Hopeahainol A attenuates memory deficits by targeting β‐amyloid in APP/PS1 transgenic mice

Increasing evidence demonstrates that amyloid beta (Aβ) elicits mitochondrial dysfunction and oxidative stress, which contributes to the pathogenesis of Alzheimer's disease (AD). Identification of the molecules targeting Aβ is thus of particular significance in the treatment of AD. Hopeahainol A (HopA), a polyphenol with a novel skeleton obtained from Hopea hainanensis, is potentially acetylcholinesterase‐inhibitory and anti‐oxidative in H₂O₂‐treated PC12 cells. In this study, we reported that HopA might bind to Aβ_1–42 directly and inhibit the Aβ_1–42 aggregation using a combination of molecular dynamics simulation, binding assay, transmission electron microscopic analysis and staining technique. We also demonstrated that HopA decreased the interaction between Aβ_1–42 and Aβ‐binding alcohol dehydrogenase, which in turn reduced mitochondrial dysfunction and oxidative stress in vivo and in vitro. In addition, HopA was able to rescue the long‐term potentiation induction by protecting synaptic function and attenuate memory deficits in APP/PS1 mice. Our data suggest that HopA might be a promising drug for therapeutic intervention in AD.     

Brain oxidative stress in a triple-transgenic mouse model of Alzheimer disease

Alzheimer disease (AD) is a neurodegenerative disease which is characterized by the presence of extracellular senile plaques mainly composed of amyloid-beta peptide (Abeta), intracellular neurofibrillary tangles, and selective synaptic and neuronal loss. AD brains revealed elevated levels of oxidative stress markers which have been implicated in Abeta-induced toxicity. In the present work we addressed the hypothesis that oxidative stress occurs early in the development of AD and evaluated the extension of the oxidative stress and the levels of antioxidants in an in vivo model of AD, the triple-transgenic mouse, which develops plaques, tangles, and cognitive impairments and thus mimics AD progression in humans. We have shown that in this model, levels of antioxidants, namely, reduced glutathione and vitamin E, are decreased and the extent of lipid peroxidation is increased. We have also observed increased activity of the antioxidant enzymes glutathione peroxidase and superoxide dismutase. These alterations are evident during the Abeta oligomerization period, before the appearance of Abeta plaques and neurofibrillary tangles, supporting the view that oxidative stress occurs early in the development of the disease.     

Triterpenoid CDDO-methylamide improves memory and decreases amyloid plaques in a transgenic mouse model of Alzheimer's disease

Oxidative stress is one of the earliest events in the pathogenesis of Alzheimer's disease (AD) and can markedly exacerbate amyloid pathology. Modulation of antioxidant and anti-inflammatory pathways represents an important approach for AD therapy. Synthetic triterpenoids have been found to facilitate antioxidant response and reduce inflammation in several models. We investigated the effect of the triterpenoid, 2-Cyano-3,12-Dioxooleana-1,9-Dien-28-Oic acid-MethylAmide (CDDO-MA) in Tg19959 mice, which carry the human amyloid precursor protein with two mutations. These mice develop memory impairments and amyloid plaques as early as 2–3 months of age. CDDO-MA was provided with chow (800 mg/kg) from 1 to 4 months of age. CDDO-MA significantly improved spatial memory retention and reduced plaque burden, Aβ42 levels, microgliosis, and oxidative stress in Tg19959 mice.     

Calmodulin‐like skin protein protects against spatial learning impairment in a mouse model of Alzheimer disease

Humanin and calmodulin‐like skin protein (CLSP) inhibits Alzheimer disease (AD)‐related neuronal cell death via the heterotrimeric humanin receptor in vitro . It has been suggested that CLSP is a central agonist of the heterotrimeric humanin receptor in vivo . To investigate the role of CLSP in the AD pathogenesis in vivo , we generated mouse CLSP‐1 transgenic mice, crossed them with the APPswe/PSEN1dE9 mice, a model mouse of AD, and examined the effect of CLSP over‐expression on the pathological phenotype of the AD mouse model. We found that over‐expression of the mouse CLSP‐1 gene attenuated spatial learning impairment, the loss of a presynaptic marker synaptophysin, and the inactivation of STAT3 in the APPswe/PSEN1dE9 mice. On the other hand, CLSP over‐expression did not affect levels of Aβ, soluble Aβ oligomers, or gliosis. These results suggest that the CLSP‐mediated attenuation of memory impairment and synaptic loss occurs in an Aβ‐independent manner. The results of this study may serve as a hint to the better understanding of the AD pathogenesis and the development of AD therapy.

     

Surface enhanced Raman spectroscopy distinguishes amyloid Β‐protein isoforms and conformational states

Amyloid β‐protein (Aβ) self‐association is one process linked to the development of Alzheimer's disease (AD). Aβ peptides, including its most abundant forms, Aβ40 and Aβ42, are associated with the two predominant neuropathologic findings in AD, vascular and parenchymal amyloidosis, respectively. Efforts to develop therapies for AD often have focused on understanding and controlling the assembly of these two peptides. An obligate step in these efforts is the monitoring of assembly state. We show here that surface‐enhanced Raman spectroscopy (SERS) coupled with principal component analysis (PCA) readily distinguishes Aβ40 and Aβ42. We show further, through comparison of assembly dependent changes in secondary structure and morphology, that the SERS/PCA approach unambiguously differentiates closely related assembly stages not readily differentiable by circular dichroism spectroscopy, electron microscopy, or other techniques. The high discriminating power of SERS/PCA is based on the rich structural information present in its spectra, which comprises not only on interatomic resonances between covalently associated atoms and hydrogen bond interactions important in controlling secondary structure, but effects of protein orientation relative to the substrate surface. Coupled with the label‐free, single molecule sensitivity of SERS, the approach should prove useful for determining structure activity relationships, suggesting target sites for drug development, and for testing the effects of such drugs on the assembly process. The approach also could be of value in other systems in which assembly dependent changes in protein structure correlate with the formation of toxic peptide assemblies.     

Aquaporin‐4 deficiency reduces TGF‐β1 in mouse midbrains and exacerbates pathology in experimental Parkinson's disease

Aquaporin‐4 (AQP4), the main water‐selective membrane transport protein in the brain, is localized to the astrocyte plasma membrane. Following the establishment of a 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP)‐induced Parkinson's disease (PD) model, AQP4‐deficient (AQP4^?/?) mice displayed significantly stronger microglial inflammatory responses and remarkably greater losses of tyrosine hydroxylase (TH⁺)‐positive neurons than did wild‐type AQP4 (AQP4^+/+) controls. Microglia are the most important immune cells that mediate immune inflammation in PD. However, recently, few studies have reported why AQP4 deficiency results in more severe hypermicrogliosis and neuronal damage after MPTP treatment. In this study, transforming growth factor‐β1 (TGF‐β1), a key suppressive cytokine in PD onset and development, failed to increase in the midbrain and peripheral blood of AQP4^?/? mice after MPTP treatment. Furthermore, the lower level of TGF‐β1 in AQP4^?/? mice partially resulted from impairment of its generation by astrocytes; reduced TGF‐β1 may partially contribute to the uncontrolled microglial inflammatory responses and subsequent severe loss of TH⁺ neurons in AQP4^?/? mice after MPTP treatment. Our study provides not only a better understanding of both aetiological and pathogenical factors implicated in the neurodegenerative mechanism of PD but also a possible approach to developing new treatments for PD via intervention in AQP4‐mediated immune regulation.     

RTN4B‐mediated suppression of Sirtuin 2 activity ameliorates β‐amyloid pathology and cognitive impairment in Alzheimer's disease mouse model

Sirtuin 2 (SIRT2) is an NAD+ dependent deacetylase that is the most abundant sirtuin protein in the brain. Accumulating evidence revealed the role of SIRT2 in a wide range of biological processes and age‐related diseases. However, the pivotal mechanism of SIRT2 played in Alzheimer's disease (AD) remains unknown. Here, we report that pharmacological inactivation of SIRT2 has a beneficial effect in AD. The deacetylase inhibitor of SIRT2 rescued the cognitive impairment in amyloid precursor protein/presenilin 1 transgenic mouse (APP/PS1 mouse), and the BACE1 cleavage was weakened to reduce the β‐amyloid (Aβ) production in the hippocampus. Moreover, we firstly identified that Reticulon 4B (RTN4B) played a crucial role between SIRT2/BACE1 regulation in AD. RTN4B, as a deacetylation substrate for SIRT2, the deacetylation by SIRT2 drived the ubiquitination and degradation of RTN4B and then the disturbed RTN4B interacted with and influenced the expression of BACE1. When we overexpressed RTN4B in neurons of the hippocampus in the AD mouse model, the abnormal Aβ accumulation and cognitive impairment were ameliorated, consistent with the results of SIRT2 inhibition in vivo. Moreover, we showed that the regulatory effect of SIRT2 on BACE1 is dependent on RTN4B. When RTN4B was knocked down, the effects of SIRT2 inhibition on the BACE1 level, Aβ pathology, and AD‐liked behaviors were also blocked. Collectively, we provide evidence that SIRT2 may be a potential target for AD; the new found SIRT2/RTN4B/BACE1 pathological pathway is one of the critical mechanisms for the improvement of SIRT2 on AD.     

A synthetic peptide corresponding to a region of the human pericentriolar material 1 (PCM‐1) protein binds β‐amyloid (Aβ1‐42) oligomers

We have recently reported that a ~19‐kDa polypeptide, rPK‐4, is a protein kinase Cs inhibitor that is 89% homologous to the 1171–1323 amino acid region of the 228‐kDa human pericentriolar material‐1 (PCM‐1) protein (Chakravarthy et al. 2012). We have now discovered that rPK‐4 binds oligomeric amyloid‐β peptide (Aβ)_1‐42 with high affinity. Most importantly, a PCM‐1‐selective antibody co‐precipitated Aβ and amyloid β precursor protein (AβPP) from cerebral cortices and hippocampi from AD (Alzheimer's disease) transgenic mice that produce human AβPP and Aβ_1‐42, suggesting that PCM‐1 may interact with amyloid precursor protein/Aβ in vivo. We have identified rPK‐4′s Aβ‐binding domain using a set of overlapping synthetic peptides. We have found with ELISA, dot‐blot, and polyacrylamide gel electrophoresis techniques that a ~ 5 kDa synthetic peptide, amyloid binding peptide (ABP)‐p4‐5 binds Aβ_1‐42 at nM levels. Most importantly, ABP‐p4‐5, like rPK‐4, appears to preferentially bind Aβ_1‐42 oligomers, believed to be the toxic AD‐drivers. As expected from these observations, ABP‐p4‐5 prevented Aβ_1‐42 from killing human SH‐SY5Y neuroblastoma cells via apoptosis. These findings indicate that ABP‐p4‐5 is a possible candidate therapeutic for AD.     

Conditional inactivation of nicastrin restricts amyloid deposition in an Alzheimer's disease mouse model

Production of Aβ by γ‐secretase is a key event in Alzheimer's disease (AD). The γ‐secretase complex consists of presenilin (PS) 1 or 2, nicastrin (ncstn), Pen‐2, and Aph‐1 and cleaves type I transmembrane proteins, including the amyloid precursor protein (APP). Although ncstn is widely accepted as an essential component of the complex required for γ‐secretase activity, recent in vitro studies have suggested that ncstn is dispensable for APP processing and Aβ production. The focus of this study was to answer this controversy and evaluate the role of ncstn in Aβ generation and the development of the amyloid‐related phenotype in the mouse brain. To eliminate ncstn expression in the mouse brain, we used a ncstn conditional knockout mouse that we mated with an established AD transgenic mouse model (5XFAD) and a neuronal Cre‐expressing transgenic mouse (CamKIIα‐iCre), to generate AD mice (5XFAD/CamKIIα‐iCre/ncstn^f/f mice) where ncstn was conditionally inactivated in the brain. 5XFAD/CamKIIα‐iCre/ncstn^f/f mice at 10 week of age developed a neurodegenerative phenotype with a significant reduction in Aβ production and formation of Aβ aggregates and the absence of amyloid plaques. Inactivation of nctsn resulted in substantial accumulation of APP‐CTFs and altered PS1 expression. These results reveal a key role for ncstn in modulating Aβ production and amyloid plaque formation in vivo and suggest ncstn as a target in AD therapeutics.     

Evaluation of oxidative stress in the brain of a transgenic mouse model of Alzheimer disease by in vivo electron paramagnetic resonance imaging

Alzheimer disease (AD) is a neurodegenerative disease clinically characterized by progressive cognitive dysfunction. Deposition of amyloid-β (Aβ) peptides is the most important pathophysiological hallmark of AD. Oxidative stress induced by reactive oxygen species is prominent in AD, and several reports suggest the relationship between a change in redox status and AD pathology containing progressive Aβ deposition, the activation of glial cells, and mitochondrial dysfunction. Therefore, we performed immunohistochemical analysis using a transgenic mouse model of AD (APdE9) and evaluated the activity of superoxide dismutase in brain tissue homogenates of APdE9 mice in vitro. Together with those analyses, in vivo changes in redox status with age in both wild-type (WT) and APdE9 mouse brains were measured noninvasively by three-dimensional electron paramagnetic resonance (EPR) imaging using nitroxide (3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine-1-yloxy) as a redox-sensitive probe. Both methods found similar changes in redox status with age, and in particular a significant change in redox status in the hippocampus was observed noninvasively by EPR imaging between APdE9 mice and age-matched WT mice from 9 to 18 months of age. EPR imaging clearly visualized the accelerated change in redox status of APdE9 mouse brain compared with WT. The evaluation of the redox status in the brain of AD model rodents by EPR imaging should be useful for diagnostic study of AD.     

Blocking the apolipoprotein E/Amyloid β interaction in triple transgenic mice ameliorates Alzheimer's disease related amyloid β and tau pathology

Inheritance of the apolipoprotein E4 (apoE4) genotype has been identified as the major genetic risk factor for late‐onset Alzheimer's disease (AD). Studies have shown that the binding between apoE and amyloid‐β (Aβ) peptides occurs at residues 244–272 of apoE and residues 12–28 of Aβ. ApoE4 has been implicated in promoting Aβ deposition and impairing clearance of Aβ. We hypothesized that blocking the apoE/Aβ interaction would serve as an effective new approach to AD therapy. We have previously shown that treatment with Aβ12‐28P can reduce amyloid plaques in APP/PS1 transgenic (Tg) mice and vascular amyloid in TgSwDI mice with congophilic amyloid angiopathy. In the present study, we investigated whether the Aβ12‐28P elicits a therapeutic effect on tau‐related pathology in addition to amyloid pathology using old triple transgenic AD mice (3xTg, with PS1_M146V, APP_Swe and tau_P30IL transgenes) with established pathology from the ages of 21 to 26 months. We show that treatment with Aβ12‐28P substantially reduces tau pathology both immunohistochemically and biochemically, as well as reducing the amyloid burden and suppressing the activation of astrocytes and microglia. These affects correlate with a behavioral amelioration in the treated Tg mice.

     

Cyclooxygenase‐1 inhibition reduces amyloid pathology and improves memory deficits in a mouse model of Alzheimer's disease

Several epidemiological and preclinical studies suggest that non‐steroidal anti‐inflammatory drugs (NSAIDs), which inhibit cyclooxygenase (COX), reduce the risk of Alzheimer's disease (AD) and can lower β‐amyloid (Aβ) production and inhibit neuroinflammation. However, follow‐up clinical trials, mostly using selective cyclooxygenase (COX)‐2 inhibitors, failed to show any beneficial effect in AD patients with mild to severe cognitive deficits. Recent data indicated that COX‐1, classically viewed as the homeostatic isoform, is localized in microglia and is actively involved in brain injury induced by pro‐inflammatory stimuli including Aβ, lipopolysaccharide, and interleukins. We hypothesized that neuroinflammation is critical for disease progression and selective COX‐1 inhibition, rather than COX‐2 inhibition, can reduce neuroinflammation and AD pathology. Here, we show that treatment of 20‐month‐old triple transgenic AD (3 × Tg‐AD) mice with the COX‐1 selective inhibitor SC‐560 improved spatial learning and memory, and reduced amyloid deposits and tau hyperphosphorylation. SC‐560 also reduced glial activation and brain expression of inflammatory markers in 3 × Tg‐AD mice, and switched the activated microglia phenotype promoting their phagocytic ability. The present findings are the first to demonstrate that selective COX‐1 inhibition reduces neuroinflammation, neuropathology, and improves cognitive function in 3 × Tg‐AD mice. Thus, selective COX‐1 inhibition should be further investigated as a potential therapeutic approach for AD.     

A third-generation mouse model of Alzheimer's disease shows early and increased cored plaque pathology composed of wild-type human amyloid β peptide

We previously developed single App knock-in mouse models of Alzheimer''s disease (AD) harboring the Swedish and Beyreuther/Iberian mutations with or without the Arctic mutation (App^NL-G-F and App^NL-F mice, respectively). These models showed Aβ pathology, neuroinflammation, and cognitive impairment in an age-dependent manner. The former model exhibits extensive pathology as early as 6 months, but is unsuitable for investigating Aβ metabolism and clearance because the Arctic mutation renders Aβ resistant to proteolytic degradation and prone to aggregation. In particular, it is inapplicable to preclinical immunotherapy studies due to its discrete affinity for anti-Aβ antibodies. The latter model may take as long as 18 months for the pathology to become prominent, which leaves an unfulfilled need for an Alzheimer''s disease animal model that is both swift to show pathology and useful for antibody therapy. We thus utilized mutant Psen1 knock-in mice into which a pathogenic mutation (P117L) had been introduced to generate a new model that exhibits early deposition of wild-type human Aβ by crossbreeding the App^NL-F line with the Psen1^P117L/WT line. We show that the effects of the pathogenic mutations in the App and Psen1 genes are additive or synergistic. This new third-generation mouse model showed more cored plaque pathology and neuroinflammation than App^NL-G-F mice and will help accelerate the development of disease-modifying therapies to treat preclinical AD.     

Passage of amyloid beta protein antibody across the blood-brain barrier in a mouse model of Alzheimer's disease

Vaccinations against amyloid β protein (AβP) reduce amyloid deposition and reverse learning and memory deficits in mouse models of Alzheimer’s disease. This has raised the question of whether circulating antibodies, normally restricted by the blood–brain barrier (BBB), can enter the brain [Nat. Med. 7 (2001) 369–372]. Here, we show that antibody directed against AβP does cross the BBB at a very low rate. Entry is by way of the extracellular pathways with about 0.11% of an intravenous (i.v.) dose entering the brain by 1 h. Clearance of antibody from brain increasingly dominates over time, but antibody is still detectable in brain 72 h after i.v. injection. Uptake and clearance is not altered in mice overexpressing AβP. This ability to enter and exit the brain even in the presence of increased brain ligand supports the use of antibody in the treatment of Alzheimer’s and other diseases of the brain.     

Seed‐induced Aβ deposition is modulated by microglia under environmental enrichment in a mouse model of Alzheimer's disease

Alzheimer's disease (AD) is characterized by severe neuronal loss as well as the accumulation of amyloid‐β (Aβ), which ultimately leads to plaque formation. Although there is now a general agreement that the aggregation of Aβ can be initiated by prion‐like seeding, the impact and functional consequences of induced Aβ deposits (Aβ seeding) on neurons still remain open questions. Here, we find that Aβ seeding, representing early stages of plaque formation, leads to a dramatic decrease in proliferation and neurogenesis in two APP transgenic mouse models. We further demonstrate that neuronal cell death occurs primarily in the vicinity of induced Aβ deposits culminating in electrophysiological abnormalities. Notably, environmental enrichment and voluntary exercise not only revives adult neurogenesis and reverses memory deficits but, most importantly, prevents Aβ seeding by activated, phagocytic microglia cells. Our work expands the current knowledge regarding Aβ seeding and the consequences thereof and attributes microglia an important role in diminishing Aβ seeding by environmental enrichment.     

Dipeptidyl peptidase 4 contributes to Alzheimer’s disease–like defects in a mouse model and is increased in sporadic Alzheimer’s disease brains

The amyloid cascade hypothesis, which proposes a prominent role for full-length amyloid β peptides in Alzheimer’s disease, is currently being questioned. In addition to full-length amyloid β peptide, several N-terminally truncated fragments of amyloid β peptide could well contribute to Alzheimer’s disease setting and/or progression. Among them, pyroGlu3–amyloid β peptide appears to be one of the main components of early anatomical lesions in Alzheimer’s disease–affected brains. Little is known about the proteolytic activities that could account for the N-terminal truncations of full-length amyloid β, but they appear as the rate-limiting enzymes yielding the Glu3–amyloid β peptide sequence that undergoes subsequent cyclization by glutaminyl cyclase, thereby yielding pyroGlu3–amyloid β. Here, we investigated the contribution of dipeptidyl peptidase 4 in Glu3–amyloid β peptide formation and the functional influence of its genetic depletion or pharmacological blockade on spine maturation as well as on pyroGlu3–amyloid β peptide and amyloid β 42–positive plaques and amyloid β 42 load in the triple transgenic Alzheimer’s disease mouse model. Furthermore, we examined whether reduction of dipeptidyl peptidase 4 could rescue learning and memory deficits displayed by these mice. Our data establish that dipeptidyl peptidase 4 reduction alleviates anatomical, biochemical, and behavioral Alzheimer’s disease–related defects. Furthermore, we demonstrate that dipeptidyl peptidase 4 activity is increased early in sporadic Alzheimer’s disease brains. Thus, our data demonstrate that dipeptidyl peptidase 4 participates in pyroGlu3–amyloid β peptide formation and that targeting this peptidase could be considered as an alternative strategy to interfere with Alzheimer’s disease progression.     

Mitochondrial accumulation of APP and Aβ: significance for Alzheimer disease pathogenesis

Accumulating evidence suggest that alterations in energy metabolism are among the earliest events that occur in the Alzheimer disease (AD) affected brain. Energy consumption is drastically decreased in the AD-affected regions of cerebral cortex and hippocampus pointing towards compromised mitochondrial function of neurons within specific brain regions. This is accompanied by an elevated production of reactive oxygen species contributing to increased rates of neuronal loss in the AD-affected brain regions. In this review, we will discuss the role of mitochondrial function and dysfunction in AD. We will focus on the consequences of amyloid precursor protein and amyloid-β peptide accumulation in mitochondria and their involvement in AD pathogenesis.     

The relationship between oxidative/nitrative stress and pathological inclusions in Alzheimer's and Parkinson's diseases

Alzheimer's (AD) and Parkinson's diseases (PD) are late-onset neurodegenerative diseases that have tremendous impact on the lives of affected individuals, their families, and society as a whole. Remarkable efforts are being made to elucidate the dominant factors that result in the pathogenesis of these disorders. Extensive postmortem studies suggest that oxidative/nitrative stresses are prominent features of these diseases, and several animal models support this notion. Furthermore, it is likely that protein modifications resulting from oxidative/nitrative damage contribute to the formation of intracytoplasmic inclusions characteristic of each disease. The frequent presentation of both AD and PD in individuals and the co-occurrence of inclusions characteristic of AD and PD in several other neurodegenerative diseases suggests the involvement of a common underlying aberrant process. It can be surmised that oxidative/nitrative stress, which is cooperatively influenced by environmental factors, genetic predisposition, and senescence, may be a link between these disorders.