Mutations In Presenilin Genes Research Articles

Aberrant accentuation of neurofibrillary degeneration in the hippocampus of Alzheimer's disease with amyloid precursor protein 717 and presenilin-1 gene mutations

This study reports correlation of the hippocampal neurofibrillary tangles (NFT) density with beta-amyloid (Aβ) precursor protein (APP) 717 mutation, presenilin (PS)-1 mutation and apolipoprotein E (Apo-E) e4 alleles (E4), being graded as 3 forms (no-E4, one-E4 and two-E4) in autopsied brains from patients with familial and non-familial Alzheimer's disease (AD). We studied the density of NFT-free neurons, intracellular NFT (I-NFT), extracellular NFT (E-NFT) and total NFT (I-NFT plus E-NFT) in the six hippocampal subdivisions: cornu ammonis (CA) 1–CA4, subiculum and entorhinal cortex. The APP mutation cases showed significantly higher total NFT density in the CA1–CA2 region, and the PS-1 mutation cases also showed higher density of total NFT in the CA1–CA3 than non-familial cases. Moreover, high densities of the E-NFT contributed to these high total NFT densities. Non-familial AD cases showed a stereotypical NFT distribution with entorhinal accentuation in the hippocampus irrespective of E4 frequency. Thus, APP and PS-1 mutations predominantly affect the CA regions with profound neurodegeneration, which contributes early and severe clinical features of familial AD.

One novel presenilin-1 gene mutation in a Chinese pedigree of familial Alzheimer's disease

This study is to explore whether there is presenilin 1 (PS1) gene mutation in Chinese familial Alzheimer's disease (FAD). There has been no such systemic research before in China. Using polymerase chain reaction, single strand conformation polymorphism (PCR-SSCP), followed by denaturing high performance liquid chromatograph (DHPLC) and DNA sequencing, we analyzed a Chinese family with early onset AD. The patients in this family showed a novel missense mutation in exon 4 of the PS1 gene (G to T change in codon 97), altering valine to leucine acid substitution. Because the change occurred in conserved domains of this gene, and is not present in normal controls, this novel mutation is likely to be causative of Chinese FAD.

Somatostatin regulates brain amyloid β peptide Aβ42 through modulation of proteolytic degradation

Expression of somatostatin in the brain declines during aging in various mammals including apes and humans. A prominent decrease in this neuropeptide also represents a pathological characteristic of Alzheimer disease. Using in vitro and in vivo paradigms, we show that somatostatin regulates the metabolism of amyloid beta peptide (Abeta), the primary pathogenic agent of Alzheimer disease, in the brain through modulating proteolytic degradation catalyzed by neprilysin. Among various effector candidates, only somatostatin upregulated neprilysin activity in primary cortical neurons. A genetic deficiency of somatostatin altered hippocampal neprilysin activity and localization, and increased the quantity of a hydrophobic 42-mer form of Abeta, Abeta(42), in a manner similar to presenilin gene mutations that cause familial Alzheimer disease. These results indicate that the aging-induced downregulation of somatostatin expression may be a trigger for Abeta accumulation leading to late-onset sporadic Alzheimer disease, and suggest that somatostatin receptors may be pharmacological-target candidates for prevention and treatment of Alzheimer disease.

Clinical, Pathological, and Biochemical Spectrum of Alzheimer Disease Associated With PS-1 Mutations

Three genes have been implicated in the etiology of early-onset autosomal-dominant Alzheimer disease (AD): the amyloid precursor protein, the presenilin-1, and presenilin-2 genes. Approximately half of autosomal-dominant AD cases are associated with mutations in the presenilin-1 (PS-1) gene on the long arm of Chromosome 14. Marked allelic heterogeneity characterizes families with PS-1 gene mutations; more than 100 different mutations have been found in independent families thus far. With the exception of age at onset, the clinical phenotype is similar to late-onset AD, although some rare specific phenotypes have been described. These mutations lead to enhanced deposition of total Abeta and Abeta42 (but not Abeta40) in the brain, compared with sporadic AD. There is a considerable heterogeneity in the histological profiles among brains from patients with different mutations, and although some lead to predominantly parenchymal deposition of Abeta in the form of diffuse and cored plaques, others show predominantly vascular deposition, with severe amyloid angiopathy. Only some mutations are associated with enhanced neurofibrillary tangle formation and increased neuronal loss compared with sporadic AD. However, there is an important clinical and pathological variability even among family members with the same mutation, which suggests the involvement of other genetic or environmental factors that modulate the clinical expression of the disease. This represents a valuable model for identifying such factors and has potential implications for the development of new therapeutic strategies for delaying disease onset.

Mutations in presenilin 1, presenilin 2 and amyloid precursor protein genes in patients with early-onset Alzheimer's disease in Poland

Mutations in three causative genes have been identified in patients with an autosomal-dominant form of early-onset Alzheimer's disease (EOAD). To determine the spectrum of mutations in a group consisting of 40 Polish patients with clinically diagnosed familial EOAD and 1 patient with mild cognitive impairment (MCI) and family history of AD, we performed a screening for mutations in the presenilin 1 ( PSEN1), presenilin 2 ( PSEN2) and amyloid precursor protein ( APP) genes. Four previously recognized pathogenic mutations in PSEN1 gene (H163R, M139V) and APP gene (T714A, V715A), and three novel putative mutations in PSEN1 gene (P117R and I213F) and PSEN2 gene (Q228L) were identified. The 34 patients with no mutations detected were older than the patients with mutations. A frequency of APOE4 allele was higher in this group. Frequency of mutations is relatively low (17%), possibly due to used operational definition of a patient with familial EOAD (a patient having at least one relative with early-onset dementia). It could be concluded that screening for mutations in the three genes could be included in a diagnostic program directed at patients with a positive family history or age of onset before 55 years.

Transient hypoxia causes Alzheimer-type molecular and biochemical abnormalities in cortical neurons: potential strategies for neuroprotection.

Familial Alzheimer's Disease (AD) has been linked to amyloid beta protein precursor (AbetaPP) and presenilin gene mutations. In sporadic AD, which accounts for the vast majority of cases, the pathogenesis of neurodegeneration is unknown; however, recent evidence suggests a role for oxidative stress. The present study demonstrates that transient hypoxic injury to cortical neurons causes several of the molecular and biochemical abnormalities that occur in AD including, mitochondrial dysfunction, impaired membrane integrity, increased levels of DNA damage, reactive oxygen species, phospho-tau, phospho-MAP-1B, and ubiquitin immunoreactivity, and AbetaPP cleavage with accumulation of Abeta-immunoreactive products. These abnormalities were associated with activation of kinases that phosphorylate tau, including glycogen synthase kinase 3beta (GSK-3beta), mitogen-activated protein kinase (MAPK), and cyclin-dependent kinase 5 (Cdk-5). Further studies showed that significant neuro-protection with sparing of mitochondrial function and membrane integrity could be achieved by pre-treating the cortical neurons with N-acetyl cysteine, glutathione, or inhibitors of GSK-3beta, MAP kinase, or AbetaPP gamma-secretase. Therefore, in the absence of underlying gene mutations, oxidative stress can cause AD-type abnormalities, including aberrant post-translational processing of neuronal cytoskeletal proteins and APP. Our results also suggest that pre-treatment with agents that block specific components of the AD neurodegeneration cascade may provide neuroprotection against oxidative stress-induced impairments in membrane integrity, mitochondrial function, and viability.

COX-2 expression in brains of patients with familial Alzheimer's disease

Epidemiological evidence has demonstrated that nonsteroidal anti-inflammatory drugs (NSAIDs) may lower the risk of developing Alzheimer's disease (AD). Cyclooxygenase (COX)-2, a target enzyme of NSAIDs which is involved in inflammatory reactions as well as physiological brain functions, has been found to be upregulated in AD brains. In the present study, we used immunohistochemistry to determine the localization and the cellular type of the COX-2-expressing cells as well as the spatial relationship between these cells and the characteristic β-amyloid (Aβ 1–42)-immunopositive senile plaques (SP) or tau-positive neurofibrillary tangles (NFT) which are found in the brains of patients with a Presenilin-1 gene mutation. Numerous Aβ 1–42-positive SP were detected throughout the brains. COX-2 immunoreactivity was present between SP mainly in pyramidal neurons in the cerebral cortex and the hippocampal formation. COX-2 was also found in small round cells in close association with SP. The COX-2-positive neurons were most dense in the superficial cortical layers, while tau-positive NFT were densest in the deeper layers. In the frontal and temporal cortices that were most severely affected, the COX-2-positive neurons were damaged and decreased in number. The COX-2 expression in these cells might contribute to Aβ accumulation or represent a cytoprotective reaction against the Aβ-related inflammatory process.

Two suppressors of sel-12 encode C2H2 zinc-finger proteins that regulate presenilin transcription in Caenorhabditis elegans.

Mutations in presenilin genes are associated with familial Alzheimer's disease in humans and affect LIN-12/Notch signaling in all organisms tested so far. Loss of sel-12 presenilin activity in Caenorhabditis elegans results in a completely penetrant egg-laying defect. In screens for extragenic suppressors of the sel-12 egg-laying defect, we have isolated mutations in at least five genes. We report the cloning and characterization of spr-3 and spr-4, which encode large basic C(2)H(2) zinc-finger proteins. Suppression of sel-12 by spr-3 and spr-4 requires the activity of the second presenilin gene, hop-1. Mutations in both spr-3 and spr-4 de-repress hop-1 transcription in the early larval stages when hop-1 expression is normally nearly undetectable. As sel-12 and hop-1 are functionally redundant, this suggests that mutations in spr-3 and spr-4 bypass the need for one presenilin by stage-specifically de-repressing the transcription of the other. Both spr-3 and spr-4 code for proteins similar to the human REST/NRSF (Re1 silencing transcription factor/neural-restrictive silencing factor) transcriptional repressors. As other Spr genes encode proteins homologous to components of the CoREST co-repressor complex that interacts with REST, and the INHAT (inhibitor of acetyltransferase) co-repressor complex, our data suggest that all Spr genes may function through the same mechanism that involves transcriptional repression of the hop-1 locus.

The role of presenilin cofactors in the gamma-secretase complex.

Mutations in presenilin genes account for the majority of the cases of the familial form of Alzheimer's disease (FAD). Presenilin is essential for gamma-secretase activity, a proteolytic activity involved in intramembrane cleavage of Notch and beta-amyloid precursor protein (betaAPP). Cleavage of betaAPP by FAD mutant presenilin results in the overproduction of highly amyloidogenic amyloid beta42 peptides. gamma-Secretase activity requires the formation of a stable, high-molecular-mass protein complex that, in addition to the endoproteolysed fragmented form of presenilin, contains essential cofactors including nicastrin, APH-1 (refs 15-18) and PEN-2 (refs 16, 19). However, the role of each protein in complex formation and the generation of enzymatic activity is unclear. Here we show that Drosophila APH-1 (Aph-1) increases the stability of Drosophila presenilin (Psn) holoprotein in the complex. Depletion of PEN-2 by RNA interference prevents endoproteolysis of presenilin and promotes stabilization of the holoprotein in both Drosophila and mammalian cells, including primary neurons. Co-expression of Drosophila Pen-2 with Aph-1 and nicastrin increases the formation of Psn fragments as well as gamma-secretase activity. Thus, APH-1 stabilizes the presenilin holoprotein in the complex, whereas PEN-2 is required for endoproteolytic processing of presenilin and conferring gamma-secretase activity to the complex.

A novel mutation (L250V) in the presenilin 1 gene in a Japanese familial Alzheimer's disease with myoclonus and generalized convulsion

This study reports a novel presenilin 1 (PS1) gene mutation in a Japanese family with Alzheimer's disease (AD). Two patients developed progressive memory disorder with disorientation around 50 years of age and showed myoclonus with frequent tonic–clonic seizures several years later. Direct sequencing of the proband's PS1 gene revealed a novel mis-sense mutation (leucine-to-valine at residue 250 (L250V)). This mutation was found in both patients, but not in a normal family member or normal Japanese control subjects. Thus, L250V is a novel PS1 gene mutation responsible for familial AD (FAD) in Japan.

High sensitive detection of presenilin-1 point mutation based on electrochemiluminescence

Electrochemiluminescence (ECL) is a highsensitive detection method with broad biological applications. Ruthenium (II) tris (bipyridyl) (Ru(bpy) 3 2+ ) and tripropylamine (TPA) are most commomly used combination of the reactive species in ECL. The redox of Ru(bpy) 3 2+ with excess TPA at the surface of an electrode produces a highly efficient and stable light emission due to a rapid cycle of the reactions. This rapid, highly sensitive and accurate method has been applied to gene quantification. In this work, an ECL detection system is designed and applied in detection of presenilin-1 (PS-1) gene mutation. The results show that given the same polymerase chain reaction (PCR) cycle number, the ECL intensities from the digested and the undigested wild-type samples are nearly identical, but the difference in ECL intensities between the digested and the undigested mutant samples is distinctive. This detection technology connects PCR with ECL and allows a reliable discrimination between the wild-type and the mutant genes.

Oxidative stress and reduced antioxidant defenses in peripheral cells from familial Alzheimer's patients.

We have measured the levels of typical end products of the processes of lipid peroxidation, protein oxidation, and total antioxidant capacity (TAC) in skin fibroblasts and lymphoblasts taken from patients with familial Alzheimer's disease (FAD), sporadic Alzheimer's disease (AD), and age-matched healthy controls. Compared to controls, the fibroblasts and lymphoblasts carrying amyloid precursor protein (APP) and presenilin-1 (PS-1) gene mutations showed a clear increase in lipoperoxidation products, malondialdehyde (MDA), and 4-hydroxynonenal (4-HNE). In contrast, the antioxidant defenses of cells from FAD patients were lower than those from normal subjects. Lipoperoxidation and antioxidant capacity in lymphoblasts from patients affected by sporadic AD were virtually indistinguishable from the basal values of normal controls. An oxidative attack on protein gave rise to greater protein carbonyl content in FAD patients than in age-matched controls. Furthermore, ADP ribosylation levels of poly(ADP-ribose) polymerase (PARP) nuclear substrates were significantly raised, whereas the PARP content did not differ significantly between fibroblasts carrying gene mutations and control cells. These results indicate that peripheral cells carrying APP and PS-1 gene mutations show altered levels of oxidative markers even though they are not directly involved in the neurodegenerative process of AD. These results support the hypothesis that oxidative damage to lipid, protein, and DNA is an important early event in the pathogenesis of AD.

The Caenorhabditis elegans Presenilin sel-12 Is Required for Mesodermal Patterning and Muscle Function

Mutations in presenilin genes impair Notch signalling and, in humans, have been implicated in the development of familial Alzheimer's disease. We show here that a reduction of the activity of the Caenorhabditis elegans presenilin sel-12 results in a late defect during sex muscle development. The morphological abnormalities and functional deficits in the sex muscles contribute to the egg-laying defects seen in sel-12 hermaphrodites and to the severely reduced mating efficiency of sel-12 males. Both defects can be rescued by expressing sel-12 from the hlh-8 promoter that is active during the development of the sex muscle-specific M lineage, but not by expressing sel-12 from late muscle-specific promoters. Both weak and strong sel-12 mutations cause defects in the sex muscles that resemble the defects we found in lin-12 hypomorphic alleles, suggesting a previously uncharacterised LIN-12 signalling event late in postembryonic mesoderm development. Together with a previous study indicating a role of lin-12 and sel-12 during the specification of the π cell lineage required for proper vulva–uterine connection, our data suggest that the failure of sel-12 animals to lay eggs properly is caused by defects in at least two independent signalling events in different tissues during development.

Characterization of the amphioxus presenilin gene in a high gene-density genomic region illustrates duplication during the vertebrate lineage

Molecular studies on the pathology of Alzheimer's disease (AD) have revealed that mutations in presenilin genes ( PS) account for some familial cases. Although the contribution of these genes to the etiology is clear, their biological function remains obscure. Approaches using model organisms have been hampered by the fact that rodents contain two PS copies in the genome and do not develop the hallmark features associated with AD upon aging. To understand PS function and evolution, we have searched for PS homologous sequences in the genome of a lower chordate, Branchiostoma floridae. We report the structure of a single copy Branchiostoma floridae PS gene, named BfPS, and describe new features at the molecular level. Moreover, molecular phylogenetic analysis suggests that BfPS is orthologous to the vertebrate PS-1 and PS-2 forms. Finally, the analysis of more than 16 kb of genomic DNA encompassing BfPS identified three novel genes, which cluster with BfPS in a high gene-density region.

Presenilin-1-associated abnormalities in regional cerebral perfusion.

To investigate the influence of the presenilin-1 gene (PS-1) mutation on regional cerebral perfusion, SPECT was evaluated in 57 individuals. The subjects were members of a large pedigree from Colombia, South America, many of whom carry a PS-1 mutation for early-onset AD. Members of this large kindred who were cognitively normal and did not carry the PS-1 mutation (n = 23) were compared with subjects who were carriers of the mutation but were asymptomatic (n = 18) and with individuals with the mutation and a clinical diagnosis of AD (n = 16). Cerebral perfusion was measured in each subject using hexamethylpropyleneamine oxime SPECT. The data were analyzed in two ways: 1) Mean cerebral perfusion in each of 4320 voxels in the brain was compared among the groups using t-tests (t-maps); and 2) each individual received a weighted score on 20 vectors (factors), based on a large normative sample (n = 200), using a method known as singular value decomposition (SVD). Based on t-maps, subjects with the PS-1 mutation who were asymptomatic demonstrated reduced perfusion in comparison with the normal control subjects in the hippocampal complex, anterior and posterior cingulate, posterior parietal lobe, and anterior frontal lobe. The AD patients demonstrated decreased perfusion in the posterior parietal and superior frontal cortex in comparison with the normal control subjects. Discriminant function analysis of the vector scores derived from SVD (adjusted for age and gender) accurately discriminated 86% of the subjects in the three groups (p < 0.0005). Regional cerebral perfusion abnormalities based on SPECT are detectable before development of the clinical symptoms of AD in carriers of the PS-1 mutation.

Alzheimer's disease presenilin-1 expression modulates the assembly of neurofilaments.

Mutations in presenilin-1 gene are responsible for the majority of early-onset familial Alzheimer's disease cases. The function of this protein and the mechanism underlying the pathogenicity of its mutations are still unclear. To elucidate the role of presenilin-1 in the Alzheimer's disease pathology, we tested two such mutations (P117L and M146L) for their effect in stably transfected mouse neuroblastoma cell lines. Over-expression of the wild-type presenilin-1 gene induced formation of a well-extended, orderly organized network consisting of neurofilaments assembled from the L and H subunits, while in cells with the mutant gene this network was markedly reduced to short filaments concentrated in structures resembling cups. Cells expressing the mutant gene displayed altered processing of the transgene protein and neurofilament-H, suggesting that presenilin-1 is the mediator of changes targeted at neurofilaments. The two different mutations produced similar alterations, implying that this is a common pathogenic mechanism. Presenilin-1, neurofilament-H and tau proteins showed co-localization as evidenced by confocal microscopy, suggesting a possible physiological connection between these three proteins. Presenilin-1 appears to influence assembly of the H subunit into neurofilaments and the subsequent formation of new neurites. Mutations impair this function of presenilin-1, resulting in inhibition of neurite outgrowth. That presenilin-1 influences the assembly of neurofilaments may represent a novel pathway through which presenilin-1 mutations are involved in Alzheimer's disease pathology. In this hypothesis, presenilin-1 mutations will be associated with aberrant sprouting leading to synaptic loss, a key neuropathological feature of Alzheimer's disease.

Presenilin-1 Mutations Reduce Cytoskeletal Association, Deregulate Neurite Growth, and Potentiate Neuronal Dystrophy and Tau Phosphorylation

Mutations in presenilin genes are linked to early onset familial Alzheimer's disease (FAD). Previous work in non-neuronal cells indicates that presenilin-1 (PS1) associates with cytoskeletal elements and that it facilitates Notch1 signaling. Because Notch1 participates in the control of neurite growth, cultured hippocampal neurons were used to investigate the cytoskeletal association of PS1 and its potential role during neuronal development. We found that PS1 associates with microtubules (MT) and microfilaments (MF) and that its cytoskeletal association increases dramatically during neuronal development. PS1 was detected associated with MT in the central region of neuronal growth cones and with MF in MF-rich areas extending into filopodia and lamellipodia. In differentiated neurons, PS1 mutations reduced the interaction of PS1 with cytoskeletal elements, diminished the nuclear translocation of the Notch1 intracellular domain (NICD), and promoted a marked increase in total neurite length. In developing neurons, PS1 overexpression increased the nuclear translocation of NICD and inhibited neurite growth, whereas PS1 mutations M146V, I143T, and deletion of exon 9 (D9) did not facilitate NICD nuclear translocation and had no effect on neurite growth. In cultures that were treated with amyloid beta (Abeta), PS1 mutations significantly increased neuritic dystrophy and AD-like changes in tau such as hyperphosphorylation, release from MT, and increased tau protein levels. We conclude that PS1 participates in the regulation of neurite growth and stabilization in both developing and differentiated neurons. In the Alzheimer's brain PS1 mutations may promote neuritic dystrophy and tangle formation by interfering with Notch1 signaling and enhancing pathological changes in tau.

Presenilin-1 P264L Knock-In Mutation: Differential Effects on Aβ Production, Amyloid Deposition, and Neuronal Vulnerability

The pathogenic mechanism linking presenilin-1 (PS-1) gene mutations to familial Alzheimer's disease (FAD) is uncertain, but has been proposed to include increased neuronal sensitivity to degeneration and enhanced amyloidogenic processing of the beta-amyloid precursor protein (APP). We investigated this issue by using gene targeting with the Cre-lox system to introduce an FAD-linked P264L mutation into the endogenous mouse PS-1 gene, an approach that maintains normal regulatory controls over expression. Primary cortical neurons derived from PS-1 homozygous mutant knock-in mice exhibit basal neurodegeneration similar to their PS-1 wild-type counterparts. Staurosporine and Abeta1-42 induce apoptosis, and neither the dose dependence nor maximal extent of cell death is altered by the PS-1 knock-in mutation. Similarly, glutamate-induced neuronal necrosis is unaffected by the PS-1P264L mutation. The lack of effect of the PS-1P264L mutation is confirmed by measures of basal- and toxin-induced caspase and calpain activation, biochemical indices of apoptotic and necrotic signaling, respectively. To analyze the influence of the PS-1P264L knock-in mutation on APP processing and the development of AD-type neuropathology, we created mouse lines carrying mutations in both PS-1 and APP. In contrast to the lack of effect on neuronal vulnerability, cortical neurons cultured from PS-1P264L homozygous mutant mice secrete Abeta42 at an increased rate, whereas secretion of Abeta40 is reduced. Moreover, the PS-1 knock-in mutation selectively increases Abeta42 levels in the mouse brain and accelerates the onset of amyloid deposition and its attendant reactive gliosis, even as a single mutant allele. We conclude that expression of an FAD-linked mutant PS-1 at normal levels does not generally increase cortical neuronal sensitivity to degeneration. Instead, enhanced amyloidogenic processing of APP likely is critical to the pathogenesis of PS-1-linked FAD.

Alzheimer's disease presenilin-1 exon 9 deletion and L250S mutations sensitiZe SH-SY5Y neuroblastoma cells to hyperosmotic stress-induced apoptosis

Mutations in the presenilin-1 (PS1) and presenilin-2 (PS2) genes account for the majority of early-onset familial Alzheimer's disease cases. Recent studies suggest that presenilin gene mutations predispose cells to apoptosis by mechanisms involving altered calcium homeostasis and oxidative damage. In the present study, we determined whether PS1 mutations also sensitize cells to hyperosmotic stress-induced apoptosis. For this, we established SH-SY5Y neuroblastoma cell lines stably transfected with wild-type PS1 or either the PS1 exon 9 deletion (ΔE9) or PS1 L250S mutants. Cultured cells were exposed to an overnight (17 h) serum deprivation, followed by a 30 min treatment with either 20 mM glucose, 10 nM insulin-like growth factor-1 or 20 mM glucose +10 nM insulin-like growth factor-1. Cells were then cultured for a further 3, 6 or 24 h and stained for apoptotic condensed nuclei using propidium iodide. Confirmation that cells were undergoing an active apoptotic process was achieved by labelling of DNA strand breaks using the terminal dUTP nick end labelling (TUNEL) technique. We also determined cell viability using 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction. Propidium iodide staining revealed that all cell lines and controls showed an increased number of apoptotic cells appearing with condensed nuclei at 24 h compared with 6 h and 3 h. High glucose-induced hyperosmotic stress resulted in significantly more apoptotic cells in the PS1 ΔE9 and PS1 L250S mutation cell lines at 24 h, compared with the wild-type PS1 lines ( P<0.001, ANOVA for both comparisons). Mean values (±S.D.) for the percentage number of apoptotic cells at 24 h following high glucose treatment were 16.1±3.5%, 26.7±5.5% and 31.0±5.7% for the wild-type PS1, PS1 ΔE9 and PS1 L250S lines, respectively. The pro-apoptotic effects of high glucose treatment were reversed by 10 nM insulin-like growth factor-1, although to a lesser extent in the mutation cell lines (5.8±2.4%, 15.2±7.3% and 13.2±2.0% for the wild-type PS1, PS1 ΔE9 ( P<0.01 for comparison with wild-type PS1) and PS1 L250S ( P<0.01 for comparison with wild-type PS1) transfected lines, respectively. TUNEL labelling of cells at 24 h following treatment gave essentially the same results pattern as obtained using propidium iodide. The percentage number of apoptotic cells with DNA strand breaks (means±S.D.) following high glucose treatment was 15.4±2.6% for the wild-type PS1, 26.8±3.2% for the PS1 ΔE9 ( P<0.001 for comparison with wild-type PS1) and 29.7±6.1% for the PS1 L250S transfected lines ( P<0.001 for comparison with wild-type PS1). The PS1 ΔE9 and PS1 L250S transfected lines also showed a higher number of apoptotic cells with DNA strand breaks at 24 h following high glucose plus insulin-like growth factor-1 treatment (11.4±2.0% and 14.3±2.8%, respectively), compared with values for the wild-type PS1 lines (8.5±2.4%). These differences were significant ( P<0.01) for the comparision of wild-type PS1 and PS1 L250S, but not PS1 ΔE9 lines. The mutation-related increases in number of apoptotic cells at 24 h following high glucose treatment were not accompanied by significant differences in cell viability at this time-point. Our results indicate that PS1 mutations predispose to hyperosmotic stress-induced apoptosis and that the anti-apoptotic effects of insulin-like growth factor-1 are compromised by these mutations. Perturbations of insulin-like growth factor-1 signalling may be involved in PS1 mutation-related apoptotic neuronal cell death in Alzheimer's disease.

Effects of amyloid precursor protein derivatives andoxidative stress on basal forebrain cholinergic systems inALZHEIMERS disease

The dysfunction and degeneration of cholinergic neuronal circuits in the brain is aprominent feature of Alzheimers disease. Increasing data suggest that age-related oxidative stresscontributes to degenerative changes in basal forebrain cholinergic systems. Experimental studieshave shown that oxidative stress, and membrane lipid peroxidation in particular, can disruptmuscarinic cholinergic signaling by impairing coupling of receptors to GTP-binding proteins.Altered proteolytic processing of the β-amyloid precursor protein (APP) may contributeto impaired cholinergic signaling and neuronal degeneration in at least two ways. First, levels ofcytotoxic forms of amyloid β-peptide (Aβ) are increased ; Aβdamages and kills neurons by inducing membrane lipid peroxidation resulting in impairment ofion-motive ATPases, and glucose and glutamate transporters, thereby rendering neuronsvulnerable to excitotoxicity. The latter actions of Aβ may be mediated by4-hydroxynonenal, an aldehydic product of membrane lipid peroxidation that covalently modifiesand inactivates the various transporter proteins. Subtoxic levels of Aβ can also suppresscholine acetyltransferase levels, and may thereby promote dysfunction of intact cholinergiccircuits. A second way in which altered APP processing may endanger cholinergic neurons is byreducing levels of a secreted form of APP which has been shown to modulate neuronalexcitability, and to protect neurons against excitotoxic, metabolic and oxidative insults. Mutationsin presenilin genes, which are causally linked to many cases of early-onset inherited Alzheimersdisease, may increase vulnerability of cholinergic neurons to apoptosis. The underlying mechanismappears to involve perturbed calcium regulation in the endoplasmic reticulum, which promotesloss of cellular calcium homeostasis, mitochondrial dysfunction and oxyradical production.Knowledge of the cellular and molecular underpinnings of dysfunction and degeneration ofcholinergic circuits is leading to the development of novel preventative and therapeuticapproaches for Alzheimers disease and related disorders.