Amyloid Precursor Protein Synthesis Research Articles

Sequential combined Treatment of Pifithrin-α and Posiphen Enhances Neurogenesis and Functional Recovery After Stroke

Objective: Although cerebral ischemia can activate endogenous reparative processes, such as proliferation of endogenous neural stem cells (NSCs) in the subventricular zone (SVZ) and subgranular zone (SGZ), the majority of these new cells die shortly after injury and do not appropriately differentiate into neurons, or migrate and functionally integrate into the brain. The purpose of this study was to examine a novel strategy for treatment of stroke after injury by optimizing the survival of ischemia-induced endogenous NSCs in the SVZ and SGZ. Methods: Adult SVZ and SGZ NSCs were grown as neurospheres in culture and treated with a p53 inactivator, pifithrin-α (PFT-α), and an amyloid precursor protein (APP)-lowering drug, posiphen, and effects on neurosphere number, size and neuronal differentiation were evaluated. This combined sequential treatment approach was then evaluated in mice challenged with middle cerebral artery occlusion (MCAo). Locomotor behavior and cognition were evaluated at 4 weeks, and the number of new surviving neurons was quantified in nestin creERT2-YFP mice. Results: PFT-α and posiphen enhanced the self-renewal, proliferation rate and neuronal differentiation of adult SVZ and SGZ NSCs in culture. Their sequential combination in mice challenged with MCAo-induced stroke mitigated locomotor and cognitive impairments and increased the survival of SVZ and SGZ NSCs cells. PFT-α and the combined posiphen+PFT-α treatment similarly improved locomotion behavior in stroke challenged mice. Notably, however, the combined treatment provided significantly more potent cognitive function enhancement in stroke mice, as compared with PFT-α single treatment. Interpretation: Delayed combined sequential treatment with an inhibitor of p53 dependent apoptosis (PFT-α) and APP synthesis (posiphen) proved able to enhance stroke-induced endogenous neurogenesis and improve the functional recovery in stroke animals. Whereas the combined sequential treatment provided no further improvement in locomotor function, as compared with PFT-α alone treatment, suggesting a potential ceiling in the locomotion behavioral outcome in stroke animals, combined treatment more potently augmented cognitive function recovery after stroke.

Alzheimer-related decrease in CYFIP2 links amyloid production to tau hyperphosphorylation and memory loss.

Characteristic features of Alzheimer's disease are memory loss, plaques resulting from abnormal processing of amyloid precursor protein (APP), and presence of neurofibrillary tangles and dystrophic neurites containing hyperphosphorylated tau. Currently, it is not known what links these abnormalities together. Cytoplasmic FMR1 interacting protein 2 (CYFIP2) has been suggested to regulate mRNA translation at synapses and this may include local synthesis of APP and alpha-calcium/calmodulin-dependent kinase II, a kinase that can phosphorylate tau. Further, CYFIP2 is part of the Wiskott-Aldrich syndrome protein-family verprolin-homologous protein complex, which has been implicated in actin polymerization at synapses, a process thought to be required for memory formation. Our previous studies on p25 dysregulation put forward the hypothesis that CYFIP2 expression is reduced in Alzheimer's disease and that this contributes to memory impairment, abnormal APP processing and tau hyperphosphorylation. Here, we tested this hypothesis. First, in post-mortem tissue CYFIP2 expression was reduced by ∼50% in severe Alzheimer's hippocampus and superior temporal gyrus when normalized to expression of a neuronal or synaptic marker protein. Interestingly, there was also a trend for decreased expression in mild Alzheimer's disease hippocampus. Second, CYFIP2 expression was reduced in old but not in young Tg2576 mice, a model of familial Alzheimer's disease. Finally, we tested the direct impact of reduced CYFIP2 expression in heterozygous null mutant mice. We found that in hippocampus this reduced expression causes an increase in APP and β-site amyloid precursor protein cleaving enzyme 1 (BACE1) protein, but not mRNA expression, and elevates production of amyloid-β42 Reduced CYFIP2 expression also increases alpha-calcium/calmodulin-dependent kinase II protein expression, and this is associated with hyperphosphorylation of tau at serine-214. The reduced expression also impairs spine maturity without affecting spine density in apical dendrites of CA1 pyramidal neurons. Furthermore, the reduced expression prevents retention of spatial memory in the water maze. Taken together, our findings indicate that reduced CYFIP2 expression triggers a cascade of change towards Alzheimer's disease, including amyloid production, tau hyperphosphorylation and memory loss. We therefore suggest that CYFIP2 could be a potential hub for targeting treatment of the disease.

A role for amyloid precursor protein translation to restore iron homeostasis and ameliorate lead (Pb) neurotoxicity.

Iron supplementation ameliorates the neurotoxicity of the environmental contaminant lead (Pb); however, the mechanism remains undefined. Iron is an essential nutrient but high levels are toxic due to the catalytic generation of destructive hydroxyl radicals. Using human neuroblastoma SH-SY5Y cells to model human neurons, we investigated the effect of Pb on proteins of iron homeostasis: the Alzheimer's amyloid precursor protein (APP), which stabilizes the iron exporter ferroportin 1; and, the heavy subunit of the iron-storage protein, ferritin (FTH). Lead (Pb(II) and Pb(IV) inhibited APP translation and raised cytosolic iron(II). Lead also increased iron regulatory protein-1 binding to the cognate 5'untranslated region-specific iron-responsive element (IRE) of APP and FTH mRNAs. Concurrent iron treatment rescued cells from Pb toxicity by specifically restoring APP synthesis, i.e. levels of the APP-related protein, APLP-2, were unchanged. Significantly, iron/IRE-independent over-expression of APP695 protected SH-SY5Y cells from Pb toxicity, demonstrating that APP plays a key role in maintaining safe levels of intracellular iron. Overall, our data support a model of neurotoxicity where Pb enhances iron regulatory protein/IRE-mediated repression of APP and FTH translation. We propose novel treatment options for Pb poisoning to include chelators and the use of small molecules to maintain APP and FTH translation. We propose the following cascade for Lead (Pb) toxicity to neurons; by targeting the interaction between Iron regulatory protein-1 and Iron-responsive elements, Pb caused translational repression of proteins that control intracellular iron homeostasis, including the Alzheimer's amyloid precursor protein (APP) that stabilizes the iron exporter ferroportin, and the ferroxidase heavy subunit of the iron-storage protein, ferritin. When unregulated, IRE-independent over-expression of APP695 protected SH-SY5Y neurons from Pb toxicity. There is a novel and key role for APP in maintaining safe levels of intracellular iron pertinent to lead toxicity.

Effects and possible mechanisms of action of acacetin on the behavior and eye morphology of Drosophila models of Alzheimer's disease.

The human β-amyloid (Aβ) cleaving enzyme (BACE-1) is a target for Alzheimer’s disease (AD) treatments. This study was conducted to determine if acacetin extracted from the whole Agastache rugosa plant had anti-BACE-1 and behavioral activities in Drosophila melanogaster AD models and to determine acacetin’s mechanism of action. Acacetin (100, 300, and 500 μM) rescued amyloid precursor protein (APP)/BACE1-expressing flies and kept them from developing both eye morphology (dark deposits, ommatidial collapse and fusion, and the absence of ommatidial bristles) and behavioral (motor abnormalities) defects. The reverse transcription polymerase chain reaction analysis revealed that acacetin reduced both the human APP and BACE-1 mRNA levels in the transgenic flies, suggesting that it plays an important role in the transcriptional regulation of human BACE-1 and APP. Western blot analysis revealed that acacetin reduced Aβ production by interfering with BACE-1 activity and APP synthesis, resulting in a decrease in the levels of the APP carboxy-terminal fragments and the APP intracellular domain. Therefore, the protective effect of acacetin on Aβ production is mediated by transcriptional regulation of BACE-1 and APP, resulting in decreased APP protein expression and BACE-1 activity. Acacetin also inhibited APP synthesis, resulting in a decrease in the number of amyloid plaques.

Evaluating the role of hnRNP-C and FMRP in the cAMP-induced APP metabolism.

Cyclic adenosine monophosphate (cAMP) modulates synaptic plasticity and memory and manipulation of the cAMP/protein kinase A/cAMP responsive element binding protein pathway significantly affects cognitive functions. Notably, cAMP can increase the expression of the amyloid precursor protein (APP), whose proteolytic processing gives rise to amyloid beta (Aβ) peptides. Despite playing a pathogenic role in Alzheimer's disease, physiological concentrations of Aβ are necessary for the cAMP-mediated regulation of long-term potentiation, supporting the existence of a novel cAMP/APP/Aβ cascade with a crucial role in memory formation. However, the molecular mechanisms by which cAMP stimulates APP expression and Aβ production remain unclear. Here, we investigated whether hnRNP-C and FMRP, two RNA-binding proteins largely involved in the expression of APP, are the cAMP effectors inducing the protein synthesis of APP. Using RNA immunoprecipitation and RNA-silencing approaches, we found that neither hnRNP-C nor FMRP is required for cAMP to stimulate APP and Aβ production.

Blocking IRES-mediated translation pathway as a new method to treat Alzheimer’s disease

Scientists theorized that β-amyloid (Aβ) plaques and tau tangles are involved in the development of Alzheimer’s disease (AD), and amyloid precursor protein (APP) produces Aβ to trigger the disease process. However, the normal synaptic function of APP itself is not fully understood. Several findings cast APP as a potential key player in learning and memory under normal condition. Nevertheless, the regular operation of APP will be disrupted by abnormal accumulation of Aβ under cellular pathological conditions. Herein, there is a hypothesis that AD could be treated by attenuating APP synthesis during cellular pathophysiological stress. In virtue of a previous study, it was speculated that cells could not decrease APP synthesis via self-protection maybe because APP is synthesized via internal ribosome entry segment (IRES)-mediated translation. Consequently, the blockage of this translation might be a new inoffensive and high-level specificity treatment.

A novel mechanism for cyclic adenosine monophosphate–mediated memory formation: Role of amyloid beta

Cyclic adenosine monophosphate (cAMP) regulates long-term potentiation (LTP) and ameliorates memory in healthy and diseased brain. Increasing evidence shows that, under physiological conditions, low concentrations of amyloid β (Aβ) are necessary for LTP expression and memory formation. Here, we report that cAMP controls amyloid precursor protein (APP) translation and Aβ levels, and that the modulatory effects of cAMP on LTP occur through the stimulation of APP synthesis and Aβ production.

Improved Neuroprotective Effects by CombiningBacopa monnieriandRosmarinus officinalisSupercritical CO2Extracts

Ethnobotanical evidence suggests that herbs such as brahmi (Bacopa monnieri) and rosemary (Rosmarinus officinalis) may possess antioxidant and neuroprotective properties. We compared the antioxidant and neuroprotective effects of supercritical extract of Bacopa monnieri and rosemary antioxidant extract obtained from Rosmarinus officinalis as well as their combination to examine the effects on human glial (U-87 MG) and embryonic mouse hypothalamus cells. Bacopa monnieri extract, rosemary antioxidant extract, and their combination (1:1) are not cytotoxic in both glial and embryonic mouse hypothalamus cell lines up to 200 μg/mL concentration. The combination of extracts of Bacopa monnieri + rosemary antioxidant has better antioxidant potential and antilipid peroxidation activity than either agent alone. Although the extract of Bacopa monnieri + rosemary antioxidant showed almost similar inhibition of phospho tau expression as Bacopa monnieri or rosemary antioxidant extract alone, the combination has better inhibitory effect on amyloid precursor protein synthesis and higher brain-derived neurotrophic factor production in hypothalamus cells than single agents. These results suggest that the extract of Bacopa monnieri + rosemary antioxidant is more neuroprotective than Bacopa monnieri or rosemary antioxidant extract.

Synthesis of the Alzheimer Drug Posiphen into its Primary Metabolic Products (+)-N1-norPosiphen, (+)-N8-norPosiphen and (+)-N1, N8-bisnorPosiphen, their Inhibition of Amyloid Precursor Protein, α -Synuclein Synthesis, Interleukin-1β Release, and Cholinergic Action.

A major pathological hallmark of Alzheimer disease (AD) is the appearance in the brain of senile plaques that are primarily composed of aggregated forms of β-amyloid peptide (Aβ) that derive from amyloid precursor protein (APP). Posiphen (1) tartrate is an experimental AD drug in current clinical trials that reduces Aβ levels by lowering the rate of APP synthesis without toxicity. To support the clinical development of Posiphen (1) and elucidate its efficacy, its three major metabolic products, (+)-N1-norPosiphen (15), (+)-N8-norPosiphen (17) and (+)-N1, N8-bisnorPosiphen (11), were required in high chemical and optical purity. The efficient transformation of Posiphen (1) into these metabolic products, 15, 17 and 11, is described. The biological activity of these metabolites together with Posiphen (1) and its enantiomer, the AD drug candidate (-)-phenserine (2), was assessed against APP,α-synuclein and classical cholinergic targets. All the compounds potently inhibited the generation of APP and α-synuclein in neuronal cultures. In contrast, metabolites 11 and 15, and (-)-phenserine (2) but not Posiphen (1) or 17, possessed acetyl cholinesterase inhibitory action and no compounds bound either nicotinic or muscarinic receptors. As Posiphen (1) lowered CSF markers of inflammation in a recent clinical trial, the actions of 1 and 2 on proinflammatory cytokine interleukin (IL)-1β release human peripheral blood mononuclear cells was evaluated, and found to be potently inhibited by both agents.

Enhanced Aβ1–40 Production in Endothelial Cells Stimulated with Fibrillar Aβ1–42

Amyloid accumulation in the brain of Alzheimer’s patients results from altered processing of the 39- to 43-amino acid amyloid β protein (Aβ). The mechanisms for the elevated amyloid (Aβ1–42) are considered to be over-expression of the amyloid precursor protein (APP), enhanced cleavage of APP to Aβ, and decreased clearance of Aβ from the central nervous system (CNS). We report herein studies of Aβ stimulated effects on endothelial cells. We observe an interesting and as yet unprecedented feedback effect involving Aβ1–42 fibril-induced synthesis of APP by Western blot analysis in the endothelial cell line Hep-1. We further observe an increase in the expression of Aβ1–40 by flow cytometry and fluorescence microscopy. This phenomenon is reproducible for cultures grown both in the presence and absence of serum. In the former case, flow cytometry reveals that Aβ1–40 accumulation is less pronounced than under serum-free conditions. Immunofluorescence staining further corroborates these observations. Cellular responses to fibrillar Aβ1–42 treatment involving eNOS upregulation and increased autophagy are also reported.

Early-Onset Familial Alzheimer's Disease (EOFAD)

Early-onset familial Alzheimer's disease (EOFAD) is a condition characterized by early onset dementia (age at onset < 65 years) and a positive family history for dementia. To date, 230 mutations in presenilin (PS1, PS2) and amyloid precursor protein (APP) genes have been identified in EOFAD. The mutations within these three genes (PS1/PS2/APP) affect a common pathogenic pathway in APP synthesis and proteolysis, which lead to excessive production of amyloid β. Compared with sporadic Alzheimer's disease (AD), EOFAD has some distinctive features including early age at onset, positive familial history, a variety of non-cognitive neurological symptoms and signs, and a more aggressive course. There is marked phenotypic heterogeneity among different mutations of EOFAD. Studies in presymptomatic mutation carriers reveal biomarkers abnormalities. EOFAD diagnosis is based on clinical and family history, neurological symptoms and examination, biomarker features, as well as genotyping in some cases. New therapeutic agents targeting amyloid formation may benefit EOFAD individuals.

Liver X receptor agonists and altered cellular cholesterol metabolism modulate APP expression in cerebromicrovascular endothelial cells

Several lines of evidence suggest a close relationship between dysregulated lipid and lipoprotein metabolism and the development of Alzheimer's disease (AD). Impaired clearance of cerebral amyloid beta (Aß) across the blood-brain barrier (BBB) may facilitate the onset and progression of AD. However, regulatory pathways governing amyloid precursor protein (APP) synthesis, secretion, and processing by brain capillary endothelial cells (BCEC; the anatomical basis of the BBB) are largely unknown. Therefore, we aimed to investigate whether BCEC are capable of endogenous APP synthesis and whether to what extent APP synthesis is under control of cellular cholesterol homeostasis. Primary porcine brain capillary endothelial cells (pBCEC) were used as an in vitro model of the BBB. Gene and protein expression levels of APP and cholesterol metabolism genes were determined by real-time PCR and immunoblotting, respectively. Effects of either liver X receptor (LXR) activation or altered cholesterol biosynthesis on APP metabolism were investigated in pBCEC treated with 24(S), 27-hydroxycholesterol (24OH-C, 27OH-C), non steroidal LXR ligand TO901317, cholesterol or simvastatin. Cholesterol biosynthesis and cholesterol ester formation from radiolabeled precursor [14C]-acetate was examined in pBCEC in the absence or presence of the conditions described above. APP secretion was investigated in a polarized in vitro model mimicking the BBB. Using the in vitro model of the BBB consisting of pBCEC, we demonstrate that endogenous APP synthesis by pBCEC is significantly increased in response to 24OH-C or 27OH-C, TO901317, cholesterol, or simvastatin, while secretion of APP into the culture medium was attenuated (25-43%). In parallel, LXR agonists reduced cholesterol biosynthesis by 30-80% while stimulating its esterification up to 2.5-fold by modifying HMG-CoA reductase (HMGCR), sterol regulatory element-binding protein (SREBP-2), andacyl-coenzyme A: cholesterol acyltransferase (ACAT-2) expression levels. In a polarized in vitro model mimicking the BBB, pBCEC secreted APP/Aß peptides in a polarized manner - preferentially to the basolateral compartment. Our data demonstrate the BBB as an additional source of cerebral APP/Aß. An imbalance in its cholesterol and APP metabolism might therefore contribute to the pathogenesis of AD, whereas modulation of cellular cholesterol metabolism could contribute to regulate APP synthesis by cerebromicrovascular endothelial cells.

EIF4A Inhibition Allows Translational Regulation of mRNAs Encoding Proteins Involved in Alzheimer's Disease

Alzheimer's disease (AD) is the main cause of dementia in our increasingly aging population. The debilitating cognitive and behavioral symptoms characteristic of AD make it an extremely distressing illness for patients and carers. Although drugs have been developed to treat AD symptoms and to slow disease progression, there is currently no cure. The incidence of AD is predicted to increase to over one hundred million by 2050, placing a heavy burden on communities and economies, and making the development of effective therapies an urgent priority. Two proteins are thought to have major contributory roles in AD: the microtubule associated protein tau, also known as MAPT; and the amyloid-beta peptide (A-beta), a cleavage product of amyloid precursor protein (APP). Oxidative stress is also implicated in AD pathology from an early stage. By targeting eIF4A, an RNA helicase involved in translation initiation, the synthesis of APP and tau, but not neuroprotective proteins, can be simultaneously and specifically reduced, representing a novel avenue for AD intervention. We also show that protection from oxidative stress is increased upon eIF4A inhibition. We demonstrate that the reduction of these proteins is not due to changes in mRNA levels or increased protein degradation, but is a consequence of translational repression conferred by inhibition of the helicase activity of eIF4A. Inhibition of eIF4A selectively and simultaneously modulates the synthesis of proteins involved in Alzheimer's disease: reducing A-beta and tau synthesis, while increasing proteins predicted to be neuroprotective.

Regulating amyloid precursor protein synthesis through an internal ribosomal entry site.

Expression of amyloid precursor protein (APP) is critical to the etiology of Alzheimer's disease (AD). Consequently, regulating APP expression is one approach to block disease progression. To this end, APP can be targeted at the levels of transcription, translation, and protein stability. Little is currently known about the translation of APP mRNA. Here, we report that endogenous APP mRNA is translated in neural cell lines via an internal ribosome entry site (IRES) located in the 5′-untranslated leader. The functional unit of the APP IRES is located within the 5′ 50 nucleotides of the 5′-leader. In addition, we found that the APP IRES is positively regulated by two conditions correlated with AD, increased intracellular iron concentration and ischemia. Interestingly, the enhancement of APP IRES activity is dependent upon de novo transcription. Taken together, our data suggest that internal initiation of translation of the APP mRNA is an important mode for synthesis of APP, a mechanism which is regulated by conditions that also contribute to AD.

The Cholesterol Transport Inhibitor U18666a Regulates Amyloid Precursor Protein Metabolism and Trafficking in N2aAPP &amp;#x201C;Swedish&amp;#x201D; Cells

Cholesterol transport is a key regulator of amyloid precursor protein (APP) processing and beta-amyloid (Abeta production, implicated in Alzheimer's disease. Perturbation of cholesterol transport can be pharmacologically induced by the class II amphiphile 3-beta-[2-(diethylamino)ethoxy]androst-5-en-17-one, U18666a; however, the mechanisms by which U18666a controls APP metabolism and trafficking have not been elucidated. We proposed to determine how U18666a regulates APP holoprotein metabolism and trafficking in N2a mouse neuroblastoma cells stably expressing the human APP protein. Secretion of Abeta1-40 was reduced in U18666a-treated cells. U18666a elevated the steady state level of the APP holoprotein but not APP mRNA levels. U18666a increased sAPPalpha secretion and intracellular alpha-CTF/C83 levels but intracellular betaCTF/C99 levels were reduced. The increase in APP protein level was due to decreased catabolism rather than increased APP synthesis. Interestingly, U18666a regulated APP trafficking and increased the level of the holoprotein at the cell surface for alpha-secretase processing and reduced internalization for beta-secretase processing. These data demonstrate that U18666a effects on cholesterol transport function to regulate amyloid precursor protein metabolism and trafficking.

Protein–RNA Interaction Links Fragile X Syndrome and Alzheimer Disease

Protein–RNA Interaction Links Fragile X Syndrome and Alzheimer Disease

Effects of huperzine A on amyloid precursor protein processing and β‐amyloid generation in human embryonic kidney 293 APP Swedish mutant cells

The amyloid precursor protein (APP) is cleaved enzymatically by nonamyloidogenic and amyloidogenic pathways. alpha-Secretase (alpha-secretase), cleaves APP within the beta-amyloid (Abeta) sequence, resulting in the release of a secreted fragment of APP (alphaAPPs) and precluding Abeta generation. In this study, we investigated the effects of an acetylcholinesterase inhibitor, huperzine A (Hup A), on APP processing and Abeta generation in human embryonic kidney 293 cells transfected with human APP bearing the Swedish mutation (HEK293 APPsw). Hup A dose dependently (0-10 microM) increased alphaAPPs release and membrane-coupled APP CTF-C83, suggesting increased APP metabolism toward the nonamyloidogenic alpha-secretase pathway. The metalloprotease inhibitor TAPI-2 inhibited the Hup A-induced increase in alphaAPPs release, further suggesting a modulatory effect of Hup A on alpha-secretase activity. The synthesis of full-length APP and cell viability were unchanged after Hup A incubation, whereas the level of Abeta(Total) was significantly decreased, suggesting an inhibitory effect of Hup A on Abeta production. Hup A-induced alphaAPPs release was significantly reduced by the protein kinase C (PKC) inhibitors GF109203X and Calphostin C. These data, together with the finding that the PKCalpha level was enhanced prior to the increase of alphaAPPs secretion, indicate that PKC may be involved in Hup A-induced alphaAPPs secretion by HEK293 APPsw cells. Our data suggest alternative pharmacological mechanisms of Hup A relevant to the treatment of Alzheimer's disease.

Decoy mRNAs reduce β-amyloid precursor protein mRNA in neuronal cells

Overproduction of amyloid precursor protein (APP) and β-amyloid likely contribute to neurodegeneration in Alzheimer's disease (AD). In an effort to understand neuronal APP gene regulation, we identified a 52 base element (52sce) immediately downstream from the stop codon that stabilizes APP mRNA. Deletion of this domain drastically destabilized APP mRNAs and reduced APP synthesis in vitro. Chimeric globin-APP mRNAs containing the globin coding sequence fused to the entire APP 3′-UTR, showed regulation similar to full-length APP mRNA. A variety of cytoplasmic lysates contain 52sce RNA binding activity, suggesting cis– trans interactions regulate the element's functionality. Finally, the overexpression of chimeric mRNAs, containing the GFP coding sequence and APP 3′-UTR, dramatically reduced endogenous APP steady-state levels in SH-SY5Y neuroblastoma cells and suggests a novel approach to reduce the amyloid burden in AD patients.

Persistent Amyloidosis following Suppression of Aβ Production in a Transgenic Model of Alzheimer Disease

BackgroundThe proteases (secretases) that cleave amyloid-β (Aβ) peptide from the amyloid precursor protein (APP) have been the focus of considerable investigation in the development of treatments for Alzheimer disease. The prediction has been that reducing Aβ production in the brain, even after the onset of clinical symptoms and the development of associated pathology, will facilitate the repair of damaged tissue and removal of amyloid lesions. However, no long-term studies using animal models of amyloid pathology have yet been performed to test this hypothesis.Methods and FindingsWe have generated a transgenic mouse model that genetically mimics the arrest of Aβ production expected from treatment with secretase inhibitors. These mice overexpress mutant APP from a vector that can be regulated by doxycycline. Under normal conditions, high-level expression of APP quickly induces fulminant amyloid pathology. We show that doxycycline administration inhibits transgenic APP expression by greater than 95% and reduces Aβ production to levels found in nontransgenic mice. Suppression of transgenic Aβ synthesis in this model abruptly halts the progression of amyloid pathology. However, formation and disaggregation of amyloid deposits appear to be in disequilibrium as the plaques require far longer to disperse than to assemble. Mice in which APP synthesis was suppressed for as long as 6 mo after the formation of Aβ deposits retain a considerable amyloid load, with little sign of active clearance.ConclusionThis study demonstrates that amyloid lesions in transgenic mice are highly stable structures in vivo that are slow to disaggregate. Our findings suggest that arresting Aβ production in patients with Alzheimer disease should halt the progression of pathology, but that early treatment may be imperative, as it appears that amyloid deposits, once formed, will require additional intervention to clear.

Estrogen stimulates release of secreted amyloid precursor protein from primary rat cortical neurons via protein kinase C pathway1

To investigate the mechanism of the action of estrogen, which stimulates the release of secreted amyloid precursor protein alpha (sAPP(alpha)) and decreases the generation of amyloid-beta protein (A(beta)), a dominant component in senile plaques in the brains of Alzheimer's disease patients. Experiments were carried out in primary rat cortical neurons, and Western blot was used to detect sAPP(alpha) in a culture medium and the total amount of cellular amyloid precursor protein (APP) in neurons. 17beta-Estradiol (but not 17alpha-estradiol) and beta-estradiol 6-(O-carboxymethyl) oxime: BSA increased the secretion of sAPP(alpha) and this effect was blocked by protein kinase C (PKC) inhibitor calphostin C, but not by the classical estrogen receptor antagonist ICI 182,780. Meanwhile, 17beta-estradiol did not alter the synthesis of cellular APP. The effect of 17beta-estradiol on sAPP(alpha) secretion is likely mediated through the membrane binding sites, and needs molecular configuration specificity of the ligand. Furthermore, the action of the PKC-dependent pathway might be involved in estrogen-induced sAPP(alpha) secretion.