Volume 11, Number 3, June 2007

Pages 257-260
Zaven S. Khachaturian
Obituary: Leon J. Thal, M.D.

Pages 261-274
K. Sathishkumar, Xiaochun Xi, Roy Martin, Rao M. Uppu
Cholesterol Secoaldehyde, an Ozonation Product of Cholesterol, Induces Amyloid Aggregation and Apoptosis in Murine GT1-7 Hypothalamic Neurons
Abstract: Aldehydic products from ozonation of cholesterol and peroxidation of phospholipids have been shown to accelerate aggregation of amyloid β (Aβ) in vitro. Here, we show that 3β-hydroxy-5-oxo-5,6-secocholestan-6-al (ChSeco), an ozonation product of cholesterol, induces Aβ aggregation, generation of reactive oxygen species (ROS), and cytotoxicity in murine GT1-7 hypothalamic neurons. The formation of Aβ aggregates in situ was dose-dependent at ChSeco concentrations ranging from 1 to 20 uM. The increase in insoluble Aβ aggregates at increasing concentrations of ChSeco was accompanied by a decrease in soluble Aβ as evidenced by Western blot analysis. The formation of ROS in neuronal cells was found to be dose- and time-dependent with the magnitude being higher at 20 uM compared to 10 uM ChSeco or untreated controls. The increase in ROS was associated with depletion of GSH. The cytotoxicity induced by ChSeco involved changes in phosphatidylserine translocation, DNA fragmentation, and caspase 3/7 activity that are characteristic of apoptosis. Pretreatment of neuronal cells with Trolox, a water-soluble analog of alpha-tocopherol offered partial, but significant protection against ChSeco-induced cell death, whereas N-acetyl-L-cysteine (NAC) completely prevented the cytotoxic effects of ChSeco. NAC and Trolox were without any effects on ChSeco-induced Aβ aggregation. Fibrillogenesis inhibitors, which inhibited Aβ aggregation, did not inhibit cell death induced by ChSeco, implying that ROS generation, and not Aβ aggregation, plays a major role in the observed cytotoxicity. However, since Alzheimer’s and other neurodegenerative diseases are slow and progressive, the formation of Aβ aggregates in vivo by ChSeco may have long- term pathological consequences.

Pages 275-290
Andrea Fuso, Rosaria A. Cavallaro, Alessandro Zampelli, Fabrizio D’Anselmi, Paola Piscopo, Annamaria Confaloni, Sigfrido Scarpa
Gamma-secretase is differentially modulated by alterations of Homocysteine cycle in neuroblastoma and glioblastoma cells
Abstract: Multiple aspects of homocysteine metabolism were studied to understand the mechanism responsible for hyperhomocysteinemia toxicity in Alzheimer disease. Besides oxidative stress and vascular damage, homocysteine has also a great importance in regulating DNA methylation through S-adenosylmethionine, the main methyl donor in eukaryotes. Alterations of S-adenosylmethionine and methylation were evidenced in Alzheimer disease and in elderly. In order to clarify whether DNA methylation can provide the basis for amyloid-β overproduction, we used human SK-N-BE neuroblastoma and A172 glioblastoma cell lines. We tested the effects of folate, B12 and B6 deprivation and S-adenosylmethionine addition on methylation metabolism. Our results indicate that homocysteine accumulation induced through vitamin B deprivation could impair the “Methylation Potential” with consequent presenilin 1, BACE and amyloid-β upregulation. Moreover, we found that homocysteine alterations had an effect on neuroblastoma but not on glioblastoma cells; this suggests a possible differential role of the two cell types in Alzheimer disease.

Pages 291-300
Orrie M. Friedman, Paul J. Matsudaira, Arthur H. Reis, Jr., Neil Simister, Ivan Correia, David Kew, Julie Y. Wei, Thomas Pochapsky
Substituted Organosiloxanes as Potential Therapeutics for Treatment and Prevention of Neurodegenerative Diseases
Abstract: Extensive testing of hydrolysates of commercially available organosilanes has identified a number of bifunctional organosiloxane compounds that show potential as therapeutics for treatment of diseases characterized by amyloid deposition such as Alzheimer’s disease (AD). All of these compounds protect from and/or reverse the metal-induced aggregation of amyloid Aß(1-42) peptide in dynamic light scattering (DLS) assays in trifluoroethanol (TFE) solutions, protect from and/or reverse the metal-induced loss of alpha-helical structure in TFE solutions of amyloid Aß (1-42) as measured by circular dichroism (CD), and are able to cross blood-brain barrier models at rates above background using Caco-2 and MDCK cell permeation assays. Based on these studies, we conclude that members of this class of bifunctional organosiloxanes are promising candidates for testing in treatment and/or prevention of AD and other diseases characterized by amyloid deposition.

Commentary
   Pages 301-302
   Christopher Exley
   Organosilicon therapy in Alzheimer's disease?

   Pages 303-304
   Orrie M. Friedman, Paul J. Matsudaira, Arthur H. Reis, Jr., Neil Simister, Thomas Pochapsky
   Authors' reply to "Organosilicon therapy in Alzheimer's disease?"

Pages 305-311
Lakshmi Thirumangalakudi, Linda Yin, Haripriya Vittal Rao, Paula Grammas
IL-8 induces expression of matrix metalloproteinases, cell cycle and pro-apoptotic proteins, and cell death in cultured neurons
Abstract: Neuronal cell loss is a critical feature of age-related neurodegenerative diseases such as Alzheimer’s disease (AD).  In the AD brain, a marked increase in pro-inflammatory cytokines and chemokines, including IL-8, has been documented. The objective of this study was to determine the effect of IL-8 on cell viability and expression of neurotoxic, apoptotic, and cell cycle proteins in cultured neurons. Incubation of cultured neurons with IL-8 for 24 h resulted in neuronal cell death. RT-PCR analysis of primary rat neuronal cultures treated with IL-8 for 24 h showed induction of genes for matrix metalloproteinases (MMP-2 and MMP-9), proinflammatory proteases with neurotoxic properties.  Gelatin zymography demonstrated IL-8 induced MMP-2 and MMP-9 activity. Western blot analysis showed that IL-8 also increased levels of the pro-apoptotic protein Bim (Bcl-2-interacting mediator of cell death). In addition, message levels of the cell cycle protein cyclin D1, an early marker for G1/S transition and a protein implicated as a regulator of neuronal apoptosis, were elevated after IL-8 exposure. These results suggest that IL-8 could be an important mediator of neuronal death in AD both via its effects on release of neurotoxins such as MMPs as well as by induction of cell cycle and pro-apoptotic proteins.

Pages 313-321
Sara M Debanne, Roger A Bielefeld, Vinay K Cheruvu, Thomas Fritsch, Douglas Y Rowland
Alzheimer’s Disease and Smoking: Bias in Cohort Studies
Abstract: The discrepancy between cohort and case-control studies regarding the association between smoking and Alzheimer’s disease (AD) has been attributed to the competing risk of early mortality of smokers. A simulation study was conducted to show that the bias favoring smokers acts also on cohort studies. In the model, individuals grow older and have smoking habits according to published year-age-gender-specific patterns, with morbidity and mortality according to their demographic and smoking profiles. Those individuals dying of smoking-related causes (“phantoms”) remain at risk of AD and of death from other causes. Three scenarios were considered: no association of AD and smoking, increased risk for smokers, and decreased risk for smokers. For each simulation of a cohort study, two incidence density ratios (IDR) were computed: one including the phantoms that developed AD (thus ignoring smoking-related deaths) and another excluding them (thus mimicking real-life studies). For all scenarios, the simulations show that smoking-related death creates a bias, resulting in smokers having an understated risk of AD compared to non-smokers. The speculation that the conflicting results of case-control and cohort studies are solely due to the increased mortality in smokers thus appears unjustified. Other factors must also be considered to explain the discrepancy in results.

Pages 323-335
Douglas L. Trenkle, William R. Shankle, Stanley P. Azen (Communicated by Marwan Sabbagh)
Detecting Cognitive Impairment in Primary Care: Performance Assessment of Three Screening Instruments
Abstract: Early detection of Alzheimer’s disease and related disorders (ADRD) is important, especially in primary care settings. We compared performances of two common screening tests, the Mini-Mental State Exam (MMSE) and Clock Drawing Test (CDT), with that of the MCI Screen (MCIS) in 254 patients over 65.  None had previous diagnosis of ADRD, and 81% were asymptomatic by Functional Assessment Staging Test (FAST) (FAST=1).  215 patients completed all screening tests -141 had ≥1 abnormal result, 121/141 completed standardized diagnostic assessment, and the remaining 74/215 (34%) screened entirely normally and weren’t further evaluated.  Potential bias due to unevaluated cases was statistically adjusted. Among diagnosed cases: AD=43%, cerebrovascular disease=36%, other causes=21%. Bias-adjusted MCI prevalence for FAST stages 1 and 1-3 were 13.9-20.3% and 23.0-28.3%. Bias-adjusted results for the CDT, MMSE and MCIS were: clinical diagnosis validity (kappa statistic) = {-0.02 (p=.61), 0.06 (p=.23), 0.92 (p<.0001)}; sensitivity = {59%, 71%, 94%}; specificity = {39%, 36%, 97%}; overall accuracy = {54%, 62%, 96%}; positive predictive value = {16%, 17%, 86%}; and negative predictive value = {83%, 87%, 96%}. The MMSE and CDT were not valid for early detection, while the MCIS had high validity and accuracy in the primary care cohort.

Pages 337-341
Grace J. Petot, Ursula Vega, Fatoumata Traore, Thomas Fritsch, Sara M. Debanne, Robert P. Friedland, Alan J. Lerner
Height and Alzheimer’s disease:  Findings from a case-control study
Abstract: The purpose of this study is to examine the relationship of height, Apolipoprotein E genotype (APOE) and Alzheimer’s disease (AD). Using a case-control design, subjects were recruited from the research registry of the University Memory and Aging Center of Case Western Reserve University and University Hospitals of Cleveland.  On entry to the study, Height was measured on 239 probable or possible AD patients and 341 healthy controls living in northeast Ohio.  Risk of AD was modeled as a function of quartile of height, APOE genotype, years of education and year of birth.  Analyses were stratified by gender.  For men, cases were more likely to be shorter when compared to controls (p = 0.001).  There was only a small difference in mean height between AD cases and controls among women (p = 0.05).  For men, height in the highest quartile [>179.7 cm (70.75 in)] had a 59% lower risk of developing AD that in the lowest quartile [<169.5 cm (66.75 in)], controlling for year of birth, and education (p = 0.03).  For women without an APOE epsilon4 allele, increasing height was associated with lower risk for AD (OR = 0.88; p = 0.01) but no significant association was found for women with at least one epsilon4 allele (OR = 1.03; p = 0.56).

Pages 343-358
Mithu Raychaudhuri, Debashis Mukhopadhyay
AICD and Its Adaptors – In Search of New Players
Abstract: In view of the emerging evidence that amyloid-β load in the brain and neuronal deficits are possibly independent events and the increasing importance of downstream molecular cascades in Alzheimer’s disease (AD) pathogenesis, the role of Amyloid Intracellular C-terminal Domain (AICD) is evaluated. This C-terminal fragment of amyloid-β protein precursor (AβPP) is cytotoxic and is a major component of AD brain. Different portions of AICD bind to different ‘adaptors’ and are seen to take part in various cellular events including AβPP processing and trafficking, apoptosis, neuronal growth and regulation of gene transcription. Phosphorylation also plays an important role in terms of choice of binding partners. The review emphasizes the dynamics of the network created by AICD interactions and points to possible alternative routs of AD like neurodegeneration.

Pages 359-370
Nicoletta Berardi, Chiara Braschi, Simona Capsoni, Antonino Cattaneo, Lamberto Maffei
Environmental enrichment delays the onset of memory deficits and reduces neuropathological hallmarks in a mouse model of Alzheimer-like neurodegeneration
Abstract: Alzheimer’s disease (AD) is a devastating neurodegenerative disorder characterized by progressive memory deficits and cognitive decline. We explored the possibility that Environmental Enrichment (EE) may reduce the disease progression in a comprehensive mouse model for AD like neurodegeneration, the AD11 mice. AD11 mice, which express anti nerve growth factor (NGF) antibodies, develop an age dependent neurodegeneration which encompasses all hallmarks of human AD. We have tested the efficacy of EE starting from 2 months of age, that is before the onset of behavioural deficits in AD11 mice. At 7 months of age, visual recognition memory was tested with the Object Recognition Test (ORT), spatial memory with the Morris Water Maze (MWM) and the presence of AD pathological hallmarks (Aβ clusters, presence of hyperphosphorylated tau and cholinergic deficit) was assessed immunohistochemically. We found that in AD11 mice exposed to EE from 2 to 7 months of age performance in both memory tests is significantly better than in non EE AD11 mice and indistinguishable from that in wt mice of the same age. Exposure to EE from 2 to 7 months significantly reduces the appearance of AD neuropathological hallmarks. A group of AD11 mice was tested also at 12 months of age: we found that 12 months old AD11 mice exposed to EE from 2 to 7 months of age performed significantly better than non EE AD11 mice of the same age and did not differ from 12 months old wt mice. Thus, EE is able to prevent the onset of memory deficits up to at least 12 months of age and to restrain the progression of neurodegeneration in a mouse model of AD.

Pages 371-383
Diana S. Woodruff-Pak, Alexis Agelan, Luis Del Valle (Communicated by Othman Ghribi)
A Rabbit Model of Alzheimer’s Disease: Valid at Neuropathological, Cognitive, and Therapeutic Levels
Abstract: Supplementing a rabbit’s diet with 2% cholesterol alone or with a trace amount of copper created neuropathological changes that resembled those seen in Alzheimer’s disease (AD). AD model rabbits were impaired in eyeblink classical conditioning; a form of learning severely impaired in AD. Our aim was to replicate AD rabbit model neuropathology, test eyeblink conditioning in this model, and determine if galantamine (Razadyne) would ameliorate impaired conditioning.  In Experiment 1 rabbit chow with 2% cholesterol and drinking water with 0.12 mg/liter copper sulfate were administered for 10 weeks. Control rabbits received normal food and water.  Rabbit brains were probed for neuropathology. AD model rabbits had significant neuronal loss in frontal cortex, hippocampus and cerebellum.  Changes in neurons in the hippocampus were consistent with neurofibrillary degeneration and cytoplasmic immunoreactivity for amyloid-β and tau. In Experiment 2 AD model rabbits were injected daily with vehicle or 3.0 mg/kg galantamine and tested on 750 ms trace and delay eyeblink conditioning.  Galantamine improved eyeblink conditioning significantly over vehicle.  The AD rabbit model has validity from neuropathological to cognitive levels and offers a promising addition to the available animal models of AD. Galantamine ameliorated impaired eyeblink conditioning, extending the validity of the AD rabbit model to treatment modalities.

Pages 385-397
HyeSook Youn, MyoungKun Jeoung, YongBum Koo, Hanlee Ji, William R. Markesbery, Inhae Ji, Tae H. Ji (Communicated by Mark Lovell)
Kalirin is under-expressed in Alzheimer’s Disease Hippocampus
Abstract: To identify genes aberrantly expressed in the brain of individuals with Alzheimer’s Disease (AD), we analyzed RNA extracts from the hippocampus and cerebellum from 19 AD patients and 15 age- and sex-matched control subjects. Our analysis identified a number of genes that were over-expressed or under-expressed specifically in AD hippocampus.  Among these genes, kalirin was the most consistently under-expressed in AD hippocampus, which was verified by semi-quantitative RT-PCR and real time PCR. Kalirin is predominantly expressed in the brain, particularly in the hippocampus, and plays crucial roles in neuronal stability and growth. Our observation is the first to relate kalirin to AD and a human disease. In addition to kalirin, the genes for voltage-gated Ca++ channel γ subunit 3 and visinin-like protein 1 (a Ca++ sensor protein) were under-expressed, whereas inositol 1,4,5-triphosphate 3-kinase B was over-expressed in AD hippocampus. Collectively, these differential expressions could severely impair calcium homeostasis. Remarkably, these aberrant gene expressions in AD hippocampus were not observed in AD cerebellum. Furthermore, housekeeping genes such as ribosomal protein genes are not affected by AD. These results provide new insights into the biochemistry of AD.

Pages 399-417
Discussion: Alzheimer Research Forum
Gain or Loss of Function—Time to Shake up Assumptions on γ-Secretase in Alzheimer Disease?

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