Oligomer-specific Abeta toxicity in cell models is mediated by selective uptake

Alzheimer's disease (AD) is characterized by the aggregation and subsequent deposition of misfolded beta-amyloid (Abeta) peptide. Previous studies show that aggregated Abeta is more toxic in oligomeric than in fibrillar form, and that each aggregation form activates specific molecular pathways in the cell. We hypothesize that these differences between oligomers and fibrils are related to their different accessibility to the intracellular space. To this end we used fluorescently labelled Abeta1-42 and demonstrate that Abeta1-42 oligomers readily enter both HeLa and differentiated SKNSH cells whereas fibrillar Abeta1-42 is not internalized. Oligomeric Abeta1-42 is internalized by an endocytic process and is transported to the lysosomes. Inhibition of uptake specifically inhibits oligomer but not fibril toxicity. Our study indicates that selective uptake of oligomers is a determinant of oligomer specific Abeta toxicity.     

Amino-terminally truncated Abeta peptide species are the main component of cotton wool plaques

Cotton wool plaques (CWPs) are round lesions that lack a central amyloid core. CWPs have been observed in individuals affected by early-onset familial Alzheimer disease (FAD) associated with mutations in the presenilin 1 (PSEN1) gene. Here we present the characterization of the amyloid-beta (Abeta) peptides deposited in the brain of an individual affected by FAD carrying the novel missense (V261I) mutation in the PSEN1 gene. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry was used to determine the Abeta peptide species present in the cerebral and cerebellar cortices, in leptomeningeal vessels, and in CWPs isolated by laser microdissection (LMD). Our results indicate that amino-terminally truncated Abeta peptide species ending at residues 42 and 43 are the main Abeta peptides deposited in brain parenchyma and LMD-CWPs in association with the PSEN1 V261I mutation. Full-length Abeta1-42 and Abeta1-43 peptide species were underrepresented. CWPs were not found to be associated with vessels and did not contain Abeta1-40 peptides, the main component of the vascular deposits. Although Abeta deposits were present mostly in the form of CWPs in the cerebral cortex and as diffuse deposits in the cerebellar cortex, a similar array of amino-terminally truncated Abeta peptide species was seen in both cases. The biochemical data support the concept that parenchymal and vascular amyloid deposits are associated with a different array of Abeta peptide species. The generation and parenchymal deposition of highly insoluble amino-terminally truncated Abeta peptides may play an important role in the pathogenesis of AD and must be taken into consideration in developing new diagnostic and therapeutic strategies.     

FAD mutants unable to increase neurotoxic Abeta 42 suggest that mutation effects on neurodegeneration may be independent of effects on Abeta

Strong support for a primary causative role of the Abeta peptides in the development of Alzheimer's disease (AD) neurodegeneration derives from reports that presenilin familial AD (FAD) mutants alter amyloid precursor protein processing, thus increasing production of neurotoxic Abeta 1-42 (Abeta 42). This effect of FAD mutants is also reflected in an increased ratio of peptides Abeta 42 over Abeta 1-40 (Abeta 40). In the present study, we show that several presenilin 1 FAD mutants failed to increase production of Abeta 42 or the Abeta 42/40 ratio. Our data suggest that the mechanism by which FAD mutations promote neurodegeneration and AD may be independent of their effects on Abeta production.     

Characterization of chemical exchange between soluble and aggregated states of beta-amyloid by solution-state NMR upon variation of salt conditions

Alzheimer's disease (AD) is characterized by the accumulation of insoluble fibrillar aggregates of beta-amyloid peptides (Abeta), a 39-42 residue peptide, in the brain of AD patients. It is hypothesized that the disease causing form is not the fibrillar species but an oligomeric Abeta molecule, which is often referred to as the "critical oligomer" of Abeta. We show in this paper that Abeta(1-40) undergoes chemical exchange between a monomeric, soluble state and an oligomeric, aggregated state under physiological conditions. In circular dichroism spectroscopy, we observe for this intermediate an alpha-helical structure. The oligomer is assigned a molecular weight of >100 kDa by diffusion-ordered spectroscopy-solution-state NMR spectroscopy (NMR). We can show by saturation transfer difference NMR experiments that the oligomer is related to monomeric Abeta. This experiment also allows us to identify the chemical groups that are involved in interactions between mono- and oligomeric Abeta molecules. Variation of the anionic strength in the buffer induces a shift of equilibrium between mono- and oligomeric states and possibly allows for the stabilization of these intermediate structures.     

Curcumin inhibits formation of amyloid beta oligomers and fibrils, binds plaques, and reduces amyloid in vivo

Alzheimer's disease (AD) involves amyloid beta (Abeta) accumulation, oxidative damage, and inflammation, and risk is reduced with increased antioxidant and anti-inflammatory consumption. The phenolic yellow curry pigment curcumin has potent anti-inflammatory and antioxidant activities and can suppress oxidative damage, inflammation, cognitive deficits, and amyloid accumulation. Since the molecular structure of curcumin suggested potential Abeta binding, we investigated whether its efficacy in AD models could be explained by effects on Abeta aggregation. Under aggregating conditions in vitro, curcumin inhibited aggregation (IC(50) = 0.8 microM) as well as disaggregated fibrillar Abeta40 (IC(50) = 1 microM), indicating favorable stoichiometry for inhibition. Curcumin was a better Abeta40 aggregation inhibitor than ibuprofen and naproxen, and prevented Abeta42 oligomer formation and toxicity between 0.1 and 1.0 microM. Under EM, curcumin decreased dose dependently Abeta fibril formation beginning with 0.125 microM. The effects of curcumin did not depend on Abeta sequence but on fibril-related conformation. AD and Tg2576 mice brain sections incubated with curcumin revealed preferential labeling of amyloid plaques. In vivo studies showed that curcumin injected peripherally into aged Tg mice crossed the blood-brain barrier and bound plaques. When fed to aged Tg2576 mice with advanced amyloid accumulation, curcumin labeled plaques and reduced amyloid levels and plaque burden. Hence, curcumin directly binds small beta-amyloid species to block aggregation and fibril formation in vitro and in vivo. These data suggest that low dose curcumin effectively disaggregates Abeta as well as prevents fibril and oligomer formation, supporting the rationale for curcumin use in clinical trials preventing or treating AD.     

Characterization of amyloid beta peptides from brain extracts of transgenic mice overexpressing the London mutant of human amyloid precursor protein

Alzheimer's disease (AD) is marked by the presence of neurofibrillary tangles and amyloid plaques in the brain of patients. To study plaque formation, we report on further quantitative and qualitative analysis of human and mouse amyloid beta peptides (Abeta) from brain extracts of transgenic mice overexpressing the London mutant of human amyloid precursor protein (APP). Using enzyme-linked immunosorbant assays (ELISAs) specific for either human or rodent Abeta, we found that the peptides from both species aggregated to form plaques. The ratios of deposited Abeta1-42/1-40 were in the order of 2-3 for human and 8-9 for mouse peptides, indicating preferential deposition of Abeta42. We also determined the identity and relative levels of other Abeta variants present in protein extracts from soluble and insoluble brain fractions. This was done by combined immunoprecipitation and mass spectrometry (IP/MS). The most prominent peptides truncated either at the carboxyl- or the amino-terminus were Abeta1-38 and Abeta11-42, respectively, and the latter was strongly enriched in the extracts of deposited peptides. Taken together, our data indicate that plaques of APP-London transgenic mice consist of aggregates of multiple human and mouse Abeta variants, and the human variants that we identified were previously detected in brain extracts of AD patients.     

Domains 16 and 17 of tropoelastin in elastic fibre formation

AD (Alzheimer's disease) is a neurodegenerative disorder characterized by self-assembly and amyloid formation of the 39-43 residue long Abeta (amyloid-beta)-peptide. The most abundant species, Abeta(1-40) and Abeta(1-42), are both present within senile plaques, but Abeta(1-42) peptides are considerably more prone to self-aggregation and are also essential for the development of AD. To understand the molecular and pathological mechanisms behind AD, a detailed knowledge of the amyloid structures of Abeta-peptides is vital. In the present study we have used quenched hydrogen/deuterium-exchange NMR experiments to probe the structure of Abeta(1-40) fibrils. The fibrils were prepared and analysed identically as in our previous study on Abeta(1-42) fibrils, allowing a direct comparison of the two fibrillar structures. The solvent protection pattern of Abeta(1-40) fibrils revealed two well-protected regions, consistent with a structural arrangement of two beta-strands connected with a bend. This protection pattern partly resembles the pattern found in Abeta(1-42) fibrils, but the Abeta(1-40) fibrils display a significantly increased protection for the N-terminal residues Phe4-His14, suggesting that additional secondary structure is formed in this region. In contrast, the C-terminal residues Gly37-Val40 show a reduced protection that suggests a loss of secondary structure in this region and an altered filament assembly. The differences between the present study and other similar investigations suggest that subtle variations in fibril-preparation conditions may significantly affect the fibrillar architecture.     

Abeta42 is essential for parenchymal and vascular amyloid deposition in mice

Considerable circumstantial evidence suggests that Abeta42 is the initiating molecule in Alzheimer's disease (AD) pathogenesis. However, the absolute requirement for Abeta42 for amyloid deposition has never been demonstrated in vivo. We have addressed this by developing transgenic models that express Abeta1-40 or Abeta1-42 in the absence of human amyloid beta protein precursor (APP) overexpression. Mice expressing high levels of Abeta1-40 do not develop overt amyloid pathology. In contrast, mice expressing lower levels of Abeta1-42 accumulate insoluble Abeta1-42 and develop compact amyloid plaques, congophilic amyloid angiopathy (CAA), and diffuse Abeta deposits. When mice expressing Abeta1-42 are crossed with mutant APP (Tg2576) mice, there is also a massive increase in amyloid deposition. These data establish that Abeta1-42 is essential for amyloid deposition in the parenchyma and also in vessels.     

Behavioral assessment of Alzheimer's transgenic mice following long-term Abeta vaccination: task specificity and correlations between Abeta deposition and spatial memory.

Long-term vaccinations with human beta-amyloid peptide 1-42 (Abeta1-42) have recently been shown to prevent or markedly reduce Abeta deposition in the PDAPP transgenic model of Alzheimer's disease (AD). Using a similar protocol to vaccinate 7.5-month-old APP (Tg2576) and APP+PS1 transgenic mice over an 8-month period, we previously reported modest reductions in brain Abeta deposition at 16 months. In these same mice, Abeta vaccinations had no deleterious behavioral effects and, in fact, benefited the mice by providing partial protection from age-related deficits in spatial working memory in the radial arm water maze task (RAWM) at 15.5 months. By contrast, control-vaccinated transgenic mice exhibited impaired performance throughout the entire RAWM test period at 15.5 months. The present study expands on our initial report by presenting additional behavioral results following long-term Abeta vaccination, as well as correlational analyses between cognitive performance and Abeta deposition in vaccinated animals. We report that 8 months of Abeta vaccinations did not reverse an early-onset balance beam impairment in transgenic mice. Additionally, in Y-maze testing at 16 months, all mice showed comparable spontaneous alternation irrespective of genotype or vaccination status. Strong correlations were nonetheless present between RAWM performance and extent of "compact" Abeta deposition in both the hippocampus and the frontal cortex of vaccinated APP+PS1 mice. Our results suggest that the behavioral protection of long-term Abeta vaccinations is task specific, with preservation of hippocampal-associated working memory tasks most likely to occur. In view of the early short-term memory deficits exhibited by AD patients, Abeta vaccination of presymptomatic AD patients could be an effective therapeutic to protect against such cognitive impairments.     

Differential physiologic responses of alpha7 nicotinic acetylcholine receptors to beta-amyloid1-40 and beta-amyloid1-42

The beta-amyloid peptides (Abeta), Abeta(1-40) and Abeta(1-42), have been implicated in Alzheimer's disease (AD) pathology. Although Abeta(1-42) is generally considered to be the pathological peptide in AD, both Abeta(1-40) and Abeta(1-42) have been used in a variety of experimental models without discrimination. Here we show that monomeric or oligomeric forms of the two Abeta peptides, when interact with the neuronal cation channel, alpha7 nicotinic acetylcholine receptors (alpha7nAChR), would result in distinct physiologic responses as measured by acetylcholine release and calcium influx experiments. While Abeta(1-42) effectively attenuated these alpha7nAChR-dependent physiology to an extent that was apparently irreversible, Abeta(1-40) showed a lower inhibitory activity that could be restored upon washings with physiologic buffers or treatment with alpha7nAChR antagonists. Our data suggest a clear pharmacological distinction between Abeta(1-40) and Abeta(1-42).     

Amyloid beta-peptide (1-42)-induced oxidative stress and neurotoxicity: implications for neurodegeneration in Alzheimer's disease brain. A review

Oxidative stress, manifested by protein oxidation, lipid peroxidation, DNA oxidation and 3-nitrotyrosine formation, among other indices, is observed in Alzheimer's disease (AD) brain. Amyloid beta-peptide (1-42) [Abeta(1-42)] may be central to the pathogenesis of AD. Our laboratory and others have implicated Abeta(1-42)-induced free radical oxidative stress in the neurodegeneration observed in AD brain. This paper reviews some of these studies from our laboratory. Recently, we showed both in-vitro and in-vivo that methionine residue 35 (Met-35) of Abeta(1-42) was critical to its oxidative stress and neurotoxic properties. Because the C-terminal region of Abeta(1-42) is helical, and invoking the i + 4 rule of helices, we hypothesized that the carboxyl oxygen of lle-31, known to be within a van der Waals distance of the S atom of Met-35, would interact with the latter. This interaction could alter the susceptibility for oxidation of Met-35, i.e. free radical formation. Consistent with this hypothesis, substitution of lle-31 by the helix-breaking amino acid, proline, completely abrogated the oxidative stress and neurotoxic properties of Abeta(1-42). Removal of the Met-35 residue from the lipid bilayer by substitution of the negatively charged Asp for Gly-37 abrogated oxidative stress and neurotoxic properties of Abeta(1-42). The free radical scavenger vitamin E prevented A(beta (1-42)-induced ROS formation, protein oxidation, lipid peroxidation, and neurotoxicity in hippocampal neurons, consistent with our model for Abeta-associated free radical oxidative stress induced neurodegeneration in AD. ApoE, allele 4, is a risk factor for AD. Synaptosomes from apoE knock-out mice are more vulnerable to Abeta-induced oxidative stress (protein oxidation, lipid peroxidation, and ROS generation) than are those from wild-type mice. We also studied synaptosomes from allele-specific human apoE knock-in mice. Brain membranes from human apoE4 mice have greater vulnerability to Abeta(1-42)-induced oxidative stress than brain membranes from apoE2 or E3, assessed by the same indices, consistent with the notion of a coupling of the oxidative environment in AD brain and increased risk of developing this disorder. Using immunoprecipitation of proteins from AD and control brain obtained no longer than 4h PMI, selective oxidized proteins were identified in the AD brain. Creatine kinase (CK) and beta-actin have increased carbonyl groups, an index of protein oxidation, and Glt-1, the principal glutamate transporter, has increased binding of the lipid peroxidation product, 4-hydroxy-2-nonenal (HNE). Abeta inhibits CK and causes lipid peroxidation, leading to HNE formation. Implications of these findings relate to decreased energy utilization, altered assembly of cytoskeletal proteins, and increased excitotoxicity to neurons by glutamate, all reported for AD. Other oxidatively modified proteins have been identified in AD brain by proteomics analysis, and these oxidatively-modified proteins may be related to increased excitotoxicity (glutamine synthetase), aberrant proteasomal degradation of damaged or aggregated proteins (ubiquitin C-terminal hydrolase L-1), altered energy production (alpha-enolase), and diminished growth cone elongation and directionality (dihydropyrimindase-related protein 2). Taken together, these studies outlined above suggest that Met-35 is key to the oxidative stress and neurotoxic properties of Abeta(1-42) and may help explain the apoE allele dependence on risk for AD, some of the functional and structural alterations in AD brain, and strongly support a causative role of Abeta(1-42)-induced oxidative stress and neurodegeneration in AD.     

Antiamyloidogenic and neuroprotective functions of cathepsin B: implications for Alzheimer's disease

Alzheimer's disease (AD) may result from the accumulation of amyloid-beta (Abeta) peptides in the brain. The cysteine protease cathepsin B (CatB) is associated with amyloid plaques in AD brains and has been suspected to increase Abeta production. Here, we demonstrate that CatB actually reduces levels of Abeta peptides, especially the aggregation-prone species Abeta1-42, through proteolytic cleavage. Genetic inactivation of CatB in mice with neuronal expression of familial AD-mutant human amyloid precursor protein (hAPP) increased the relative abundance of Abeta1-42, worsening plaque deposition and other AD-related pathologies. Lentivirus-mediated expression of CatB in aged hAPP mice reduced preexisting amyloid deposits, even thioflavin S-positive plaques. Under cell-free conditions, CatB effectively cleaved Abeta1-42, generating C-terminally truncated Abeta peptides that are less amyloidogenic. Thus, CatB likely fulfills antiamyloidogenic and neuroprotective functions. Insufficient CatB activity might promote AD; increasing CatB activity could counteract the neuropathology of this disease.     

Methionine residue 35 is critical for the oxidative stress and neurotoxic properties of Alzheimer's amyloid beta-peptide 1-42

Amyloid beta-peptide 1-42 [Abeta(1-42)] is central to the pathogenesis of Alzheimer's disease (AD), and the AD brain is under intense oxidative stress. Our laboratory combined these two aspects of AD into the Abeta-associated free radical oxidative stress model for neurodegeneration in AD brain. Abeta(1-42) caused protein oxidation, lipid peroxidation, reactive oxygen species formation, and cell death in neuronal and synaptosomal systems, all of which could be inhibited by free radical antioxidants. Recent studies have been directed at discerning molecular mechanisms by which Abeta(1-42)-associated free radical oxidative stress and neurotoxicity arise. The single methionine located in residue 35 of Abeta(1-42) is critical for these properties. This review presents the evidence supporting the role of methionine in Abeta(1-42)-associated free radical oxidative stress and neurotoxicity. This work is of obvious relevance to AD and provides a coupling between the centrality of Abeta(1-42) in the pathogenesis of AD and the oxidative stress under which the AD brain exists.     

Beta-amyloid treatment of two complementary P301L tau-expressing Alzheimer's disease models reveals similar deregulated cellular processes

Alzheimer's disease (AD) is characterized by Abeta peptide-containing plaques and tau-containing neurofibrillary tangles (NFTs). Both pathologies have been combined by crossing Abeta plaque-forming APP mutant mice with NFT-forming P301L tau mutant mice or by stereotaxically injecting beta-amyloid peptide 1-42 (Abeta42) into brains of P301L tau mutant mice. In cell culture, Abeta42 induces filamentous tau aggregates. To understand which processes are disrupted by Abeta42 in the presence of tau aggregates, we applied comparative proteomics to Abeta42-treated P301L tau-expressing neuroblastoma cells and the amygdala of P301L tau transgenic mice stereotaxically injected with Abeta42. Remarkably, a significant fraction of proteins altered in both systems belonged to the same functional categories, i.e. stress response and metabolism. We also identified model-specific effects of Abeta42 treatment such as differences in cell signaling proteins in the cellular model and of cytoskeletal and synapse associated proteins in the amygdala. By Western blotting (WB) and immunohistochemistry (IHC), we were able to show that 72% of the tested candidates were altered in human AD brain with a major emphasis on stress-related unfolded protein responsive candidates. These data highlight these processes as potentially important initiators in the Abeta42-mediated pathogenic cascade in AD and further support the role of unfolded proteins in the course of AD.     

Highly conserved and disease-specific patterns of carboxyterminally truncated Abeta peptides 1-37/38/39 in addition to 1-40/42 in Alzheimer's disease and in patients with chronic neuroinflammation

Human lumbar CSF patterns of Abeta peptides were analysed by urea-based beta-amyloid sodium dodecyl sulphate polyacrylamide gel electrophoresis with western immunoblot (Abeta-SDS-PAGE/immunoblot). A highly conserved pattern of carboxyterminally truncated Abeta1-37/38/39 was found in addition to Abeta1-40 and Abeta1-42. Remarkably, Abeta1-38 was present at a higher concentration than Abeta1-42, being the second prominent Abeta peptide species in CSF. Patients with Alzheimer's disease (AD, n = 12) and patients with chronic inflammatory CNS disease (CID, n = 10) were differentiated by unique CSF Abeta peptide patterns from patients with other neuropsychiatric diseases (OND, n = 37). This became evident only when we investigated the amount of Abeta peptides relative to their total Abeta peptide concentration (Abeta1-x%, fractional Abeta peptide pattern), which may reflect disease-specific gamma-secretase activities. Remarkably, patients with AD and CID shared elevated Abeta1-38% values, whereas otherwise the patterns were distinct, allowing separation of AD from CID or OND patients without overlap. The presence of one or two ApoE epsilon4 alleles resulted in an overall reduction of CSF Abeta peptides, which was pronounced for Abeta1-42. The severity of dementia was significantly correlated to the fractional Abeta peptide pattern but not to the absolute Abeta peptide concentrations.     

Modulation of beta-amyloid metabolism by non-steroidal anti-inflammatory drugs in neuronal cell cultures

Alzheimer disease (AD) is characterized by cerebral deposits of beta-amyloid (Abeta) peptides, which are surrounded by neuroinflammatory cells. Epidemiological studies have shown that prolonged use of non-steroidal anti-inflammatory drugs (NSAIDs) reduces the risk of developing AD. In addition, biological data indicate that certain NSAIDs specifically lower Abeta42 levels in cultures of peripheral cells independently of cyclooxygenase (COX) activity and reduce cerebral Abeta levels in AD transgenic mice. Whether other NSAIDs, including COX-selective compounds, modulate Abeta levels in neuronal cells remains unexploited. Here, we investigated the effects of compounds from every chemical class of NSAIDs on Abeta40 and Abeta42 secretion using both Neuro-2a cells and rat primary cortical neurons. Among non-selective NSAIDs, flurbiprofen and sulindac sulfide concentration-dependently reduced the secretion not only of Abeta42 but also of Abeta40. Surprisingly, both COX-2 (celecoxib; sc-125) or COX-1 (sc-560) selective compounds significantly increased Abeta42 secretion, and either did not alter (sc-560; sc-125) or reduced (celecoxib) Abeta40 levels. The levels of betaAPP C-terminal fragments and Notch cleavage were not altered by any of the NSAIDs, indicating that gamma-secretase activity was not overall changed by these drugs. The present findings show that only a few non-selective NSAIDs possess Abeta-lowering properties and therefore have a profile potentially relevant to their clinical use in AD.     

The many faces of amyloid beta in Alzheimer's disease

The 'amyloid cascade hypothesis' links amyloid beta peptide (Abeta) with the pathological process of Alzheimer's disease (AD) and it still awaits universal acceptance. Amyloid precursor protein (APP), through the actions of the gamma-secretase complex, eventually becomes a different Abetaspecies. The various Abeta species have proven to be difficult to investigate under physiological conditions, and the species of Abeta responsible for neurotoxicity has yet to be unequivocally identified. The two important Abeta peptides involved are Abeta(1-40) and Abeta(1-42), and each has been ascribed both toxic and beneficial attributes. The ratio between the two species can be important in AD etiology. Additionally, shorter variants of Abeta peptides such as Abeta(1-8), Abeta(9-16) and Abeta(16) have also been shown to be potential participants in AD pathology. Interestingly, a new 56-kDa Abeta peptide (Abeta*56) disrupts memory when injected into the brains of young rats. Transgenic mice models are complicated by the interplay between various human Abeta types and the mouse Abeta types in the mouse brains. However, the accumulation of Abeta(1-42) in the brains of transgenic C. elegans worms and Drosophila is indeed detrimental. A less investigated aspect of AD is epigenetics, but in time the investigation of the role of epigenetics in AD may add to our understanding of the development of AD.     

Pyroglutamate-modified amyloid beta-peptides--AbetaN3(pE)--strongly affect cultured neuron and astrocyte survival

N-terminally truncated amyloid-beta (Abeta) peptides are present in early and diffuse plaques of individuals with Alzheimer's disease (AD), are overproduced in early onset familial AD and their amount seems to be directly correlated to the severity and the progression of the disease in AD and Down's syndrome (DS). The pyroglutamate-containing isoforms at position 3 [AbetaN3(pE)-40/42] represent the prominent form among the N-truncated species, and may account for more than 50% of Abeta accumulated in plaques. In this study, we compared the toxic properties, fibrillogenic capabilities, and in vitro degradation profile of Abeta1-40, Abeta1-42, AbetaN3(pE)-40 and AbetaN3(pE)-42. Our data show that fibre morphology of Abeta peptides is greatly influenced by the C-terminus while toxicity, interaction with cell membranes and degradation are influenced by the N-terminus. AbetaN3(pE)-40 induced significantly more cell loss than the other species both in neuronal and glial cell cultures. Aggregated AbetaN3(pE) peptides were heavily distributed on plasma membrane and within the cytoplasm of treated cells. AbetaN3(pE)-40/42 peptides showed a significant resistance to degradation by cultured astrocytes, while full-length peptides resulted partially degraded. These findings suggest that formation of N-terminally modified peptides may enhance beta-amyloid aggregation and toxicity, likely worsening the onset and progression of the disease.     

Inhibition of Aggregation of Amyloid β42 by Arginine-Containing Small Compounds

Aggregations of proteins are in many cases associated with neurodegenerative diseases such as Alzheimer’s (AD). Small compounds capable of inhibiting protein aggregation are expected to be useful for not only in the treatment of disease but also in probing the structures of aggregated proteins. In previous studies using phage display, we found that arginine-rich short peptides consisting of four or seven amino acids bound to soluble 42-residue amyloid β (Aβ42) and inhibited globulomer (37/48 kDa oligomer) formation. In the present study, we searched for arginine-containing small molecules using the SciFinder searching service and tested their inhibitory activities against Aβ42 aggregation, by sodium dodecyl sulfate (SDS)-PAGE and thioflavine T binding assay. Commercially available Arg-Arg-7-amino-4-trifluoromethylcoumarin was found to exhibit remarkable inhibitory activities to the formation of the globulomer and the fibril of Aβ42. This chimera-type tri-peptide is expected to serve as the seed molecule of a potent inhibitor of the Aβ aggregation process.     

BACE1 deficiency rescues memory deficits and cholinergic dysfunction in a mouse model of Alzheimer's disease

beta-site APP cleaving enzyme 1 (BACE1) is the beta-secretase enzyme required for generating pathogenic beta-amyloid (Abeta) peptides in Alzheimer's disease (AD). BACE1 knockout mice lack Abeta and are phenotypically normal, suggesting that therapeutic inhibition of BACE1 may be free of mechanism-based side effects. However, direct evidence that BACE1 inhibition would improve cognition is lacking. Here we show that BACE1 null mice engineered to overexpress human APP (BACE1(-/-).Tg2576(+)) are rescued from Abeta-dependent hippocampal memory deficits. Moreover, impaired hippocampal cholinergic regulation of neuronal excitability found in the Tg2576 AD model is ameliorated in BACE1(-/-).Tg2576(+) bigenic mice. The behavioral and electrophysiological rescue of deficits in BACE1(-/-).Tg2576(+) mice is correlated with a dramatic reduction of cerebral Abeta40 and Abeta42 levels and occurs before amyloid deposition in Tg2576 mice. Our gene-based approach demonstrates that lower Abeta levels are beneficial for AD-associated memory impairments, validating BACE1 as a therapeutic target for AD.