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Cellular actions of beta-amyloid precursor protein and its soluble and fibrillogenic derivatives
Abstract
beta-Amyloid precursor protein (beta-APP), the source of the fibrillogenic amyloid beta-peptide (A beta) that accumulates in the brain of victims of Alzheimer's disease, is a multifunctional protein that is widely expressed in the nervous system. beta-Amyloid precursor protein is axonally transported and accumulates in presynaptic terminals and growth cones. A secreted form of beta-APP (sAPP alpha) is released from neurons in response to electrical activity and may function in modulation of neuronal excitability, synaptic plasticity, neurite outgrowth, synaptogenesis, and cell survival. A signaling pathway involving guanosine 3',5'-cyclic monophosphate is activated by sAPP alpha and modulates the activities of potassium channels, N-methyl-D-aspartate receptors, and the transcription factor NF kappa B. Additional functions of beta-APP may include modulation of cell adhesion and regulation of proliferation of nonneuronal cells. Alternative enzymatic processing of beta-APP liberates A beta, which has a propensity to form amyloid fibrils; A beta can damage and kill neurons and increase their vulnerability to excitotoxicity. The mechanism involves generation of oxyradicals and impairment of membrane transport systems (e.g., ion-motive ATPases and glutamate and glucose transporters). Genetic mutations or age-related metabolic changes may promote neuronal degeneration in Alzheimer's disease by increasing production of A beta and/or decreasing levels of neuroprotective sAPP alpha.
... Memory loss, called "blackout", a kind of temporary anterograde amnesia [16] is frequently experienced by teenagers [16] and, one predictive factor of "blackout" experiences is female sex [16] which is in accordance with a higher vulnerability of female to ethanol-induced cognitive impairment [9,12] . Preclinical studies described ethanol-induced memory impairments in males or females [17][18][19][20] , with a possible role of estrogen in female [21,22] . However, there is a lack of demonstration that higher vulnerability to memory impairments in female rodents includes a role for circulating estrogen. ...
... Among these targets, estrogen increases GluN2B-containing NMDA receptor current and mRNA levels [23] while GluN2B protein or level of phosphorylation were neither increased [25] , leading to the suggestion that estrogen recruits GluN2B-containing NMDA receptor at the synapse. In our previous study performed in adolescent/young adult male rats, we measured an overexpression of GluN2B 48h after two binge-like ethanol exposures [26] while LTD (Longterm Depression) was blocked leading to cognitive deficit [20] . Therefore, we tested whether estrogen modulates ethanol-induced LTD abolition after binge-like ethanol exposure in female rats. ...
... Post-pubertal female rats received 2 i.p. injections of ethanol (3g/kg; 20%v/v) given 9h apart [20,26] in the different phases of the estrous cycle. Recording was performed 24 or 48h later. ...
Le binge drinking (BD) est un mode de consommation d'alcool très répandu chez les jeunes adultes et qui se caractérise par une prise massive et épisodique d'alcool dans un laps de temps court afin d'atteindre rapidement l'état d’ivresse. Chez ces jeunes, le BD est responsable de perturbations des capacités cognitives et la consommation des femmes rattrape celle des hommes alors qu'elles seraient plus sensibles aux effets délétères de l'éthanol (EtOH) sur le cerveau. Dans ce contexte, il nous est apparu important de comprendre les différences liées au sexe dans les effets du BD sur le fonctionnement cérébral, et notamment la plasticité neuronale d’e l'hippocampe, mécanisme cellulaire de la mémoire et des apprentissages. Ainsi, les objectifs de ma thèse ont été i) d'explorer l'impact du cycle ovarien sur les effets du BD sur la plasticité neuronale de l'hippocampe, ii) de mesurer la modulation des effets de l'éthanol par les œstrogènes pendant l'adolescence chez la femelle et chez le mâle, et iii) de chercher les mécanismes d'action impliqués. Ainsi, nous avons enregistré chez la ratte adolescente la plasticité du réseau hippocampique ex vivo après une exposition courte à l'éthanol après 2 binges d'éthanol. Nous avons mis en évidence une abolition de la plasticité de type dépression à long terme (LTD) à 24h de délai, seulement lorsque l'administration d'éthanol était effectuée pendant le pic d'œstrogènes endogènes, suggérant une interaction entre éthanol et œstrogènes (E2). Pour vérifier cette hypothèse, nous avons mesuré chez des rattes pubères, prépubères, et chez des mâles adolescents les effets sur la DLT d'un traitement exogène d’E2+EtOH. L'abolition de la DLT a été observé à chaque fois. L'utilisation d'antagonistes spécifiques nous a permis de mettre en évidence l'implication des récepteurs aux œstrogènes, ainsi que de la sous-unité GluN2B des récepteurs NMDA dans ces résultats. Nous concluons que les œstrogènes pourrait être un facteur de vulnérabilité liés au genre vis-à-vis des effets de l'alcool sur les capacités cognitives chez les femelles, pendant l'adolescence
... La voie faisant intervenir l'α-sécrétase est dite « non amyloïdogène » [6] . Celle-ci prévient la production de peptide Aβ et libère un fragment sAPPα («s» pour soluble) qui est neuroprotecteur [7]. La voie dite « amyloïdogène » fait intervenir une β-sécrétase clivant l'APP et aboutissant à la sécrétion d'un fragment N-terminal (sAPPβ). ...
... Dans la MA deux processus pathologiques post-traductionnels interviennent : l'hyperphosphorylation et la phosphorylation anormale des protéines Tau. Ces protéines Tau anormalement phosphorylées s'agrègent en paquets de filaments pathologiques déstabilisant les microtubules à l'origine de la perte du transport axonal [7]. ...
German neuropsychiatrist Alois Alzheimer is less well known than the disease that bears his name, which he first described over a century ago. Indeed, it was in 1906, after the death of one of his patients with memory problems, that he studied her brain and discovered characteristic lesions. Even today, the definitive diagnosis of Alzheimer's disease is based on the histological study of post-mortem brain sections and the identification of these lesions. Yet medicine has made many advances in our understanding of the disease. In particular, knowledge of the basic anatomopathological lesions of the disease has enabled the identification of pathophysiological mechanisms. These discoveries have opened up new perspectives in the development of diagnostic and therapeutic tools. Nevertheless, these efforts have not yet borne fruit, since the entire pathophysiological process leading to the disease is not yet fully understood. What's more, Alzheimer's disease remains an incurable disease: the management of patients and their families, from diagnostic suspicion to treatment, is far from resolved. The one goes hand in hand with the other, and it is perhaps by diagnosing patients better and, above all, earlier that therapies will have the best chance of being effective. In the absence of a reliable disease marker, the current diagnostic approach combines several clinical and paraclinical arguments, leading to a diagnosis of probability at a stage of the disease that is often too advanced. Biomarkers of Alzheimer's disease in cerebrospinal fluid, the biological and pathophysiological elements of this set of arguments, are currently used routinely in most university hospitals. However, while these standard biomarkers provide invaluable assistance and have revolutionized the diagnostic approach, they still have limitations in the differential diagnosis of dementia. In order to improve the performance of these tests, various combinations of biomarkers have been developed to overcome these difficulties.
... Amyloid plaques are formed by the accumulation of extracellular aggregates of β-amyloid (Aβ) peptides, resulting from the sequential proteolysis of the amyloid precursor protein (APP) by β and γ secretases. In contrast to Aβ peptides, the soluble fragment of APP (sAPPα), generated by α-secretase, appears to have neurotrophic and neuroprotective properties [18]. Neurofibrillary lesions consist of intraneuronal fibrillar aggregates of hyperphosphorylated and abnormally phosphorylated Tau proteins. ...
... One of the characteristic lesions found in AD, amyloid plaques, is specifically composed of Aβ peptides. Aβ peptides derive from the amyloidogenic processing of APP by β and γ secretases, while alternative cleavage by α secretase produces a soluble neurotrophic and neuroprotective fragment, sAPPα, which prevents the formation of Aβ peptides [18]. P2X7 can affect Aβ build-up via this and other pathways. ...
Alzheimer’s disease (AD) is a progressive neurodegenerative disease characterized by impaired episodic memory and two pathological lesions: amyloid plaques and neurofibrillary tangles. In AD, damaged neurons and the accumulation of amyloid β (Aβ) peptides cause a significant release of high amounts of extracellular ATP, which acts as a danger signal. The purinergic receptor P2X7 is the main sensor of high concentrations of ATP, and P2X7 has been shown to be upregulated in the brains of AD patients, contributing to the disease’s pathological processes. Further, there are many polymorphisms of the P2X7 gene that impact the risk of developing AD. P2X7 can directly modulate Aβ plaques and Tau protein lesions as well as the inflammatory response by regulating NLRP3 inflammasome and the expression of several chemokines. The significant role of microglial P2X7 in AD has been well established, although other cell types may also be important in P2X7-mediated mechanisms. In this review, we will discuss the different P2X7-dependent pathways involved in the development of AD.
... In both humans (Zott et al., 2018) and animals, amyloid proximity correlates with neuronal hyperactivity (Busche, 2012;Gurevicius et al., 2013;Busche and Konnerth, 2016;Zott et al., 2019). Downstream consequences of amyloid accumulation, such as glial activation (Barger and Harmon, 1997;Pasqualetti et al., 2015;Hansen et al., 2018) or neuronal death (Klyubin, 2008;Mattson, 1997;Smale et al., 1995;Kobayashi et al., 2002;Kobayashi et al., 2005), might also shape pain perception. ...
... Amyloid exposure alone can lead to increased neuronal excitability (Busche, 2012;Busche et al., 2008). Amyloid presence can induce both glial activation and neuronal cell death (Klyubin, 2008;Mattson, 1997;Fig. 8. Mechanical sensitivity. ...
Pain and cognitive decline increase with age. In particular, there is a troubling relationship between dementia and pain, with some studies showing higher prevalence and inadequate treatment of pain in this population. Alzheimer’s disease (AD) is one of the most common causes of dementia in older adults. Amyloid plaques are a hallmark of AD. The downstream processes these plaques promote are believed to affect neuronal and glial health and activity. There is a need to better understand how the neuropathological changes of AD shape neural activity and pain sensitivity. Here, we use the 5XFAD mouse model, in which dense amyloid accumulations occur at early ages, and in which previous studies reported signs of cognitive decline. We hypothesized that 5XFAD mice develop sensory and pain processing dysfunctions. Although amyloid burden was high throughout the brain, including in regions involved with sensory processing, we identified no functionally significant differences in reflexive or spontaneous signs of pain. Furthermore, expected signs of cognitive decline were modest; a finding consistent with variable results in the literature. These data suggest that models recapitulating other pathological features of Alzheimer’s disease might be better suited to studying differences in pain perception in this disease.
... APP is an integral plasma membrane protein with only one membrane traversing helix and its amino-terminus is large extracellular and glycosylated while its carboxyl-terminus is shorter and it is in the cytoplasm. There are several different isoforms of APP but the most common isoform is (APP695) which is located in the brain and this isoform is produced primarily in neurons (Mattson, 1997). ...
Alzheimer’s disease is the leading predominant demyelinating and degenerative ailment, described by the loss of cognitive function because of advanced neuronal loss in the brain. There are two pathological hallmarks in the brains of AD sufferers. One is the buildup of abnormal Tau proteins in neurons and the other is the formation of amyloid plaques. Although we still don’t know the comprehensive mechanism that is involved in AD pathophysiology; there are enormous studies that suggested thatmalfunctioning of mitochondria plays a substantial function in the pathology of AD. As we know that mitochondria are very dynamic organelles and the powerhouse of the cell (generate ATP). And it is said that a healthy pool of mitochondria is very essential because it provides energy to the neurons to perform the most important functions and it also protects the neurons by reducing the oxidative damage related to mitochondria. These organelles also play many important cellular functions such asregulation of intracellular calcium ions, bioenergetics processes, a scavenging system for free radicals, and stimulation of cell death that is mediated by caspases. But these functions can be adversely affected by amyloid beta-mediated mitochondrial dysfunction. In this article, I recapitulated the current advancement that highlights the starring role of mitochondrial impairment in AD pathology and summarize how different types of mechanisms are involved in mitochondrial impairment in thepathophysiology of AD.
... The b segment produces A-b and g-secretases present in amyloid plaques in AD patients' brains. When a-secretase processes APP in a non-amyloidogenic manner, it leads to hydrolysis of A-b peptide sequences and shedding of sAPPa, which has neuroprotective and neurotrophic effects (141). ...
The P2X7 receptor (P2X7R), a non-selective cation channel modulated by adenosine triphosphate (ATP), localizes to microglia, astrocytes, oligodendrocytes, and neurons in the central nervous system, with the most incredible abundance in microglia. P2X7R partake in various signaling pathways, engaging in the immune response, the release of neurotransmitters, oxidative stress, cell division, and programmed cell death. When neurodegenerative diseases result in neuronal apoptosis and necrosis, ATP activates the P2X7R. This activation induces the release of biologically active molecules such as pro-inflammatory cytokines, chemokines, proteases, reactive oxygen species, and excitotoxic glutamate/ATP. Subsequently, this leads to neuroinflammation, which exacerbates neuronal involvement. The P2X7R is essential in the development of neurodegenerative diseases. This implies that it has potential as a drug target and could be treated using P2X7R antagonists that are able to cross the blood-brain barrier. This review will comprehensively and objectively discuss recent research breakthroughs on P2X7R genes, their structural features, functional properties, signaling pathways, and their roles in neurodegenerative diseases and possible therapies.
... For a neuropathological diagnosis of AD, it becomes imperative to determine the density of neuritic plaques and the topographic distribution of NFTs [74]. Oxidative stress emerge as the prime causative factor in the pathophysiology of diseases like Alzheimer's, and research has established a linkage between Aβ and oxidative stress [75,76]. Aβ triggers an increase in levels of hydrogen peroxide and lipid peroxides both in vitro and in vivo, suggesting its role as a potent inducer of oxidative stress, leading to impaired synaptic function and neuronal demise. ...
Purpose of Review
Phloroglucinol (PG) is a potent secondary metabolite found in various organisms, with extensive applications in medicine since its discovery in the nineteenth century. Research has demonstrated its diverse properties, including anti-inflammatory, antioxidant, anti-viral, anti-cancer, and anthelmintic effects. In clinical settings, it alleviates pain by relaxing the gastrointestinal, biliary, and urinary systems. Despite its broad pharmacological potential, its use in treating neurodegenerative diseases (ND) remains underexplored. This study systematically explores PG’s role in ND management, utilizing data exclusively from reputable sources like PubMed, Google Scholar, Scopus, ScienceDirect, and SpringerLink, and adhering to PRISMA guidelines to provide structured insights.
Recent findings
The review emphasizes PG’s significant role in treating various neurodegenerative diseases (NDs) such as Alzheimer’s disease (AD), Parkinson’s disease (PD), Huntington’s disease (HD), and amyotrophic lateral sclerosis (ALS). PG offers cytoprotective effects by preventing dendritic spine density loss in AD, reducing ROS levels in PD and ALS, and inhibiting amyloid aggregate formation in HD. These benefits stem from its powerful antioxidant properties, which alleviate cerebral oxidative stress. Overall, PG stands out as an affordable and readily accessible resource with promising pharmaceutical applications for treating NDs.
Summary
This study underscores the prospect of PG serving as a wellspring for innovative drugs catering to the intricate demands of neurodegenerative conditions. Nevertheless, further research is necessary to thoroughly assess its clinical effectiveness in human subjects to ensure its widespread acceptance in clinical practice.
... Both Aβ40 and Aβ42 are soluble helical peptides that undergo a conformational transition from α-helix to β-sheet structures and eventually form β-sheet-enriched fibril structures [6]. Previous studies of Aβ toxicity revealed that fibrillar Aβ could directly kill neurons or initiate a cascade of events leading to neuronal cell death [7][8][9]. The lack of cures for Alzheimer's disease poses a major challenge because many available AD drugs only reduce the symptoms of dementia and are not capable of preventing the progression of the disease, and do not halt the slow deterioration of the brain [10,11]. ...
Chlorella sp. is green freshwater microalgae have drawn great attention as a promising sustainable source of lipids and carotenoids. As neuroprotective natural products, carotenoids have shown promising preventive activity, as well as helping in slowing down Alzheimer's disease (AD) progression. However, the detailed information on the inhibition mechanism of amyloid beta (Aβ) fibril, one of the hallmarks of AD, by carotenoid compounds is poorly discussed in both experimental and computational studies. Thus, in this study, molecular docking simulations were performed to investigate the binding interactions between nine carotenoid compounds derived from Chlorella sp. against full sequence Aβ42 fibril. The results reveal that the binding energies ranged from -5.3 to -6.5 kcal/mol with binding interactions were dominated by hydrophobic interactions via π-alkyl, and only two carotenoid compounds, fucoxanthin and zeaxanthin, formed hydrogen bonds. In comparison, donepezil showed binding energy of -6.7 kcal/mol and interacted with Aβ42 residues through a hydrogen bond and hydrophobic interaction that involves alkyl, π- alkyl, π-sigma and π-π T-shape interactions. Our result showed that donepezil, fucoxanthin and zeaxanthin compounds disrupted the Aβ42 fibril by disaggregation pathway, which primarily interacted with residues within the hydrophobic (Phe19, Phe20, Val24, Ala30, and Ile32) and N-terminal (Tyr10, Val12, His13, and His14) regions. This study provides theoretical insights into the inhibitory mechanism of antioxidant compounds against Aβ fibril, which is beneficial for AD drug design.
... Thereafter, the majority of Aβ is produced in the Golgi complex and endosomes. The exact function of APP is still not fully understood, but several studies suggest that APP participates in important neuronal functions including neuronal survival, neurite outgrowth, enhanced synaptogenesis, and synaptic plasticity [23]. It is also suggested that APP could act as a receptor, transducing signals extracellularly, but this hypothesis is not supported by strong evidence. ...
After several years of research in the field of Alzheimer’s disease (AD), it is still unclear how amyloid-beta (Aβ) and Tau, two key hallmarks of the disease, mediate the neuropathogenic events that lead to AD. Current data challenge the “Amyloid Cascade Hypothesis” that has prevailed in the field of AD, stating that Aβ precedes and triggers Tau pathology that will eventually become the toxic entity in the progression of the disease. This perspective also led the field of therapeutic approaches towards the development of strategies that target Aβ or Tau. In the present review, we discuss recent literature regarding the neurotoxic role of both Aβ and Tau in AD, as well as their physiological function in the healthy brain. Consequently, we present studies suggesting that Aβ and Tau act independently of each other in mediating neurotoxicity in AD, thereafter, re-evaluating the “Amyloid Cascade Hypothesis” that places Tau pathology downstream of Aβ. More recent studies have confirmed that both Aβ and Tau could propagate the disease and induce synaptic and memory impairments via the amyloid precursor protein (APP). This finding is not only interesting from a mechanistic point of view since it provides better insights into the AD pathogenesis but also from a therapeutic point of view since it renders APP a common downstream effector for both Aβ and Tau. Subsequently, therapeutic strategies that act on APP might provide a more viable and physiologically relevant approach for targeting AD.
... Aβ protein is a peptide fragment derived from its precursor protein, amyloid precursor protein (APP). APP plays crucial role in regulating neuronal excitability, synaptic plasticity, neuronal dendritic growth, synaptogenesis, and cell survival [24]. In the brain, β-secretase and γ-secretase cleave APP into Aβ40 and Aβ42. ...
Alzheimer's disease (AD) is a chronic neurodegenerative disorder that is distinguished by impairments in learning and memory. The microbiota-gut-brain axis has emerged as a significant factor in the pathogenesis of Alzheimer's disease. Eucommiae cortex polysaccharides (EPs), one of the most abundant substances in the Eucommiae cortex , exhibit potential immunomodulatory and neuroprotective effects. In our study, we discovered that long-term intervention with EPs (0.25%, w/w in food) significantly improved learning and memory in 5×FAD mice, a well recognized mouse model for Alzheimer's disease. Additionally, EPs substantially reduce the generation and deposition of amyloid-beta (Aβ) plaques in AD Mice. EPs reshaped the composition of gut microbiota, increased the abundance of Erysipelotrichaceae , Allobaculum , Bacteroidota , and Muribaculaceae , which are related to gut immunity and short-chain fatty acids(SCFs) production that exert neuroprotective effects through the gut-brain axis. Furthermore, EPs administration was able to correct metabolic disorders in AD mice, particularly glutamate metabolism disturbances. Correlation analysis demonstrates a robust association between the restructuring of gut microbiota and alterations in glutamate metabolism. Importantly, EPs administration significantly inhibited neuroinflammation and restored the oxidative-antioxidative balance in the brains of AD mice, which are potential factors contributing to learning and memory impairments. Interestingly, EPs administration significantly activated autophagy. Our results demonstrate that EPs reshaped gut microbiota, corrected glutamate metabolism disturbances, and significantly alleviated learning and memory impairments in 5xFAD mice through the gut-brain axis. The natural extract EPs holds promising potential for the therapeutic management of neurodegenerative disorders.
... The production of Aβ primarily occurs through the proteolytic cleavage of the transmembrane precursor protein (AβPP) by βand γ-secretases. 13,14 The two predominant monomeric forms of Aβ peptides are Aβ40 and Aβ42, with Aβ42 being more neurotoxic and prone to fibrillogenesis. 15,16 Given the important role of Aβ42 and its aggregates in AD, extensive studies have focused on investigating the structure of Aβ42 and the underlying mechanism of Aβ42 peptide selfassembly. ...
... APP is produced in neurons, released from presynaptic terminals, and available at the dendritic synapse (Ishida et al., 1997;Weingarten et al., 2017). Constitutional APP (before processing) contributes to neurite growth (Mattson, 1997;Turner et al., 2003), gliogenesis (Kwak et al., 2010), synaptogenesis (Turner et al., 2003), neuroproliferation (Mattson, 1994), migration (Priller et al., 2006), suppression of cell adhesion (Chen and Bodles, 2007), and participates in translational pathways (Wang et al., 2004;Westmark and Malter, 2007). ...
Recent studies promote new interest in the intersectionality between autism spectrum disorder (ASD) and Alzheimer’s Disease. We have reported high levels of Amyloid-β Precursor Protein (APP) and secreted APP-alpha (sAPP a ) and low levels of amyloid-beta (Aβ) peptides 1–40 and 1–42 (Aβ40, Aβ42) in plasma and brain tissue from children with ASD. A higher incidence of microcephaly (head circumference less than the 3 rd percentile) associates with ASD compared to head size in individuals with typical development. The role of Aβ peptides as contributors to acquired microcephaly in ASD is proposed. Aβ may lead to microcephaly via disruption of neurogenesis, elongation of the G1/S cell cycle, and arrested cell cycle promoting apoptosis. As the APP gene exists on Chromosome 21, excess Aβ peptides occur in Trisomy 21-T21 (Down’s Syndrome). Microcephaly and some forms of ASD associate with T21, and therefore potential mechanisms underlying these associations will be examined in this review. Aβ peptides’ role in other neurodevelopmental disorders that feature ASD and acquired microcephaly are reviewed, including dup 15q11.2-q13, Angelman and Rett syndrome.
... Bu nedenle, bağışıklık sisteminin anti-inflamatuar ilaçlarla bastırılması, Tau oligomerlerinin sağlıklı nöronlara yayılmasını artırabileceğinden, uzun vadede AH tedavisi için yararlı olmayabilir. Pek çok araştırma, Aβ'nın neden olduğu nörotoksisitede oksidatif stresi işaret etmektedir (Mattson, 1997). Hücre modellerini kullanan in vitro deneyler, Aβ tedavisinin hidrojen peroksit ve lipid peroksit seviyelerini artırabileceğini göstermiştir (Behl vd., 1994 (Yao, vd., 2005;Nunomura, vd., 2001). ...
... Mutations in the APP gene are linked to rare familial forms of Alzheimer's disease [14]. According to [15], it includes, for instance, D678N (D7N in Aβ) [16], E693G (E22G in Ab) [17], and V717I or V717F [18,19]. Specifically, the L723P mutation of APP is found in the Australian early onset AD pedigree [20]. ...
One of the hallmarks of Alzheimer’s disease (AD) is the accumulation of amyloid beta (Aβ) peptides in the brain. The processing of amyloid precursor protein (APP) into Aβ is dependent on the location of APP in the membrane, membrane lipid composition and, possibly, presence of lipid rafts. In this study, we used atomic force microscopy (AFM) to investigate the interaction between transmembrane fragment APP672–726 (corresponding to Aβ1–55) and its amyloidogenic mutant L723P with membranes combining liquid-ordered and liquid-disordered lipid phases. Our results demonstrated that most of the APP672–726 is located either in the liquid-disordered phase or at the boundary between ordered and disordered phases, and hardly ever in rafts. We did not notice any major changes in the domain structure induced by APP672–726. In membranes without cholesterol APP672–726, and especially its amyloidogenic mutant L723P formed annular structures and clusters rising above the membrane. Presence of cholesterol led to the appearance of concave membrane regions up to 2 nm in depth that were deeper for wild type APP672–726. Thus, membrane cholesterol regulates changes in membrane structure and permeability induced by APP that might be connected with further formation of membrane pores.
... Interestingly, a study published during the 2000s established that insulin might induce the release of soluble APPα fragments through PI3K activation in SH-SY5Y cells [38]. Soluble APPα is hypothesized to have a neuroprotective effect by modulating neuronal excitability, synaptic plasticity, neurite outgrowth, synaptogenesis, and cell survival [39]. ...
Insulin resistance as a hallmark of type 2 DM (T2DM) plays a role in dementia by promoting pathological lesions or enhancing the vulnerability of the brain. Numerous studies related to insulin/insulin-like growth factor 1 (IGF-1) signaling are linked with various types of dementia. Brain insulin resistance in dementia is linked to disturbances in Aβ production and clearance, Tau hyperphosphorylation, microglial activation causing increased neuroinflammation, and the breakdown of tight junctions in the blood–brain barrier (BBB). These mechanisms have been studied primarily in Alzheimer’s disease (AD), but research on other forms of dementia like vascular dementia (VaD), Lewy body dementia (LBD), and frontotemporal dementia (FTD) has also explored overlapping mechanisms. Researchers are currently trying to repurpose anti-diabetic drugs to treat dementia, which are dominated by insulin sensitizers and insulin substrates. Although it seems promising and feasible, none of the trials have succeeded in ameliorating cognitive decline in late-onset dementia. We highlight the possibility of repositioning anti-diabetic drugs as a strategy for dementia therapy by reflecting on current and previous clinical trials. We also describe the molecular perspectives of various types of dementia through the insulin/IGF-1 signaling pathway.
... The neurons damage in AD patients is initiated by extracellular accumulation of amyloid-β (Aβ) containing plaques which is produced from the Aβ precursor protein (APP) proteolysis that is accompanied by failure of the clearance mechanisms. In addition, there are accumulation of neurofibrillary tangles that are formed of aggregates of hyperphosphorylated tau protein intracellularly (which is considered a primary biomarker of AD), as well as signs of proteolysis mediated by increased Ca2+ levels and oxidative stress (Mattson 1997). ...
Neurons are postmitotic cells that may remain functional for years and cannot be replaced easily, so irreversible neuronal damage could result in destructive diseases like neurodegenerative illnesses. The mechanism of neuronal cell death represents a fundamental factor in determining the pathway of many diseases, and since the neurons take a long time to be lost, it might be challenging to recognize the modes of cell death and the initiating events.
Neurological studies have become more interested recently in the molecular mechanisms of apoptotic and excitotoxic cell death in the central nervous system lesions and their relevance to different neurodegenerative disorders. As well, the interaction between apoptosis and excitotoxicity has drawn attention to shed light on the postulation that excitotoxic cell death pathways could be mediated by apoptotic processes.
Numerous findings demonstrate increased extracellular concentrations of glutamate neurotransmitters and overstimulation of postsynaptic glutamate receptors in most neurodegenerative diseases. The resultant excitotoxic response is likely to elicit some cytotoxic events including increased calcium overload, accumulation of reactive oxygen species, mitochondrial dysfunction, endoplasmic reticulum stress, deficiency in energy supply, and DNA damage, which result in the induction of neuronal death pathways mostly by apoptosis, autophagy, necrosis, or by a combination of different death modes depending on different factors like the insult severity and duration.
Evidence of the crucial role of the apoptotic-excitotoxic pathway in neurodegenerative illnesses is growing, although additional research into the underlying mechanism by which it contributes to neurodegeneration is still needed.
... Hcy is an important inducer of prooxidative-antioxidative imbalance associated with increased intracellular calcium ion concentrations and DNA damage [98,135]. Hcy contributes to oxidative stress by reducing glutathione peroxidase activity and lowering the level of vitamins A, E and C [136]. ...
Homocysteine is an organic compound that can be measured in the blood of humans and animals. High levels of homocysteine in human blood are associated with an increased risk of heart disease, diseases of blood vessels, formation of blood clots and brain damage. However, the role of homocysteine in the health and disease of domestic animals is poorly understood. This review critically appraises the literature concerning homocysteine in animals, focusing on horses. It aims to clearly define the existing knowledge gap to path an avenue for future research into homocysteine as a potential diagnostic marker of health and disease in this species.
... Therefore, a loss of sAPPα neuroprotective and neurotrophic functions could be involved in the decreased neuronal plasticity and the increased neuronal susceptibility to cellular stress observed in aging and neurodegeneration. Experimental evidence suggests that sAPPα neuroprotective actions are correlated with effects on ion channel function followed by late transcription-dependent events [140], as well as pleiotropic effects on cell survival via the PI3K/Akt/NF-κB pathway as we previously demonstrated [141]. In this context, a deeper understanding of the sAPPα-correlated modulation of genes of interest for both AD and TBI could be significant in terms of basic research and future pharmacologic intervention [142]. ...
Amyloid Precursor Protein (APP) and its cleavage processes have been widely investigated in the past, in particular in the context of Alzheimer’s Disease (AD). Evidence of an increased expression of APP and its amyloidogenic-related cleavage enzymes, β-secretase 1 (BACE1) and γ-secretase, at the hit axon terminals following Traumatic Brain Injury (TBI), firstly suggested a correlation between TBI and AD. Indeed, mild and severe TBI have been recognised as influential risk factors for different neurodegenerative diseases, including AD. In the present work, we describe the state of the art of APP proteolytic processing, underlining the different roles of its cleavage fragments in both physiological and pathological contexts. Considering the neuroprotective role of the soluble APP alpha (sAPPα) fragment, we hypothesised that sAPPα could modulate the expression of genes of interest for AD and TBI. Hence, we present preliminary experiments addressing sAPPα-mediated regulation of BACE1, Isthmin 2 (ISM2), Tetraspanin-3 (TSPAN3) and the Vascular Endothelial Growth Factor (VEGFA), each discussed from a biological and pharmacological point of view in AD and TBI. We finally propose a neuroprotective interaction network, in which the Receptor for Activated C Kinase 1 (RACK1) and the signalling cascade of PKCβII/nELAV/VEGF play hub roles, suggesting that vasculogenic-targeting therapies could be a feasible approach for vascular-related brain injuries typical of AD and TBI.
... Nevertheless, in diseases withimpaired cognition, β and γ secretases cleave most of APP, and release Aβ. Several reports showed that insulin therapy reduces Aβ level and increases sAPPα production in the CNS, suggesting that sAPP-α secretion is mediated by insulin [38,39], hence stimulating the insulinsignaling pathway [40,41]. A cohort study has shown an association between Aβ plaques and cognitive impairment on one hand and serine-phosphorylated IRS-1 on the other hand [42]. ...
... sAPPα ist im Gegensatz zu Aβ neuroprotektiv, indem es die Nervenzelle vor Exzitotoxizität schützt. (Mattson 1997 (Ring u. a. 2007). ...
... Alternatively, increased oxidation [89] and/or changes in dimer active conformation [90] by changes in redox status are thought to alter SR activity, especially in the aging brain in which potent oxidative stress occurs (see [4]). Considering that sAPPα activates signaling pathways that protect synapses against excitotoxicity linked to increased oxidation and impaired energy metabolism [91][92][93][94], this antioxidative property may help sAPPα to optimize SR activity and D-serine production. ...
Impaired activation of the N-methyl-D-aspartate subtype of glutamate receptors (NMDAR) by D-serine is linked to cognitive aging. Whether this deregulation may be used to initiate pharmacological strategies has yet to be considered. To this end, we performed electrophysiological extracellular recordings at CA3/CA1 synapses in hippocampal slices from young and aged mice. We show that 0.1 nM of the soluble N-terminal recombinant fragment of the secreted amyloid-protein precursor-α (sAPPα) added in the bath significantly increased NMDAR activation in aged but not adult mice without impacting basal synaptic transmission. In addition, sAPPα rescued the age-related deficit of theta-burst-induced long-term potentiation. Significant NMDAR improvement occurred in adult mice when sAPPα was raised to 1 nM, and this effect was drastically reduced in transgenic mice deprived of D-serine through genetic deletion of the synthesizing enzyme serine racemase. Altogether, these results emphasize the interest to consider sAPPα treatment targeting D-serine-dependent NMDAR deregulation to alleviate cognitive aging.
... Previous studies have identified relationships between cancer and AD in which the amyloid precursor protein (APP) plays an important role. APP is a transmembrane protein, source of β-amyloid aggregation which is one of the major causes of AD, is expressed in various neuron cells and may be involved in development of cells (2,3). In cancer cells, it has been reported that APP is a primary androgen-responsive gene, found in breast and prostate cancer; it is also implicated in various human cancers including colon, lung, breast, parathyroid, prostate, thyroid and breast cancers, and its high immunoreactivity is related with poor prognoses for prostate cancer and estrogen-receptor-positive breast cancer patients (4)(5)(6)(7)(8)(9)(10). ...
7‑Methoxyheptaphylline (7‑MH) is a carbazole extracted from Clausena harmandiana, a medicinal plant that is used to treat headaches and stomachaches. The aim of the present study was to examine the neuroprotective effects and anticancer activity of 7‑MH. Cell death was assessed using an MTT assay and flow cytometry. The expression of apoptosis‑related proteins was determined by western blot analysis. An animal model was used to test anti‑metastasis. The interactions between 7‑MH and the molecular target were observed using molecular docking. The results revealed that 7‑MH provided protection against hydrogen peroxide (H2O2)‑induced neuronal cell death. In cancer cells, 7‑MH induced SH‑SY5Y, 4T1, HT29, HepG2, and LNCaP cell death. 7‑MH inhibited metastasis of HT29 cells in vitro and 4T1‑Luc cells in vitro and in vivo. 7‑MH inhibited proteins, including P‑glycogen synthase kinase (GSK)‑3, and cleaved caspase‑3, but it activated anti‑apoptotic proteins in H2O2‑induced SH‑SY5Y cell death. By contrast, 7‑MH activated the cleaving of caspase‑3 and GSK‑3, but it suppressed anti‑apoptotic proteins in SH‑SY5Y cells. 7‑MH reduced the levels of NF‑κB and STAT3 in 4T1 cells; phospho‑p65, Erk, and MAPK13 in LNCaP cells; and phospho‑Erk and matrix metalloproteinase‑9 in HT29 cells. Molecular docking analysis showed that 7‑MH targets TAK1 kinase. The present study indicated that 7‑MH induced apoptosis of cancer cells and provided protection against H2O2‑induced neuron cell death via TAK1 kinase.
... Conversely, α-secretase can cleave within the Aβ domain, thereby precluding Aβ generation and producing a fragment (sAPPα) that is believed to be neuroprotective [47]. sAPPα is believed to promote neurite outgrowth, synaptogenesis and cell adhesion [48,49] while APPβ acts as ligand for DR6, promoting caspase 6 activation resulting in axonal pruning [50]. ...
The genetic study of multi-incident families is a powerful tool to investigate genetic contributions to the development of Parkinson’s disease. In this study, we identified the rare PTPRA p.R223W variant as one of three putative genetic factors potentially contributing to disease in an Australian family with incomplete penetrance. Whole exome sequencing identified these mutations in three affected cousins. The rare PTPRA missense variant was predicted to be damaging and was absent from 3,842 alleles from PD cases. Overexpression of the wild-type RPTPα and R223W mutant in HEK293T cells identified that the R223W mutation did not impair RPTPα expression levels or alter its trafficking to the plasma membrane. The R223W mutation did alter proteolytic processing of RPTPα, resulting in the accumulation of a cleavage product. The mutation also resulted in decreased activation of Src family kinases. The functional consequences of this variant, either alone or in concert with the other identified genetic variants, highlights that even minor changes in normal cellular function may increase the risk of developing PD.
... AD is characterized by the pathological aggregation of amyloid- and tau. Amyloid- peptide is generated from the APP by the concerted action of two proteases on the expense of the production of neuroprotective and neurotrophic soluble APP fragment (55). The endocytic receptor sortilin-related receptor SORL1 binds to APP and regulates its intracellular trafficking and amyloidogenic processing (56). ...
Aging is a prominent risk factor for neurodegenerative disorders (NDDs); however, the molecular mechanisms rendering the aged brain particularly susceptible to neurodegeneration remain unclear. Here, we aim to determine the link between physiological aging and NDDs by exploring protein turnover using metabolic labeling and quantitative pulse-SILAC proteomics. By comparing protein lifetimes between physiologically aged and young adult mice, we found that in aged brains protein lifetimes are increased by ~20% and that aging affects distinct pathways linked to NDDs. Specifically, a set of neuroprotective proteins are longer-lived in aged brains, while some mitochondrial proteins linked to neurodegeneration are shorter-lived. Strikingly, we observed a previously unknown alteration in proteostasis that correlates to parsimonious turnover of proteins with high biosynthetic costs, revealing an overall metabolic adaptation that preludes neurodegeneration. Our findings suggest that future therapeutic paradigms, aimed at addressing these metabolic adaptations, might be able to delay NDD onset.
... Aβ peptides are produced through the sequential cleavage of the amyloid precursor protein (APP). APP has several physiological functions including neuronal survival, synaptogenesis, synaptic plasticity, and neuronal excitability [118], and can be cleaved by two different pathways. The first is considered to be non-amyloidogenic and takes place at the cell surface membrane by an α-secretase. ...
Rab GTPases (Rabs) are small proteins that play crucial roles in vesicle transport and membrane trafficking. Owing to their widespread functions in several steps of vesicle trafficking, Rabs have been implicated in the pathogenesis of several disorders, including cancer, diabetes, and multiple neurodegenerative diseases. As treatments for neurodegenerative conditions are currently rather limited, the identification and validation of novel therapeutic targets, such as Rabs, is of great importance. This review summarises proof-of-concept studies, demonstrating that modulation of Rab GTPases in the context of Alzheimer’s disease (AD) can ameliorate disease-related phenotypes, and provides an overview of the current state of the art for the pharmacological targeting of Rabs. Finally, we also discuss the barriers and challenges of therapeutically targeting these small proteins in humans, especially in the context of AD.
... Given that it interacts with several extracellular proteins in common with full-length APP, sAPPα has similar important role in CNS development: it acts as a growth factor (Herzog et al., 2004), and regulates cell proliferation (Guillot-Sestier et al., 2012), neurite growth and synaptogenesis (Milward et al., 1992;Mattson, 1997;Gakhar-Koppole et al., 2008). sAPPα is also involved in synaptic signaling as it facilitates long-term potentiation (LTP) (Ishida et al., 1997) and regulates synaptic activity (Furukawa et al., 1996) through combined interactions with NMDA (Taylor et al., 2008), GABA B R1a (Rice et al., 2019) and K + synaptic receptors (Furukawa et al., 1996). ...
A critical challenge in current research of Alzheimer’s disease (AD) is to clarify the relationship between early neuropathology and network dysfunction. In the present work, the new generation AppNL-FxMAPT double knock in (dKI) model was used to evaluate early stages of AD. The initial step of tau pathology was restricted to the perirhinal-entorhinal region, sparing the hippocampus. This discrete neuropathological sign coincided with deficits in object-place associative memory, one of the earliest recognition memory forms affected in individuals at risk for developing AD. Analyses of task-dependent c-Fos activation were carried out in regions susceptible to early AD pathology, and revealed decreased network efficiency during memory retrieval, likely due to reduced information flow through the retrosplenial cortex. Our results suggest that early perirhinal-entorhinal tau pathology is associated with local hyper-activity which spreads towards connected regions such as the claustrum, the medial prefrontal cortex and ultimately the key retrosplenial hub which is needed to relay information flow from frontal to temporal lobes.
... In addition to its role in cancer, hnRNP C plays an important role in other human diseases. In the nervous system, hnRNP C binds with a 29-nt sequence in the 3′-UTR of amyloid precursor protein (APP) mRNA, whose cleavage product Aβ is highly correlated with degenerative neuropathy, such as Alzheimer's disease, and regulates neuronal synapse growth [84][85][86][87][88]. These results proved its effects in stabilizing and enhancing the translation of mRNA. ...
Heterogeneous nuclear ribonucleoproteins C (HnRNP C) is part of the hnRNP family of RNA-binding proteins. The relationship between hnRNP C and cancers has been extensively studied, and dysregulation of hnRNP C has been found in many cancers. According to existing public data, hnRNP C could promote the maturation of new heterogeneous nuclear RNAs (hnRNA s, also referred to as pre-mRNAs) into mRNAs and could stabilize mRNAs, controlling their translation. This paper reviews the regulation and dysregulation of hnRNP C in cancers. It interacts with some cancer genes and other biological molecules, such as microRNAs (miRNAs), long noncoding RNAs (lncRNAs), and double-stranded RNAs (dsRNAs). Even directly binds to them. The effects of hnRNP C on biological processes such as alternative cleavage and polyadenylation (APA) and N6-methyladenosine (m6A) modification differ among cancers. Its main function is regulating stability and level of translation of cancer genes, and the hnRNP C is regarded as a candidate biomarker and might be valuable for prognosis evaluation.
... It is found in the somatodendritic and axonal compartments of neurons, and in the presynaptic zone, the latter being reached by fast axoplasmic transport. APP is an acute phase protein, promoting survival under metabolically challenging conditions that produce cellular stress, such as ischaemia-hypoxia, traumatic brain injury, and inflammation (Mattson 1997;Finnie et al. 2010;Kogel et al. 2012;Plummer et al. 2016;Hefter and Draguhn 2017). Upregulation of APP in neurotrauma has been found to be due to increased mRNA expression ( Van den Heuvel et al. 1999). ...
The signature pathological feature of the pseudolaminar cerebrocortical necrosis found in polioencephalomalacia (PEM) of ruminants is the development of red (eosinophilic) neurons. These neurons are generally considered to be irredeemably injured but we have shown, for the first time, in ovine PEM cases, that most strongly express amyloid precursor protein (APP), which has a neuroprotective role in the brain. By contrast, neurons in unaffected cerebral cortices from control sheep were APP immunonegative. This finding suggests that, rather than being inevitably destined to die, some of these APP immunoreactive cortical neurons may survive and regain structural and functional integrity.
... The amyloid precursor protein (APP), a type of glycoprotein, is produced by the neuron and has several roles in the development and function of the neuron (Mattson, 1997). APP can undergo cleavage by either non-amyloidogenic processing (α-secretase and γ-secretase) or amyloidogenic processing (βsecretase and γ-secretase). ...
Endocytosis is a process essential to the health and well-being of cell. It is required for the internalisation and sorting of “cargo”—the macromolecules, proteins, receptors and lipids of cell signalling. Clathrin mediated endocytosis (CME) is one of the key processes required for cellular well-being and signalling pathway activation. CME is key role to the recycling of synaptic vesicles [synaptic vesicle recycling (SVR)] in the brain, it is pivotal to signalling across synapses enabling intracellular communication in the sensory and nervous systems. In this review we provide an overview of the general process of CME with a particular focus on two key proteins: clathrin and dynamin that have a central role to play in ensuing successful completion of CME. We examine these two proteins as they are the two endocytotic proteins for which small molecule inhibitors, often of known mechanism of action, have been identified. Inhibition of CME offers the potential to develop therapeutic interventions into conditions involving defects in CME. This review will discuss the roles and the current scope of inhibitors of clathrin and dynamin, providing an insight into how further developments could affect neurological disease treatments.
... Hypoxia drives the metabolism of APP, leading to the amyloidogenic pathway, with Aβ protein as the end product [69]. Mattson [70] reported that this pathway could be a defense mechanism by increasing soluble neuroprotective APPα production. However, hypoxia favors APP metabolism through the amyloidogenic pathway, causing an increase in Aβ levels and not APPα levels [71]. ...
Neurodegenerative diseases (NDs) like Alzheimer’s disease, multiple sclerosis, amyotrophic lateral sclerosis, Parkinson’s disease, and Huntington’s disease predominantly pose a significant socioeconomic burden. Characterized by progressive neural dysfunction coupled with motor or intellectual impairment, the pathogenesis of ND may result from contributions of certain environmental and molecular factors. One such condition is hypoxia, characterized by reduced organ/tissue exposure to oxygen. Reduced oxygen supply often occurs during the pathogenesis of ND and the aging process. Despite the well-established relationship between these two conditions (i.e., hypoxia and ND), the underlying molecular events or mechanisms connecting hypoxia to ND remain ill-defined. However, the relatedness may stem from the protective or deleterious effects of the transcription factor, hypoxia-inducible factor 1-alpha (HIF-1α). The upregulation of HIF-1α occurs in the pathogenesis of most NDs. The dual function of HIF-1α in acting as a “killer factor” or a “protective factor” depends on the prevailing local cellular condition. The kynurenine pathway is a metabolic pathway involved in the oxidative breakdown of tryptophan. It is essential in neurotransmission and immune function and, like hypoxia, associated with ND. Thus, a good understanding of factors, including hypoxia (i.e., the biochemical implication of HIF-1α) and kynurenine pathway activation in NDs, focusing on Alzheimer’s disease could prove beneficial to new therapeutic approaches for this disease, thus the aim of this review.
... Indeed, Aβ production can be avoided by an alternate APP cleavage pathway mediated by the α-secretases in the normal brain [11]. Interestingly, α-secretase cleavage of APP generates a secreted form of APP (sAPP), which has been shown to play a role of neurotrophy and neuroprotection in AD models [12,13]. ...
Amyloid-β (Aβ) accumulating is considered as a causative factor for formation of senile plaque in Alzheimer's disease (AD), but its mechanism is still elusive. The Nicotinamide mononucleotide adenylyltransferase 2 (Nmnat2), a key redox cofactor for energy metabolism, is reduced in AD. Accumulative evidence has shown that the decrease of α-secretase activity, a disintegrin and metalloprotease domain 10 (ADAM10), is responsible for the increase of Aβ productions in AD patient's brain. Here, we observe that the activity of α-secretase ADAM10 and levels of Nmnat2 are significantly decreased, meanwhile there is a simultaneous elevation of Aβ in Tg2576 mice. Over-expression of Nmnat2 increases the mRNA expression of α-secretase ADAM10 and its activity and inhibits Aβ production in N2a/APPswe cells, which can be abolished by Compound C, an AMPK antagonist, suggesting that AMPK is involved in over-expression of Nmnat2 against Aβ production. The further assays demonstrate that Nmnat2 activates AMPK by up-regulating the ratio of NAD+/NADH, moreover AMPK agonist AICAR can also increase ADAM10 activity and reduces Aβ1-40/1-42. Taken together, Nmnat2 suppresses Aβ production and up-regulates ADAM10 in AMPK activity-dependent manner, suggesting that Nmnat2 may serve as a new potential target in arresting AD.
... Aβ1-42 is longer, hydrophobic, and fibrillogenic that is less produced than Aβ1-40. It is the principal species deposited in the brain [12,13] and favors oligomerization and subsequent fibril formation [14,15]. Aβ1-42 is deposited as the main component of senile plaques, and these oligo-mers are collectively produced by the activities of neurons and related astrocytes [16]. ...
Alzheimer's disease (AD) is a neurodegenerative disorder with no clear causative event making the disease difficult to diagnose and treat. The pathological hallmarks of AD include amyloid plaques, neurofibrillary tangles, and widespread neuronal loss. Amyloid-beta has been extensively studied and targeted to develop an effective disease-modifying therapy, but the success rate in clinical practice is minimal. Recently, neuroinflammation has been focused on as the event in AD progression to be targeted for therapies. Various mechanistic pathways including cytokines and chemokines, complement system, oxidative stress, and cyclooxygenase pathways are linked to neuroinflammation in the AD brain. Many cells including microglia, astrocytes, and oligodendrocytes work together to protect the brain from injury. This review is focused to better understand the AD inflammatory and immunoregulatory processes to develop novel anti-inflammatory drugs to slow down the progression of AD.
... In AD, it is well known that amyloid formation can cause neurotoxicity and lead to an increase in oxidative radicals and other free radicals (51). Other potential mechanisms include mitochondria dysfunction and metal homeostasis caused by amyloid formation (52)(53)(54). ...
We report a novel single-cell whole-genome amplification method (LCS-WGA) that can efficiently capture spontaneous DNA damage existing in single cells. We refer to these damage-associated single-nucleotide variants as “damSNVs,” and the whole-genome distribution of damSNVs as the damagenome. We observed that in single human neurons, the damagenome distribution was significantly correlated with three-dimensional genome structures. This nonuniform distribution indicates different degrees of DNA damage effects on different genes. Next, we identified the functionals that were significantly enriched in the high-damage genes. Similar functionals were also enriched in the differentially expressed genes (DEGs) detected by single-cell transcriptome of both Alzheimer’s disease (AD) and autism spectrum disorder (ASD). This result can be explained by the significant enrichment of high-damage genes in the DEGs of neurons for both AD and ASD. The discovery of high-damage genes sheds new lights on the important roles of DNA damage in human diseases and disorders.
Alois Alzheimer described the first patient with Alzheimer’s disease (AD) in 1907 and today AD is the most frequently diagnosed of dementias. AD is a multi-factorial neurodegenerative disorder with familial, life style and comorbidity influences impacting a global population of more than 47 million with a projected escalation by 2050 to exceed 130 million. In the USA the AD demographic encompasses approximately six million individuals, expected to increase to surpass 13 million by 2050, and the antecedent phase of AD, recognized as mild cognitive impairment (MCI), involves nearly 12 million individuals. The economic outlay for the management of AD and AD-related cognitive decline is estimated at approximately 355 billion USD. In addition, the intensifying prevalence of AD cases in countries with modest to intermediate income countries further enhances the urgency for more therapeutically and cost-effective treatments and for improving the quality of life for patients and their families. This narrative review evaluates the pathophysiological basis of AD with an initial focus on the therapeutic efficacy and limitations of the existing drugs that provide symptomatic relief: acetylcholinesterase inhibitors (AChEI) donepezil, galantamine, rivastigmine, and the N-methyl-D-aspartate receptor (NMDA) receptor allosteric modulator, memantine. The hypothesis that amyloid-β (Aβ) and tau are appropriate targets for drugs and have the potential to halt the progress of AD is critically analyzed with a particular focus on clinical trial data with anti-Aβ monoclonal antibodies (MABs), namely, aducanumab, lecanemab and donanemab. This review challenges the dogma that targeting Aβ will benefit the majority of subjects with AD that the anti-Aβ MABs are unlikely to be the “magic bullet”. A comparison of the benefits and disadvantages of the different classes of drugs forms the basis for determining new directions for research and alternative drug targets that are undergoing pre-clinical and clinical assessments. In addition, we discuss and stress the importance of the treatment of the co-morbidities, including hypertension, diabetes, obesity and depression that are known to increase the risk of developing AD.
Background:
Alzheimer's disease (AD) is an age-related neurodegenerative disease that is clinically characterized by progressive cognitive decline. Glucagon-like peptide-1 (GLP-1) is a hormone that belongs to the incretin family and is released in response to nutrient intake. It plays a role in maintaining metabolic homeostasis and has been suggested to be involved in maintaining the brain microenvironment. However, the role of GLP-1 in AD pathogenesis has not been fully illustrated.
Objective:
This study aims to investigate the clinical relevance of GLP-1 in AD and the effects of GLP-1 in amyloid-β (Aβ) metabolism in vitro.
Methods:
In this study, 39 AD patients and 120 cognitively intact controls were included. Plasma levels of GLP-1 were measured using ELISA. SH-SY5Y cells overexpressing human amyloid precursor protein (APP) were treated with GLP-1. Western blot analysis was used to assess the effects of GLP-1 on the metabolism of Aβ.
Results:
Plasma GLP-1 levels were decreased with aging. Plasma GLP-1 levels were lower in AD patients in comparison with healthy older adults. Plasma GLP-1 levels were positively associated with Mini-Mental State Examination scores but negatively associated with plasma pTau181 levels. GLP-1 dose-dependently increased the area fraction of mitochondrial staining in vitro. Furthermore, GLP-1 dose-dependently promoted the α-cleavage of APP, thus reducing the generation of Aβ.
Conclusions:
GLP-1 has neuroprotective effects in AD, and therefore the decrease in GLP-1 levels during aging might contribute to the development of AD.
Purinergic receptors (Rs) of the ATP/ADP, UTP/UDP (P2X, P2Y) and adenosine (A1, A2A)-sensitive classes broadly interfere with cognitive processes both under quasi normal and disease conditions. During neurodegenerative illnesses, high concentrations of ATP are released from the damaged neuronal and non-neuronal cells of the brain; then, this ATP is enzymatically degraded to adenosine. Thus, the primary injury in neurodegenerative diseases appears to be caused by various protein aggregates on which a superimposed damage mediated by especially P2X7 and A2AR activation develops; this can be efficiently prevented by small molecular antagonists in animal models of the above diseases, or are mitigated in the respective knockout mice. Dementia is a leading symptom in Alzheimer's disease (AD), and accompanies Parkinson's disease (PD) and Huntington's disease (HD), especially in the advanced states of these illnesses. Animal experimentation suggests that P2X7 and A2ARs are also involved in a number of psychiatric diseases, such as major depressive disorder (MDD), obsessive compulsive behavior, and attention deficit hyperactivity disorder. In conclusion, small molecular antagonists of purinergic receptors are expected to supply us in the future with pharmaceuticals which are able to combat in a range of neurological/psychiatric diseases the accompanying cognitive deterioration.
Insulin, a well-known hormone, has been implicated as a regulator of blood glucose levels for almost a century now. Over the past few decades, the non-glycemic actions of insulin i.e. neuronal growth and proliferation have been extensively studied. In 2005, Dr. Suzanne de La Monte and her team reported that insulin might be involved in the pathogenesis of Alzheimer's Disease (AD) and thus coined a term "Type-3 diabetes" This hypothesis was supported by several subsequent studies. The nuclear factor erythroid 2- related factor 2 (Nrf2) triggers a cascade of events under the regulation of distinct mechanisms including protein stability, phosphorylation and nuclear cytoplasmic shuttling, finally leading to the protection against oxidative damage. The Nrf2 pathway has been investigated extensively in relevance to neurodegenerative disorders, particularly AD. Many studies have indicated a strong correlation between insulin and Nrf2 signalling pathways both in the periphery and the brainbut merely few of them have focused on elucidating their inter-connective role in AD. The present review emphasizes key molecular pathways that correlate the role of insulin with Nrf2 during AD. The review has also identified key unexplored areas that could be investigated in future to further establish the insulin and Nrf2 influence in AD.
Amyloid-β (Aβ) aggregation and β-amyloid precursor protein cleaving enzyme 1 (BACE1) are the potential therapeutic drug targets for Alzheimer’s disease (AD). A recent study highlighted that tacrine-benzofuran hybrid C1 displayed anti-aggregation activity against Aβ42 peptide and inhibit BACE1 activity. However, the inhibition mechanism of C1 against Aβ42 aggregation and BACE1 activity remains unclear. Thus, molecular dynamics (MD) simulations of Aβ42 monomer and BACE1 with and without C1 were performed to inspect the inhibitory mechanism of C1 against Aβ42 aggregation and BACE1 activity. In addition, a ligand-based virtual screening followed by MD simulations was employed to explore potent new small-molecule dual inhibitors of Aβ42 aggregation and BACE1 activity. MD simulations highlighted that C1 promotes the non aggregating helical conformation in Aβ42 and destabilizes D23–K28 salt bridge that plays a vital role in the self-aggregation of Aβ42. C1 displays a favourable binding free energy (–50.7 ± 7.3 kcal/mol) with Aβ42 monomer and preferentially binds to the central hydrophobic core (CHC) residues. MD simulations highlighted that C1 strongly interacted with the BACE1 active site (Asp32 and Asp228) and active pockets. The scrutiny of interatomic distances among key residues of BACE1 highlighted the close flap (non-active) position in BACE1 on the incorporation of C1. The MD simulations explain the observed high inhibitory activity of C1 against Aβ aggregation and BACE1 in the in vitro studies. The ligand-based virtual screening followed by MD simulations identified CHEMBL2019027 (C2) as a promising dual inhibitor of Aβ42 aggregation and BACE1 activity.
Communicated by Ramaswamy H. Sarma
Cerebral palsy (CP) is part of a group of nonprogressive motor disorders. The disease affects movement and posture and constitutes the most frequent cause of motor disability in childhood. CP is characterized by spasticity, reflecting lesions in the pyramidal pathway. Treatment is currently focused on physical rehabilitation, and the annual progression of the disease is 2-3%. About 60% of these patients present severe degrees of malnutrition associated with dysphagia, gastrointestinal abnormalities, malabsorption, increased metabolism, and depression. These alterations promote sarcopenia functional dependence and affect the quality of life and delay the evolution of motor skills. Currently, there is evidence that the supplementation of several nutrients, dietary correction, and probiotics can improve neurological response by stimulating neuroplasticity, neuroregeneration, neurogenesis, and myelination. This therapeutic strategy could shorten the response period to treatment and increase both gross and fine motor skills. The interaction of nutrients and functional foods integrating a Nutritional Support System (NSS) has shown greater efficiency in neurological stimulation than when nutrients are supplied separately. The most studied elements in the neurological response are glutamine, arginine, zinc, selenium, cholecalciferol, nicotinic acid, thiamine, pyridoxine, folate, cobalamin, Spirulina, omega-3 fatty acids, ascorbic acid, glycine, tryptophan, and probiotics. The NSS represents a therapeutic alternative that will restore neurological function in patients with spasticity and pyramidal pathway lesions, both characteristics of patients with CP.
Alzheimer’s disease (AD) is a neurodegenerative disorder associated with cognitive decline. Despite worldwide efforts to find a cure, no proper treatment has been developed yet, and the only effective countermeasure is to prevent the disease progression by early diagnosis. The reason why new drug candidates fail to show therapeutic effects in clinical studies may be due to misunderstanding the cause of AD. Regarding the cause of AD, the most widely known is the amyloid cascade hypothesis, in which the deposition of amyloid beta and hyperphosphorylated tau is the cause. However, many new hypotheses were suggested. Among them, based on preclinical and clinical evidence supporting a connection between AD and diabetes, insulin resistance has been pointed out as an important factor in the development of AD. Therefore, by reviewing the pathophysiological background of brain metabolic insufficiency and insulin insufficiency leading to AD pathology, we will discuss how can insulin resistance cause AD.
According to clinical studies, the development of Alzheimer’s disease (AD) is linked to the abnormal aggregation of amyloid-β (Aβ) peptides into toxic soluble oligomers, protofibrils as well as mature fibrils....
Alzheimer’s disease (AD) is a common age-related neurodegenerative disease that leads to memory loss and cognitive function damage due to intracerebral neurofibrillary tangles (NFTs) and amyloid-β (Aβ) protein deposition. The phosphoinositide-dependent protein kinase (PDK1)/protein kinase B (Akt) signaling pathway plays a significant role in neuronal differentiation, synaptic plasticity, neuronal survival, and neurotransmission via the axon–dendrite axis. The phosphorylation of PDK1 and Akt rises in the brain, resulting in phosphorylation of the TNF-α-converting enzyme (TACE) at its cytoplasmic tail (the C-terminal end), changing its internalization as well as its trafficking. The current review aimed to explain the mechanisms of the PDK1/Akt/TACE signaling axis that exerts its modulatory effect on AD physiopathology. We provide an overview of the neuropathological features, genetics, Aβ aggregation, Tau protein hyperphosphorylation, neuroinflammation, and aging in the AD brain. Additionally, we summarized the phosphoinositide 3-kinase (PI3K)/PDK1/Akt pathway-related features and its molecular mechanism that is dependent on TACE in the pathogenesis of AD. This study reviewed the relationship between the PDK1/Akt signaling pathway and AD, and discussed the role of PDK1/Akt in resisting neuronal toxicity by suppressing TACE expression in the cell membrane. This work also provides a perspective for developing new therapeutics targeting PDK1/Akt and TACE for the treatment of AD.
Alzheimer’s disease (AD) is a progressive neurodegeneration in vulnerable regions of the hippocampus and cerebral cortex that leads to loss in thinking abilities and dementia. The main pathological features of AD include loss of neurons and synapses, extracellular amyloid plaques composed primarily of amyloid β peptide 1–42, and intracellular neurofibrillary tangles which are mostly abnormally paired helical filaments of hyperphosphorylated tau protein. The precise mechanisms of neuronal cell deaths in AD are yet to be established. The amyloid hypothesis of AD states that the aberrant processing of the β-amyloid precursor protein (APP) and aggregation of amyloid β-peptide 1-42 (Aβ 1-42) plays a central role in neurodegeneration in AD. Normally, APP is cleaved by α-secretase to release an extracellular fragment known as sAPPα. In contrast, Aβ is produced when APP is cleaved by β-secretase (BACE1) and gamma-secretase. Cleavage of APP by β-secretase generates a soluble NH2-terminal fragment (sAPPβ) and a membrane-bound COOH-terminal fragment C99. Cleavage of C99 by gamma-secretase produces Aβ and an APP intracellular domain (AICD). It is generally believed that Aβ 1–42 is neurotoxic while sAPPα confers neuroprotection. Mutations in familial Alzheimer’s disease (FAD) genes, including those in APP and presenilins (which include presenilin-1, PS-1, and presenilin-2, PS-2), are associated with early-onset AD cases. Par-4 (prostate apoptosis response-4) is a cell death-promoting protein that was initially isolated as an apoptosis-associated protein by differential screening for genes upregulated in prostate cancer cells undergoing apoptosis. Par-4 is expressed in neurons, and it is found in both cytoplasmic and nucleus compartments. In this chapter, we provide emerging evidence that Par-4 is involved in neuronal cell death in a variety of neurodegenerative diseases. Specifically, we will focus on the cellular and molecular mechanisms by which Par-4 sensitizes neurons to apoptosis or necroptosis in different experimental models of AD, including the first mouse “knock-in” model of a naturally occurring presenilin-1 mutation responsible for an early-onset form of Alzheimer disease described by our laboratory. Par-4 significantly increases production of the neurotoxic Aβ species while decreases the release of the neuroprotective sAPP by altering cell death signaling, disrupting intracellular calcium homeostasis, and enhancing amyloidogenic processing of APP. Par-4 also interacts with ACID to promote neurodegeneration in AD by regulating AICD-mediated transcriptional activity. Of importance, we identified AATF (apoptosis antagonizing transcription factor), another leucine zipper domain containing protein, to be an endogenous interaction partner and potent inhibitor of Par-4 activity in neurons. AATF confers neuroprotection by interacting with Par-4 via the leucine zipper domain and interfering with binding of Par-4 to AICD. Of importance, AATF is secreted extracellularly by cortical neurons under neurodegenerative conditions, and secreted AATF (sAATF) blocks TLR4-mediated, RIPK3/MLKL-dependent necroptosis of cortical neurons. Surprisingly, a small core peptide from TRL-4 binding region of AATF, termed as SAP-12, provides a much greater neuroprotective potency and broader effective dose range than the full-length sAATF. Participation of Par-4 in other neurodegenerative diseases and neurological disorders as well as future directions is also discussed.
The secreted form of Alzheimer amyloid beta/A4 protein precursor (APP) has been shown to be involved in cell growth regulation (Saitoh, T., Sundsmo, M., Roch, J.-M., Kimura, N., Cole, G., Schubert, D., Oltersdorf, T., and Schenk, D.B. (1989) Cell 58, 615-622). Using a strong prokaryotic expression system, we expressed, in Escherichia coli, peptide fragments covering different regions of the secreted form of APP-695. The longest of these fragments (KB75, 572 amino acids from Val-20 to Ile-591), which contained neither the Kunitz-type protease inhibitor (KPI) domain nor the amyloid beta/A4-protein domain, was purified and shown to be biologically active in terms of growth regulation. Two other APP fragments (KB48, 316 amino acids from Val-20 to Met-335; and RB17, 150 amino acids from Thr-296 to Pro-445), overlapping by only 40 amino acids at a close site C-terminal to the KPI insertion site, were also active. Furthermore, a chemically synthesized 40-residue peptide corresponding to this region of overlap also stimulated the growth of A-1 fibroblasts. These results establish the presence of growth-promoting activity in the secreted form of APP-695 and suggest that the site of this activity of APP-695 lies within a 40-amino acid domain next to the KPI insertion site.
We have analyzed the metabolic pathway of maturation of APP751 in stably transfected 293 cells, in the presence of either of the cysteine protease inhibitors leupeptin or E-64. Metabolic labeling, followed by immunoprecipitation at various times in the chase with a rabbit polyclonal antibody (anti-BX6) specific to the carboxyl-terminal end of amyloid precursor protein (APP), revealed the accumulation of a novel approximately 22-kDa carboxyl-terminal fragment (22-CTF) in the inhibitor-treated cells. This fragment, which was not detectable in untreated cells, was immunoprecipitated by four separate antibodies to the carboxyl-terminal region of APP as well as by polyclonal and monoclonal antibodies specific to the first 16 amino acids of the beta-peptide domain. Antibodies to the amino-terminal end of APP do not, however, recognize the fragment. Co-treatment of the inhibitor-treated cells with either of the lysosomotropic agents chloroquine or ammonium chloride completely blocked the generation of this fragment but did not significantly affect APP maturation or secretion. All, however, slowed the intracellular turnover of the cell-associated, approximately 9-kDa carboxyl-terminal fragment (c-CTF) produced during constitutive secretion. Densitometric analyses of these results suggest that this non-secretory pathway of APP degradation, mediated by cysteine proteases in an intracellular acidic compartment, accounts for approximately 70% of total APP metabolism and that a key processing intermediate in this pathway is a 22-kDa, beta-peptide-containing APP carboxyl-terminal fragment. It is possible that inefficient degradation of such an intermediate leads to the formation of aggregating beta-peptide.
The human beta-amyloid protein is deposited in senile plaques and in the cerebro-vasculature of people with Alzheimer's disease and Down's syndrome. The precise role of beta-amyloid in Alzheimer's disease pathology is presently unknown. To study the properties of beta-amyloid in vivo, we generated transgenic mice that harbor the gene for the carboxyl-terminal 100 amino acids of the human amyloid precursor protein, beginning with the beta-amyloid region, under control of the JC viral early region promoter. The mRNA is expressed exclusively in brain tissue. Further, we demonstrate increased levels of beta-amyloid immunoreactivity on fixed brain tissue. These animals will be useful as a model to study beta-amyloid deposition and its consequences.
The Alzheimer's disease related protein, amyloid beta-protein precursor (A beta PP), contains a domain homologous to Kunitz-type serine protease inhibitors (KPI). The recombinant KPI domain of A beta PP is a potent inhibitor of coagulation factors XIa and IXa and functions as an anticoagulant in vitro. Here we report the expression, purification, and characterization of a reactive center lysine mutant of the KPI domain of A beta PP (KPI-Lys17). An expression plasmid for the KPI-Lys17 domain of A beta PP encoded amino acids 285-345 of the A beta PP cDNA containing a lysine substitution at arginine 17 in the KPI domain. The secreted 61-amino acid product was purified to homogeneity and functionally characterized. The protease inhibitory properties of the KPI-Lys17 domain were compared to those of the native KPI domain of A beta PP. Both KPI domains equally inhibited trypsin, chymotrypsin, and coagulation factors IXa and Xa. However, the KPI-Lys17 domain was an approximately 25-fold less effective inhibitor of coagulation factor XIa resulting in markedly less prolongation of the activated partial thromboplastin time compared to the native KPI domain of A beta PP. On the other hand, the KPI-Lys17 domain was an approximately 10- and 5-fold better inhibitor of plasmin in a chromogenic substrate assay and in a fibrinolytic assay, respectively, than the native KPI domain of A beta PP. Together, these studies suggest that the KPI-Lys17 domain has enhanced anti-fibrinolytic and diminished factor XIa inhibitory properties compared to the native KPI domain of A beta PP.
The effects of the ionotropic glutamate receptor (iGluR) selective agonist N-methyl-D-aspartate (NMDA) on amyloid precursor protein (APP) levels were investigated in primary cultures of rat cerebellar granule cells. Both immunocytochemistry and immunoblotting techniques showed increased APP levels 4 h after a 15 min pulse with NMDA. This effect was completely prevented by incubating the neurones in the presence of the selective metabotropic GluR (mGluR) agonist 1S,3R-ACPD. This phenomenon was related, in terms of doses and time, with the observed 1S,3R-ACPD-mediated protection on NMDA-induced granule cell death. Our findings indicate that APP metabolism is differentially regulated by the stimulation of various GluR subtypes. The GluR-mediated changes in APP content might participate in the control of neuronal viability.
The 4-kDa beta-amyloid protein that forms fibrillar deposits in Alzheimer's diseased brains is derived from a large precursor, the beta-amyloid precursor protein (beta-APP). Recently, it has been reported that beta-amyloid is normally produced and secreted by cultured mammalian cells. In our studies involving recombinant expression of beta-APP, increased yields of beta-amyloid were associated with expression of aberrant beta-APP molecules. Deletion mutations within the beta-amyloid domain, incorrect beta-APP isoform expression in fibroblasts or neuronal cells, or excess amounts of beta-APP all led to increases in beta-amyloid production. Aberrant beta-APP appears to be diverted from the secretory pathway and then degraded to beta-amyloid.
The major pathological change in Alzheimer's disease is the deposition of 39-42-amino acid beta-amyloid peptide (BAP) in the brain. Since BAP begins at the aspartate residue (Asp1, or codon 672 of the amyloid precursor protein (APP)770 transcript), the ability of several proteases to cleave the peptide bond methionine-Asp1 (M/D) was evaluated by using peptides and recombinant APP molecules as substrates. Cathepsin G and chymotrypsin cleave the synthetic peptide HSEVKMDAEF at M/D under acidic conditions, whereas cleavage at lysine-methionine (K/M) predominates when the pH is alkaline. Trypsin and cathepsins B, D, and L are unable to cleave the synthetic peptide at M/D. Peptide SEVNLDAEF, representing the mutation found in early onset Alzheimer's disease families from Sweden, is cleaved by cathepsin G and chymotrypsin at leucine-aspartate (L/D). Incubation of cathepsin G with soluble protease nexin-2 obtained from recombinant APP (APP-REP) derivatives resulted in proteolytic cleavage at or near the amino terminus of BAP. Cathepsin G-mediated cleavage was also observed in the domain representing the amino terminus of BAP when mature plasma membrane-associated APP-REP molecules were used as substrates. Our results strongly suggest the involvement of a chymotrypsin-like serine protease in the generation of the amino terminus of BAP beginning at Asp1.
A formulation of the pathophysiology of dementia of the Alzheimer’s type, particularly that of later onset, proposes that it is a convergence syndrome in which a variety of genetic and environmental abnormalities can contribute to characteristic brain damage. This is consistent with currently available epidemiologic and risk factor data, molecular neuropathologic findings, and data on the brain-behavior relationship in the syndrome. This model can accommodate the typical findings in dementia of the Alzheimer’s type, including mitochondrial damage, selective loss of neurons and synapses, cytoskeletal abnormalities, localized inflammatory reaction, and amyloidosis, implicating loss of synapses as the proximal cause of clinical dementia and emphasizing the mechanistic role of mitochondrial damage in nerve cell damage and metabolic dysfunction. This model implies that different therapies may be directed at different contributing causes of dementia of the Alzheimer’s type in the individual patient and that subgroups of patients can be expected to respond differently to different treatments.
The presence ofprothrontbin, thrombin receptors and thrombin inhibitors in the brain, together with recent evidence that thrombin can affect neuronal outgrowth and survival, suggests that thrombin signaling may be involved in neuronal plasticity and injury. In Alzheimer 's disease, thrombin is associated with plaques comprised largely of amyloid β-peptide (Aβ). Because recent studies have shown that 4β can destabilize neuronal calcium homeostasis, and because thrombin receptors are linked to inositol phospholipid hydrolysis and elevation of [Ca2+]i, we tested the hypothesis that Aβ modifies [Ca2+]i responses to thrombin. Studies using thrombin receptor antibodies and antisense oligodeoxynucleotide technology to suppress expression of thrombin receptors demonstrated that human SH-SY5Y neuroblastoma cells expressed thrombin receptors linked to Ca2+ release from intracellular stores. Relatively short term pretreatment (1 to 3 h) of the SH-SY5Y cells with Aβ25–35 or 4β1–40 resulted in a significant two-to three fold enhancement of thrombin-induced elevation of [Ca2+]i. In contrast, chronic pretreatment with Aβs (8 to 16 h) resulted in an attenuation or complete abrogation of' [Ca2+]i responses to thrombin. Imaging of thiobarbituric acid fluorescence demonstrated that Aβ induced lipid peroxidation, and the effects of both short and long term exposure to 4β on [Ca2+]i responses, were largely abrogated in cultures pretreated with antioxidants. Collectively, these data suggest that Aβ induces lipid peroxidation which impairs thrombin receptor-mediated Ca2+ signaling. Taken together with an increasing amount of data suggesting that thrombin plays roles in neuronal plasticity and neurodegenerative processes, our data suggest that Aβ may induce aberrant thrombin signal transduction which could contribute to the pathogenesis of AD.
Amyloid β-peptide (Aβ) has been shown to activate the classical complement pathway in vitro. Here, we demonstrate that this interaction is fully capable of killing cells and damaging cellular processes in mixed hippocampal cultures from embryonic day 18 rat fetuses. Lactic acid dehydrogenase (LDH) release and morphologic changes were used to evaluate toxicity.
Overproduction or aberrant catabolism of the predicted amyloid β-protein precursor (APP) is suspected as the cause of amyloid deposition in Alzheimer's disease and Down's syndrome brains. For possible in vitro experiments of amyloid formation, we have examined the expression of APP in various cultured cells. We found two types of APP producing cell lines. PC12h (rat pheochromocytoma) and HL-60 (human acute promyelocytic leukemia) cells produce a secretory form that is released into the culture medium, while Bu-17 (human glioma) cells synthesize only a non-secretory form that accumulates at the cell surface. APP immunoreactivity on the latter cells was detected at the tips of cell processes or growth cones. These observations indicate that the nonsecretory form of APP may play a role in cell contact or adhesion.
A heightened production of interleukin 1β(IL-1β) has been reported in microglial-associated amyloid deposits in Alzheimer's disease (AD) brains. These plaques are composed predominantly of β/A4 peptide derived from β-amyloid precursor protein (βAPP). We demonstrate that short-term (1 h) IL-1β-treatment increases βAPPs secretion and concomitantly decreases cell-associated βAPP in human H4 neuroglioma cells. Long-term (5 h) IL-1β treatment did not alter secreted or cell-associated βAPP content. In contrast, the secretion of β/A4-containing epitope was not affected by short-term IL-1β stimulation; however, long-term IL-1β treatment decreased the amount of β/A4-containing epitope secreted from the cells. These results show that IL-1β modifies the processing and secretion of βAPP to exacerbate perhaps the neuropathology of AD.
Cleavage after Met596 of the β-amyloid precursor protein to generate the N-terminus of β-protein indicates the activity of a protease having chymotrypsin-like specificity. A chymotrypsin-like protease is further implicated in Alzheimer's disease by the increased synthesis of the protease inhibitor α1-antichymotrypsin in pathologically affected brain regions and by the presence in the amyloid deposits of inactivated forms of α1-antichymotrypsin (indicating irreversible binding to a target chymotrypsin-like protease). In the present report, we have purified from rat brain a chymotrypsin-like protease that (a) binds with high affinity to human α1-antichymotrypsin, (b) proteolytically generates a β-protein-containing C-terminal fragment from full-length recombinant human β-amyloid precursor protein, and (c) selectively cleaves methoxysuccinyl-Glu-Val-Lys-Met-p-nitroanilide (a substrate modeling the protease recognition domain for the β-protein N-terminal cleavage site). Amino acid sequences of tryptic fragments of the purified rat brain chymotrypsin-like protease indicate an identity with rat mast cell protease I. Moreover, the ontogeny and compartmentalization of rat brain chymotrypsin-like protease are consistent with those of connective tissue-type mast cells in the meningeal and intracortical perivasculature. Because these areas in human brain form extensive β-amyloid deposits in Alzheimer's disease, Down's syndrome, and hereditary cerebral hemorrhage with amyloidosis of Dutch origin, the present findings suggest that a brain mast cell chymotrypsin-like protease may participate in generating perivascular β-protein, which ultimately aggregates into β-amyloid deposits.
Human fetal spinal cords and other non-neural tissues from cases with gestational age from 6 to 21 weeks were examined with a panel of antibodies to different domains of β-amyloid precursor proteins (β-AMPs). In the early developmental stages, the β-APPs were expressed in three distinct layers, i.e., primitive neuroepithelial cell layer, mantle layer and marginal layer. β-APP immunoreactivity was most prominent in cell bodies of putative neuroblasts located in the outer ventral part of the mantle layer. β-APP expression diminished as the spinal cord matured and a weak residual immunoreactivity was detected exclusively in a subset of the anterior horn cells by 21 weeks gestational age. Throughout the gestational ages examined, no convincing β / A4 immunostaining was seen in any of the spinal cord regions. Outside the spinal cord, β-APP immunostaining was consistently present in (1) cell bodies and proximal nerves of immature neurons of dorsal root ganglia and in (2) myotubules, although these cells were devoid of β / A4 immunoreactivity. Western blot analysis of fetal spinal cord revealed immunoreactive bands with apparant molecular weight between 100 and 140 kDa in the membrane-associated fraction, while soluble proteins with a molecular mass centered on 115 kDa were detected in the cytosolic fraction. Our results indicate that: (1) one or more isoforms of full length β-APPs are expressed at very early gestational ages in the developing human spinal cord; (2) the normal metabolism of β-APPs does not result in accumulations of β / A4 fragments.
The amyloid deposited in Alzheimer's disease (AD) is composed primarily of a 39–42 residue polypeptide (βAP) that is derived from a larger β amyloid protein precursor (βAPP). In previous studies, we and others identified full-length, membrane-associated forms of the βAPP and showed that these forms are processed into soluble derivatives that lack the carboxyl-terminus of the full-length forms. In this report, we demonstrate that the soluble ∼125 and ∼105 kDa forms of the βAPP found in human cerebrospinal fluid are specifically labeled by several different antisera to the βAP. This finding indicates that both soluble derivatives contain all or part of the βAP sequence, and it suggests that one or both of these forms may be the immediate precursor of the amyloid deposited in AD.
The role of β-amyloid in Alzheimer's disease and its cellular mechanism of action on neurons are still unclear. There is growing evidence that β-amyloid or its fragment, 25–35, influence neuronal calcium regulation. To investigate the effects of β-amyloid on calcium homeostasis in man we used peripheral human lymphocytes as a model system for central neurons. β-Amyloid fragment 25–35 exposed to lymphocytes for 60 s elevates the phytohemagglutinin (PHA)-induced Ca2+ rise in a dose-dependent manner. Small effects were already seen at concentrations as low as 50 . Similar effects were also observed with fragment 1–40, whereas fragments 1–28 or 12–28 did not affect the Ca2+ response after PHA stimulation. Our findings support the hypothesis of an enhanced calcium response as a general feature of β-amyloid's neurotoxicity. The lymphocyte seems to be a valuable model to study this effect in man.
Temporal lobes from 25 human brains, obtained at random post mortem have been examined for biochemical and morphological changes. On the basis of the histological and autopsy findings and clinical presentation the brains have been subdivided. One group contained 11 neurological specimens; all but one of the patients had at least circumstantial evidence of memory loss, and 6 of these were definitive cases of organic 'senile-type' dementia (that is, senile dementia and mixed senile and vascular dementia). The controls had no histological evidence of 'senile' morphological changes and were matched to the neurological specimens with respect to age, interval between death and autopsy, the agonal state and place of death. Aliquots of homogenates prepared from the entire temporal lobe were assayed for the content of 15 biochemical constituents chosen as potential indices of neural cell type and subcellular structure. The biochemical constituents measured were: 7 enzymes (GAD, AChE, β-galactosidase, phosphohydrolase, succinate dehydrogenase, β-glucuronidase, and carbonic anhydrase activities), total protein and 3 specific proteins (neuronin S-6, neuronin S-5 or 14.3.2, and mitochondrial light fraction protein), 2 classes of brain lipid (ganglioside NANA and galactolipid galactose), and nucleic acids (DNA and RNA). In all cases the intensity of senile plaque formation, neurofibrillary degeneration, neuronal loss, glial increase, cerebrovascular disease, and atrophy was assessed. The biochemical results were expressed per whole lobe. Possible age-related changes in the controls (see preceding paper) and correlations between variables were examined by regression analysis. Differences between the groups were established using Students 't' test. A 'computer technique' was used to reveal possible interrelationships between the variables. On the assumption that the morphological changes in the dements reflect ongoing neuronal loss, it is assumed that the estimated reductions in the total amount of nerve cell constituents are an estimate of total neuronal loss. Furthermore, since the estimate of the loss is based on age-matched comparisons the neuronal loss can be considered as the disease-related loss. By these criteria in senile dementia the mean total disease-related loss was 53%. As far as could be established, the course of the disease in these cases was 7 to 8 yr. Thus, nerve cells appear to have been lost from the lobes at a rate that exceeds the normal ageing rat by perhaps 5% or more per year. In comparison with results for mixed senile and vascular dementia the biochemical changes in senile dementia appear more extensive than would have been predicted from the histopathology. Thus, in non-vascular senile dementia, alterations in the metabolism of neurons may well precede slowly progressive changes in morphology.
The amyloid precursor protein (APP) is a glycoprotein consisting of at least four isoforms derived from a single gene by a process of alternative splicing. The membrane-bound forms of APP have been suggested to have adhesive properties and to mediate neural cell adhesion. Previous studies have demonstrated the ability of Fab' fragments of antibodies to extracellular domains of APP to inhibit neural cell binding to a collagen substrate, suggesting a physiological role for the collagen-binding properties of APP. The binding of APP has been demonstrated to be specific for type IV collagen, and no binding to other extracellular matrix components, including fibronectin and laminin, was detected. The APP-collagen binding appeared to be mediated by a heparin-bridge mechanism, since the binding was abolished by the addition of excess heparan or heparinase. These results were observed by both a homogenate-collagen binding assay and a cell-surface adhesion assay, thus providing further evidence for the adhesion role of APP. They also pose the question of the possible role of the heparin-binding properties of APP in the genesis of the neuritic plaques characteristic of Alzheimer's disease.
The synthetic peptide acetyl-Glu-Val-Lys-Met-Asp-Ala-Glu-Phe-NH2, which spans the cleavage site (Met-Asp) required to generate the N-terminus of the Alzheimer beta-amyloid protein from its precursor, was used to search for human brain peptidases which may be involved in this potentially amyloidogenic process. In both soluble and particulate fractions from human brain the primary cleavage point of the peptide was the Met-Asp bond. Purification and characterization of the activity from the soluble fraction showed that it was metalloendopeptidase 24.15 (EC3.4.24.15). This enzyme is therefore a candidate for the generation of the N-terminus of beta-amyloid protein from its precursor.
The beta-amyloid protein (beta-AP) derived from a beta-amyloid protein precursor (APP) is a hallmark of Alzheimer's disease. The abundant generation of beta-AP suggests the abnormal processing of APP, but the molecular mechanism remains unclear. The main APP-processing enzyme was purified from the rat brain and identified to be a macropain-like multicatalytic proteinase. The purified enzyme cleaved the Gln15-Lys16 bond of beta-AP, but altered to cleave at the N-terminus of beta-AP to release the extracellular domain of beta-AP in the presence of Ca2+. These findings suggest that the functional change in this multicatalytic proteinase may result in abnormal processing of APP.
Treatment of PC12 and C6 cell cultures with recombinant basic fibroblast growth factor results in approximately a five to ten-fold stimulation of beta-amyloid precursor mRNA in the C6 astrocytoma cell line but only a slight induction of precursor mRNA in the PC-12 neuronal cell line. Stimulation of expression occurred at a hormone concentration of approximately 0.5 to 1 nM and was seen after 2 days. These results suggest that basic fibroblast growth factor may contribute to amyloidosis of Alzheimer's disease.
Neurofibrillary tangles (NFT) are abnormal filamentous inclusions that develop in neurons in Alzheimer disease and other disorders. When neurons die, the neurofibrillary tangles that persist in the extracellular space show ultrastructural and antigenic changes. Both intra- and extracellular NFT have recently been shown to contain heparan sulfate proteoglycans (HSPGs). HSPGs are also present in other amyloid deposits in the brain and in systemic amyloidoses. Basic fibroblast growth factor (bFGF) is a heparin binding growth factor which is involved in angiogenesis and also has neurite promoting activity. We now report that bFGF binds avidly to extracellular NFT. Alz-50, a monoclonal antibody (MAb) to an abnormal form of tau and bFGF binding label mutually exclusive subpopulations of neurofibrillary tangles. bFGF binding is abolished by heparinase or heparitinase treatment and therefore is most likely based on the presence of HSPG. Binding of bFGF is a specific and sensitive morphological method to distinguish intra- from extracellular NFT. As intracellular NFT, which also contain HSPGs, are not labeled by bFGF binding, this finding also suggests that HSPGs are modified when the NFT become extracellular.
The amyloid precursor protein (APP) is thought to be processed aberrantly to yield the major constituent of the amyloid plaques observed in the brains of patients with Alzheimer disease and Down syndrome. However, the gene encoding APP is expressed widely in normal human tissues and in adult and fetal mouse tissues and is alternatively spliced in a tissue-specific pattern in the adult. There is evidence that APP may function as a growth factor and as a mediator of cell adhesion and in these roles could be important in morphogenesis. As a step toward determining the role of APP in development and in determining how the adult pattern of tissue-specific splicing is established, we have used reverse transcription and the polymerase chain reaction to demonstrate APP expression in mouse oocytes, preimplantation embryos, and postimplantation embryonic stages to the late embryonic period. All three splicing forms described in mouse were present at each stage, although there were changes in the ratios of the splicing forms at different stages. Screens for APP clones in embryonic cDNA libraries from the egg cylinder stage and the early somite stage were used to confirm the results of the polymerase chain reaction, and APP clone abundance was found to increase 10-fold between the two stages.
One of the major clinical findings in Alzheimer's disease (AD) is the formation of deposits of beta-amyloid protein in amyloid plaques, derived from the beta-amyloid precursor protein (beta-APP). To determine the possible use of beta-APP as a diagnostic marker for AD in CSF, a monoclonal antibody-based immunoassay specific for this protein was developed. The assay does not differentiate between beta-APP695 and beta-APP751 forms but does preferentially recognize beta-APP751 complexed with a protease. Of the two sets of CSF samples tested, one set, obtained from living patients, gave a slightly lower level of beta-APP in AD and Parkinson's disease patients relative to controls, whereas the other set, composed of postmortem samples, showed no significant differences between the AD and control groups.
Two classes of amyloid beta-protein precursors which differ by the presence of a serine protease inhibitor domain have been described. We have used synthetic oligonucleotide probes to investigate the tissue distribution and cellular localization of mRNAs encoding the two classes of amyloid beta-protein precursors. RNA blot analysis showed that transcripts encoding the protease inhibitor sequence are ubiquitously expressed in peripheral and central tissues. By contrast, transcripts lacking the protease inhibitor domain were only found in the central nervous system. By in situ hybridization on cerebral cortex and hippocampal formation both types of transcripts were present exclusively in nerve cells and they appeared to be produced by the same cells. A reduction in the transcript lacking the protease inhibitor domain was observed in frontal cortex from Alzheimer's disease patients. The present results indicate that there exists no correlation between the distribution of amyloid amyloid beta-protein precursor mRNAs and the tissue and cellular pathology of Alzheimer's disease; they also suggest that an overproduction of amyloid beta-protein precursor mRNA is unlikely to be responsible for amyloid beta-protein deposition in Alzheimer's disease.
The biophysical properties of hippocampal membrane preparations from patients with Alzheimer's disease were examined by fluorescence spectroscopy using the membrane lipid probe 1,6-diphenyl-1,3,5-hexatriene (DPH) and its cationic derivative 1-[4-(trimethylamino)-phenyl]-6-phenyl-1,3,5-hexatriene (TMA-DPH). Results of these experiments suggest that Alzheimer's disease is associated with a biophysical alteration in superficial regions of brain cell membranes, as reflected by the mobility of TMA-DPH. In contrast, no change in the mobility of DPH, which preferentially localizes to the hydrocarbon core, was observed. Although a trend was observed for TMA-DPH mobility to parallel histopathologic severity in hippocampal specimens, the biophysical changes did not appear to reflect a loss of neuronal membranes relative to glial membranes or the presence of senile plaques or neurofibrillary tangles.
Electron spin resonance, enzymatic, and SDS-polyacrylamide gel electrophoretic investigations of erythrocyte membranes from patients with Alzheimer's disease were performed. Alterations in the physical state of membrane proteins in Alzheimer's disease erythrocytes were found by spin labeling studies. However, no alterations in membrane lipid fluidity or in the activities of membrane-bound sodium plus potassium-stimulated, magnesium-dependent adenosine triphosphatase or acetylcholinesterase could be demonstrated. Also, no changes in staining profiles of AD erythrocyte membrane proteins subjected to electrophoresis were observed. The altered conformation and/or organization of extraneural membrane proteins in Alzheimer's disease suggests the possibility that this disorder may have more widespread membrane involvement than was originally thought.
1. This study is an attempt to examine in vitro the cyto chemical changes in hippocampal neurones induced by beta-amyloid protein (β-AP).
2. The mechanism of cell death, and the vulnerability of different regions of the hippocampus to b-AP toxicity, has also been explored using TUNEL staining to locate fragmented DNA in both dissociated and organotypic cultures.
3. Apoptotic cell profiles and the detection by immunocytochemistry of ubiquitin and tau protein confirmed the acute neurodegenerative effects of b-AP, and organotypic cultures revealed the dentate gyrus to be especially vulnerable.
4. A scrambled sequence of b-AP, a peptide with similar hydrophobic groups to b-AP, and islet pancreatic amyloidogenic peptide also showed neurodegenerative effects, although less severely than b-AP.
5. It is concluded that organotypic cultures provide a valuable in vitro model with which to observe and characterize the neurotoxic effects of b-AP. These effects, however, may be non-specific and related more to the general amyloidogenicity of the b-AP molecule.
Amyloid beta-protein (A beta) deposits in the white matter were investigated by the double immunohistochemical staining for A beta and neuritic, glial or vascular components. Reactive astroglia and neurite abnormality were absent around A beta deposits in the white matter (w-A beta) even those with a core. The association of w-A beta with blood vessels was not consistent. Aggregates of activated microglia were found to be the sole but a consistent accompaniment of A beta deposits even in the absence of other components such as neuron, synapse, neurite abnormality and reactive astroglia, as observed in the white matter. This suggests that the aggregates of activated microglia most likely represent one of the factors promoting the process of A beta deposition.
Three genetic loci for Alzheimer's disease have been identified. These are the amyloid precursor gene on chromosome 21, a gene for early-onset autosomal dominant Alzheimer's disease on chromosome 14, and the risk-modifying gene APOE on chromosome 19. Additional Alzheimer's disease genes remain to be found. The genes identified by studying inherited forms of Alzheimer's disease are now being used to understand the initiating steps in the pathogenesis of the disease.
beta-amyloid protein (A beta) is produced from amyloid precursor protein (APP) is mainly secreted after cleavage within the beta-amyloid protein (A beta) sequence precluding A beta production. Stimulation of APP secretion inhibits A beta production in some cultured cells, which suggests that the relationship between these two processes is alternate. In this study, we investigated the effect of the inhibition of APP secretion on A beta production using stable transformants of glioma U251 which express the mutant APP695 with a lysine-to-valine substitution at residue 612. Immunoprecipitation analysis showed that the mutant APP695 was secreted much less compared to the wild protein. The respective ratios of the amount of secreted APP to that of cellular APP for the mutant and wild forms were 0.11 and 1.01. A beta production by the mutant APP also was suppressed. The respective ratios of the amount of secreted A beta to that of cellular APP for the mutant and wild forms were 0.022 and 0.13. Both processes of APP secretion and A beta production could be inhibited simultaneously by the mutation, which suggests that they are not always alternate. Immunocytochemistry showed that the mutant APP was not transported to the cell membrane. Both APP secretory processes may require the transportation of APP to the cell membrane.
The heat shock or stress response may play a role in the pathogenesis of Alzheimer's disease. We conducted experiments to visualize microscopically the distribution of wild type amyloid precursor protein (APP) and the behavior of an APP deletion mutant under stress. This was achieved by heat-shock treatment of cells expressing fusion recombinant APP proteins tagged with secreted placental alkaline phosphatase (SEAP). The fusion proteins were cleaved and secreted in a manner similar to wild type APP in unstressed control cells. SEAP activity was detected by cytochemical methods within the cytoplasm in less than 10% of transfected unstressed cells. Heat shocked cells showed a striking difference from the control cells in that over 90% of the stressed cells displayed strong intracytoplasmic SEAP activity occurring with Golgi-like pattern and/or membranous distribution. The effects of heat shock were not due to a peculiar behavior of the clones and depended on the APP portion of the constructs. This study shows miscompartmentalization of APP under stress. Such cellular changes may bear important implications in the processing of APP.
Amyloid beta-protein (A beta) and secreted beta-amyloid precursor protein (sAPP), derived from beta-amyloid precursor protein (APP), are normally released by cultured mammalian cells. We investigated by pulse-chase analysis the secretion kinetics of these two APP derivatives using mouse cholinergic SN49 cell lines stably transfected with mouse APP695 cDNA. After both A beta and sAPP peaked at about the second hour, sAPP decreased with a half-life of approximately 5 hours, but A beta remained almost unchanged for at least 14 hours. These results indicate that A beta is more stable than sAPP in the SN49 conditioned medium.
The influence of serum amyloid P component (SAP) on the survival of rat cerebrocortical cultures was tested. Cytotoxic cell death was examined on 8-9-day-old cell cultures by phase contrast microscopy and quantified by the measurement of lactate dehydrogenase (LDH) leakage. SAP (16-48 nM) evoked a concentration-dependent cell death within 24 h exposure. Our results suggest that SAP, as a constituent of cerebral amyloid deposits, may play a role in the pathomechanism of Alzheimer's disease.
It has been suggested that the vulnerability of the aged brain to Alzheimer's disease (AD) pathogenesis depends on a number of risk factors, including abnormal glycolytic metabolism and beta-amyloid accumulation. Intrahippocampal injections of beta-amyloid and related peptides were administered to chronically hyperglycemic rats to examine beta-amyloid toxicity and the interaction with imbalances of glucose metabolism. Chronic hyperglycemia was induced by systemic injection of streptozotocin (STZ) which selectively destroys pancreatic beta-islet cells. Ten days after intrahippocampal injection of synthetic beta-amyloid peptides (beta 1-42, beta 25-35, scrambled beta 25-35), lesion volume, blood glucose, and plasma corticosterone concentrations, beta 1-42 immunoreactivity and gliosis were assessed to determine peptide toxicity in the normoglycemic and hyperglycemic conditions. Glucose levels correlated with plasma corticosterone concentrations (r = 0.85) and increased lesion volume size (r = 0.36). Intrahippocampal peptide injections in normoglycemic subjects did not induce significant damage as compared to control injections of vehicle alone. STZ-treated groups demonstrated a trend for increased lesion volume size following injection of either vehicle, beta 1-42, or beta 25-35. The combination of the beta 1-42 peptide and streptozotocin-induced hyperglycemia was toxic and induced significantly larger lesions (p < 0.01) of the dorsal blade of the dentate gyrus as compared to injections of beta 1-42 into normoglycemic subjects.
To investigate the role of Na(+)-Ca2+ exchange in the regulation of cytosolic free Ca2+ and the pathogenesis of primary hypertension.
Cytosolic free Ca2+ ([Ca2+]i) in cultured vascular smooth muscle cells from normotensive and spontaneously hypertensive rats of the Münster strain was measured using the fluorescent dye fura-2 after inhibition of Na+,K+ATPase by ouabain and after addition of angiotensin II.
[Ca2+]i showed a rapid increase together with a depolarization of membrane potential as measured by merocyanine 540. The ouabain-induced increase in [Ca2+]i was blocked in Ca(2+)-free medium and by nifedipine, but incubation with the inhibitor of the Na(+)-Ca2+ exchange, NiCl2, did not diminish the effect of ouabain. Likewise, in Na(+)-free medium the response to ouabain was not suppressed. The angiotensin II-induced changes in [Ca2+]i were diminished in Ca(2+)-free medium and by nifedipine, but enhanced by NiCl2.
The increase in [Ca2+]i after Na+,K+ ATPase inhibition is not due to a modulation of Na(+)-Ca2+ exchange, but to a Ca2+ influx through Ca2+ channels. Changes in Na(+)-Ca2+ exchange caused by Na+,K+ ATPase inhibition may not play an important role in vascular smooth muscle cells of spontaneously hypertensive rats.
We conducted immunochemical measurements of soluble amyloid beta-protein precursor (beta PP) in cerebrospinal fluid (CSF) from three monozygous twin pairs. Two of the twin pairs are discordant for Alzheimer's disease and one pair showed concordance for Alzheimer's disease, which was confirmed neuropathologically. All affected individuals displayed substantially lower levels of soluble beta PP in CSF compared with the unaffected individuals. There were no differences in total protein levels in CSF samples from the affected twins compared with those of the unaffected twins. These studies suggest that decreased soluble beta PP in CSF may reflect neuropathological processes in Alzheimer's disease involving beta PP.
Based upon recent evidence that the secreted form of APP can cause the release of cytokines and elicit other biological activities, we sought to identify whether a receptor could be identified on the surface of cells. The secreted amyloid precursor protein containing the Kunitz domain (scAPP751) is identical to protease nexin II, a protease inhibitor which has been shown to form complexes with labeled EGF binding protein that subsequently binds to cells. Results of [125I]scAPP751-trypsin complex incubated with intact fibroblast cells show that the complex appears to bind in a saturable time-dependent and reversible manner. The kinetic constants from the binding studies demonstrate a k1 = 2.5 x 10(7) M-1 s-1 and k2 = 4.7 x 10(-4) s-1 and thus a KD (= k2/k1) = 20 pM. Furthermore, the complex formation of [125I]scAPP751 with a protease appears to be a requirement for optimal binding. The binding affinity of secreted APP demonstrated in this study is consistent with its potency in eliminating a range of biological efforts that have been documented.
The main component of amyloid plaques in Alzheimer's disease (AD) is the beta-amyloid peptide (beta/A4), derived from beta-amyloid precursor proteins (beta-APPs). In order to identify proteases possibly involved in the cleavage at the N-terminal site of beta/A4 a chromogenic peptide corresponding to this region of beta-APP was used. Here the purification and characterization of a new human brain protease with the ability to cleave the beta-APP peptide as well as beta-APP in vitro are described. The enzyme has a molecular mass of 100 kDa and belongs likely to the class of metalloproteases. It should further be named "MP100". The enzyme has a very broad substrate specificity in vitro.
Ionic currents through the N-methyl-D-aspartate (NMDA) receptor channel are modulated by sulfhydryl redox reagents. We report here a novel form of redox modulation that alters NMDA channel kinetics in a voltage-dependent manner. The effects of the thiol reductant dithiothreitol (DTT) and the oxidizing agent 5,5'-dithio-bis(2-nitrobenzoic acid) (DTNB) on NMDA-activated whole-cell currents were examined at various transmembrane voltages in cultured rat cortical neurons. DTT produced a similar level of potentiation of NMDA-induced currents at both -60 mV and +30 mV. However, the reversal of this potentiation by a sulfhydryl-oxidizing agent was dependent on the holding potential, because DTNB decreased the DTT-potentiated NMDA responses more effectively at negative voltages. Interestingly, the NMDA peak current-voltage relationship became substantially outwardly rectifying when sequential DTT/DTNB treatments took place at a positive holding potential, but not under any other circumstances. Single-channel recordings from outside-out patches revealed that this phenomenon was likely produced by a significant and long-lasting 2.3-fold prolongation of the mean open time of NMDA channels at a positive holding potential. Thus, a voltage-dependent chemical alteration in NMDA receptor structure modified the kinetic properties of the associated ion channel.
Rat pheochromocytoma PC12 cells are killed by amyloid beta protein (ABP), a component of plaques and other amyloid deposits in Alzheimer's disease. To investigate the possibility of protection from ABP toxicity by induced heat shock proteins (hsps), PC12 cells were heat treated for 60 min at 42 degrees C. The stress response of the PC12 cells was examined at the protein level using hsp72-specific monoclonal antibodies. Hsp72 immunoreactivity was highly induced following heat treatment. The toxic effect of the beta 25-35 ABP peptide fragment, the major cytotoxic sequence within ABP, was reduced in heat pretreated PC12 cultures. These data indicate a protective role of hsps in neuronal cells exposed to ABP.
Various mechanisms have been identified by which hormones and neurotransmitters interacting with seven transmembrane alpha-helical spanning segments receptors modulate the activity of ion channels. All of the mechanisms involve heterotrimeric G-proteins; the best documented are hormonal modulations of voltage-dependent Ca2+ channels in cardiac, neuronal and endocrine cells. Recent studies using antisense oligonucleotide probes allow the exact identification of the G-proteins involved in these signal transduction pathways.
It has been reported that a discrete peptide fragment of beta-amyloid protein, beta A(25-35), and neuropeptide substance P (SP) possessed sequence homology and could bind to the serine protease inhibitor (serpin) enzyme complex (SEC) receptor. Thus, it has been thought that these peptides and SEC receptor ligand might have similar biological activities. In the present study, we found that C-terminal amidated beta A(25-35)-NH2, SP, and the SEC receptor ligand, Phe-Val-Phe-Leu-Met(FVFLM), could induce an increase in the intracellular free Ca2+ concentration ([Ca2+]i) in neutrophil-like human leukemic (HL-60) cells. Pretreatment with pertussis toxin (PTX) potently inhibited the increase in [Ca2+]i stimulated by these peptides, suggesting that these responses might be mediated by PTX-sensitive G-proteins. Furthermore, we examined the effect on these responses of t-butyloxycarbonyl-methionyl-leucyl-phenylalanine (BocMLF), which is a competitive antagonist of chemotactic peptide N-formyl-methionyl-leucyl-phenylalanine (fMLF) at its receptor. BocMLF scarcely inhibited the [Ca2+]i increase stimulated by beta A(25-35)-NH2. However, the increase in FVFLM-induced [Ca2+]i was potently inhibited by BocMLF. The results suggest that the [Ca2+]i activation of beta A(25-35)-NH2 may have a different mechanism from that of FVFLM in neutrophil-like HL-60 cells, which is not mediated by the SEC-receptor.
Alternative splicing of beta-amyloid precursor protein (APP) RNA generates APP isoforms with or without a Kunitz protease inhibitor (KPI) domain. Previously, we showed that KPI (+) APP RNA, but not KPI (-) APP RNA, is upregulated in response to experimental lesions in which neurotoxicity is dependent on NMDA receptor activation and in Alzheimer's disease hippocampus. Recent studies by Mucke et al. (1995) showed that neuronal expression of human KPI (+) APP, but not KPI (-) APP, in transgenic mice is neuroprotective against experimental lesions. In this study we examined the direct effects of the excitotoxic amino acid Glu on alternatively, spliced APP RNAs and the corresponding protein isoforms in cultured rat cortical neurons. Glu treatment rapidly induced (4.5 h) KPI (+) APP RNA but not KPI (-) APP RNA. Induction of KPI (+) RNA preceded Glu-induced neuronal cell death and was partially blocked by an NMDA-receptor antagonist. In contrast to the RNA, cellular levels of KPI (+) APP were not changed by 4.5 h of Glu treatment. Instead, the cellular full-length form of the protein KPI (-) APP was reduced by approximately 50% after 2 h of Glu treatment and remained depleted after 24 h of treatment. Cellular levels of KPI (+) forms of amyloid precursor-like protein 2 (APLP2) were not changed by Glu treatment. Our data are consistent with the hypothesis that sustained NMDA-receptor activation can regulate alternative splicing of the APP pre-mRNA in neurons.