Swedish Mutant Amyloid Precursor Protein Research Articles

MT5-MMP controls APP and β-CTF/C99 metabolism through proteolytic-dependent and -independent mechanisms relevant for Alzheimer's disease.

We previously discovered the implication of membrane-type 5-matrix metalloproteinase (MT5-MMP) in Alzheimer's disease (AD) pathogenesis. Here, we shed new light on pathogenic mechanisms by which MT5-MMP controls the processing of amyloid precursor protein (APP) and the fate of amyloid beta peptide (Aβ) as well as its precursor C99, and C83. We found in human embryonic kidney cells (HEK) carrying the APP Swedish familial mutation (HEKswe) that deleting the C-terminal non-catalytic domains of MT5-MMP hampered its ability to process APP and release the soluble 95kDa form (sAPP95). Catalytically inactive MT5-MMP variants increased the levels of Aβ and promoted APP/C99 sorting in the endolysosomal system, likely through interactions of the proteinase C-terminal portion with C99. Most interestingly, the deletion of the C-terminal domain of MT5-MMP caused a strong degradation of C99 by the proteasome and prevented Aβ accumulation. These discoveries reveal new control of MT5-MMP over APP by proteolytic and non-proteolytic mechanisms driven by the C-terminal domains of the proteinase. The targeting of these non-catalytic domains of MT5-MMP could, therefore, provide new insights into the therapeutic regulation of APP-related pathology in AD.

Hepcidin contributes to Swedish mutant APP-induced osteoclastogenesis and trabecular bone loss

Patients with Alzheimer’s disease (AD) often have lower bone mass than healthy individuals. However, the mechanisms underlying this change remain elusive. Previously, we found that Tg2576 mice, an AD animal model that ubiquitously expresses Swedish mutant amyloid precursor protein (APPswe), shows osteoporotic changes, reduced bone formation, and increased bone resorption. To understand how bone deficits develop in Tg2576 mice, we used a multiplex antibody array to screen for serum proteins that are altered in Tg2576 mice and identified hepcidin, a master regulator of iron homeostasis. We further investigated hepcidin’s function in bone homeostasis and found that hepcidin levels were increased not only in the serum but also in the liver, muscle, and osteoblast (OB) lineage cells in Tg2576 mice at both the mRNA and protein levels. We then generated mice selectively expressing hepcidin in hepatocytes or OB lineage cells, which showed trabecular bone loss and increased osteoclast (OC)-mediated bone resorption. Further cell studies suggested that hepcidin increased OC precursor proliferation and differentiation by downregulating ferroportin (FPN) expression and increasing intracellular iron levels. In OB lineage cells, APPswe enhanced hepcidin expression by inducing ER stress and increasing OC formation, in part through hepcidin. Together, these results suggest that increased hepcidin expression in hepatocytes and OB lineage cells in Tg2576 mice contributes to enhanced osteoclastogenesis and trabecular bone loss, identifying the hepcidin-FPN-iron axis as a potential therapeutic target to prevent AD-associated bone loss.

Increased β-site APP cleaving enzyme 1-mediated insulin receptor cleavage in type 2 diabetes mellitus with cognitive impairment.

Patients with type 2 diabetes mellitus (T2DM) are at a high risk of cognitive impairment, with insulin resistance playing a pivotal role. β-site amyloid precursor protein cleaving enzyme 1 (BACE1) is considered a predictor of Alzheimer's disease. However, the potential roles of BACE1 in insulin resistance and the risk of cognitive impairment in T2DM remain unclear. We measured plasma BACE1 levels, BACE1 cleavage activities for Swedish mutant amyloid precursor protein (APPsw) and insulin receptor β subunit (INSR-β), and soluble INSR (sINSR) levels in a clinical cohort study. T2DM patients with or without cognitive impairment exhibited elevated plasma BACE1 levels and BACE1 enzymatic activities for APPsw and INSR-β, and sINSR levels. Moreover, the glycemic status correlated with elevated BACE1 levels and BACE1-mediated INSR cleavage, which was associated with insulin resistance. The elevated BACE1 levels in T2DM may contribute to increasing the cognitive impairment risk through both amyloidogenesis and insulin resistance.

The behavioral effects of virally induced human tau expression in the locus coeruleus of TgF344‐AD rats

AbstractBackgroundAberrant tau in the locus coeruleus (LC) is one of the first detectable neuropathological changes in Alzheimer’s disease (AD). Ubiquitous expression of mutant tau exacerbates pathological and cognitive impairments in rodent models of AD, but the consequences of wild‐type (WT) human tau expression in the LC is unknown. LC dysfunction has been linked to neuropsychiatric disorders and cognitive impairments that are relevant to AD, including sleep disorders, depression, anxiety, and memory loss. TgF344 AD rats are a transgenic rat model of AD that express APPSwe and PS1DE9 mutations and develop age‐dependent amyloid and tau pathology, neuroinflammation, cell death, and memory impairments that closely resemble humans with AD. We took advantage of this clinically‐relevant AD rat model to investigate the pathological and behavioral consequences of early human WT tau expression in the LC of TgF344 AD rats.MethodAt 2 months of age, TgF344 AD rats and WT littermates received bilateral LC infusions of human tau or mCherry control virus driven by the LC‐specific PRSx8 promotor. When rats were 6‐7 months of age, behavioral assays were administered to assess arousal (sleep latency), anxiety (open field, elevated plus maze, and novelty suppressed feeding), depression (forced swim task), and memory (Morris water maze and fear conditioning). Tau pathology in the LC was also evaluated by immunohistochemistry.ResultPreliminary analyses suggest that the behavioral consequence of human tau expression in the LC may interact with genotype. Specifically, TgF344‐AD rats that received human tau virus infusions had reduced freezing during cued and contextual fear conditioning tests compared to TgF344 AD rats that received control virus. By contrast, freezing behavior in WT rats was not influenced by human tau infusions in the LC. Additional analyses to evaluate group differences in measures of arousal, anxiety, and depression tasks are still being analyzed. Rats that received human tau virus showed tau pathology in the LC that was reminiscent of that observed in early human AD.ConclusionTau in the LC may contribute to early behavioral symptoms of AD that occur before widespread neuropathology and neurodegeneration occurs.

Increases of iASPP-Keap1 interaction mediated by syringin enhance synaptic plasticity and rescue cognitive impairments via stabilizing Nrf2 in Alzheimer's models

Oxidative stress is an important pathogenic manifestation of Alzheimer's disease (AD) that contributes to synaptic dysfunction, which precedes Aβ accumulation and neurofibrillary tangle formation. However, the molecular machineries that govern the decline of antioxidative defence in AD remains to be elucidated, and effective candidate for AD treatment is limited. Here, we showed that the decreases in the inhibitor of apoptosis-stimulating protein of p53 (iASPP) was associated with the vulnerability to oxidative stress in the amyloid precursor protein (APP)/presenilin 1 (PS1) mouse brain. Treatment with an antioxidant, syringin, could ameliorate AD-related pathologic and behavioural impairments. Interestingly, syringin treatment resulted in an upregulation of iASPP and the increase in the interaction of iASPP with Kelchlike ECH-associating protein 1 (Keap1). Syringin reduced neuronal apoptosis independently of p53. We confirmed that syringin-induced enhancement of antioxidant defenses involved the stabilization of Nrf2 in overexpressing human Swedish mutant APP (APPswe) cells in vitro. Syringin-mediated Nrf2 nuclear translocation facilitated the activation of the Nrf2 downstream genes via iASPP/Nrf2 axis. Our results demonstrate that syringin-mediated increases of iASPP-Keap1 interaction restore cellular redox balance. Further study on the syringin-iASPP interactions may help in understanding the regulatory mechanism and designing novel potent modulators for AD treatment.

Combination therapy with cilostazol, aripiprazole, and donepezil protects neuronal cells from β-amyloid neurotoxicity through synergistically enhanced SIRT1 expression

Alzheimer’s disease (AD) is a multi-faceted neurodegenerative disease. Thus, current therapeutic strategies require multitarget-drug combinations to treat or prevent the disease. At the present time, single drugs have proven to be inadequate in terms of addressing the multifactorial pathology of AD, and multitarget-directed drug design has not been successful. Based on these points of views, it is judged that combinatorial drug therapies that target several pathogenic factors may offer more attractive therapeutic options. Thus, we explored that the combination therapy with lower doses of cilostazol and aripiprazole with add-on donepezil (CAD) might have potential in the pathogenesis of AD. In the present study, we found the superior efficacies of donepezil add-on with combinatorial mixture of cilostazol plus aripiprazole in modulation of expression of AD-relevant genes: Aβ accumulation, GSK-3β, P300, acetylated tau, phosphorylated-tau levels, and activation of α-secretase/ADAM 10 through SIRT1 activation in the N2a Swe cells expressing human APP Swedish mutation (N2a Swe cells). We also assessed that CAD synergistically raised acetylcholine release and choline acetyltransferase (CHAT) expression that were declined by increased β-amyloid level in the activated N2a Swe cells. Consequently, CAD treatment synergistically increased neurite elongation and improved cell viability through activations of PI3K, BDNF, β-catenin and a7-nicotinic cholinergic receptors in neuronal cells in the presence of Aβ1-42. This work endorses the possibility for efficient treatment of AD by supporting the synergistic therapeutic potential of donepezil add-on therapy in combination with lower doses of cilostazol and aripiprazole.

A Novel Dual Fluorochrome Near-Infrared Imaging Probe for Potential Alzheimer's Enzyme Biomarkers-BACE1 and Cathepsin D.

A molecular imaging probe to fluorescently image the β-site of the amyloid precursor protein (APP) cleaving enzyme 1 (BACE1) and cathepsin D (CatD) enzymes associated with Alzheimer’s disease (AD) was designed and synthesized. This imaging probe was built upon iron oxide nanoparticles (cross-linked dextran iron oxide nanoparticles, or CLIO). Peptide substrates containing a terminal near-infrared fluorochrome (fluorophore emitting at 775 nm for CatD or fluorophore emitting at 669 nm for BACE1) were conjugated to the CLIO nanoparticles. The CatD substrate contained a phenylalanine-phenylalanine cleavage site more specific to CatD than BACE1. The BACE1 substrate contained the sequence surrounding the leucine-asparagine cleavage site of the BACE1 found in the Swedish mutation of APP, which is more specific to BACE1 than CatD. These fluorescently-labeled peptide substrates were then conjugated to the nanoparticle. The nanoparticle probes were purified by gel filtration, and their fluorescence intensities were determined using a fluorescence plate reader. The CatD peptide substrate demonstrated a 15.5-fold increase in fluorescence when incubated with purified CatD enzyme, and the BACE1 substrate exhibited a 31.5-fold increase in fluorescence when incubated with purified BACE1 enzyme. Probe specificity was also demonstrated in the human H4 neuroglioma cells and the H4 cells stably transfected with BACE1 in which the probe monitored enzymatic cleavage. In the H4 and H4-BACE1 cells, BACE1 and active CatD activity increased, an occurrence that was reflected in enzyme expression levels as determined by immunoblotting. These results demonstrate the applicability of this probe for detecting potential Alzheimer’s enzyme biomarkers.

Orexin-A aggravates cytotoxicity and mitochondrial impairment in SH-SY5Y cells transfected with APPswe via p38 MAPK pathway.

Alzheimer's disease (AD) is one of the common neurodegenerative diseases and is characterized by the accumulation of amyloid-β (Aβ). Orexin-A is a neuropeptide produced in the hypothalamus and thought to be involved in the pathogenesis of AD. However, its underlying mechanism and signaling pathway remains unclear. The aim of this work was to investigate the effect of Orexin-A on AD, and to explore its potential mechanism and signaling pathway. SH-SY5Y cells that were stably transfected with the Swedish mutant amyloid precursor protein (APPswe), a cell model of AD with excessive Aβ production, were used in this study. Cells were treated with Orexin-A, and with or without SB203580, an inhibitor of the p38 mitogen-activated protein kinase (MAPK) pathway, one of the key MAPK pathways associated with cell death. Following treatment, cells were collected and analyzed by western blotting, ELISA, electron microscopy, real-time PCR, fluorescence microscopy, and other biochemical assays. Orexin-A increased the level of Aβ1-40 and Aβ1-42 in the cell medium, and activated the p38 MAPK pathway. As evidenced by the CCK-8 and ELISA BrdU assays, Orexin-A decreased cell viability and cell proliferation. Electron microscopic analysis used to observe the morphology of mitochondria, showed that Orexin-A increased the percentage of abnormal mitochondria. Further, decreased activity of cytochrome c oxidase (CCO), level of ATP, and mitochondrial DNA (mtDNA) copy number following Orexin-A treatment showed that Orexin-A exacerbated mitochondrial dysfunction. The level of intracellular reactive oxygen species (ROS), which is mainly generated in mitochondria and reflects mitochondrial dysfunction, was also increased by Orexin-A. SB203580 blocked the cytotoxicity and mitochondrial impairment aggravated by Orexin-A. These findings demonstrate that Orexin-A aggravates cytotoxicity and mitochondrial impairment in SH-SY5Y cells transfected with APPswe through the p38 MAPK pathway, and suggest that Orexin-A participates in the pathogenesis of AD, which may provide a new treatment target in the future.

Activation of PKA/SIRT1 signaling pathway by photobiomodulation therapy reduces Aβ levels in Alzheimer's disease models.

A hallmark of Alzheimer's disease (AD) is the accumulation of amyloid‐β (Aβ), which correlates significantly with progressive cognitive deficits. Although photobiomodulation therapy (PBMT), as a novel noninvasive physiotherapy strategy, has been proposed to improve neuronal survival, decrease neuron loss, ameliorate dendritic atrophy, and provide overall AD improvement, it remains unknown whether and how this neuroprotective process affects Aβ levels. Here, we report that PBMT reduced Aβ production and plaque formation by shifting amyloid precursor protein (APP) processing toward the nonamyloidogenic pathway, thereby improving memory and cognitive ability in a mouse model of AD. More importantly, a pivotal protein, SIRT1, was involved in this process by specifically up‐regulating ADAM10 and down‐regulating BACE1, which is dependent on the cAMP/PKA pathway in APP/PS1 primary neurons and SH‐SY5Y cells stably expressing human APP Swedish mutation (APPswe). We further found that the activity of the mitochondrial photoacceptor cytochrome c oxidase (CcO) was responsible for PBMT‐induced activation of PKA and SIRT1. Together, our research suggests that PBMT as a viable therapeutic strategy has great potential value in improving cognitive ability and combatting AD.

Depletion of Caveolin-1 in Type 2 Diabetes Model Induces Alzheimer's Disease Pathology Precursors.

Type 2 diabetes mellitus (T2DM) is a risk factor for the development of late-onset Alzheimer's disease (AD). However, the mechanism underlying the development of late-onset AD is largely unknown. Here we show that levels of the endothelial-enriched protein caveolin-1 (Cav-1) are reduced in the brains of T2DM patients compared with healthy aging, and inversely correlated with levels of β-amyloid (Aβ). Depletion of Cav-1 is recapitulated in the brains of db/db (Leprdb ) diabetic mice and corresponds with recognition memory deficits as well as the upregulation of amyloid precursor protein (APP), BACE-1, a trending increase in β-amyloid Aβ42/40 ratio and hyperphosphorylated tau (p-tau) species. Importantly, we show that restoration of Cav-1 levels in the brains of male db/db mice using adenovirus overexpressing Cav-1 (AAV-Cav-1) rescues learning and memory deficits and reduces pathology (i.e., APP, BACE-1 and p-tau levels). Knocking down Cav-1 using shRNA in HEK cells expressing the familial AD-linked APPswe mutant variant upregulates APP, APP carboxyl terminal fragments, and Aβ levels. In turn, rescue of Cav-1 levels restores APP metabolism. Together, these results suggest that Cav-1 regulates APP metabolism, and that depletion of Cav-1 in T2DM promotes the amyloidogenic processing of APP and hyperphosphorylation of tau. This may suggest that depletion of Cav-1 in T2DM underlies, at least in part, the development of AD and imply that restoration of Cav-1 may be a therapeutic target for diabetic-associated sporadic AD.SIGNIFICANCE STATEMENT More than 95% of the Alzheimer's patients have the sporadic late-onset form (LOAD). The cause for late-onset Alzheimer's disease is unknown. Patients with Type 2 diabetes mellitus have considerably higher incidence of cognitive decline and AD compared with the general population, suggesting a common mechanism. Here we show that the expression of caveolin-1 (Cav-1) is reduced in the brain in Type 2 diabetes mellitus. In turn, reduced Cav-1 levels induce AD-associated neuropathology and learning and memory deficits. Restoration of Cav-1 levels rescues these deficits. This study unravels signals underlying LOAD and suggests that restoration of Cav-1 may be an effective therapeutic target.

PSEN1ΔE9, APPswe, and APOE4 Confer Disparate Phenotypes in Human iPSC-Derived Microglia

SummaryHere we elucidate the effect of Alzheimer disease (AD)-predisposing genetic backgrounds, APOE4, PSEN1ΔE9, and APPswe, on functionality of human microglia-like cells (iMGLs). We present a physiologically relevant high-yield protocol for producing iMGLs from induced pluripotent stem cells. Differentiation is directed with small molecules through primitive erythromyeloid progenitors to re-create microglial ontogeny from yolk sac. The iMGLs express microglial signature genes and respond to ADP with intracellular Ca2+ release distinguishing them from macrophages. Using 16 iPSC lines from healthy donors, AD patients and isogenic controls, we reveal that the APOE4 genotype has a profound impact on several aspects of microglial functionality, whereas PSEN1ΔE9 and APPswe mutations trigger minor alterations. The APOE4 genotype impairs phagocytosis, migration, and metabolic activity of iMGLs but exacerbates their cytokine secretion. This indicates that APOE4 iMGLs are fundamentally unable to mount normal microglial functionality in AD.

Anagliptin protects neuronal cells against endogenous amyloid β (Aβ)-induced cytotoxicity and apoptosis

Excessive generation and accumulation of amyloid-β (Aβ) fragments by familial mutations of amyloid precursor protein (APP) and presenilin 1 (PS1) play a key role in causing oxidative stress, mitochondrial abnormalities and neuronal apoptosis in Alzheimer’s disease (AD). Anagliptin, a novel DPP-4 inhibitor, is a clinical drug for the management of type II diabetes approved for use in 2012. Little on the pharmacological function of anagliptin against Aβ-induced cytotoxicity in neuronal cells is known. Here, we examined the protective capacities of anagliptin against cytotoxicity in N2a neuronal cells overexpressing APP Swedish mutant and PS1 exon 9 deletion mutant (N2a/Swe.D9). Our results demonstrate that anagliptin reduced the production of ROS and the expression of NADPH oxidase 4 (NOX-4) in N2a/Swe.D9 cells. We also reported that anagliptin activates the antioxidant system by increasing the level of reduced glutathione (GSH) and glutathione peroxidase (GPx) activity. Notably, anagliptin is able to improve mitochondrial function by elevating mitochondrial membrane potential (MMP) and adenosine triphosphate (ATP) production. Additionally, our results demonstrate that anagliptin decreased the vulnerability of cells to hydrogen peroxide (H2O2)-induced secondary insult by increasing cell viability and reducing the secretion of lactate dehydrogenase (LDH) and high mobility group box 1 protein (HMGB-1). Importantly, we found that treatment with anagliptin suppressed the mitochondrial-dependent apoptosis pathway by preventing the translocation of cytochrome C, reducing cleavage of caspase-3, and the inhibiting expression of Bax. These results implicate that anagliptin may have potential as a therapeutic agent for AD treatment.

PHluorin-BACE1-mCherry Acts as a Reporter for the Intracellular Distribution of Active BACE1 In Vitro and In Vivo.

β-site APP-cleaving enzyme 1 (BACE1) initiates amyloid precursor protein (APP) cleavage and β-amyloid (Aβ) production, a critical step in the pathogenesis of Alzheimer’s disease (AD). It is thus of considerable interest to investigate how BACE1 activity is regulated. BACE1 has its maximal activity at acidic pH and GFP variant—pHluorin—displays pH dependence. In light of these observations, we generated three tandem fluorescence-tagged BACE1 fusion proteins, named pHluorin-BACE1-mCherry, BACE1-mCherry-pHluorin and BACE1-mCherry-EGFP. Comparing the fluorescence characteristics of these proteins in response to intracellular pH changes induced by chloroquine or bafilomycin A1, we found that pHluorin-BACE1-mCherry is a better pH sensor for BACE1 because its fluorescence intensity responds to pH changes more dramatically and more quickly. Additionally, we found that (pro)renin receptor (PRR), a subunit of the v-ATPase complex, which is critical for maintaining vesicular pH, regulates pHluorin’s fluorescence and BACE1 activity in pHluorin-BACE1-mCherry expressing cells. Finally, we found that the expression of Swedish mutant APP (APPswe) suppresses pHluorin fluorescence in pHluorin-BACE1-mCherry expressing cells in culture and in vivo, implicating APPswe not only as a substrate but also as an activator of BACE1. Taken together, these results suggest that the pHluorin-BACE1-mCherry fusion protein may serve as a useful tool for visualizing active/inactive BACE1 in culture and in vivo.

Electro-acupuncture therapy to improve spatial learning and memory in APPswe/PS1dE9 transgenic mice through the inhibition of the TLR4/MyD88 signaling pathway

Abstract Objective To determine whether electro-acupuncture (EA) therapy could improve the cognitive functions of amyloid precursor protein Swedish mutation (APPswe)/presenilin 1 deleted in exon 9 (PS1dE9) mice and examine whether EA treatment could attenuate neuroinflammation by targeting the toll-like receptor 4 (TLR4)/myeloid differentiation primary response factor 88 (MyD88) signaling pathway. Methods Twenty-seven double transgenic APPswe/PS1dE9 mice were randomly allocated into three groups: an Alzheimer's disease model group (AD group), a medication group (M group) and an EA treatment group (EA group). Each group contained nine mice, and nine wild-type mice were used in a normal group (N group). The animals in the M group were treated with oral administrations of 0.92 mg/kg donepezil hydrochloride for 15 days. For animals in the EA group, EA treatments were used on the Yintang (GV 29) and Baihui (GV 20) acupoints for 20 minutes, and the Shuigou (GV 26) acupoint was pricked without needle retention following EA treatments. Following treatments, the spatial learning and memory of the mice were measured using the Morris water maze test. The expression levels of TLR4, MyD88, nuclear factor kappa B (NF-κB) and inducible nitric oxide synthase (iNOS) were analyzed by immunohistochemical staining and western blot. Results The escape latencies of the M and EA groups were significantly lower than those of the AD group (vs M, P = .002; vs EA, P Conclusion EA treatment enhanced the memory and learning abilities of APPswe/PS1dE9 mice by regulating the TLR4/MyD88 inflammatory signaling pathway.

Increased Insoluble Amyloid-β Induces Negligible Cognitive Deficits in Old AppNL/NL Knock-In Mice.

Commonly used Alzheimer’s disease mouse models are based on the ectopic overexpression of the human amyloid precursor protein (APP) gene, together with a mutant presenilin gene. Surprisingly, humanized APP knock-in mouse models carrying a single APP Swedish mutation (AppNL), failed to develop amyloid plaque aggregation or cognitive deficits. Here we characterized the effect of this mutation in more advanced ages. We show that 24-month-old AppNL/NL mice, despite presenting an age dependent increase in insoluble amyloid-β oligomers in the prefrontal cortex, they do not develop amyloid plaque deposition, reactive gliosis, or cognitive deficits.

APP promotes osteoblast survival and bone formation by regulating mitochondrial function and preventing oxidative stress

Amyloid precursor protein (APP) is ubiquitously expressed in various types of cells including bone cells. Mutations in App gene result in early-onset Alzheimer’s disease (AD). However, little is known about its physiological function in bone homeostasis. Here, we provide evidence for APP’s role in promoting bone formation. Mice that knocked out App gene (APP−/−) exhibit osteoporotic-like deficit, including reduced trabecular and cortical bone mass. Such a deficit is likely due in large to a decrease in osteoblast (OB)-mediated bone formation, as little change in bone resorption was detected in the mutant mice. Further mechanical studies of APP−/− OBs showed an impairment in mitochondrial function, accompanied with increased reactive oxygen species (ROS) and apoptosis. Intriguingly, these deficits, resemble to those in Tg2576 animal model of AD that expresses Swedish mutant APP (APPswe), were diminished by treatment with an anti-oxidant NAC (n-acetyl-l-cysteine), uncovering ROS as a critical underlying mechanism. Taken together, these results identify an unrecognized physiological function of APP in promoting OB survival and bone formation, implicate APPswe acting as a dominant negative factor, and reveal a potential clinical value of NAC in treatment of AD-associated osteoporotic deficits.

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.

The Isoquinoline Alkaloid Dauricine Targets Multiple Molecular Pathways to Ameliorate Alzheimer-Like Pathological Changes In Vitro.

Alzheimer's disease (AD), the most common neurodegenerative disease, has no effective treatment. Dauricine (DAU), a benzyl tetrahydroisoquinoline alkaloid isolated from the root of Menispermum dauricum DC, reportedly has neuroprotective effects in cerebral ischemia. Here, we investigated the effects of DAU on N2a cells stably transfected with Swedish mutant amyloid precursor protein (N2a/APP), an AD-like cell model. ELISA and Western blot analysis revealed that DAU inhibited APP processing and reduced Aβ accumulation. In addition, DAU ameliorated tau hyperphosphorylation via PP2A, p35/25, and CDK5 pathways in N2a/APP cells. The amelioration of tau hyperphosphorylation by DAU was also validated in HEK293/Tau cells, another cell line with tau hyperphosphorylation. Proteomic analysis revealed 85 differentially expressed proteins in the lysates between the wild-type N2a cells (N2a/WT) and the N2a/APP cells in the presence or absence of DAU; these were classified into 6 main categories according to their functions: endoplasmic reticulum (ER) stress-associated proteins, oxidative stress-associated proteins, cytoskeleton proteins, molecular chaperones, mitochondrial respiration and metabolism-related proteins, and signaling proteins. Taken together, we demonstrated that DAU treatment reduces AD-like pathology, thereby suggesting that DAU has potential therapeutic utility in AD.

Full-length amyloid precursor protein regulates lipoprotein metabolism and amyloid-β clearance in human astrocytes

Mounting evidence suggests that alterations in cholesterol homeostasis are involved in Alzheimer's disease (AD) pathogenesis. Amyloid precursor protein (APP) or multiple fragments generated by proteolytic processing of APP have previously been implicated in the regulation of cholesterol metabolism. However, the physiological function of APP in regulating lipoprotein homeostasis in astrocytes, which are responsible for de novo cholesterol biosynthesis and regulation in the brain, remains unclear. To address this, here we used CRISPR/Cas9 genome editing to generate isogenic APP-knockout (KO) human induced pluripotent stem cells (hiPSCs) and differentiated them into human astrocytes. We found that APP-KO astrocytes have reduced cholesterol and elevated levels of sterol regulatory element-binding protein (SREBP) target gene transcripts and proteins, which were both downstream consequences of reduced lipoprotein endocytosis. To elucidate which APP fragments regulate cholesterol homeostasis and to examine whether familial AD mutations in APP affect lipoprotein metabolism, we analyzed an isogenic allelic series harboring the APP Swedish and APP V717F variants. Only astrocytes homozygous for the APP Swedish (APPSwe/Swe) mutation, which had reduced full-length APP (FL APP) due to increased β-secretase cleavage, recapitulated the APP-KO phenotypes. Astrocytic internalization of β-amyloid (Aβ), another ligand for low-density lipoprotein (LDL) receptors, was also impaired in APP-KO and APPSwe/Swe astrocytes. Finally, impairing cleavage of FL APP through β-secretase inhibition in APPSwe/Swe astrocytes reversed the LDL and Aβ endocytosis defects. In conclusion, FL APP is involved in the endocytosis of LDL receptor ligands and is required for proper cholesterol homeostasis and Aβ clearance in human astrocytes.

Findings from the Swedish Study on Familial Alzheimer's Disease Including the APP Swedish Double Mutation.

This is a brief summary of the findings from the Swedish study on familial Alzheimer's disease (FAD). Similar to other FAD studies, it includes prospective assessments of cognitive function, tissue sampling, and technical analyses such as MRI and PET. This 24-year-old study involves 69 individuals with a 50% risk of inheriting a disease-causing mutation in presenilin 1 (PSEN1 H163Y or I143T), or amyloid precursor protein (the Swedish APP or the arctic APP mutation) who have made a total of 169 visits. Our results show the extraordinary power in this study design to unravel the earliest changes in preclinical AD. The Swedish FAD study will continue and future research will focus on disentangling the order of pathological change using longitudinal data as well as modeling the changes in patient derived cell systems.