Investigation of anti-Alzheimer's activity of aqueous extract of areca nuts (Areca catechu L): in vitro and in vivo studies / Investigación de la actividad anti-Alzheimer del extracto acuoso de nueces de areca (Areca catechu L): estudios in vitro e in vivo

Alzheimer's disease (AD) is an age-related neurodegenerative disorder. Sever cognitive and memory impairments, huge increase in the prevalence of the disease, and lacking definite cure have absorbed worldwide efforts to develop therapeutic approaches. Since many drugs have failed in the clinical trials due to multifactorial nature of AD, symptomatic treatments are still in the center attention and now, nootropic medicinal plants have been found as versatile ameliorators to reverse memory disorders. In this work, anti-Alzheimer's activity of aqueous extract of areca nuts (Areca catechu L.) was investigated via in vitro and in vivo studies. It depicted good amyloid ß (Aß) aggregation inhibitory activity, 82% at 100 µg/mL. In addition, it inhibited beta-secretase 1 (BACE1) with IC50 value of 19.03 µg/mL. Evaluation of neuroprotectivity of the aqueous extract of the plant against H2O2-induced cell death in PC12 neurons revealed 84.5% protection at 1 µg/mL. It should be noted that according to our results obtained from Morris Water Maze (MWM) test, the extract reversed scopolamine-induced memory deficit in rats at concentrations of 1.5 and 3 mg/kg.

La enfermedad de Alzheimer (EA) es un trastorno neurodegenerativo relacionado con la edad. Los severos deterioros cognitivos y de la memoria, el enorme aumento de la prevalencia de la enfermedad y la falta de una cura definitiva han absorbido los esfuerzos mundiales para desarrollar enfoques terapéuticos. Dado que muchos fármacos han fallado en los ensayos clínicos debido a la naturaleza multifactorial de la EA, los tratamientos sintomáticos siguen siendo el centro de atención y ahora, las plantas medicinales nootrópicas se han encontrado como mejoradores versátiles para revertir los trastornos de la memoria. En este trabajo, se investigó la actividad anti-Alzheimer del extracto acuoso de nueces de areca (Areca catechu L.) mediante estudios in vitro e in vivo. Representaba una buena actividad inhibidora de la agregación de amiloide ß (Aß), 82% a 100 µg/mL. Además, inhibió la beta-secretasa 1 (BACE1) con un valor de CI50 de 19,03 µg/mL. La evaluación de la neuroprotección del extracto acuoso de la planta contra la muerte celular inducida por H2O2 en neuronas PC12 reveló una protección del 84,5% a 1 µg/mL. Cabe señalar que, de acuerdo con nuestros resultados obtenidos de la prueba Morris Water Maze (MWM), el extracto revirtió el déficit de memoria inducido por escopolamina en ratas a concentraciones de 1,5 y 3 mg/kg.

The role of GLP-1/GIP receptor agonists in Alzheimer's disease.

BACKGROUND: New glucagon-like peptide-1 (GLP-1) analogues developed in recent years have a long half-life and offer further prospects for clinical application. At present, the neuroprotection of GLP-1 analogues in Alzheimer's disease (AD) has just begun to be explored. OBJECTIVES: To investigate how glucagon-like peptide-1 (liraglutide) plays a protective role in AD by regulating tau activation and BACE1 expression. MATERIAL AND METHODS: Human neuroblastoma cell line SH-SY5Y cells were cultured in vitro and pretreated with different concentrations of liraglutide, and then treated with different concentrations of okadaic acid (OA) in order to observe the apoptosis of the SH-SY5Y cells. After liraglutide treatment, the apoptosis of neurons in AD rats was detected using flow cytometry, and tau activation and ß-site APP cleaving enzyme 1 (BACE1) expression were detected using western blot. RESULTS: Different concentrations of OA were able to induce apoptosis of SH-SY5Y cells in a dose-dependent manner. Different concentrations of liraglutide were used to pretreat SH-SY5Y cells, which were able to protect the SH-SY5Y cells from apoptosis induced by OA. Okadaic acid significantly increased tau activation and BACE1 expression in the SH-SY5Y cells, which was blocked with liraglutide pretreatment. The results of a water maze experiment showed that liraglutide had significant protective effects on memory and cognitive ability in AD rats induced with OA, inhibited apoptosis of neural cells in AD rats, and inhibited tau activation and BACE1 expression of neural cells in AD rats induced with OA. CONCLUSIONS: Liraglutide has a protective effect on AD in vivo and in vitro, which may be mediated by preventing neuronal apoptosis and inhibiting the activation of tau and the expression of BACE1.

δ-secretase in neurodegenerative diseases: mechanisms, regulators and therapeutic opportunities.

Mammalian asparagine endopeptidase (AEP) is a cysteine protease that cleaves its protein substrates on the C-terminal side of asparagine residues. Converging lines of evidence indicate that AEP may be involved in the pathogenesis of several neurological diseases, including Alzheimer's disease, Parkinson's disease, and frontotemporal dementia. AEP is activated in the aging brain, cleaves amyloid precursor protein (APP) and promotes the production of amyloid-ß (Aß). We renamed AEP to δ-secretase to emphasize its role in APP fragmentation and Aß production. AEP also cleaves other substrates, such as tau, α-synuclein, SET, and TAR DNA-binding protein 43, generating neurotoxic fragments and disturbing their physiological functions. The activity of δ-secretase is tightly regulated at both the transcriptional and posttranslational levels. Here, we review the recent advances in the role of δ-secretase in neurodegenerative diseases, with a focus on its biochemical properties and the transcriptional and posttranslational regulation of its activity, and discuss the clinical implications of δ-secretase as a diagnostic biomarker and therapeutic target for neurodegenerative diseases.

Activation of peroxisome proliferator-activated receptor delta suppresses BACE1 expression by up-regulating SOCS1 in a JAK2/STAT1-dependent manner.

Neuronal expression of beta-secretase 1 (BACE1) has been implicated in the progression of Alzheimer's disease. However, the mechanisms that regulate BACE1 expression are unclear. Here, we show that peroxisome proliferator-activated receptor delta (PPARδ) decreases BACE1 expression by up-regulating suppressor of cytokine signaling 1 (SOCS1) in SH-SY5Y neuroblastoma cells. The activation of PPARδ by GW501516, a specific PPARδ agonist, inhibited expression of BACE1. This effect was abrogated by shRNA-mediated knockdown of PPARδ and by treatment with the PPARδ antagonist GSK0660, indicating that PPARδ is involved in GW501516-mediated suppression of BACE1 expression. On the other hand, GW501516-activated PPARδ induced expression of SOCS1, which is a negative regulator of cytokine signal transduction, at the transcriptional level by binding to a PPAR response element in its promoter. This GW501516-mediated induction of SOCS1 expression led to down-regulation of BACE1 expression via inactivation of signal transducer and activator of transcription 1. GW501516-activated PPARδ suppressed the generation of neurotoxic amyloid beta (Aß) in accordance with the decrease in BACE1 expression. Taken together, these results indicate that PPARδ attenuates BACE1 expression via SOCS1-mediated inhibition of signal transducer and activator of transcription 1 signaling, thereby suppressing BACE1-associated generation of neurotoxic Aß.

Multidisciplinary approaches for targeting the secretase protein family as a therapeutic route for Alzheimer's disease.

The continual increase of the aging population worldwide renders Alzheimer's disease (AD) a global prime concern. Several attempts have been focused on understanding the intricate complexity of the disease's development along with the on- andgoing search for novel therapeutic strategies. Incapability of existing AD drugs to effectively modulate the pathogenesis or to delay the progression of the disease leads to a shift in the paradigm of AD drug discovery. Efforts aimed at identifying AD drugs have mostly focused on the development of disease-modifying agents in which effects are believed to be long lasting. Of particular note, the secretase enzymes, a group of proteases responsible for the metabolism of the ß-amyloid precursor protein (ßAPP) and ß-amyloid (Aß) peptides production, have been underlined for their promising therapeutic potential. This review article attempts to comprehensively cover aspects related to the identification and use of drugs targeting the secretase enzymes. Particularly, the roles of secretases in the pathogenesis of AD and their therapeutic modulation are provided herein. Moreover, an overview of the drug development process and the contribution of computational (in silico) approaches for facilitating successful drug discovery are also highlighted along with examples of relevant computational works. Promising chemical scaffolds, inhibitors, and modulators against each class of secretases are also summarized herein. Additionally, multitarget secretase modulators are also taken into consideration in light of the current growing interest in the polypharmacology of complex diseases. Finally, challenging issues and future outlook relevant to the discovery of drugs targeting secretases are also discussed.

Ganglioside Hp-s1 Analogue Inhibits Amyloidogenic Toxicity in Alzheimer's Disease Model Cells.

Alzheimer's disease (AD) is characterized by extracellular deposition of amyloid plaques, which are predominantly composed of amyloid-ß (Aß) peptide derived from amyloid precursor protein (APP) cleavage. APP interacts with tropomyosin receptor kinase A, a neurotrophic receptor associated with gangliosides and mediating neuronal survival and differentiation through the extracellular signal-regulated protein kinase (ERK) pathway. The ganglioside Hp-s1's analogue Hp-s1A exerts neuritogenic activity; however, its effect on AD pathology remains unknown. To test the hypothesis that Hp-s1A is a potential candidate to treat AD, we established the AD-modeled cell line by expressing human Swedish and Indiana APP gene (APP-Swe/Ind) in N2a mouse neuroblastoma cells. The cells were treated with Hp-s1A or monosialoganglioside GM1 for comparison. The AD model cells expressing APP-Swe/Ind exhibited a significant reduction in viability, as well as neurite outgrowth rate, in comparison to the control cells expressing APP-695. APP C-terminal fragment-ß (CTFß) and Aß42 were increased in the AD cell lysates and the culture media, respectively. With the treatment of either Hp-s1A or GM1 at 1 µM, the AD model cells showed a significant increase in viability; however, only Hp-s1A reduced CTFß levels in these cells. Further analysis of the culture media revealed that Hp-s1A also reduced Aß42 production from AD model cells. The phosphorylation of ERK was elevated and the neurite outgrowth rate was restored with Hp-s1A treatment. In conclusion, the ganglioside analogue Hp-s1A inhibited amyloidogenic processing of APP and promoted neurotrophic activity and survival of AD model cells. Hp-s1A has great potential in AD therapeutic development.

In Silico Identification and Experimental Validation of Novel Anti-Alzheimer's Multitargeted Ligands from a Marine Source Featuring a "2-Aminoimidazole plus Aromatic Group" Scaffold.

Multitargeting or polypharmacological approaches, looking for single chemical entities retaining the ability to bind two or more molecular targets, are a potentially powerful strategy to fight complex, multifactorial pathologies. Unfortunately, the search for multiligand agents is challenging because only a small subset of molecules contained in molecular databases are bioactive and even fewer are active on a preselected set of multiple targets. However, collections of natural compounds feature a significantly higher fraction of bioactive molecules than synthetic ones. In this view, we searched our library of 1175 natural compounds from marine sources for molecules including a 2-aminoimidazole+aromatic group motif, found in known compounds active on single relevant targets for Alzheimer's disease (AD). This identified two molecules, a pseudozoanthoxanthin (1) and a bromo-pyrrole alkaloid (2), which were predicted by a computational approach to possess interesting multitarget profiles on AD target proteins. Biochemical assays experimentally confirmed their biological activities. The two compounds inhibit acetylcholinesterase, butyrylcholinesterase, and ß-secretase enzymes in high- to sub-micromolar range. They are also able to prevent and revert ß-amyloid (Aß) aggregation of both Aß1-40 and Aß1-42 peptides, with 1 being more active than 2. Preliminary in vivo studies suggest that compound 1 is able to restore cholinergic cortico-hippocampal functional connectivity.

Regulation of ADAM10 and ADAM17 by Sorafenib Inhibits Epithelial-to-Mesenchymal Transition in Epstein-Barr Virus-Infected Retinal Pigment Epithelial Cells.

PURPOSE: The a-disintegrin-and-metalloprotease (ADAM) family proteins are widely expressed in the different layers of the retina throughout development. The effect of ADAM proteins on the epithelial-to-mesenchymal transition (EMT) in proliferative vitreoretinopathy (PVR) or AMD is yet to be elucidated. In this study we used Epstein-Barr virus (EBV)-transformed adult retinal pigment epithelial (ARPE) cells to investigate how sorafenib, a multikinase inhibitor, modulates ADAM proteins to control EMT. METHODS: Epithelial to mesenchymal transition and related mechanisms in EBV-infected ARPE cells were determined by RT-PCR, Western blot, invasion assay, ELISA assay, and gene silencing with siRNA. RESULTS: Mesenchymal-like ARPE/EBV cells exhibited considerably increased cellular migration and invasion compared with ARPE cells and produced EMT-related cytokines. Sorafenib significantly inhibited production of TGF-ß1, VEGF, IL-6, IL-8, MCP-1, and TNF-α and blocked the activation of migration-related signaling molecules, such as HIF-1α, p-STAT3, MMP2, and Ang-1. The expression of mature ADAM10, ADAM17, and cleaved Notch 1 proteins in ARPE/EBV cells was downregulated after treatment with sorafenib through the regulatory activity of nardilysin (NRD-1). Gene silencing of NRD-1 in ARPE/EBV cells attenuated secretion of EMT-related cytokines and expression of ADAM10 and 17 and upregulated epithelial markers. CONCLUSIONS: Sorafenib controls the mesenchymal characteristics of EBV-infected ARPE cells. Nardilysin and ADAM family proteins might be new targets for the prevention or control of EMT in retinal diseases.

[(3) H]-L685,458 binding sites are abundant in multiple peripheral organs in rats: implications for safety assessment of putative γ-secretase targeting drugs.

γ-Secretase is a multimeric enzyme complex that carries out proteolytic processing to a variety of cellular proteins. It is currently explored as a therapeutic target for Alzheimer's disease (AD) and cancer. Mechanism-based toxicity needs to be thoroughly evaluated for γ-secretase inhibitory and/or modulatory drugs. This study comparatively assessed putative γ-secretase catalytic sites in rat peripheral tissues relative to brain and explored an effort of its pharmacological inhibition on hair regeneration. Using [(3) H]-labelled L685,458, a potent γ-secretase inhibitor, as probe, we found more abundant presence of γ-secretase binding sites in the liver, gastrointestinal tract, hair follicle, pituitary gland, ovary and testis, as compared to the brain. Local application of L658,458 delayed vibrissal regrowth following whisker removal. These results suggest that γ-secretase may execute important biological functions in many peripheral systems, as in the brain. The development of γ-secretase inhibitors/modulators for AD and cancer therapy should include close monitoring of toxicological panels for hepatic, gastrointestinal, endocrinal and reproductive functions.

Ubiquilin-1 modulates γ-secretase-mediated ε-site cleavage in neuronal cells.

Ubiquilin-1 is an Alzheimer's disease-associated protein, which is known to modulate amyloid precursor protein (APP) processing, amyloid-ß (Aß) secretion, and presenilin-1 (PS1) accumulation. Here, we aim to elucidate the molecular mechanisms by which full-length transcript variant 1 of ubiquilin-1 (TV1) affects APP processing and γ-secretase function in human neuroblastoma cells stably overexpressing APP (SH-SY5Y-APP751). We found that TV1 overexpression significantly increased the level of APP intracellular domain (AICD) generation. However, there was no increase in the levels of secreted Aß40, Aß42, or total Aß, suggesting that ubiquilin-1 in particular enhances γ-secretase-mediated ε-site cleavage. This is supported by the finding that TV1 also significantly increased the level of intracellular domain generation of another γ-secretase substrate, leukocyte common antigen-related (LAR) phosphatase. However, in these cells, the increase in AICD levels was abolished, suggesting a preference of the γ-secretase for LAR over APP. TV2, another ubiquilin-1 variant that lacks the protein fragment encoded by exon 8, did not increase the level of AICD generation like TV1 did. The subcellular and plasma membrane localization of APP or γ-secretase complex components PS1 and nicastrin was not altered in TV1-overexpressing cells. Moreover, the effects of TV1 were not mediated by altered expression or APP binding of FE65, an adaptor protein thought to regulate AICD generation and stability. These data suggest that ubiquilin-1 modulates γ-secretase-mediated ε-site cleavage and thus may play a role in regulating γ-secretase cleavage of various substrates.

Steroids as γ-secretase modulators.

Aggregation and accumulation of Aß42 play an initiating role in Alzheimer's disease (AD); thus, selective lowering of Aß42 by γ-secretase modulators (GSMs) remains a promising approach to AD therapy. Based on evidence suggesting that steroids may influence Aß production, we screened 170 steroids at 10 µM for effects on Aß42 secreted from human APP-overexpressing Chinese hamster ovary cells. Many acidic steroids lowered Aß42, whereas many nonacidic steroids actually raised Aß42. Studies on the more potent compounds showed that Aß42-lowering steroids were bonafide GSMs and Aß42-raising steroids were inverse GSMs. The most potent steroid GSM identified was 5ß-cholanic acid (EC50=5.7 µM; its endogenous analog lithocholic acid was virtually equipotent), and the most potent inverse GSM identified was 4-androsten-3-one-17ß-carboxylic acid ethyl ester (EC50=6.25 µM). In addition, we found that both estrogen and progesterone are weak inverse GSMs with further complex effects on APP processing. These data suggest that certain endogenous steroids may have the potential to act as GSMs and add to the evidence that cholesterol, cholesterol metabolites, and other steroids may play a role in modulating Aß production and thus risk for AD. They also indicate that acidic steroids might serve as potential therapeutic leads for drug optimization/development.

Modulation of γ-secretase by EVP-0015962 reduces amyloid deposition and behavioral deficits in Tg2576 mice.

BACKGROUND: A hallmark of Alzheimer's disease is the presence of senile plaques in human brain primarily containing the amyloid peptides Aß42 and Aß40. Many drug discovery efforts have focused on decreasing the production of Aß42 through γ-secretase inhibition. However, identification of γ-secretase inhibitors has also uncovered mechanism-based side effects. One approach to circumvent these side effects has been modulation of γ-secretase to shift Aß production to favor shorter, less amyloidogenic peptides than Aß42, without affecting the overall cleavage efficiency of the enzyme. This approach, frequently called γ-secretase modulation, appears more promising and has lead to the development of new therapeutic candidates for disease modification in Alzheimer's disease. RESULTS: Here we describe EVP-0015962, a novel small molecule γ-secretase modulator. EVP-0015962 decreased Aß42 in H4 cells (IC50 = 67 nM) and increased the shorter Aß38 by 1.7 fold at the IC50 for lowering of Aß42. AßTotal, as well as other carboxyl-terminal fragments of amyloid precursor protein, were not changed. EVP-0015962 did not cause the accumulation of other γ-secretase substrates, such as the Notch and ephrin A4 receptors, whereas a γ-secretase inhibitor reduced processing of both. A single oral dose of EVP-0015962 (30 mg/kg) decreased Aß42 and did not alter AßTotal peptide levels in a dose-dependent manner in Tg2576 mouse brain at an age when overt Aß deposition was not present. In Tg2576 mice, chronic treatment with EVP-0015962 (20 or 60 mg/kg/day in a food formulation) reduced Aß aggregates, amyloid plaques, inflammatory markers, and cognitive deficits. CONCLUSIONS: EVP-0015962 is orally bioavailable, detected in brain, and a potent, selective γ-secretase modulator in vitro and in vivo. Chronic treatment with EVP-0015962 was well tolerated in mice and lowered the production of Aß42, attenuated memory deficits, and reduced Aß plaque formation and inflammation in Tg2576 transgenic animals. In summary, these data suggest that γ-secretase modulation with EVP-0015962 represents a viable therapeutic alternative for disease modification in Alzheimer's disease.

5-lipoxygenase as an endogenous modulator of amyloid ß formation in vivo.

OBJECTIVE: The 5-lipoxygenase (5-LO) enzymatic pathway is widely distributed within the central nervous system, and is upregulated in Alzheimer's disease. However, the mechanism whereby it may influence the disease pathogenesis remains elusive. METHODS: We evaluated the molecular mechanism by which 5-LO regulates amyloid ß (Aß) formation in vitro and in vivo by pharmacological and genetic approaches. RESULTS: Here we show that 5-LO regulates the formation of Aß by activating the cAMP-response element binding protein (CREB), which in turn increases transcription of the γ-secretase complex. Preventing CREB activation by pharmacologic inhibition or dominant negative mutants blocks the 5-LO-dependent elevation of Aß formation and the increase of γ-secretase mRNA and protein levels. Moreover, 5-LO targeted gene disruption or its in vivo selective pharmacological inhibition results in a significant reduction of Aß, CREB and γ-secretase levels. INTERPRETATION: These data establish a novel functional role for 5-LO in regulating endogenous formation of Aß levels in the central nervous system. Thus, 5-LO pharmacological inhibition may be beneficial in the treatment and prevention of Alzheimer's disease.

Oleic acid ameliorates amyloidosis in cellular and mouse models of Alzheimer's disease.

Several lines of evidence support protective as well as deleterious effects of oleic acid (OA) on Alzheimer's disease (AD) and other neurological disorders; however, the bases of these effects are unclear. Our investigation demonstrates that amyloid precursor protein (APP) 695 transfected Cos-7 cells supplemented with OA have reduced secreted amyloid-beta (Aß) levels. An early-onset AD transgenic mouse model expressing the double-mutant form of human APP, Swedish (K670N/M671L) and Indiana (V717F), corroborated our in vitro findings when they were fed a high-protein, low-fat (18% reduction), cholesterol-free diet enriched with OA. These mice exhibited an increase in Aß40/Aß42 ratio, reduced levels of beta-site APP cleaving enzyme (BACE) and reduced presenilin levels along with reduced amyloid plaques in the brain. The decrease in BACE levels was accompanied by increased levels of a non-amyloidogenic soluble form of APP (sAPPα). Furthermore, the low-fat/+OA diet resulted in an augmentation of insulin-degrading enzyme and insulin-like growth factor-II. These results suggest that OA supplementation and cholesterol intake restriction in a mouse model of AD reduce AD-type neuropathology.

Efficient four-drug cocktail therapy targeting amyloid-ß peptide for Alzheimer's disease.

Cocktail treatment is an effective multidrug medication therapy for some diseases, such as cancer and AIDS, because of the additive or synergistic effect of each medicine and relief from adverse effects. Amyloid-ß peptide (Aß), which is now recognized as central to the development of Alzheimer's disease (AD), is derived from the sequential proteolysis of amyloid precursor protein (APP) by ß- and γ-secretases. Secretase inhibitors are one of most attractive targets for therapeutic intervention in AD. However, because ß- and γ-secretases cleave not only APP but also other substrate proteins, strong inhibition of these secretases leads to severe adverse effects. Some nonsteroidal antiinflammatory drugs (NSAIDs) and cholesterol-lowering drugs (statins) can modify the production of Aß. Here, we report that a cocktail treatment with four drugs (NSAID, statin, and ß- and γ-secretase inhibitors) had additive effects on the reduction of Aß levels in cultured cells without competing with each other. Moreover, the four-drug cocktail treatment caused no changes in processing of the γ-secretase substrate Notch. This is suggests that this cocktail treatment could be a new therapeutic approach for AD.

Nanolipidic particles improve the bioavailability and alpha-secretase inducing ability of epigallocatechin-3-gallate (EGCG) for the treatment of Alzheimer's disease.

Prevention of amyloidogenic processing of amyloid precursor protein with the use of natural phytochemicals capable of enhancing alpha-secretase activity may be a therapeutic approach for treatment of neurodegenerative diseases including Alzheimer's disease (AD) and HIV-associated dementia (HAD). We have recently shown promising preclinical results with the use of green tea polyphenol, (-)-epigallocatechin-3-gallate (EGCG) in mouse models of both diseases, however the translation into clinical use has been problematic primarily as a result of poor bioavailability and inefficient delivery to the central nervous system (CNS). While the antioxidant properties of EGCG are well known, we have shown that it is able to promote non-amyloidogenic processing of amyloid precursor protein (APP) by upregulating alpha-secretase, thus preventing brain beta amyloid plaque formation, a hallmark of AD pathology and common finding in HIV infection. In this preliminary study, we investigated the ability of one preformulation method to improve the oral bioavailability of EGCG. We found that forming nanolipidic EGCG particles improves the neuronal (SweAPP N2a cells) alpha-secretase enhancing ability in vitro by up to 91% (P<001) and it's oral bioavailability in vivo by more than two-fold over free EGCG.

Vascular endothelial growth factor (VEGF) affects processing of amyloid precursor protein and beta-amyloidogenesis in brain slice cultures derived from transgenic Tg2576 mouse brain.

The up-regulation of the angiogenic vascular endothelial growth factor (VEGF) in brains of Alzheimer patients in close relationship to beta-amyloid (Abeta) plaques, suggests a link of VEGF action and processing of the amyloid precursor protein (APP). To reveal whether VEGF may affect APP processing, brain slices derived from 17-month-old transgenic Tg2576 mice were exposed with 1ng/ml VEGF for 6, 24, and 72h, followed by assessing cytosolic and membrane-bound APP expression, level of both soluble and fibrillar Abeta-peptides, as well as activities of alpha- and beta-secretases in brain slice tissue preparations. Treatment of brain slices with VEGF did not significantly affect the expression level of APP, regardless of the exposure time studied. In contrast, VEGF exposure of brain slices for 6h reduced the formation of soluble, SDS extractable Abeta(1-40) and Abeta(1-42) as compared to brain slice cultures incubated in the absence of any drug, while the fibrillar Abeta peptides did not change significantly. This effect was less pronounced 24h after VEGF exposure, but was no longer detectable when brain slices were exposed by VEGF for 72h, which indicates an adaptive response to chronic VEGF exposure. The VEGF-mediated reduction in Abeta formation was accompanied by a transient decrease in beta-secretase activity peaking 6h after VEGF exposure. To reveal whether the VEGF-induced changes in soluble Abeta-level may be due to actions of VEGF on Abeta fibrillogenesis, the fibrillar status of Abeta was examined using the thioflavin-T binding assay. Incubation of Abeta preparations obtained from Tg2576 mouse brain cortex, in the presence of VEGF slightly decreased the fibrillar content with increasing incubation time up to 72h. The data demonstrate that VEGF may affect APP processing, at least in vitro, suggesting a role of VEGF in the pathogenesis of Alzheimer's disease.

ADAM10 as a therapeutic target for cancer and inflammation.

Both cancer and chronic inflammatory diseases are often marked by homeostatic signal transduction pathways run amok. Cleavage of membrane-bound substrates by extracellular metalloproteinases is frequently the rate limiting step in activating many of these pathways, resulting either in liberation of active ligands (shedding) or initiating further processing into bioactive cytoplasmic domains (regulated intramembrane proteolysis or RIP). ADAM10 is a member of the ADAM (A Disintegrin And Metalloproteinase) family of transmembrane metalloproteinases implicated in the RIPing and shedding of dozens of substrates that drive cancer progression and inflammatory disease, including Notch, E-cadherin, EGF, ErbB2 and inflammatory cytokines. ADAM10's emerging role as a significant contributor to these pathologies has led to intense interest in it as a potential drug target for disease treatment. Here we discuss some of the established functions of ADAM10 and the implications of its inhibition in disease progression.

gamma-Secretase in biology and medicine.

gamma-Secretase is a membrane-embedded proteolytic complex composed of presenilin and three other subunits. The gamma-secretase complex generates the amyloid beta-peptide of Alzheimer's disease but also plays important roles in normal physiology, especially in signaling from the Notch receptor. How this hydrolytic enzyme works in a hydrophobic environment is largely unanswered, but mutagenesis and chemical probes have offered insight. gamma-Secretase is an important therapeutic target, although mechanism-based toxicity presents a serious obstacle. Agents that lower amyloid beta-peptide production while leaving important normal functions of gamma-secretase intact are promising therapeutic leads. Inhibition of Notch signaling by gamma-secretase inhibitors, which is undesirable for the prevention or treatment of Alzheimer's disease, may be beneficial for the treatment of a variety of cancers.

1,3-Capryloyl-2-arachidonoyl glycerol activates alpha-secretase activity and suppresses Abeta40 secretion in A172 cells.

Alzheimer's disease (AD) is characterized by the deposition of amyloid beta-peptide (Abeta), derived from amyloid precursor protein (APP). Membrane states, such as lipid components or membrane fluidity, are important for enzymes related to APP processing in meeting their substrates efficiently. We analyzed the effects of triglycerides combined with polyunsaturated fatty acids (PUFAs) and/or caprylic acids on APP proteolysis. Our results showed that 1,3-capryloyl-2-arachidonoyl glycerol (8A8) moderately increased alpha-secretase activity (18%) in A172 cells. beta-Secretase activity was not statistically significantly changed in HEK293 cells stably expressing BACE1. However, Abeta40 secretion decreased by 22%. Thus, we conclude that 8A8 is a useful lipid compound for activating alpha-secretase activity and suppressing Abeta40 secretion.