Aβ was only 1.5% at 19 months, suggesting that APP/PS1-dKI, despite less total Aβ compared to Tg2576 at comparable ages, more closely mimics AD brain pathology. Conclusion: This report supports a significant role for pE3Aβ in AD pathogenesis by confirming that pE3Aβ represents a large fraction of Aβ within the AD brain. Compared to the age-matched control brain, pE3Aβ increased to a greater extent compared to Aβ species without this N-terminal modification. Further, the APP/PS1-dKI model more closely resembles the AD brain in this regard, compared to the Tg2576 model.

PET Approaches for Diagnosis of Dementia

There is increasing use of neuroimaging modalities, including PET, for diagnosing dementia. For example, FDG-PET demonstrates hypometabolic regions in the posterior cingulate gyri, precuneus, and parietotemporal association cortices, while amyloid PET indicates amyloid deposition in Alzheimer disease and mild cognitive impairment due to Alzheimer disease. Furthermore, the use of combination PET with structural MR imaging can improve the diagnostic accuracy of dementia. In other neurodegenerative dementias, each disease exhibits a specific metabolic reduction pattern. In dementia with Lewy bodies, occipital glucose metabolism is decreased, while in frontotemporal dementia, frontal and anterior temporal metabolism is predominantly decreased. These FDG-PET findings and positive or negative amyloid deposits are important biomarkers for various neurodegenerative dementias.

Charged surfactants induce a non‐fibrillar aggregation pathway of amyloid‐beta peptide

The amyloid β-peptide with a sequence of 42 amino acids is the major constituent of extracellular amyloid deposits in Alzheimer's disease plaques. The control of the peptide self-assembly is difficult to achieve because the process is fast and is affected by many variables. In this paper, we describe the effect of different charged and non-charged surfactants on Aβ(₁₋₄₂) fibrillation to define common alternate aggregation pathways. The characterization of the peptide-surfactant interactions by ultra-structural analysis, thioflavin T assay and secondary structure analysis, suggested that charged surfactants interact with Aβ(₁₋₄₂) through electrostatic interactions. Charged micelles slow down the aggregation process and stabilize the peptide in the oligomeric state, whereas non-charged surfactants promote the Aβ(₁₋₄₂) fibril formation.

Matrix Metalloproteinase-9 Reduces Islet Amyloid Formation by Degrading Islet Amyloid Polypeptide

Deposition of islet amyloid polypeptide (IAPP) as amyloid is a pathological hallmark of the islet in type 2 diabetes, which is toxic to β-cells. We previously showed that the enzyme neprilysin reduces islet amyloid deposition and thereby reduces β-cell apoptosis, by inhibiting fibril formation. Two other enzymes, matrix metalloproteinase (MMP)-2 and MMP-9, are extracellular gelatinases capable of degrading another amyloidogenic peptide, Aβ, the constituent of amyloid deposits in Alzheimer disease. We therefore investigated whether MMP-2 and MMP-9 play a role in reducing islet amyloid deposition. MMP-2 and MMP-9 mRNA were present in mouse islets but only MMP-9 activity was detectable. In an islet culture model where human IAPP (hIAPP) transgenic mouse islets develop amyloid but nontransgenic islets do not, a broad spectrum MMP inhibitor (GM6001) and an MMP-2/9 inhibitor increased amyloid formation and the resultant β-cell apoptosis. In contrast, a specific MMP-2 inhibitor had no effect on either amyloid deposition or β-cell apoptosis. Mass spectrometry demonstrated that MMP-9 degraded amyloidogenic hIAPP but not nonamyloidogenic mouse IAPP. Thus, MMP-9 constitutes an endogenous islet protease that limits islet amyloid deposition and its toxic effects via degradation of hIAPP. Because islet MMP-9 mRNA levels are decreased in type 2 diabetic subjects, islet MMP-9 activity may also be decreased in human type 2 diabetes, thereby contributing to increased islet amyloid deposition and β-cell loss. Approaches to increase islet MMP-9 activity could reduce or prevent amyloid deposition and its toxic effects in type 2 diabetes.

APP mutations in the Aβ coding region are associated with abundant cerebral deposition of Aβ38

Aβ is the main component of amyloid deposits in Alzheimer disease (AD) and its aggregation into oligomers, protofibrils and fibrils is considered a seminal event in the pathogenesis of AD. Aβ with C-terminus at residue 42 is the most abundant species in parenchymal deposits, whereas Aβ with C-terminus at residue 40 predominates in the amyloid of the walls of large vessels. Aβ peptides with other C-termini have not yet been thoroughly investigated. We analysed Aβ38 in the brains of patients with Aβ deposition linked to sporadic and familial AD, hereditary cerebral haemorrhage with amyloidosis, or Down syndrome. Immunohistochemistry, confocal microscopy, immunoelectron microscopy, immunoprecipitation and the electrophoresis separation of low molecular weight aggregates revealed that Aβ38 accumulates consistently in the brains of patients carrying APP mutations in the Aβ coding region, but was not detected in the patients with APP mutations outside the Aβ domain, in the patients with presenilin mutations or in subjects with Down syndrome. In the patients with sporadic AD, Aβ38 was absent in the senile plaques, but it was detected only in the vessel walls of a small subset of patients with severe cerebral amyloid angiopathy. Our results suggest that APP mutations in the Aβ coding region favour Aβ38 accumulation in the brain and that the molecular mechanisms of Aβ deposition in these patients may be different from those active in patients with familial AD associated with other genetic defects and sporadic AD.

Neuroinflammation and Common Mechanism in Alzheimer’s Disease and Prion Amyloidosis: Amyloid-Associated Proteins, Neuroinflammation and Neurofibrillary Degeneration

Background: In cases with a long (>1 year) clinical duration of prion disease, the prion protein can form amyloid deposits. These cases do not show accumulation of 4-kDa β-amyloid, which is observed in amyloid deposits in Alzheimer’s disease (AD). In AD, amyloid is associated with inflammation and neurofibrillary degeneration, and it is elusive whether prion amyloid is associated with these changes as well. Objectives: The presence of inflammation and neurofibrillary degeneration was evaluated in prion amyloidosis. Material and Methods: Cortical areas of variant Creutzfeldt-Jakob disease (CJD; n = 3), young sporadic CJD (n = 4), different Gerstmann-Sträussler-Scheinker’s disease patients (n = 5) and AD cases (n = 5) were examined using immunohistochemistry and specific stainings for amyloid. Results: In both AD and prion disease cases, which were negative for 4-kDa β-amyloid, parenchymal and vascular amyloid deposits were positive for amyloid-associated proteins such as complement protein and were associated with microglia clusters. Tau and ubiquitin were found near prion plaques in some of the Gerstmann-Sträussler-Scheinker’s disease and sporadic CJD cases and also near vascular prion amyloid deposits. In variant CJD cases, occasionally, microglia clustering was found in plaques but no ubiquitin or complement proteins and hardly tau protein. Conclusions: In both AD and prion disease amyloid formation, irrespective of the protein involved, there seems to be a neuroinflammatory response with secondary neurofibrillary degeneration.

Biochemistry of amyloid β-protein and amyloid deposits in Alzheimer disease.

Progressive cerebral deposition of the amyloid β-protein (Aβ) in brain regions serving memory and cognition is an invariant and defining feature of Alzheimer disease. A highly similar but less robust process accompanies brain aging in many nondemented humans, lower primates, and some other mammals. The discovery of Aβ as the subunit of the amyloid fibrils in meningocerebral blood vessels and parenchymal plaques has led to innumerable studies of its biochemistry and potential cytotoxic properties. Here we will review the discovery of Aβ, numerous aspects of its complex biochemistry, and current attempts to understand how a range of Aβ assemblies, including soluble oligomers and insoluble fibrils, may precipitate and promote neuronal and glial alterations that underlie the development of dementia. Although the role of Aβ as a key molecular factor in the etiology of Alzheimer disease remains controversial, clinical trials of amyloid-lowering agents, reviewed elsewhere in this book, are poised to resolve the question of its pathogenic primacy.

Amyloid pathway-based candidate gene analysis of [11C]PiB-PET in the Alzheimer’s Disease Neuroimaging Initiative (ADNI) cohort

Amyloid imaging with [11 C]Pittsburgh Compound-B (PiB) provides in vivo data on plaque deposition in those with, or at risk for, Alzheimer’s disease (AD). We performed a gene-based association analysis of 15 quality-controlled amyloid-pathway associated candidate genes in 103 Alzheimer’s Disease Neuroimaging Initiative participants. The mean normalized PiB uptake value across four brain regions known to have amyloid deposition in AD was used as a quantitative phenotype. The minor allele of an intronic SNP within DHCR24 was identified and associated with a lower average PiB uptake. Further investigation at whole-brain voxel-wise level indicated that non-carriers of the minor allele had higher PiB uptake in frontal regions compared to carriers. DHCR24 has been previously shown to confer resistance against beta-amyloid and oxidative stress-induced apoptosis, thus our findings support a neuroprotective role. Pathway-based genetic analysis of targeted molecular imaging phenotypes appears promising to help elucidate disease pathophysiology and identify potential therapeutic targets.

POD14 Amyloid PET using 18F-AV-45 in Alzheimer's disease and frontotemporal dementia: first UK results

Introduction18F-AV-45 (Florbetapir F-18) is a novel 18F-labelled positron emission tomography (PET) tracer for in vivo imaging of cerebral amyloid deposition. Here we present preliminary results for differentiation between Alzheimer's disease...

Functional alterations in memory networks in early Alzheimer's disease.

The hallmark clinical symptom of early Alzheimer's disease (AD) is episodic memory impairment. Recent functional imaging studies suggest that memory function is subserved by a set of distributed networks, which include both the medial temporal lobe (MTL) system and the set of cortical regions collectively referred to as the default network. Specific regions of the default network, in particular, the posteromedial cortices, including the precuneus and posterior cingulate, are selectively vulnerable to early amyloid deposition in AD. These regions are also thought to play a key role in both memory encoding and retrieval, and are strongly functionally connected to the MTL. Multiple functional magnetic resonance imaging (fMRI) studies during memory tasks have revealed alterations in these networks in patients with clinical AD. Similar functional abnormalities have been detected in subjects at-risk for AD, including those with genetic risk and older individuals with mild cognitive impairment. Recently, we and other groups have found evidence of functional alterations in these memory networks even among cognitively intact older individuals with occult amyloid pathology, detected by PET amyloid imaging. Taken together, these findings suggest that the pathophysiological process of AD exerts specific deleterious effects on these distributed memory circuits, even prior to clinical manifestations of significant memory impairment. Interestingly, some of the functional alterations seen in prodromal AD subjects have taken the form of increases in activity relative to baseline, rather than a loss of activity. It remains unclear whether these increases in fMRI activity may be compensatory to maintain memory performance in the setting of early AD pathology or instead, represent evidence of excitotoxicity and impending neuronal failure. Recent studies have also revealed disruption of the intrinsic connectivity of these networks observable even during the resting state in early AD and asymptomatic individuals with high amyloid burden. Research is ongoing to determine if these early network alterations will serve as sensitive predictors of clinical decline, and eventually, as markers of pharmacological response to potential disease-modifying treatments for AD.

Alterations in microglial activation and amyloid deposits in Alzheimer's disease

Alterations in microglial activation and amyloid deposits in Alzheimer's disease

Principal component analysis with pre-normalization improves the signal-to-noise ratio and image quality in positron emission tomography studies of amyloid deposits in Alzheimer's disease

This study introduces a new approach for the application of principal component analysis (PCA) with pre-normalization on dynamic positron emission tomography (PET) images. These images are generated using the amyloid imaging agent N-methyl [11C]2-(4′-methylaminophenyl)-6-hydroxy-benzothiazole ([11C]PIB) in patients with Alzheimer's disease (AD) and healthy volunteers (HVs). The aim was to introduce a method which, by using the whole dataset and without assuming a specific kinetic model, could generate images with improved signal-to-noise and detect, extract and illustrate changes in kinetic behavior between different regions in the brain. Eight AD patients and eight HVs from a previously published study with [11C]PIB were used. The approach includes enhancement of brain regions where the kinetics of the radiotracer are different from what is seen in the reference region, pre-normalization for differences in noise levels and removal of negative values. This is followed by slice-wise application of PCA (SW-PCA) on the dynamic PET images. Results obtained using the new approach were compared with results obtained using reference Patlak and summed images. The new approach generated images with good quality in which cortical brain regions in AD patients showed high uptake, compared to cerebellum and white matter. Cortical structures in HVs showed low uptake as expected and in good agreement with data generated using kinetic modeling. The introduced approach generated images with enhanced contrast and improved signal-to-noise ratio (SNR) and discrimination power (DP) compared to summed images and parametric images. This method is expected to be an important clinical tool in the diagnosis and differential diagnosis of dementia.

In vivo mapping of amyloid toxicity in Alzheimer disease

To study the relationship between gray matter atrophy and amyloid deposition in Alzheimer disease (AD). Volumetric magnetic resonance (MR) and [11C]-PIB PET were acquired from 23 patients with AD and 17 healthy older persons. Standardized [11C]-PIB uptake values were coregistered to MR scans in a standard space. Decreased density of and increased [11C]-PIB uptake in the gray matter of patients with AD vs controls were assessed with both voxel-based (p < 0.05 corrected) and region-of-interest (ROI) analyses. The relationship between decreased density of and increased [11C]-PIB uptake in the gray matter was investigated with voxel-based Pearson r maps (thresholded at p < 0.05) and ROI linear regression plots. Atrophy mapped to the hippocampus and increased [11C]-PIB uptake to large frontal, parietal, and posterior cingulate cortical areas. ROI analysis showed the largest effect size for atrophy in the hippocampus (2.01) and amygdala (1.27) and the highest effect size for [11C]-PIB uptake in frontal (2.66), posterior cingulate/retrosplenial (2.43), insular (2.41), and temporal (2.23) regions. In the hippocampus, [11C]-PIB uptake was significantly increased, but effect size was milder (1.72). Significant correlations between atrophy and increased [11C]-PIB uptake were found in the hippocampal (r = -0.54) and amygdalar ROIs (r = -0.40) but not in the frontal, temporal, posterior cingulate/retrosplenial, insular, and caudate ROIs (r between 0.04 and 0.25). The medial temporal lobe might be highly susceptible to amyloid toxicity, whereas neocortical areas might be more resilient.

Molecular Dynamics Simulations to Investigate the Aggregation Behaviors of the Aß(17–42) Oligomers

The amyloid β-peptides (Aßs) are the main protein components of amyloid deposits in Alzheimer's disease (AD). Detailed knowledge of the structure and assembly dynamics of Aß is important for the development of properly targeted AD therapeutics. So far, the process of the monomeric Aß assembling into oligomeric fibrils and the mechanism underlying the aggregation process remain unclear. In this study, several molecular dynamics simulations were conducted to investigate the aggregation behaviors of the Aß(17–42) oligomers associated with various numbers of monomers (dimer, trimer, tetramer, and pentamer). Our results showed that the structural stability of the Aß(17–42) oligomers increases with increasing the number of monomer. We further demonstrated that the native hydrophobic contacts are positive correlated with the ß-sheet contents, indicating that hydrophobic interaction plays an important role in maintaining the structural stability of the Aß(17–42) oligomers, particularly for those associated with more monomers. Our results also showed that the stability of the C-terminal hydrophobic segment 2 (residues 30–42) is higher than that of the N-terminal hydrophobic segment 1 (residues 17–21), suggesting that hydrophobic segment 2 may act as the nucleation site for aggregation. We further identified that Met35 residue initiates the hydrophobic interactions and that the intermolecular contact pairs, Gly33-Gly33 and Gly37-Gly37, form a stable “molecular notch”, which may mediate the packing of the ß-sheet involving many other hydrophobic residues during the early stage of amyloid-like fibril formation.

Effects of docosahexaenoic acid on in vitro amyloid beta peptide 25–35 fibrillation

Amyloid β peptide 25–35 (Aβ 25–35) encompasses one of the neurotoxic domains of full length Aβ 1–40/42, the major proteinaceous component of amyloid deposits in Alzheimer's disease (AD). We investigated the effect of docosahexaenoic acid (DHA, 22:6, n-3), an essential brain polyunsaturated fatty acid, on the in vitro fibrillation of Aβ 25–35 and found that it significantly reduced the degree of fibrillation, as shown by a decrease in the intensity of both the thioflavin T and green fluorescence in confocal microscopy. Transmission electron microscopy revealed that DHA-incubated samples were virtually devoid of structured fibrils but had an amorphous-like consistency, whereas the controls contained structured fibers of various widths and lengths. The in vitro fibrillation of Aβ 25–35 appeared to be pH-dependent, with the strongest effect seen at pH 5.0. DHA inhibited fibrillation at all pHs, with the strongest effect at pH 7.4. It also significantly decreased the levels of Aβ 25–35 oligomers. Nonreductive gradient gel electrophoresis revealed that the molecular size of the oligomers of Aβ 25–35 was 10 kDa (equivalent to decamers of Aβ 25–35) and that DHA dose-dependently reduced these decamers. These results suggest that DHA decreases the in vitro fibrillation of Aβ 25–35 by inhibiting the oligomeric amyloid species and, therefore, Aβ 25–35-related neurotoxicity or behavioral impairment could be restrained by DHA.