Abnormal Processing Of Amyloid Precursor Protein Research Articles

Abnormal amyloid precursor protein processing in periodontal tissue in a murine model of periodontitis induced by Porphyromonas gingivalis.

The study aimed to investigate the change of amyloid precursor protein (APP) processing and amyloid β (Aβ) metabolites in linking periodontitis to Alzheimer's disease (AD). Aβ is one of the main pathological features of AD, and few studies have discussed changes in its expression in peripheral tissues or analyzed the relationship between the peripheral imbalance of Aβ production and clearance. A murine model of periodontitis was established by oral infection with Porphyromonas gingivalis (P. gingivalis). Micro-computed tomography (Micro-CT) was used to observe the destruction of the alveolar bone. Nested quantitative polymerase chain reaction (qPCR) was used to measure small quantities of P.gingivalis DNA in different tissues. Behavioral experiments were performed to measure cognitive function in the mice. The mRNA levels of TNF-α, IL-6, IL-8, RANKL, OPG, APP695, APP751, APP770, and BACE1 in the gingival tissues or cortex were detected by RT-PCR. The levels of Aβ1-40 and Aβ1-42 in gingival crevicular fluid (GCF) and plasma were tested by ELISA. P. gingivalis oral infection was found to cause alveolar bone resorption and impaired learning and memory. P.gingivalis DNA was detected in the gingiva, blood and cortex of the P.gingivalis group by nested qPCR (p < .05). The mRNA expression of TNF-α, IL-6, IL-8, RANKL/OPG, and BACE1 in the gingival tissue was significantly higher than that in the control group (p < .05). Similarly, upregulated mRNA levels of APP695 and APP770 were observed in the gingival tissuses and cortex of the P. gingivalis group (p < .05). The levels of Aβ1-40 and Aβ1-42 in the GCF and plasma of the P. gingivalis group were significantly higher than those in the control group (p < .05). P. gingivalis can directly invade the brain via hematogenous infection. The invasion of P. gingivalis could trigger an immune response and lead to an imbalance between Aβ production and clearance in peripheral tissues, which may trigger an abnormal Aβ metabolite in the brain, resulting in the occurrence and development of AD.

Activated STAT3 signaling pathway by ligature-induced periodontitis could contribute to neuroinflammation and cognitive impairment in rats

BackgroundIncreasing evidence suggests a causal link between periodontitis and cognitive disorders. Systemic inflammation initiated by periodontitis may mediate the development of cognitive impairment. Our study aims to investigate the effect of ligature-induced periodontitis on cognitive function and the role of signal transducers and activators of transcription 3 (STAT3) in this process.Materials and methodsLigature-induced periodontitis was established, and the rats were treated intraperitoneally with/without the pSTAT3 inhibitor cryptotanshinone (CTS). Alveolar bone resorption and periodontal inflammation were detected by micro-computed tomography analysis and histopathological evaluation. Locomotor activity and cognitive function were evaluated by the open field test and the Morris water maze test, respectively. The activation of microglia and astrocytes in the hippocampus and cortex was assessed by immunohistochemistry (IHC). The expression of interleukins (IL-1β, IL-6, IL-8, IL-21) in both the periphery and cortex was evaluated by RT-PCR and ELISA. The expression of TLR/NF-κB and ROS cascades was evaluated by RT-PCR. The expression of pSTAT3 and the activation of the STAT3 signaling pathway (JAK2, STAT3, and pSTAT3) in the periodontal tissue and cortex were assessed by IHC and Western blot. The expression of amyloid precursor protein (APP) and its key secretases was evaluated by RT-PCR. The level of amyloid β-protein (Aβ) and the ratio of Aβ1-40/1-42 were measured via ELISA in the plasma and cortex while IHC was used to detect the level of Aβ1-42 in the brain.ResultsIn periodontal ligature rats, significant alveolar bone resorption and local inflammatory cell infiltration were present. Apparent increases in inflammatory cytokines (IL-1β, IL-6, IL-8, and IL-21) were detected in peripherial blood and brain. Additionally, spatial learning and memory ability was impaired, while locomotor activity was not affected. Activated microglia and astrocytes were found in the cortex and hippocampus, presenting as enlarged cell bodies and irregular protrusions. Levels of TLR/NF-kB, PPAR and ROS were altered. The STAT3 signaling pathway was activated in both the periodontal tissue and cortex, and the processing of APP by β- and γ-secretases was promoted. The changes mentioned above could be relieved by the pSTAT3 inhibitor CTS.ConclusionsLigature-induced periodontitis in rats resulted in systemic inflammation and further abnormal APP processing, leading to cognitive impairments. In this progress, the activation of the STAT3 signaling pathway may play an important role by increasing inflammatory load and promoting neuroinflammation.

Periodontitis Induced by P. gingivalis-LPS Is Associated With Neuroinflammation and Learning and Memory Impairment in Sprague-Dawley Rats.

BackgroundPeriodontitis is one of the most common oral diseases and is a potential risk factor for systemic diseases. In this study, we aimed to investigate the association between periodontitis and learning and memory impairment.MethodsWe established a periodontitis model by topical application of Porphyromonas gingivalis lipopolysaccharide (P. gingivalis-LPS) into the palatal gingival sulcus of the maxillary first molars of 10-week-old male rats for a 10-week period. We assessed alveolar bone resorption using micro–computed tomography analysis and learning and memory ability using the Morris water maze test. We determined the levels of cytokines [interleukin (IL)-1β, IL-6, IL-8, and IL-21] and LPS in the peripheral blood and cortex, as well as toll-like receptor 4 (TLR4)/NF-κB signaling pathway activation, using reverse transcription-polymerase chain reaction (RT-PCR), enzyme-linked immunosorbent assay (ELISA), and western blot. We determined activation of microglia and astrocytes, expression of Aβ1-42, APP and Tau by immunohistochemistry. Finally, we measured the expression of amyloid precursor protein (APP) and its key secretases, as well as the Aβ1-40/1-42 ratio, by RT-PCR, western blot, and ELISA.ResultsWe found that periodontitis induced learning and memory impairment in the rats. Further, we observed that it induced significant alveolar bone resorption. There was an increase in the levels of inflammatory cytokines and LPS. Moreover, we confirmed TLR4/NF-κB signaling pathway activation. We also observed activated microglia and astrocytes with enlarged cell bodies and irregular protrusions. Finally, we observed the promotion of β- and γ-secretases APP processing.ConclusionOur findings indicated that periodontitis was associated with learning and memory impairment, probably induced by neuroinflammation via activating the TLR4/NF-κB signaling pathway. Furthermore, abnormal APP processing could be involved in this progress.

Involvement of Lipids in Alzheimer's Disease Pathology and Potential Therapies.

Lipids constitute the bulk of the dry mass of the brain and have been associated with healthy function as well as the most common pathological conditions of the brain. Demographic factors, genetics, and lifestyles are the major factors that influence lipid metabolism and are also the key components of lipid disruption in Alzheimer’s disease (AD). Additionally, the most common genetic risk factor of AD, APOE ϵ4 genotype, is involved in lipid transport and metabolism. We propose that lipids are at the center of Alzheimer’s disease pathology based on their involvement in the blood-brain barrier function, amyloid precursor protein (APP) processing, myelination, membrane remodeling, receptor signaling, inflammation, oxidation, and energy balance. Under healthy conditions, lipid homeostasis bestows a balanced cellular environment that enables the proper functioning of brain cells. However, under pathological conditions, dyshomeostasis of brain lipid composition can result in disturbed BBB, abnormal processing of APP, dysfunction in endocytosis/exocytosis/autophagocytosis, altered myelination, disturbed signaling, unbalanced energy metabolism, and enhanced inflammation. These lipid disturbances may contribute to abnormalities in brain function that are the hallmark of AD. The wide variance of lipid disturbances associated with brain function suggest that AD pathology may present as a complex interaction between several metabolic pathways that are augmented by risk factors such as age, genetics, and lifestyles. Herewith, we examine factors that influence brain lipid composition, review the association of lipids with all known facets of AD pathology, and offer pointers for potential therapies that target lipid pathways.

Protective effects of BACE1 inhibitory ligand molecules against amyloid beta-induced synaptic and mitochondrial toxicities in Alzheimer's disease.

Amyloid-β (Aβ) peptides are the major drivers of Alzheimer's disease (AD) pathogenesis, and are formed by successive cleavage of the amyloid precursor protein (APP) by the beta and gamma secretases. Mounting evidence suggests that Aβ and mitochondrial structural and functional abnormalities are critically involved in the loss of synapses and cognitive decline, in patients with AD. In AD brain, state the sequential proteolytic cleavage of APP by beta secretase 1 enzyme (BACE1) and γ-secretase leads to the production and release of Aβ40 and 42. BACE1 expression and activity increased in the brains of AD patients. Structurally, β-secretase has a very large binding site (1000Å) with fewer hydrophobic domains that makes a challenge to identify the specific targets/binding sites of BACE1. In the present study, we constructed a BACE1 pharmacophore with pepstatin and screened through molecular docking studies. We found one potential candidate (referred as ligand 1) that binds to the key catalytic residues of BACE1 and predicts to inhibit abnormal APP processing and reduce Aβ levels in AD neurons. Using biochemical, molecular, transmission electron microscopy, immunoblotting and immunofluorescence analyses, we studied the protective effects of ligand 1 against Aβ-induced synaptic and mitochondrial toxicities in mouse neuroblastoma (N2a) cells that express mutant APP. We found interaction between ligand 1 and BACE1 and this interaction decreased BACE1 activity, Aβ40 and 42 levels. We also found increased mitochondrial biogenesis, mitochondrial fusion and synaptic activity and reduced mitochondrial fission in ligand 1-treated mutant APP cells. Based on these results, we cautiously conclude that ligand 1 reduces Aβ-induced mitochondrial and synaptic toxicities, and maintains mitochondrial dynamics and neuronal function in AD.

Novel MicroRNA-455-3p and its protective effects against abnormal APP processing and amyloid beta toxicity in Alzheimer's disease

The purpose of our study is to understand the protective role of miR-455-3p against abnormal amyloid precursor protein (APP) processing, amyloid beta (Aβ) formation, defective mitochondrial biogenesis/dynamics and synaptic damage in AD progression. In-silico analysis of miR-455-3p has identified the APP gene as a putative target. Using mutant APP cells, miR-455-3p construct, biochemical and molecular assays, immunofluorescence and transmission electron microscopy (TEM) analyses, we studied the protective effects of miR-455-3p on – 1) APP regulation, amyloid beta (Aβ)(1–40) & (1–42) levels, mitochondrial biogenesis & dynamics; 3) synaptic activities and 4) cell viability & apoptosis. Our luciferase reporter assay confirmed the binding of miR-455-3p at the 3’UTR of APP gene. Immunoblot, sandwich ELISA and immunostaining analyses revealed that the reduced levels of the mutant APP, Aβ(1–40) & Aβ(1–42), and C99 by miR-455-3p. We also found the reduced levels of mRNA and proteins of mitochondrial biogenesis (PGC1α, NRF1, NRF2, and TFAM) and synaptic genes (synaptophysin and PSD95) in mutant APP cells; on the other hand, mutant APP cells that express miR-455-3p showed increased mRNA and protein levels of biogenesis and synaptic genes. Additionally, expression of mitochondrial fission proteins (DRP1 and FIS1) were decreased while the fusion proteins (OPA1, Mfn1 and Mfn2) were increased by miR-455-3p. Our TEM analysis showed a decrease in mitochondria number and an increase in the size of mitochondrial length in mutant APP cells transfected with miR-455-3p. Based on these observations, we cautiously conclude that miR-455-3p regulate APP processing and protective against mutant APP-induced mitochondrial and synaptic abnormalities in AD.

Analyzing the Behavior of Neuronal Pathways in Alzheimer's Disease Using Petri Net Modeling Approach.

Alzheimer's Disease (AD) is the most common neuro-degenerative disorder in the elderly that leads to dementia. The hallmark of AD is senile lesions made by abnormal aggregation of amyloid beta in extracellular space of brain. One of the challenges in AD treatment is to better understand the mechanism of action of key proteins and their related pathways involved in neuronal cell death in order to identify adequate therapeutic targets. This study focuses on the phenomenon of aggregation of amyloid beta into plaques by considering the signal transduction pathways of Calpain-Calpastatin (CAST) regulation system and Amyloid Precursor Protein (APP) processing pathways along with Ca2+ channels. These pathways are modeled and analyzed individually as well as collectively through Stochastic Petri Nets for comprehensive analysis and thorough understating of AD. The model predicts that the deregulation of Calpain activity, disruption of Calcium homeostasis, inhibition of CAST and elevation of abnormal APP processing are key cytotoxic events resulting in an early AD onset and progression. Interestingly, the model also reveals that plaques accumulation start early (at the age of 40) in life but symptoms appear late. These results suggest that the process of neuro-degeneration can be slowed down or paused by slowing down the degradation rate of Calpain-CAST Complex. In the light of this study, the suggestive therapeutic strategy might be the prevention of the degradation of Calpain-CAST complexes and the inhibition of Calpain for the treatment of neurodegenerative diseases such as AD.

Distinct amyloid precursor protein processing machineries of the olfactory system

Processing of amyloid precursor protein (APP) occurs through sequential cleavages first by β-secretase and then by the γ-secretase complex. However, abnormal processing of APP leads to excessive production of β-amyloid (Aβ) in the central nervous system (CNS), an event which is regarded as a primary cause of Alzheimer's disease (AD). In particular, gene mutations of the γ-secretase complex—which contains presenilin 1 or 2 as the catalytic core—could trigger marked Aβ accumulation.Olfactory dysfunction usually occurs before the onset of typical AD-related symptoms (eg, memory loss or muscle retardation), suggesting that the olfactory system may be one of the most vulnerable regions to AD. To date however, little is known about why the olfactory system is affected so early by AD prior to other regions. Thus, we examined the distribution of secretases and levels of APP processing in the olfactory system under either healthy or pathological conditions.Here, we show that the olfactory system has distinct APP processing machineries. In particular, we identified higher expressions levels and activity of γ-secretase in the olfactory epithelium (OE) than other regions of the brain. Moreover, APP c-terminal fragments (CTF) are markedly detected. During AD progression, we note increased expression of presenilin2 of γ-secretases in the OE, not in the OB, and show that neurotoxic Aβ*56 accumulates more quickly in the OE.Taken together, these results suggest that the olfactory system has distinct APP processing machineries under healthy and pathological conditions. This finding may provide a crucial understanding of the unique APP-processing mechanisms in the olfactory system, and further highlights the correlation between olfactory deficits and AD symptoms.

Alzheimer-related decrease in CYFIP2 links amyloid production to tau hyperphosphorylation and memory loss.

Characteristic features of Alzheimer's disease are memory loss, plaques resulting from abnormal processing of amyloid precursor protein (APP), and presence of neurofibrillary tangles and dystrophic neurites containing hyperphosphorylated tau. Currently, it is not known what links these abnormalities together. Cytoplasmic FMR1 interacting protein 2 (CYFIP2) has been suggested to regulate mRNA translation at synapses and this may include local synthesis of APP and alpha-calcium/calmodulin-dependent kinase II, a kinase that can phosphorylate tau. Further, CYFIP2 is part of the Wiskott-Aldrich syndrome protein-family verprolin-homologous protein complex, which has been implicated in actin polymerization at synapses, a process thought to be required for memory formation. Our previous studies on p25 dysregulation put forward the hypothesis that CYFIP2 expression is reduced in Alzheimer's disease and that this contributes to memory impairment, abnormal APP processing and tau hyperphosphorylation. Here, we tested this hypothesis. First, in post-mortem tissue CYFIP2 expression was reduced by ∼50% in severe Alzheimer's hippocampus and superior temporal gyrus when normalized to expression of a neuronal or synaptic marker protein. Interestingly, there was also a trend for decreased expression in mild Alzheimer's disease hippocampus. Second, CYFIP2 expression was reduced in old but not in young Tg2576 mice, a model of familial Alzheimer's disease. Finally, we tested the direct impact of reduced CYFIP2 expression in heterozygous null mutant mice. We found that in hippocampus this reduced expression causes an increase in APP and β-site amyloid precursor protein cleaving enzyme 1 (BACE1) protein, but not mRNA expression, and elevates production of amyloid-β42 Reduced CYFIP2 expression also increases alpha-calcium/calmodulin-dependent kinase II protein expression, and this is associated with hyperphosphorylation of tau at serine-214. The reduced expression also impairs spine maturity without affecting spine density in apical dendrites of CA1 pyramidal neurons. Furthermore, the reduced expression prevents retention of spatial memory in the water maze. Taken together, our findings indicate that reduced CYFIP2 expression triggers a cascade of change towards Alzheimer's disease, including amyloid production, tau hyperphosphorylation and memory loss. We therefore suggest that CYFIP2 could be a potential hub for targeting treatment of the disease.

Early striatal amyloid deposition distinguishes down syndrome and autosomal dominant Alzheimer’s disease from late-onset amyloid deposition

To evaluate the early PiB PET amyloid deposition patterns in three different groups of cerebral amyloidosis: 1) amyloid precursor protein (APP) overproduction [i.e., Down's syndrome (DS)]; abnormal processing of APP [i.e., Autosomal dominant Alzheimer disease (ADAD)]; and 3) presumed alterations in amyloid-beta (Aβ) clearance [i.e., late-onset AD (LOAD)]. We performed a whole brain, voxel-wise comparison of cerebral amyloid between 23 DS, 10 ADAD and 16 elderly controls, all nondemented, that had evidence of cerebral amyloid deposition, based on PiB-PET. Compared to the elderly control group we found both DS and ADAD shared a distinct pattern of increased bilateral striatal and thalamic amyloid deposition. Additionally, we found our ADAD group to have increased amyloid in the lingual gyrus bilaterally compared to the DS subjects. There was a trend towards increased parietal amyloid in the elderly controls compared to the other two groups.

Mitochondria-targeted catalase reduces abnormal APP processing, amyloid production and BACE1 in a mouse model of Alzheimer's disease: implications for neuroprotection and lifespan extension

The purpose of this study was to investigate the protective effects of the mitochondria-targeted antioxidant catalase (MCAT) and lifespan extension in mice that express amyloid beta (Aβ). Using immunoblotting and immunostaining analyses, we measured the production of full-length amyloid precursor protein (APP), soluble APPα, C-terminal fragments CTF99 and CTF83, monomeric and oligomeric Aβ, Aβ deposits and beta site amyloid precursor protein cleaving enzyme 1 (BACE1), in different stages of disease progression in MCAT/AβPP and AβPP mice. Using quantitative reverse transcriptase polymerase chain reaction and immunostaining analyses, we studied the expression of catalase, BACE1, the Alzheimer's disease (AD) markers, synaptophysin, APP, neprilysin, insulin-degrading enzymeand transthyretin in MCAT, AβPP, MCAT/AβPP and wild-type (WT) mice. Using the high pressure liquid chromatography analysis of 8-hydroxy-2-deoxyguanosine, we measured oxidative DNA damage in the cerebral cortical tissues from MCAT, AβPP, MCAT/AβPP and WT mice. We found that the AβPP transgenic mice that carried the human MCAT gene lived 5 months longer than did the AβPP mice. We also found that the overexpression of MCAT in the brain sections from the MCAT/AβPP transgenic mice significantly correlated with a reduction in the levels of full-length APP, CTF99, BACE1, Aβ levels (40 and 42), Aβ deposits and oxidative DNA damage relative to the brain sections from the AβPP mice. Interestingly, we found significantly increased levels of soluble APPα and CTF83 in the MCAT/AβPP mice, relative to the AβPP mice. These data provide direct evidence that oxidative stress plays a primary role in AD etiopathology and that in MCAT mice express Aβ, MCAT prevents abnormal APP processing, reduces Aβ levels and enhances Aβ-degrading enzymes in mice at different ages, corresponding to different stages of disease progression. These findings indicate that mitochondria-targeted molecules may be an effective therapeutic approach to treat patients with AD.

Delineating Alzheimer‘s Disease Progression with MGAT3, a Biomarker for Improved Prognosis and Personalized Therapy

Blood biomarkers have been based either on biochemical tests, such as plasma cytokine antibody assays, which require ana lysis of 18 signaling proteins [4], or functional tests, such as Ab phagocytosis and mRNA transcription by flow cytometry [5]. ApoE4 is a well-known risk factor for AD [6], and plasma apoE levels are decreased but not in all studies [7]. The multiple mechanisms common to neurodegenerative diseases, including inflammation, oxidative stress and dyslipidemia, render the goal of a specific protein blood biomarker elusive. In the brain, the defects in neuronal autophagy [8] and the retromer sorting pathway [9] are considered the crucial neuronal mechanisms leading to AD. The mechanisms are postulated to include the disruption of substrate proteolysis within autolysosomes in the former and abnormal amyloid-precursor protein processing in the latter. However, these mechanisms cannot be examined in living patients by biochemical ana lysis, limiting their diagnostic value. Here, we discuss an alternative approach based on a hypothesis that the immune system has a high sensitivity for the disease process, including abnormal degradation of Ab in lysosomes. Peripheral blood mononuclear cells (PBMCs) are defective in phagocytosis and degradation of Ab [5,10]. In the AD brain, the distinction between macrophages and microglia is not clear but both appear to originate from the blood in pathological states (histochemistry shows macro phages migrating across a vessel wall [11] and mouse studies show the microglia to have bone-marrow origin [12]). The defects of AD macrophages also involve their trafficking – inability to emigrate across the blood–brain barrier and apoptotic death of macrophages engorged with Ab around brain vessels contributing to congophilic angiopathy [13]. According to this Studies of the neuronal mechanisms of Alzheimer’s disease (AD) have discovered new therapeutic targets but not biomarkers for personalized medicine of AD. Exploitation of immune responses to amyloid-b (Ab) by peripheral blood mononuclear cells led to the discovery of two biomarkers: phagocytosis of amyloid-b and transcription of MGAT3 (GnT‐III). A pilot study suggests these two markers will be valuable for AD diagnosis, prognosis and therapy by vitamin D3 and curcuminoids [1].

The amyloid cascade hypothesis has misled the pharmaceutical industry

The pharmaceutical industry has invested a great deal of time and finance in the development of therapeutics targeting amyloid generation, signalling and plaque stability. This has been based on the amyloid cascade hypothesis which states that abnormal amyloid precursor protein processing and the formation of amyloid plaques is the central process in the development of the symptoms of Alzheimer's disease. However, most clinical trials in this area have been disappointing; therefore the attendees of the Models of Dementia: the Good, the Bad and the Future meeting were given the opportunity to openly debate the proposal 'the amyloid cascade has misled the pharmaceutical industry', with the main contributions from Professor John Hardy and Professor John Mayer. The present article is a representation of the debate.

Nicastrin Is Dispensable for γ-Secretase Protease Activity in the Presence of Specific Presenilin Mutations

γ-Secretase is a multisubunit membrane protein complex consisting of presenilin (PS1), nicastrin (NCT), anterior pharynx-1, and presenilin enhancer 2. To analyze the activity of familial Alzheimer disease mutants and to understand the roles of the subunits, we established a yeast transcriptional activator Gal4p system with artificial γ-secretase substrates containing amyloid precursor protein or Notch fragments. The γ-secretase activities were evaluated by transcriptional activation of reporter genes upon Gal4p release from the membrane-bound substrates, i.e. growth of yeast on histidine and adenine, or β-galactosidase assay. We screened and evaluated γ-secretase mutants using this reconstitution system in yeast, which does not possess endogenous γ-secretase activity. When we introduced familial Alzheimer mutants of PS1 in this system, their activities were shown to be loss of function. Although the protease activity of wild type PS1 depends on the other three subunits introduced, we obtained 15 new PS1 mutants, which are active in the absence of NCT. They possessed a S438P mutation at the ninth transmembrane domain (TM9) together with one missense mutation distributed through transmembrane and loop regions. These mutations were not related to familial Alzheimer mutations of PS1 as identified so far. The S438P mutant was partially active but required other mutations for full activation. Results of the β-galactosidase assay suggested that they have wild type protease activities, which were further confirmed by the endoproteolysis of PS1, amyloid β peptides, and Notch intracellular domain production in mammalian cells. These results suggest that NCT is dispensable for the protease activity of γ-secretase.

Hyperhomocysteinemia Increases β-Amyloid by Enhancing Expression of γ-Secretase and Phosphorylation of Amyloid Precursor Protein in Rat Brain

Hyperhomocysteinemia and beta-amyloid (Abeta) overproduction are critical etiological and pathological factors in Alzheimer disease, respectively; however, the intrinsic link between them is still missing. Here, we found that Abeta levels increased and amyloid precursor protein (APP) levels simultaneously decreased in hyperhomocysteinemic rats after a 2-week induction by vena caudalis injection of homocysteine. Concurrently, both the mRNA and protein levels of presenilin-1, a component of gamma-secretase, were elevated, whereas the expression levels of beta-secretase and presenilin-2 were not altered. We also observed that levels of phosphorylated APP at threonine-668, a crucial site facilitating the amyloidogenic cleavage of APP, increased in rats with hyperhomocysteinemia, although the phosphorylation per se did not increase the binding capacity of pT668-APP to the secretases. The enhanced phosphorylation of APP in these rats was not relevant to either c-Jun N-terminal kinase or cyclin-dependent kinase-5. A prominent spatial memory deficit was detected in rats with hyperhomocysteinemia. Simultaneous supplementation of folate and vitamin-B12 attenuated the hyperhomocysteinemia-induced abnormal processing of APP and improved memory. Our data revealed that hyperhomocysteinemia could increase Abeta production through the enhanced expression of gamma-secretase and APP phosphorylation, causing memory deficits that could be rescued by folate and vitamin-B12 treatment in these rats. It is suggested that hyperhomocysteinemia may serve as an upstream factor for increased Abeta production as seen in patients with Alzheimer disease.

Biogenesis of γ-secretase early in the secretory pathway

γ-Secretase is responsible for proteolytic maturation of signaling and cell surface proteins, including amyloid precursor protein (APP). Abnormal processing of APP by γ-secretase produces a fragment, Aβ42, that may be responsible for Alzheimer's disease (AD). The biogenesis and trafficking of this important enzyme in relation to aberrant Aβ processing is not well defined. Using a cell-free reaction to monitor the exit of cargo proteins from the endoplasmic reticulum (ER), we have isolated a transient intermediate of γ-secretase. Here, we provide direct evidence that the γ-secretase complex is formed in an inactive complex at or before the assembly of an ER transport vesicle dependent on the COPII sorting subunit, Sec24A. Maturation of the holoenzyme is achieved in a subsequent compartment. Two familial AD (FAD)–linked PS1 variants are inefficiently packaged into transport vesicles generated from the ER. Our results suggest that aberrant trafficking of PS1 may contribute to disease pathology.

SorLA/LR11 Regulates Processing of Amyloid Precursor Protein via Interaction with Adaptors GGA and PACS-1

SorLA has been recognized as a novel sorting receptor that regulates trafficking and processing of the amyloid precursor protein (APP) and that represents a significant risk factor for sporadic Alzheimer disease. Here, we investigated the cellular mechanisms that control intracellular trafficking of sorLA and their relevance for APP processing. We demonstrate that sorLA acts as a retention factor for APP in trans-Golgi compartments/trans-Golgi network, preventing release of the precursor into regular processing pathways. Proper localization and activity of sorLA are dependent on functional interaction with GGA and PACS-1, adaptor proteins involved in protein transport to and from the trans-Golgi network. Aberrant targeting of sorLA to the recycling compartment or the plasma membrane causes faulty APP trafficking and imbalance in non-amyloidogenic and amyloidogenic processing fates. Thus, our findings identified altered routing of sorLA as a major cellular mechanism contributing to abnormal APP processing and enhanced amyloid beta-peptide formation.

TNF and Alzheimer’s Disease

Alzheimer’s disease (AD) is associated with the accumulation in the brain of plaques of amyloid β protein (Aβ), which is produced by the processing of amyloid precursor protein (APP) by the enzymes β-secretase (BACE1) and γ-secretase. He et al . pursued their earlier finding that overexpression of tumor necrosis factor (TNF) receptor 1 (TNFR1) promoted Aβ-induced neuronal death by examining the role of TNFR1 in Aβ production. They crossed transgenic APP23 mice, which express a mutant APP and develop Aβ plaques, with TNFR1 –/– mice (APP23/ TNFR1 –/– ). Immunhistochemical analyses of the brains of 14-month-old animals showed a decreased number and size of Aβ plaques in APP23/ TNFR1 –/– mice compared with those in APP23 mice. Real-time reverse transcription polymerase chain reaction assays, together with Western blotting, showed that the abundance of BACE1 mRNA and protein were lower in APP23/ TNFR1 –/– mice compared with APP23 mice, which suggests that TNFR1 signaling promotes the production of BACE1. Experiments in 293 cells transfected with a reporter plasmid showed that TNF-α treatment induced BACE1 promoter activity, which was blocked by a specific inhibitor of the transcription factor nuclear factor κB (NF-κB). Immunohistochemistry demonstrated the presence of fewer neurons in the brains of APP23 mice than in brains from APP23/ TNFR1 –/– mice. In learning and object recognition tests, APP23 mice performed poorly; APP23/ TNFR1 –/– mice performed better, with abilities similar to those of wild-type mice. Together, these data implicate TNFR1 in promoting abnormal APP processing, Aβ plaque formation, and learning defects and suggest that TNFR1 might be an effective therapeutic target in treating AD. P. He, Z. Zhong, K. Lindholm, L. Berning, W. Lee, C. Lemere, M. Staufenbiel, R. Li, Y. Shen, Deletion of tumor necrosis factor death receptor inhibits amyloid β generation and prevents learning and memory deficits in Alzheimer’s mice. J. Cell Biol. 178 , 829-841 (2007). [Abstract] [Full Text]

Abnormal APP processing in platelets of patients with Alzheimer’s disease: correlations with membrane fluidity and cognitive decline

Previous studies have implicated platelet amyloid precursor protein (APP) as a candidate biomarker for Alzheimer's disease (AD). Platelets contain more than 95% of the circulating APP and enclose the enzymatic machinery for the APP metabolism yielding both soluble APP and amyloid-beta peptides. The objective of this study is to compare the ratio of 130- to 110-kDa fragments of APP in platelets from patients with AD, mild cognitive impairment (MCI), and elderly controls. After subjects were grouped according to diagnosis, APP ratio in platelets was evaluated by means of Western blot analysis. The APP ratio was significantly lower in AD patients (1.01 +/- 0.21) as compared to controls (1.24 +/- 0.21, p = 0.001) and MCI patients (1.18 +/- 0.21, p = 0.027), but no significant differences were found between MCI and controls (p = 0.904). In addition, we found positive correlations between the APP ratio and 1,6-diphenyl-1,3,5-hexatriene anisotropy (r = 0.3, p = 0.01), as well as with certain parameters of cognitive decline, namely, the mini-mental state examination score (r = 0.33, p = 0.003), the total Cambridge cognitive test (CAMCOG) score (r = 0.37, p = 0.001), and the score on the memory subscale of the CAMCOG (r = 0.38, p = 0.001). The pattern of platelet APP fragments was altered in patients with AD but not in patients with MCI. The alteration of APP fragments was correlated with membrane fluidity and the cognitive decline.

Distinct destructive signal pathways of neuronal death in Alzheimer's disease

Abundant neuron loss is a major feature of Alzheimer's disease (AD). Hypotheses for this loss include abnormal amyloid precursor protein processing (i.e. excess Abeta production, protein aggregation or misfolding), oxidative stress, excitotoxicity and inflammation. Neuron loss is a major cause of dementia in AD; however, it seems that there is no definitive pathway that causes cell death in the AD brain. Here, we examine the hypotheses for neuron loss in AD and pose the argument that the means by which neurons degenerate is irrelevant for cognitive decline. The best treatment for cognitive decline is to prevent the toxicity that first sets the neuron on its path to destruction, which is the production of Abeta peptide.