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Volume 56, Number 3, IN PRESS

Hypothesis
Efraim Jaul, Oded Meiron
Dementia and Pressure Ulcers: Is There a Close Pathophysiological Interrelation?
Abstract: The current theoretical investigation aimed to explore common pathophysiological mechanisms underlying dementia and pressure ulcers (PU). Along with the increased longevity, especially in frail elderly patients, there is a higher rate of functional and cognitive impairment with dementia coinciding with disabilities and immobility resulting in a higher rate of PU. Understanding common etiological paths resulting in pressure ulcers and dementia is likely to produce new treatment strategies that could lead to the prevention of comorbid complications. Data collected from elderly dementia patients indicate a deterioration of several neurophysiological subsystems associated with motor, sensory, autonomic, cognitive, or behavioral pathways, supporting a “close pathophysiological interrelation” perspective linking PU with dementia progression. Overall, the authors’ theoretical systemic-model of disease progression and PU comorbidity proposes that increased clinician awareness to PU in mild to moderate dementia may suppress the accelerated development of PU, resulting in less patient suffering, reduced long-term care hospitalization, and hopefully PU prevention.

Hypothesis
Frank O. Bastian
Combined Creutzfeldt-Jakob-Jacob/Alzheimer’s Disease Cases are Important in Search for Microbes in Alzheimer’s Disease
Abstract: The question whether Alzheimer’s disease is infectious as brought up in the recent editorial published in the Journal of Alzheimer’s Disease is complicated by the controversy whether the causal agent is a microbe or a misfolded host protein (amyloid). The replicating amyloid (prion) theory, based upon data from studies of Creutzfeldt-Jakob disease (CJD) and other transmissible spongiform encephalopathies (TSEs), has been challenged since the prion can be separated from TSE infectivity, and spiroplasma, a wall-less bacterium, has been shown to be involved in the pathogenesis of CJD. Further support for a microbial cause for AD comes from occurrence of mixed CJD/AD cases involving up to 15% of AD brains submitted to brain banks. The association of CJD with AD suggests a common etiology rather than simply being a medical curiosity. A co-infection with the transmissible agent of CJD, which we propose to be a Spiroplasma sp., would explain the diversity of bacteria shown to be associated with cases of AD.

Short Communication
Hao Wang, Xiaoyu Hong, Yong Wang
Mitochondrial Repair Effects of Oxygen Treatment on Alzheimer’s Disease Model Mice Revealed by Quantitative Proteomics
Abstract: Mitochondrial dysfunction plays a pivotal role in Alzheimer’s disease (AD), even before signs of AD pathology are evident. Our previous research has shown that oxygen treatment can improve cognitive function in AD model mice. To address whether oxygen treatment is beneficial to mitochondrial biology, we analyzed differential expressions of hippocampal mitochondrial proteins in AD model mice given supplementary oxygen. Numerous respiratory chain, Kreb’s cycle, and glycolysis proteins were upregulated significantly after oxygen treatment, suggesting that oxygen therapy can alleviate mitochondrial damage. Furthermore, the treatment was associated with decreased expressions of some AD biomarkers, suggesting oxygen treatment to be a potential therapy for AD.

Short Communication
Jamie Toombs, Martha S. Foiani, Ross W. Paterson, Amanda Heslegrave, Selina Wray, Jonathan M. Schott, Nick C. Fox, Michael P. Lunn, Kaj Blennow, Henrik Zetterberg
Effect of Spinal Manometers on Cerebrospinal Fluid Amyloid-β Concentration
Abstract: The effect of spinal manometers on cerebrospinal fluid (CSF) amyloid-β (Aβ) concentration was investigated. Pooled human CSF samples were divided in two, one half passed through a manometer into a collection tube, the other transferred directly to a collection tube. CSF was analyzed for Aβ38/40/42 using an electrochemiluminescence immunoassay. Relative to control, use of a manometer decreased Aβ38/40/42 concentration by 5.6% (± 1.5SE), 4.4% (± 1.7SE), and 4.3% (± 2.4SE), respectively. The ratios of Aβ42:40, Aβ42:38, and Aβ40:38 were not affected by manometer treatment. Factors which artificially lower CSF Aβ concentrations are relevant to clinical diagnosis for AD and study design.

Short Communication
Liyong Wu*, Jia Liu*, Longze Sha, Xianling Wang, Jieying Li, Jing Dong, Jianping Jia *These authors contributed equally to this work.
Sporadic Cases with Novel Mutations and Pedigree in Hereditary Leukoencephalopathy with Axonal Spheroids
Abstract: Hereditary diffuse leukoencephalopathy with spheroids (HDLS) is an autosomal dominant hereditary disease, featured by cerebral white matter degeneration with demyelination and axonal spheroids. We collected three gene-confirmed HDLS cases in our neurodegenerative clinic. Two HDLS cases were sporadic with novel mutations, while another case had a family history with previously described mutations. All three cases suffered memory problems with white matter lesions and pyramid signs. No obvious clinical differences were observed between sporadic and familial HDLS cases. Distinct features, such as subcortical calcification in brain computed tomography and asymmetric abnormal MRI signal along the pyramid tracts throughout brainstem and spinal cord (cervical, thoracic, and lumbar segments), were observed in one sporadic case with novel mutation. Therefore, the interactions of genotype-phenotype still need to be further investigated.

Kim E. Innes, Terry Kit Selfe, Dharma Singh Khalsa, Sahiti Kandati (Handling Associate Editor: J. Wesson Ashford)
Meditation and Music Improve Memory and Cognitive Function in Adults with Subjective Cognitive Decline: A Pilot Randomized Controlled Trial
Abstract: Background: While effective therapies for preventing or slowing cognitive decline in at-risk populations remain elusive, evidence suggests mind-body interventions may hold promise. Objectives: In this study, we assessed the effects of Kirtan Kriya meditation (KK) and music listening (ML) on cognitive outcomes in adults experiencing subjective cognitive decline (SCD), a strong predictor of Alzheimer's disease. Methods: Sixty participants with SCD were randomized to a KK or ML program and asked to practice 12 minutes/day for 3 months, then at their discretion for the ensuing 3 months. At baseline, 3 months, and 6 months we measured memory and cognitive functioning [Memory Functioning Questionnaire (MFQ), Trail-making Test (TMT-A/B), and Digit-Symbol Substitution Test (DSST)]. Results: The 6-month study was completed by 53 participants (88%). Participants performed an average of 93% (91% KK, 94% ML) of sessions in the first 3 months, and 71% (68% KK, 74% ML) during the 3-month, practice-optional, follow-up period. Both groups showed marked and significant improvements at 3 months in memory and cognitive performance (MFQ, DSST, TMT-A/B; p’s≤0.04). At 6 months, overall gains were maintained or improved (p’s≤0.006), with effect sizes ranging from medium (DSST, ML group) to large (DSST, KK group; TMT-A/B, MFQ). Changes were unrelated to treatment expectancies and did not differ by age, gender, baseline cognition scores, or other factors. Conclusions: Findings of this preliminary randomized controlled trial suggest practice of meditation or ML can significantly enhance both subjective memory function and objective cognitive performance in adults with SCD, and may offer promise for improving outcomes in this population.

María José González-Muñoz, Alba Garcimartín, Isabel Meseguer, Carmen José Mateos-Vega, José María Orellana, Antonio Peña-Fernández, Juana Benedí, Francisco J. Sánchez-Muniz
Silicic Acid and Beer Consumption Reverses the Metal Imbalance and the Prooxidant Status Induced by Aluminum Nitrate in Mouse Brain
Abstract: Background: Emerging evidence suggests that by affecting mineral balance, aluminum (Al) may enhance some events associated with neurodegenerative diseases. Objective: To examine the effect of Al(NO3)3 exposure on brain Al, cooper (Cu), iron (Fe), magnesium (Mg), manganese (Mn), silicon (Si), and zinc (Zn) levels, and the metal-change implication in brain oxidant and inflammatory status. Methods: Four groups of six-week-old male NMRI mice were treated for three months: i) controls, administrated with deionized water; ii) Al, which received Al(NO3)3; iii) Al+silicic acid, which were given Al(NO3)3 plus silicic acid; and iv) Al+beer, which received Al(NO3)3 plus beer. Results: Brain Al and TBARS levels and TNFα and GPx expressions increased, while Cu, Mn, and Zn levels, and catalase and CuZn-SOD expression decreased (at least, p<0.05) in Al versus control animals. Al, Si, and TBARS levels and TNFα expression decreased (p<0.05) in Al+silicic acid and Al+beer specimens while Cu, Mn, and Zn levels and antioxidant expression increased versus the Al group. Brain Al levels correlated negatively with those of Cu, Fe, Mn, and Zn, and catalase, CuZn-SOD, and GPx enzyme expressions but positively with Si and TBARS levels and TNFα expression. Two components of the principal component analysis (PCA) explained 71.2% of total data variance (p<0.001). PCA connected the pro-oxidant markers with brain Al content, while brain Zn and Cu levels were closer to antioxidant enzyme expression. Conclusion: Administration of Al(NO3)3 induced metal imbalance, inflammation, and antioxidant status impairment in the brain. Those effects were blocked to a significant extent by silicic acid and beer administration.

Zahinoor Ismail, Luis Agüera-Ortiz, Henry Brodaty, Alicja Cieslak, Jeffrey Cummings, Corinne E. Fischer, Serge Gauthier, Yonas E. Geda, Nathan Herrmann, Jamila Kanji, Krista L. Lanctôt, David S. Miller, Moyra E. Mortby, Chiadi U. Onyike, Paul B. Rosenberg, Eric E. Smith, Gwenn S. Smith, David L. Sultzer, Constantine Lyketsos for the NPS Professional Interest Area of the International Society of to Advance Alzheimer’s Research and Treatment (NPS-PIA of ISTAART)
The Mild Behavioral Impairment Checklist (MBI-C): A Rating Scale for Neuropsychiatric Symptoms in Pre-Dementia Populations
Abstract: Background: Mild behavioral impairment (MBI) is a construct that describes the emergence at 50 years of age of sustained and impactful neuropsychiatric symptoms (NPS), as a precursor to cognitive decline and dementia. MBI describes NPS of any severity, which are not captured by traditional psychiatric nosology, persist for at least 6 months, and occur in advance of or in concert with mild cognitive impairment. While the detection and description of MBI has been operationalized in the International Society to Advance Alzheimer’s Research and Treatment – Alzheimer’s Association (ISTAART-AA) research diagnostic criteria, there is no instrument that accurately reflects MBI as described. Objective: To develop an instrument based on ISTAART-AA MBI criteria. Methods: Eighteen subject matter experts participated in development using a modified Delphi process. An iterative process ensured items reflected the five MBI domains of 1) decreased motivation; 2) emotional dysregulation; 3) impulse dyscontrol; 4) social inappropriateness; and 5) abnormal perception or thought content. Instrument language was developed a priori to pertain to non-demented functionally independent older adults. Results: We present the Mild Behavioral Impairment Checklist (MBI-C), a 34-item instrument, which can easily be completed by a patient, close informant, or clinician. Conclusion: The MBI-C provides the first measure specifically developed to assess the MBI construct as explicitly described in the criteria. Its utility lies in MBI case detection, and monitoring the emergence of MBI symptoms and domains over time. Studies are required to determine the prognostic value of MBI for dementia development, and for predicting different dementia subtypes.

Joshua D. Grill, Andrew Holbrook, Aimee Pierce, Dan Hoang, Daniel L. Gillen (Handling Associate Editor: Russell Swerdlow)
Attitudes toward Potential Participant Registries
Abstract: Difficult participant recruitment is a consistent barrier to successful medical research. Potential participant registries represent an increasingly common intervention to overcome this barrier. A variety of models for registries exist, but few data are available to instruct their design and implementation. To provide such data, we surveyed 110 cognitively normal research participants enrolled in a longitudinal study of aging and dementia. Seventy-four (67%) individuals participated in the study. Most (78%, CI: 0.67, 0.87) participants were likely to enroll in a registry. Willingness to participate was reduced for registries that required enrollment through the Internet using a password (26%, CI: 0.16, 0.36) or through email (38%, CI: 0.27, 0.49). Respondents acknowledged their expectations that researchers share information about their health and risk for disease and their concerns that their data could be shared with for-profit companies. We found no difference in respondent preferences for registries that shared contact information with researchers, compared to honest broker models that take extra precautions to protect registrant confidentiality (28% versus 30%; p = 0.46). Compared to those preferring a shared information model, respondents who preferred the honest broker model or who lacked model preference voiced increased concerns about sharing registrant data, especially with for-profit organizations. These results suggest that the design of potential participant registries may impact the population enrolled, and hence the population that will eventually be enrolled in clinical studies. Investigators operating registries may need to offer particular assurances about data security to maximize registry enrollment but also must carefully manage participant expectations.

Erin E. Sundermann, Anat Biegon, Leah H. Rubin, Richard B. Lipton, Susan Landau, Pauline M. Maki for the Alzheimer’s Disease Neuroimaging Initiative (Handling Associate Editor: Kerryn Pike)
Does the Female Advantage in Verbal Memory Contribute to Underestimating Alzheimer’s Disease Pathology in Women versus Men?
Abstract: There is a growing recognition of sex differences in Alzheimer’s disease (AD). Females show an advantage over males on tests of verbal memory, which are used to diagnose AD and its precursor, amnestic mild cognitive impairment (aMCI). Women retain this advantage in aMCI despite reduced hippocampal volume and temporal lobe glucose metabolism. Here we examined whether this female advantage endures despite evidence of AD-specific pathology, cortical amyloid-β (Aβ) deposition measured with [18F]AV45 (florbetapir) positron emission tomography. Participants with normal cognition (N=304), aMCI (N=515), and AD dementia (N=175) were drawn from the Alzheimer's Disease Neuroimaging Initiative (ADNI). Across and within diagnostic groups, we conducted linear regressions to examine the interaction of sex with cortical Aβ burden on immediate and delayed recall on the Rey Auditory Verbal Learning Test (RAVLT) adjusting for age, education, and APOE4. In the overall group, sex by cortical Aβ interaction was significant for delayed recall only. Overall, delayed recall performance was significantly better in women versus men among those with low to moderate Aβ burden, but women and men performed similarly among those with high Aβ burden. In diagnosis-stratified analyses, a significant sex by cortical Aβ interaction was observed for delayed recall in the aMCI group, but not in the normal or AD dementia groups. Thus, women maintain a verbal memory advantage over men in aMCI despite similar levels of AD pathology. Although this advantage may benefit women by delaying verbal memory impairment until more advanced pathology, it may also delay diagnosis of aMCI and treatment intervention.

Tian-Syuan Lin*, Han-Jung Tsai*, Chih-Han Lee, Yan-Qing Song, Rih-Sheng Huang, Hsiu-Mei Hsieh-Li, Mei-Chih Liang, Yenshou Lin *These authors contributed equally to this work.
An Improved Drugs Screening System Reveals that Baicalein Ameliorates the Aβ/AMPA/NMDA-Induced Depolarization of Neurons
Abstract: The presence of amyloid-β (Aβ) plaque and tau protein hyperphosphorylation in brain tissue is the pathological hallmark of Alzheimer’s disease (AD). At least some Aβ neurotoxicity is caused by the presence of excess glutamate that has been induced by Aβ accumulation. Memantine is currently the only NMDA receptor inhibitor approved for treating moderate-to-severe AD patients. We utilized primary cortical neurons and DiBAC4(3), a slow-response voltage sensitive fluorescence dye, to create a novel system for screening herbal medicines that allows the identification of pure compounds able to ameliorate Aβ-induced abnormal depolarization. The intensity of DiBAC4(3) fluorescence was increased when primary neurons were stimulated by Aβ; furthermore, pre-treatment with memantine abolished this change. Using this system, we identified six crude extracts made from herbal medicines that effectively alleviated this Aβ-induced abnormal depolarization. Among these herbal medicines, one pure compound, baicalein, which was known to be present in Scutellaria baricalensis and is known to improve memory using an AD mouse model, was identified by our assay. However, the compound's molecular mechanism remained unknown. We found that baicalein, in addition to inhibiting Aβ-induced depolarization, possibly functions as an antagonist of AMPA and NMDA receptors. Taken together, we have established a system/platform to identify herbal medicines that ameliorate Aβ-induced depolarization of neurons. Equally important, baicalein is a candidate drug with great potential for the treatment of AD patients.

Alberto Gomez-Ramos, Angel J. Picher, Esther García, Patricia Garrido, Felix Hernandez, Eduardo Soriano, Jesús Avila
Validation of Suspected Somatic Single Nucleotide Variations in the Brain of Alzheimer’s Disease Patients
Abstract: Next-generation sequencing techniques and genome-wide association study analyses have provided a huge amount of data, thereby enabling the identification of DNA variations and mutations related to disease pathogenesis. New techniques and software tools have been developed to improve the accuracy and reliability of this identification. Most of these tools have been designed to discover and validate single nucleotide variants (SNVs). However, in addition to germ-line mutations, human tissues bear genomic mosaicism, which implies that somatic events are present only in low percentages of cells within a given tissue, thereby hindering the validation of these variations using standard genetic tools. Here we propose a new method to validate some of these somatic mutations. We combine a recently developed software with a method that cuts DNA by using restriction enzymes at the sites of the variation. The non-cleaved molecules, which bear the SNV, can then be amplified and sequenced using Sanger's technique. This procedure, which allows the detection of alternative alleles present in as few as 10% of cells, could be of value for the identification and validation of low frequency somatic events in a variety of tissues and diseases.

Romain Salza, Claire Lethias, Sylvie Ricard-Blum
The Multimerization State of the Amyloid-β42 Amyloid Peptide Governs its Interaction Network with the Extracellular Matrix
Abstract: The goals of this work were i) to identify the interactions of amyloid-β (Aβ)42 under monomeric, oligomeric, and fibrillar forms with the extracellular matrix (ECM) and receptors, ii) to determine the influence of Aβ42 supramolecular organization on these interactions, and iii) to identify the molecular functions, biological processes, and pathways targeted by Aβ42 in the ECM. The ECM and cell surface partners of Aβ42 and its supramolecular forms were identified with protein and glycosaminoglycan (GAG) arrays (81 molecules in triplicate) probed by surface plasmon resonance imaging. The number of partners of Aβ42 increased upon its multimerization, ranging from 4 for the peptide up to 53 for the fibrillar aggregates. The peptide interacted only with ECM proteins but their percentage among Aβ42 partners decreased upon multimerization. Aβ42 and its supramolecular forms recognized different molecular features on their partners, and the partners of Aβ42 fibrillar forms were enriched in laminin IV-A, N-terminal, and EGF-like domains. Aβ42 oligomerization triggered interactions with receptors, whereas Aβ42 fibrillogenesis promoted binding to GAGs, proteoglycans, enzymes, and growth factors and the ability to interact with perineuronal nets. Fibril aggregation bind to further membrane proteins including tumor endothelial marker-8, syndecan-4, and discoidin-domain receptor-2. The partners of the Aβ42 supramolecular forms are enriched in proteins contributing to cell growth and/or maintenance, involved in integrin cell surface interactions and expressed in kidney cancer, preadipocytes, and dentin. In conclusion, the supramolecular assembly of Aβ42 governs its ability to interact in vitro with ECM proteins, remodeling and crosslinking ECM enzymes, proteoglycans, and receptors.

You Joung Kim*, Hoyoung An*, Binna Kim, Young Shin Park, Ki Woong Kim *These authors contributed equally to this work.
An International Comparative Study on Driving Regulations on People with Dementia
Abstract: Over 40% of people with dementia drive, with a two to five times greater accident risk than controls. This has fueled public concerns about the risk of traffic accidents by drivers with dementia (DWD). We compared driving regulations on seniors and DWD between ten European and Asia-Pacific countries to identify key implications for national strategies. Moderate to severe dementia was a reason for driver’s license revocation in all countries. However, regulations on mild dementia varied considerably, with most basing their decisions on severity, rather than simply the presence of dementia. Most used validated assessments, but responsibility for triggering the administrative process fell on drivers in some countries and on physicians in others. Administrations should consider the following when developing driving policies: 1) ideal regulations on DWD should ensure that restrictions are implemented only when needed; 2) fitness to drive should be assessed using validated instruments; 3) the use of processes that automatically initiate driving competency examinations following a diagnosis of dementia should be explored; and 4) restrictions should be delicately tailored to a range of driving competence levels, and assistive incentives compensating for lost driving privileges should be provided.

Han-Kyu Lee, Bumsup Kwon, Cynthia A. Lemere, Suzanne de la Monte, Kyohei Itamura, Austin Y. Ha, Henry W. Querfurth (Handling Associate Editor: Othman Ghribi)
mTORC2 (Rictor) in Alzheimer’s Disease and Reversal of Amyloid-β Expression-Induced Insulin Resistance and Toxicity in Rat Primary Cortical Neurons
Abstract: Mammalian target of rapamycin complex 1 (mTORC1), a nutrient sensor and central controller of cell growth and proliferation, is altered in various models of Alzheimer’s disease (AD). Even less studied or understood in AD is mammalian target of rapamycin complex 2 (mTORC2) that influences cellular metabolism, in part through the regulations of Akt/PKB and SGK. Dysregulation of insulin/PI3K/Akt signaling is another important feature of AD pathogenesis. We found that both total mTORC1 and C2 protein levels and individual C1 and C2 enzymatic activities were decreased in human AD brain samples. In two rodent AD models, mTORC1 and C2 activities were also decreased. In a neuronal culture model of AD characterized by accumulation of cellular amyloid-β (Aβ)42, mTORC1 activity was reduced. Autophagic vesicles and markers were correspondingly increased and new protein synthesis was inhibited, consistent with mTORC1 hypofunction. Interestingly, mTORC2 activity in neural culture seemed resistant to the effects of intracellular amyloid. In various cell lines, Aβ expression provoked insulin resistance, characterized by inhibition of stimulated Akt phosphorylation, and an increase in negative mTORC1 regular, p-AMPK, itself a nutrient sensor. Rapamycin decreased phospho-mTOR and to lesser degree p-Rictor. This further suppression of mTORC1 activity protected cells from Aβ-induced toxicity and insulin resistance. More striking, Rictor over-expression fully reversed the Aβ-effects on primary neuronal cultures. Finally, using in vitro assay, Rictor protein addition completely overcame oligomeric Aβ-induced inhibition of the PDK-Akt activation step. We conclude that striking a new balance by restoring mTORC2 abundance and/or inhibition of mTORC1 has therapeutic potential in AD.

Patrick C.G. Haddick*, Jessica L. Larson*, Nisha Rathore*, Tushar R. Bhangale*, Qui T. Phung, Karpagam Srinivasan, David V. Hansen, Jennie R. Lill, Alzheimer's Disease Genetic Consortium (ADGC), Alzheimer's Disease Neuroimaging Initiative (ADNI), Margaret A. Pericak-Vance, Jonathan Haines, Lindsay A. Farrer, John S. Kauwe, Gerard D. Schellenberg, Carlos Cruchaga, Alison M. Goate, Timothy W. Behrens, Ryan J. Watts, Robert R. Graham, Joshua S. Kaminker, Marcel van der Brug *These authors contributed equally to this work.
A Common Variant of IL-6R is associated with Elevated IL-6 Pathway Activity in Alzheimer’s Disease Brains
Abstract: The common p.D358A variant (rs2228145) in IL-6R is associated with risk for multiple diseases and with increased levels of soluble IL-6R in the periphery and central nervous system (CNS). Here, we show that the p.D358A allele leads to increased proteolysis of membrane bound IL-6R and demonstrate that IL-6R peptides with A358 are more susceptible to cleavage by ADAM10 and ADAM17. IL-6 responsive genes were identified in primary astrocytes and microglia and an IL-6 gene signature was increased in the CNS of late onset Alzheimer’s disease subjects in an IL6R allele dependent manner. We conducted a screen to identify variants associated with the age of onset of Alzheimer’s disease in APOE ε4 carriers. Across five datasets, p.D358A had a meta P=3 x10-4 and an odds ratio = 1.3, 95% confidence interval 1.12 – 1.48. Our study suggests that a common coding region variant of the IL-6 receptor results in neuroinflammatory changes that may influence the age of onset of Alzheimer’s disease in APOE ε4 carriers.

Rosanna Squitti, Armando J. Mendez, Ilaria Simonelli, Camillo Ricordi
Diabetes and Alzheimer’s Disease: Can Elevated Free Copper Predict the Risk of the Disease?
Abstract: Background: Defective copper regulation, primarily referred to as chelatable redox active Cu(II), has been involved in the etiology of diabetes, and Alzheimer’s disease (AD). Objectives: However, no study has determined levels of labile copper non-bound to ceruloplasmin (non-Cp Cu, also known as ‘free’ copper) in the blood of subjects with diabetes compared with that of AD patients. Methods: To this aim, values of non-Cp Cu were measured in 25 Type 1 (T1D) and 31 Type 2 (T2D) subjects and in 28 healthy controls, along with measurements of C-reactive protein, glycated hemoglobin A1c, cholesterol, and triglycerides. Non-Cp Cu levels were compared with those of an AD group previously studied. Results: T2D subjects had significantly higher non-Cp Cu levels than Controls and T1D subjects (both p < 0.001 after adjusting for age, sex, and body mass index). A multinomial logistic model revealed that a one unit standard deviation increase of non-Cp Cu increased the relative risk of having T2D by 9.64 with respect to Controls (95% CI: 2.86-32.47). The comparison of non-Cp Cu levels in T2D with those of an AD population previously studied shows rising blood non-Cp Cu copper levels from Controls to T2D and AD. Conclusion: These results suggest the involvement of catalytically-active Cu(II) and glucose dysregulation in oxidative stress reactions leading to tissue damage in both diseases.

Gemma Tell-Marti, Joan Anton Puig-Butille, Miriam Potrony, Estel Plana, Celia Badenas, Anna Antonell, Raquel Sanchez-Valle, José L Molinuevo, Alberto Lleó, Daniel Alcolea, Juan Fortea, Ruben Fernandez-Santiago, Jordi Clarimón, Albert Lladó, Susana Puig
A Common Variant in the MC1R Gene (p.V92M) is associated with Alzheimer’s Disease Risk
Abstract: Despite the recent identification of some novel risk genes for Alzheimer’s disease (AD), the genetic etiology of late-onset Alzheimer’s disease (LOAD) remains largely unknown. The inclusion of these novel risk genes to the risk attributable to the APOE gene accounts for roughly half of the total genetic variance in LOAD. The evidence indicates that undiscovered genetic factors may contribute to AD susceptibility. In the present study, we sequenced the MC1R gene in 525 Spanish LOAD patients and in 160 controls. We observed that a common MC1R variant p.V92M (rs2228479), not related to pigmentation traits, was present in 72 (14%) patients and 15 (9%) controls and confers increased risk of developing LOAD (OR: 1.99, 95% CI: 1.08-3.64, p = 0.026), especially in those patients whose genetic risk could not be explained by APOE genotype. This association remains and even increased in the subset of 69 patients with typical AD cerebrospinal fluid profile (OR: 3.40 95% CI: 1.40-8.27, p = 0.007). We did not find an association between p.V92M and age of onset of AD. Further studies are necessary to elucidate the role of MC1R in brain cells through the different MC1R pathways.

Nicholas F. Fitz*,**, Alexis Y. Carter*, Victor Tapias, Emilie L. Castranio, Ravindra Kodali, Iliya Lefterov, Radosveta Koldamova** (Handling Associate Editor: Rakez Kayed) *These authors contributed equally to this work. **Co-senior authors
Abca1 Deficiency Affects Basal Cognitive Deficits and Dendritic Density in Mice
Abstract: ATP-binding cassette transporter A1 (ABCA1) mediates cholesterol efflux to lipid-free apolipoproteins and regulates the generation of high density lipoproteins. Previously, we have shown that lack of Abca1 significantly increases amyloid deposition and cognitive deficits in Alzheimer’s disease model mice expressing human amyloid-β protein precursor (APP). The goal of this study was to determine if ABCA1 plays a role in memory deficits caused by amyloid-β (Aβ) oligomers and examine neurite architecture of pyramidal hippocampal neurons. Our results confirm previous findings that Abca1 deficiency significantly impairs spatial memory acquisition and retention in the Morris water maze and long-term memory in novel object recognition of APP transgenic mice at a stage of early amyloid pathology. Neither test demonstrated a significant difference between Abca1ko and wild-type (WT) mice. We also examined the effect of intra-hippocampal infused Aβ oligomers on cognitive performance of Abca1ko mice, compared to control infusion of scrambled Aβ peptide. Age-matched WT mice undergoing the same infusions were also used as controls. In this model system, we found a statistically significant difference between WT and Abca1ko mice infused with scrambled Aβ, suggesting that Abca1ko mice are vulnerable to the effect of mild stresses. Moreover, examination of neurite architecture in the hippocampi revealed a significant decrease in neurite length, number of neurite segments, and branches in Abca1ko mice when compared to WT mice. We conclude that mice lacking ABCA1 have basal cognitive deficits that prevent them from coping with additional stressors, which is in part due to impairment of neurite morphology of the hippocampus.

Loqman A. Mohamed, Haihao Zhu, Youssef M. Mousa, Erming Wang, Wei Qiao Qiu, Amal Kaddoumi (Handling Associate Editor: Eva Carro)
Amylin Enhances Amyloid-β Peptide Brain to Blood Efflux Across the Blood-Brain Barrier
Abstract: Findings from Alzheimer's disease (AD) mouse models showed that amylin treatment improved AD pathology and enhanced amyloid-β (Aβ) brain to blood clearance; however, the mechanism was not investigated. Using the Tg2576 AD mouse model, a single intraperitoneal injection of amylin significantly increased Aβ serum levels, and the effect was abolished by AC253, an amylin receptor antagonist, suggesting that amylin effect could be mediated by its receptor. Subsequent mechanistic studies showed amylin enhanced Aβ transport across a cell-based model of the blood-brain barrier (BBB), an effect that was abolished when the amylin receptor was inhibited by two amylin antagonists and by siRNA knockdown of amylin receptor Ramp3. To explain this finding, amylin effect on Aβ transport proteins expressed at the BBB was evaluated. Findings indicated that cells treated with amylin induced LRP1 expression, a major receptor involved in brain Aβ efflux, in plasma membrane fraction, suggesting intracellular translocation of LRP1 from the cytoplasmic pool. Increased LRP1 in membrane fraction could explain, at least in part, the enhanced uptake and transport of Aβ across the BBB. Collectively, our findings indicated that amylin induced Aβ brain to blood clearance through amylin receptor by inducing LRP1 subcellular translocation to the plasma membrane of the BBB endothelium.

Laura Ferrari, Su-Chun Huang, Giuseppe Magnani, Alessandro Ambrosi, Giancarlo Comi, Letizia Leocani
Optical Coherence Tomography Reveals Retinal Neuroaxonal Thinning in Frontotemporal Dementia as in Alzheimer's Disease
Abstract: Background: Alzheimer’s disease (AD) and frontotemporal dementia (FTD) are leading causes of cognitive decline. Optical coherence tomography (OCT) allows the measurement of thickness of retinal neuroaxonal layers. While in AD and mild cognitive impairment (MCI), retinal nerve fiber layer (RNFL) thinning is frequently reported, less information is available on ganglion cell layer-inner plexiform layer (GCL-IPL). Data on FTD are lacking. Objective: To obtain cross-sectional information on RNFL and GCL-IPL thickness among MCI, AD, FTD, and healthy controls (HC), and their correlations with dementia severity. Methods: Peripapillary OCT scans were obtained in 27 MCI, 39 AD, 17 FTD, 49 HC using high-definition Heidelberg Spectral-domain OCT, with RNFL and GCL-IPL thickness measurement. Statistical analysis tested group effects and correlation with gender, disease duration and severity (Mini-Mental State Examination, MMSE). Results: RNFL showed a significant group effect [F(4,132)=3.786, p=0.006], being reduced versus controls in MCI (p=0.033), moderate AD (p=0.025), and FTD (p<0.001), and versus mild AD in FTD (p=0.042). GCL-IPL showed a significant group effect as well [F(4,121)=5.104, p<0.001], with reduction in moderate AD versus HC (p<0.001), MCI (p=0.037), and mild AD (p=0.009); in FTD versus HC (p=0.002) and mild AD (p=0.038). In AD, GCL-IPL correlated with MMSE (r=0.487, p=0.003), without significant effects of age, gender, or disease duration. Conclusion: Retinal neuroaxonal thinning occurs in MCI/AD consistently with previous reports, as well as in FTD. Correlation with disease severity in AD suggests that retinal and brain neurodegeneration may occur in parallel to some extent, and prompts larger studies aimed at providing surrogate endpoints for clinical trials in AD.

Cassandra Kaizik, Jashelle Caga, Julieta Camino, Claire M O’Connor, Colleen McKinnon, Jan R. Oyebode, Olivier Piguet, John R. Hodges, Eneida Mioshi
Factors Underpinning Caregiver Burden in Frontotemporal Dementia Differ in Spouses and their Children
Abstract: The objectives of this observational study were to (1) compare spousal and child caregiver burden; (2) compare co-resident and live-out child caregiver burden; and (3) investigate factors influencing spousal and child caregiver burden. Data was collected from 90 caregivers of people with frontotemporal degeneration (FTD) recruited from the Frontotemporal Dementia Research Group (Frontier) at Neuroscience Research, Australia. Of this caregiver group, 43 were spousal caregivers and 47 were child caregivers. Caregiver burden and emotional state were evaluated using the short Zarit Burden Interview and the short version of the Depression, Anxiety and Stress Scale-21. The Social Network Index was applied to ascertain the social network of the caregiver, while the Intimate Bond Measure was used to evaluate the current quality of the relationship between the caregiver and the person with dementia. The Frontotemporal Dementia Rating Scale was used to assess severity of dementia. Spousal and child caregivers experienced similar levels of burden, depression, anxiety, and stress, regardless of disease severity. Co-resident child caregivers had smaller social networks and greater burden than live-out caregivers. Dementia severity was key in spousal caregiver burden, whereas caregiver depression was most important in child caregiver burden. Child and spousal caregivers of individuals with FTD share similar levels of burden, influenced by different factors. Future interventions need to account for these differences.

David A. Loewenstein, Rosie E. Curiel, Clinton Wright, Xiaoyan Sun, Noam Alperin, Elzabeth Crocco, Sara J. Czaja, Arlene Raffo, Ailyn Penate, Jose Melo, Kimberly Capp, Monica Gamez, Ranjan Duara
Recovery from Proactive Semantic Interference in Mild Cognitive Impairment and Normal Aging: Relationship to Atrophy in Brain Regions Vulnerable to Alzheimer’s Disease
Abstract: Background: There is growing evidence that proactive semantic interference (PSI) and failure to recover from PSI may represent early features of Alzheimer’s disease (AD). Objective: This study investigated the association between PSI, recovery from PSI, and reduced MRI volumes in AD signature regions among cognitively impaired and unimpaired older adults. Methods: Performance on the LASSI-L (a novel test of PSI and recovery from PSI) and regional brain volumetric measures were compared between 38 cognitively normal (CN) elders and 29 older participants with amnestic mild cognitive impairment (MCI). The relationship between MRI measures and performance on the LASSI-L as well as traditional memory and non-memory cognitive measures was also evaluated in both diagnostic groups. Results: Relative to traditional neuropsychological measures, MCI patients’ failure to recover from PSI was associated with reduced volumes in the hippocampus (rs=0.48), precuneus (rs=0.50); rostral middle frontal lobules (rs=0.54); inferior temporal lobules (rs=0.49), superior parietal lobules (rs=0.47), temporal pole (rs=0.44), and increased dilatation of the inferior lateral ventricle (rs=-0.49). For CN elders, only increased inferior lateral ventricular size was associated with vulnerability to PSI (rs=-0.49), the failure to recover from PSI (rs=-0.57), and delayed recall on the Hopkins Verbal Learning Test-Revised (rs=-0.48). Discussion: LASSI-L indices eliciting failure to recover from PSI were more highly associated with more MRI regional biomarkers of AD than other traditional cognitive measures. These results as well as recent amyloid imaging studies with otherwise cognitively normal subjects, suggest that recovery from PSI may be a sensitive marker of preclinical AD and deserves further investigation.

Kirsten E. Peters, Wendy A. Davis, Kevin Taddei, Ralph N. Martins, Colin L. Masters, Timothy M.E. Davis, David G. Bruce
Plasma Amyloid-β Peptides in Type 2 Diabetes: A Matched Case-Control Study
Abstract: Background: Plasma amyloid-β (Aβ) levels have rarely been investigated in type 2 diabetes despite its known associations with Alzheimer’s disease. Objective: To compare blood plasma Aβ concentrations (Aβ40 and Aβ42) in cognitively normal individuals with and without type 2 diabetes. Methods: Plasma Aβ40 and Aβ42 were measured in 194 participants with diabetes recruited from the community-based Fremantle Diabetes Study Phase II cohort (mean age 71 years, 59% males) and 194 age-, sex-, and APOE ε4 allele-matched, control subjects without diabetes from the Australian Imaging, Biomarkers and Lifestyle Study using a multiplex microsphere-based immunoassay. Results: Plasma Aβ40 and Aβ42 were normally distributed in the controls but were bimodal in the participants with diabetes. Median Aβ40 and Aβ42 concentrations were significantly lower in those with type 2 diabetes (Aβ40 median [inter-quartile range]: 125.0 [52.6-148.3] versus 149.3 [134.0-165.6] pg/mL; Aβ42: 26.9 [14.5-38.3] versus 33.6 [28.0-38.9] pg/mL, both p<0.001) while the ratio Aβ42:Aβ40 was significantly higher (0.26 [0.23-0.32] versus 0.22 [0.19-0.25], p<0.001). After adjustment, participants with diabetes and plasma Aβ40 levels in the low peak of the bimodal distribution were significantly more likely to have normal to high estimated glomerular filtration rates (odds ratio (95% CI): 2.40 (1.20-4.80), p=0.013) although the group with diabetes had lower renal function overall. Conclusion: Type 2 diabetes is associated with altered plasma concentrations of Aβ peptides and is an important source of variation that needs to be taken into account when considering plasma Aβ peptides as biomarkers for Alzheimer’s disease.

Lukas Kunz, Martin Reuter, Nikolai Axmacher, Christian Montag
Conscientiousness is Negatively Associated with Grey Matter Volume in Young APOE ε4-Carriers
Abstract: The etiology of late onset Alzheimer’s disease (LOAD) depends on multiple factors, among which the APOE ε4 allele is the most adverse genetic determinant and conscientiousness represents an influential personality trait. A potential association of both factors with brain structure in young adulthood may constitute a constellation that sets the course toward or against the subtle disease progression of LOAD that starts decades before clinical manifestation. Hence, in the present study, we examined the modulating effects of APOE ε4 on the relation between personality dimensions, including conscientiousness, and total grey matter volume (GMV) in young healthy adults using an a priori genotyping design. 105 participants completed an inventory assessing the Five Factor Model of Personality (NEO-FFI) and a structural MRI scan. Total GMV was estimated using both Freesurfer as well as VBM8. Across all participants, total GMV was positively associated with extraversion and negatively related to age. In APOE ε4-carriers—but not in APOE ε4-non-carriers—conscientiousness was negatively associated with total GMV. In line with the hypothesis of antagonistic pleiotropy of the APOE ε4 allele, this result suggests that young APOE ε4-carriers with increased total GMV may particularly benefit from cognitive advantages and thus have a lower need to engage in conscientious behavior. In this subset of young APOE ε4-carriers, the reduction in conscientiousness could then bring along adverse health behavior in the long run, potentiating the risk for LOAD. Hence, young APOE ε4-carriers with increased total GMV may be at a particularly high risk for LOAD.

Marlena Zyśk, Hanna Bielarczyk, Sylwia Gul-Hinc, Aleksandra Dyś, Beata Gapys, Anna Ronowska, Monika Sakowicz-Burkiewicz, Andrzej Szutowicz (Handling Associate Editor: Martin Kleinschmidt)
Phenotype-Dependent Interactions between N-acetyl-L-Aspartate and Acetyl-CoA in Septal SN56 Cholinergic Cells Exposed to an Excess of Zinc
Abstract: Pyruvate dehydrogenase reaction utilizing glucose-derived pyruvate is an almost exclusive source of acetyl-CoA in different cell mitochondrial compartments of the brain. In neuronal mitochondria, the largest fraction of acetyl-CoA is utilized for energy production and the much smaller one for N-acetyl-L-aspartate (NAA) synthesis. Cholinergic neurons, unlike others, require additional amounts of acetyl-CoA for acetylcholine synthesis. Therefore, several neurotoxic signals, which inhibit pyruvate dehydrogenase, generate deeper shortages of acetyl-CoA and greater mortality of cholinergic neurons than noncholinergic ones. NAA is considered to be a marker of neuronal energy status in neuropathic brains. However, there is no data on putative differential fractional distribution of the acetyl-CoA pool between energy producing and NAA or acetylcholine synthesizing pathways in noncholinergic and cholinergic neurons, respectively. Therefore, the aim of this study was to investigate whether zinc-excess, a common excitotoxic signal, may evoke differential effects on the NAA metabolism in neuronal cells with low and high expression of the cholinergic phenotype. Differentiated SN56 neuronal cells, displaying a high activity of choline acetyltransferase and rates of acetylcholine synthesis, contained lower levels of acetyl-CoA and NAA, being more susceptible to ZnCl2 exposition that the nondifferentiated SN56 or differentiated dopaminergic SHSY5Y neuronal and astroglial C6 cells. Differentiated SN56 accumulated greater amounts of Zn2+ from extracellular space than the other ones, and displayed a stronger suppression of pyruvate dehydrogenase complex activity and acetyl-CoA, NAA, ATP, acetylcholine levels, and loss of viability. These data indicate that the acetyl-CoA synthesizing system in neurons constitutes functional unity with energy generating and NAA or acetylcholine pathways of its utilization, which are uniformly affected by neurotoxic conditions.

Wasim Khan, Vincent Giampietro, Tobias Banaschewski, Gareth J. Barker, Arun L.W. Bokde, Christian Büchel, Patricia Conrod, Herta Flor, Vincent Frouin, Hugh Garavan, Penny Gowland, Anreas Heinz, Bernd Ittermann, Hervé Lemaître, Frauke Nees, Tomas Paus, Zdenka Pausova, Marcella Rietschel, Michael N. Smolka, Andreas Ströhle, Jeurgen Gallinat, Bruno Vellas, Hilkka Soininen, Iwona Kloszewska, Magda Tsolaki, Patrizia Mecocci, Christian Spenger, Victor L. Villemagne, Colin L. Masters, J-Sebastian Muehlboeck, Lars Bäckman, Laura Fratiglioni, Grégoria Kalpouzos, Lars-Olof Wahlund, Gunther Schumann, Simon Lovestone, Steven C.R. Williams, Eric Westman, Andrew Simmons; Alzheimer’s Disease Neuroimaging Initiative, AddNeuroMed Consortium, Australian, Imaging, Biomarkers, and Lifestyle Study Research Group, and the IMAGEN consortium (Handling Associate Editor: Maheen Adamson)
A Multi-Cohort Study of ApoE ε4 and Amyloid-β Effects on the Hippocampus in Alzheimer’s Disease
Abstract: The apolipoprotein E (APOE) gene has been consistently shown to modulate the risk of Alzheimer’s disease (AD). Here, using an AD and normal aging dataset primarily consisting of three AD multi-center studies (n=1,781), we compared the effect of APOE and amyloid-β (Aβ) on baseline hippocampal volumes in AD patients, mild cognitive impairment (MCI) subjects, and healthy controls. A large sample of healthy adolescents (n=1,387) was also used to compared hippocampal volumes between APOE groups. Subjects had undergone a magnetic resonance imaging (MRI) scan and APOE genotyping. Hippocampal volumes were processed using FreeSurfer. In the AD and normal aging dataset, hippocampal comparisons were performed in each APOE group and in ε4 carriers with positron emission tomography Aβ who were dichotomized (Aβ+/Aβ-) using previous cut-offs. We found a linear reduction in hippocampal volumes with ε4 carriers possessing the smallest volumes, ε3 carriers possessing intermediate volumes, and ε2 carriers possessing the largest volumes. Moreover, AD and MCI ε4 carriers possessed the smallest hippocampal volumes and control ε2 carriers possessed the largest hippocampal volumes. Subjects with both APOE ε4 and Aβ+ had the lowest hippocampal volumes when compared to Aβ- ε4 carriers, suggesting a synergistic relationship between APOE ε4 and Aβ. However, we found no hippocampal volume differences between APOE groups in healthy 14-year-old adolescents. Our findings suggest that the strongest neuroanatomic effect of APOE ε4 on the hippocampus is observed in AD and groups most at risk of developing the disease, whereas hippocampi of old and young healthy individuals remain unaffected.

András Horváth, Anna Szűcs, Gábor Barcs, Anita Kamondi (Handling Associate Editor: Brian Lebowitz)
Sleep EEG Detects Epileptiform Activity in Alzheimer’s Disease with High Sensitivity
Abstract: Background: The reported prevalence of epilepsy in Alzheimer’s disease (AD) is variable, probably due to the different methodological approaches. Objective: We aimed to define the optimal electroencephalogram (EEG) settings for reliable detection of epileptiform discharges in AD patients. Methods: We analyzed 24-h EEGs of 5 patients living with AD and epilepsy. The sensitivity of various length EEGs in detecting epileptiform discharges in different periods of the day, the diurnal distribution of the discharges, and their relation to sleep-stages were calculated. Results: Significant high correlation was identified between the sensitivity of EEG and the length of recordings (r=0.972, p=0.005). The sensitivity of a 30-min EEG-epoch recorded between 8:00 and 16:00 was 0.0375 compared to 0.7 between 0:00 and 8:00 (p=0.005). The average sensitivity of an 8-h EEG-epoch was ≥0.8. 82% of epileptiform discharges occurred during sleep, mainly related to non-REM sleep (p<0.001). Conclusion: 8-h awake-, or 1-h sleep-EEG provide sufficient sensitivity in detecting epileptiform activity in AD. This needs to be considered in studies on AD-related epilepsy. Recognizing epilepsy in AD patients is essential because it might compromise cognitive functions and accelerate the progression of the disease.

Alexandra Polcher, Ingo Frommann, Alexander Koppara, Steffen Wolfsgruber, Frank Jessen, Michael Wagner
Face-Name Associative Recognition Deficits in Subjective Cognitive Decline and Mild Cognitive Impairment
Abstract: Background: There is a need for more sensitive neuropsychological tests to detect subtle cognitive deficits emerging in the preclinical stage of Alzheimer’s disease (AD). Associative memory is a cognitive function supported by the hippocampus and affected early in the process of AD. Objective: We developed a short computerized face-name-associative-recognition test (FNART) and tested whether it would detect memory impairment in memory-clinic patients with mild cognitive impairment (MCI) and subjective cognitive decline (SCD). Methods: We recruited 61 elderly patients with either SCD (n=32) or MCI (n=29) and 28 healthy controls (HC) and compared performance on FNART, self-reported cognitive deterioration in different domains (ECog-39), and, in a reduced sample (n=46), performance on the visual paired associates test of the CANTAB battery. Results: A significant effect of group on FNART test performance in the total sample was found (p<0.001). Planned contrasts indicated a significantly lower associative memory performance in the SCD (p=0.001, d=0.82) and MCI group (p<0.001, d=1.54), as compared to HCs, respectively. The CANTAB-PAL discriminated only between HC and MCI, possibly because of reduced statistical power. Adjusted for depression, performance on FNART was significantly related to ECog-39 Memory in SCD patients (p=0.024) but not in MCI patients. Conclusions: Associative memory is substantially impaired in memory-clinic patients with SCD and correlates specifically with memory complaints at this putative preclinical stage of AD. Further studies will need to examine the predictive validity of the FNART in SCD patients with regard to longitudinal (i.e., conversion to MCI/AD) and biomarker outcomes.

Dalton L. Ebel*, Christopher G. Torkilsen*, Tim D. Ostrowski *These authors contributed equally to this work.
Blunted Respiratory Responses in the Streptozotocin-Induced Alzheimer’s Disease Rat Model
Abstract: Alzheimer’s disease (AD) is known for the progressive decline of cognition and memory. In addition to these disease-defining symptoms, impairment of respiratory function is frequently observed and often expressed by sleep-disordered breathing or reduced ability to adjust respiration when oxygen demand is elevated. The mechanisms for this are widely unknown. Postmortem analysis from the brainstem of AD patients reveals pathological alterations, including in nuclei responsible for respiratory control. In this study, we analyzed respiratory responses and morphological changes in brainstem nuclei following intracerebroventricular (ICV) injections of streptozotocin (STZ), a rat model commonly used to mimic sporadic AD. ICV-STZ induced significant astrogliosis in the commissural part of the nucleus tractus solitarii, an area highly involved in respiration control. The astrogliosis was identified by a significant increase in S100B-immunofluorescence that is similar to the astrogliosis found in the CA1 region of the hippocampus. Using plethysmography, the control group displayed a typical age-dependent decrease of ventilation that was absent in the STZ rat group. This is indicative of elevated minute ventilation at rest after STZ treatment. Peripheral chemoreflex responses were significantly blunted in STZ rats as seen by a reduced respiratory rate and minute ventilation to hypoxia. Central chemoreflex responses to hypercapnia, on the other hand, only decreased in respiratory rate following STZ treatment. Overall, our results show that ICV-STZ induces respiratory dysfunction at rest and in response to hypoxia. This provides a new tool to study the underlying mechanisms of breathing disorders in clinical AD.