Non-amyloidogenic Processing Of Amyloid Precursor Protein Research Articles

Inhibition of ACAT as a Therapeutic Target for Alzheimer's Disease Is Independent of ApoE4 Lipidation.

APOE4, encoding apolipoprotein E4 (apoE4), is the greatest genetic risk factor for Alzheimer's disease (AD), compared to the common APOE3. While the mechanism(s) underlying APOE4-induced AD risk remains unclear, increasing the lipidation of apoE4 is an important therapeutic target as apoE4-lipoproteins are poorly lipidated compared to apoE3-lipoproteins. ACAT (acyl-CoA: cholesterol-acyltransferase) catalyzes the formation of intracellular cholesteryl-ester droplets, reducing the intracellular free cholesterol (FC) pool. Thus, inhibiting ACAT increases the FC pool and facilitates lipid secretion to extracellular apoE-containing lipoproteins. Previous studies using commercial ACAT inhibitors, including avasimibe (AVAS), as well as ACAT-knock out (KO) mice, exhibit reduced AD-like pathology and amyloid precursor protein (APP) processing in familial AD (FAD)-transgenic (Tg) mice. However, the effects of AVAS with human apoE4 remain unknown. In vitro, AVASinduced apoE efflux at concentrations of AVASmeasured in the brains of treated mice. AVAS treatment of male E4FAD-Tg mice (5xFAD+/-APOE4+/+) at6-8months had no effect on plasma cholesterol levels or distribution, the original mechanism forAVAS treatment of CVD. In the CNS, AVAS reduced intracellular lipid droplets, indirectly demonstrating target engagement. Surrogate efficacy was demonstrated by an increase in Morris water maze measures of memory and postsynaptic protein levels. Amyloid-beta peptide (Aβ) solubility/deposition and neuroinflammation were reduced, critical components of APOE4-modulated pathology. However, there was no increase in apoE4 levels or apoE4 lipidation, while amyloidogenic and non-amyloidogenic processing of APP were significantly reduced. This suggests that the AVAS-induced reduction in Aβ via reduced APP processing was sufficient to reduce AD pathology, as apoE4-lipoproteins remained poorly lipidated.

Inhibition of SIRT2 promotes APP acetylation and ameliorates cognitive impairment in APP/PS1 transgenic mice.

Aging is a primary risk factor for neurodegenerative diseases, such as Alzheimer's disease (AD). SIRT2, an NAD+(nicotinamide adenine dinucleotide)-dependent deacetylase, accumulates in the aging brain. Here, we report that, in the amyloid precursor protein (APP)/PS1 transgenic mouse model of AD, genetic deletion of SIRT2 or pharmacological inhibition of SIRT2 ameliorates cognitive impairment. We find that suppression of SIRT2 enhances acetylation of APP, which promotes non-amyloidogenic processing of APP at the cell surface, leading to increased soluble APP-α (sAPPα). We discover that lysines 132 and 134 of the major pathogenic protein β-amyloid (Aβ) precursor are acetylated and that these residues are deacetylated by SIRT2. Strikingly, exogenous expression of wild-type or an acetylation-mimic APP mutant protects cultured primary neurons from Aβ42 challenge. Our study identifies SIRT2-mediated deacetylation of APP on K132 and K134 as a regulated post-translational modification (PTM) and suggests inhibition of SIRT2 as a potential therapeutic strategy for AD.

Probiotic Bifidobacterium breve decreases Aβ production via the upregulation of ADAM10 level and attenuates microglia activation in an APP knock‐in mouse model of Alzheimer’s disease

Probiotics supplementation reinstates microbiome diversity and improves brain function, including memory and learning abilities, although the molecular mechanisms have not been fully elucidated. In the current study, we investigated the effects of orally supplemented Bifidobacterium breve MCC1274 (B. breve MCC1274, synonym B. breve A1) on memory impairment and Alzheimer's disease (AD) pathogenesis in amyloid precursor protein knock-in (APP-KI) mice, (AppNL-G-F ).Three-month-old AppNL-G-F mice were orally supplemented with B. breve MCC1274 for four months. Novel object recognition test was used to evaluate memory-ameliorating effect of B. breve MCC1274. Amyloid plaques, Aβ levels, amyloid precursor protein (APP) and its processing enzymes, its metabolic products, and glial activity were assessed by immunohistochemistry, Aβ ELISA, Western blotting, and immunofluorescence staining. The mRNA expression levels of neuroinflammatory cytokines determined by quantitative RT-PCR analysis.Oral supplementation of B. breve MCC1274 rescued cognitive deficits in AppNL-G-F mice and down-regulated hippocampal Aβ40 and Aβ42 levels via the enhancement of non-amyloidogenic processing of APP, resulting from the enhancement of a-disintegrin and metalloproteinase 10 (ADAM10) protein level. Moreover, oral supplementation of B. breve MCC1274 activated the ERK/HIF-1α signaling pathway, which is responsible for the enhancement of ADAM10 level. In addition, oral supplementation ofB. breve MCC1274 attenuated microglial activation, which in turn led to down-regulation of the neuroinflammatory cytokines such as interleukin-6 (IL-6) and IL-1β mRNA expression levels in the brain. Moreover, B. breve MCC1274 supplementation increased postsynaptic protein levels in the hippocampus.Our study sheds light on the mechanism by which oral supplementation of B. breve MCC1274 reduced Aβ production and deposition through the enhancement of ERK-HIF-1α-ADAM10 signaling pathway, consequently rescuing cognitive deficits. Therefore, our findings suggest that oral supplementation of B. breve MCC1274 could be used as a potential therapy for preventing AD progression.

Levetiracetam Treatment Normalizes Levels of Presynaptic Endocytosis Machinery and Restores Nonamyloidogenic APP Processing in App Knock-in Mice.

Toxic amyloid-beta (Aβ) peptides, produced by sequential proteolytic cleavage of the amyloid precursor protein (APP), play a key role in the initial stage of Alzheimer’s disease (AD). Increasing evidence indicates that Aβ42 induces neuronal circuit hyperexcitability in the early stages of AD pathology. As a result, researchers have investigated treatments that modulate the excitatory/inhibitory imbalance as potential AD therapies. For example, levetiracetam, an atypical antiepileptic drug used to quell hyperexcitability, has garnered recent interest in the AD field, even though its exact mechanism(s) of action remains elusive. Here, we show that in APP knock-in mouse models of amyloid pathology, chronic levetiracetam administration decreases cortical Aβ42 levels and lowers the amyloid plaque burden. In addition, using multiplexed tandem mass tag-quantitative mass spectrometry-based proteomic analysis, we determined that chronic levetiracetam administration selectively normalizes levels of presynaptic endocytic proteins. Finally, we found that levetiracetam treatment selectively lowers beta carboxyl-terminal fragment levels, while the abundance of full-length APP remains unchanged. In summary, this work reports that chronic treatment with levetiracetam serves as a useful therapeutic in AD by normalizing levels of presynaptic endocytic proteins and altering APP cleavage preference, leading to a decrease in both Aβ42 levels and the amyloid plaque burden. These novel findings provide novel evidence for the previously documented therapeutic value of levetiracetam to mitigate AD pathology.

Acute Interleukin‐6 Injection Influences Key Enzymes Involved in Amyloid Precursor Protein Processing in C57BL/6J Mice

Background Amyloid-beta plaques are a hallmark feature of Alzheimer's disease and are the product of aggregated amyloid-beta peptides. These peptides originate from amyloid precursor protein (APP) which is cleaved by beta secretase (BACE1), a key enzyme in the amyloidogenic cascade. Cleavage of APP by alpha secretase (ADAM10) however, does not result in amyloid-beta peptides and is a key enzyme in the non-amyloidogenic cascade. It is widely accepted that exercise reduces amyloid-beta production and plaque build-up in the brain, although it is unclear how exercise may be driving this observation. Interleukin-6 (IL-6) is an exercise induced cytokine and recent work in this area has demonstrated a role for acute exercise induced IL-6 in tissues such as muscle, adipose, and liver, however the role of acute IL-6 in the brain remains ill defined. The purpose of this study is to investigate if acute IL-6 injection will modulate key enzymes involved in APP processing, namely BACE1 and ADAM10. It is hypothesized that acute IL-6 injection will result in a shift towards the non-amyloidogenic processing of APP. Methods Male 10-week-old C57BL/6J mice were intraperitoneally injected with a weight-adjusted dose of recombinant murine IL-6 (3ng/g body mass) or an equivalent volume of saline. Brain tissue was acquired 15 minutes post-injection and the pre-frontal cortex and hippocampus regions were dissected. Results Acute IL-6 injection resulted in lower BACE1 activity in both the cortex (p=0.011) and the hippocampus (p=0.027) compared to the saline injected group. This was accompanied by lower BACE1 (p=0.007) and sAPPb(p=0.019) protein content in the hippocampus of mice injected with IL-6. No differences for BACE1 or sAPPb were observed in the cortex. ADAM10 activity was higher with IL-6 injection compared to saline in the hippocampus (p=0.018) and the cortex (p=0.055). A change in ADAM10 protein content was also observed with the ratio of pro to mature ADAM10 being higher in the cortex (p=0.033) of the IL-6 injected mice as compared to the saline injected control. Similarly, both pro (p=0.017) and mature (p=0.035) ADAM10 protein content were lower in the hippocampus of the IL-6 injected mice as compared to the saline injected control. Conclusions These results demonstrate that acute IL-6 modulates important enzymes involved in APP processing; pushing the cascade towards the non-amyloidogenic arm. This work further highlights regional brain differences in response to acute IL-6 and extends our knowledge of the function of acute IL-6 as a signalling molecule in the brain. Future work should determine if exercise induced IL-6 is in fact the main mediator of non-amyloidogenic APP processing.

Autophagy-Dependent Increased ADAM10 Mature Protein Induced by TFEB Overexpression Is Mediated Through PPARα.

Nonamyloidogenic processing of amyloid precursor protein (APP) by augmenting ADAM10 is a promising therapeutic strategy for Alzheimer's disease (AD). Therefore identification of molecular pathways that regulate ADAM10 expression is crucial. Autophagy is strongly dysregulated in AD, and TFEB was recently shown to be a master regulator of autophagy-lysosome pathway (ALP). Here, we report that TFEB expression in HeLa cells increased ADAM10 mature form by 72% (p < 0.01, n = 4), while TFEB knockdown by CRISPR strategy reduced ADAM10 mature form by 36% (p < 0.05, n = 4). Autophagy inhibition by 3-methyladenine (3-MA), but not bafilomycin A1 (BAF1), reduced ADAM10 mature form by 49% (p < 0.05, n = 4) in the TFEB expressing HeLa cells. Autophagy activation by 3h of starvation increased ADAM10 to 91% (p < 0.001, n = 6) relative to 51% (p < 0.01, n = 6) in the nutrient-fed cells. Further, siRNAs targeted against PPARα in HeLa cells decreased ADAM10 levels by 28% (p < 0.05, n = 6) relative to the cells treated with scrambled siRNAs. Further, incubation of EGFP-TFEB expressing HeLa cells with PPARα antagonist, but not PPARβ or PPARγ antagonists, prevented TFEB-induced increase in ADAM10 levels. Importantly, flag-TFEB expression in the brain also increased ADAM10 by 60% (p < 0.05, n = 3) in the cortical and 34% (p < 0.001, n = 3) in the hippocampal homogenates. ADAM10 activity also increased by 57% (p < 0.01, n = 3) in the HeLa cells. Finally, TFEB-induced ADAM10 potentiation led to increased secretion of sAPPα by 154% (p < 0.001, n = 3) in the cortex and 62% (p < 0.001, n = 3) in the hippocampus. Thus, TFEB expression enhances nonamyloidogenic processing of APP. In conclusion, TFEB expression induces ADAM10 in an autophagy-dependent manner through PPARα.

The protective mutation A673T in amyloid precursor protein gene decreases Aβ peptides production for 14 forms of Familial Alzheimer's Disease in SH-SY5Y cells.

The deposition of Aβ plaques in the brain leads to the onset and development of Alzheimer's disease. The Amyloid precursor protein (APP) is cleaved by α-secretase (non-amyloidogenic processing of APP), however increased cleavage by β-secretase (BACE1) leads to the accumulation of Aβ peptides, which forms plaques. APP mutations mapping to exons 16 and 17 favor plaque accumulation and cause Familial Alzheimer Disease (FAD). However, a variant of the APP gene (A673T) originally found in an Icelandic population reduces BACE1 cleavage by 40%. A series of plasmids containing the APP gene, each with one of 29 different FAD mutations mapping to exon 16 and exon 17 was created. These plasmids were then replicated with the addition of the A673T mutation. Combined these formed the library of plasmids that was used in this study. The plasmids were transfected in neuroblastomas to assess the effect of this mutation on Aβ peptide production. The production of Aβ peptides was decreased for some FAD mutations due to the presence of the co-dominant A673T mutation. The reduction of Aβ peptide concentrations for the London mutation (V717I) even reached the same level as for A673T control in SH-SY5Y cells. These preliminary results suggest that the insertion of A673T in APP genes containing FAD mutations might confer a clinical benefit in preventing or delaying the onset of some FADs.

Centrally Active Multitarget Anti-Alzheimer Agents Derived from the Antioxidant Lead CR-6.

Oxidative stress is a major pathogenic factor in Alzheimer's disease, but it should not be tackled alone rather together with other key targets to derive effective treatments. The combination of the scaffold of the polar antioxidant lead 7-methoxy-2,2-dimethylchroman-6-ol (CR-6) with that of the lipophilic cholinesterase inhibitor 6-chlorotacrine results in compounds with favorable brain permeability and multiple activities in vitro (acetylcholinesterase, butyrylcholinesterase, β-site amyloid precursor protein (APP) cleaving enzyme-1 (BACE-1), and Aβ42 and tau aggregation inhibition). In in vivo studies on wild-type and APP/presenilin 1 (PS1) mice, two selected compounds were well tolerated and led to positive trends, albeit statistically nonsignificant in some cases, on memory performance, amyloid pathology (reduced amyloid burden and potentiated non-amyloidogenic APP processing), and oxidative stress (reduced cortical oxidized proteins and increased antioxidant enzymes superoxide dismutase 2 (SOD2), catalase, glutathione peroxidase 1 (GPX1), and heme oxygenase 1 (Hmox1) and transcription factor nuclear factor-erythroid 2-related factor 2 (Nrf2)). These compounds emerge as interesting brain-permeable multitarget compounds, with a potential as anti-Alzheimer agents beyond that of the original lead CR-6.

Phosphatidylinositol-4-phosphate 5-kinase type 1α attenuates Aβ production by promoting non-amyloidogenic processing of amyloid precursor protein.

Alzheimer's disease (AD) is characterized by a chronic decline in cognitive function and is pathologically typified by cerebral deposition of amyloid-β peptide (Aβ). The production of Aβ is mediated by sequential proteolysis of amyloid precursor protein (APP) by β- and γ-secretases, and has been implicated as the essential determinant of AD pathology. Previous studies have demonstrated that the level of phosphatidylinositol-4,5-bisphosphate [PI(4,5)P2] in the membrane may potentially modulate Aβ production. Given that PI(4,5)P2 is produced by type 1 phosphatidylinositol-4-phosphate 5-kinases (PIP5Ks), we sought to determine whether the level of PIP5K type Iα (PIP5K1A) can affect production of Aβ by modulating the lipid composition of the membrane. Using a HEK-derived cell line that constitutively expresses yellow fluorescent protein-tagged APP (APP-YFP), we demonstrated that overexpression of PIP5K1A results in significant enhancement of non-amyloidogenic APP processing and a concomitant suppression of the amyloidogenic pathway, leading to a marked decrease in secreted Aβ. Consistently, cells overexpressing PIP5K1A exhibited a significant redistribution of APP-YFP from endosomal compartments to the cell surface. Our findings suggest that PIP5K1A may play a critical role in governing Aβ production by modulating membrane distribution of APP, and as such, the pathway may be a valuable therapeutic target for AD.

Abstract TP486: Role of Beta-Site Amyloid Precursor Protein-Cleaving Enzyme 2 in the Metabolism of Amyloid Precursor Protein in Human Brain Microvascular Endothelial Cells

Under physiological conditions, β-site amyloid precursor protein (APP)-cleaving enzyme 2 (BACE2) cleaves APP within Aβ sequence thereby functioning like a α-secretase. However, BACE2 could also function as a conditional β-secretase during aging, contributing to Alzheimer’s disease pathogenesis. To date the physiological functions of BACE2 in endothelium are largely unknown. The present study is therefore designed to investigate the role of BACE2 in APP metabolism in human BMECs. Cultured human BMECs (passage 5-6 or passage 22) were treated with BACE2siRNA (30 nM, for 3 days), levels of soluble APPα (sAPPα, a neurotrophic product of non-amyloidogenic processing of APP) and Aβ40 in the supernatant were measured. In human BMECs (passage 5-6), genetic inactivation of BACE2 significantly decreased production of sAPPα (n=12, P&lt;0.05), but had no effect on production of Aβ40 (n=9, P&gt;0.05). BACE2siRNA treatment significantly suppressed APP protein expression (n=7, P&lt;0.05), but augmented protein levels of BACE1 (n=7, P&lt;0.05). Genetic inactivation of BACE2 did not change protein levels of mature ADAM10 (n=7, P&gt;0.05). Thus, reduced sAPPα secretion by BACE2siRNA treatment is likely caused not only by decreased α-secretase-like function of BACE2, but also by reduced APP expression. We further examined the effects of BACE2siRNA in senescent human BMECs. In cultured human BMECs (passage 22), protein expressions of senescent markers (p 21Cip1 and p 16INK4a ) were significantly increased (n=4, P&lt;0.05). Genetic inactivation of BACE2 in senescent human BMECs also significantly suppressed secretion of sAPPα (n=8, P&lt;0.05), but did not affect Aβ40 production (n=8, P&gt;0.05). BACE2-siRNA treatment significantly inhibited protein expressions of APP and mature ADAM10 (n=7, P&lt;0.05), but did not change BACE1 protein expression (n=7, P&gt;0.05). Thus in senescent human BMECs, reduced APP expression and impaired α-processing may play important roles in the decreased sAPPα production. Since our previous studies have demonstrated that endothelial production of sAPPα significantly contributes to the sAPPα content in the hippocampus, our current findings suggests that inhibition of BACE2 could impair protective function of sAPPα in the hippocampus.

α-Tocopherol Modulates Non-Amyloidogenic Pathway and Autophagy in an In Vitro Model of Alzheimer's Disease: A Transcriptional Study.

Alzheimer’s disease (AD) is the most common form of dementia worldwide. The hallmarks of AD are the extracellular amyloid plaques, which are formed by amyloid β (Aβ) aggregates derived from the processing of the amyloid precursor protein (APP), and the intraneuronal neurofibrillary tangles, which are formed by the hyperphosphorylated tau protein. The aim of this work was to study the effects of α-tocopherol in retinoic acid differentiated SH-SY5Y neuroblastoma cells exposed to Aβ1-42 evaluating the transcriptional profile by next-generation sequencing. We observed that α-tocopherol was able to reduce the cytotoxicity induced by Aβ treatment, as demonstrated by Thiazolyl Blue Tetrazolium Bromide (MTT) assay. Moreover, the transcriptomic analysis evidenced that α-tocopherol treatment upregulated genes involved in the non-amyloidogenic processing of APP, while it downregulated the amyloidogenic pathway. Moreover, α-tocopherol modulated the expression of the genes involved in autophagy and the cell cycle, which are both known to be altered in AD. The treatment with α-tocopherol was also able to reduce oxidative stress, restoring nuclear factor erythroid-derived 2-like 2 (Nrf2) and decreasing inducible nitric oxide synthase (iNOS) levels, as demonstrated by immunocytochemistry.

LRP1 Has a Predominant Role in Production over Clearance of A\u03b2 in a Mouse Model of Alzheimer\u2019s Disease

The low-density lipoprotein receptor-related protein-1 (LRP1) has a dual role in the metabolism of the amyloid precursor protein (APP). In cellular models, LRP1 enhances amyloid-β (Aβ) generation via APP internalization and thus its amyloidogenic processing. However, conditional knock-out studies in mice define LRP1 as an important mediator for the clearance of extracellular Aβ from brain via cellular degradation or transcytosis across the blood-brain barrier (BBB). In order to analyze the net effect of LRP1 on production and clearance of Aβ in vivo, we crossed mice with impaired LRP1 function with a mouse model of Alzheimer’s disease (AD). Analysis of Aβ metabolism showed that, despite reduced Aβ clearance due to LRP1 inactivation in vivo, less Aβ was found in cerebrospinal fluid (CSF) and brain interstitial fluid (ISF). Further analysis of APP metabolism revealed that impairment of LRP1 in vivo shifted APP processing from the Aβ-generating amyloidogenic cleavage by beta-secretase to the non-amyloidogenic processing by alpha-secretase as shown by a decrease in extracellular Aβ and an increase of soluble APP-α (sAPP-α). This shift in APP processing resulted in overall lower Aβ levels and a reduction in plaque burden. Here, we present for the first time clear in vivo evidence that global impairment of LRP1’s endocytosis function favors non-amyloidogenic processing of APP due to its reduced internalization and subsequently, reduced amyloidogenic processing. By inactivation of LRP1, the inhibitory effect on Aβ generation overrules the simultaneous impaired Aβ clearance, resulting in less extracellular Aβ and reduced plaque deposition in a mouse model of AD.

Proteolytic shedding of the prion protein via activation of metallopeptidase ADAM10 reduces cellular binding and toxicity of amyloid-β oligomers

The cellular prion protein (PrPC) is a key neuronal receptor for β-amyloid oligomers (AβO), mediating their neurotoxicity, which contributes to the neurodegeneration in Alzheimer's disease (AD). Similarly to the amyloid precursor protein (APP), PrPC is proteolytically cleaved from the cell surface by a disintegrin and metalloprotease, ADAM10. We hypothesized that ADAM10-modulated PrPC shedding would alter the cellular binding and cytotoxicity of AβO. Here, we found that in human neuroblastoma cells, activation of ADAM10 with the muscarinic agonist carbachol promotes PrPC shedding and reduces the binding of AβO to the cell surface, which could be blocked with an ADAM10 inhibitor. Conversely, siRNA-mediated ADAM10 knockdown reduced PrPC shedding and increased AβO binding, which was blocked by the PrPC-specific antibody 6D11. The retinoic acid receptor analog acitretin, which up-regulates ADAM10, also promoted PrPC shedding and decreased AβO binding in the neuroblastoma cells and in human induced pluripotent stem cell (iPSC)-derived cortical neurons. Pretreatment with acitretin abolished activation of Fyn kinase and prevented an increase in reactive oxygen species caused by AβO binding to PrPC. Besides blocking AβO binding and toxicity, acitretin also increased the nonamyloidogenic processing of APP. However, in the iPSC-derived neurons, Aβ and other amyloidogenic processing products did not exhibit a reciprocal decrease upon acitretin treatment. These results indicate that by promoting the shedding of PrPC in human neurons, ADAM10 activation prevents the binding and cytotoxicity of AβO, revealing a potential therapeutic benefit of ADAM10 activation in AD.

Amyloid Precursor Protein Mediates Neuronal Protection from Rotenone Toxicity

Mitochondrial complex I dysfunction is the most common respiratory chain defect in human disorders and a hotspot for neurodegenerative diseases. Amyloid precursor protein (APP) and its non-amyloidogenic processing products, in particular soluble APP α (sAPPα), have been shown to provide neuroprotection in models of neuronal injury; however, APP-mediated protection from acute mitochondrial injury has not been previously reported. Here, we use the plant-derived pesticide rotenone, a potent complex I-specific mitochondrial inhibitor, to discover neuroprotective effects of APP and sAPPα in vitro, in neuronal cell lines over-expressing APP, and in vivo, in a retinal neuronal rotenone toxicity mouse model. Our results show that APP over-expression is protective against rotenone toxicity in neurons via sAPPα through an autocrine/paracrine mechanism that involves the Pi3K/Akt pro-survival pathway. APP−/− mice exhibit greater susceptibility to retinal rotenone toxicity, while intravitreal delivery of sAPPα reduces inner retinal neuronal death in wild-type mice following rotenone challenge. We also show a significant decrease in human retinal expression of APP with age. These findings provide insights into the therapeutic potential of non-amyloidogenic processing of APP in complex I-related neurodegeneration.

Amyloid precursor protein-mediated mitochondrial regulation and Alzheimer's disease.

Despite clear evidence of a neuroprotective physiological role of amyloid precursor protein (APP) and its non-amyloidogenic processing products, APP has been investigated mainly in animal and cellular models of amyloid pathology in the context of Alzheimer's disease. The rare familial mutations in APP and presenilin-1/2, which sometimes drive increased amyloid β (Aβ) production, may have unduly influenced Alzheimer's disease research. APP and its cleavage products play important roles in cellular and mitochondrial metabolism, but many studies focus solely on Aβ. Mitochondrial bioenergetic metabolism is essential for neuronal function, maintenance and survival, and multiple reports indicate mitochondrial abnormalities in patients with Alzheimer's disease. In this review, we focus on mitochondrial abnormalities reported in sporadic Alzheimer's disease patients and the role of full-length APP and its non-amyloidogenic fragments, particularly soluble APPα, on mitochondrial bioenergetic metabolism. We do not review the plethora of animal and in vitro studies using mutant APP/presenilin constructs or experiments using exogenous Aβ. In doing so, we aim to invigorate research and discussion around non-amyloidogenic APP processing products and the mechanisms linking mitochondria and complex neurodegenerative disorders such as sporadic Alzheimer's disease. LINKED ARTICLES: This article is part of a themed section on Therapeutics for Dementia and Alzheimer's Disease: New Directions for Precision Medicine. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v176.18/issuetoc.

Distinct invivo roles of secreted APP ectodomain variants APPsα and APPsβ in regulation of spine density, synaptic plasticity, and cognition.

Increasing evidence suggests that synaptic functions of the amyloid precursor protein (APP), which is key to Alzheimer pathogenesis, may be carried out by its secreted ectodomain (APPs). The specific roles of APPsα and APPsβ fragments, generated by non-amyloidogenic or amyloidogenic APP processing, respectively, remain however unclear. Here, we expressed APPsα or APPsβ in the adult brain of conditional double knockout mice (cDKO) lacking APP and the related APLP2. APPsα efficiently rescued deficits in spine density, synaptic plasticity (LTP and PPF), and spatial reference memory of cDKO mice. In contrast, APPsβ failed to show any detectable effects on synaptic plasticity and spine density. The C-terminal 16 amino acids of APPsα (lacking in APPsβ) proved sufficient to facilitate LTP in a mechanism that depends on functional nicotinic α7-nAChRs. Further, APPsα showed high-affinity, allosteric potentiation of heterologously expressed α7-nAChRs in oocytes. Collectively, we identified α7-nAChRs as a crucial physiological receptor specific for APPsα and show distinct invivo roles for APPsα versus APPsβ. This implies that reduced levels of APPsα that might occur during Alzheimer pathogenesis cannot be compensated by APPsβ.

Impairment of amyloid precursor protein alpha-processing in cerebral microvessels of type 1 diabetic mice

The mechanisms underlying dysfunction of cerebral microvasculature induced by type 1 diabetes (T1D) are not fully understood. We hypothesized that in cerebral microvascular endothelium, α-processing of amyloid precursor protein (APP) is impaired by T1D. In cerebral microvessels derived from streptozotocin (STZ)-induced T1D mice protein levels of APP and its α-processing enzyme, a disintegrin and metalloprotease 10 (ADAM10) were significantly decreased, along with down-regulation of adenylate cyclase 3 (AC3) and enhanced production of thromboxane A2 (TXA2). In vitro studies in human brain microvascular endothelial cells (BMECs) revealed that knockdown of AC3 significantly suppressed ADAM10 protein levels, and that activation of TXA2 receptor decreased APP expression. Furthermore, levels of soluble APPα (sAPPα, a product of α-processing of APP) were significantly reduced in hippocampus of T1D mice. In contrast, amyloidogenic processing of APP was not affected by T1D in both cerebral microvessels and hippocampus. Most notably, studies in endothelial specific APP knockout mice established that genetic inactivation of APP in endothelium was sufficient to significantly reduce sAPPα levels in the hippocampus. In aggregate, our findings suggest that T1D impairs non-amyloidogenic processing of APP in cerebral microvessels. This may exert detrimental effect on local concentration of neuroprotective molecule, sAPPα, in the hippocampus.

M344 promotes nonamyloidogenic amyloid precursor protein processing while normalizing Alzheimer’s disease genes and improving memory

Alzheimer's disease (AD) comprises multifactorial ailments for which current therapeutic strategies remain insufficient to broadly address the underlying pathophysiology. Epigenetic gene regulation relies upon multifactorial processes that regulate multiple gene and protein pathways, including those involved in AD. We therefore took an epigenetic approach where a single drug would simultaneously affect the expression of a number of defined AD-related targets. We show that the small-molecule histone deacetylase inhibitor M344 reduces beta-amyloid (Aβ), reduces tau Ser396 phosphorylation, and decreases both β-secretase (BACE) and APOEε4 gene expression. M344 increases the expression of AD-relevant genes: BDNF, α-secretase (ADAM10), MINT2, FE65, REST, SIRT1, BIN1, and ABCA7, among others. M344 increases sAPPα and CTFα APP metabolite production, both cleavage products of ADAM10, concordant with increased ADAM10 gene expression. M344 also increases levels of immature APP, supporting an effect on APP trafficking, concurrent with the observed increase in MINT2 and FE65, both shown to increase immature APP in the early secretory pathway. Chronic i.p. treatment of the triple transgenic (APPsw/PS1M146V/TauP301L) mice with M344, at doses as low as 3 mg/kg, significantly prevented cognitive decline evaluated by Y-maze spontaneous alternation, novel object recognition, and Barnes maze spatial memory tests. M344 displays short brain exposure, indicating that brief pulses of daily drug treatment may be sufficient for long-term efficacy. Together, these data show that M344 normalizes several disparate pathogenic pathways related to AD. M344 therefore serves as an example of how a multitargeting compound could be used to address the polygenic nature of multifactorial diseases.

Amyloid precursor protein drives down-regulation of mitochondrial oxidative phosphorylation independent of amyloid beta

Amyloid precursor protein (APP) and its extracellular domain, soluble APP alpha (sAPPα) play important physiological and neuroprotective roles. However, rare forms of familial Alzheimer’s disease are associated with mutations in APP that increase toxic amyloidogenic cleavage of APP and produce amyloid beta (Aβ) at the expense of sAPPα and other non-amyloidogenic fragments. Although mitochondrial dysfunction has become an established hallmark of neurotoxicity, the link between Aβ and mitochondrial function is unclear. In this study we investigated the effects of increased levels of neuronal APP or Aβ on mitochondrial metabolism and gene expression, in human SH-SY5Y neuroblastoma cells. Increased non-amyloidogenic processing of APP, but not Aβ, profoundly decreased respiration and enhanced glycolysis, while mitochondrial DNA (mtDNA) transcripts were decreased, without detrimental effects to cell growth. These effects cannot be ascribed to Aβ toxicity, since higher levels of endogenous Aβ in our models do not cause oxidative phosphorylation (OXPHOS) perturbations. Similarly, chemical inhibition of β-secretase decreased mitochondrial respiration, suggesting that non-amyloidogenic processing of APP may be responsible for mitochondrial changes. Our results have two important implications, the need for caution in the interpretation of mitochondrial perturbations in models where APP is overexpressed, and a potential role of sAPPα or other non-amyloid APP fragments as acute modulators of mitochondrial metabolism.

Iron dysregulates APP processing accompanying with sAPPα cellular retention and β-secretase inhibition in rat cortical neurons.

Amyloid precursor protein (APP) and iron both play pivotal roles in the central nervous system, but whether and how iron influences the processing of endogenous APP in neurons remain unclear. Here, we investigated the regulatory effects and underlying mechanisms of iron on non-amyloidogenic and amyloidogenic processing of APP in rat primary cortical neurons. Treatment of the neurons with ferric ammonium citrate (FAC, 100 μmol/L) markedly facilitated the non-amyloidogenic processing of APP, as evidenced by a robust increase in α-secretase-derived carboxy-terminal fragment α (CTFα). Furthermore, the distribution of sAPPα was altered after iron treatment, and sAPPα remained in the cellular lysates instead of being secreted into the extracellular milieu. Moreover, the levels of APP amyloidogenic products, including sAPPβ and Aβ were both decreased. We further revealed that FAC did not alter the expression of β-secretase, but significantly suppressed its enzymatic activity in iron-treated neurons. In a cell-free β-secretase activity assay, FAC dose-dependently inhibited the activity of purified β-secretase with an IC50 value of 21.67 μmol/L. Our data provide the first evidence that iron overload alters the neuronal sAPPα distribution and directly inhibits β-secretase activity. These findings shed light on the regulatory mechanism of bio-metals on APP processing.