Beta-amyloid Plaque Formation Research Articles

Cathepsin D SNP associated with increased risk of variant Creutzfeldt-Jakob disease

BackgroundVariant Creutzfeldt-Jakob disease (vCJD) originally resulted from the consumption of foodstuffs contaminated by bovine spongiform encephalopathy (BSE) material, with 163 confirmed cases in the UK to date. Many thousands are likely to have been exposed to dietary infection and so it is important (for surveillance, epidemic modelling, public health and understanding pathogenesis) to identify genetic factors that may affect individual susceptibility to infection. This study looked at a polymorphism in the cathepsin D gene (refSNP ID: rs17571) previously examined in Alzheimer's disease (AD).MethodsBlood samples taken from 110 vCJD patients were tested for the C-T base change, and genotype data were compared with published frequencies for a control population using multiple logistic regression.ResultsThere was a significant excess of the cathepsin D polymorphism TT genotype in the vCJD cohort compared to controls. The TT genotype was found to have a 9.75 fold increase in risk of vCJD compared to the CT genotype and a 10.92 fold increase compared to the CC genotype.ConclusionThis mutation event has been observed to alter the protease activity of the cathepsin D protein and has been linked to an increase in amyloid beta plaque formation in AD. vCJD neuropathology is characterised by the presence of amyloid plaques, formed from the prion protein, and therefore alterations in the amyloid processing activity of cathepsin D may affect the neuropathogenesis of this disease.

Neurogenic responses to amyloid-beta plaques in the brain of Alzheimer’s disease-like transgenic (pPDGF-APPSw,Ind) mice

Formation and accumulation of amyloid-beta (A beta) plaques are associated with declined memory and other neurocognitive function in Alzheimer's disease (AD) patients. However, the effects of A beta plaques on neural progenitor cells (NPCs) and neurogenesis from NPCs remain largely unknown. The existing data on neurogenesis in AD patients and AD-like animal models remain controversial. For this reason, we utilized the nestin second-intron enhancer controlled LacZ (pNes-LacZ) reporter transgenic mice (pNes-Tg) and Bi-transgenic mice (Bi-Tg) containing both pPDGF-APPSw,Ind and pNes-LacZ transgenes to investigate the effects of A beta plaques on neurogenesis in the hippocampus and other brain regions of the AD-like mice. We chose transgenic mice at 2, 8 and 12 months of age, corresponding to the stages of A beta plaque free, plaque onset and plaque progression to analyze the effects of A beta plaques on the distribution and de novo neurogenesis of (from) NPCs. We demonstrated a slight increase in the number of NPCs in the hippocampal regions at the A beta plaque free stage, while a significant decrease in the number of NPCs at A beta plaque onset and progression stages. On the other hand, we showed that A beta plaques increase neurogenesis, but not gliogenesis from post-mitotic NPCs in the hippocampus of Bi-Tg mice compared with age-matched control pNes-Tg mice. The neurogenic responses of NPCs to A beta plaques suggest that experimental approaches to promote de novo neurogenesis may potentially improve neurocognitive function and provide an effective therapy for AD.

Selective ablation of bone marrow‐derived dendritic cells increases amyloid plaques in a mouse Alzheimer's disease model

We have recently shown that the ability of microglia to effectively fight off aggregated beta-amyloid plaque formation and cognitive loss in transgenic mouse models of Alzheimer's disease (Tg-AD), is augmented in response to T-cell-based immunization, using glatiramer acetate (GA). The immunization increases incidence of local CD11c+ dendritic-like cells. It is unclear, however, whether these dendritic cells are derived from resident microglia or from the bone marrow. To determine the origin of this dendritic-cell population, we used chimeric mice whose bone marrow-derived cells express a transgene that allows the cells to be specifically ablated by diphtheria toxin. We show here that T-cell-based immunization of these mice, using GA, induced the recruitment of bone marrow-derived dendritic cells. Depletion of the dendritic cells by systemic injection of diphtheria toxin resulted in significantly increased formation of amyloid plaques. Thus, recruitment of bone marrow-derived dendritic cells evidently plays a role in reducing plaque formation in a mouse model of Alzheimer's disease.

Behavioral consequences of ovarian atrophy and estrogen replacement in the APPswe mouse

Cognitive performance was evaluated in a longitudinal study of APPswe2576 transgenic mice (APP) and a wildtype (WT) comparison group. Subgroups of the APP mice were treated with the ovarian toxicant 4-vinylcyclo-hexene diepoxide (VCD) at 60–75 days of age to induce ovarian atrophy and/or given estrogen (estradiol, 4 μg/day) continuously by pellet from 76 days of age. APP mice had a generally poorer radial maze performance than WT at 4.5, 7.5, 10.5 and 15 months of age. In separate tests, APP mice had a slight motor impairment, higher incidence of homecage stereotypy, hyperactivity in an open field and reduced object exploration relative to the WT group. Ovarian atrophy led to better maze performance at 7.5 months. The effect of estrogen on maze performance with aging could not be effectively evaluated due to poor survival (30%) of these mice. No effects of ovarian atrophy or estrogen treatment were identified for amyloid-beta accumulation or plaque formation at 15 months. Long-term longitudinal studies in animal models are needed to explore the consequences of menopause and hormone replacement on Alzheimer's disease, but they are complicated by considerations of survival, pre-aging deficits, testing experience and selection of appropriate estrogen treatment levels.

Association of Increased Cortical Soluble Aβ42 Levels With Diffuse Plaques After Severe Brain Injury in Humans

Traumatic brain injury (TBI) is an environmental risk factor for developing Alzheimer disease. This may be due, in part, to changes associated with beta-amyloid (Abeta) plaque formation, which can occur within hours after injury, regardless of the patient's age. In addition to being precursors of toxic fibrils that deposit into plaques, soluble (nonfibrillar) Abeta peptides are posited to disrupt synaptic function and are associated with cognitive decline in Alzheimer disease. Changes in soluble Abeta levels and their relationship to Abeta plaque formation following TBI are unknown. To quantify brain tissue levels of soluble Abeta peptides and their precursor protein in relation to Abeta plaque formation after TBI in humans. Surgically resected temporal cortex tissue from patients with severe TBI was processed for biochemical assays of soluble Abeta peptides with COOH-termini ending in amino acid 40 (Abeta(40)) or 42 (Abeta(42)) and Abeta precursor protein to compare patients with cortical Abeta plaques and those without. Patients Nineteen subjects admitted to the University of Pittsburgh Medical Center for treatment of severe closed head injury. Patients with severe TBI and cortical plaques had higher levels of soluble Abeta(1-42) but not Abeta(1-40); half of them were apolipoprotein E (APOE) epsilon4 allele carriers. The lowest Abeta levels were in 1 patient without plaques who was the only subject with an APOE epsilon2 allele. beta-Amyloid precursor protein levels were comparable in the 2 TBI groups. Selective increases in soluble Abeta(1-42) after TBI may predispose individuals with a brain injury to Alzheimer disease pathology. This may be influenced by the APOE genotype, and it may confer increased risk for developing Alzheimer disease later in life.

Macromolecules involved in production and metabolism of beta-amyl oid at the brain barriers

One of the notable features of Alzheimer's disease (AD) is the overabundance of beta-amyloid peptides in brain fluids, leading to the formation and deposition of insoluble amyloid plaques. Previous work in this lab demonstrates that the normal choroid plexus, a primary component of the blood–cerebrospinal fluid barrier, has the capacity to remove beta-amyloid from the cerebrospinal fluid, potentially preventing the formation of beta-amyloid plaques. The purpose of this work was to determine whether the choroid plexus and/or the brain capillaries, a primary component of the blood–brain barrier, possessed the capacity to produce or degrade beta-amyloid peptides. Using quantitative real-time RT-PCR, immunodetection and enzyme activity assays, we demonstrated the presence in brain barriers of several key enzymes involved in beta-amyloid production, namely, amyloid precursor protein and beta-secretase, and in beta-amyloid metabolism and alternate processing, such as insulin degrading enzyme, endothelin-converting enzyme-1, neprilysin and alpha-secretase. Furthermore, beta-amyloid presence, in the absence of its application in culture media, was detected in an immortalized choroidal epithelial cell line, known as Z310 cells. The ability of the choroid plexus to produce and degrade beta-amyloid, in addition to its transport function, suggests a vital role of this tissue in maintaining beta-amyloid homeostasis. Disruption of this homeostasis due to aging, injury or toxicant exposure may contribute to accumulation of beta-amyloid peptides in the brain fluids, leading to AD.

Canine cognitive dysfunction and the cerebellum: Acetylcholinesterase reduction, neuronal and glial changes

The specific functional and pathological alterations observed in Alzheimer's disease are less severe in the cerebellum than in other brain areas, particularly the entorhinal cortex and hippocampus. Since dense core amyloid-beta plaque formation has been associated with an acetylcholinesterase heterogeneous nucleator action, we examined if an acetylcholinesterase imbalance was involved in cerebellum plaque deposition. By using the canine counterpart of senile dementia of the Alzheimer's type, a promising model of human brain aging and early phases of Alzheimer's disease, we investigated how cerebellar pathology and acetylcholinesterase density could be related with cognitive dysfunction. As in Alzheimer's disease, the late affectation of the cerebellum was evidenced by its lack of amyloid-beta plaque and the presence of diffuse deposition throughout all cortical grey matter layers. The highest acetylcholinesterase optic density corresponded to cerebellar islands of the granular layer and was predominantly associated with synaptic glomeruli and the somata of Golgi cells. Its reduction correlated with aging and loss of granule cells, whereas cognitive deficit only correlated with loss of Purkinje cells. The observed Bergmann glia alterations may correspond to a reactive response to the loss and damage of the Purkinje cells, their specific neuronal partner. Regarding the role of acetylcholinesterase mediation in amyloid-beta deposition, our data argue against an interaction between these two proteins because acetylcholinesterase reduction correlates with aging but not with cognitive deficit. Finally, our data support the use of companion dogs of all breeds to study aging and early phases of Alzheimer's disease.

Radioligand Development for PET Imaging of β-Amyloid (Aβ)-Current Status

Two of the main pathological hallmarks of Alzheimer's disease (AD) are neuritic plaques and neurofibrillary tangles. Significant evidence supports a critical and probable causative role of beta amyloid (Abeta) plaque formation. Since neuroprotective treatments are typically most effective at early stages of injury, the detection and measurement of Abeta load in living brain should be performed at early and perhaps even presymptomatic stages of AD. Two primary targets of molecular imaging research with positron emission tomography (PET) are to develop surrogate markers (radioligands) for assessing disease progression and for monitoring the efficacy of developmental therapeutics. Here, we review the current status of radioligand development for PET imaging of Abeta aggregates. General structure-activity relationships have emerged, including the identification of at least three different ligand binding sites in various Abeta aggregates and recognition of the general structural requirements for ligand binding at each site. Also a few radioligands applicable to imaging Abeta plaques in living human brain with positron emission tomography (PET) have emerged, including [(11)C]PIB, [(11)C]SB-13 and [(18)F]FDDNP.

Association between the Polymorphism of CCR5 and Alzheimer's Disease

Alzheimer's disease (AD) is the most common cause of dementia in Western society. The prevalence of AD is greater in women than in men, largely due to longevity and survival differences favoring women. However, some studies suggest that incidence rates may really be increased in women. One possible factor influencing AD incidence in women is the loss of ovarian estrogens production after menopause, which might be involved in AD pathogenesis. Estrogens seem to influence some neuronal functions. Many of these actions appear beneficial (i.e., neuroprotective action against a variety of insults, as oxidative stress, and reduction of beta-amyloid plaques formation). Furthermore, several studies have shown that proinflammatory genotypes seem to significantly contribute to AD risk. In the present study, we evaluated whether the anti-inflammatory allele of chemokine receptor CCR5 is a component of the genetic protective background versus AD neuronal degeneration. We genotyped for Delta32 (a 32-bp deletion of the CCR5 gene that causes a frameshift at amino acid 185) in 191 AD patients (133 women and 58 men; age range: 53-98 years; mean age: 74.88 +/- 8.44) and 182 controls (98 women and 84 men; age range: 65-93; mean age 73.21 +/- 8.24) from northern Italy. No different distribution of the CCRDelta32 deletion in the two cohorts was clearly evident. Statistical analysis by gender stratification, demonstrated no differences in genotype distribution and allelic frequency both in women and in men. Further, studies should focus on identification of proinflammatory genetic variants involved in AD pathogenesis in women.

Aluminium Mediated Oxidative Stress : Possible Relationship to Cognitive Impairment of Alzheimer's Type

Aluminum (Al) is a trivalent cation that does not undergo redox change and it may cause a variety of neurological disorders. In this review, we discuss evidence for a significant role of aluminum in Alzheimer's disease (AD). Al has been implicated as a potential risk factor in AD and for elderly cognitive impairment by epidemiological studies of drinking water, food, medicine as well as natural resources. Neurotoxicity from excess brain exposure to aluminum is documented from both clinical observations and animal experiments. The pathology of Alzheimer's disease is related to the alteration in neurotransmission, beta amyloid production, plaques formation and cytoskeletal abnormalities. The exact mechanism of Al toxicity is still unclear; however accumulating evidences suggested that Al could potentiate oxidative and inflammatory events leading to tissue damage. In conclusion it is proposed that a combination of anticholinergic drugs and micronutrients including antioxidants may be more effective than the individual agents in prevention and treatment of Alzheimer's disease. doi: 10.5214/ans.0972.7531.2006.130104

Discovery of Acetylcholinesterase Peripheral Anionic Site Ligands through Computational Refinement of a Directed Library

The formation of beta-amyloid plaques in the brain is a key neurodegenerative event in Alzheimer's disease. Small molecules capable of binding to the peripheral anionic site of acetylcholinesterase (AChE) have been shown to inhibit the AChE-induced aggregation of the beta-amyloid peptide. Using the combination of a computational docking model and experimental screening, five compounds that completely blocked the amyloidogenic effect of AChE were rapidly identified from an approximately 200-member library of compounds designed to disrupt protein-protein interactions. Critical to this docking model was the inclusion of two explicit water molecules that are tightly bound to the enzyme. Interestingly, none of the tested compounds inhibited the related enzyme butyrylcholinesterase (BuChE) up to their aqueous solubility limits. These compounds are among the most potent inhibitors of amyloid beta-peptide aggregation and are equivalent only to propidium, a well-characterized AChE peripheral anionic site binder and aggregation inhibitor.

Agrin binds α-synuclein and modulates α-synuclein fibrillation

Recent studies have begun to investigate the role of agrin in brain and suggest that agrin's function likely extends beyond that of a synaptogenic protein. Particularly, it has been shown that agrin is associated with the pathological lesions of Alzheimer's disease (AD) and may contribute to the formation of beta-amyloid (Abeta) plaques in AD. We have extended the analysis of agrin's function in neurodegenerative diseases to investigate its role in Parkinson's disease (PD). Alpha-synuclein is a critical molecular determinant in familial and sporadic PD, with the formation of alpha-synuclein fibrils being enhanced by sulfated macromolecules. In the studies reported here, we show that agrin binds to alpha-synuclein in a heparan sulfate-dependent (HS-dependent) manner, induces conformational changes in this protein characterized by beta-sheet structure, and enhances insolubility of alpha-synuclein. We also show that agrin accelerates the formation of protofibrils by alpha-synuclein and decreases the half-time of fibril formation. The association of agrin with PD lesions was also explored in PD human brain, and these studies shown that agrin colocalizes with alpha-synuclein in neuronal Lewy bodies in the substantia nigra of PD brain. These studies indicate that agrin is capable of accelerating the formation of insoluble protein fibrils in a second common neurodegenerative disease. These findings may indicate shared molecular mechanisms leading to the pathophysiology in these two neurodegenerative disorders.

Does beta-amyloid plaque formation cause structural injury to neuronal processes?

The precise role of beta-amyloid plaque formation in the cascade of brain cell changes that lead to neurodegeneration and dementia in Alzheimer's disease has been unclear. Studies have indicated that neuronal processes surrounding and within plaques undergo a series of biochemical and morphological alterations. Morphological alterations include reactive, degenerative and sprouting-related 'dystrophic' neuritic structures, derived principally from axons, which involve specific changes in cytoskeletal proteins such as tau and NF triplet proteins. More compact and fibrous plaques are associated with more extensive neuritic pathology than non-fibrillar, diffuse beta-amyloid deposits. Cortical apical dendritic processes are either 'clipped' by plaque formation or are bent around more compact plaques. Examination of cases of 'pathological' brain ageing, which may represent a preclinical form of Alzheimer's disease, demonstrated that the earliest neuritic pathology associated with plaques was similar to the reactive changes that follow structural injury to axons. In vivo and in vitro experimental models of structural injury to axons produce identical reactive changes that subsequently lead to an attempt at regenerative sprouting by damaged axons. Thus, beta-amyloid plaque formation may cause structural injury to axons that is subsequently followed by an aberrant sprouting response that presages neurodegeneration and dementia. Identification of the key neuronal alterations underlying the pathology of Alzheimer's disease may provide new avenues for therapeutic intervention.

Alzheimer's disease β‐secretase BACE1 is not a neuron‐specific enzyme

The brains of Alzheimer's disease (AD) patients are morphologically characterized by neurofibrillar abnormalities and by parenchymal and cerebrovascular deposits of beta-amyloid peptides. The generation of beta-amyloid peptides by proteolytical processing of the amyloid precursor protein (APP) requires the enzymatic activity of the beta-site APP cleaving enzyme 1 (BACE1). The expression of this enzyme has been localized to the brain, in particular to neurons, indicating that neurons are the major source of beta-amyloid peptides in brain. Astrocytes, on the contrary, are known to be important for beta-amyloid clearance and degradation, for providing trophic support to neurons, and for forming a protective barrier between beta-amyloid deposits and neurons. However, under certain conditions related to chronic stress, the role of astrocytes may not be beneficial. Here we present evidence demonstrating that astrocytes are an alternative source of BACE1 and therefore may contribute to beta-amyloid plaque formation. While resting astroyctes in brain do not express BACE1 at detectable levels, cultured astrocytes display BACE1 promoter activity and express BACE1 mRNA and enzymatically active BACE1 protein. Additionally, in animal models of chronic gliosis and in brains of AD patients, there is BACE1 expression in reactive astrocytes. This would suggest that the mechanism for astrocyte activation plays a role in the development of AD and that therapeutic strategies that target astrocyte activation in brain may be beneficial for the treatment of AD. Also, there are differences in responses to chronic versus acute stress, suggesting that one consequence of chronic stress is an incremental shift to different phenotypic cellular states.

α-Internexin immunoreactivity reflects variable neuronal vulnerability in Alzheimer's disease and supports the role of the β-amyloid plaques in inducing neuronal injury

This study investigated the role of alpha-internexin in the neuronal alterations associated with beta-amyloid plaque formation in Alzheimer's disease (AD). Cortical neurons could be defined by their variable content of neurofilament (NF) triplet and alpha-internexin proteins, with a distinct population of supragranular pyramidal cells containing alpha-internexin alone. Both NF triplet and alpha-internexin were localized to reactive axonal structures in physically damaged neurons in experimental trauma models. Similarly, NF triplet and alpha-internexin immunoreactive neurites were localized to plaques densely packed with beta-amyloid fibrils in preclinical AD cases, indicating that certain plaques may cause structural injury or impediment of local axonal transport. However, alpha-internexin, and not NF triplet, ring-like reactive neurites were present in end-stage AD cases, indicating the relatively late involvement of neurons that selectively contain alpha-internexin. These results implicate the expression of specific intermediate filament proteins in a distinct hierarchy of differential neuronal vulnerability to AD.

Aging-related increase in oxidative stress correlates with developmental pattern of beta-secretase activity and beta-amyloid plaque formation in transgenic Tg2576 mice with Alzheimer-like pathology

The molecular mechanisms of β-amyloidogenesis in sporadic Alzheimer’s disease are still poorly understood. To reveal whether aging-associated increases in brain oxidative stress and inflammation may trigger onset or progression of β-amyloid deposition, a transgenic mouse (Tg2576) that express the Swedish double mutation of human amyloid precursor protein (APP) was used as animal model to study the developmental pattern of markers of oxidative stress and APP processing. In Tg2576 mouse brain, cortical levels of soluble β-amyloid (1–40) and (1–42) steadily increased with age, but significant deposition of fibrillary β-amyloid in cortical areas did not occur before postnatal age of 10 months. The slope of increase in cerebral cortical β-secretase (BACE1) activities in Tg2576 mice between ages of 9 and 13 months was significantly higher as compared to that of the α-secretase, while the expression level of BACE1 protein and mRNA did not change with age. The activities of superoxide dismutase and glutathione peroxidase in cortical tissue from Tg2576 mice steadily increased from postnatal age 9–12 months. The levels of cortical nitric oxide, and reactive nitrogen species demonstrated peak values around 9 months of age, while the level of interleukin-1β steadily increased from postnatal month 13 onwards. The developmental temporal coincidence of increased levels of reactive nitrogen species and antioxidative enzymes with the onset of β-amyloid plaque deposition provides further evidence that developmentally and aging-induced alterations in brain oxidative status exhibit a major factor in triggering enhanced production and deposition of β-amyloid, and potentially predispose to Alzheimer’s disease.

Immunotherapy of Alzheimer's disease. Results of experimental investigations and treatment of perspectives

This review discusses the molecular basis and current status of immunotherapeutic strategies for prevention and treatment of Alzheimer's disease (AD). From the molecular view-point AD belongs to the group of conformational diseases. In-vitro studies demonstrated that monoclonal antibodies could modulate the conformation of Abeta peptides with subsequent inhibition of amyloid fibrils formation and aggregation. The efficacy of this approach was then successfully proved in the murine models of AD using predominantly Abeta (42) peptide as immunogen. Immunisation of the young animals essentially prevented the development of beta-amyloid plaques formation and of concomittant neuropathology. Treatment of the older animals markedly reduced the pre-existing AD-like neuropathology. Immunisation was capable of preventing cognitive deficits in the young transgenic animals and improve the memory and behavioural disturbances in the older animals. Measurement of specific murine immunoglobulines in Abeta-vaccinated mice demonstrated a predominant IgG1 and IgG2b isotypes, suggesting a type 2 (T (H)2) T-helper cell immune response, which drives humoral immunity. The intensity of the immune response depended on transgenic animals genotype, dose, frequency and route of immunogen administration. The mechanism of antibodies action in transgenic animals consists of inducing conformational and solubility changes in Abeta peptides as well as their peripheral sink. Lymphocyte proliferation assays using Abeta peptides and splenocytes from vaccinated mice demonstrated that vaccine specifically stimulated T-cell epitopes present within the Abeta-peptide. Extensive quantitative morphological, histochemical and molecular analysis of brain tissue from several species of Abeta-immunised transgenic and non-transgenic animals showed no evidence of autoimmune reaction, complement activation or cross-reaction. No pathological changes were found in all other organs, including the kidney. Neuropathologic examination in a patient treated with vaccine revealed similar vaccination effects as in experimental animals. An aseptic meningo-encephalitis was reported in 5 % of patients included in a clinical trial in which a vaccine containing Abeta (42) peptide (AN1792) was administered intramuscularly. The causal relationship to the vaccine administration cannot be excluded since in transgenic mice a transient microglia activation was seen. However, this relatively infrequent although severe adverse effect points to a possible participation of some actually unknown risk factors in the treated patients. With regard to the rapid progress in biotechnology, especially in the vaccines technology, the development of efficacious and safe immunogens as well as of new vaccination techniques for immuntherapy of AD can be expected in the next future.

Estrogen treatment improves spatial learning in APP + PS1 mice but does not affect beta amyloid accumulation and plaque formation

We investigated the effects of ovariectomy (OVX) and 17β-estradiol (0.18 mg per pellet) treatment on spatial learning and memory, hippocampal beta amyloid (Aβ) levels, and amyloid plaque counts in double transgenic mice (A/P) carrying mutated amyloid precursor protein (APPswe) and presenilin-1 (PS1-A246E). After OVX at 3 months of age, the mice received estrogen treatment for the last 3 months of their lifetime before they were killed at 6, 9, or 12 months of age. Estrogen treatment in A/P OVX mice increased the number of correct choices in a position discrimination task in the T-maze, and slightly improved their performance in a win-stay task (1/8 arms baited) in the radial arm maze (RAM). However, estrogen treatment did not reverse the Aβ-dependent cognitive deficits of A/P mice in the water maze (WM) spatial navigation task. Furthermore, ovariectomy or estrogen treatment in OVX and sham-operated A/P mice had no effect on hippocampal amyloid accumulation. These results show that the estrogen treatment in a transgenic mouse model of Alzheimer's disease (AD) improves performance in the same learning and memory tasks as in the normal C57BL/6J mice. However, the estrogen effects in these mice appeared to be unrelated to Aβ-induced cognitive deficits. Our results do not support the idea that estrogen treatment decreases the risk or alleviates the symptoms of Alzheimer's disease by inhibiting the accumulation of Aβ or formation of amyloid plaques.

Guinea pigs as a nontransgenic model for APP processing in vitro and in vivo.

Alzheimer's disease (AD) is characterized, amongst others, by the appearance of vascular and parenchymal beta-amyloid deposits in brain. Such aggregates are mainly composed of beta-amyloid peptides, which are derived by proteolytic processing of a larger amyloid precursor protein (APP). APP is highly conserved among mammalian species, but experimental studies in rodents are often hampered by the humble APP-processing in the amyloidogenic pathway and by the inability of rodent beta-amyloid peptides to form higher molecular aggregates such as soluble oligomers and insoluble beta-amyloid plaques. Thus, there is need for in vitro and in vivo model systems that allow identification of factors that increase amyloidogenic APP processing and accelerate beta-amyloid plaque formation and testing the potency of pharmacological manipulations to ameliorate beta-amyloid load in brain. Transgenic mice that overexpress human APP containing AD-associated mutations that favor the amyloidogenic pathway of APP processing represent such a model. However, mutations of the APP gene are not frequent in AD and, therefore, the mechanisms of beta-amyloid plaque formation, the composition of beta-amyloid plaques, and the accompanying tissue response in brain of these animals may be different from that in AD. In contrast, guinea pigs express beta-amyloid peptides of the human sequence and appear to represent a more physiological model to examine the long-term effects of experimental manipulations on APP processing and beta-amyloid plaque formation in vivo. Additionally, APP processing in guinea pig primary neuronal cultures has been shown to be similar to cultures of human origin. In this article we highlight the advantages and limitations of using guinea pigs as experimental models to study APP processing.

Multiple-peptide conjugates for binding beta-amyloid plaques of Alzheimer's disease.

Formation of beta-amyloid plaques in Alzheimer's disease is initiated by intermolecular contact of the 5-amino acid sequence, KLVFF, in beta-amyloid peptides ranging in size from 40 to 43 residues. Through optimization of binding avidity using structure/function studies, we have found that the retro-inverso peptide, ffvlk, binds artificial fibrils made from Abeta(1)(-)(40) with moderate affinity (K(d) = 5 x 10(-)(7) M). Conjugates having two copies of this peptide, whether connected by a long poly(ethylene glycol) (PEG) spacer or just two amino acids, display about 100-fold greater affinity for fibrils. Placing six copies of ffvlk on a branched PEG resulted in a 10 000-fold greater affinity (K(d) = 1 x 10(-)(10) M) than the monomer peptide. This increased affinity was accompanied by more effective inhibition of the thioflavin T fluorescence signal, which correlates with neurotoxicity of plaques and fibrils. We propose that conjugates bearing several copies of ffvlk may be useful as diagnostic and therapeutic agents for Alzheimer's disease.