与阿尔兹海默症相关的G 蛋白偶联受体研究进展

G蛋白偶联受体(GPCRs)在大脑信号传递中至关重要,而在阿尔兹海默症(AD)中,G蛋白偶联受体通过调控α-、β-及γ-分泌酶分泌、淀粉样前体蛋白(APP)生成及β-淀粉样蛋白(Aβ)降解,直接影响β-淀粉样蛋白在神经系统信号级联反应;另外,阿尔兹海默症中β-淀粉样蛋白的生成可以扰乱G蛋白偶联受体功能.因此,阐明G蛋白偶联受体与阿尔兹海默症发病之间的关联有助于开发以G蛋白偶联受体为靶点的阿尔兹海默症治疗药物.     

GPCR, a rider of Alzheimer's disease

Alzheimer's disease (AD) is the most common type of dementia that affects thinking,learning,memory and behavior of older people.Based on the previous studies,three pathogenic pathways are now commonly accepted as the culprits of this disease namely,amyloid-β pathway,tauopathology and cholinergic dysfunction.This review focuses on the current findings on the regulatory roles of G protein-coupled receptors (GPCRs) in the pathological progression of AD and discusses the potential of the GPCRs as novel therapeutic targets for AD.     

A Novel Compound YS-5-23 Exhibits Neuroprotective Effect by Reducing β-Site Amyloid Precursor Protein Cleaving Enzyme 1’s Expression and H2O2-Induced Cytotoxicity in SH-SY5Y Cells

Neurochemical Research - The abnormally accumulated amyloid-β (Aβ) and oxidative stress contribute to the initiation and progression of Alzheimer’s disease (AD). β-site amyloid...     

β-Arrestins as Potential Therapeutic Targets for Alzheimer’s Disease

β-arrestins represent a small family of G protein-coupled receptors (GPCRs) regulators, which provide modulating effects by facilitating desensitization and internalization of GPCRs as well as initiating their own signalings. Recent reports have demonstrated that β-arrestins levels were correlated with amyloid-β peptide (Aβ) pathology in brains of Alzheimer’s disease (AD) patients and animal models. β-arrestins could enhance the activity of γ-secretase via interacting with anterior pharynx defective 1 subunit, which increased Aβ production and contributed to the pathogenesis of AD. In addition, Aβ-induced internalization of β2-adrenergic receptor internalization and loss of dendritic spine in neurons were proven to be mediated by β-arrestins, further establishing their pathogenic role in AD. More importantly, deletion of β-arrestins markedly attenuated AD pathology, without causing any gross abnormality. Here, we review the evidence about the roles of β-arrestins in the progression of AD. In addition, the established and postulated mechanisms by which β-arrestins mediated in AD pathogenesis are also discussed. Based on the role of β-arrestins in AD pathogenesis, genetically or pharmacologically targeting β-arrestins might provide new opportunities for AD treatment.     

Beta-amyloid peptide variants in brains and cerebrospinal fluid from amyloid precursor protein (APP) transgenic mice: comparison with human Alzheimer amyloid

In this study, we report a detailed analysis of the different variants of amyloid-β (Aβ) peptides in the brains and the cerebrospinal fluid from APP23 transgenic mice, expressing amyloid precursor protein with the Swedish familial Alzheimer disease mutation, at different ages. Using one- and two-dimensional gel electrophoresis, immunoblotting, and mass spectrometry, we identified the Aβ peptides Aβ(1-40), -(1-42), -(1-39), -(1-38), -(1-37), -(2-40), and -(3-40) as well as minor amounts of pyroglutamate-modified Aβ (Aβ(N3pE)) and endogenous murine Aβ in brains from 24-month-old mice. Chemical modifications of the N-terminal amino group of Aβ were identified that had clearly been introduced during standard experimental procedures. To address this issue, we additionally applied amyloid extraction in ultrapure water. Clear differences between APP23 mice and Alzheimer disease (AD) brain samples were observed in terms of the relative abundance of specific variants of Aβ peptides, such as Aβ(N3pE), Aβ(1-42), and N-terminally truncated Aβ(2/3-42). These differences to human AD amyloid were also noticed in a related mouse line transgenic for human wild type amyloid precursor protein. Taken together, our findings suggest different underlying molecular mechanisms driving the amyloid deposition in transgenic mice and AD patients.     

Reassessing the amyloid cascade hypothesis of Alzheimer's disease

Since its inception, the amyloid cascade hypothesis has dominated the field of Alzheimer's disease (AD) research and has provided the intellectual framework for therapeutic intervention. Although the details of the hypothesis continue to evolve, its core principle has remained essentially unaltered. It posits that the amyloid-β peptides, derived from amyloid precursor protein (APP), are the root cause of AD. Substantial genetic and biochemical data support this view, and yet a number of findings also run contrary to its tenets. The presence of familial AD mutations in APP and presenilins, demonstration of Aβ toxicity, and studies in mouse models of AD all support the hypothesis, whereas the presence of Aβ plaques in normal individuals, the uncertain nature of the pathogenic Aβ species, and repeated disappointments with Aβ-centered therapeutic trials are inconsistent with the hypothesis. The current state of knowledge does not prove nor disprove the amyloid hypothesis, but rather points to the need for its reassessment. A view that Aβ is one of the factors, as opposed to the factor, that causes AD is more consistent with the present knowledge, and is more likely to promote comprehensive and effective therapeutic strategies.     

Abnormal platelet amyloid-β precursor protein metabolism in SAMP8 mice: Evidence for peripheral marker in Alzheimer's disease

Senescence-accelerated mouse strains have proved to be an accelerated-aging model, which mimics numerous features with Alzheimer's disease (AD). Three, six, and nine-month senescence-accelerated resistant 1 and senescence-accelerated prone 8 (SAMP8) mice were used in the current study, to unravel potential mechanisms for dementia and explore new diagnostic approaches for AD. The amyloid-β (Aβ40) and Aβ42 levels were elevated in hippocampi and platelets from SAMP8, along with a reduced α-secretase expression and an enhanced β-secretase expression extent with age, compared to control mice. Furthermore, hippocampal Aβ40 and Aβ42 of SAMP8 were positively correlated with platelet of these mice with aging progression. In addition, β-γ-secretase-modulated proteolytic proceeding of amyloid precursor protein in platelet might work through the PI3K/Akt/GSK3β pathway. These results indicate that platelet could be a potential early marker in the periphery to study the age-correlative aggregation of the amyloid-β peptide in patients with AD, while still requiring the considerable study.     

Cellular cholesterol homeostasis and Alzheimer's disease: Thematic Review Series: ApoE and Lipid Homeostasis in Alzheimer's Disease

Alzheimer's disease (AD) is the most common form of dementia in older adults. Currently, there is no cure for AD. The hallmark of AD is the accumulation of extracellular amyloid plaques composed of amyloid-β (Aβ) peptides (especially Aβ1-42) and neurofibrillary tangles, composed of hyperphosphorylated tau and accompanied by chronic neuroinflammation. Aβ peptides are derived from the amyloid precursor protein (APP). The oligomeric form of Aβ peptides is probably the most neurotoxic species; its accumulation eventually forms the insoluble and aggregated amyloid plaques. ApoE is the major apolipoprotein of the lipoprotein(s) present in the CNS. ApoE has three alleles, of which the Apoe4 allele constitutes the major risk factor for late-onset AD. Here we describe the complex relationship between ApoE4, oligomeric Aβ peptides, and cholesterol homeostasis. The review consists of four parts: 1) key elements involved in cellular cholesterol metabolism and regulation; 2) key elements involved in intracellular cholesterol trafficking; 3) links between ApoE4, Aβ peptides, and disturbance of cholesterol homeostasis in the CNS; 4) potential lipid-based therapeutic targets to treat AD. At the end, we recommend several research topics that we believe would help in better understanding the connection between cholesterol and AD for further investigations.     

Platelet Proteomic Analysis Revealed Differential Pattern of Cytoskeletal- and Immune-Related Proteins at Early Stages of Alzheimer’s Disease

Platelets are considered a good model system to study a number of elements associated with neuronal pathways as they share biochemical similarities. Platelets represent the major source of amyloid-β (Aβ) in blood contributing to the Aβ accumulation in the brain parenchyma and vasculature. Peripheral blood platelet alterations including cytoskeletal abnormalities, abnormal cytoplasmic calcium fluxes or increased oxidative stress levels have been related to Alzheimer’s disease (AD) pathology. Therefore, platelets can be considered a peripheral model to study metabolic mechanisms occurring in AD. To investigate peripheral molecular alterations, we examined platelet protein expression in a cohort of 164 subjects, including mild cognitive impairment (MCI), and AD patients, and healthy aged-matched controls. A two-dimensional difference gel electrophoresis (2D-DIGE) discovery phase revealed significant differences between patients and controls in five proteins: talin, vinculin, moesin, complement C3b and Rho GDP, which are known to be involved in cytoskeletal regulation including focal adhesions, inflammation and immune functions. Western blot analysis verified that talin was found to be increased in mild and moderate AD groups versus control, while the other three were found to be decreased. We also analysed amyloid precursor protein (APP), amyloid-β 1-40 (Aβ₄₀) and 1-42 (Aβ₄₂) levels in platelets from the same groups of subjects. Upregulation of platelet APP and Aβ peptides was found in AD patients compared to controls. These findings complement and expand previous reports concerning the morphological and functional alterations in AD platelets, and provide more insights into possible mechanisms that participate in the multifactorial and systemic damage in AD.     

Characterization of 1-(2-[18F] fluoro-3-pyridyl)-4-(2-isopropyl-1-oxo- isoindoline-5-yl)-5-methyl-1H-1,2,3-triazole, a PET ligand for imaging the metabotropic glutamate receptor type 1 in rat and monkey brains

Neurovascular degeneration contributes to the pathogenesis of Alzheimer's disease (AD). Because erythropoietin (EPO) promotes endothelial regeneration, we investigated the therapeutic effects of EPO in animal models of AD. In aged Tg2576 mice, EPO receptors (EPORs) were expressed in the cortex and hippocampus. Tg2576 mice were treated with daily injection of EPO (5000 IU/kg/day) for 5 days. At 14 days, EPO improved contextual memory as measured by fear-conditioning test. EPO enhanced endothelial proliferation and the level of synaptophysin expression in the brain. EPO also increased capillary density, and decreased the level of the receptor for advanced glycation endproducts (RAGE) in the brain, while decreasing in the amount of amyloid plaque and amyloid-β (Aβ). In cultured human endothelial cells, EPO enhanced angiogenesis and suppressed the expression of the RAGE. These results show that EPO improves memory and ameliorates endothelial degeneration induced by Aβ in AD models. This pre-clinical evidence suggests that EPO may be useful for the treatment of AD.     

Curcumin promotes A-beta fibrillation and reduces neurotoxicity in transgenic Drosophila

The pathology of Alzheimer's disease (AD) is characterized by the presence of extracellular deposits of misfolded and aggregated amyloid-β (Aβ) peptide and intraneuronal accumulation of tangles comprised of hyperphosphorylated Tau protein. For several years, the natural compound curcumin has been proposed to be a candidate for enhanced clearance of toxic Aβ amyloid. In this study we have studied the potency of feeding curcumin as a drug candidate to alleviate Aβ toxicity in transgenic Drosophila. The longevity as well as the locomotor activity of five different AD model genotypes, measured relative to a control line, showed up to 75% improved lifespan and activity for curcumin fed flies. In contrast to the majority of studies of curcumin effects on amyloid we did not observe any decrease in the amount of Aβ deposition following curcumin treatment. Conformation-dependent spectra from p-FTAA, a luminescent conjugated oligothiophene bound to Aβ deposits in different Drosophila genotypes over time, indicated accelerated pre-fibrillar to fibril conversion of Aβ(1-42) in curcumin treated flies. This finding was supported by in vitro fibrillation assays of recombinant Aβ(1-42). Our study shows that curcumin promotes amyloid fibril conversion by reducing the pre-fibrillar/oligomeric species of Aβ, resulting in a reduced neurotoxicity in Drosophila.     

Development of a Small D-Enantiomeric Alzheimer's Amyloid-β Binding Peptide Ligand for Future In Vivo Imaging Applications

Alzheimer's disease (AD) is a devastating disease affecting predominantly the aging population. One of the characteristic pathological hallmarks of AD are neuritic plaques, consisting of amyloid-β peptide (Aβ). While there has been some advancement in diagnostic classification of AD patients according to their clinical severity, no fully reliable method for pre-symptomatic diagnosis of AD is available. To enable such early diagnosis, which will allow the initiation of treatments early in the disease progress, neuroimaging tools are under development, making use of Aβ-binding ligands that can visualize amyloid plaques in the living brain. Here we investigate the properties of a newly designed series of D-enantiomeric peptides which are derivatives of ACI-80, formerly called D1, which was developed to specifically bind aggregated Aβ1-42. We describe ACI-80 derivatives with increased stability and Aβ binding properties, which were characterized using surface plasmon resonance and enzyme-linked immunosorbent assays. The specific interactions of the lead compounds with amyloid plaques were validated by ex vivo immunochemistry in transgenic mouse models of AD. The novel compounds showed increased binding affinity and are promising candidates for further development into in vivo imaging compounds.     

Tau Proteins and Tauopathies in Alzheimer’s Disease

Alzheimer’s disease (AD) is characterized by progressive memory loss and cognitive function deficits. There are two major pathological hallmarks that contribute to the pathogenesis of AD which are the presence of extracellular amyloid plaques composed of amyloid-β (Aβ) and intracellular neurofibrillary tangles composed of hyperphosphorylated tau. Despite extensive research that has been done on Aβ in the last two decades, therapies targeting Aβ were not very fruitful at treating AD as the efficacy of Aβ therapies observed in animal models is not reflected in human clinical trials. Hence, tau-directed therapies have received tremendous attention as the potential treatments for AD. Tauopathies are closely correlated with dementia and immunotherapy has been effective at reducing tau pathology and improving cognitive deficits in animal models. Thus, in this review article, we discussed the pathological mechanism of tau proteins, the key factors contributing to tauopathies, and therapeutic approaches for tauopathies in AD based on the recent progress in tau-based research.     

A kinetic aggregation assay allowing selective and sensitive amyloid-β quantification in cells and tissues

The process of amyloid-β (Aβ) fibril formation is genetically and pathologically linked to Alzheimer's disease (AD). Thus, a selective and sensitive method for quantifying Aβ fibrils in complex biological samples allows a variety of hypotheses to be tested. Herein, we report the basis for a quantitative in vitro kinetic aggregation assay that detects seeding-competent Aβ aggregates in mammalian cell culture media, in Caenorhabditis elegans lysate, and in mouse brain homogenate. Sonicated, proteinase K-treated Aβ fibril-containing tissue homogenates or cell culture media were added to an initially monomeric Aβ(1-40) reporter peptide to seed an in vitro nucleated aggregation reaction. The reduction in the half-time (t(50)) of the amyloid growth phase is proportional to the quantity of seeding-competent Aβ aggregates present in the biological sample. An ion-exchange resin amyloid isolation strategy from complex biological samples is demonstrated as an alternative for improving the sensitivity and linearity of the kinetic aggregation assay.     

Mitochondrial import and degradation of amyloid-β peptide

Mitochondrial dysfunctions associated with amyloid-β peptide (Aβ) accumulation in mitochondria have been observed in Alzheimer's disease (AD) patients' brains and in AD mice models. Aβ is produced by sequential action of β- and γ-secretases cleaving the amyloid precursor protein (APP). The γ-secretase complex was found in mitochondria-associated endoplasmic reticulum membranes (MAM) suggesting that this could be a potential site of Aβ production, from which Aβ is further transported into the mitochondria. In vitro, Aβ was shown to be imported into the mitochondria through the translocase of the outer membrane (TOM) complex. The mitochondrial presequence protease (PreP) is responsible for Aβ degradation reducing toxic effects of Aβ on mitochondrial functions. The proteolytic activity of PreP is, however, lower in AD brain temporal lobe mitochondria and in AD transgenic mice models, possibly due to an increased reactive oxygen species (ROS) production. Here, we review the intracellular mechanisms of Aβ production, its mitochondrial import and the intra-mitochondrial degradation. We also discuss the implications of a reduced efficiency of mitochondrial Aβ clearance for AD. Understanding the underlying mechanisms may provide new insights into mitochondria related pathogenesis of AD and development of drug therapy against AD. This article is part of a Special Issue entitled: 18th European Bioenergetic Conference.     

β-amyloid PET neuroimaging: A review of radiopharmaceutical development

Alzheimer's disease (AD) is a neurodegenerative disease characterized by deposition of amyloid-β plaques that occurs even before symptoms of brain failure are clinically detectable. Whereas previously the diagnosis of AD was only routinely based on clinical assessment, an improvement over the past few years in imaging biomarkers has now led to reconsider the core of the AD diagnostic pathway. Therefore, positron emission tomography (PET) radiotracers for the in vivo imaging of amyloid plaques have been the focus of intense research. Many chemical compounds, mostly derived from the chemistry of amyloid histological staining dyes, have permitted to obtain promising brain amyloid radiopharmaceuticals. Three of them have been approved by the FDA and European regulatory bodies. The present review focuses on the development of these compounds not only as a suitable imaging biomarker to improve AD diagnosis, but also to evaluate new potential therapy.     

Amyloid-β forms fibrils by nucleated conformational conversion of oligomers

Amyloid-β amyloidogenesis is reported to occur via a nucleated polymerization mechanism. If this is true, the energetically unfavorable oligomeric nucleus should be very hard to detect. However, many laboratories have detected early nonfibrillar amyloid-β oligomers without observing amyloid fibrils, suggesting that a mechanistic revision may be needed. Here we introduce Cys-Cys-amyloid-β(1-40), which cannot bind to the latent fluorophore FlAsH as a monomer, but can bind FlAsH as an nonfibrillar oligomer or as a fibril, rendering the conjugates fluorescent. Through FlAsH monitoring of Cys-Cys-amyloid-β(1-40) aggregation, we found that amyloid-β(1-40) rapidly and efficiently forms spherical oligomers in vitro (85% yield) that are kinetically competent to slowly convert to amyloid fibrils by a nucleated conformational conversion mechanism. This methodology was used to show that plasmalogen ethanolamine vesicles eliminate the proteotoxicity-associated oligomerization phase of amyloid-β amyloidogenesis while allowing fibril formation, rationalizing how low concentrations of plasmalogen ethanolamine in the brain are epidemiologically linked to Alzheimer's disease.     

Mechanistic involvement of the calpain-calpastatin system in Alzheimer neuropathology

The mechanism by which amyloid-β peptide (Aβ) accumulation causes neurodegeneration in Alzheimer's disease (AD) remains unresolved. Given that Aβ perturbs calcium homeostasis in neurons, we investigated the possible involvement of calpain, a calcium-activated neutral protease. We first demonstrated close postsynaptic association of calpain activation with Aβ plaque formation in brains from both patients with AD and transgenic (Tg) mice overexpressing amyloid precursor protein (APP). Using a viral vector-based tracer, we then showed that axonal termini were dynamically misdirected to calpain activation-positive Aβ plaques. Consistently, cerebrospinal fluid from patients with AD contained a higher level of calpain-cleaved spectrin than that of controls. Genetic deficiency of calpastatin (CS), a calpain-specific inhibitor protein, augmented Aβ amyloidosis, tau phosphorylation, microgliosis, and somatodendritic dystrophy, and increased mortality in APP-Tg mice. In contrast, brain-specific CS overexpression had the opposite effect. These findings implicate that calpain activation plays a pivotal role in the Aβ-triggered pathological cascade, highlighting a target for pharmacological intervention in the treatment of AD.     

Oleocanthal-rich extra-virgin olive oil enhances donepezil effect by reducing amyloid-β load and related toxicity in a mouse model of Alzheimer’s disease

Previous evidence suggested that extra-virgin olive oil (EVOO) is linked to attenuating amyloid-β (Aβ) pathology and improving cognitive function in Alzheimer’s disease (AD) mouse models. In addition, we recently reported the beneficial effect of oleocanthal, a phenolic compound in EVOO, against AD pathology. Currently, medications available to target AD pathology are limited. Donepezil is an acetylcholine esterase inhibitor approved for use for all AD stages. Donepezil has been reported to have limited Aβ-targeting mechanisms beside its acetylcholine esterase inhibition. The aim of this study was to investigate the consumption of EVOO rich with oleocanthal (hereafter EVOO) as a medical food on enhancing the effect of donepezil on attenuating Aβ load and related toxicity in 5xFAD mouse model of AD. Our results showed that EVOO consumption in combination with donepezil significantly reduced Aβ load and related pathological changes. Reduced Aβ load could be explained, at least in part, by enhancing Aβ clearance pathways including blood–brain barrier (BBB) clearance and enzymatic degradation, and shifting amyloid precursor protein processing toward the nonamyloidogenic pathway. Furthermore, EVOO combination with donepezil up-regulated synaptic proteins, enhanced BBB tightness and reduced neuroinflammation associated with Aβ pathology. In conclusion, EVOO consumption as a medical food combined with donepezil offers an effective therapeutic approach by enhancing the noncholinergic mechanisms of donepezil and by providing additional mechanisms to attenuate Aβ-related pathology in AD patients.     

Targeted proteomics in Alzheimer’s disease: focus on amyloid-β

Diagnosis and monitoring of sporadic Alzheimer’s disease (AD) have long depended on clinical examination of individuals with end-stage disease. However, upcoming anti-AD therapies are optimally initiated when individuals show very mild signs of neurodegeneration. There is a developing consensus for cerebrospinal fluid amyloid-β (Aβ) as a core biomarker for the mild cognitive impairment stage of AD. Aβ is directly involved in the pathogenesis of AD or tightly correlated with other primary pathogenic factors. It is produced from amyloid precursor protein (APP) by proteolytic processing that depends on the β-site APP-cleaving enzyme 1 and the γ-secretase complex, and is degraded by a broad range of proteases. This review summarizes targeted proteomic studies of Aβ in biological fluids and identifies clinically useful markers of disrupted Aβ homeostasis in AD. The next 5 years will see a range of novel assays developed on the basis of these results. From a longer perspective, establishment of the most effective combinations of different biomarkers and other diagnostic modalities may be foreseen.