Differential gene expression in human brain pericytes induced by amyloid-beta protein

Cerebral amyloid angiopathy is one of the characteristics of Alzheimer's disease (AD) and this accumulation of fibrillar amyloid-beta (Alphabeta) in the vascular wall is accompanied by marked vascular damage. In vitro, Abeta1-40 carrying the 'Dutch' mutation (DAbeta1-40) induces degeneration of cultured human brain pericytes (HBP). To identify possible intracellular mediators of Abeta-induced cell death, a comparative cDNA expression array was performed to detect differential gene expression of Abeta-treated vs. untreated HBP. Messenger RNA expression of cyclin D1, integrin beta4, defender against cell death-1, neuroleukin, thymosin beta10, and integrin alpha5 were increased in DAbeta1-40-treated HBP, whereas insulin-like growth factor binding protein-2 mRNA expression was decreased. Corresponding protein expression was investigated in AD and control brains to explore a potential role for these proteins in pathological lesions of the AD brain. Cyclin D1 expression was increased in cerebral amyloid angiopathy and cells in a perivascular position, suggesting that the cell cycle may be disturbed during Abeta-mediated degeneration of cerebrovascular cells. Moreover, cyclin D1 expression, but also that of integrin beta4, defender against cell death-1, neuroleukin and thymosin beta10 was found in a subset of senile plaques, suggesting a role for these proteins in the pathogenesis of senile plaques.     

Lack of neprilysin suffices to generate murine amyloid-like deposits in the brain and behavioral deficit in vivo

Accumulation of the beta-amyloid peptide (Abeta) in the brain is a major pathological hallmark of Alzheimer's disease (AD), leading to synaptic dysfunction, neuronal death, and memory impairment. The levels of neprilysin, a major Abeta-degrading enzyme, are decreased in AD brains and during aging. Because neprilysin cleaves Abeta in vivo, its down-regulation may contribute to the pathophysiology of AD. The aim of this study was to assess the consequences of neprilysin deficiency on accumulation of murine Abeta in brains and associated pathologies in vivo by investigating neprilysin-deficient mice on biochemical, morphological, and behavioral levels. Aged neprilysin-deficient mice expressed physiological amyloid precursor protein (APP) levels and exhibited elevated brain Abeta concentrations and amyloid-like deposits in addition to signs of neuronal degeneration in their brains. Behaviorally, neprilysin-deficient mice acquired a significantly weaker conditioned taste aversion that extinguished faster than the aversion of age-matched controls. Our data establish that, under physiological APP expression levels, neprilysin deficiency is associated with increased Abeta accumulation in the brain and leads to deposition of amyloid-like structures in vivo as well as with signs of AD-like pathology and with behavioral deficits.     

Beta-amyloid racemized at the Ser26 residue in the brains of patients with Alzheimer disease: implications in the pathogenesis of Alzheimer disease

Oligomeric and fibrillar beta-amyloid (Abeta) may be toxic in Alzheimer disease (AD), especially after post-translation modification cumulative over time. Racemization of Ser and Asp residues of Abeta in senile plaques (SPs) occurs as an age-dependent process in AD. We previously reported that Abeta1-40 racemized at Ser26 is soluble and susceptible to proteolysis yielding toxic [D-Ser26]Abeta25-35/40 fragments in vitro and in vivo. Here, we focus on the localization of racemized Ser26 residues in AD brains within the limbic system, the earliest site of AD histopathology. We developed antisera (20.1 and 22.7). each with epitopes within [D-Ser26]Abeta25-40. Two forms of truncated [D-Ser26]Abeta were detected either in SPs or within neurons in all 11 AD-affected brains, but not in age-matched controls. [D-Ser26]Abeta25/26-35 (detected by 20.1) was localized to plaque cores, extracellular neurofibrillary "ghost" tangles and vascular amyloid deposits. In contrast, [D-Ser26]Abeta25-40 (detected by 22.7) was observed in most neurons containing intracellular neurofibrillary tangles, but not in SPs. These results suggest [D-Ser26]Abeta]1-40, formed during aging, becomes soluble and diffuses from SPs. It is then proteolyzed to [D-Ser26]Abeta25-35/40, which is toxic and may contribute to the neurodegeneration. This hypothesis may explain the long lag between SP formation and neurofibrillary degeneration in AD brains.     

Apoptotic neurons in Alzheimer's disease frequently show intracellular Abeta42 labeling

It is widely accepted that Abeta plays a pivotal role in the pathogenesis of Alzheimer's disease (AD) [27]. Attention has been focused mainly on how extracellular Abeta exerts its effects on neuronal cells [7,11,16,32]. However, neuronal degeneration from an accumulation of intracellular Abetax-42 (iAbeta42) occurs in presenilin 1 (PS1) mutant mice without extracellular Abeta deposits [5]. In the present study, intracellular deposits of iAbeta42 are correlated with apoptotic cell death in AD and PS-1 familial AD (PS1 FAD) brains by means of triple staining with antibodies to Abeta, TUNEL, and staining with Hoechst 33342. Neurons simultaneously positive for iAbeta42 and the TUNEL assay were significantly more abundant in AD brains than in controls. The number of apoptotic neurons with intracellular neurofibrillary tangles (iNFTs) was insignificant. Our results indicate that intraneuronal deposition of a neurotoxic form of Abeta seems to be an early event in the neurodegeneration of AD.     

Expression of LIF and LIF receptor beta in Alzheimer's and Parkinson's diseases

Background –  Signaling through the leukemia inhibitory factor (LIF) receptor (LIFR) is crucial for nervous system development. There are few studies concerning the expression of LIF and LIFR in normal and degenerating adult human brain.
Objectives –  To study the expression of LIF and LIFR in Alzheimer's disease (AD), Parkinson's disease (PD), and control brains.
Patients and methods –  LIF and LIFR mRNA copy numbers were determined by quantitative real-time RT-PCR from four brain regions of 34 patients with AD, 40 patients with PD, and 40 controls. Immunohistochemistry was performed in seven PD and in four AD patients and in seven normal controls.
Results –  In general, the LIF copy numbers were 1 log higher than the LIFR copy numbers. In the AD brains, LIF expression was higher than in the controls in the hippocampus and in the temporal cortex, and in the PD brains in the hippocampus and in the anterior cingulated cortex. Expressions of LIF and LIFR in different brain regions were opposite except for the AD hippocampus and PD anterior cingulated cortex, where the expression patterns were parallel.
Conclusions –  Co-operative expression of LIF and LIFR in AD hippocampus and PD anterior cingulated cortex may indicate a role for LIF in neuronal damage or repair in these sites.     

Amyloid-beta causes apoptosis of neuronal cells via caspase cascade, which can be prevented by amyloid-beta-derived short peptides

Amyloid beta 1-42 (Abeta42) and Abeta17-42 are major constituents of diffuse plaque in brains with Alzheimer's disease (AD). We demonstrate the potent cytotoxicity of Abeta42 and Abeta17-42, lesser toxicity of Abeta1-40 (Abeta40) and lack of toxicity of Abeta1-16 (Abeta16) in neuronal cells as measured by inhibition of cell proliferative response using thymidine incorporation assay and that this cytotoxicity can be reduced with Abeta16 and eight-residue Abeta derivatives such as Abeta1-8 and Abeta9-16. FACS analysis also revealed that Abeta16 could dramatically protect against the apoptosis induced by Abeta17-42 with over 80% viable cells. We determined the caspases involved in the Abeta-mediated apoptotic pathway using caspase-specific inhibitors in MTT assays. For all Abetas, the executor was caspase 3, while the initiator was caspase 9 for Abeta42 and caspase 8 for Abeta40 and Abeta17-42. Microscopic observation of lucifer-yellow-labeled neuronal cells demonstrated the occurrence of lysosomal membrane injury of the cells, corresponding to the severe cytotoxic effects of Abeta42. Our findings suggest that the apoptosis of neuronal cells due to Abeta42, Abeta40 and Abeta17-42 is mediated by the different caspase pathways and that this apoptosis can be reduced with the eight-residue Abeta-derived fragments Abeta1-8, Abeta9-16 and Abeta16.     

Cerebral ischemia and Alzheimer's disease: the expression of amyloid-beta and apolipoprotein E in human hippocampus

Growing evidence suggests a synergistic and perhaps etiological relationship between vascular disease and Alzheimer's disease (AD), which is characterized by the progressive accumulation of amyloid-beta peptide (Abeta). Moreover, apolipoprotein E (ApoE) has also been shown to be associated with AD and cerebral ischemia. It seems that cerebral ischemia may play an important, both direct and indirect, role in the pathogenesis of AD. We investigated the expression and distribution of Abeta1-40, beta1-42 and ApoE in human hippocampus after cerebral ischemia in this study to determine the role of cerebral ischemia in Alzheimer's disease. Our study has demonstrated that the accumulation of both Abeta1-40 and beta1-42 were increased dramatically and consistently after cerebral ischemia. Neuronal ApoE immunoreactivity was also significantly increased in all ischemic groups compared with controls. The most likely stimulus for the increased Abeta1-40, Abeta1-42 and ApoE immunoreactivity in the CA1 and CA3 neurons is the ischemic conditions, and their upregulation, in turn, may partly explain the contribution of cerebral ischemia to the pathogenesis of AD. Therefore our observations provide a basis for establishing therapeutic strategies aimed at preventing ischemic insults and subsequent neurodegeneration in AD.     

The levels of soluble versus insoluble brain Abeta distinguish Alzheimer's disease from normal and pathologic aging.

The abundance and solubility of Abeta peptides are critical determinants of amyloidosis in Alzheimer's disease (AD). Hence, we compared levels of total soluble, insoluble, and total Abeta1-40 and Abeta1-42 in AD brains with those in age-matched normal and pathologic aging brains using a sandwich enzyme-linked immunosorbent assay (ELISA). Since the measurement of Abeta1-40 and Abeta1-42 depends critically on the specificity of the monoclonal antibodies used in the sandwich ELISA, we first demonstrated that each assay is specific for Abeta1-40 or Abeta1-42 and the levels of these peptides are not affected by the amyloid precursor protein in the brain extracts. Thus, this sandwich ELISA enabled us to show that the average levels of total cortical soluble and insoluble Abeta1-40 and Abeta1-42 were highest in AD, lowest in normal aging, and intermediate in pathologic aging. Remarkably, the average levels of insoluble Abeta1-40 were increased 20-fold while the average levels of insoluble Abeta1-42 were increased only 2-fold in the AD brains compared to pathologic aging brains. Further, the soluble pools of Abeta1-40 and Abeta1-42 were the largest fractions of total Abeta in the normal brain (i.e., 50 and 23%, respectively), but they were the smallest in the AD brain (i.e., 2.7 and 0.7%, respectively) and intermediate (i.e., 8 and 0.8%, respectively) in pathologic aging brains. Thus, our data suggest that pathologic aging is a transition state between normal aging and AD. More importantly, our findings imply that a progressive shift of brain Abeta1-40 and Abeta1-42 from soluble to insoluble pools and a profound increase in the levels of insoluble Abeta1-40 plays mechanistic roles in the onset and/or progression of AD.     

Close association of water channel AQP1 with amyloid-β deposition in Alzheimer disease brains

Aquaporin-1 (AQP1), a membrane water channel protein, is expressed exclusively in the choroid plexus epithelium in the central nervous system under physiological conditions. However, AQP1 expression is enhanced in reactive astrocytes, accumulating in brain lesions of Creutzfeldt-Jakob disease and multiple sclerosis, suggesting a role of AQP1-expressing astrocytes in brain water homeostasis under pathological conditions. To clarify a pathological implication of AQP1 in Alzheimer disease (AD), we investigated the possible relationship between amyloid-beta (Abeta) deposition and astrocytic AQP1 expression in the motor cortex and hippocampus of 11 AD patients and 16 age-matched other neurological disease cases. In all cases, AQP1 was expressed exclusively in a subpopulation of multipolar fibrillary astrocytes. The great majority of AQP1-expressing astrocytes were located either on the top of or in close proximity to Abeta plaques in AD brains but not in non-AD cases, whereas those independent of Abeta deposition were found predominantly in non-AD brains. By Western blot, cultured human astrocytes constitutively expressed AQP1, and the levels of AQP1 protein expression were not affected by exposure to Abeta(1-42) peptide, but were elevated by hypertonic sodium chloride. By immunoprecipitation, the C-terminal fragment-beta (CTFbeta) of amyloid precursor protein interacted with the N-terminal half of AQP1 spanning the transmembrane helices H1, H2 and H3. These observations suggest the possible association of astrocytic AQP1 with Abeta deposition in AD brains.     

Amyloid beta up-regulates brain-derived neurotrophic factor production from astrocytes: rescue from amyloid beta-related neuritic degeneration

Astrocytes, the most abundant type of glia in the brain, are considered to play a key role in Alzheimer's disease (AD) pathologies. In a cell culture study, we have previously shown that astroglial responses against amyloid beta (Abeta) occur before obvious neuronal damage could be detected, suggesting the possibility that astrocytes might be an attractive therapeutic target for treating AD. In the present study, we investigated astroglial gene expression changes in response to Abeta to elucidate further the role of astrocytes in Abeta toxicity. By using real-time PCR and ELISA analyses, we found that Abeta rapidly induced astrocytes to produce brain-derived neurotrophic factor (BDNF). Abeta42 was more effective than Abeta40 in increasing astroglial BDNF production. Moreover, BDNF treatment rescued the neuronally differentiated human neuroblastoma cells from neuritic degeneration caused by Abeta toxicity. This is the first study to demonstrate that astrocytes are capable of increasing the production of a particular neurotrophic factor in response to Abeta. Our findings also identify BDNF as a potential therapeutic agent for preventing Abeta-related neuritic degeneration.     

Protective effects of EUK4010 on beta-amyloid(1-42) induced degeneration of neuronal cells

EUK4010 has been identified to exhibit an inhibitory effect on beta-amyloid (Abeta)(1-42)-induced loss of neuronal cell viability. Further studies demonstrated that EUK4010 attenuated the Abeta(1-42)-induced degeneration in both cultured rat hippocampal neurons and human neuroblastoma cells, as demonstrated by typical morphological changes, cell viability and the chip-based flow cytometric assay. Gene expression analysis using DNA microarray showed that the senescence marker calcium-binding protein, regucalcin (Rgn), GABA-A receptor pi subunit (Gabrp), the huntingtin binding protein, optineurin (Optn) and a semaphorin family plexin A3 similar protein (Plex-similar) changed their expression levels significantly in cultured neurons after Abeta(1-42) treatment. In this report, we have undertaken a chemical genetic approach to study the molecular basis of Abeta(1-42) effects on the neuronal degeneration. Our results demonstrate that EUK4010 completely blocked the Abeta(1-42)-induced up-regulation of GABA-A receptor pi subunit and the semaphorin family plexin A3 similar protein, and partially attenuated the down-regulation of senescence marker calcium-binding protein, regucalcin. These observations suggest that EUK4010 may prevent or reduce the Abeta toxicity by regulating the expression of genes involved in the Abeta induced neuronal degeneration. These genes may represent a promising target for the therapeutic drug development for Alzheimer's disease (AD) and other neurological disorders. Furthermore, EUK4010 and its analogues could potentially be developed as neuronal protective agents for the treatment of these diseases.     

Involvement of microglial receptor for advanced glycation endproducts (RAGE) in Alzheimer's disease: identification of a cellular activation mechanism

Receptor-mediated interactions with amyloid beta-peptide (Abeta) could be important in the evolution of the inflammatory processes and cellular dysfunction that are prominent in Alzheimer's disease (AD) pathology. One candidate receptor is the receptor for advanced glycation endproducts (RAGE), which can bind Abeta and transduce signals leading to cellular activation. Data are presented showing a potential mechanism for Abeta activation of microglia that could be mediated by RAGE and macrophage colony-stimulating factor (M-CSF). Using brain tissue from AD and nondemented (ND) individuals, RAGE expression was shown to be present on microglia and neurons of the hippocampus, entorhinal cortex, and superior frontal gyrus. The presence of increased numbers of RAGE-immunoreactive microglia in AD led us to further analyze RAGE-related properties of these cells cultured from AD and ND brains. Direct addition of Abeta(1-42) to the microglia increased their expression of M-CSF. This effect was significantly greater in microglia derived from AD brains compared to those from ND brains. Increased M-CSF secretion was also demonstrated using a cell culture model of plaques whereby microglia were cultured in wells containing focal deposits of immobilized Abeta(1-42). In each case, the Abeta stimulation of M-CSF secretion was significantly blocked by treatment of cultures with anti-RAGE F(ab')2. Treatment of microglia with anti-RAGE F(ab')2 also inhibited the chemotactic response of microglia toward Abeta(1-42). Finally, incubation of microglia with M-CSF and Abeta increased expression of RAGE mRNA. These microglia also expressed M-CSF receptor mRNA. These data suggest a positive feedback loop in which Abeta-RAGE-mediated microglial activation enhances expression of M-CSF and RAGE, possibly initiating an ascending spiral of cellular activation.     

Identification of peptides that specifically bind Abeta1-40 amyloid in vitro and amyloid plaques in Alzheimer''s disease brain using phage display

The accumulation of the amyloid-beta (Abeta) peptides in amyloid plaques correlates with pathologic changes that occur in the brains of patients with Alzheimer's disease (AD). The ability to directly target reagents to the amyloid form of the Abeta peptide may allow the delivery of neuroprotective agents to make amyloid plaques less toxic, the delivery of amyloid-destroying molecules to eliminate plaques, or the delivery of reagents to prevent amyloid plaque formation. In addition, such reagents may be useful as diagnostic tools to quantitate the extent of amyloid plaque formation in AD patients. As a step toward these goals, we have used phage peptide display technology to identify peptides that bind specifically to the amyloid form of the Abeta(1-40) peptide. Here we identify two 20-amino acid peptides with similar structural features that bind to the amyloid form of Abeta(1-40) but not to monomeric Abeta(1-40). A recombinant form of one of these peptides was produced in Escherichia coli as a fusion protein with thioredoxin. After purification, this reagent bound Abeta(1-40) amyloid in vitro with a K(d) of 60 nM and specifically labeled amyloid plaques in AD brains. A chemically synthesized version of this peptide also bound Abeta(1-40) amyloid and specifically stained amyloid plaques in AD brain. These peptide sequences represent new potential carrier molecules to deliver medicines to amyloid plaques in AD patients and to image plaques in AD brains.     

The severity of cortical Alzheimer's type changes is positively correlated with increased amyloid-beta Levels: Resolubilization of amyloid-beta with transition metal ion chelators

The most consistent diagnostic neuropathological lesion in Alzheimer's disease (AD) is the senile plaque of which the 4 kD amyloid-beta (Abeta) peptide is the major proteinaceous component. In this study cortical Abeta levels were immunochemically measured in 70 post-mortem human brains and compared against their neuropathological grading as determined by the densities of amyloid plaques and neurofibrillary tangles. The mean concentration of cortical Abeta/mg protein increased with the severity of the cortical degenerative changes (AD0 < AD1 < AD2 < AD3). Brains with the severe degenerative changes (AD3), corresponded to definite AD cases and exhibited significantly increased concentrations of Abeta (11.1+/-3.08 ng/mg total protein, n=17) when compared with control brains without any degenerative changes (AD0; 0.06+/-0.06 ng/mg total protein, n=14,P=0.003). The extraction of Abeta from the cortex of AD3 brains was significantly enhanced in a dose dependent manner by the presence of the metal ion chelator N,N,N',N'-tetrakis(2-pyridylmethyl) ethylenediamine (5 mM TPEN, P < 0.0001). The chelator/antioxidant 1,2-dithiolane-3-pentanoic acid (lipoic acid), also resolubilized Abetain a dose-dependant manner. Both chelators also enhanced the extraction of Abeta from the frontal cortex of AbetaPP-transgenic mice suggesting this animal model of amyloidosis may be useful for evaluating the biochemical and therapeutic effects of chelators/antioxidants on Abeta deposition. In summary our results indicate that increased Abeta load is correlated with the severity of the cortical AD-type changes and that chelators/antioxidants may be useful in reducing neuronal amyloid burden.     

Codeposition of cystatin C with amyloid-beta protein in the brain of Alzheimer disease patients

Immunohistochemical analysis of brains of patients with Alzheimer disease (AD) revealed that the cysteine proteinase inhibitor cystatin C colocalizes with amyloid beta-protein (Abeta) in parenchymal and vascular amyloid deposits. No evidence of cerebral hemorrhage was observed in any of the brains studied. Immunoelectron microscopy demonstrated dual staining of amyloid fibrils with anti-Abeta and anti-cystatin C antibodies. Cystatin C immunoreactivity was also observed in amyloid deposits in the brain of transgenic mice overexpressing human beta amyloid precursor protein. Massive deposition of the variant cystatin C in the cerebral vessels of patients with the Icelandic form of hereditary cerebral hemorrhage with amyloidosis is thought to be responsible for the pathological processes leading to stroke. Anti-cystatin C antibodies strongly labeled pyramidal neurons within cortical layers most prone to amyloid deposition in the brains of AD patients. Immunohistochemistry with antibodies against the carboxyl-terminus of Abeta(x-42) showed intracellular immunoreactivity in the same neuronal subpopulation. It remains to be established whether the association of cystatin C to Abeta plays a primary role in amyloidogenesis of AD or is a late event in which the protein is bound to the previously formed Abeta amyloid fibrils.     

RAGE-Abeta interactions in the pathophysiology of Alzheimer's disease

RAGE is a cell surface molecule primarily identified for its capacity to bind advanced glycation end-products and amphoterin. Immunocytochemical studies demonstrated that in Alzheimer's Disease (AD) the expression of RAGE is elevated in neurons close to neuritic plaque beta-amyloid (Abeta) deposits and in the cells of Abeta containing vessels. Cross-linking of surface bound Abeta 1-40 to endothelial cells, yielded a band of 50 kDa identified as RAGE. Using the soluble extracellular domain of recombinant human RAGE, we found that Abeta binds to RAGE with a Kd = 57 +/- 14 nM, a value close to those found for mouse brain endothelial cells and rat cortical neurons. The interaction of Abeta with RAGE in neuronal, endothelial, and RAGE-transfected COS-1 cells induced oxidative stress, as assessed by the TBARS and MTT assays. ELISA demonstrated a 2.5 times increase of RAGE in AD over control brains. Activated microglia also showed elevated expression of RAGE. In the BV-2 microglial cell line, RAGE bound Abeta in dose dependent manner with a Kd of 25 +/- 9 nM. Soluble Abeta induced the migration of microglia along a concentration gradient, while immobilized Abeta arrested this migration. Abeta-RAGE interaction also activated NF-kappaB, resulting in neuronal up-regulation of macrophage-colony stimulating factor (M-CSF) which also induced microglial migration. Taken together, our data suggest that RAGE-Abeta interactions play an important role in the pathophysiology of Alzheimer's Disease.     

A possible model of senile plaques using synthetic amyloid beta-protein and rat glial culture

The senile plaque (SP) is one of the pathological hallmarks in the brains of patients with Alzheimer's disease (AD), but the mechanism of its formation and its role in AD progression are not yet fully understood. Synthetic amyloid beta-protein (Abeta)1-40 is known to aggregate in vitro, and the aggregated Abeta has been widely used for in vitro experiments, in which its peculiar effects on neuronal and glial cells have been shown. To date, however, the formation of a SP-like structure in a culture system using synthetic Abeta has not been demonstrated. In this study, we established a possible SP model using synthetic Abeta1-40 and rat glial cultures as follows: (1) large spherical aggregates of synthetic Abeta (sAmys) were produced from synthetic Abeta1-40 (10-50 microm in diameter), (2) the sAmys were added to a glial culture, and (3) the characteristics of the sAmys and the reactions of glial cells (microglia and astrocytes) around the sAmys were analyzed. We found that the sAmys exhibited the same features as the dense amyloid core in SPs, including the intense green birefringence under polarized light with Congo red, and induced reactive features in glial cells, including induction of major histocompatibility complex class II antigen in the microglia and interleukin-1beta in the astrocytes, similar to those seen in SPs in the brain in AD. Given our findings, we consider that this glial culture system with the sAmys is a possible in vitro SP model and useful for investigating the effects of massive amyloid deposition on neuronal and glial cells.     

Copper levels are increased in the cerebral cortex and liver of APP and APLP2 knockout mice

The pathological process in Alzheimer's disease (AD) involves amyloid beta (Abeta) deposition and neuronal cell degeneration. The neurotoxic Abeta peptide is derived from the amyloid precursor protein (APP), a member of a larger gene family including the amyloid precursor-like proteins, APLP1 and APLP2. The APP and APLP2 molecules contain metal binding sites for copper and zinc. The zinc binding domain (ZnBD) is believed to have a structural rather than a catalytic role. The activity of the copper binding domain (CuBD) is unknown, however, APP reduces copper (II) to copper (I) and this activity could promote copper-mediated neurotoxicity. The expression of APP and APLP2 in the brain suggests they could have an important direct or indirect role in neuronal metal homeostasis. To examine this, we measured copper, zinc and iron levels in the cerebral cortex, cerebellum and selected non-neuronal tissues from APP (APP(-/-)) and APLP2 (APLP2(-/-)) knockout mice using atomic absorption spectrophotometry. Compared with matched wild-type (WT) mice, copper levels were significantly elevated in both APP(-/-) and APLP2(-/-) cerebral cortex (40% and 16%, respectively) and liver (80% and 36%, respectively). Copper levels were not significantly different between knockout and WT cerebellum, spleen or serum samples. There were no significant differences observed between APP(-/-), APLP2(-/-) and WT mice zinc or iron levels in any tissue examined. These findings indicate APP and APLP2 expression specifically modulates copper homeostasis in the liver and cerebral cortex, the latter being a region of the brain particularly involved in AD. Perturbations to APP metabolism and in particular, its secretion or release from neurons may alter copper homeostasis resulting in increased Abeta accumulation and free radical generation. These data support a novel mechanism in the APP/Abeta pathway which leads to AD.     

α7 nicotinic acetylcholine receptor expression in Alzheimer's disease

The brains of people with Alzheimer's disease (AD) display several characteristic pathological features, including deposits (plaques) of beta-amyloid 1-42 (Abeta1-42), intraneuronal accumulations (tangles) of hyperphosphorylated tau, degeneration of the basal forebrain cholinergic pathway, and gliosis. Abeta1-42 plaques develop in specific brain regions, including hippocampus and cortex, as well as in the vasculature. Abeta1-42 might promote neurodegeneration through the induction of free radicals and disruption of Ca2+ homeostasis, giving rise to the symptoms of AD. Abeta1-42 interacts with the alpha7 subtype of the nicotinic acetylcholine receptor (alpha7 nAChR), which is widely expressed throughout the central and peripheral nervous systems, as well as in several nonneuronal loci, such as epithelial cells, lymphoid tissues, and peripheral blood lymphocytes. Western blot and autoradiographic analyses indicate that the alpha7 nAChR subunit protein is up-regulated in human brain samples from Alzheimer patients, as well as in animal models of AD (Dineley et al., 2001; Bednar et al., 2002), and might be involved in nicotine-mediated reduction of Abeta1-42 deposition (Hellstrom et al., 2004), although the nature of this relationship remains ill-defined. We have undertaken a semiquantitative histological evaluation of alpha7 nAChR expression in a mouse model of AD pathology, as well as a comparison of alpha7 nAChR levels in lymphocytes from AD patients and control subjects.