Secretion Of Amyloid Precursor Protein Research Articles

Secreted Amyloid Precursor Protein Alpha (sAPPα) Regulates the Cellular Proteome and Secretome of Mouse Primary Astrocytes.

Secreted amyloid precursor protein alpha (sAPPα), processed from a parent mammalian brain protein, amyloid precursor protein, can modulate learning and memory. Recently it has been shown to modulate the transcriptome and proteome of human neurons, including proteins with neurological functions. Here, we analysed whether the acute administration of sAPPα facilitated changes in the proteome and secretome of mouse primary astrocytes in culture. Astrocytes contribute to the neuronal processes of neurogenesis, synaptogenesis and synaptic plasticity. Cortical mouse astrocytes in culture were exposed to 1 nM sAPPα, and changes in both the whole-cell proteome (2 h) and the secretome (6 h) were identified with Sequential Window Acquisition of All Theoretical Fragment Ion Spectra-Mass Spectrometry (SWATH-MS). Differentially regulated proteins were identified in both the cellular proteome and secretome that are involved with neurologically related functions of the normal physiology of the brain and central nervous system. Groups of proteins have a relationship to APP and have roles in the modulation of cell morphology, vesicle dynamics and the myelin sheath. Some are related to pathways containing proteins whose genes have been previously implicated in Alzheimer's disease (AD). The secretome is also enriched in proteins related to Insulin Growth Factor 2 (IGF2) signaling and the extracellular matrix (ECM). There is the promise that a more specific investigation of these proteins will help to understand the mechanisms of how sAPPα signaling affects memory formation.

Enhancing Neurological Competence of Nanoencapsulated Cordyceps/Turmeric Extracts in Human Neuroblastoma SH-SY5Y Cells.

Neurological diseases, including Alzheimer's, Parkinson's diseases, and brain cancers, are reportedly caused by genetic aberration and cellular malfunction. Herbs with bioactive compounds that have anti-oxidant effects such as cordyceps and turmeric, are of interest to clinical applications due to their minimal adverse effects. The aim of study is to develop the nanoencapsulated cordyceps and turmeric extracts and investigate their capability to enhance the biological activity and improve neuronal function. Human neuroblastoma SH-SY5Y cells were utilized as a neuronal model to investigate the properties of nanoencapsulated cordyceps or turmeric extracts, called CMP and TEP, respectively. SH-SY5Y cells were treated with either CMP or TEP and examined the biological consequences, including neuronal maturation and neuronal function. The results showed that both CMP and TEP improved cellular uptake efficiency within 6h by 2.3 and 2.8 times, respectively. Besides, they were able to inhibit cellular proliferation of SH-SY5Y cells up to 153- and 218-fold changes, and increase the expression of mature neuronal markers (TUJ1, PAX6, and NESTIN). Upon the treatment of CMP and TEP, the expression of dopaminergic-specific genes (LMX1B, FOXA2, EN1, and NURR1), and the secretion level of dopamine were significantly improved up to 3.3-fold and 3.0-fold, respectively, while the expression of Alzheimer genes (PSEN1, PSEN2, and APP), and the secretion of amyloid precursor protein were significantly reduced by 32-fold and 108-fold, respectively. Importantly, the autophagy activity was upregulated by CMP and TEP at 6.3- and 5.5-fold changes, respectively. This finding suggested that the nanoencapsulated cordyceps and turmeric extracts accelerated neuronal maturation and alleviated neuronal pathology in human neural cells. This paves the way for nanotechnology-driven drug delivery systems that could potentially be used as an alternative medicine in the future for neurological diseases.

Secreted Amyloid Precursor Protein Alpha, a Neuroprotective Protein in the Brain Has Widespread Effects on the Transcriptome and Proteome of Human Inducible Pluripotent Stem Cell-Derived Glutamatergic Neurons Related to Memory Mechanisms.

Secreted amyloid precursor protein alpha (sAPPα) processed from a parent human brain protein, APP, can modulate learning and memory. It has potential for development as a therapy preventing, delaying, or even reversing Alzheimer’s disease. In this study a comprehensive analysis to understand how it affects the transcriptome and proteome of the human neuron was undertaken. Human inducible pluripotent stem cell (iPSC)-derived glutamatergic neurons in culture were exposed to 1 nM sAPPα over a time course and changes in the transcriptome and proteome were identified with RNA sequencing and Sequential Window Acquisition of All THeoretical Fragment Ion Spectra-Mass Spectrometry (SWATH-MS), respectively. A large subset (∼30%) of differentially expressed transcripts and proteins were functionally involved with the molecular biology of learning and memory, consistent with reported links of sAPPα to memory enhancement, as well as neurogenic, neurotrophic, and neuroprotective phenotypes in previous studies. Differentially regulated proteins included those encoded in previously identified Alzheimer’s risk genes, APP processing related proteins, proteins involved in synaptogenesis, neurotransmitters, receptors, synaptic vesicle proteins, cytoskeletal proteins, proteins involved in protein and organelle trafficking, and proteins important for cell signalling, transcriptional splicing, and functions of the proteasome and lysosome. We have identified a complex set of genes affected by sAPPα, which may aid further investigation into the mechanism of how this neuroprotective protein affects memory formation and how it might be used as an Alzheimer’s disease therapy.

Regressive Autism Spectrum Disorder: High Levels of Total Secreted Amyloid Precursor Protein and Secreted Amyloid Precursor Protein-α in Plasma.

Autism spectrum disorder (ASD) is a complex neurodevelopmental disorder characterized by social communication difficulties, repetitive behaviors, and parochial interests. Individuals with regressive ASD (RA), a unique subtype, have poor outcomes. Moreover, there are currently no validated blood-based biomarkers for ASD, hindering early diagnosis and treatment. This study was the first to examine plasma levels of total secreted amyloid precursor protein (sAPPtotal), secreted amyloid precursor protein-α (sAPPα), and secreted amyloid precursor protein-β (sAPPβ) in children diagnosed with RA (n = 23) and compare them with the levels in age-matched children with non-regressive ASD (NRA) (n = 23) and typically developing (TD) controls (n = 23). We found that sAPPtotal and sAPPα levels were significantly higher in children with RA than in children with NRA or in TD controls. In contrast, no difference was observed in sAPPβ levels. In conclusion, increased plasma levels of sAPPtotal and sAPPα may be valuable biomarkers for the early identification of ASD regression. Prospective studies will be conducted using a larger sample to further investigate these differences.

Carbon Dots: From Lab Synthesis to Unique Applications

Carbon dots (CDs) are a group of relatively new carbon-based spherical nanoparticles (NPs) with diameters less than 10 nm. They are widely present in nature and can also be synthesized using various carbon-based substances as precursors by either top-down or bottom-up approaches. They are well characterized for tunable surface functionality, excellent photoluminescence (PL), high photostability and water dispersity, good biocompatibility, and nontoxicity. Also, they display different sizes and surface chemistry depending on the preparation methods, and precursors applied. Applications of CDs in drug delivery, bioimaging, sensing, optics, photocatalysis, and other nanotechnology fields are rapidly rising due to their aforementioned unique properties. In this presentation, I will focus on the preparation, characterization and several significant applications of the CDs developed in our group. Specifically, I will introduce one type of CDs in yellow powder (Y-CDs) together with their amphiphilicity, tunable PL, and nontoxicity. In addition, Y-CDs showed an excellent permeability of body tissues, even including the blood-brain barrier (BBB) via passive diffusion using a zebrafish model. Furthermore, Y-CDs exhibited a strong inhibitory effect on the secretion of amyloid precursor protein (APP) and beta-amyloid (Aβ), which is significant for the future treatment of Alzheimer’s disease (AD). Considering their excellent permeability, Y-CDs are also promising drug nanocarriers for the treatment of diverse viral diseases such as COVID-19. Another work I would like to share with the public is that CDs have proved to be promising Lego-like building blocks for the assembly of novel versatile drug nanocarriers to simultaneously fulfill multitasks.

Alix and Syntenin-1 direct amyloid precursor protein trafficking into extracellular vesicles

BackgroundEndosomal trafficking and amyloidogenic cleavage of amyloid precursor protein (APP) is believed to play a role in the neurodegeneration observed in Alzheimer’s disease (AD). Recent evidence has suggested that packaging and secretion of APP and its amyloidogenic cleaved products into small extracellular vesicles (EVs) may facilitate uptake of these neurotoxic factors during disease progression. However, the molecular mechanisms underlying trafficking of APP into EVs are poorly understood.ResultsIn this study, the mechanism and impact of APP trafficking into extracellular vesicles (EVs) were assessed by a series of inducible gene knockdowns. We demonstrate that vesicle-associated proteins Alix and Syntenin-1 are essential for proper subcellular localization and efficient EV secretion of APP via an endosomal sorting complexes required for transport (ESCRT)-independent pathway. The neurotoxic C-terminal fragment (CTFβ) of APP is similarly secreted in association with small vesicles. These mechanisms are conserved in terminally differentiated neuron-like cells. Furthermore, knockdown of Alix and Syntenin-1 alters the subcellular localization of APP, sequestering the precursor protein to endoplasmic reticulum and endolysosomal compartments, respectively. Finally, transfer of small EVs containing mutant APP confers an increase in reactive oxygen species production and neurotoxicity to human induced pluripotent stem cell-derived cortical neurons and naïve primary neurons, an effect that is ameliorated by Alix and Syntenin-1 depletion.ConclusionsAltogether these findings elucidate a novel mechanism for understanding the intracellular trafficking of APP and CTFβ into secreted extracellular vesicles, and the resultant potential impact on neurotoxicity in the context of Alzheimer’s disease amyloidopathy.

Tetraspanin 7 and its closest paralog tetraspanin 6: membrane organizers with key functions in brain development, viral infection, innate immunity, diabetes and cancer.

Tetraspanin (TSPAN) protein family forms a family of transmembrane proteins that act as organizers/scaffold for other proteins. TSPANs are primarily present on plasma membranes although they are also found in other biological membranes. They are organized in tetraspanin-enriched microdomains (TEMs), which allow spatiotemporal tuning of protein functions through the control of their membrane localization. TSPAN6 and TSPAN7 are close paralogs expressed in different tissues, TSPAN7 being highly expressed in the brain. Their functions only started to be unveiled in the late 2000's and are still poorly understood. Here, we introduce how TSPAN7 was first highlighted has a protein mutated in some forms of X-linked mental retardation, which waslater proposed to be caused by defects in neuronal morphogenesis and synaptic transmission. We then discuss the impacts TSPAN7 has on cell morphology of dendritic cells and osteoclasts, through rearrangement of actin cytoskeleton and how TSPAN7 was shown to be a target of autoantibody in patients suffering from type 1 diabetes. Finally, we are addressing the double edge sword that is TSPAN7 in cancer. In the second part of this review, we address the known roles of TSPAN6 and how this protein was shown to participate in synaptic transmission and in amyloid precursor protein secretion, which may contribute to Alzheimer's disease pathology. We conclude this review by discussing the anti-inflammatory effect of TSPAN6.

Icaritin, an inhibitor of beta-site amyloid cleaving enzyme-1, inhibits secretion of amyloid precursor protein in APP-PS1-HEK293 cells by impeding the amyloidogenic pathway.

BackgroundIcaritin (ICT) is a prenylflavonoid derivative from Epimedium brevicornum Maxim. ICT has been shown to have neuroprotective effects. We investigate how ICT affects secretion of amyloid precursor protein (APP).MethodsWe exposed APP-PS1-HEK293 cells to ICT to investigate its effect on beta-site amyloid cleaving enzyme (BACE)1. Cell viability was evaluated by MTT and lactate dehydrogenase (LDH) assays. The half-maximal inhibitory concentration (IC50) of ICT for BACE1 was measured using fluorescence resonance energy transfer. Effects of ICT on the mRNA expression of APP were assessed by quantitative polymerase chain reaction, and protein expression was measured by western blotting and immunofluorescence.ResultsIcaritin inhibited BACE1 activity and IC50 was 5.70 ± 1.09 μM. Compared with the control group, at ICT concentrations of 5 μM and 10 μM, the viability increased and LDH leakage decreased in APP-PS1-293 cells. Also, mRNA expression of A disintegrin and metalloproteinase domain-containing protein 10 (ADAM10) increased, while that of BACE1 and presenilin-1 (PS1) decreased, upon ICT treatment. Western blotting and immunofluorescence confirmed that protein expression of ADAM10, BACE1 and PS1 showed the same trend. Expression of the APP fragments sAPPβ and C-terminal fragment β decreased, while that of sAPPα increased, upon ICT treatment. Expression of amyloid β peptides in APP-PS1-HEK293 cells was lower in ICT-treated groups compared with that in the control group.ConclusionsIcaritin, as a BACE1 inhibitor, inhibits APP secretion in APP-PS1-HEK293 cells by impeding the amyloidogenic pathway.

CORRIGENDUM.

CorrigendumCORRIGENDUMPublished Online:05 Feb 2019https://doi.org/10.1152/ajpendo.zh1-8043-corr.2019Original articleMoreSectionsPDF (28 KB)Download PDF ToolsExport citationAdd to favoritesGet permissionsTrack citations ShareShare onFacebookTwitterLinkedInWeChat Kulas JA, Franklin WF, Smith NA, Manocha GD, Puig KL, Nagamoto-Combs K, Hendrix RD, Taglialatela G, Barger SW, Combs CK. Ablation of amyloid precursor protein increases insulin-degrading enzyme levels and activity in brain and peripheral tissues. Am J Physiol Endocrinol Metab 316: E106–E120, 2019. First published November 13, 2018; doi:10.1152/ajpendo.00279.2018.—In disclosures, the following statement was omitted in error: “S. W. Barger receives royalties from Merck/Sigma-Aldrich for the sales of secreted amyloid precursor protein.”This article has no references to display. Download PDF Previous Back to Top FiguresReferencesRelatedInformation Related ArticlesAblation of amyloid precursor protein increases insulin-degrading enzyme levels and activity in brain and peripheral tissues 05 Jan 2019American Journal of Physiology-Endocrinology and Metabolism More from this issue > Volume 316Issue 2February 2019Pages E345-E345 Copyright & PermissionsCopyright © 2019 the American Physiological Societyhttps://doi.org/10.1152/ajpendo.zh1-8043-corr.2019PubMed30721099History Published online 5 February 2019 Published in print 1 February 2019 Metrics

Coconut oil decreases expression of amyloid precursor protein (APP) and secretion of amyloid peptides through inhibition of ADP-ribosylation factor 1 (ARF1)

Alzheimer’s disease (AD), affecting 5.3 million people in the U.S., impairs portions of the brain controlling memories. In humans, mutations in the amyloid precursor protein (APP) gene has been implicated in increased plaque formation, which can block the communication between nerve cells, decrease dendritic formation and increase cell death, and promote neuroinflammation. As coconut oil has been suggested to alleviate the symptoms in AD patients, we examined the impact of coconut oil on APP expression and secretion of amyloid peptides in N2a cells expressing the human APP gene (N2a/APP695). We found that coconut oil treatment decreased APP expression in N2a cells and reduced the secretion of amyloid peptides Aβ40 and Aβ42. Moreover, coconut oil treatment promoted differentiation of N2a cells. Our data suggest that ADP-Ribosylation Factor 1 (ARF1) may contribute to the effects of coconut oil on APP expression and secretion of Aβ. A high ARF1 expression was also detected in the primary neuronal cells from the mice overexpressing the Swedish mutant APP. Immunostaining results revealed that APP is co-localized with ARF1 in the Golgi apparatus and this interaction is impaired after coconut oil treatment. Furthermore, knockdown of ARF-1 using siRNA decreased secretion of amyloid peptides, confirming the impact of ARF1 on the secretion of amyloid peptides. ConclusionThese results suggest that coconut oil decreases intracellular ARF1 expression, thereby resulting in an inhibition of APP and amyloid β secretion. This study reveals a novel mechanism for intracellular APP processing in neuronal cells.

Secreted amyloid precursor protein alpha activates neuronal insulin receptors and prevents diabetes-induced encephalopathy

Secreted amyloid precursor protein alpha (sAPPα) is a potent neurotrophin in the CNS but a dedicated receptor has not been found. However, protein interactions involving amyloid beta (Aβ), a peptide cleaved from the same parent peptide as sAPPα, indicate that insulin receptors (IRs) could be a target of amyloid peptides. In this study, in vitro analysis of cortical neuronal cultures revealed that exogenous sAPPα increased IR phosphorylation in the absence of insulin. Furthermore, in an APP overexpressing mouse model, sAPPα bound IRs in the cortex with significantly greater binding in hypoinsulinemic animals. To further examine the effects of sAPPα on the diabetic brain, we next rendered sAPPα overexpressing mice insulin depleted and found that sAPPα blocked aberrant tau phosphorylation (T231) in cortical tissue after 16 weeks diabetes. sAPPα overexpression also prevented hyperphosphorylation of AKT/GSK3 and activation of the unfolded protein response (UPR). In total, these data show sAPPα binds and activates neuronal IRs and that sAPPα has a protective effect on diabetic brain tissue.

Therapeutic Potential of Secreted Amyloid Precursor Protein APPsα.

Cleavage of the amyloid precursor protein (APP) by α-secretase generates an extracellularly released fragment termed secreted APP-alpha (APPsα). Not only is this process of interest due to the cleavage of APP within the amyloid-beta sequence, but APPsα itself has many physiological properties that suggest its great potential as a therapeutic target. For example, APPsα is neurotrophic, neuroprotective, neurogenic, a stimulator of protein synthesis and gene expression, and enhances long-term potentiation (LTP) and memory. While most early studies have been conducted in vitro, effectiveness in animal models is now being confirmed. These studies have revealed that either upregulating α-secretase activity, acutely administering APPsα or chronic delivery of APPsα via a gene therapy approach can effectively treat mouse models of Alzheimer’s disease (AD) and other disorders such as traumatic head injury. Together these findings suggest the need for intensifying research efforts to harness the therapeutic potential of this multifunctional protein.

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

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

Astrocytes regulate α-secretase-cleaved soluble amyloid precursor protein secretion in neuronal cells: Involvement of group IIA secretory phospholipase A2

Astrocytes are major supportive cells in brains with important functions including providing nutrients and regulating neuronal activities. In this study, we demonstrated that astrocytes regulate amyloid precursor protein (APP) processing in neuronal cells through secretion of group IIA secretory phospholipase A2 (sPLA2-IIA). When astrocytic cells (DITNC) were mildly stimulated with the pro-inflammatory cytokines, such as TNF α and IL-1β, sPLA2-IIA was secreted into the medium. When conditioned medium containing sPLA2-IIA was applied to human neuroblastoma (SH-SY5Y) cells, there was an increase in both cell membrane fluidity and secretion of α-secretase-cleaved soluble amyloid precursor protein (sAPPα). These changes were abrogated by KH064, a selective inhibitor of sPLA2-IIA. In addition, exposing SH-SY5Y cells to recombinant human sPLA2-IIA also increased membrane fluidity, accumulation of APP at the cell surface, and secretion of sAPPα, but without altering total expressions of APP, α-secretases and β-site APP cleaving enzyme (BACE1). Taken together, our results provide novel information regarding a functional role of sPLA2-IIA in astrocytes for regulating APP processing in neuronal cells.

Acute function of secreted amyloid precursor protein fragment APPsα in synaptic plasticity.

The key role of APP in the pathogenesis of Alzheimer disease is well established. However, postnatal lethality of double knockout mice has so far precluded the analysis of the physiological functions of APP and the APLPs in the brain. Previously, APP family proteins have been implicated in synaptic adhesion, and analysis of the neuromuscular junction of constitutive APP/APLP2 mutant mice showed deficits in synaptic morphology and neuromuscular transmission. Here, we generated animals with a conditional APP/APLP2 double knockout (cDKO) in excitatory forebrain neurons using NexCre mice. Electrophysiological recordings of adult NexCre cDKOs indicated a strong synaptic phenotype with pronounced deficits in the induction and maintenance of hippocampal LTP and impairments in paired pulse facilitation, indicating a possible presynaptic deficit. These deficits were also reflected in impairments in nesting behavior and hippocampus-dependent learning and memory tasks, including deficits in Morris water maze and radial maze performance. Moreover, while no gross alterations of brain morphology were detectable in NexCre cDKO mice, quantitative analysis of adult hippocampal CA1 neurons revealed prominent reductions in total neurite length, dendritic branching, reduced spine density and reduced spine head volume. Strikingly, the impairment of LTP could be selectively rescued by acute application of exogenous recombinant APPsα, but not APPsβ, indicating a crucial role for APPsα to support synaptic plasticity of mature hippocampal synapses on a rapid time scale. Collectively, our analysis reveals an essential role of APP family proteins in excitatory principal neurons for mediating normal dendritic architecture, spine density and morphology, synaptic plasticity and cognition.

Axonal amyloid precursor protein and its fragments undergo somatodendritic endocytosis and processing.

Deposition of potentially neurotoxic Aβ fragments derived from amyloid precursor protein (APP) at synapses may be a key contributor to Alzheimer's disease. However, the location(s) of proteolytic processing and subsequent secretion of APP fragments from highly compartmentalized, euploid neurons that express APP and processing enzymes at normal levels is not well understood. To probe the behavior of endogenous APP, particularly in human neurons, we developed a system using neurons differentiated from human embryonic stem cells, cultured in microfluidic devices, to enable direct biochemical measurements from axons. Using human or mouse neurons in these devices, we measured levels of Aβ, sAPPα, and sAPPβ secreted solely from axons. We found that a majority of the fragments secreted from axons were processed in the soma, and many were dependent on somatic endocytosis for axonal secretion. We also observed that APP and the β-site APP cleaving enzyme were, for the most part, not dependent on endocytosis for axonal entry. These data establish that axonal entry and secretion of APP and its proteolytic processing products traverse different pathways in the somatodendritic compartment before axonal entry.

Platelet-Derived Secreted Amyloid-Precursor Protein-β as a Marker for Diagnosing Alzheimer’s Disease

A marker of Alzheimer's disease (AD) with a high sensitivity and specificity would facilitate a diagnosis at early stages. Blood platelets may be of particular interest in search of biomarkers, because they express amyloid-precursor protein (APP), and display a dysfunctional processing in AD. The aim of the present study is to establish and validate an assay for secreted amyloid-precursor protein (sAPP)-α and -β in platelets of AD and mild cognitively impaired (MCI) subjects, compared to healthy young and old controls. Freshly isolated platelet extracts (25 µg) were incubated with or without recombinant BACE1 (beta-site APP-Cleaving Enzyme; β-secretase, 8U) at 37°C and low pH and the levels of sAPP-α and sAPP-b were measured by specific ELISAs. Our data show that sAPP-α levels were not different between AD, MCI and control subjects. However, sAPP-β levels in MCI and AD were significantly elevated relative to controls. When recombinant BACE1 was added, no changes were seen in sAPP-α levels, but the processed sAPP-β levels were again markedly increased. The sAPP-β processing was specific and selective after 2.5 hours at 37°C, and was possibly mediated by exogenous BACE1, because it was blocked by a BACE1 inhibitor and BACE1 enzyme levels were enhanced in AD patients. Our data reveal that quantitive analysis of platelet sAPP-β assay by ELISA may be a novel diagnostic biomarker for MCI and AD.

Effects of rivastigmine on secreted amyloid precursor protein and beta-amyloid secretion in neuroblastoma SK-N-SH cells

Alzheimer’s disease (AD) is a neurodegenerative disorder characterized clinically by progressive impairment of memory and cognition. The currently available pharmacological treatment of AD consists mainly of cholinesterase inhibitors. Rivastigmine is one of the cholinesterase inhibitors clinically used to treat this disease, and many clinical trials have indicated that it did alleviate some AD symptoms without causing apparent side effects. Since the amyloid precursor protein (APP) processing imbalance plays a crucial role in AD pathogenesis, the effects of rivastigmine on APP processing were investigated. In neuroblastoma SK-N-SH cells, rivastigmine significantly increased the secretion of sAPPα and decreased the release of Aβ40 and Aβ42 as compared with control group, but it has no effect on cellular full length APP expression. Rivastigmine significantly increased α-secretase activity and decreased β-secretase activity as compared with control group. The increased sAPPα can be partially blocked by muscarinic receptor inhibitor scopolamine but not by nicotinic receptor antagonist α-Bungarotoxin. The effect of rivastigmine on sAPPα can be partially reversed by PKC inhibitor GF109203X, ERK inhibitor PD98059 and JNK inhibitor SP600125. The data present here indicated that rivastigmine can regulate APP processing in vitro by increasing sAPPα secretion and decreasing Aβ release and this pharmacological property may underlie the clinical effect of the drug in the treatment of AD patients.

Effects of Rivastigmine on Secreted Amyloid Precursor Protein and Beta-Amyloid Secretion in Neuroblastoma SK–N–SH Cells

Effects of Rivastigmine on Secreted Amyloid Precursor Protein and Beta-Amyloid Secretion in Neuroblastoma SK–N–SH Cells

Munc13 genotype regulates secretory amyloid precursor protein processing via postsynaptic glutamate receptors

The amyloid precursor protein (APP) can be proteolytically degraded via non-amyloidogenic α-secretase and amyloidogenic β-secretase pathways. Previously, we have identified the presynaptic protein Munc13-1 as a diacylglycerol/phorbolester (DAG/PE) receptor that contributes to secretory, non-amyloidogenic APP processing after PE stimulation. Here, we used organotypic brain slice cultures from wild-type mice and from Munc13-1 knock-out (KO), Munc13-2 KO and Munc13-1/2 double KO (DKO) mice for pharmacological stimulation experiments. First, we demonstrate that neuronal populations and synaptic components important for secretory APP processing develop normally in organotypic brain slice cultures of all genotypes analyzed. Blockade of voltage-gated Na+ channels by tetrodotoxin reduced the PE-stimulated secretory APP processing, whereas depolarization by high extracellular K+ concentration evoked APP secretion. Additionally, the PE-stimulated APP secretion from Munc13-1 KO brain slices was significantly lower than that from wild-type brain slices. This effect was not observed in brain slices from Munc13-2 KO mice, which is consistent with the lower abundance and subpopulation-specific distribution of Munc13-2 in presynaptic elements. In Munc13-1/2 DKO brain slices, the deficiency of Munc13-1 dominated the effect of APP processing. The Munc13-1 KO effect on APP processing could be rescued by the stimulation of postsynaptic glutamatergic receptors. This indicates that lack of postsynaptic glutamate receptor stimulation in Munc13-1 KO brain slice cultures but not presynaptic mechanisms account for compromised APP processing. We conclude that organotypic brain slices cultures are a valuable tool for studying APP processing pathways in intact neuronal circuits and that neuronal activity is important for maintenance of the non-amyloidogenic APP processing.