Expression Of Amyloid-β Precursor Protein Research Articles

APP-CD74 axis mediates endothelial cell-macrophage communication to promote kidney injury and fibrosis.

Kidney injuries often carry a grim prognosis, marked by fibrosis development, renal function loss, and macrophage involvement. Despite extensive research on macrophage polarization and its effects on other cells, like fibroblasts, limited attention has been paid to the influence of non-immune cells on macrophages. This study aims to address this gap by shedding light on the intricate dynamics and diversity of macrophages during renal injury and repair. During the initial research phase, the complexity of intercellular communication in the context of kidney injury was revealed using a publicly available single-cell RNA sequencing library of the unilateral ureteral obstruction (UUO) model. Subsequently, we confirmed our findings using an independent dataset from a renal ischemia-reperfusion injury (IRI) model. We treated two different types of endothelial cells with TGF-β and co-cultured their supernatants with macrophages, establishing an endothelial cell and macrophage co-culture system. We also established a UUO and an IRI mouse model. Western blot analysis, flow cytometry, immunohistochemistry and immunofluorescence staining were used to validate our results at multiple levels. Our analysis revealed significant changes in the heterogeneity of macrophage subsets during both injury processes. Amyloid β precursor protein (APP)-CD74 axis mediated endothelial-macrophage intercellular communication plays a dominant role. In the in vitro co-culture system, TGF-β triggers endothelial APP expression, which subsequently enhances CD74 expression in macrophages. Flow cytometry corroborated these findings. Additionally, APP and CD74 expression were significantly increased in the UUO and IRI mouse models. Immunofluorescence techniques demonstrated the co-localization of F4/80 and CD74 in vivo. Our study unravels a compelling molecular mechanism, elucidating how endothelium-mediated regulation shapes macrophage function during renal repair. The identified APP-CD74 signaling axis emerges as a promising target for optimizing renal recovery post-injury and preventing the progression of chronic kidney disease.

APPlications of amyloid-β precursor protein metabolites in macrocephaly and autism spectrum disorder.

Metabolites of the Amyloid-β precursor protein (APP) proteolysis may underlie brain overgrowth in Autism Spectrum Disorder (ASD). We have found elevated APP metabolites (total APP, secreted (s) APPα, and α-secretase adamalysins in the plasma and brain tissue of children with ASD). In this review, we highlight several lines of evidence supporting APP metabolites' potential contribution to macrocephaly in ASD. First, APP appears early in corticogenesis, placing APP in a prime position to accelerate growth in neurons and glia. APP metabolites are upregulated in neuroinflammation, another potential contributor to excessive brain growth in ASD. APP metabolites appear to directly affect translational signaling pathways, which have been linked to single gene forms of syndromic ASD (Fragile X Syndrome, PTEN, Tuberous Sclerosis Complex). Finally, APP metabolites, and microRNA, which regulates APP expression, may contribute to ASD brain overgrowth, particularly increased white matter, through ERK receptor activation on the PI3K/Akt/mTOR/Rho GTPase pathway, favoring myelination.

Does amyloid β precursor protein gene expression have a role in diagnosis of biliary atresia?

Biliary atresia (BA) is an important cause of surgical jaundice. Although the precise etiology is unknown, β-amyloid (Aβ) has been observed around the bile ducts in BA livers. It is unclear whether Aβ plays a role in the pathogenesis of this disease. This study aims to assess the amyloid β precursor protein (APP) gene expression in infants with BA in comparison with other causes of neonatal cholestasis. This could help explore the role of Aβ in the pathogenesis and diagnosis of BA. A prospective study was conducted at the outpatient clinic of Paediatric Hepatology, Gastroenterology, and Nutrition Department, National Liver Institute, Menoufia University, Shebin El Kom, Menoufia, Egypt during the period March 2022 to December 2022. Clinical data were gathered and laboratory and radiological investigations were conducted including β precursor protein gene expression measured in liver biopsies of the three groups using quantitative real-time PCR (qPCR). Gene expression of APP was considerably higher in the BA group (p = 0.0001) compared to neonatal cholestasis (NC) patients. Gamma glutamyl transferase (GGT) and APP had a positive correlation (p = 0.001). No significant association was found between APP and fibrosis. APP was noticeably higher in BA than NC other than BA. Also, APP in BA was higher (statistically significant, p = 0.0001) than the control. There was no statistically significant difference among NC, BA, and control groups regarding APP (p = 0.07). Both males and females did not show significant differences as regards APP (p = 0.851). Age did not have a statistically significant correlation with APP (p = 0.532). Also, there were no correlations between APP and alkaline phosphatase (ALP), aspartate transaminase (AST), or total bilirubin (TB) (p > 0.05). We concluded that the development and identification of BA may depend on the liver expression of serum APP. Surgeons may be able to carry out early intraoperative cholangiography for BA confirmation if the combination of APP with GGT and other hepatic function parameters exhibits a high predictive potential as a diagnostic test for BA. To evaluate this hypothesis, more research with sizable sample numbers is necessary.

An Adequate Supply of Bis(ethylmaltolato)oxidovanadium(IV) Remarkably Reversed the Pathological Hallmarks of Alzheimer's Disease in Triple-Transgenic Middle-Aged Mice.

Alzheimer's disease (AD) is a complex and progressive neurodegenerative disease with impaired synapse, imbalanced mineral metabolism, protein mis-folding and aggregation. Bis(ethylmaltolato)oxidovanadium(IV) (BEOV), an organic bioactive vanadium compound with low toxicity and high bioavailability, has been studied as therapeutic agent against tuberculosis and diabetes. However, its neuroprotective effects have rarely been reported. Therefore, in this study, the potential application of BEOV in intervening AD cognitive dysfunction and neuropathology was evaluated. Both low- and high-dose of BEOV (0.2mmol/L and 1.0mmol/L) supplementation for 2months improved the spatial learning and memory deficits of the triple-transgenic AD (3 × Tg AD) mice and mitigated the loss of synaptic proteins and synaptic dysfunction. By inhibiting the expression of amyloid-β precursor protein and β-secretase, and the phosphorylation of tau protein at Ser262, Ser396, Ser404, and Ser202/Thr205 residues, BEOV reduced the amyloid-β deposition and neurofibrillary tangle formation in AD mouse brains and primarily cultured neurons. Further analysis of the brain ionome revealed that BEOV supplementation could significantly affect the concentrations of a variety of metals, most of which, including several AD risk metals, showed reduced levels, particularly with a high-dose intake. Additionally, the elemental correlation network identified both conserved and specific elemental correlations, implying a highly complex and dynamic crosstalk between vanadium and other elements during long-term BEOV supplementation. Overall, our results suggest that BEOV is effective in AD intervention via both ameliorating the disease related pathology and regulating metal homeostasis.

Transcriptional and Post-Transcriptional Regulations of Amyloid-β Precursor Protein (APP) mRNA.

Alzheimer’s disease (AD) is an age-associated neurodegenerative disorder characterized by progressive impairment of memory, thinking, behavior, and dementia. Based on ample evidence showing neurotoxicity of amyloid-β (Aβ) aggregates in AD, proteolytically derived from amyloid precursor protein (APP), it has been assumed that misfolding of Aβ plays a crucial role in the AD pathogenesis. Additionally, extra copies of the APP gene caused by chromosomal duplication in patients with Down syndrome can promote AD pathogenesis, indicating the pathological involvement of the APP gene dose in AD. Furthermore, increased APP expression due to locus duplication and promoter mutation of APP has been found in familial AD. Given this background, we aimed to summarize the mechanism underlying the upregulation of APP expression levels from a cutting-edge perspective. We first reviewed the literature relevant to this issue, specifically focusing on the transcriptional regulation of APP by transcription factors that bind to the promoter/enhancer regions. APP expression is also regulated by growth factors, cytokines, and hormone, such as androgen. We further evaluated the possible involvement of post-transcriptional regulators of APP in AD pathogenesis, such as RNA splicing factors. Indeed, alternative splicing isoforms of APP are proposed to be involved in the increased production of Aβ. Moreover, non-coding RNAs, including microRNAs, post-transcriptionally regulate the APP expression. Collectively, elucidation of the novel mechanisms underlying the upregulation of APP would lead to the development of clinical diagnosis and treatment of AD.

Amyloid-β Precursor Protein APP Down-Regulation Alters Actin Cytoskeleton-Interacting Proteins in Endothelial Cells.

The amyloid-β precursor protein (APP) is a ubiquitous membrane protein often associated with Alzheimer’s disease (AD) and cerebral amyloid angiopathy (CAA). Despite its role in the development of the pathogenesis, APP exerts several physiological roles that have been mainly investigated in neuronal tissue. To date, the role of APP in vasculature and endothelial cells has not been fully elucidated. In this study, we used molecular and proteomic approaches to identify and investigate major cellular targets of APP down-regulation in endothelial cells. We found that APP is necessary for endothelial cells proliferation, migration and adhesion. The loss of APP alters focal adhesion stability and cell–cell junctions’ expression. Moreover, APP is necessary to mediate endothelial response to the VEGF-A growth factor. Finally, we document that APP propagates exogenous stimuli and mediates cellular response in endothelial cells by modulating the Scr/FAK signaling pathway. Thus, the intact expression and processing of APP is required for normal endothelial function. The identification of molecular mechanisms responsible for vasoprotective properties of endothelial APP may have an impact on clinical efforts to preserve and protect healthy vasculature in patients at risk of the development of cerebrovascular disease and dementia including AD and CAA.

Supplementation with Nicotinamide Riboside Reduces Brain Inflammation and Improves Cognitive Function in Diabetic Mice.

The purpose of this study is to investigate whether nicotinamide riboside (NR) can improve inflammation and cognitive function in diabetic mice. ICR male mice were fed for 14 weeks with either high-fat chow diet (HF, 60% kcal fat) or standard chow diet (CON, 10% kcal fat). HF, streptozotocin, and nicotinamide were used to induce hyperglycemia. NR or vehicle was delivered via stomach gavage for six weeks. Oral glucose tolerance test, Y-maze test, and nest construction test were conducted before and after the NR treatment period. NR treatment induced down-regulation of NLRP3, ASC, and caspase-1. NR reduced IL-1 expression significantly by 50% in whole brains of hyperglycemic mice. Other inflammatory markers including TNF-α and IL-6 were also attenuated by NR. Brain expression of amyloid-β precursor protein and presenilin 1 were reduced by NR. In addition, NR induced significant reduction of amyloid-β in whole brains of diabetic mice. NR treatment restored hyperglycemia-induced increases in brain karyopyknosis to the levels of controls. Nest construction test showed that NR improved hippocampus functions. Spatial recognition memory and locomotor activity were also improved by NR supplementation. These findings suggest that NR may be useful for treating cognitive impairment by inhibiting amyloidogenesis and neuroinflammation.

Amyloid β precursor protein silencing attenuates epithelial‑mesenchymal transition of nasopharyngeal carcinoma cells via inhibition of the MAPK pathway.

Advances in the treatment of nasopharyngeal carcinoma (NPC) have significantly improved the local control rate; however, distant metastasis remains a principal cause of mortality. Previous studies have demonstrated that the expression levels of amyloid β precursor protein (APP) are increased in NPC. The present study aimed to investigate the association between APP and the development of NPC. In order to knockdown APP expression, an APP-small interfering RNA vector was synthesized and transfected into SUNE-1 cells. Cell Counting Kit-8 assay was performed to assess cell viability. The migratory and invasive abilities of SUNE-1 cells were examined by wound healing and Transwell assays, respectively. Reverse transcription-quantitative polymerase chain reaction and western blotting were performed to measure the mRNA and protein expression levels of APP, and additional factors involved in epithelial-mesenchymal transition (EMT) and in the mitogen-activated protein kinase (MAPK) signaling pathway. APP silencing significantly suppressed cell viability, migration and invasion. In addition, APP interference downregulated the expression levels of metastasis-associated 1, matrix metalloproteinase (MMP)-2 and MMP-9; however, knockdown of APP led to upregulation of tissue inhibitor of metalloproteinases 2 and inhibited EMT. The phosphorylation levels of p38, extracellular signal-regulated kinases 1/2 and c-Jun N-terminal kinases 1/2 were decreased following downregulation of APP. The present results suggested that APP knockdown may significantly inhibit the development of NPC by suppressing cell viability, migration and invasion, and by inhibiting the EMT process via downregulation of the MAPK signaling pathway. Therefore, APP may facilitate the development of a novel gene therapy for the treatment of NPC.

Identification of APP As a Novel Candidate Familial Eosinophilia Predisposition Gene

Eosinophilia, such as familial eosinophilia (FE), idiopathic eosinophilia and reactive eosinophilia, is characterized by a sustained overproduction of peripheral blood eosinophils with or without end-organ damage. Germline variants in cytokine and eosinophil function genes are known to contribute to the development of the disease, suggesting common genetic basis may be shared among FE and sporadic eosinophilia. For example, FE is a rare autosomal dominant disorder with susceptible locus mapped to the chromosome 5q region in one American famliy (Rioux et al., 1998). This region (5q31-33) contains several cytokine genes which play functional roles in the development of eosinophils. Germline mutation in JAK1, which is involved in cytokine signaling, has been found in inherited eosinophilia (Kate et al., 2016). Furthermore, genetic variants in genes associated with eosinophilopoietic function (CCR3,CCL5 etc) have been identified in several idiopathic eosinophilia (Lee et al.,2017). Despite of these findings, most cases of eosinophilia have no identifiable mutations. In this new study, we collected a Chinese three-generation pedigree with documented FE. We aimed to investigate novel genetic variants of FE in this pedigree, and found that APP (amyloid-β precursor protein) might be a novel predisposition gene associated with FE.Firstly, we performed Whole-Genome sequencing analysis on isolated eosinophils from peripheral blood of 3 patients and 3 unaffected family members. Point mutations and small indels were selected as potential pathogenic variants if they met the following criteria: 1. heterozygous mutations 2. not previously described 3. predicted to be deleterious or located in regulatory region. As a result of this analysis, 109 mutated genes were considered as candidates, of which APP was the only gene carrying nonsynonymous mutation in the coding region. APP encodes amyloid-β precursor protein that can generate a number of functional peptides. A deleterious missense mutation (c.C470G, p.T157R) was found in the protein binding domain of APP. This mutation was verified by Sanger sequencing in all 3 FE patients. APP has been shown to regulate gene expression (Kögel et al., 2005). As APP knock-out mouse exhibits decreased mRNA expression levels of several cytokines (IL-6, IL-10 and IL-1β) (Carrano et al., 2015), and IL-1β is associated with eosinophil activation (Esnault et al., 2013), we pursued further on APP as a potential eosinophilia predisposition gene.Secondly, we explored transcriptional changes that may be associated with germline mutated APP in eosinophilia patients. We adopted RNA-seq analysis on isolated eosinophils from peripheral blood of 3 patients and 3 controls. A total of 2035 (up:1249, down:786) differentially expressed genes (DEGs) were identified in sorted patient eosinophils as compared to controls (FDR < 0.01). To functionally classify these genes, we conducted pathway analysis and identified “Lysosome”, “Chemokine signaling”, “Cytokine-cytokine receptor interaction” as significant pathways associated with DEGs. In addition, we found that 51 DEGs were overlapped with previously documented eosinophil-associated genes based on IPA Knowledge Base (N=324). We further determined whether these 51 eosinophil-associated DEGs are interacted with 109 mutated genes by constructing the protein-protein interaction network (PPI). Remarkably, APP was the only gene that was functional linked to 10 eosinophil-associated proteins, including several cytokines (CXCRs, CCRs), to form a sub-network (Score > 0.9). In contrast, randomly picked 51 genes failed to form PPI network. These results indicated that the interactions between APP and eosinophil-associated proteins were potentially of biological significance. Since the analysis of PPI was based on the prior knowledge, we further conducted co-expression analysis on APP and its interaction proteins using our transcriptome data. Interestingly, we found that APP expression was highly correlated with the mRNA level of several eosinophilia-related cytokines including IL-1β, CCR2,CCR3 and CXCR4 (PCC > 0.8).Collectively, our data show that APP is a promising predisposition gene that may contributes to FE as a cytokine modulator. This study furthers our understanding of the genetic basis of familial eosinophilia and other eosinophilia and provides novel therapeutic targets for this heterogenous disease. DisclosuresNo relevant conflicts of interest to declare.

Endothelium-specific amyloid precursor protein deficiency causes endothelial dysfunction in cerebral arteries

The exact physiological function of amyloid-β precursor protein (APP) in endothelial cells is unknown. Endothelium-specific APP-deficient (eAPP−/−) mice were created to gain new insights into the role of APP in the control of vascular endothelial function. Endothelium-dependent relaxations to acetylcholine were significantly impaired in basilar arteries of global APP knockout (APP−/−) and eAPP−/− mice (P < 0.05). In contrast, endothelium-independent relaxations to nitric oxide (NO)-donor diethylamine-NONOate were unchanged. Western blot analysis revealed that protein expression of endothelial nitric oxide synthase (eNOS) was significantly downregulated in large cerebral arteries of APP−/− mice and eAPP−/− mice as compared to respective wild-type littermates (P < 0.05). Furthermore, basal levels of cyclic guanosine monophosphate (cGMP) were also significantly reduced in large cerebral arteries of APP-deficient mice (P < 0.05). In contrast, protein expression of prostacyclin synthase as well as levels of cyclic adenosine monophosphate (cAMP) was not affected by genetic inactivation of APP in endothelial cells. By using siRNA to knockdown APP in cultured human brain microvascular endothelial cells we also found a significant downregulation of eNOS mRNA and protein expressions in APP-deficient endothelium (P < 0.05). These findings indicate that under physiological conditions, expression of APP in cerebral vascular endothelium plays an important protective function by maintaining constitutive expression of eNOS.

Selective Acetyl- and Butyrylcholinesterase Inhibitors Reduce Amyloid-&amp;#946; Ex Vivo Activation of Peripheral Chemo-cytokines From Alzheimer’s Disease Subjects: Exploring the Cholinergic Anti-inflammatory Pathway

Increasing evidence suggests that elevated production and/or reduced clearance of amyloid-β peptide (Aβ) drives the early pathogenesis of Alzheimer's disease (AD). Aβ soluble oligomers trigger a neurotoxic cascade that leads to neuronal dysfunction, neurodegeneration and, ultimately, clinical dementia. Inflammation, both within brain and systemically, together with a deficiency in the neurotransmitter acetylcholine (ACh) that underpinned the development of anticholinesterases for AD symptomatic treatment, are invariable hallmarks of the disease. The inter-relation between Aβ, inflammation and cholinergic signaling is complex, with each feeding back onto the others to drive disease progression. To elucidate these interactions plasma samples and peripheral blood mononuclear cells (PBMCs) were evaluated from healthy controls (HC) and AD patients. Plasma levels of acetylcholinesterase (AChE), butyrylcholinesterase (BuChE) and Aβ were significantly elevated in AD vs. HC subjects, and ACh showed a trend towards reduced levels. Aβ challenge of PBMCs induced a greater release of inflammatory cytokines interleukin-1β (IL-1β), monocyte chemotactic protein-1 (MCP-1) and tumor necrosis factor-alpha (TNF-α) from AD vs. HC subjects, with IL-10 being similarly affected. THP-1 monocytic cells, a cell culture counterpart of PBMCs and brain microglial cells, responded similarly to Aβ as well as to phytohaemagglutinin (PHA) challenge, to allow preliminary analysis of the cellular and molecular pathways underpinning Aβ-induced changes in cytokine expression. As amyloid-β precursor protein expression, and hence Aβ, has been reported regulated by particular cytokines and anticholinesterases, the latter were evaluated on Aβ- and PHA-induced chemocytokine expression. Co-incubation with selective AChE/BuChE inhibitors, (-)-phenserine (AChE) and (-)-cymserine analogues (BuChE), mitigated the rise in cytokine levels and suggest that augmentation of the cholinergic anti-inflammatory pathway may prove valuable in AD.

Oxidative stress induces DNA demethylation and histone acetylation in SH-SY5Y cells: potential epigenetic mechanisms in gene transcription in Aβ production

Overwhelming evidence has suggested that enhanced oxidative stress is involved in the pathogenesis and/or progression of Alzheimer's disease (AD). Amyloid-β (Aβ) that composes senile plaques plays a causal role in AD, and its abnormal deposition in brains is the typical neuropathologic hallmark of AD. Recent studies have suggested that epigenetic mechanisms play an important role in the initiation and development of AD. In the present study, we investigated the epigenetic mechanisms, such as DNA methylation and histone acetylation, involved in the transcription of AD-related genes with Aβ production under oxidative stress. Human neuroblastoma SH-SY5Y cells were treated with hydrogen peroxide (H2O2) and used as the cell model. The intracellular Aβ level was significantly increased in H2O2-treated SH-SY5Y cells. The expression of amyloid-β precursor protein and β-site amyloid-β precursor protein-cleaving enzyme 1 was upregulated by demethylation in the gene promoters associated with the reduction of methyltransferases. Meanwhile, H2O2 induced the upregulation of histone acetyltransferases p300/cAMP-response element binding protein (p300/CBP) and downregulation of histone deacetylases. DNA hypomethylation induced by DNA methyltransferase inhibitor could activate the DNA binding activity of transcription factor nuclear factor-κB, whereas no significant effect was observed on specific protein 1. DNA binding activities of nuclear factor-κB and specific protein 1 were activated by histone hyperacetylation induced by histone deacetylase inhibitor. These findings suggested that oxidative stress resulted in an imbalance between DNA methylation and demethylation and histone acetylation and deacetylation associated with the activation of transcription factors, leading to the AD-related gene transcription in the Aβ overproduction. This could be a potential mechanism for oxidative stress response, which might contribute to the pathogenesis and development of AD.

MicroRNA-153 Physiologically Inhibits Expression of Amyloid-β Precursor Protein in Cultured Human Fetal Brain Cells and Is Dysregulated in a Subset of Alzheimer Disease Patients

Regulation of amyloid-β (Aβ) precursor protein (APP) expression is complex. MicroRNAs (miRNAs) are expected to participate in the molecular network that controls this process. The composition of this network is, however, still undefined. Elucidating the complement of miRNAs that regulate APP expression should reveal novel drug targets capable of modulating Aβ production in AD. Here, we investigated the contribution of miR-153 to this regulatory network. A miR-153 target site within the APP 3'-untranslated region (3'-UTR) was predicted by several bioinformatic algorithms. We found that miR-153 significantly reduced reporter expression when co-transfected with an APP 3'-UTR reporter construct. Mutation of the predicted miR-153 target site eliminated this reporter response. miR-153 delivery in both HeLa cells and primary human fetal brain cultures significantly reduced APP expression. Delivery of a miR-153 antisense inhibitor to human fetal brain cultures significantly elevated APP expression. miR-153 delivery also reduced expression of the APP paralog APLP2. High functional redundancy between APP and APLP2 suggests that miR-153 may target biological pathways in which they both function. Interestingly, in a subset of human AD brain specimens with moderate AD pathology, miR-153 levels were reduced. This same subset also exhibited elevated APP levels relative to control specimens. Therefore, endogenous miR-153 inhibits expression of APP in human neurons by specifically interacting with the APP 3'-UTR. This regulatory interaction may have relevance to AD etiology, where low miR-153 levels may drive increased APP expression in a subset of AD patients.

Pro-inflammatory interleukin-18 increases Alzheimer’s disease-associated amyloid-β production in human neuron-like cells

BackgroundAlzheimer’s disease (AD) involves increased accumulation of amyloid-β (Aβ) plaques and neurofibrillary tangles as well as neuronal loss in various regions of the neocortex. Neuroinflammation is also present, but its role in AD is not fully understood. We previously showed increased levels of pro-inflammatory cytokine interleukin-18 (IL-18) in different regions of AD brains, where it co-localized with Aβ-plaques, as well as the ability of IL-18 to increase expression of glycogen synthase kinase-3β (GSK-3β) and cyclin dependent kinase 5, involved in hyperphosphorylation of tau-protein. Elevated IL-18 has been detected in several risk conditions for AD, including obesity, type-II diabetes, and cardiovascular diseases as well as in stress.MethodsWe differentiated SH-SY5Y neuroblastoma cells as neuron-like and exposed them to IL-18 for various times. We examined the protein levels of amyloid-β precursor protein (APP) and its processing products, its cleaving enzymes, involved in amyloidogenic processing of APP, and markers of apoptosis.ResultsIL-18 increased protein levels of the β-site APP-cleaving enzyme BACE-1, the N-terminal fragment of presenilin-1 and slightly presenilin enhancer 2, both of which are members of the γ-secretase complex, as well as Fe65, which is a binding protein of the C-terminus of APP and one regulator for GSK-3β. IL-18 also increased APP expression and phosphorylation, which preceded increased BACE-1 levels. Further, IL-18 altered APP processing, increasing Aβ40 production in particular, which was inhibited by IL-18 binding protein. Increased levels of soluble APPβ were detected in culture medium after the IL-18 exposure. IL-18 also increased anti-apoptotic bcl-xL levels, which likely counteracted the minor increase of the pro-apoptotic caspase-3. Lactate dehydrogenase activity in culture medium was unaffected.ConclusionsThe IL-18 induction of BACE-1, APP processing, and Aβ is likely to be linked to stress-associated adaptations in neurons during the course of normal functioning and development. However, in the course of wider changes in the aging brain, and particularly in AD, the effects of heightened or prolonged levels of IL-18 may contribute to the process of AD, including via increased Aβ.

Contributions of Brain Insulin Resistance and Deficiency in Amyloid-Related Neurodegeneration in Alzheimerʼs Disease

Alzheimer's disease (AD) is the most common cause of dementia in North America. Growing evidence supports the concept that AD is fundamentally a metabolic disease that results in progressive impairment in the brain's capacity to utilize glucose and respond to insulin and insulin-like growth factor (IGF) stimulation. Moreover, the heterogeneous nature of AD is only partly explained by the brain's propensity to accumulate aberrantly processed, misfolded and aggregated oligomeric structural proteins, including amyloid-β peptides and hyperphosphorylated tau. Evidence suggests that other factors, including impaired energy metabolism, oxidative stress, neuroinflammation, insulin and IGF resistance, and insulin/IGF deficiency in the brain should be incorporated into an overarching hypothesis to develop more realistic diagnostic and therapeutic approaches to AD. In this review, the interrelationship between impaired insulin and IGF signalling and amyloid-β pathology is discussed along with potential therapeutic approaches. Impairments in brain insulin/IGF signalling lead to increased expression of amyloid-β precursor protein (AβPP) and accumulation of AβPP-Aβ. In addition, they promote oxidative stress and deficits in energy metabolism, leading to the activation of pro-AβPP-Aβ-mediated neurodegeneration cascades. Although brain insulin/IGF resistance and deficiency can be induced by primary or secondary disease processes, the soaring rates of peripheral insulin resistance associated with obesity, diabetes mellitus and metabolic syndrome quite likely play major roles in the current AD epidemic. Both clinical and experimental data have linked chronic hyperinsulinaemia to cognitive impairment and neurodegeneration with increased AβPP-Aβ accumulation/reduced clearance in the CNS. Correspondingly, both the restoration of insulin responsiveness and the use of insulin therapy can lead to improved cognitive performance, although with variable effects on brain AβPP-Aβ load. On the other hand, experimental evidence supports the concept that the toxic effects of AβPP-Aβ can promote insulin resistance. Together, these findings suggest that a positive feedback loop of progressive neurodegeneration can develop whereby insulin resistance drives AβPP-Aβ accumulation, and AβPP-Aβ fibril toxicity drives brain insulin resistance. This phenomenon could explain why measuring AβPP-Aβ levels in cerebrospinal fluid or imaging of the brain has proven to be inadequate as a stand-alone biomarker for diagnosing AD, and why the clinical trial results of anti-AβPP-Aβ monotherapy have been disappointing. Instead, the aggregate data suggest that brain insulin resistance and deficiency must also be therapeutically targeted to halt AD progression or reverse its natural course. The positive therapeutic effects of different treatments that address the role of brain insulin/IGF resistance and deficiency, including the use of intranasal insulin delivery, incretins and insulin sensitizer agents are discussed along with potential benefits of lifestyle changes to modify risk for developing mild cognitive impairment or AD. Altogether, the data strongly support the notion that we must shift toward the implementation of multimodal rather than unimodal diagnostic and therapeutic strategies for AD.

Epigenetic mechanisms of amyloid-β production in anisomycin-treated SH-SY5Y cells

Oxidative stress and inflammation as the pathological components of Alzheimer's disease (AD) have been well understood. Among a diversity of mitogen-activated protein kinase (MAPK) family members, JNK and p38 MAPK subfamilies are relevant to the response of environmental stress, inflammatory stimuli, or other insults. Recent studies have demonstrated that epigenetic mechanisms may play a pivotal role in AD pathogenesis and development. In the present study, we have investigated epigenetic mechanisms such as DNA methylation and histone acetylation involved in the activation of stress-related signaling pathways for amyloid-β (Aβ) production. Human neuroblastoma SH-SY5Y cells were treated by anisomycin, an activator of stress-related MAPKs (JNK and p38 MAPK). A significant increase of intracellular Aβ level in anisomycin-treated SH-SY5Y cells was observed. The expression of amyloid-β precursor protein (APP), β-site APP-cleaving enzyme 1 (BACE1), and presenilin 1 (PS1) was upregulated by demethylation in three gene promoters associated with the reduction of methyltransferases (DNMTs). Meanwhile, an enhanced level of global histone H3 acetylation accompanied with upregulation of histone acetyltransferases p300/CREB-binding protein (CBP) and downregulation of histone deacetylases (HDACs) was also observed. These findings indicated that the activation of stress-related signaling pathways could result in the increased transcription of APP, BACE1, and PS1 genes through DNMT-dependent hypomethylation and histone H3 hyperacetylation, thus leading to Aβ overproduction. Moreover, our findings provided a novel insight into epigenetic mechanisms by which oxidative stress contributes to the pathogenesis of AD.

MicroRNA-101 downregulates Alzheimer’s amyloid-β precursor protein levels in human cell cultures and is differentially expressed

The full repertoire of regulatory interactions utilized by human cells to control expression of amyloid-β precursor protein (APP) is still undefined. We investigated here the contribution of microRNA (miRNA) to this regulatory network. Several bioinformatic algorithms predicted miR-101 target sites within the APP 3′-untranslated region (3′-UTR). Using reporter assays, we confirmed that, in human cell cultures, miR-101 significantly reduced the expression of a reporter under control of APP 3′-UTR. Mutation of predicted site 1, but not site 2, eliminated this reporter response. Delivery of miR-101 directly to human HeLa cells significantly reduced APP levels and this effect was eliminated by co-transfection with a miR-101 antisense inhibitor. Delivery of a specific target protector designed to blockade the interaction between miR-101 and its functional target site within APP 3′-UTR enhanced APP levels in HeLa. Therefore, endogenous miR-101 regulates expression of APP in human cells via a specific site located within its 3′-UTR. Finally, we demonstrate that, across a series of human cell lines, highest expression of miR-101 levels was observed in model NT2 neurons.

Divalent metal transporter 1 is involved in amyloid precursor protein processing and Aβ generation

The amyloid-beta precursor protein (APP) and its pathogenic byproduct beta-amyloid peptide (Abeta) play central roles in the pathogenesis of Alzheimer's disease (AD). Reduction in levels of the potentially toxic Abeta is one of the most important therapeutic goals in AD. Recent studies have shown that bivalent metals such as iron, copper, and zinc are involved in APP expression, Abeta deposition, and senile plaque formation in the AD brain. However, the underlying mechanisms involved in abnormal homeostasis of bivalent metals in AD brain remain unclear. In the present study, we found that two isoforms of the divalent metal transporter 1 (DMT1), DMT1-IRE, and DMT1-nonIRE, were colocalized with Abeta in the plaques of postmortem AD brain. Using the APP/PS1 transgenic mouse model, we found that the levels of both DMT1-IRE and DMT1-nonIRE were significantly increased in the cortex and hippocampus compared with wild type-control. We further verified the proposed mechanisms by which DMT1 might be involved in APP processing and Abeta secretion by using the SH-SY5Y cell line stably overexpressing human APP Swedish mutation (APPsw) as a cell model. We found that overexpression of APPsw resulted in increased expression levels of both DMT1-IRE and DMT1-nonIRE in SH-SY5Y cells. Interestingly, silencing of endogenous DMT1 by RNA interference, which reduced bivalent ion influx, led to reductions of APP expression and Abeta secretion. These findings suggest both that DMT1 plays a critical role in ion-mediated neuropathogenesis in AD and that pharmacological blockage of DMT1 may provide novel therapeutic strategies against AD.

Systemic pathology in aged mouse models of Down's syndrome and Alzheimer's disease

Down's syndrome (DS) in humans is caused by trisomy of chromosome 21 (HSA 21). DS patients have a variety of pathologies, including mental retardation and an unusually high incidence of leukemia or lymphoma such as megakaryocytic leukemia. Individuals with DS develop the characteristic neuropathological hallmarks of Alzheimer's disease (AD) in early adulthood, generally by the fourth decade of life. There are several mouse models of DS that have a segmental trisomy of mouse chromosome 16 (MMU 16) with triplicated genes orthologous to HSA 21. These mice display neurodegeneration similar to DS. Although brain pathology in DS models is known, little information is available about other organs. We studied the extraneural pathology in aged DS mice (Ts65Dn, Ts2 and Ts1Cje aged 8 to 24 months) as well as other mouse models of neurodegeneration, including presenilin (PS), amyloid-β precursor protein (APP), and tau (hTau and JNPL) transgenic mice. An increased incidence of peripheral amyloidosis, positive for amyloid A (AA) but not amyloid-β peptide (Aβ), was found in APP over-expressing and tauopathic mice as compared to non-transgenic (ntg) littermates or to DS mouse models. A higher incidence of lymphoma was found in the DS models, including Ts1Cje that is trisomic for a small segment of MMU 16 not including the App gene, but not in the APP over-expressing mice, suggesting that high APP expression is not the cause of lymphoma in DS. The occurrence of lymphomas in mouse DS models is of interest in relation to the increased incidence of malignant conditions in human DS.

Differentially expressed genes in a porcine adult hepatic stem-like cell line and their expression in developing and regenerating liver

To identify differentially expressed genes in adult hepatic stem cells, we performed suppression-subtractive hybridization (SSH) between adult porcine hepatic stem-like cells (HSLCs) and hepatocytes, and the expression of selected genes was assessed in porcine fetal livers and regenerating liver in an 80% hepatectomy model. SSH and subsequent differential screening selected 39 clones that were expressed differentially in HSLCs, including six known genes, 10 unknown genes, one unidentified gene and some chimeric fragments. Four of these genes showed significantly higher expression in HSLCs than in mature hepatocytes: anti-leukoproteinase, matrix Gla protein, amyloid-β precursor protein (APP) and dickkopf-3 (DKK-3). Among them, the mRNA expression of APP and DKK-3 was significantly higher in fifth GW fetal liver than in seventh and thirteenth GW fetal and adult livers, unlike the expression patterns of α-fetoprotein (AFP) or albumin. These mRNAs were detected in the parenchyma of fifth GW fetal liver, whereas in normal adult liver possible expression was limited to the periportal area. On the other hand, immunohistochemistry, Masson's trichrome staining and silver impregnation demonstrated APP and DKK-3 proteins in fifth GW fetal liver in which intralobular bile ducts and hepatic plates had not completely developed. DKK-3 and AFP mRNAs were upregulated on the seventh day (7D) after 80% hepatectomy. In the liver tissue, DKK-3 and AFP proteins were detected in mesenchymal cells in the periportal area and parenchyma, respectively. These data for DKK-3 expression in adult livers suggest the possible presence of adult HSLCs in the periportal area. The pattern of histological staining suggested that 7D liver was in the process of regeneration, showing a character similar to the fifth GW fetal liver. It is speculated that DKK-3 is upregulated in immature and developing livers, and has possible involvement in hepatic differentiation and liver regeneration.