Neprilysin regulates amyloid β peptide levels

That neprilysin (NEP) is a major Aβ peptide-degrading enzyme in vivo is shown by higher Aβ peptide levels in the brain of an NEP knockout mouse. In addition, we show that infusion of an NEP inhibitor, but not inhibitors of other peptidases, into the brains of an APP transgenic mouse elevates Aβ levels. We have investigated the use of NEP as a potential therapeutic agent to prevent the accumulation of Aβ peptides in the brain. Lentivirus expressing NEP was initially used to demonstrate the ability of the enzyme to reduce Aβ levels in a model CHO cell line and to make primary hippocampal neurons resistant to Aβ-mediated neurotoxicity. Injection of NEP-expressing lentivirus, but not inactive NEP-expressing lentivirus, GFP-expressing lentivirus, or vehicle, into the hippocampus of 12–20-mo-old hAPP transgenic mice led to an approx 50% reduction in the number of amyloid plaques. These studies provide the impetus for further investigating of the use of NEP in a gene transfer therapy paradigm to prevent the accumulation of Aβ and prevent or delay the onset of Alzheimer’s disease.     

HIV and Alzheimer’s disease: complex interactions of HIV-Tat with amyloid β peptide and Tau protein

Journal of NeuroVirology - In patients infected with the human immunodeficiency virus (HIV), the HIV-Tat protein may be continually produced despite adequate antiretroviral therapy. As the...     

Probucol, a lipid-lowering drug, prevents cognitive and hippocampal synaptic impairments induced by amyloid β peptide in mice

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by synaptic loss and cognitive impairments. The presence of extracellular senile plaques (mainly composed of amyloid-β (Aβ) peptide) is an important molecular hallmark in AD and neuronal damage has been attributed, at least in part, to Aβ-mediated toxicity. Although the molecular mechanisms involved in the pathogenesis of AD are not yet completely understood, several lines of evidence indicate that oxidative stress and cholesterol dyshomeostasis play crucial roles in mediating the synaptic loss and cognitive deficits observed in AD patients. This study evaluated the effects of Probucol, a phenolic lipid-lowering agent with anti-inflammatory and antioxidant properties, on biochemical parameters related to oxidative stress and synaptic function (hippocampal glutathione and synaptophysin levels; glutathione peroxidase, glutathione reductase and acetylcholinesterase activities; lipid peroxidation), as well as on behavioral parameters related to the cognitive function (displaced and new object recognition tasks) in Aβ-exposed mice. Animals were treated with a single intracerebroventricular (i.c.v.) injection of aggregated Aβ(1-40) (400 pmol/site) and, subsequently, received Probucol (10 mg/kg, i.p.) once a day, during the following 2 weeks. At the end of treatments, Aβ(1-40)-exposed animals showed a significant impairment on learning-memory ability, which was paralleled by a significant decrease in hippocampal synaptophysin levels, as well as by an increase in hippocampal acetylcholinesterase activity. Importantly, Probucol treatment blunted the deleterious effects of Aβ(1-40) on learning-memory ability and hippocampal biochemistry. Although Aβ(1-40) treatment did not change hippocampal glutathione levels and glutathione peroxidase (GPx) and glutathione reductase (GR) activities, Aβ(1-40)-exposed animals showed increased hippocampal lipid peroxidation and this event was completely blunted by Probucol treatment. These findings reinforce and extend the notion of the hazardous effects of Aβ(1-40) toward hippocampal synaptic homeostasis and cognitive functions. In addition, the present results indicate that Probucol is able to counteract the cognitive and biochemical impairments induced by i.c.v. Aβ(1-40) administration in mice. The study is the first to report the protective effects of Probucol (a "non-statin cholesterol-lowering drug") against Aβ(1-40)-induced synaptic and behavioral impairments, rendering this compound a promising molecule for further pharmacological studies on the search for therapeutic strategies to treat or prevent AD.     

First evidence for helical transitions in supercoiled DNA by amyloid β peptide (1–42) and aluminum

Previously, we evidenced a B → Z helical change in Alzheimer’s brain genomic DNA, leading to a hypothesis that Alzheimer’s disease (AD) etiological factors such as aluminum (Al), amyloid β (Aβ) peptide, and Tau might play a role in modulating DNA topology. In the present study, we investigated the interaction of Al and Aβ with DNA. Our results show that Aβ(1–42) could induce a B → ψ (Psi) conformational change in pUC 18 supercoiled DNA (scDNA), Aβ(1–16) caused an altered B-form, whereas Al induced a complex B-C-A mixed conformation. Ethidium bromide binding and agarose gel electrophoresis studies revealed that Al uncoiled the DNA to a fully relaxed form, whereas Aβ(1–42) and Aβ(1–16) effected a partial uncoiling and also showed differential sensitivity toward chloroquine-induced topoisomer separation. Our findings show for the first time that Aβ and Al modulate both helicity and superhelicity in scDNA. A new hypothetical model explaining the potential toxicity of Aβ and Al in terms of their DNA binding properties leading to DNA conformational alteration is proposed. This paper is dedicated to the memory of the late Prof. M. A. Viswamitra, Indian Institute of Science, Bangalore, India. M. L. Hegde and S. Anitha have contributed equally to this paper.     

Peripheral complement interactions with amyloid β peptide: Erythrocyte clearance mechanisms

Introduction

Although amyloid β peptide (Aβ) is cleared from the brain to cerebrospinal fluid and the peripheral circulation, mechanisms for its removal from blood remain unresolved. Primates have uniquely evolved a highly effective peripheral clearance mechanism for pathogens, immune adherence, in which erythrocyte complement receptor 1 (CR1) plays a major role.

Regulation of β cleavage of amyloid precursor protein

Alzheimer’s disease ranks the first cause for senile dementia. The amyloid cascade is proposed to contribute to the pathogenesis of this disease. In this cascade, amyloid β peptide (Aβ) is produced through a sequential cleavage of amyloid precursor protein (APP) by β and γ secretases, while its cleavage by α secretase precludes Aβ production and generates neurotrophic sAPPα. Thus, enhancing αsecretase activity or suppressing βand γcleavage may reduce A βformation and ameliorate the pathological process of the disease. Several regulatory mechanisms of APP cleavage have been established. The present review mainly summarizes the signaling pathways pertinent to the regulation of APP β cleavage.     

Peripheral complement interactions with amyloid β peptide in Alzheimer's disease: 2. Relationship to amyloid β immunotherapy

Introduction

Our previous studies have shown that amyloid β peptide (Aβ) is subject to complement-mediated clearance from the peripheral circulation, and that this mechanism is deficient in Alzheimer's disease. The mechanism should be enhanced by Aβ antibodies that form immune complexes (ICs) with Aβ, and therefore may be relevant to current Aβ immunotherapy approaches.

Increased expression of β amyloid precursor gene in the hippocampus of streptozotocin-induced diabetic mice with memory deficit and anxiety induction

Diabetes has been associated with memory and behavioral dysfunctions such as anxiety. However, exact mechanisms of how diabetes affect such changes remain to be characterized. The purpose of present study is to search for streptozotocin-regulated genes in hippocampus of the mice using a differential display PCR technique, in the hope of type I diabetes-related hippocampal gene(s). It has been found that expression of a PCR product was increased by streptozotocin treatment and it was identified as β amyloid precursor protein. These results were further confirmed by performing RT-PCR analysis. In addition, the protein expression of β amyloid precursor protein as evidenced by Western blot analysis was increased in the hippocampus of streptozotocin-induced diabetic mice. To explore if the changes in amyloid β precursor protein could be related with functional changes in the brain regarding memory activity and anxiety, passive avoidance test and elevated plus maze test were performed, respectively. There is significant reduction of memory formation and marked induction of anxiety in the streptozotocin-induced diabetic mice. These results suggest that increase of β amyloid precursor protein may play a role in the memory loss and anxiety induction in type I diabetic mice.     

Overexpression of human cystatin C in transgenic mice does not affect levels of endogenous brain amyloid β peptide

Cystatin C, an inhibitor of cysteine proteases, colocalizes with amyloid β (Aβ) in parenchymal and vascular amyloid deposits in brains of Alzheimer’s disease (AD) patients, suggesting that cystatin C has a role in AD. Cystatin C also colocalizes with β amyloid precursor protein (βAPP) in transfected cultured cells. In vitro analysis of the association between the two proteins revealed that binding of cystatin C to full-length βAPP does not affect the level of Aβ secretion. Here we studied the effect of in vivo overexpression of cystatin C on the levels of endogenous brain Aβ. We have generated lines of transgenic mice expressing either wild-type human cystatin C or the Leu68Gln variant that forms amyloid deposits in the cerebral vessels of Icelandic patients with hereditary cerebral hemorrhage, under control sequences of the human cystatin C gene. Western blot analysis of brain homogenates was used to select lines of mice expressing various levels of the transgene. Analysis of Aβ40 and Aβ42 concentrations in the brain showed no difference between transgenic mice and their nontransgenic littermates. Thus, in vivo overexpression of human cystatin C does not affect Aβ levels in mice that do not deposit Aβ.     

Neutralization of soluble, synaptotoxic amyloid β species by antibodies is epitope specific

Several anti-amyloid β (Aβ) antibodies are under evaluation for the treatment of Alzheimer's disease (AD). Clinical studies using the N-terminal-directed anti-Aβ antibody bapineuzumab have demonstrated reduced brain PET-Pittsburg-B signals, suggesting the reduction of Aβ plaques, and reduced levels of total and phosphorylated tau protein in the CSF of treated AD patients. Preclinical studies using 3D6 (the murine form of bapineuzumab) have demonstrated resolution of Aβ plaque and vascular burdens, neuritic dystrophy, and preservation of synaptic density in the transgenic APP mouse models. In contrast, few studies have evaluated the direct interaction of this antibody with synaptotoxic soluble Aβ species. In the current report, we demonstrated that 3D6 binds to soluble, synaptotoxic assemblies of Aβ(1-42) and prevents multiple downstream functional consequences in rat hippocampal neurons including changes in glutamate AMPA receptor trafficking, AD-type tau phosphorylation, and loss of dendritic spines. In vivo, we further demonstrated that 3D6 prevents synaptic loss and acutely reverses the behavioral deficit in the contextual fear conditioning task in transgenic mouse models of AD, two endpoints thought to be linked to synaptotoxic soluble Aβ moieties. Importantly C-terminal anti-Aβ antibodies were ineffective on these endpoints. These results, taken with prior studies, suggest that N-terminal anti-Aβ antibodies effectively interact with both soluble and insoluble forms of Aβ and therefore appear particularly well suited for testing the Aβ hypothesis of AD.     

Amyloid β peptide 1–42 highly correlates with capillary cerebral amyloid angiopathy and Alzheimer disease pathology

Recent studies reported both positive [Thal et al. (2003) J Neuropathol Exp Neurol 62:1287–1301] and negative [Tian et al. (2003) Neurosci Lett 352:137–140] correlations between cerebral amyloid angiopathy (CAA) and Alzheimers disease (AD) pathology. We have recently shown high correlations between neuritic AD pathology and amyloid peptide (A) deposits in the capillary/pericapillary compartment (CapCAA) with only low correlations to general CAA (non-capillary). We have now studied the relationship between CapCAA and AD pathology with respect to the distribution of A40 and 42 in the frontal cortex of 100 human postmortem brains from both male and female, demented and non-demented patients (mean age ± SD 84.3±9.3 years). Using polyclonal antibodies to A40 and 42, capillary and plaques positivity were assessed semiquantiatively on a four-point scale. A42 deposits in capillaries correlated highly with both A42 deposits in plaques and morphological AD criteria (CERAD, Braak stages, and NIA-Reagan-Institute criteria), while only a low correlation with CAA was observed. A40 deposits in capillaries differed morphologically from A42 ones: they were limited to capillary walls, were significantly less frequent in both capillaries and plaques compared to A42 (P<0.01), and showed a low correlation with morphological AD criteria (P<0.05) and general CAA (P<0.01). By contrast, A42 deposits were seen in the glia limitans rather than in capillary walls themselves, and showed high correlation with morphological AD criteria (P<0.01). These data indicate that CapCAA is characterized by A42 deposits in pericapillary spaces or in the glia limitans. A low correlation between CAA and CapCAA, but high correlations between morphological AD criteria and CapCAA suggest different pathomechanisms for both types of CAA, and a close relation between CapCAA and AD pathology (both neuritic and plaque type). These data support the concept of a neuronal origin of A via drainage from interstitial fluid from the central nervous system along basement membranes to capillaries.List of Abbreviations AD Alzheimer disease - A beta amyloid peptide - A 40/42 CapS score of deposits of A 1–40/42 in capillaries - A 40/42 C number of A 1–40/42 positive cortical vessels - A 40/42 PS score of deposits of A 1–40/42 in plaques - A 40/42 TS total score of A 1–40/42 deposits - A 40/42 Csev severity of A 1–40/42 affection of cortical vessels - A 40/42 CS A 1–40/42 cortical score - A 40/42 L percentage of A 40/42 positive leptomeningeal vessels - A 40/42 Lsev severity of A 40/42 affection of leptomeningeal vessels - A 40/42 LS A 40/42 leptomeningeal score - ACTS A cortical total score - ALTS A leptomeningeal total score - CAA cerebral amyloid angiopathy - CAATS CAA total score - CapCAA capillary CAA - CERAD Consortium to Establish a Registry of Alzheimers Disease - NFT neurofibrillary tangle - NIA National Institute of Aging - NIA-RI National Institute of Aging and Reagan Institute - NP neuritic plaque - SP senile plaque - TS total scoreAn erratum to this article can be found at     

Colocalization of cholinesterases with β amyloid protein in aged and Alzheimer's brains

Summary The colocalization of amyloid protein with the enzymes acetyl- and butyrylcholinesterase was assessed using immunocytochemistry for amyloid protein and a sensitive histochemical technique for cholinesterases. In non-demented aged and Alzheimer's disease brains, double-stained sections for cholinesterases and thioflavin-S showed that all thioflavin-S-positive plaques were also positive for cholinesterases, indicating the presence of these enzymes in all plaques with -pleated amyloid protein. When amyloid angiopathy was present, cholinesterases were also observed in amyloid-laden vessels walls. Comparison of series of adjacent sections alternatively stained for acetylcholinesterase, amyloid protein and butyrylcholinesterase, as well as by double histo-immunocytochemical staining, showed either cholinesterase in a proportion of the preamyloid diffuse plaques. These data indicate that cholinesterases are associated with the amyloid protein from very early stages, when the -pleated structure is being formed. Novel functions attributed to acetyl- and butyrylcholinesterase, such us their proteolytic activity either by themselves or in association with heparan sulfate proteoglycans, may play a role in the aggregation or the consolidation processes taking place at the early stages of diffuse plaque formation.Supported by Fondo de Investigaciones Sanitarias de la Seguridad Social grant no. 90/0259 (Spain), NIA AG09466 and AG10161 (USA)     

Monoclonal antibody to β peptide,recognizing amyloid deposits,neuronal cells and lipofuscin pigments in systemic organs

Summary A monoclonal antibody (AmT-1) produced against synthetic amyloid peptide (1–28 residues) was revealed to be reactive with amyloid peptide blotted on nitrocellulose membrane, but not with that dissolved in sodium dodecyl sulfate and electrophoresed. AmT-1 immunostained senile plaques of typical, primitive and diffuse type, as well as amyloid deposits in cerebral vessels. It also reacted with neuronal and glial cells of normal and Alzheimer's disease (AD) brains. In addition, AmT-1 was also reactive strongly with lipofuscin pigments of adrenal reticular cells, and weakly with those of eccrine glands and liver cells. A rat neural cell line (PC12h) was reactive with AmT-1. By immunoelectron microscopy, a positive reaction was seen in ribosomes along the rough endoplasmic reticulum of nerve cells and PC12h cells. By immunoprecipitation, AmT1 reacted with a band at 36 kDa in the brain homogenates from Ad patients as well as from normal aged subjects. By immunoblotting analysis, AmT1 reacted with a band at 36 kDa in the cytosolic fraction of PC12 cells, and three bands (12–17 kDa) in the lipopigment fraction of the adrenal gland. These findings suggest that the cerebral amyloid deposits contain substance(s) having an epitope common to neuronal cells and lipofuscin pigments. The possible relationship between cerebral amyloid deposits and lipofuscin pigments in systemic organs is discussed.     

A novel study on amyloid β peptide 40, 42 and 40/42 ratio in Saudi autistics

Objectives  

We examined whether plasma concentrations of amyloid beta (Aβ) as protein derivatives play a central role in the etiology of autistic features.     

Inhibition of the NGF and IL-1β-induced expression of alzheimer’s amyloid precursor protein by antisense oligonucleotides

Alzheimer’s disease (AD) is a neurodegenerative disorder of the brain characterized by the extracellular deposition of amyloid in senile plaques and along the walls of the cerebral vasculature. The principal constituent of amyloid deposit is amyloid β peptide (Aβ) derived from its larger precursor protein, amyloid precursor protein (APP). The overexpression of APP is known to be a risk factor for Aβ deposit in AD and in Down syndrome (DS). The inhibition of APP expression has been thought to be beneficial to patients with AD and DS. In this study, we investigated the effects of antisense oligonucleotide (AO) on the overexpression of APP induced by IL-1β and NGF. Using phosphorothioate-oligonucleotides against initiation codon significantly reduced the protein levels of APP induced by NGF and IL-1β to basal level in PC12 cell culture systems. These results showed that these antisense oligonucleotides may have a potential to be a therapeutic agent for some patients with AD and DS.     

Differential deposition of amyloid β peptides in cerebral amyloid angiopathy associated with Alzheimer’s disease and vascular dementia

Cerebral amyloid angiopathy (CAA) caused by deposition of amyloid β (Aβ) peptides in the cerebrovasculature, involves degeneration of normal vascular components and increases the risk of infarction and cerebral hemorrhage. Accumulating evidence suggests that sporadic CAA is also a significant contributor to cognitive decline and dementia in the elderly. However, the mechanisms by which CAA arises are poorly understood. While neuronal sources of Aβ peptides are sufficient to cause CAA in transgenic mice overexpressing the amyloid precursor protein, there is reason to believe that in aging man, vascular disease modulates the disease process. To better understand CAA mechanisms in dementia, we assessed the frontal cortex of 62 consecutive cases of Alzheimer’s disease (AD), vascular dementia (VaD), and mixed dementia (MD) using immunohistochemistry with antibodies to Aβ, smooth muscle actin and the carboxyl-terminal peptides to detect Aβ(40) and Aβ(42). While vascular Aβ deposition was invariably associated with smooth muscle degeneration as indicated by absence of smooth muscle cell actin reactivity, VaD/MD cases exhibited markedly more vascular Aβ(42) deposits and smooth muscle actin loss compared to AD cases with similar degrees of CAA and Aβ(40) deposition. This suggests that distinct mechanisms are responsible for the differential deposition of Aβ in CAA associated with AD and that associated with ischemic/cerebrovascular disease. It is plausible that experimental studies on the effects of cerebrovascular disease on Aβ production and elimination will yield important clues on the pathogenesis of CAA.     

Intracellular biology of Alzheimer’s disease amyloid beta peptide

Strong evidence links excess production of a small peptide and the pathogenesis of Alzheimer’s disease (AD). Originally this peptide, beta-amyloid 42 (Aβ42), was assumed to be released by a pathogenic event; it is now well established that Aβ42 is released from cells during normal cellular metabolism of the Alzheimer amyloid precursor protein. Recently, in a series of surprising reports it was discovered that Aβ42 is produced intracellularly, and what might have been regarded first as a strange abnormality of a few selected cell lines has now been recognized as an important cellular pathway for Aβ production. Moreover, the differences between secretory and intracellular Aβ production might hold the clues for brain specificity and cellular mechanisms of AD pathogenesis.     

Spherulites in human brain tissue are composed of β sheets of amyloid and resemble senile plaques

Recent evidence showed that amyloid-β, Aβ(42), formed spherulites in vitro and, possibly, in vivo in Alzheimer's disease brain tissue. We now confirm the presence of spherulites in human brains and that they are composed of β sheets of amyloid. The spherulites were identical in appearance to spherulites of Aβ(42) formed in vitro which suggested that they may too be composed of Aβ. The physiological significance of this finding may be in its support of previous speculation that spherulites in human brain tissue are the 3-dimensional manifestations of what are otherwise identified as senile or neuritic plaques.