Expression of Novel Alzheimer’s Disease Risk Genes in Control and Alzheimer’s Disease Brains

Late onset Alzheimer’s disease (LOAD) etiology is influenced by complex interactions between genetic and environmental risk factors. Large-scale genome wide association studies (GWAS) for LOAD have identified 10 novel risk genes: ABCA7, BIN1, CD2AP, CD33, CLU, CR1, EPHA1, MS4A6A, MS4A6E, and PICALM. We sought to measure the influence of GWAS single nucleotide polymorphisms (SNPs) and gene expression levels on clinical and pathological measures of AD in brain tissue from the parietal lobe of AD cases and age-matched, cognitively normal controls. We found that ABCA7, CD33, and CR1 expression levels were associated with clinical dementia rating (CDR), with higher expression being associated with more advanced cognitive decline. BIN1 expression levels were associated with disease progression, where higher expression was associated with a delayed age at onset. CD33, CLU, and CR1 expression levels were associated with disease status, where elevated expression levels were associated with AD. Additionally, MS4A6A expression levels were associated with Braak tangle and Braak plaque scores, with elevated expression levels being associated with more advanced brain pathology. We failed to detect an association between GWAS SNPs and gene expression levels in our brain series. The minor allele of rs3764650 in ABCA7 is associated with age at onset and disease duration, and the minor allele of rs670139 in MS4A6E was associated with Braak tangle and Braak plaque score. These findings suggest that expression of some GWAS genes, namely ABCA7, BIN1, CD33, CLU, CR1 and the MS4A family, are altered in AD brains.     

The Preventive and Therapeutic Effects of Intravenous Human Adipose-Derived Stem Cells in Alzheimer’s Disease Mice

Alzheimer’s disease (AD) is characterized by the accumulation of amyloid plaques and neurofibrillary tangles accompanied by cognitive dysfunction. The aim of the present study was to elucidate preventive and therapeutic potential of stem cells for AD. Among stem cells, autologous human adipose-derived stem cells (hASCs) elicit no immune rejection responses, tumorigenesis, or ethical problems. We found that intravenously transplanted hASCs passed through the BBB and migrated into the brain. The learning, memory and pathology in an AD mouse model (Tg2576) mice greatly improved for at least 4 months after intravenous injection of hASC. The number of amyloid plaques and Aβ levels decreased significantly in the brains of hASC-injected Tg mice compared to those of Tg-sham mice. Here, we first report that intravenously or intracerebrally transplanted hASCs significantly rescues memory deficit and neuropathology, in the brains of Tg mice by up-regulating IL-10 and VEGF and be a possible use for the prevention and treatment of AD.     

Effects of Long-Term Treatment with Quercetin on Cognition and Mitochondrial Function in a Mouse Model of Alzheimer’s Disease

Amyloid-β (Aβ)-induced mitochondrial dysfunction has been recognized as a prominent, early event in Alzheimer’s disease (AD). Therefore, therapeutics targeted to improve mitochondrial function could be beneficial. Quercetin, a bioflavanoid, has been reported to have potent neuro-protective effects, but its preventive effects on Aβ-induced mitochondrial dysfunction and cognitive impairment have not been well characterised. Three-month-old APPswe/PS1dE9 transgenic mice were randomly assigned to a vehicle group, two quercetin (either 20 or 40 mg kg^?1 day^?1) groups, or an Aricept (2 mg kg^?1 day^?1) group. After 16 weeks of treatment, we observed beneficial effects of quercetin (40 mg kg^?1 day^?1), including lessening learning and memory deficits, reducing scattered senile plaques, and ameliorating mitochondrial dysfunction, as evidenced by restoration of mitochondrial membrane potential, reactive oxygen species and ATP levels in mitochondria isolated from the hippocampus compared to control. Furthermore, the AMP-activated protein kinase (AMPK) activity significantly increased in the quercetin-treated (40 mg kg^?1 day^?1) group. These findings suggest that a reduction in plaque burden and mitochondrial dysfunction through the activation of AMPK may be one of the mechanisms by which quercetin improves cognitive functioning in the APPswe/PS1dE9 transgenic mouse model of AD.     

Characterization of Novel CSF Tau and ptau Biomarkers for Alzheimer’s Disease

Cerebral spinal fluid (CSF) Aβ42, tau and p181tau are widely accepted biomarkers of Alzheimer’s disease (AD). Numerous studies show that CSF tau and p181tau levels are elevated in mild-to-moderate AD compared to age-matched controls. In addition, these increases might predict preclinical AD in cognitively normal elderly. Despite their importance as biomarkers, the molecular nature of CSF tau and ptau is not known. In the current study, reverse-phase high performance liquid chromatography was used to enrich and concentrate tau prior to western-blot analysis. Multiple N-terminal and mid-domain fragments of tau were detected in pooled CSF with apparent sizes ranging from <20 kDa to ~40 kDa. The pattern of tau fragments in AD and control samples were similar. In contrast, full-length tau and C-terminal-containing fragments were not detected. To quantify levels, five tau ELISAs and three ptau ELISAs were developed to detect different overlapping regions of the protein. The discriminatory potential of each assay was determined using 20 AD and 20 age-matched control CSF samples. Of the tau ELISAs, the two assays specific for tau containing N-terminal sequences, amino acids 9-198 (numbering based on tau 441) and 9-163, exhibited the most significant differences between AD and control samples. In contrast, CSF tau was not detected with an ELISA specific for a more C-terminal region (amino acids 159-335). Significant discrimination was also observed with ptau assays measuring amino acids 159-p181 and 159-p231. Interestingly, the discriminatory potential of p181 was reduced when measured in the context of tau species containing amino acids 9-p181. Taken together, these results demonstrate that tau in CSF occurs as a series of fragments and that discrimination of AD from control is dependent on the subset of tau species measured. These assays provide novel tools to investigate CSF tau and ptau as biomarkers for other neurodegenerative diseases.     

Multifaceted Alzheimer’s Disease: Building a Roadmap for Advancement of Novel Therapies

Neurochemical Research - Alzheimer's disease (AD) is one of the most prevailing neurodegenerative disorders of elderly humans associated with cognitive damage. Biochemical, epigenetic, and...     

Metalloproteinase’s Activity and Oxidative Stress in Mild Cognitive Impairment and Alzheimer’s Disease

Matrix metalloproteinases (MMPs) and oxidative stress have been implicated in neurological diseases such as Alzheimer’s disease (AD). Plasma MMP-2 and MMP-9 activities were assessed in Mild Cognitive Impairment (MCI) and AD subjects compared with aged-matched controls, and subsequently analysed in relation to oxidative stress markers. Both MMP-2 and MMP-9 showed no significant changes versus control subjects. Plasma glutathione peroxidase Se-dependent (GPx-Se) activity and malondialdehyde (MDA) levels were higher in AD than in controls (P < 0.05), suggesting a role for GPx-Se in controlling oxidative stress in AD. Negative correlations were observed between MMPs and MDA in AD and MCI patients (P < 0.05). In conclusion, oxidative stress events did not include activation of MMPs and this similar pattern in AD and MCI suggests that both are biochemically equivalent.     

Effects of Physical Activity Training in Patients with Alzheimer’s Dementia: Results of a Pilot RCT Study

Background

There is evidence that physical activity (PA) is of cognitive benefit to the ageing brain, but little is known on the effect in patients with Alzheimer’s disease (AD). The present pilot study assessed the effect of a home-based PA training on clinical symptoms, functional abilities, and caregiver burden after 12 and 24 weeks.

Methods

In an RCT thirty patients (aged 72.4±4.3 years) with AD (MMSE: 20.6±6.5 points) and their family caregivers were allocated to a home-based 12-week PA intervention program or the usual care group. The program changed between passive, motor-assisted or active resistive leg training and changes in direction on a movement trainer in order to combine physical and cognitive stimuli.

Results

Analysis of activities of daily living in the patients (ADCS ADL total score) revealed a significant group × time interaction effect (95% CI of the difference between both groups at T2: 5.01–10.51). The control group experienced decreases in ADL performance at week 12 and 24 whereas patients in the intervention group remained stable. Analyses of executive function and language ability revealed considerable effects for semantic word fluency with a group × time interaction (95% CI of the difference between both groups at T2: 0.18–4.02). Patients in the intervention group improved during the intervention and returned to initial performance at week 12 whereas the controls revealed continuous worsening. Analyses of reaction time, hand-eye quickness and attention revealed improvement only in the intervention group. Caregiver burden remained stable in the intervention group but worsened in the control group.

Conclusions

This study suggests that PA in a home-based setting might be an effective and intrinsically attractive way to promote PA training in AD and modulate caregiver burden. The results demonstrate transfer benefits to ADL, cognitive and physical skill in patients with AD.

Trial Registration

ClinicalTrials.gov NCT02196545     

Peripheral and Central Effects of Memantine in a Mixed Preclinical Mice Model of Obesity and Familial Alzheimer’s Disease

There is growing evidence that obesity associated with type 2 diabetes mellitus (T2DM) and aging are risk factors for the development of Alzheimer’s disease (AD). However, the molecular mechanisms through which obesity interacts with β-amyloid (Aβ) to promote cognitive decline remains poorly understood. Memantine (MEM), a N-methyl-d-aspartate receptor antagonist, is currently used for the treatment of AD. Nonetheless, few studies have reported its effects on genetic preclinical models of this neurodegenerative disease exacerbated with high-fat diet (HFD)-induced obesity. Therefore, the present research aims to elucidate the effects of MEM on familial AD HFD-induced insulin resistance and learning and memory impairment. Furthermore, it aspires to determine the possible underlying mechanisms that connect AD to T2DM. Wild type and APPswe/PS1dE9 mice were used in this study. The animals were fed with either chow or HFD until 6 months of age, and they were treated with MEM-supplemented water (30 mg/kg) during the last 12 weeks. Our study demonstrates that MEM improves the metabolic consequences produced by HFD in this model of familial AD. Behavioural assessments confirmed that the treatment also improves animals learning abilities and decreases memory loss. Moreover, MEM treatment improves brain insulin signalling upregulating AKT, as well as cyclic adenosine monophosphate response element binding (CREB) expression, and modulates the amyloidogenic pathway, which, in turn, reduced the accumulation of Aβ. Moreover, this drug increases the activation of molecules involved with insulin signalling in the liver, such as insulin receptor substrate 2 (IRS2), which is a key protein regulating hepatic resistance to insulin. These results provide new insight into the role of MEM not only in the occurrence of AD treatment, but also in its potential application on peripheral metabolic disorders where Aβ plays a key role, as is the case of T2DM.     

Problems Associated with Biological Markers of Alzheimer’s Disease

The etiopathogenesis of Alzheimer’s disease (AD) is still unclear, although clinical diagnostic criteria exist and the neuropathology of AD has been studied in great detail during the last 20 years. The present study addresses certain problems in the search for biological markers for the diagnosis, as well as in the follow-up of the course of AD and its differential diagnosis and reports some of our own observations in comparison with other studies. These include protein, genetic and neuroimaging markers. The definitions of biological markers and search strategies are also discussed. Special issue dedicated to Dr. Simo S. Oja     

Sporadic Alzheimer’s Disease Begins as Episodes of Brain Ischemia and Ischemically Dysregulated Alzheimer’s Disease Genes

The study of sporadic Alzheimer’s disease etiology, now more than ever, needs an infusion of new concepts. Despite ongoing interest in Alzheimer’s disease, the basis of this entity is not yet clear. At present, the best-established and accepted “culprit” in Alzheimer’s disease pathology by most scientists is the amyloid, as the main molecular factor responsible for neurodegeneration in this disease. Abnormal upregulation of amyloid production or a disturbed clearance mechanism may lead to pathological accumulation of amyloid in brain according to the “amyloid hypothesis.” We will critically review these observations and highlight inconsistencies between the predictions of the “amyloid hypothesis” and the published data. There is still controversy over the role of amyloid in the pathological process. A question arises whether amyloid is responsible for the neurodegeneration or if it accumulates because of the neurodegeneration. Recent evidence suggests that the pathophysiology and neuropathology of Alzheimer’s disease comprises more than amyloid accumulation, tau protein pathology and finally brain atrophy with dementia. Nowadays, a handful of researchers share a newly emerged view that the ischemic episodes of brain best describe the pathogenic cascade, which eventually leads to neuronal loss, especially in hippocampus, with amyloid accumulation, tau protein pathology and irreversible dementia of Alzheimer type. The most persuasive evidences come from investigations of ischemically damaged brains of patients and from experimental ischemic brain studies that mimic Alzheimer-type dementia. This review attempts to depict what we know and do not know about the triggering factor of the Alzheimer’s disease, focusing on the possibility that the initial pathological trigger involves ischemic episodes and ischemia-induced gene dysregulation. The resulting brain ischemia dysregulates additionally expression of amyloid precursor protein and amyloid-processing enzyme genes that, in addition, ultimately compromise brain functions, leading over time to the complex alterations that characterize advanced sporadic Alzheimer’s disease. The identification of the genes involved in Alzheimer’s disease induced by ischemia will enable to further define the events leading to sporadic Alzheimer’s disease-related abnormalities. Additionally, knowledge gained from the above investigations should facilitate the elaboration of the effective treatment and/or prevention of Alzheimer’s disease.     

Unintended Effects of Cardiovascular Drugs on the Pathogenesis of Alzheimer’s Disease

Alzheimer’s disease (AD) is rapidly becoming one of the leading causes of disability and mortality in the elderly. As life-expectancy increases, an increasing number of people will rely on modern medicines to treat age-associated disorders. Among these medications, some might benefit, while others might exacerbate, the pathogenesis of AD. We screened 1,600 FDA approved drugs for β-amyloid (Aβ)-modifying activity and identified drugs that can potentially influence amyloid precursor protein processing. In this study, we focused on cardiovascular drugs and demonstrated that some hypertensive medication can differentially modulate Aβ, both in vitro and in vivo. Our study suggests that some commonly prescribed drugs might exert unintended effects and modulate AD and provides the basis for continuing investigation of the role of individual drugs on a case-by-case basis. This line of investigation will lead to the identification of common medications that are potentially beneficial or detrimental to AD as a reference for physicians to consider when prescribing the most appropriate drugs for their patients, particularly for treating chronic disorders among the growing geriatric population.     

Neuroprotective Effects of Protocatechuic Aldehyde against Neurotoxin-Induced Cellular and Animal Models of Parkinson’s Disease

Protocatechuic aldehyde (PAL) has been reported to bind to DJ-1, a key protein involved in Parkinson’s disease (PD), and exerts potential neuroprotective effects via DJ-1 in SH-SY5Y cells. In this study, we investigated the neuroprotective pharmacological effects of PAL against neurotoxin-induced cell and animal models of PD. In cellular models of PD, PAL markedly increased cell viability rates, mitochondrial oxidation-reduction activity and mitochondrial membrane potential, and reduced intracellular ROS levels to prevent neurotoxicity in PC12 cells. In animal models of PD, PAL reduced the apomorphine injection, caused turning in 6-OHDA treated rats, and increased the motor coordination and stride decreases in MPTP treated mice. Meanwhile, in an MPTP mouse model, PAL prevented a decrease of the contents of dopamine (DA) and its metabolites in the striatum and TH-positive dopaminergic neuron loss in the substantia nigra (SN). In addition, PAL increased the protein expression of DJ-1 and reduced the level of α-synuclein in the SN of MPTP lesioned mice. PAL also increased the spine density in hippocampal CA1 neurons. The current study demonstrates that PAL can efficiently protect dopaminergic neurons against neurotoxin injury in vitro and in vivo, and that the potential mechanisms may be related to its effects in increasing DJ-1, decreasing α-synuclein and its growth-promoting effect on spine density.     

Effects of Polyphenols on Brain Ageing and Alzheimer’s Disease: Focus on Mitochondria

The global trend of the phenomenon of population ageing has dramatic consequences on public health and the incidence of neurodegenerative diseases. Physiological changes that occur during normal ageing of the brain may exacerbate and initiate pathological processes that may lead to neurodegenerative disorders, especially Alzheimer's disease (AD). Hence, the risk of AD rises exponentially with age. While there is no cure currently available, sufficient intake of certain micronutrients and secondary plant metabolites may prevent disease onset. Polyphenols are highly abundant in the human diet, and several experimental and epidemiological evidences indicate that these secondary plant products have beneficial effects on AD risks. This study reviews current knowledge on the potential of polyphenols and selected polyphenol-rich diets on memory and cognition in human subjects, focusing on recent data showing in vivo efficacy of polyphenols in preventing neurodegenerative events during brain ageing and in dementia. Concentrations of polyphenols in animal brains following oral administration have been consistently reported to be very low, thus eliciting controversial discussion on their neuroprotective effects and potential mechanisms. Whether polyphenols exert any direct antioxidant effects in the brain or rather act by evoking alterations in regulatory systems of the brain or even the body periphery is still unclear. To understand the mechanisms behind the protective abilities of polyphenol-rich foods, an overall understanding of the biotransformation of polyphenols and identification of the various metabolites arising in the human body is also urgently needed.     

A Novel Approach to Delayed-Start Analyses for Demonstrating Disease-Modifying Effects in Alzheimer’s Disease

One method for demonstrating disease modification is a delayed-start design, consisting of a placebo-controlled period followed by a delayed-start period wherein all patients receive active treatment. To address methodological issues in previous delayed-start approaches, we propose a new method that is robust across conditions of drug effect, discontinuation rates, and missing data mechanisms. We propose a modeling approach and test procedure to test the hypothesis of noninferiority, comparing the treatment difference at the end of the delayed-start period with that at the end of the placebo-controlled period. We conducted simulations to identify the optimal noninferiority testing procedure to ensure the method was robust across scenarios and assumptions, and to evaluate the appropriate modeling approach for analyzing the delayed-start period. We then applied this methodology to Phase 3 solanezumab clinical trial data for mild Alzheimer’s disease patients. Simulation results showed a testing procedure using a proportional noninferiority margin was robust for detecting disease-modifying effects; conditions of high and moderate discontinuations; and with various missing data mechanisms. Using all data from all randomized patients in a single model over both the placebo-controlled and delayed-start study periods demonstrated good statistical performance. In analysis of solanezumab data using this methodology, the noninferiority criterion was met, indicating the treatment difference at the end of the placebo-controlled studies was preserved at the end of the delayed-start period within a pre-defined margin. The proposed noninferiority method for delayed-start analysis controls Type I error rate well and addresses many challenges posed by previous approaches. Delayed-start studies employing the proposed analysis approach could be used to provide evidence of a disease-modifying effect. This method has been communicated with FDA and has been successfully applied to actual clinical trial data accrued from the Phase 3 clinical trials of solanezumab.     

Characterization of a Novel Mouse Model of Alzheimer’s Disease—Amyloid Pathology and Unique β-Amyloid Oligomer Profile

Amyloid plaques composed of β-amyloid (Aβ) protein are a pathological hallmark of Alzheimer’s disease. We here report the generation and characterization of a novel transgenic mouse model of Aβ toxicity. The rTg9191 mice harbor a transgene encoding the 695 amino-acid isoform of human amyloid precursor protein (APP) with the Swedish and London mutations (APP_NLI) linked to familial Alzheimer’s disease, under the control of a tetracycline-response element, as well as a transgene encoding the tetracycline transactivator, under the control of the promoter for calcium-calmodulin kinase IIα. In these mice, APP_NLI is expressed at a level four-fold that of endogenous mouse APP and its expression is restricted to forebrain regions. Transgene expression was suppressed by 87% after two months of doxycycline administration. Histologically, we showed that (1) Aβ plaques emerged in cerebral cortex and hippocampus as early as 8 and 10.5-12.5 months of age, respectively; (2) plaque deposition progressed in an age-dependent manner, occupying up to 19% of cortex at ~25 months of age; and (3) neuropathology—such as abnormal neuronal architecture, tau hyperphosphorylation and misfolding, and neuroinflammation—was observed in the vicinity of neuritic plaques. Biochemically, we determined total Aβ production at varied ages of mice, and we showed that mice produced primarily fibrillar Aβ assemblies recognized by conformation-selective OC antibodies, but few non-fibrillar oligomers (e.g., Aβ*56) detectable by A11 antibodies. Finally, we showed that expression of the tetracycline transactivator resulted in reduced brain weight and smaller dentate-gyrus size. Collectively, these data indicate that rTg9191 mice may serve as a model for studying the neurological effects of the fibrillar Aβ assemblies in situ.     

The Distinct Role of ADAM17 in APP Proteolysis and Microglial Activation Related to Alzheimer’s Disease

Alzheimer’s disease (AD) is a progressive neurodegenerative disease with the symptom of cognitive impairment. The deposition of amyloid β (Aβ) peptide is believed to be the primary cause to neuronal dystrophy and eventually dementia. Aβ is the proteolytic product from its precursor amyloid precursor protein (APP) by β- and γ- secretase. An optional cleavage by α-secretase happens inside the Aβ domain. ADAM17 is supposed to be the regulated α-secretase of APP. Enhanced activity of ADAM17 leads to the increasing secretion of neuroprotective soluble APP α fragment and reduction of Aβ generation, which may be benefit to the disease. ADAM17 is then considered the potential therapeutic target for AD. Microglia activation and neuroinflammation is another important event in AD pathogenesis. Interestingly, ADAM17 also participates in the cleavage of many other membrane-bound proteins, especially some inflammatory factors related to microglia activation. The facilitating role of ADAM17 in inflammation and further neuronal damage has also been illustrated. In results, the activation of ADAM17 as the solution to AD may be a tricky task. The comprehensive consideration and evaluation has to be carried out carefully before the final treatment. In the present review, the distinct role of ADAM17 in AD-related APP shedding and neuroinflammatory microglial activation will be carefully discussed.     

Inhibition of BACE1 Activity by a DNA Aptamer in an Alzheimer’s Disease Cell Model

An initial step in amyloid-β (Aβ) production includes amyloid precursor protein (APP) cleavage via β-Site amyloid precursor protein cleaving enzyme 1 (BACE1). Increased levels of brain Aβ have been implicated in the pathogenesis of Alzheimer’s disease (AD). Thus, β-secretase represents a primary target for inhibitor drug development in AD. In this study, aptamers were obtained from combinatorial oligonucleotide libraries using a technology referred to as systematic evolution of ligands by exponential enrichment (SELEX). A purified human BACE1 extracellular domain was used as a target to conduct an in vitro selection process using SELEX. Two DNA aptamers were capable of binding to BACE1 with high affinity and good specificity, with Kd values in the nanomolar range. We subsequently confirmed that one aptamer, A1, exhibited a distinct inhibitory effect on BACE1 activity in an AD cell model. We detected the effects of M17-APPsw cells that stably expressed Swedish mutant APP after aptamer A1 treatment. Aβ40 and Aβ42 concentrations secreted by M17-APPsw cells decreased intracellularly and in culture media. Furthermore, Western blot analysis indicated that sAPPβ expression significantly decreased in the A1 treated versus control groups. These findings support the preliminary feasibility of an aptamer evolved from a SELEX strategy to function as a potential BACE1 inhibitor. To our knowledge, this is the first study to acquire a DNA aptamer that exhibited binding specificity to BACE1 and inhibited its activity.     

Effects of Vitamin C on Fibroblasts from Sporadic Alzheimer’s Disease Patients

Several therapies for Alzheimer’s Disease (AD) are currently under investigation. Some studies have reported that concentration of vitamins in biological fluids are lower in AD patients compared to control subjects and clinical evidence has shown the therapeutic potential of vitamin C and E in delaying AD progression. However, the molecular mechanism(s) that are engaged upon their administration in the APP metabolism in vitro or in vivo still need clarifying. Here, we investigate the effects of vitamin C supplementation, at physiological concentration, in skin fibroblasts obtained from SAD and FAD patients. This study shows that SAD patients’ fibroblasts exhibited the exclusive appearance of C-terminal fragments, derived from APP processing, without giving rise to the β-amyloid peptide, other than corresponding decreased levels of lysosomal enzymes, such as β-hexosaminidase, α-mannosidase and cathepsins B, L, and D. Special issue article in honor of Anna Maria Giuffrida-Stella.