Beta-amyloid plaques in a model for sporadic Alzheimer's disease based on transgenic anti-nerve growth factor antibodies

Cerebral deposition of beta-amyloid (Abeta) is an invariant event of Alzheimer's disease (AD). We recently described that the brain of aged transgenic mice expressing anti-nerve growth factor (NGF) antibodies (AD11 mice) show a dramatic neurodegenerative phenotype, reminiscent of AD, which includes neuronal loss, cholinergic deficit, and tau hyperphosphorylation, associated with neurofibrillary pathology. We now report that brains of aged transgenic mice contain large amounts of beta-amyloid plaques and describe their morphology by a variety of approaches. In conclusion, the chronic deprivation of NGF leads to the formation and deposition of Abeta in AD11 mice, suggesting a direct link between NGF signaling and abnormal processing of amyloid precursor protein.     

Amyloid-induced neurofibrillary tangle formation in Alzheimer''s disease: insight from transgenic mouse and tissue-culture models

Of all forms of dementia, Alzheimer's disease is the most prevalent. It is histopathologically characterized by beta-amyloid-containing plaques, tau-containing neurofibrillary tangles, reduced synaptic density and neuronal loss in selected brain areas. For the rare familial forms of Alzheimer's disease, pathogenic mutations have been identified in both the gene encoding the precursor of the Abeta peptide, APP, itself and in the presenilin genes which encode part of the APP-protease complex. For the more frequent sporadic forms of Alzheimer's disease, the pathogenic trigger has not been unambiguously identified. Whether Abeta is again the main cause remains to be heavily discussed. In a related disorder termed frontotemporal dementia, which is characterized by tangles in the absence of beta-amyloid deposition, mutations have been identified in tau which also lead to neurodegeneration and dementia. For Alzheimer's disease the existence of familial forms lead to the proposition of the amyloid cascade hypothesis, which claims that beta-amyloid causes or enhances the tangle pathology. In this review, we describe tau transgenic mouse models in which aspects of the tau-associated pathology, including tangle formation, has been achieved. Moreover, tau transgenic mouse and tissue-culture models were used to test the amyloid cascade hypothesis. In addition, we discuss alternative hypotheses to explain the sporadic forms. The animal and tissue-culture models will provide insight into the underlying biochemical mechanisms of tau aggregation and nerve cell degeneration. These mechanisms may be partially shared between sporadic Alzheimer's disease, the familial forms and frontotemporal dementia. Eventually, Alzheimer's disease may be redefined based on biochemical events rather than phenotype.     

Mouse models of Alzheimer's disease: the long and filamentous road

Alzheimer's disease (AD) is characterized by memory impairment leading to dementia, deposition of amyloid plaques and neurofibrillary tangles (NFTs), and neuronal loss. The major component of plaques is the amyloid beta peptide, A beta, whereas NFTs contain hyperphosphorylated forms of the microtubule-associated protein tau (tau). Familial AD (FAD) mutations either elevate A beta synthesis by favoring 'secretase' of the Alzheimer beta-amyloid precursor protein (APP) or enhance the fibrillogenic properties of this peptide. Mutations in the tau gene cause a different disease denoted FTPD-17, but suggest that the aberrant forms of tau seen in AD are unlikely to be benign. These findings imply a complex pathogenic cascade in AD and important goals of transgenic modeling are to capture and stratify this pathogenic process. Several laboratories have created APP transgenic (Tg) mice that exhibit AD-like amyloid pathology and A beta burdens. These Tg lines also exhibit deficits in spatial reference and/or working memory, with immunization against A beta attenuating both AD-associated phenotypes. Tangle-like pathologies are observed in mice expressing FTPD-17 mutant forms of tau, but florid tau pathologies based upon the wild type (wt) tau isoforms present in AD have proven more elusive. Creation of animal models with robust amyloid and tau pathologies, yet free of irrelevant confounding pathologies, remains a major objective in this field.     

Triple-transgenic Alzheimer's disease mice exhibit region-specific abnormalities in brain myelination patterns prior to appearance of amyloid and tau pathology

Alzheimer's disease (AD) is a progressively debilitating brain disorder pathologically defined by extracellular amyloid plaques, intraneuronal neurofibrillary tangles, and synaptic disintegrity. AD has not been widely considered a disease of white matter, but more recent evidence suggests the existence of abnormalities in myelination patterns and myelin attrition in AD-afflicted human brains. Herein, we demonstrate that triple-transgenic AD (3xTg-AD) mice, which harbor the human amyloid precursor protein Swedish mutant transgene, presenilin knock-in mutation, and tau P301L mutant transgene, exhibit significant region-specific alterations in myelination patterns and in oligodendrocyte marker expression profiles at time points preceding the appearance of amyloid and tau pathology. These immunohistochemical signatures are coincident with age-related alterations in axonal and myelin sheath ultrastructure as visualized by comparative electron microscopic examination of 3xTg-AD and nontransgenic mouse brain tissue. Overall, these findings indicate that 3xTg-AD mice represent a viable model in which to examine mechanisms underlying AD-related myelination and neural transmission defects that occur early during presymptomatic stages of the disease process.     

Head injury and dementia

PURPOSE OF REVIEW: The link between head injury and dementia/Alzheimer's disease is controversial. This review discusses some recent epidemiological, human autopsy and experimental studies on the relationship between traumatic head injury and dementia. RECENT FINDINGS: Recent epidemiological studies have shown that head injury is a risk factor for the development of dementia/Alzheimer's disease, whereas others have not. After experimental brain trauma the long-term accumulation of amyloid beta peptide suggests that neurodegeneration is influenced by apolipoprotein E epsilon 4, and after human brain injury both amyloid beta peptide deposition and tau pathology are seen, even in younger patients. Amyloid beta peptide levels in the cerebrospinal fluid and the overproduction of beta amyloid precursor protein in humans and animals after traumatic brain injury are increased. Repeated mild head trauma in both animals and humans accelerates amyloid beta peptide accumulation and cognitive impairment. Retrospective autopsy data support clinical studies suggesting that severe traumatic brain injury with long-lasting morphological residuals are a risk factor for the development of dementia/Alzheimer's disease. The influence of the apolipoprotein E genotype on the prognosis of traumatic brain injury is under discussion. SUMMARY: Although epidemiological studies and retrospective autopsy data provide evidence that a later cognitive decline may occur after severe traumatic brain injury, the relationship between dementia after head/brain trauma and apolipoprotein E status is still ambiguous. Both human postmortem and experimental studies showing apolipoprotein beta deposition and tau pathology after head injury support the link between traumatic brain injury and dementia, and further studies are warranted to clarify this relationship.     

Stereological analysis of microvascular parameters in a double transgenic model of Alzheimer's disease

Morphological alterations in microvasculature occur as a common finding in the brains of non-demented aged persons and patients with Alzheimer's disease. Quantifying the extent of this vascular pathology, however, has been complicated by systematic error (bias) associated with the applications of assumption- and model-based morphometric techniques to human and animal tissues. The current study used novel assumption- and model-free stereological approaches to quantify capillary parameters in the corpus callosum of a double amyloid precursor protein/presenilin-1 transgenic murine model of Alzheimer's disease. The results revealed significant reductions in the total number of capillary segments in white matter of transgenic mice compared to non-transgenic littermates, with no differences in total capillary length. These findings support the view that the expression of mutant human genes for beta-amyloid peptides alters the normal architecture of cerebral capillary vessels in the white matter of mouse brain, which may model microvasculature changes reported in Alzheimer's disease.     

Alzheimer cortical neurons containing abundant amyloid mRNA. Relationship to amyloid deposition and senile plaques

Since the detailed molecular events leading to the formation of amyloid-containing senile plaques of the Alzheimer's disease (AD) brain are incompletely understood, the present studies were undertaken to address this issue using a combination of molecular and cytochemical approaches. Amyloid precursor protein riboprobes containing the A4 (beta-amyloid) domain were applied to cortex using the in situ hybridization method to examine the distribution of neuronal amyloid mRNA in relation to the laminar pattern of amyloid deposition and the localization of plaques. The derived data indicated that high levels of amyloid mRNA can be synthesized by AD cortical neurons that appeared to be morphologically intact. The distribution of these cells was not coincident with the cortical laminar pattern that is typical of amyloid deposits observed after immunostaining with anti-A4 monoclonal antibodies. Further, there was no obvious relationship between neurons containing abundant amyloid mRNA and the distribution of plaques identified by thioflavin S staining. While the neuronal synthesis of amyloid may be a significant factor at some point during plaque formation, it may not be the exclusive determinant. The possibility is raised that processes affecting secretion, diffusion, and/or transport of amyloid away from neuronal or non-neuronal cells of origin to sites of deposition may be meaningful aspects of the molecular pathology of Alzheimer's disease.     

Abeta deposition does not cause the aggregation of endogenous tau in transgenic mice

In Alzheimer disease, the extracellular deposition of beta-amyloid (Abeta) in the brain is accompanied by the intracellular accumulation of aggregated forms of hyperphosphorylated tau. In developing animal models of AD, the authors and others have been able to reproduce extracellular amyloid pathology in the brains of mice by expressing mutant amyloid precursor proteins (APP). The co-expression of APP with mutant presenilin leads to a dramatic acceleration in Abeta deposition, leading to very high amyloid burdens in mice. In the current study, the authors have examined whether the brains of mice with high burdens of amyloid deposition also contain aggregated forms of tau, using a cellulose acetate filter trap assay. Although discrete accumulations of phosphorylated tau immunoreactivity were apparent in neurites proximal to cored deposits of Abeta, little if any of this tau was in a SDS-resistant state of aggregation. By contrast, the brains of AD patients contained large amounts of aggregated tau. Overall, this study demonstrates that, in mice, deposition of Abeta does not cause endogenous tau to aggregate.     

A novel GSK-3β inhibitor reduces Alzheimer's pathology and rescues neuronal loss in vivo

Amyloid deposits, neurofibrillary tangles, and neuronal cell death in selectively vulnerable brain regions are the chief hallmarks in Alzheimer's (AD) brains. Glycogen synthase kinase-3 (GSK-3) is one of the key kinases required for AD-type abnormal hyperphosphorylation of tau, which is believed to be a critical event in neurofibrillary tangle formation. GSK-3 has also been recently implicated in amyloid precursor protein (APP) processing/Aβ production, apoptotic cell death, and learning and memory. Thus, GSK-3 inhibition represents a very attractive drug target in AD and other neurodegenerative disorders. To investigate whether GSK-3 inhibition can reduce amyloid and tau pathologies, neuronal cell death and memory deficits in vivo, double transgenic mice coexpressing human mutant APP and tau were treated with a novel non-ATP competitive GSK-3β inhibitor, NP12. Treatment with this thiadiazolidinone compound resulted in lower levels of tau phosphorylation, decreased amyloid deposition and plaque-associated astrocytic proliferation, protection of neurons in the entorhinal cortex and CA1 hippocampal subfield against cell death, and prevention of memory deficits in this transgenic mouse model. These results show that this novel GSK-3 inhibitor has a dual impact on amyloid and tau alterations and, perhaps even more important, on neuronal survival in vivo further suggesting that GSK-3 is a relevant therapeutic target in AD.     

Alzheimer's disease: beta-Amyloid protein and tau

Research on the molecular pathogenesis of Alzheimer's disease (AD) has made great strides over the last decade. This progress is the result of protein chemical analysis of two extracellular and intracellular fibrillary lesions in AD brain conducted during the 1980s, which identified beta-amyloid protein (A beta) and tau as their major components, respectively. Linkage analysis of familial AD identified four responsible genes: three causative genes (beta-amyloid precursor protein, presenilin 1, and presenilin 2) and one susceptibility gene (apolipoprotein E epsilon 4). All those genes causing and predisposing to AD exhibit a common phenotype: an increased production of A beta 42, a longer, more amyloidogenic A beta species, and/or its enhanced deposition. This observation was substantiated when presenilins were shown to be directly involved in A beta production. Whereas A beta deposition is relatively specific for AD, tau deposition is observed in various neurodegenerative diseases and is assumed to be intimately associated with neuronal loss. The genetic analysis of frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17) revealed the presence of mutations in the tau gene in affected members. Thus, tau can lead to intracellular tau deposits and neuronal loss, although the mechanism remains to be clarified. Taken together, A beta might exert neurotoxicity through tau, leading to neuronal loss in the AD brain.     

Localization and regional distribution of p23/TMP21 in the brain

Sequential processing of amyloid precursor protein by beta- and gamma-secretases generates Alzheimer's disease (AD)-associated beta-amyloid peptides. Recently it was reported that the transmembrane protein p23/TMP21 associates with gamma-secretase, and negatively regulates beta-amyloid production. Despite the link between p23 function and AD pathogenesis, the expression of p23 has not been examined in the brain. Here, we describe the detailed immunohistochemical characterization of p23 expression in rodent and human brain. We report that p23 is co-expressed with gamma-secretase subunits in select neuronal cell populations in rodent brain. Interestingly, the steady-state level of p23 in the brain is high during embryonic development and then declines after birth. Furthermore, the steady-state p23 levels are reduced in the brains of individuals with AD. We conclude that p23 is expressed in neurons throughout the brain and the decline in p23 expression during postnatal development may significantly contribute to enhanced beta-amyloid production in the adult brain.     

Does β-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.     

Neuropathological Characterization of Mutant Amyloid Precursor Protein Yeast Artificial Chromosome Transgenic Mice

Mutations in the amyloid precursor protein (APP) gene result in elevated production and deposition of the 42 amino acid beta-amyloid (Abeta1-42) peptide and early-onset Alzheimer's disease (AD). To accurately examine the effect of the APP FAD mutations in vivo, we introduced yeast artificial chromosomes (YACs) containing the entire genomic copy of human APP harboring FAD mutations into transgenic mice. Our current results demonstrate that mutant APP YAC transgenic mice exhibit many features characteristic of human AD, including regional deposition of Abeta with preferential deposition of Abeta1-42, extensive neuritic abnormalities as evidenced by staining with APP, ubiquitin, neurofilament, and hyperphosphorylated tau antibodies, increased markers of inflammation, and the overlapping deposition of Abeta with apolipoproteins E and J. Our results also suggest that APP YAC transgenic mice possess unique pathological attributes when compared to other transgenic mouse models of AD that may reflect the experimental design of each model.     

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 beta-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 beta-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 beta-amyloid (1-40) and (1-42) steadily increased with age, but significant deposition of fibrillary beta-amyloid in cortical areas did not occur before postnatal age of 10 months. The slope of increase in cerebral cortical beta-secretase (BACE1) activities in Tg2576 mice between ages of 9 and 13 months was significantly higher as compared to that of the alpha-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-1beta 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 beta-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 beta-amyloid, and potentially predispose to Alzheimer's disease.     

The molecular pathology of Alzheimer's disease

Neurofibrillary pathology in Alzheimer's disease consists of paired helical filaments comprising tau protein. This pathology is correlated with dementia, but can appear in the first two decades of life. Extracellular amyloid β-protein arises through proteolytic processing of a transmembrane precursor, which involves the action of several enzymes. Mutations in the genes for the precursor and presenilin proteins accelerate the deposition of Aβ. Tau mutations cause other tauopathies in the absence of amyloid deposition, indicating that amyloid deposition is not a prerequisite for dementia. An improved understanding of Alzheimer's disease awaits to be obtained by molecular imaging of these pathologies.     

Beta-amyloid precursor detected in human cerebral cortex

1. Amyloid deposition is one of the pathologic hallmarks of Alzheimer's disease. Since the isolation of the beta-amyloid gene, which revealed that the amyloid forming 4 kD protein is part of a larger precursor, interest has focused on the process by which amyloid is generated and deposited. 2. The authors have developed an immunologic means of detecting amyloid precursor proteins in human brain. 3. The method involves the expression of human beta-amyloid precursor cDNA in a recombinant vaccinia virus, so that antibodies are produced against the precursor proteins in their native forms. 4. By using this expression system, the amyloid precursor immunogens incorporate post-translational modifications that normally occur in vivo; this cannot be achieved with small synthetic peptides. 5. Using antibodies to the 695 residue amyloid precursor, we have detected using Western blot analysis a protein of approximately 120 kD in samples of cerebral cortex from three subjects with Alzheimer's disease and one control subject. 6. Additional antibodies to other amyloid-related proteins have been developed. These are being used to assess the differential expression of the various amyloid precursors and subdomains in additional cases.     

Cytokines in Alzheimer's disease.

1. Human astrocytoma cells produced biologically active interleukin-6 when treated with a variety of agents including bacterial lipopolysaccharides, viruses, and interleukin-1. 2. Both human recombinant IL-6 and IL-6 produced by stimulated astrocytes promoted differentiation of cultured neuronal cells and reduced survival time in culture. 3. Interleukin-6 and interleukin-1 stimulated the synthesis of the Alzheimer's disease beta-amyloid precursor protein. 4. Cytokines may be involved in stimulation of dystrophic neuritic sprouting, neuronal death, and amyloid deposition noted in the brains of Alzheimer's disease patients.     

Effects of Cerebrolysin on amyloid-beta deposition in a transgenic model of Alzheimer's disease

We investigated the potential mechanisms through which Cerebrolysin, a neuroprotective noothropic agent, might affect Alzheimer's disease pathology. Transgenic (tg) mice expressing mutant human (h) amyloid precursor protein 751 (APP751) cDNA under the Thy-1 promoter (mThy1-hAPP751) were treated for four weeks with this compound and analyzed by confocal microscopy to asses its effects on amyloid plaque formation and neurodegeneration. In this model, amyloid plaques in the brain are found much earlier (beginning at 3 months) than in other tg models. Quantitative computer-aided analysis with anti-amyloid-beta protein (A beta) antibodies, revealed that Cerebrolysin significantly reduced the amyloid burden in the frontal cortex of 5-month-old mice. Furthermore, Cerebrolysin treatment reduced the levels of A beta(1-42). This was accompanied by amelioration of the synaptic alterations in the frontal cortex of mThy1-hAPP751 tg mice. In conclusion, the present study supports the possibility that Cerebrolysin might have neuroprotective effects by decreasing the production of A beta(1-42) and reducing amyloid deposition.     

A calcium-stimulated serine protease from monkey brain degrades the beta-amyloid precursor protein.

Amyloid deposition, a histopathological feature of Alzheimer's disease brain, may be the underlying cause of this disease. The isolation of enzymes involved in both the normal and aberrant or alternative processing of the beta-amyloid precursor protein may lead to an understanding of how beta-protein, the major component of amyloid deposits, is formed in the brain parenchyma and vasculature of Alzheimer's disease patients and aged humans. As the same kind of deposits is also found in aged primates, the use of primates will undoubtedly help to understand the mechanisms of amyloid deposition, both spatially and temporally. Here we report the partial purification from adult monkey brain of a calcium-activated serine protease that is immunoreactive with antibodies against cathepsin G and is potentially involved in the abnormal degradation of the beta-amyloid precursor protein. Moreover, immunoreactivity with cathepsin G antibodies was localised to astrocytes in both adult and aged monkey cortex, suggesting that our protease may be expressed in astrocytes.     

Cerebral cortical amyloid protein precursor mRNA expression is similar in Alzheimer's disease and other neurodegenerative diseases.

The expression of 3 beta-amyloid protein precursor (APP) mRNAs (695, 751, and 770) in the cerebral cortex in Alzheimer's disease and other neurodegenerative diseases was analyzed by the S1 nuclease protection assay. We found no significant Alzheimer's disease-specific alteration of APP mRNA expression when compared to the other neurological diseases as controls. Since the expression of this mRNA was not correlated with amyloid deposition, it is possible that gliosis/neuronal loss may secondarily alter APP mRNA expression. However, the current study revealed no significant correlation between them.