Aβ Fibrillogenesis Research Articles

Modulation of Aβ 42 fìbrillogenesis by glycosaminoglycan structure

The role of amyloid β (Aβ) peptide in the onset and progression of Alzheimer's disease is linked to the presence of soluble Aβ species. Sulfated glycosaminoglycans (GAGs) promote Aβ fibrillogenesis and reduce the toxicity of the peptide in neuronal cell cultures, but a satisfactory rationale to explain these effects at the molecular level has not been provided yet. We have used circular dichroism, Fourier transform infrared spectroscopy, fluorescence microscopy and spectroscopy, protease digestion, atomic force microscopy (AFM), and molecular dynamics simulations to characterize the association of the 42-residue fragment Aβ(42) with sulfated GAGs, hyaluronan, chitosan, and poly(vinyl sulfate) (PVS). Our results indicate that the formation of stable Aβ(42) fibrils is promoted by polymeric GAGs with negative charges placed in-frame with the 4.8-Å separating Aβ(42) monomers within protofibrillar β-sheets. Incubation of Aβ(42) with excess sulfated GAGs and hyaluronan increased amyloid fibril content and resistance to proteolysis 2- to 5-fold, whereas in the presence of the cationic polysaccharide chitosan, Aβ(42) fibrillar species were reduced by 25% and sensitivity to protease degradation increased ∼3-fold. Fibrils of intermediate stability were obtained in the presence of PVS, an anionic polymer with more tightly packed charges than GAGs. Important structural differences between Aβ(42) fibrils induced by PVS and Aβ(42) fibrils obtained in the presence of GAGs and hyaluronan were observed by AFM, whereas mainly precursor protofibrillar forms were detected after incubation with chitosan. Computed binding energies per peptide from -11.2 to -13.5 kcal/mol were calculated for GAGs and PVS, whereas a significantly lower value of -7.4 kcal/mol was obtained for chitosan. Taken together, our data suggest a simple and straightforward mechanism to explain the role of GAGs as enhancers of the formation of insoluble Aβ(42) fibrils trapping soluble toxic forms.

Amyloid-β Fibrillogenesis Seeded by Interface-Induced Peptide Misfolding and Self-Assembly

The amphipathicity of the natively unstructured amyloid-β (Aβ40) peptide may play an important role in its aggregation into β-sheet rich fibrils, which is linked to the pathogenesis of Alzheimer's disease. Using the air/subphase interface as a model interface, we characterized Aβ's surface activity and its conformation, assembly, and morphology at the interface. Aβ readily adsorbed to the air/subphase interface to form a 20 Å thick film and showed a critical micelle concentration of ∼120 nM. Aβ adsorbed at the air/subphase exhibited in-plane ordering that gave rise to Bragg peaks in grazing-incidence x-ray diffraction measurements. Analysis of the peaks showed that the air/subphase interface induced Aβ to fold into a β-sheet conformation and to self-assemble into ∼100 Å-sized ordered clusters. The formation of these clusters at the air/subphase interface was not affected by pH, salts, or the presence of sucrose or urea, which are known to stabilize or denature native proteins, suggesting that interface-driven Aβ misfolding and assembly are strongly favored. Furthermore, Aβ at the interface seeded the growth of fibrils in the bulk with a distinct morphology compared to those formed by homogeneous nucleation. Our results indicate that interface-induced Aβ misfolding may serve as a heterogeneous, nucleation-controlled aggregation mechanism for Aβ fibrillogenesis in vivo.

Insulin inhibits Aβ fibrillogenesis through a decrease of the GM1 ganglioside‐rich microdomain in neuronal membranes

Type 2 diabetes is a risk factor for late-onset Alzheimer's disease. However, the underlying mechanisms remain unknown. To investigate whether insulin is associated with the assembly of amyloid beta-protein from the cell surface, we treated nerve growth factor (NGF)-treated rat pheochromocytoma 12 (PC12) cells with insulin, which is related to the development of diabetes. Insulin treatment induced a decrease in GM1 ganglioside (GM1) levels in detergent-resistant membrane microdomains of NGF-treated PC12 cells. The insulin-induced effects on GM1 levels were regulated by a phosphatidylinositol 3-kinase inhibitor, but not by an extracellular signal-regulated kinase inhibitor. Pre-treatment with a protein synthesis inhibitor did not inhibit the decrease in GM1 levels induced by insulin. In addition, insulin failed to induce formation of fibrils from soluble amyloid beta-protein or to accelerate GM1-induced fibril formation. Furthermore, assembly of amyloid beta-protein in cultures of NGF-treated PC12 cells was significantly decreased by insulin. These results suggest that insulin inhibits amyloid beta-protein assembly by decreasing GM1 expression in detergent-resistant membrane microdomains of neuronal membranes.

The roles of GxxxG motif and gamma-secretase components in APP processing

Alzheimer’s disease (AD) is the leading cause of dementia in the elderly. Although the pathological features of AD, including excessive amyloid deposits, neurofibrillary tangles, neuroinflammatory responses, and progressive neuronal loss, have been relatively well-characterized, the pathogenic mechanisms underlying neurodegeneration and cognitive decline in AD remain unclear. Recent findings suggest that GxxxG glycine zipper motifs in amyloid-β precursor protein (APP) and Aβ peptide play critical roles in APP dimerization as well as Aβ production and fibrillogenesis. Given the fact that the glycine zipper motif is also found in α-synuclein and prion protein, already implicated in other neurodegenerative diseases, further understanding of this motif might provide novel therapeutic targets for protein misfolding disorders. Another research area of growing interest is γ-secretase. Although the process of intramembranous endoproteolysis was initially believed to be biochemically unfeasible to happen due to the hydrophobic environment of membranes, subsequent studies convincingly demonstrated that regulated intramembrane proteolysis does occur and it plays a critical role in cellular signaling and inflammatory response. Recent studies have identified more γ-secretase substrates and provided conflicting findings on the roles of γ-secretase components. Additional investigations into γ-secretase function may enhance our understanding of AD etiology and lead to the development of safer and more effective therapeutics.

Recombinant GST-I-Aβ28-induced efficient serum antibody against Aβ42

Six recombinant proteins GST-Aβ28/Aβ35/Aβ42 and GST-I-Aβ28/Aβ35/Aβ42 [I was the abbreviation for an immunostimulatory sequence that consisted of pan HLA DR binding epitope (PADRE) and Tetanus toxin epitope (TT)] were used as antigens after expressed and purified to immunize mice. The strongest antibody response against Aβ42 (titer 1:3200) was achieved by GST-I-Aβ28 or GST-Aβ42 immunization. However, IgG1 and IgG2b were the predominant serum antibody isotype responses by GST-I-Aβ28 immunization, whereas did IgG2a by GST-Aβ42 immunization. Thus, it indicated that GST-I-Aβ28 immunization in a mouse mainly evoked a stronger Th-2-type response; whereas, GST-Aβ42 immunization mainly elicited a Th-1-type response. Moreover, GST-I-Aβ28-induced serum antibodies had higher specificity to Aβ42 monomers and oligomers than to protofibrils and mature fibrils and exhibited the highest efficacy to block Aβ42 aggregation or fibrillogenesis and to disassemble Aβ42 aggregates in vitro. GST-I-Aβ28-induced serum antibodies also showed the most protective and restorative effects on target cells in vitro by inhibiting or neutralizing Aβ42-induced cytotoxicity. All of the above results indicated that Aβ28 could be speculated to substitute for Aβ42 and would become a better antigenic peptide for Alzheimer's disease immunotherapy in the presence of additional Th-cell epitopes such as the immunostimulatory sequence (I) applied in this study.

Vascular endothelial growth factor (VEGF) affects processing of amyloid precursor protein and β-amyloidogenesis in brain slice cultures derived from transgenic Tg2576 mouse brain

The up-regulation of the angiogenic vascular endothelial growth factor (VEGF) in brains of Alzheimer patients in close relationship to β-amyloid (Aβ) plaques, suggests a link of VEGF action and processing of the amyloid precursor protein (APP). To reveal whether VEGF may affect APP processing, brain slices derived from 17-month-old transgenic Tg2576 mice were exposed with 1ng/ml VEGF for 6, 24, and 72h, followed by assessing cytosolic and membrane-bound APP expression, level of both soluble and fibrillar Aβ-peptides, as well as activities of α- and β-secretases in brain slice tissue preparations.Treatment of brain slices with VEGF did not significantly affect the expression level of APP, regardless of the exposure time studied. In contrast, VEGF exposure of brain slices for 6h reduced the formation of soluble, SDS extractable Aβ(1–40) and Aβ(1–42) as compared to brain slice cultures incubated in the absence of any drug, while the fibrillar Aβ peptides did not change significantly. This effect was less pronounced 24h after VEGF exposure, but was no longer detectable when brain slices were exposed by VEGF for 72h, which indicates an adaptive response to chronic VEGF exposure. The VEGF-mediated reduction in Aβ formation was accompanied by a transient decrease in β-secretase activity peaking 6h after VEGF exposure. To reveal whether the VEGF-induced changes in soluble Aβ-level may be due to actions of VEGF on Aβ fibrillogenesis, the fibrillar status of Aβ was examined using the thioflavin-T binding assay. Incubation of Aβ preparations obtained from Tg2576 mouse brain cortex, in the presence of VEGF slightly decreased the fibrillar content with increasing incubation time up to 72h. The data demonstrate that VEGF may affect APP processing, at least in vitro, suggesting a role of VEGF in the pathogenesis of Alzheimer's disease.

Microscopic investigation of reversible nanoscale surface size dependent protein conjugation.

Aβ1–40 coated 20 nm gold colloidal nanoparticles exhibit a reversible color change as pH is externally altered between pH 4 and 10. This reversible process may contain important information on the initial reversible step reported for the fibrillogenesis of Aβ (a hallmark of Alzheimer’s disease). We examined this reversible color change by microscopic investigations. AFM images on graphite surfaces revealed the morphology of Aβ aggregates with gold colloids. TEM images clearly demonstrate the correspondence between spectroscopic features and conformational changes of the gold colloid.

Fibril formation of Aβ (10-35) studied by UV resonance Raman Spectroscopy

In Alzheimer's disease the major pathological feature observed is the progressive deposition of insoluble senile amyloid plaques within the cerebral cortex. The major component of these plaques is amyloid beta (Aβ), a 39-43 residue peptide. This peptide can be present as monomers with soluble disordered structure, which change conformation into partially folded intermediates and further assemble to form dimers, trimers, oligomers, protofibrils and larger fibrils. Earlier studies have suggested that neurotoxicity of the disease lies not in the insoluble fibrils but in the formation of soluble oligomers, which impair and alter neurotransmision. Thus, the solution conformation of Aβ is of significant interest for understanding the molecular mechanism of Aβ fibrillogenesis. In the present study we have investigated the process of fibril formation in the 25 residue Aβ(10-35) which has a similar fibril structure as Aβ40 and Aβ42. In addition, Aβ(10-35) contains the central hydrophobic cluster (residues 17-21) suspected to initiate folding.We have employed fluorescence, circular dichroism (CD) and time-dependent UVRR spectroscopic methods to explore the fibril formation process in vitro. Thioflavin-T induced fluorescence and CD studies indicate a conformational transition from an unfolded to β-sheet structure. UVRR studies conducted with 215 nm and 230 nm excitation facilitate characterization of the two phenylalanine and one tyrosine residues in the peptide, respectively. It is observed that formation of soluble Aβ oligomers is accompanied by a decrease in the intensities of the Phe vibrational mode, v8a (1606 cm-1) and the Tyr vibrational mode, v8a (1617 cm-1) over a 150 min incubation period. These results indicate that fibril formation proceeds via phenylalanine and tyrosine π-stacking interactions, which stabilize parallel β-sheets and reduce solvent exposure. Based on these results we have proposed a probable mechanism of fibril formation in Aβ(10-35).

Design and synthesis of new trehalose‐conjugated pentapeptides as inhibitors of Aβ(1–42) fibrillogenesis and toxicity

Aggregation of the amyloid Abeta peptide and its accumulation into insoluble deposits (plaques) are believed to be the main cause of neuronal dysfunction associated with Alzheimer's disease (AD); small molecules that can interfere with the Abeta amyloid fibril formation are therefore of interest for a potential therapeutic strategy. Three new trehalose-conjugated peptides of the well known beta-sheet breaker peptide iAbeta5p, were synthesized. The disaccharide was covalently attached to different sites of the LPFFD peptide chain, i.e. at the N-terminus, C-terminus or at the Asp side chain. CD spectroscopy in different solvents was used to assess changes in the peptide conformation of these compounds. The effects of these glycopeptides on the self-assembly and morphology of Abeta aggregates were investigated by ThT fluorescence assay and dynamic Scanning Force Microscopy, respectively. All the synthesized compounds were tested as inhibitors of Abeta toxicity toward pure cultures of rat cortical neurons.

The Ratio of Monomeric to Aggregated Forms of Aβ40 and Aβ42 Is an Important Determinant of Amyloid-β Aggregation, Fibrillogenesis, and Toxicity

Aggregation and fibril formation of amyloid-beta (Abeta) peptides Abeta40 and Abeta42 are central events in the pathogenesis of Alzheimer disease. Previous studies have established the ratio of Abeta40 to Abeta42 as an important factor in determining the fibrillogenesis, toxicity, and pathological distribution of Abeta. To better understand the molecular basis underlying the pathologic consequences associated with alterations in the ratio of Abeta40 to Abeta42, we probed the concentration- and ratio-dependent interactions between well defined states of the two peptides at different stages of aggregation along the amyloid formation pathway. We report that monomeric Abeta40 alters the kinetic stability, solubility, and morphological properties of Abeta42 aggregates and prevents their conversion into mature fibrils. Abeta40, at approximately equimolar ratios (Abeta40/Abeta42 approximately 0.5-1), inhibits (> 50%) fibril formation by monomeric Abeta42, whereas inhibition of protofibrillar Abeta42 fibrillogenesis is achieved at lower, substoichiometric ratios (Abeta40/Abeta42 approximately 0.1). The inhibitory effect of Abeta40 on Abeta42 fibrillogenesis is reversed by the introduction of excess Abeta42 monomer. Additionally, monomeric Abeta42 and Abeta40 are constantly recycled and compete for binding to the ends of protofibrillar and fibrillar Abeta aggregates. Whereas the fibrillogenesis of both monomeric species can be seeded by fibrils composed of either peptide, Abeta42 protofibrils selectively seed the fibrillogenesis of monomeric Abeta42 but not monomeric Abeta40. Finally, we also show that the amyloidogenic propensities of different individual and mixed Abeta species correlates with their relative neuronal toxicities. These findings, which highlight specific points in the amyloid peptide equilibrium that are highly sensitive to the ratio of Abeta40 to Abeta42, carry important implications for the pathogenesis and current therapeutic strategies of Alzheimer disease.

How Does Shear Affect Aβ Fibrillogenesis?

The fibrillogenesis of Abeta1-40 proceeds via three main stages: (i) formation of aggregates from monomers, (ii) linear association of these aggregates to form "beaded" protofibrils, and (iii) fusion and structural reorganization of protofibrils into mature fibrils. We have studied the effect of shear on the rate of each of these steps through a combination of fluorescence, atomic force microscopy, and circular dichroism experiments. We find that shear increases the rate of the first two stages (aggregation and protofibril formation) and inhibits the third. Our hypothesis is that in the first stage shear mechanically perturbs the peptide from its native state inducing aggregation via hydrophobic interactions; in the second stage, shear enhances the linear alignment of aggregates due to minimization of drag in the shear flow field; in the third stage, exposure to constant and uniform shear inhibits the formation of mature fibrils.

Influence of fluorinated and hydrogenated nanoparticles on the structure and fibrillogenesis of amyloid beta-peptide

Peptide aggregation in amyloid fibrils is implicated in the pathogenesis of several diseases such as Alzheimer's disease. There is a strong correlation between amyloid fibril formation and a decrease in conformational stability of the native state. Amyloid-β peptide (Aβ), the aggregating peptide in Alzheimer's disease, is natively unfolded. The deposits found in Alzheimer's disease are composed of Aβ fibrillar aggregates rich in β-sheet structure. The influence of fluorinated complexes on the secondary structure and fibrillogenesis of Aβ peptide was studied by circular dichroism (CD) spectroscopy and transmission electron microscopy (TEM). CD spectra show that complexes of polyampholyte and fluorinated dodecanoic acid induce α-helix structure in Aβ, but their hydrogenated analogous lead to β-sheet formation and aggregation. The fluorinated nanoparticles with highly negative zeta potential and hydrophobic fluorinated core have the fundamental characteristics to prevent Aβ fibrillogenesis.

Simultaneous monitoring of peptide aggregate distributions, structure, and kinetics using amide hydrogen exchange: Application to Aβ(1‐40) fibrillogenesis

Increasing evidence indicates that soluble aggregates of amyloid beta protein (Abeta) are neurotoxic. However, difficulty in isolating these unstable, dynamic species impedes studies of Abeta and other aggregating peptides and proteins. In this study, hydrogen-deuterium exchange (HX) detected by mass spectrometry (MS) was used to measure Abeta(1-40) aggregate distributions without purification or modification that might alter the aggregate structure or distribution. Different peaks in the mass spectra were assigned to monomer, low molecular weight oligomer, intermediate, and fibril based on HX labeling behavior and complementary assays. After 1 h labeling, the intermediates incorporated approximately ten more deuterons relative to fibrils, indicating a more solvent exposed structure of such intermediates. HX-MS also showed that the intermediate species dissociated much more slowly to monomer than did the very low molecular weight oligomers that were formed at very early times in Abeta aggregation. Atomic force microscopy (AFM) measurements revealed the intermediates were roughly spherical with relatively homogenous diameters of 30-50 nm. Quantitative analysis of the HX mass spectra showed that the amount of intermediate species was correlated with Abeta toxicity patterns reported in a previous study under the same conditions. This study also demonstrates the potential of the HX-MS approach to characterizing complex, multi-component oligomer distributions of aggregating peptides and proteins.

Modulation of amyloid‐β aggregation and toxicity by inosose stereoisomers

Amyloid-beta (Abeta) aggregation and amyloid formation are key pathological features of Alzheimer's disease, and are considered to be two of the major contributing factors to neurodegeneration and dementia. Identification of small molecule inhibitors that are orally available, have low toxicity and high central nervous system bioavailability is one approach to the potential development of a disease-modifying treatment for Alzheimer's disease. We have previously identified inositol stereoisomers as exhibiting stereospecific inhibition of Abeta aggregation and toxicity in vitro and in vivo. We report here the effects of inosose versus inositol stereoisomers on Abeta fibrillogenesis as determined using CD and fluorescence spectroscopy and negative-stain electron microscopy. The inososes differ from inositols by the oxidation of one of the hydroxyl groups to a ketone. These molecules help in the further elucidation of the structure-activity relationships of inositol-Abeta interactions and identify both allo-inositol and epi-2-inosose as in vitro inhibitors of Abeta aggregation.

Modulation of Fibrillogenesis of Amyloid β(1−40) Peptide with Cationic Gemini Surfactant

Modulation of the fibrillogenesis of amyloid peptide Abeta(1-40) with the cationic gemini surfactant hexamethylene-1,6-bis(dodecyldimethylammonium bromide) (C(12)C(6)C(12)Br(2)) has been studied. Both UV-vis and AFM results show that C(12)C(6)C(12)Br(2) monomers can promote the fibrillogenesis of Abeta(1-40) while its micelles inhibit this process. The electrostatic/hydrophobic force balance plays important roles in determining the Abeta(1-40) aggregation style and the secondary structures. When the surfactant positive charges are close to the Abeta(1-40) negative charges in number, the hydrophobic interaction is highly enhanced in the system. Both the nucleation rate and the lateral association between fibrils are greatly promoted. However, when the surfactant positive charges are in excess of the Abeta(1-40) negative charges, the electrostatic interaction is strengthened. In this case, the lateral association is inhibited and the alpha-helix to beta-sheet transition in the secondary structure is prevented. Simultaneously, another assembly pathway is induced to give the amorphous aggregates. Moreover, the size and surface roughness of the Abeta(1-40) aggregates also vary upon increasing C(12)C(6)C(12)Br(2) concentration.

Native and oxidized low-density lipoproteins modulate the vasoactive actions of soluble β-amyloid peptides in rat aorta

Cerebrovascular accumulation of Abeta (beta-amyloid) occurs in aging and AD (Alzheimer's disease). Hypercholesterolaemia, which is associated with raised plasma LDL (low-density lipoprotein), may predispose to AD. Soluble Abeta is found in the circulation and enhances vasoconstriction. Under conditions that may favour the formation of short Abeta oligomers, as opposed to more severe polymerization leading to Abeta fibrillogenesis, we investigated the influence of LDLs on the vasoactive actions of soluble Abeta. Thus the actions of Abeta40 and Abeta42 in combination with native or oxidized LDL on vasoconstriction to NA (noradrenaline) and vasodilatation to ACh (acetylcholine) were examined in rat aortic rings. LDL, particularly when oxidized, potentiated NA-induced constriction when combined with soluble Abeta40 and, especially, Abeta42. Soluble Abeta40 reduced relaxation induced by ACh, but Abeta42 was ineffective. Native and oxidized LDL also attenuated relaxation. Synergism occurred between oxidized LDL and Abeta with respect to ACh-induced relaxation, but not between native LDL and Abeta. We have shown for the first time that, under conditions that may result in Abeta oligomer formation, LDL, particularly when oxidized, modulates the vascular actions of soluble Abeta to extents greater than those reported previously for fibrillar Abeta preparations. Mechanisms whereby a treatable condition, namely hypercholesterolaemia, might contribute to the development of the cerebrovascular component of AD are indicated.

P-195: Inhibitory effect of EGCG on beta-amyloid and lipopolysaccharide induced-cognitive dysfunction and secretase activity

Alzheimer's disease (AD) is characterized by the extracellular deposition of beta-amyloid peptide (Aβ) in cerebral plaques. Aβ is derived from the β-amyloid precursor protein (APP) by the enzymes, β and γ-secretase. Recently, secretase is an attractive drug target for AD. Compounds that alter the proteolytic cleavage of APP, including those that inhibit β- or γ-secretase activity, can reduce the production of Aβ peptides and may have therapeutic potential in the treatment of AD. We investigated possible effects of (-)-epigallocatechin-3-gallate (EGCG) on memory dysfunction caused by lipopolysaccharide (LPS) and Aβ1–42 through inhibition of secretase activities and Aβ1–42 aggregation. In vitro study To determine whether EGCG affects Aβ fibrillogenesis, we measured in vitro aggregation of Aβ in the absence and presence of EGCG by the thioflavin T method. The inhibitory effect of EGCG on the Aβ aggregation was also found by the observation of Aβ aggregation under electronic microscope. In vivo study EGCG (1 and 2%; 1.5 and 3 mg/kg) was added into the drinking water and were accessed by mouse (25∼30 g) for 3 weeks ad libitum before the induction of memory impairment mice model. The inducers (Aβ1–42 of 0.23 ng/mouse and 1 μg/mouse of LPS) and vehicle (saline) were administrated intracerebroventricularly, and the behavioral tests were started 1 day after injection. Leaning and memory capacity was determined by passive avoidance and water maze tests. Activities of β- and γ-secretase were assessed by using commercially available assay kit. EGCG reversed the LPS and Aβ1–42-induced memory dysfunction in dose dependent manner (Figure 1A and B). EGCG also dose-dependently attenuated LPS and Aβ-induced β and γ-secretase activities in cortex and hippocampus of the mice brain. These inhibitory effects are accorded well with the inhibitory effect of EGCG on the Aβ aggregation in vitro (ID50: 6.18 μg/ml) (Figure 2). Our current study shows that EGCG has recovery effect against LPS and Aβ1–42-induced memory dysfunction through inhibition of secretase activity and Aβ aggregation in addition to anti-oxidant mechanism. This study therefore suggests that EGCG may be useful agent for prevention of development or progression of AD.

Ganglioside GM1-Mediated Amyloid-beta Fibrillogenesis and Membrane Disruption

There is increasing evidence that a class of cell membrane glycolipids, gangliosides, can mediate the fibrillogenesis and toxicity of Alzheimer's disease amyloid-beta peptide (Abeta). Using lipid monolayers and vesicles as model membranes, we measured the insertion of Abeta into 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC)-ganglioside GM1 monolayers to probe Abeta-GM1 interactions, imaged the effects of Abeta insertion on monolayer morphology, and measured the rate of Abeta fibril formation when incubated with 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC)-GM1 vesicles. Furthermore, the location of Abeta association in the monolayer was assessed by dual-probe fluorescence experiments. Abeta exhibited direct and favorable interactions with GM1 as Abeta insertion monotonically increased with GM1 concentration, despite increases in monolayer rigidity at low GM1 levels. At low GM1 concentrations, Abeta preferentially inserted into the disordered, liquid expanded phase. At higher GM1 concentrations, Abeta inserted more uniformly into the monolayer, resulting in no detectable preferences for either the disordered or condensed phase. Abeta insertion led to the disruption of membrane morphology, specifically to the expansion of the disordered phase at low GM1 concentrations and significant disruption of the condensed domains at higher GM1 concentrations. During incubation with POPC vesicles containing physiological levels of GM1, the association of Abeta with vesicles seeded the formation of Abeta fibrils. In conclusion, favorable interactions between Abeta and GM1 in the cell membrane may provide a mechanism for Abeta fibrillogenesis in vivo, and Abeta-induced disruption of the cell membrane may provide a pathway by which Abeta exerts toxicity.

Chloroquine-induced endocytic pathway abnormalities: Cellular model of GM1 ganglioside-induced Aβ fibrillogenesis in Alzheimer’s disease

Endocytic pathway abnormalities were previously observed in brains affected with Alzheimer’s disease (AD). To clarify the pathological relevance of these abnormalities to assembly of amyloid β-protein (Aβ), we treated PC12 cells with chloroquine, which potently perturbs membrane trafficking from endosomes to lysosomes. Chloroquine treatment induced accumulation of GM1 ganglioside (GM1) in Rab5-positive enlarged early endosomes and on the cell surface. Notably, an increase in GM1 level on the cell surface was sufficient to induce Aβ assembly. Our results suggest that endocytic pathway abnormalities in AD brain induce GM1 accumulation on the cell surface, leading to amyloid fibril formation in brain.

P1-051: Enhancing Aβ fibrillogenesis increases plaque load but decreases behavioral deficits in human amyloid precursor protein transgenic mice

P1-051: Enhancing Aβ fibrillogenesis increases plaque load but decreases behavioral deficits in human amyloid precursor protein transgenic mice