Volume 21, Number 4, October 2010

Pages 1037
Alexandre de Mendonça and George Perry
Obituary: Edgar da Cruz e Silva 1958-2010

Pages 1039-1063
Mak Adam Daulatzai
Conversion of Elderly to Alzheimer’s Dementia: Role of Confluence of Hypothermia and Senescent stigmata—the Plausible Pathway
Abstract: Aging is a consequence of progressive decline in special and somatosensory functions and specific brain stem nuclei. Many senescent stigmata, including hypoxia, hypoxemia, depressed cerebral blood flow and glucose metabolism, diseases of senescence, and their medications all enhance hypothermia as do alcohol, cold environment, and malnutrition. Hypothermia is a critical factor having deleterious impact on brain stem and neocortical functions. Additionally, anesthesia in elderly also promotes hypothermia; anesthetics not only cause consciousness (sensory and motor) changes, but memory impairment as well. Anesthesia inhibits cholinergic pathways, reticular and thalamocortical systems, cortico-cortical connectivity, and causes post-operative delirium and cognitive dysfunction. Increasing evidence indicates that anesthetic exposures may contribute to dementia onset and Alzheimer’s disease (AD) in hypothermic elderly. Inhaled anesthetics potentiate caspases, BACE, tau hyperphosphorylation, and apoptosis. This paper addresses the important question: “Why do only some elderly fall victim to AD”? Based on information on the pathogenesis of early stages of cognitive dysfunction in elderly (i.e., due to senescent stigmata), and the effects of anesthesia superimposed, a detailed plausible neuropathological substrate (mechanism/pathway) is delineated here that reveals the possible cause(s) of AD. Basically, it encompasses several risk factors for cognitive dysfunction during senescence plus several hypothermia-enhancing routes; they all converge and tip the balance towards dementia onset. This knowledge of the confluence of heterogeneous risk factors in perpetuating dementia relentlessly is of importance in order to: (a) avoid their convergence; (b) take measures to stop/reverse cognitive dysfunction; and (c) to develop therapeutic strategies to enhance cognitive function and attenuate AD.

Pages 1065-1075
Yan-Jiang Wang, Hua-Dong Zhou, Xin-Fu Zhou
Modified Immunotherapies Against Alzheimer’s Disease: Toward Safer and Effective Amyloid-β Clearance
Abstract: Alzheimer’s disease (AD) is characterized by the deposition of amyloid plaques, loss of neurons, neuritic degeneration, accumulation of fibrillary tangles in neurons, and a progressive loss of cognitive function. Amyloid-β peptide (Aβ) appears to play a pivotal role in the development of AD. Clearance of Aβ from the brain represents an important therapeutic strategy for prevention and treatment of AD. Immunotherapy targeting Aβ is effective to remove the peptide from the brain. However, it is associated with detrimental adverse effects, such as autoimmune meningoencephalitis and microhemorrhage. These are presumably the result of brain infiltration of provoked autoimmune T lymphocytes in response to Aβ vaccination and release of proinflammatory cytokines from microglia activated by the immune complex of Aβ and antibodies. An improvement of the safety of the immunotherapy is a major goal of the immunotherapy study. Here, we review the mechanisms involved in modified immunological strategies, as well as their adverse effects. We discuss the following: the development of B epitope vaccines to avoid activation of autoimmune T lymphocytes; DNA vaccines containing appropriate immunostimulatory and immunomodulatory sequences to induce the desired humoral immune responses; antibody modifications to avoid activation of microglia and subsequent release of proinflammatory cytokines; single chain antibody-based gene therapy; immunotherapy targeting Aβ oligomers; modulation of antibody delivery approach and dose; and application of autoantibodies against Aβ. These ultimately represent future directions of therapeutic approaches toward safer and effective Aβ clearance.

Pages 1077-1087
Pedro J Modrego
Depression in Alzheimer’s Disease. Pathophysiology, Diagnosis, and Treatment.
Abstract: Depression is a comorbid condition in Alzheimer’s disease (AD) with negative consequences in patients and caregivers. Pathophysiology and optimal treatment are matters to be elucidated. A search of articles dealing with depression in AD was conducted in MEDLINE with special attention to epidemiology, pathophysiology, and treatment. Depression may predate dementia and tends to occur in up to 50% of AD patients with a decrease of noradrenalin and serotonin in the brain being the most plausible cause. Only 7 small double-blind randomized placebo-controlled clinical trials with antidepressants in AD patients with depression were found: 4 with sertraline, 1 with fluoxetine, 1 with imipramine, and another one with clomipramine. The total number of treated patients was 318. The weighted odds ratio (OR) was calculated with the method of Mantel-Haenszel. Both tricyclic antidepressants and selective serotonin reuptake inhibitors are better than placebo in treating depression in AD (weighted OR: 1.82, 95% CI: 1.13-2.96), with sertraline being one of the most used drugs. The differences were significant in 2 trials and not significant in four. The magnitude of effect is globally modest. Moreover, it is noteworthy mentioning the high rates of response to placebo in most studies. Depression is one of the most frequent behavioral symptoms in AD. Although antidepressants may work in AD, given the small number of patients treated, the effect is unclear. Further large randomized controlled clinical trials are warranted in order to know the best drug to begin with and the actual degree of efficacy.

Pages 1089-1099
Catherine H. Trepanier, Norton W. Milgram
Neuroinflammation in Alzheimer’s Disease: Are NSAIDs and Selective COX-2 Inhibitors the Next Line of Therapy?
Abstract: Alzheimer’s disease (AD) is currently treated with cholinergic and glutamatergic therapies, which provide symptomatic benefit but do not reverse the underlying pathology or cognitive deficits. The prevalence of AD is expected to triple over the next 50 years, creating an urgency to develop effective "disease-modifying" therapies to reduce the economic burden of this devastating disorder. One of the main areas of therapeutic focus has been an anti-inflammatory strategy based on an inflammatory hypothesis of AD. This hypothesis originated from epidemiological evidence that long-term exposure to nonsteroidal anti-inflammatory drugs (NSAIDs) protected against the development of AD. However, large-scale double-blind placebo-controlled clinical trials have not supported the use of NSAIDS in treating AD. The following review outlines epidemiological, preclinical, and clinical evidence evaluating the efficacy of various NSAIDs and selective COX-2 inhibitors in AD. We also review recent anecdotal data with the TNF-α inhibitor, etanercept, and discuss possible explanations for the failure of preclinical data to translate into successful clinical trials.

Pages 1101-1105
Short Communication
Emma H. Gray, Kurt J. De Vos, Colin Dingwall, Michael S. Perkinton, Christopher C.J. Miller (Handling Associate Editor: Thomas Shea)
Deficiency of the Copper Chaperone for Superoxide Dismutase Increases Amyloid-β Production
Abstract: The copper chaperone for superoxide dismutase (CCS) binds to both the β-site AβPP cleaving enzyme (BACE1) and to the neuronal adaptor protein X11α. BACE1 initiates AβPP processing to produce the amyloid-β (Aβ) peptide deposited in the brains of Alzheimer’s disease patients. X11α also interacts directly with AβPP to inhibit Aβ production. However, whether CCS affects AβPP processing and Aβ production is not known. Here we show that loss of CCS increases Aβ production in both CCS knockout neurons and CCS siRNA-treated SHSY5Y cells and that this involves increased AβPP processing at the BACE1 site.

Pages 1107-1117
Zheng-Qi Fu, Ying Yang, Jie Song, Qian Jiang, Zan-Chao Liu, Qun Wang, Ling-Qiang Zhu, Jian-Zhi Wang and Qing Tian
LiCl Attenuates Thapsigargin-Induced Tau Hyperphosphorylation by Inhibiting GSK-3β In Vivo and In Vitro
Abstract: Abnormal hyperphosphorylation of microtubule-associated protein tau is involved in the pathogenesis of several neurodegenerative diseases, including Alzheimer’s disease (AD). Endoplasmic reticulum (ER) stress is indicated to play an important role in neurodegeneration and activation of glycogen synthase kinase-3β (GSK-3β), an integral kinase in tau phosphorylation. To explore the effect of ER stress on tau phosphorylation, we treated cultured cells (HEK293 and SH-SY5Y cells) and rat brain with thapsigargin, an ER stress inducer. We found that the phosphorylation level of tau was significantly increased after thapsigargin treatment. By using a cell-free reconstitution system, we also observed that co-culture of the thapsigargin-treated ER fraction from HEK293/wt (without tau) with cytoplasm prepared from HEK293/tau induced an increased tau phosphorylation. Concurrently, activation of GSK-3β as evidenced by an increased phospho-GSK-3β at Tyr-216 and decreased phospho-GSK-3β at Ser-9 both in vitro and in vivo was detected. Application of lithium chloride, a GSK-3 inhibitor, could efficiently attenuate the thapsigargin-induced tau hyperphosphorylation with suppressed activation of GSK-3β in cell cultures and rat brains. Our data provide further evidence supporting the role of ER stress in tau hyperphosphorylation and the protective role of lithium.

Pages 1119-1128
Joakim Hertze, Lennart Minthon, Henrik Zetterberg, Eugeen Vanmechelen, Kaj Blennow, Oskar Hansson (Handling Associate Editor: Mony de Leon)
Evaluation of CSF Biomarkers as Predictors of Alzheimer’s Disease: a Clinical Follow-Up Study of 4.7 Years
Abstract: In this study, we determined the diagnostic accuracy of cerebrospinal fluid (CSF) biomarkers to predict development of Alzheimer’s disease (AD) within five years in patients with mild cognitive impairment (MCI). To do so, the levels of tau, phosphorylated tau, Aβ42, Aβ40, Aβ38, sAβPPα, and sAβPPβ were analyzed in 327 CSF samples obtained at baseline from patients with AD (n=94), MCI (n=166), depressive disorder (n=29), and cognitively healthy controls (n=38). In the cohort with MCI at baseline, 33% subsequently developed AD and 16% developed other types of dementia; however, 51% were still cognitively stable after a follow-up of 4.7 years (range 3.0-7.4). Optimal cut-offs for each biomarker or combinations of biomarkers were defined in the AD, control, and depressive disorder groups. Several combinations resulted in sensitivity and specificity levels >85% for differentiation of AD from controls and depressive disorder. Using the previously established cut-offs, a combination of Aβ42 and tau could predict future development of AD in MCI patients with a sensitivity of 88%, specificity 82%, positive predictive value 71%, and negative predictive value 94%. MCI patients with both low Aβ42 and high tau levels had a substantially increased risk of developing AD (OR 20; 95% CI 6-58), even after adjustment for confounding factors. Ultimately, CSF biomarkers can stratify MCI patients into those with very low or high risk for future development of AD. However, the specificities and positive predictive values are still too low to be able to diagnose AD before the patients fulfill the clinical criteria.

Pages 1129-1140
Michael J. Firbank, Andrew M. Blamire, Andrew Teodorczuk, Emma Teper, Emma J. Burton, Dipayan Mitra, John T. O’Brien (Handling Associate Editor: Marco Bozzali)
High Resolution Imaging of the Medial Temporal Lobe in Alzheimer’s Disease and Dementia with Lewy Bodies
Abstract: We used high resolution (0.3mm in-plane) coronal 3T magnetic resonance (MR) imaging of the medial temporal lobe in 16 subjects with Alzheimer’s disease (AD), 16 with dementia with Lewy bodies (DLB), and 16 similarly aged healthy subjects. On the anterior section of the hippocampus body, regions of interest were manually drawn blind to diagnosis on the CA1, CA2, and CA3/4 subregions, and the width of the subiculum and entorhinal cortex was measured. Controlling for intracranial volume, age, and years of education, we found the subiculum thickness was significantly reduced in AD (2.03 ± 0.29 mm) compared to both control (2.37 ± 0.28 mm, p = 0.008) and DLB (2.35 ± 0.24 mm, p = 0.001) subjects. The area of CA1 was likewise reduced in AD compared to controls and DLB. In the hippocampus images, a hypointense line is visible between CA1 and CA3/4. This line was significantly less distinct in AD, suggesting disease related changes to this region. Future studies should investigate whether subiculum thickness or the hypointense line could be a diagnostic feature to help discriminate AD from DLB.

Pages 1141-1151
Valtteri Julkunen, Eini Niskanen, Juha Koikkalainen, Sanna-Kaisa Herukka, Maija Pihlajamäki, Merja Hallikainen, Miia Kivipelto, Sebastian Muehlboeck, Alan C. Evans, Ritva Vanninen, Hilkka Soininen
Differences in Cortical Thickness in Healthy Controls, Subjects with Mild Cognitive Impairment, and Alzheimer’s Disease Patients: A Longitudinal Study
Abstract: In this study,we analyzed differences in cortical thickness (CTH) between healthy controls (HC), subjects with stable mild cognitive impairment (S-MCI), progressive MCI (P-MCI), and Alzheimer’s disease (AD), and assessed correlations between CHT and clinical disease severity, education, and apolipoprotein E4 (APOE) genotype.Automated CTH analysis was applied to baseline high-resolution structural MR images of 145 subjects with a maximum follow-up time of 7.4 years pooled from population-based study databases held in the University of Kuopio. Statistical differences in CTH between study groups and significant correlations between CTH and clinical and demographic factors were assessed and displayed on a cortical surface model. Compared to HC group (n = 26), the AD (n = 21) group displayed significantly reduced CTH in several areas of frontal and temporal cortices of the right hemisphere. Higher education and lower MMSE scores were correlated with more reduced CTH in the AD group, whereas no significant correlation was found between CDR-SB scores or APOE genotype and CTH. The P-MCI group demonstrated significantly reduced CTH compared to S-MCI in frontal, temporal and parietal cortices even after statistically adjusting for all confounding variables. Ultimately, analysis of CTH can be used to detect cortical thinning in subjects with progressive MCI several years before conversion and clinical diagnosis of AD dementia, irrespective of their cognitive performance, education level, or APOE genotype.

Pages 1153-1164
András Palotás, Helton J. Reis, Gábor Bogáts, Barna Babik, Mihály Racsmány, Linda Engvau, Éva Kecskeméti, Anna Juhász, Luciene B. Vieira, Antônio L. Teixeira, Marat A. Mukhamedyarov, Albert A. Rizvanov, Mehmet E. Yalvaç, Melissa M. Guimarães, Cláudia N. Ferreira, Andrey L. Zefirov, Andrey P. Kiyasov, Lan Wang, Zoltán Janka, János Kálmán
Coronary Artery Bypass Surgery Provokes Alzheimer’s Disease-Like Changes in the Cerebrospinal Fluid
Abstract: Several biomarkers are used in confirming the diagnosis of cognitive disorders. This study evaluates whether the level of these markers after heart surgery correlates with the development of cognitive dysfunction, which is a frequent complication of cardiac interventions. Concentrations of amyloid-β peptide, tau, and S100β in the cerebro-spinal fluid were assessed, as well as cognitive functions were evaluated before and after coronary artery bypass grafting, utilizing immuno-assays and psychometric tests, respectively. A drastic rise in the level of S100β was observed one week after the surgery, a mark of a severe generalized cerebral injury. The level of amyloid-β peptide significantly decreased, whereas the concentration of tau markedly increased six months postoperatively. Gradual cognitive decline was also present. These findings clearly demonstrate post-surgical cognitive impairment associated with changes in biomarkers similar to that seen in Alzheimer’s disease, suggesting a unifying pathognomic factor between the two disorders. A holistic approach to coronary heart disease and Alzheimer’s-type dementia is proposed.

Pages 1165-1177
Inês Baldeiras, Isabel Santana, Maria Teresa Proença, Maria Helena Garrucho, Rui Pascoal, Ana Rodrigues, Diana Duro, Catarina Resende Oliveira
Oxidative Damage and Progression to Alzheimer’s Disease in Patients with Mild Cognitive Impairment
Abstract: Recent studies show that most of the oxidative changes found in Alzheimer’s disease (AD) are already present in Mild cognitive impairment (MCI) patients. The question arises as to whether oxidative stress has a role in the progression of MCI to AD. We conducted a longitudinal study on 70 MCI patients, and the peripheral blood levels of a broad spectrum of non-enzymatic and enzymatic antioxidant defenses, as well as lipid and protein oxidation markers and nitrogen oxidative species were determined. At baseline, there were no differences in any of the indexes of oxidative damage between stable MCI patients (MCI-MCI) and patients that progressed to AD (MCI-AD). Cellular levels of lipid peroxidation markers increased in both groups and this was accompained in MCI-AD, but not in MCI-MCI patients, by a significant decrease in cellular antioxidant defenses (oxidyzed/reduced glutathione ratio and vitamin E). Among MCI-AD patients, the longitudinal decrease in cellular vitamin E was associated with the deterioration in cognitive performance. These results suggest that accumulation of oxidative damage may start in pre-symptomatic phases of AD pathology and that progression to AD might be related to depletion of antioxidant defenses.

Pages 1179-1183
Francesco Panza, Vincenzo Solfrizzi, Davide Seripa, Bruno P. Imbimbo, Alberto Pilotto, Vincenza Frisardi
Peripheral Antioxidant Markers in Mild Cognitive Impairment and its Progression to Dementia
Abstract: Mild cognitive impairment (MCI) is recognized as a prodromal phase of dementing disorders, and it has been suggested that oxidative stress may play a role in the pathogenesis of Alzheimer’s disease (AD), and in predicting progression of MCI to AD. In the present article, on the basis of increasing body of evidence from cross-sectional and longitudinal studies, we discussed the issue of the possible impact of antioxidant compounds from diet and supplementation on MCI and its progression to AD.

Pages 1185-1195
Julie Blanchard*, Muhammad Omar Chohan*, Bin Li, Fei Liu, Khalid Iqbal, Inge Grundke-Iqbal (Handling Associate Editor: Michal Novak) *These authors have equally contributed to this work
Beneficial Effect of a CNTF Tetrapeptide on Adult Hippocampal Neurogenesis, Neuronal Plasticity, and Spatial Memory in Mice
Abstract: A therapeutic strategy against cognitive disorders like Alzheimer’s disease is to take advantage of the regenerative ability of the brain and the properties of neurotrophic factors to shift the balance from neurodegeneration to neurogenesis and neuronal plasticity. Although the ciliary neurotrophic factor (CNTF) has some of the required neuroprotective characteristics, its clinical use, due to its side effects, i.e., anorexia, skeletal muscle loss, hyperalgesia, cramps, and muscle pain, has not materialized. In the present study, we report that Peptide 6c (GDDL) that corresponds to CNTF amino acid residues 147-150, enhances the dentate gyrus neurogenesis and neuronal plasticity, and improves cognition without weight loss or any other apparent side effects in mice. Normal adult C57Bl6 mice received subcutaneous implants of extended release depot pellets containing vehicle or Peptide 6c for 30 days of continuous dosing. Dentate gyrus neurogenesis was assessed by stereological analysis of cells expressing neuronal markers, doublecortin and NeuN, and BrdU uptake. We found that Peptide 6c significantly increased early neuronal commitment, differentiation, and survival of newborn progenitor cells. These newborn neurons were functionally integrated into the hippocampal network, since basal expression of c-fos was enhanced and neuronal plasticity was increased, as reflected by higher expression of MAP2a,b and synaptophysin. Consequently, Peptide 6c treatment improved encoding of hippocampal-dependent information in a spatial reference memory task in mice. Overall, these findings demonstrated the therapeutic potential of Peptide 6c for regeneration of the brain and improvement of cognition.

Pages 1197-1216
Dang Thanh Nam, Madeleine Arseneault, Neven Zarkovic, Georg Waeg, Charles Ramassamy (Handling Associate Editor: Raymond Chuen-Chung Chang)
Molecular Regulations Induced by Acrolein in Neuroblastoma SK-N-SH Cells: Relevance to Alzheimer’s Disease
Abstract: Acrolein is the most reactive aldehyde among the by-products of lipid peroxidation. Growing evidence indicates that acrolein may play an important role in the pathogenesis of Alzheimer’s disease (AD). In AD, levels of acrolein are significantly higher in hippocampus and temporal cortex regions of the brain. However, little is known about its toxicity in neuronal cells. Using the neuroblastoma cell line SK-N-SH, our results show that acrolein is toxic from 10 µM, but its toxicity does not induce the activation of caspase-3 and DNA fragmentation. Protein carbonylation and 4-hydroxy-nonenal levels were increased after 0.5 h and 1 h of treatment, respectively. Furthermore acrolein induced a biphasic effect on glutathione levels with a rapid depletion followed by a progressive increase. We have further investigated the regulation of different redox signaling pathways. A treatment with 10 µM of acrolein for 30 min activated NFκB while this activation was suppressed after a 24 h of treatment. In contrast, Nrf2 was activated only after 24 h of acrolein exposure. Consequently, the expression of heme oxygenase-1 and γ-glutamyl-cysteine-synthase were elevated after 24 h of acrolein treatment. Sirt-1 was also upregulated after 24 h of acrolein treatment. The p66Shc and ERK1/2 proteins are known to be involved in oxidative stress. Acrolein, at 10 µM, induced the phosphorylation of p66Shc and ERK1/2 only after a short period of treatment. Collectively, these data strengthen the contribution of acrolein in the induction of oxidative stress as observed in mild cognitive impairment and in AD brain.

Pages 1217-1231
Julien Couturier, Guylène Page, Milena Morel, Céline Gontier, Jean-Claude Lecron, Raymond Pontcharraud, Bernard Fauconneau, Marc Paccalin (Handling Associate Editor: Marcella Reale)
Inhibition of Double-Stranded RNA-Dependent Protein Kinase Strongly Decreases Cytokine Production and Release in Peripheral Blood Mononuclear Cells from Patients with Alzheimer’s Disease
Abstract: Alzheimer's disease (AD), a neurodegenerative disorder, is the most common form of dementia in the elderly individuals. Among the pathogenic mechanisms in AD, chronic systemic inflammation is described and characterized by massive production of proinflammatory cytokines by peripheral blood mononuclear cells (PBMCs), which may contribute to an altered immune response and exacerbated of neurodegeneration. Studies have also reported increased double-stranded RNA-dependent protein kinase (PKR) activation in the PBMCs of patients with AD. Interestingly, PKR could be involved in NF-κB activation, leading to production of a wide range of cytokines. We proposed to decrease proinflammatory cytokines production and release by treating the PBMCs in 25 patients with AD with a specific inhibitor of PKR. Our results showed that PKR inhibition greatly decreased tumor necrosis factor α, interleukin (IL)-1α, IL-1β, and IL-6 production and release but did not affect the chemokine RANTES. Moreover, inhibition of the proinflammatory factors was correlated with prevention of caspase-3 activation. These results indicated that specific inhibition of PKR at the peripheral level might decrease the inflammatory response in AD.

Supplementary Data for Couturier et al. article (PDF)

Pages 1233-1247
Zhifang Li*, Changyue Gao*, Heqing Huang, Weizhong Sun, Hongliang Yi, Xiaotang Fan, Haiwei Xu (*Both the authors contributed equally to this work.)
Neurotransmitter Phenotype Differentiation and Synapse Formation of Neural Precursors Engrafting in Amyloid-β1-40 Injured Rat Hippocampus
Abstract: Alzheimer’s disease (AD) is characterized by the dysfunction or loss of a vulnerable group of neurons. At present, only few options exist for treating neurodegenerative diseases effectively. Advances in stem cell research have raised the hope and possibility for the therapy of neurodegenerative diseases. In AD transgenic animal models, stem cell transplantation has been demonstrated to reverse behavioral deficits. Our recent study has demonstrated that neural precursor cells derived from embryonic stem (ES) cells, improve memory dysfunction in rats caused by injections of amyloid-β peptide (1-40) (Aβ1-40) in the dorsal hippocampus. However, the underlying mechanisms have remained unknown. The present study was performed to test a murine ES cell-based transplantation approach in rats subjected to Aβ1-40 injection into hippocampus dentate gyrus. Efficacy of cell therapy regarding graft survival, neuronal yield and diversity, synapse formation of the grafted cells, and the behavioral improvements were determined after transplanting ES cell-derived neural precursors into the hippocampus of adult rats. Here, we show that grafted cells can survive, and differentiate with high yield into immunohistochemically mature glial cells and neurons of diverse neurotransmitter-subtypes. More importantly, transplanted cells demonstrate characteristics of proper synapse formation between host and grafted neural cells. Thus, our observations show that ES cell-based transplantation approach may be promising in treatment of AD.

Pages 1249-1262
Chenghua Li, Juan Wang and Bing Zhou (Handling Associate Editor: Ashley Bush)
The Metal Chelating and Chaperoning Effects of Clioquinol: Insights from Yeast Studies
Abstract: Clioquinol (CQ), a once popular antibiotic, was used to inhibit the growth of microorganisms. Recently, CQ and its analog PBT2 have shown encouraging effects in the animal and clinical trials for Alzheimer’s disease (AD). However, the mechanism by which this class of molecules works remains controversial. In this work, we used the yeast Saccharomyces cerevisiae as a model to study how CQ affects molecular and cellular functions and particularly, copper, iron, and zinc homeostasis. We observed a CQ-induced inhibition of yeast growth, which could be slightly relieved by supplementation of copper or iron. Microarray results indicated that yeast cells treated with CQ sense a general deficiency in metals, despite elevated total cellular contents of copper and iron. Consistent with this, reduced activities of some metal-sensitive enzymes were observed. Intriguingly, CQ can increase the SOD1 activity, likely through Ccs1’s accessibility to CQ-bound copper ions. Further studies revealed that CQ sequestrates copper and iron at the cellular membrane, likely the plasma membrane, resulting overall metal accumulation but cytosolic metal depletion. CQ’s effects on metal-sensitive metalloenzymes were also verified in mammalian cell line SH-SY5Y. Together, our results revealed that CQ can regulate metal homeostasis by binding metal ions, resulting the cell sensing a state of deficiency of bio-available metal ions while simultaneously increasing available metals to SOD1 (via Ccs1) and possibly some other metalloproteins that can access CQ-bound metals. We hope this regulation of metal homeostasis may be helpful in explaining the therapeutic effects of CQ used in disease treatment.

Supplementary Data for Li et al. article (PDF)

Pages 1263-1269
Christoph Laske, Kateryna Sopova, Christos Gkotsis, Gerhard W. Eschweiler, Guido Straten, Meinrad Gawaz, Thomas Leyhe, Konstantinos Stellos (Handling Associate Editor: Milan Fiala)
Amyloid-β Peptides in Plasma and Cognitive Decline After 1 Year Follow-Up in Alzheimer’s Disease Patients
Abstract: Plasma levels of amyloid-β (Aβ) peptides are potential biomarkers of early cognitive impairment and of Alzheimer’s disease (AD) risk. However, the association of Aβ peptides with the rate of cognitive decline in AD patients is still unclear. In the present study we demonstrate that Aβ1-42 plasma levels show a significant correlation with the rate of cognitive decline and are significantly increased in AD patients with fast cognitive decline (decrease of Mini-Mental Status Examination (MMSE) score ≥ 5/year; n=12) compared to AD patients with slow cognitive decline (decrease of MMSE score ≤ 4/year; n=28), independent of baseline MMSE scores, age and cholinesterase inhibitor intake, but dependent on history of myocardial infarction and history of stroke in a multivariate analysis. These results suggest that Aβ1-42 plasma levels are associated with the rate of cognitive decline in AD patients and may be influenced by atherosclerotic vasculopathies such as stroke and myocardial infarction.

Pages 1271-1281
Martijn C. de Wilde, Eline M. van der Beek, Amanda J. Kiliaan, Inge Leenders, Almar A.M. Kuipers, Patrick J. Kamphuis, Laus M. Broersen
Docosahexaenoic Acid Reduces Amyloid-β1-42 Secretion in Human AβPP-Transfected CHO-Cells by Mechanisms Other than Inflammation Related to PGE2
Abstract: The effect of supplementation with the omega 3 polyunsaturated fatty acid (n-3 PUFA) docosahexaenoic acid (DHA) on membrane composition and amyloid-β1-42 (Aβ42) secretion was studied in human amyloid-β protein precursor-transfected Chinese Hamster Ovary (CHO) cells. Twenty-four hour incubation with a range of DHA concentrations resulted in a dose-dependent increase in membrane DHA and eicosapentaenoic acid content and a decrease in arachidonic acid content. In addition, DHA supplementation caused a dose-dependent reduction in the secreted Aβ42 levels and resulted in a 4-8 fold decrease in extracellular prostaglandin E2 (PGE2) levels. Tocopherol, which was added to DHA to prevent oxidation, may have contributed to the effect of DHA, since it slightly decreased extracellular Aβ42 and PGE2 levels when given alone. The addition of selective COX-2 inhibitors Celebrex and curcumin to the culture medium resulted in a significant and comparable inhibition of PGE2 release, but did not inhibit Aβ42 secretion, and even significantly increased Aβ42 production in this cell system. Together, the present data show that, whereas both DHA and COX-2 inhibitors may reduce PGE2 production, only DHA in the presence of tocopherol significantly reduced Aβ42 production and concurrently changed membrane lipid composition in CHO cells. It is concluded that in this in vitro setting DHA reduced Aβ42 secretion through membrane-related, but not PGE2-related mechanisms.

Pages 1283-1293
Kurt A. Jellinger, Johannes Attems (Handling Associate Editor: Rudy Castellani)
Prevalence and Pathology of Vascular Dementia in the Oldest-Old*
Abstract: The prevalence of both Alzheimer's disease (AD) and vascular dementia (VaD) increase with advancing age, but epidemiologic data above age 85 are imprecise and inconsistent. A retrospective hospital-based study of the prevalence and pathology of VaD was performed in 1700 consecutive autopsy cases of demented elderly in Vienna, Austria (mean age 84.3 ± 5.4 SD; 90% over age 70). It assessed clinical and general autopsy data and neuropathology including immunohistochemistry. Neuropathologic diagnosis followed current consensus criteria. Four age groups (7th to 10th decade) were evaluated. "Pure" VaD (due to cerebrovascular disease without other pathologies; neuritic Braak stages 1.2-1.6) was observed in 12.3% of the total cohort, decreasing between age 60 and 90+ from 15.0 to 8.7%. Morphologic subtypes (subcortical arteriosclerotic encephalopathy, multi-infarct encephalopathy, and strategic infarct dementia) showed no age-related differences. By contrast, AD (without concomitant pathologies; 45.6% of total), mixed dementia (AD + cerebrovascular encephalopathy; 5.5%), and AD with minor cerebrovascular lesions (22.3%) increased with age. The relative prevalence of AD + Lewy pathology (9.3%) remained fairly stable, whereas other dementias (5.0%) decreased significantly over age 90. 85% of the patients with "pure" VaD had histories of diabetes, 75% of stroke(s), 95% morphologic signs of hypertension, 65% myocardial infarction (recent and old ones), 97% cerebral hypertonic-arteriosclerotic microangiopathy (associated with cerebral amyloid angiopathy in 23%) and 90% severe atherosclerosis of large cerebral arteries. Similar autopsy findings were seen in mixed dementia (MIX) and in AD + minor cerebrovascular lesions. Major vascular lesions differed between VaD and MIX, VaD showing more than 60% subcortical infarcts, MIX only 43% such lesions. This retrograde hospital-based study using strict morphologic diagnostic criteria confirmed the existence of "pure" VaD in old age, with a tendency to decline after age 90, while AD and AD + cerebrovascular pathologies showed considerable age-related increase, and "pure" AD slightly decreasing after age 90.

Pages 1295-1301
Dag Sehlin*, Sofia Söllvander*, Staffan Paulie, RoseMarie Brundin, Martin Ingelsson, Lars Lannfelt, Frida Ekholm Pettersson, Hillevi Englund *These authors have contributed equally to the work
Interference from Heterophilic Antibodies in Amyloid-β Oligomer ELISAs
Abstract: Amyloid-β (Aβ) oligomers of different sizes and forms have recently been the focus for many Alzheimer’s disease (AD) researchers. Various immunoassays have been used to detect low concentrations of these elusive Aβ species in different forms of human samples using little or no sample dilutions. However, the possibility that positive results may be caused by interference from heterophilic antibodies (HA) is often overlooked. HA, which recognize immunoglobulins from other species, are present in human plasma and cerebrospinal fluid (CSF) and may cause interference in sandwich immunoassays like enzyme-linked immunosorbent assays (ELISAs) by cross-binding the capture and detection antibodies of the assay. They thus may generate a false positive signal. Here we show that when assessing the Aβ oligomer content in plasma samples from 44 individuals with a sandwich ELISA, none of the 21 positive signals remained when the assay was repeated in the presence of factors blocking HA. Similarly, in CSF samples from 104 individuals, the signals from the 22 positive samples were strongly reduced when analyzed after anti-HA treatment. Taken together, HA interference is a problem that needs to be addressed when measuring low levels of an antigen in human plasma and CSF samples.

Pages 1303-1309
Pravat K. Mandal, Neel Sarovar Bhavesh, Virender S. Chauhan, Vincenzo Fodale
NMR Investigations of Amyloid-β Peptide Interactions with Propofol at Clinically Relevant Concentrations with and without Aqueous Halothane Solution
Abstract: Oligomerization of amyloid-β peptide (Aβ) is an important stage in Alzheimer’s disease. Recently, it has been shown that in an experimental model, smaller sized (e.g., isoflurane, desflurane, etc.) anesthetics induce Aβ oligomerization. Using state-of-the-art solution nuclear magnetic resonance, spectroscopic studies on Aβ interaction with propofol indicated that propofol does not interact with the G29, A30, and I31 residues of Aβ peptide at a clinically relevant concentration (0.0832 mM), and no Aβ oligomerization was observed after 69 days. However, Aβ oligomerization was observed when treated with propofol (clinically relevant concentration) coadministered with aqueous halothane solution. Furthermore, dose dependence studies at various propofol concentrations (0.32 mM, 2.05 mM, and 53.4 mM) indicate the effect of propofol concentration on Aβ oligomerization revealing the hydrophobic nature of interactions between propofol with these critical residues. These experimental findings reaffirm that smaller molecular sized anesthetics (e.g., halothane) do play a leading role in Aβ oligomerization.

Pages 1311-1323
Yan Li*, Leung-Wing Chu*, Binbin Wang*, Ping-Yiu Yik, Huriletemuer, Dong-Yan Jin, Xu Ma, You-Qiang Song *contributed equally to the work
CYP46A1 Functional Promoter Haplotypes Decipher Genetic Susceptibility to Alzheimer’s Disease
Abstract: We here demonstrate that promoter polymorphisms rs8003602 and rs3783320 of cholesterol 24S-hydroxylase (CYP46A1) were significantly associated with Alzheimer’s disease (AD) in Chinese subjects. Haplotype analyses showed that haplotype CG is the risk haplotype. Either single marker or haplotypic association was found only in the APOE ε4 negative group. The association was then replicated in an independent set of case-control samples in Mongolians. We also investigated the function of promoter haplotypes and found that luciferase expression for TA promoter construct exhibited significantly higher expression level than the risk CG promoter construct. This finding might indicate individuals bearing the CG haplotype are genetically more susceptible to AD compared to those with TA haplotype. Further, we found MYT1 could be the potential transcription factor binding to the significant promoter polymorphism and mediated gene transcriptional activity. In general, our results show that promoter haplotypes could significantly affect CYP46A1 gene transcription level possibly through interacting with certain transcription factors.

Pages 1325-1334
Anna Rita Atti, Claudia Forlani, Diana De Ronchi, Katie Palmer, Paola Casadio, Edoardo Dalmonte, Laura Fratiglioni (Handling Associate Editor: Patrizia Mecocci)
Cognitive Impairment after Age 60: Clinical and Social Correlates in the “Faenza Project”
Abstract: A total of 7,389 dementia-free elderly (60-102 years old) enrolled in the “Faenza Project” (Northern Italy) were clinically evaluated by nurses and physicians with the aim of detecting the independent and combined association of medical and social factors with cognitive status. Cognitive Impairment No Dementia (CIND) was defined for MMSE scores ≤ 2 standard deviations than the age- and education-corrected mean score obtained by the non-demented persons of the Faenza cohort. Logistic Regression analysis was used to estimate Odds Ratios and 95% Confidence Intervals (OR, 95%CI) for CIND. The diagnostic procedure identified 402 (5.4%) CIND cases. Diabetes (OR, 95%CI=1.6, 1.2-2.2), stroke (OR, 95%CI=1.9, 1.4-2.6), and depressive symptoms (OR, 95%CI=1.9, 1.4-2.7) emerged as the most relevant medical comorbidities of CIND. Low education (OR, 95%CI=1.8, 1.1-2.9), low Socio Economic Status (SES) (OR, 95%CI=1.5, 1.1-2.1), and unmarried status (OR, 95%CI=1.7, 1.2-2.5) were associated with CIND. Medical and social factors were independently related to CIND occurrence. In comparison to subjects without any of the above mentioned conditions, subjects with one medical and one social factor had an OR for CIND equal to 6.0 (95%CI 2.9-12.4). The strength of the association increased when more of those conditions occurred in combination, suggesting a synergist effect. Despite some methodological limitations, data from this cross-sectional population-based Italian study show that low education, low SES, unmarried status together with diabetes, stroke, and depressive symptoms are related to cognitive impairment in the general population. The interaction of medical and social factors further increases the probability of CIND.

Pages 1335-1345
Leung-Wing Chu, Sidney Tam, Rachel LC Wong, Ping-Yiu Yik, Youqiang Song, Bernard MY Cheung, John E Morley, Karen SL Lam
Bioavailable Testosterone Predicts a Lower Risk of Alzheimer’s Disease in Older Men
Abstract: There is a paucity of data on the relationship between testosterone and Alzheimer’s disease (AD) in older men. The objective of the present study was to investigate the effects of serum total testosterone (TT), bioavailable testosterone (BT), and sex hormone binding globulin (SHBG) levels on the subsequent risk of AD in non-demented Chinese older men. This was a one-year prospective cohort study. 153 ambulatory community-living non-demented Chinese older men, aged 55 years or over, were recruited and followed for one year. Morning serum TT, BT, and SHBG levels were measured at baseline. At one-year of follow-up, assessment for dementia and AD were performed. AD was diagnosed by the NINCDS-ADRDA criteria for probable AD. Overall, the mean age of the subjects was 72.7 (SD 6.9). 6.5% (n = 10) developed dementia (converters), all having AD. 93.5% (n = 143) did not develop dementia (non-converters). Logistic regression analysis for independent predictors of AD showed that the baseline serum BT level, systolic blood pressure (SBP) and ApoE ε4 genotype were significant independent predictors, after adjustment for age, education, BMI, fasting plasma glucose, and serum HDL-C levels. The baseline serum BT level predicted a reduced risk of AD (adjusted relative risk (RR) 0.22, 95% CI: 0.07-0.69)). Baseline SBP and ApoE ε4 genotype but not SHBG were independent risk factors, with RRs of 1.04 and 5.04 respectively. In conclusion, the serum level of bioavailable testosterone in late life predicts a lower risk of future AD development in older men.

Pages 1347-1365
George Pengas, Karalyn Patterson, Robert J. Arnold, Chris M. Bird, Neil Burgess, Peter J. Nestor
Lost and Found: Bespoke Memory Testing for Alzheimer's Disease and Semantic Dementia
Abstract: The neural network activated during Topographical Memory (TM) tasks in controls overlaps with the earliest affected regions in Alzheimer’s disease (AD) but not with those of Semantic Dementia (SD). This suggests that clinical TM tests could be more bespoke to neural dysfunction in early AD and therefore more sensitive and specific. We hypothesized that TM impairment would be characteristic of AD but not of SD making it useful both for early diagnosis and differential diagnosis. TM was assessed in 69 patients (22 mild AD, 15 SD, 32 with mild cognitive impairment (MCI)) and 35 controls, using three tasks: the four mountains test and two novel tests in a virtual town (the Virtual Route Learning Test (VRLT) and the Heading Orientation Test). AD patients were impaired on all TM tasks. The VRLT was the most discriminatory; had the highest correlation with caregiver reports of navigation problems; and correlated strongly with memory, attention/executive function, and to a lesser degree, visuospatial ability. In contrast, SD patients performed well on the TM battery only becoming abnormal with very advanced dementia and performance correlated exclusively with attention/executive function. The VRLT achieved 95% sensitivity and 94% specificity in discriminating AD patients from controls; at the same cut-off, 70% of MCI patients were impaired. When combined with either naming performance or global dementia severity, there was complete separation of AD from SD. The VRLT is ecologically valid, highly sensitive to early AD, and useful in discriminating AD from the non-Alzheimer dementia, SD.

Supplementary Data for Pengas et al. article (PDF)

Pages 1367-1381
Claudia Balducci, Raffaella Tonini, Elisa Zianni, Cristiano Nazzaro, Fabio Fiordaliso, Monica Salio, Lorenzo Vismara, Fabrizio Gardoni, Monica Di Luca, Mirjana Carli, Gianluigi Forloni
Cognitive Deficits Associated with Alteration of Synaptic Metaplasticity Precede Plaque Deposition in AβPP23 Transgenic Mice
Abstract: Synaptic dysfunction is an early event in the development of Alzheimer’s disease (AD) and relates closely to the cognitive impairment characterizing this neurodegenerative process. A causative association has been proposed, largely on the basis of in vitro studies, between memory decline, soluble amyloid-β (Aβ) oligomers and alterations of glutamatergic neurotransmission. We aimed here to characterize in vivo N-methyl-D-aspartate receptor (NMDAR)-mediated signaling, at an early stage of AD, before extracellular amyloid plaques are deposited. We assessed the functional link between cognitive abilities and NMDAR-mediated pharmacological responses of six-month-old AβPP23 transgenic mice (AβPP23tg), overexpressing the human amyloid-β protein precursor carrying the Swedish double mutation. We found evidence of cognitive impairments in these mice, indicated by deficits in the delayed-non-matching-to-place task. Alterations of NMDAR-mediated signaling in this mouse model were confirmed by the reduced sensitivity of motor-activation and working memory to pharmacological inhibition of NMDAR activity. At the molecular level, AβPP23tg mice show hippocampal alterations in the trafficking of synaptic NMDAR subunits NR2A and NR2B and at an ultrastructural analysis show Aβ oligomers intracellularly localized in the synaptic compartments. Importantly, the behavioral and biochemical alterations of NMDAR signaling are associated with the inhibition of long-term synaptic potentiation and inversion of metaplasticity at CA1 synapses in hippocampal slices from AβPP23tg mice. These results indicate a general impairment of synaptic function and learning and memory in young AβPP23tg mice with Aβ oligomers but no amyloid plaques.

Pages 1383-1392
Luigi Iuliano, Roberto Monticolo, Giuseppe Straface, Ilaria Spoletini, Walter Gianni, Carlo Caltagirone, Paola Bossù, Gianfranco Spalletta
Vitamin E and Enzymatic/Oxidative Stress-Driven Oxysterols in Amnestic Mild Cognitive Impairment Subtypes and Alzheimer’s Disease
Abstract: Oxidative stress, which contributes to neuronal damage, is thought to be a pathophysiological mechanism of Alzheimer’s disease (AD). Markers of oxidative stress may appear early in the preclinical, mild cognitive impairment (MCI) phase of AD. We investigated the interaction among enzymatic-derived oxysterols (24S-hydroxycholesterol and 27-hydroxycholesterol), markers of oxidative stress, including free radical-related oxysterols (7β-hydroxycholesterol and 7-ketocholesterol), and vitamin E in AD patients and two amnestic MCI subtypes, amnestic single-domain MCI (a-MCI) subjects, and multi-domain MCI (md-MCI) subjects, compared to healthy control subjects (HC). The study included 37 patients with AD, 24 with a-MCI, 29 with md-MCI, and 24 HC. Plasma assessments were made using isotope dilution-mass spectrometry. Although we found no significant change in free radical- or enzymatic-derived oxysterol concentrations in AD or MCI patients, vitamin E levels corrected for cholesterol were reduced in AD patients compared to HC. Results suggest that AD patients have upregulated oxidative cerebral stress or a nutritional deficit of vitamin E. The oxysterols investigated here are not useful markers for diagnosing AD or MCI.

Pages 1393-1402
Thea Brennan-Krohn, Stephen Salloway, Stephen Correia, Matthew Dong, Suzanne M. de la Monte (Handling Associate Editor: Jack de la Torre)
Glial Vascular Degeneration in CADASIL
Abstract: CADASIL is a genetic vascular dementia caused by mutations in the Notch 3 gene on Chromosome 19. However, little is known about the mechanisms of vascular degeneration. We characterized upstream components of Notch signaling pathways that may be disrupted in CADASIL, by measuring expression of insulin, IGF-1, and IGF-2 receptors, Notch 1, Notch 3, and aspartyl-(asparaginyl)-β-hydroxylase (AAH) in cortex and white matter from 3 CADASIL and 6 control brains. We assessed CADASIL-associated cell loss by measuring mRNA corresponding to neurons, oligodendroglia, and astrocytes, and indices of vascular degeneration by measuring smooth muscle actin (SMA) and endothelin-1 expression in isolated vessels. Immunohistochemical staining was used to assess SMA degeneration. Significant abnormalities, including reduced cerebral white matter mRNA levels of Notch 1, Notch 3, AAH, SMA, IGF receptors, myelin-associated glycoproteins, and glial fibrillary acidic protein, and reduced vascular expression of SMA, IGF receptors, Notch 1, and Notch 3 were detected in CADASIL-lesioned brains. In addition, we found CADASIL-associated reductions in SMA, and increases in ubiquitin immunoreactivity in the media of white matter and meningeal vessels. No abnormalities in gene expression or immunoreactivity were observed in CADASIL cerebral cortex. In conclusion, molecular abnormalities in CADASIL are largely restricted to white matter and white matter vessels, corresponding to the distribution of neuropathological lesions. These preliminary findings suggest that CADASIL is mediated by both glial and vascular degeneration with reduced expression of IGF receptors and AAH, which regulate Notch expression and function.

Pages 1403-1408
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TDP-43 Models Coverage