Longitudinal Study of CSF Biomarkers in Patients with Alzheimer's Disease

Background

The CSF biomarkers tau and Aβ42 can identify patients with AD, even during the preclinical stages. However, previous studies on longitudinal changes of tau and Aβ42 in individual patients with AD and elderly controls report somewhat inconsistent results.

Methodology/Principal Findings

We investigated the levels of tau and Aβ42 at baseline and after 1 year in 100 patients with AD. In a second cohort of 45 AD patients we measured the CSF biomarkers at baseline and after 2 years. Moreover, in 34 healthy elderly controls the CSF biomarkers were followed for 4 years. The baseline levels of tau were increased with >60% in AD patients compared to controls (p<0.001), while baseline Aβ42 levels were decreased with >50% (p<0.001). In the AD group followed for 2 years, tau increased with 16% compared to the baseline levels (p<0.05). However, the levels of tau were stable over 4 years in the controls. The levels of Aβ42 did not change significantly over time in any of the groups. In the patients with AD, tau was moderately associated with worse cognitive performance already at baseline (p<0.05).

Conclusions/Significance

Tau and Aβ42 in CSF seem to reflect the underlying disease state in both early and late stages of AD. The slight increase in tau over time observed in the patients with AD is modest when compared to the relatively large difference in absolute tau levels between AD patients and controls. Therefore, these markers maintain their usefulness as state markers over time and might serve as surrogate markers for treatment efficacy in clinical trials.     

Impact of Cerebrospinal Fluid Shunting for Idiopathic Normal Pressure Hydrocephalus on the Amyloid Cascade

The aim of this study was to determine whether the improvement of cerebrospinal fluid (CSF) flow dynamics by CSF shunting, can suppress the oligomerization of amyloid β-peptide (Aβ), by measuring the levels of Alzheimer’s disease (AD)-related proteins in the CSF before and after lumboperitoneal shunting. Lumbar CSF from 32 patients with idiopathic normal pressure hydrocephalus (iNPH) (samples were obtained before and 1 year after shunting), 15 patients with AD, and 12 normal controls was analyzed for AD-related proteins and APLP1-derived Aβ-like peptides (APL1β) (a surrogate marker for Aβ). We found that before shunting, individuals with iNPH had significantly lower levels of soluble amyloid precursor proteins (sAPP) and Aβ38 compared to patients with AD and normal controls. We divided the patients with iNPH into patients with favorable (improvement ≥ 1 on the modified Rankin Scale) and unfavorable (no improvement on the modified Rankin Scale) outcomes. Compared to the unfavorable outcome group, the favorable outcome group showed significant increases in Aβ38, 40, 42, and phosphorylated-tau levels after shunting. In contrast, there were no significant changes in the levels of APL1β25, 27, and 28 after shunting. After shunting, we observed positive correlations between sAPPα and sAPPβ, Aβ38 and 42, and APL1β25 and 28, with shifts from sAPPβ to sAPPα, from APL1β28 to 25, and from Aβ42 to 38 in all patients with iNPH. Our results suggest that Aβ production remained unchanged by the shunt procedure because the levels of sAPP and APL1β were unchanged. Moreover, the shift of Aβ from oligomer to monomer due to the shift of Aβ42 (easy to aggregate) to Aβ38 (difficult to aggregate), and the improvement of interstitial-fluid flow, could lead to increased Aβ levels in the CSF. Our findings suggest that the shunting procedure can delay intracerebral deposition of Aβ in patients with iNPH.     

Cerebrospinal Fluid (CSF) Neuronal Biomarkers across the Spectrum of HIV Infection: Hierarchy of Injury and Detection

The character of central nervous system (CNS) HIV infection and its effects on neuronal integrity vary with evolving systemic infection. Using a cross-sectional design and archived samples, we compared concentrations of cerebrospinal fluid (CSF) neuronal biomarkers in 143 samples from 8 HIV-infected subject groups representing a spectrum of untreated systemic HIV progression and viral suppression: primary infection; four groups of chronic HIV infection neuroasymptomatic (NA) subjects defined by blood CD4+ T cells of >350, 200–349, 50–199, and <50 cells/µL; HAD; treatment-induced viral suppression; and ‘elite’ controllers. Samples from 20 HIV-uninfected controls were also examined. The neuronal biomarkers included neurofilament light chain protein (NFL), total and phosphorylated tau (t-tau, p-tau), soluble amyloid precursor proteins alpha and beta (sAPPα, sAPPβ) and amyloid beta (Aβ) fragments 1–42, 1–40 and 1–38. Comparison of the biomarker changes showed a hierarchy of sensitivity in detection and suggested evolving mechanisms with progressive injury. NFL was the most sensitive neuronal biomarker. Its CSF concentration exceeded age-adjusted norms in all HAD patients, 75% of NA CD4<50, 40% of NA CD4 50–199, and 42% of primary infection, indicating common neuronal injury with untreated systemic HIV disease progression as well as transiently during early infection. By contrast, only 75% of HAD subjects had abnormal CSF t-tau levels, and there were no significant differences in t-tau levels among the remaining groups. sAPPα and β were also abnormal (decreased) in HAD, showed less marked change than NFL with CD4 decline in the absence of HAD, and were not decreased in PHI. The CSF Aβ peptides and p-tau concentrations did not differ among the groups, distinguishing the HIV CNS injury profile from Alzheimer''s disease. These CSF biomarkers can serve as useful tools in selected research and clinical settings for patient classification, pathogenetic analysis, diagnosis and management.     

Drosophila Eye Model to Study Neuroprotective Role of CREB Binding Protein (CBP) in Alzheimer’s Disease

Background

The progressive neurodegenerative disorder Alzheimer’s disease (AD) manifests as loss of cognitive functions, and finally leads to death of the affected individual. AD may result from accumulation of amyloid plaques. These amyloid plaques comprising of amyloid-beta 42 (Aβ42) polypeptides results from the improper cleavage of amyloid precursor protein (APP) in the brain. The Aβ42 plaques have been shown to disrupt the normal cellular processes and thereby trigger abnormal signaling which results in the death of neurons. However, the molecular-genetic mechanism(s) responsible for Aβ42 mediated neurodegeneration is yet to be fully understood.

Methodology/Principal Findings

We have utilized Gal4/UAS system to develop a transgenic fruit fly model for Aβ42 mediated neurodegeneration. Targeted misexpression of human Aβ42 in the differentiating photoreceptor neurons of the developing eye of transgenic fly triggers neurodegeneration. This progressive neurodegenerative phenotype resembles Alzheimer’s like neuropathology. We identified a histone acetylase, CREB Binding Protein (CBP), as a genetic modifier of Aβ42 mediated neurodegeneration. Targeted misexpression of CBP along with Aβ42 in the differentiating retina can significantly rescue neurodegeneration. We found that gain-of-function of CBP rescues Aβ42 mediated neurodegeneration by blocking cell death. Misexpression of Aβ42 affects the targeting of axons from retina to the brain but misexpression of full length CBP along with Aβ42 can restore this defect. The CBP protein has multiple domains and is known to interact with many different proteins. Our structure function analysis using truncated constructs lacking one or more domains of CBP protein, in transgenic flies revealed that Bromo, HAT and polyglutamine (BHQ) domains together are required for the neuroprotective function of CBP. This BHQ domain of CBP has not been attributed to promote survival in any other neurodegenerative disorders.

Conclusions/Significance

We have identified CBP as a genetic modifier of Aβ42 mediated neurodegeneration. Furthermore, we have identified BHQ domain of CBP is responsible for its neuroprotective function. These studies may have significant bearing on our understanding of genetic basis of AD.     

No correlation between time-linked plasma and CSF Aβ levels

Plasma β-amyloid protein (Aβ) isoforms are considered potential biomarkers for Alzheimer's disease (AD) and dementia. The relation between plasma and cerebrospinal fluid (CSF) levels of Aβ isoforms remains unclear. In order to identify possible correlations between Aβ levels in plasma and CSF we determined Aβ levels in time-linked plasma and CSF samples. Aβ concentrations in plasma (Aβ_1–42 and Aβ_N–42) and CSF (Aβ_1–42) samples from 49 AD patients, 47 non-Alzheimer's disease dementia (NONAD) patients, 39 MCI patients and 29 controls were determined using a multi-parameter fluorimetric bead-based immunoassay using xMAP^® technology (for plasma) and a conventional single-parameter ELISA (for CSF). Plasma Aβ_1–42 concentrations did not correlate with CSF Aβ_1–42 concentrations in the total study population, or in the different diagnostic groups. No correlations between plasma Aβ_N–42 and CSF Aβ_1–42 levels were found either. The CSF/serum albumin index did not show any significant differences between AD, NONAD, MCI and controls.These results suggest that the Aβ levels in plasma are independent of the Aβ levels in CSF both in dementia and controls. The fact that CSF and plasma Aβ do not correlate in patients as well as controls and no significant differences in plasma Aβ_1–42 or Aβ_N–42 between patients and controls can be detected hampers the diagnostic utility of the plasma Aβ levels as biomarkers for dementia.     

Variations in the FRA10AC1 Fragile Site and 15q21 Are Associated with Cerebrospinal Fluid Aβ1-42 Level

Proteolytic fragments of amyloid and post-translational modification of tau species in Cerebrospinal fluid (CSF) as well as cerebral amyloid deposition are important biomarkers for Alzheimer’s Disease. We conducted genome-wide association study to identify genetic factors influencing CSF biomarker level, cerebral amyloid deposition, and disease progression. The genome-wide association study was performed via a meta-analysis of two non-overlapping discovery sample sets to identify genetic variants other than APOE ε4 predictive of the CSF biomarker level (Aβ_1–42, t-Tau, p-Tau_181P, t-Tau:Aβ_1–42 ratio, and p-Tau_181P:Aβ_1–42 ratio) in patients enrolled in the Alzheimer’s Disease Neuroimaging Initiative (ADNI) study. Loci passing a genome-wide significance threshold of P < 5 x 10⁻⁸ were followed-up for replication in an independent sample set. We also performed joint meta-analysis of both discovery sample sets together with the replication sample set. In the discovery phase, we identified variants in FRA10AC1 associated with CSF Aβ_1–42 level passing the genome-wide significance threshold (directly genotyped SNV rs10509663 P _FE = 1.1 x 10⁻⁹, imputed SNV rs116953792 P _FE = 3.5 x 10⁻¹⁰), rs116953792 (P _one-sided = 0.04) achieved replication. This association became stronger in the joint meta-analysis (directly genotyped SNV rs10509663 P _FE = 1.7 x 10⁻⁹, imputed SNV rs116953792 P _FE = 7.6 x 10⁻¹¹). Additionally, we identified locus 15q21 (imputed SNV rs1503351 P _FE = 4.0 x 10⁻⁸) associated with CSF Aβ_1–42 level. No other variants passed the genome-wide significance threshold for other CSF biomarkers in either the discovery sample sets or joint analysis. Gene set enrichment analyses suggested that targeted genes mediated by miR-33, miR-146, and miR-193 were enriched in various GWAS analyses. This finding is particularly important because CSF biomarkers confer disease susceptibility and may be predictive of the likelihood of disease progression in Alzheimer’s Disease.     

Apolipoprotein A-I Deficiency Increases Cerebral Amyloid Angiopathy and Cognitive Deficits in APP/PS1ΔE9 Mice

A hallmark of Alzheimer disease (AD) is the deposition of amyloid β (Aβ) in brain parenchyma and cerebral blood vessels, accompanied by cognitive decline. Previously, we showed that human apolipoprotein A-I (apoA-I) decreases Aβ₄₀ aggregation and toxicity. Here we demonstrate that apoA-I in lipidated or non-lipidated form prevents the formation of high molecular weight aggregates of Aβ₄₂ and decreases Aβ₄₂ toxicity in primary brain cells. To determine the effects of apoA-I on AD phenotype in vivo, we crossed APP/PS1ΔE9 to apoA-I^KO mice. Using a Morris water maze, we demonstrate that the deletion of mouse Apoa-I exacerbates memory deficits in APP/PS1ΔE9 mice. Further characterization of APP/PS1ΔE9/apoA-I^KO mice showed that apoA-I deficiency did not affect amyloid precursor protein processing, soluble Aβ oligomer levels, Aβ plaque load, or levels of insoluble Aβ in brain parenchyma. To examine the effect of Apoa-I deletion on cerebral amyloid angiopathy, we measured insoluble Aβ isolated from cerebral blood vessels. Our data show that in APP/PS1ΔE9/apoA-I^KO mice, insoluble Aβ₄₀ is increased more than 10-fold, and Aβ₄₂ is increased 1.5-fold. The increased levels of deposited amyloid in the vessels of cortices and hippocampi of APP/PS1ΔE9/apoA-I^KO mice, measured by X-34 staining, confirmed the results. Finally, we demonstrate that lipidated and non-lipidated apoA-I significantly decreased Aβ toxicity against brain vascular smooth muscle cells. We conclude that lack of apoA-I aggravates the memory deficits in APP/PS1ΔE9 mice in parallel to significantly increased cerebral amyloid angiopathy.     

Evaluating Amyloid-β Oligomers in Cerebrospinal Fluid as a Biomarker for Alzheimer’s Disease

The current study evaluated amyloid-β oligomers (Aβo) in cerebrospinal fluid as a clinical biomarker for Alzheimer’s disease (AD). We developed a highly sensitive Aβo ELISA using the same N-terminal monoclonal antibody (82E1) for capture and detection. CSF samples from patients with AD, mild cognitive impairment (MCI), and healthy controls were examined. The assay was specific for oligomerized Aβ with a lower limit of quantification of 200 fg/ml, and the assay signal showed a tight correlation with synthetic Aβo levels. Three clinical materials of well characterized AD patients (n = 199) and cognitively healthy controls (n = 148) from different clinical centers were included, together with a clinical material of patients with MCI (n = 165). Aβo levels were elevated in the all three AD-control comparisons although with a large overlap and a separation from controls that was far from complete. Patients with MCI who later converted to AD had increased Aβo levels on a group level but several samples had undetectable levels. These results indicate that presence of high or measurable Aβo levels in CSF is clearly associated with AD, but the overlap is too large for the test to have any diagnostic potential on its own.     

Cerebrospinal Fluid PKR Level Predicts Cognitive Decline in Alzheimer’s Disease

The cerebrospinal fluid (CSF) levels of the proapoptotic kinase R (PKR) and its phosphorylated PKR (pPKR) are increased in Alzheimer’s disease (AD), but whether CSF PKR concentrations are associated with cognitive decline in AD patients remain unknown. In this study, 41 consecutive patients with AD and 11 patients with amnestic mild cognitive impairment (aMCI) from our Memory Clinic were included. A lumbar puncture was performed during the following month of the clinical diagnosis and Mini-Mental State Examination (MMSE) evaluations were repeated every 6 months during a mean follow-up of 2 years. In AD patients, linear mixed models adjusted for age and sex were used to assess the cross-sectional and longitudinal associations between MMSE scores and baseline CSF levels of Aβ peptide (Aβ 1-42), Tau, phosphorylated Tau (p-Tau 181), PKR and pPKR. The mean (SD) MMSE at baseline was 20.5 (6.1) and MMSE scores declined over the follow-up (-0.12 point/month, standard error [SE] = 0.03). A lower MMSE at baseline was associated with lower levels of CSF Aβ 1–42 and p-Tau 181/Tau ratio. pPKR level was associated with longitudinal MMSE changes over the follow-up, higher pPKR levels being related with an exacerbated cognitive deterioration. Other CSF biomarkers were not associated with MMSE changes over time. In aMCI patients, mean CSF biomarker levels were not different in patients who converted to AD from those who did not convert.These results suggest that at the time of AD diagnosis, a higher level of CSF pPKR can predict a faster rate of cognitive decline.     

Amyloid-Related Memory Decline in Preclinical Alzheimer’s Disease Is Dependent on APOE ε4 and Is Detectable over 18-Months

High levels of β-amyloid (Aβ) in the brain and carriage of the APOE ε4 allele have each been linked to cognitive impairment in cognitively normal (CN) older adults. However, the relationship between these two biomarkers and cognitive decline is unclear. The aim of this study was to investigate the relationship between cerebral Aβ level, APOE ε4 carrier status, and cognitive decline over 18 months, in 317 cognitively healthy (CN) older adults (47.6% males, 52.4% females) aged between 60 and 89 years (Mean = 69.9, SD = 6.8). Cognition was assessed using the Cogstate Brief Battery (CBB) and the California Verbal Learning Test, Second Edition (CVLT-II). Planned comparisons indicated that CN older adults with high Aβ who were also APOE ε4 carriers demonstrated the most pronounced decline in learning and working memory. In CN older adults who were APOE ε4 non-carriers, high Aβ was unrelated to cognitive decline in learning and working memory. Carriage of APOE ε4 in CN older adults with low Aβ was associated with a significantly increased rate of decline in learning and unexpectedly, improved cognitive performance on measures of verbal episodic memory over 18 months. These results suggest that Aβ and APOE ε4 interact to increase the rate of cognitive decline in CN older adults and provide further support for the use of Aβ and APOE ε4 as biomarkers of early Alzheimer’s disease.     

Molecular insights into the primary nucleation of polymorphic amyloid β dimers in DOPC lipid bilayer membrane

Alzheimer''s disease (AD) pathology is characterized by loss of memory cognitive and behavioral deterioration. One of the hallmarks of AD is amyloid β (Aβ) plaques in the brain that consists of Aβ oligomers and fibrils. It is accepted that oligomers, particularly dimers, are toxic species that are produced extracellularly and intracellularly in membranes. It is believed that the disruption of membranes by polymorphic Aβ oligomers is the key for the pathology of AD. This is a first study that investigate the effect of polymorphic “α‐helix/random coil” and “fibril‐like” Aβ dimers on 1,2‐dioleoyl‐sn‐glycero‐3‐phosphocholine (DOPC) membrane. It has been found that the DOPC membrane promotes Aβ_1–42 “fibril‐like” dimers and impedes Aβ_1–42 “α‐helix/random coil” dimers. The N‐termini domains within Aβ_1–42 dimers play a role in Aβ aggregation in membrane milieus. In addition, the aromatic π–π interactions (involving residues F19 and F20 in Aβ_1–42) are the driving forces for the hydrophobic interactions that initiate the primary nucleation of polymorphic Aβ_1–42 dimers within DOPC membrane. Finally, the DOPC bilayer membrane thickness is locally decreased, and it is disrupted by an embedded distinct Aβ_1–42 dimer, due to relatively large contacts between Aβ_1–42 monomers and the DOPC membrane. This study reveals insights into the molecular mechanisms by which polymorphic early‐stage Aβ_1–42 dimers have distinct impacts on DOPC membrane.     

The Pathogenic Aβ43 Is Enriched in Familial and Sporadic Alzheimer Disease

The amyloid-cascade hypothesis posits that the role of amyloid β-peptide (Aβ) in Alzheimer disease (AD) involves polymerization into structures that eventually are deposited as amyloid plaques. During this process, neurotoxic oligomers are formed that induce synaptic loss and neuronal death. Several different isoforms of Aβ are produced, of which the 40 and 42 residue variants (Aβ40 and Aβ42) are the most common. Aβ42 has a strong tendency to form neurotoxic aggregates and is involved in AD pathogenesis. Longer Aβ isoforms, like the less studied Aβ43, are gaining attention for their higher propensity to aggregate into neurotoxic oligomers. To further investigate Aβ43 in AD, we conducted a quantitative study on Aβ43 levels in human brain. We homogenized human brain tissue and prepared fractions of various solubility; tris buffered saline (TBS), sodium dodecyl sulfate (SDS) and formic acid (FA). Levels of Aβ43, as well as Aβ40 and Aβ42, were quantified using ELISA. We compared quantitative data showing Aβ levels in occipital and frontal cortex from sporadic (SAD) and familial (FAD) AD cases, as well as non-demented (ND) controls. Results showed Aβ43 present in each fraction from the SAD and FAD cases, while its level was lower than the detection limit in the majority of the ND-cases. Aβ42 and Aβ43 were enriched in the less soluble fractions (SDS and FA) of SAD and FAD cases in both occipital and frontal cortex. Thus, although the total levels of Aβ43 in human brain are low compared to Aβ40 and Aβ42, we suggest that Aβ43 could initiate the formation of oligomers and amyloid plaques and thereby be crucial to AD pathogenesis.     

Prevalence of Alzheimer’s Pathologic Endophenotypes in Asymptomatic and Mildly Impaired First-Degree Relatives

Objective

A positive family history (FH) is a risk factor for late-onset Alzheimer’s disease (AD). Our aim was to examine the effects of FH on pathological and neuronal loss biomarkers across the cognitive spectrum.

Design

Cross-sectional analyses of data from a national biomarker study.

Setting

The Alzheimer’s Disease Neuroimaging Initiative national study.

Patients

257 subjects (ages 55–89), divided into cognitively normal (CN), mild cognitive impairment (MCI), and AD groups, with CSF and FH data.

Outcome Measures

Cerebrospinal fluid (CSF) Aβ42, tau, and tau/Aβ42 ratio, MRI-measured hippocampal volumes.

Statistics

Univariate and multivariate analyses.

Results

In MCI, CSF Aβ42 was lower (p = .005), t-tau was higher (p = 0.02) and t-tau/Aβ42 ratio was higher (p = 0.002) in FH+ than FH− subjects. A significant residual effect of FH on pathologic markers in MCI remained after adjusting for ApoE4 (p<0.05). Among CN, 47% of FH+ exhibited “pathologic signature of AD” (CSF t-tau/Aβ42 ratio >0.39) versus 21% of FH− controls (p = 0.03). The FH effect was not significant in AD subjects. Hippocampal and intracranial volumes did not differ between FH+ and FH− subjects in any group.

Conclusions

A positive family history of late-onset AD is associated with a higher prevalence of an abnormal cerebral beta-amyloid and tau protein phenotype in MCI. The unexplained genetic heritability in family history is about the half the size of the ApoE4 effect. Longitudinal studies are warranted to more definitively examine this issue.     

Brain Intraventricular Injection of Amyloid-β in Zebrafish Embryo Impairs Cognition and Increases Tau Phosphorylation,Effects Reversed by Lithium

Alzheimer’s disease (AD) is a devastating neurodegenerative disorder with no effective treatment and commonly diagnosed only on late stages. Amyloid-β (Aβ) accumulation and exacerbated tau phosphorylation are molecular hallmarks of AD implicated in cognitive deficits and synaptic and neuronal loss. The Aβ and tau connection is beginning to be elucidated and attributed to interaction with different components of common signaling pathways. Recent evidences suggest that non-fibrillary Aβ forms bind to membrane receptors and modulate GSK-3β activity, which in turn phosphorylates the microtubule-associated tau protein leading to axonal disruption and toxic accumulation. Available AD animal models, ranging from rodent to invertebrates, significantly contributed to our current knowledge, but complementary platforms for mechanistic and candidate drug screenings remain critical for the identification of early stage biomarkers and potential disease-modifying therapies. Here we show that Aβ1–42 injection in the hindbrain ventricle of 24 hpf zebrafish embryos results in specific cognitive deficits and increased tau phosphorylation in GSK-3β target residues at 5dpf larvae. These effects are reversed by lithium incubation and not accompanied by apoptotic markers. We believe this may represent a straightforward platform useful to identification of cellular and molecular mechanisms of early stage AD-like symptoms and the effects of neuroactive molecules in pharmacological screenings.     

Platelet biomarkers identifying mild cognitive impairment in type 2 diabetes patients

Type 2 diabetes mellitus (T2DM) is an independent risk factor of Alzheimer''s disease (AD). Therefore, identifying periphery biomarkers correlated with mild cognitive impairment (MCI) is of importance for early diagnosis of AD. Here, we performed platelet proteomics in T2DM patients with MCI (T2DM‐MCI) and without MCI (T2DM‐nMCI). Pearson analysis of the omics data with MMSE (mini‐mental state examination), Aβ1‐42/Aβ1‐40 (β‐amyloid), and rGSK‐3β(T/S9) (total to Serine‐9‐phosphorylated glycogen synthase kinase‐3β) revealed that mitophagy/autophagy‐, insulin signaling‐, and glycolysis/gluconeogenesis pathways‐related proteins were most significantly involved. Among them, only the increase of optineurin, an autophagy‐related protein, was simultaneously correlated with the reduced MMSE score, and the increased Aβ1‐42/Aβ1‐40 and rGSK‐3β(T/S9), and the optineurin alone could discriminate T2DM‐MCI from T2DM‐nMCI. Combination of the elevated platelet optineurin and rGSK‐3β(T/S9) enhanced the MCI‐discriminating efficiency with AUC of 0.927, specificity of 86.7%, sensitivity of 85.3%, and accuracy of 0.859, which is promising for predicting cognitive decline in T2DM patients.     

A Presenilin-1 Mutation Identified in Familial Alzheimer Disease with Cotton Wool Plaques Causes a Nearly Complete Loss of γ-Secretase Activity

Mutations in presenilin-1 and presenilin-2 (PS1 and PS2) are the most common cause of familial Alzheimer disease. PS1 and PS2 are the presumptive catalytic components of the multisubunit γ-secretase complex, which proteolyzes a number of type I transmembrane proteins, including the amyloid precursor protein (APP) and Notch. APP processing by γ-secretase produces β-amyloid peptides (Aβ40 and Aβ42) that accumulate in the Alzheimer disease brain. Here we identify a pathogenic L435F mutation in PS1 in two affected siblings with early-onset familial Alzheimer disease characterized by deposition of cerebral cotton wool plaques. The L435F mutation resides in a conserved C-terminal PAL sequence implicated in active site conformation and catalytic activity. The impact of PS1 mutations in and around the PAL motif on γ-secretase activity was assessed by expression of mutant PS1 in mouse embryo fibroblasts lacking endogenous PS1 and PS2. Surprisingly, the L435F mutation caused a nearly complete loss of γ-secretase activity, including >90% reductions in the generation of Aβ40, Aβ42, and the APP and Notch intracellular domains. Two nonpathogenic PS1 mutations, P433L and L435R, caused essentially complete loss of γ-secretase activity, whereas two previously identified pathogenic PS1 mutations, P436Q and P436S, caused partial loss of function with substantial reductions in production of Aβ40, Aβ42, and the APP and Notch intracellular domains. These results argue against overproduction of Aβ42 as an essential property of presenilin proteins bearing pathogenic mutations. Rather, our findings provide support for the hypothesis that pathogenic mutations cause a general loss of presenilin function.     

Vitamin D2-Enriched Button Mushroom (Agaricus bisporus) Improves Memory in Both Wild Type and APPswe/PS1dE9 Transgenic Mice

Vitamin D deficiency is widespread, affecting over 30% of adult Australians, and increasing up to 80% for at-risk groups including the elderly (age>65). The role for Vitamin D in development of the central nervous system is supported by the association between Vitamin D deficiency and incidence of neurological and psychiatric disorders including Alzheimer’s disease (AD). A reported positive relationship between Vitamin D status and cognitive performance suggests that restoring Vitamin D status might provide a cognitive benefit to those with Vitamin D deficiency. Mushrooms are a rich source of ergosterol, which can be converted to Vitamin D₂ by treatment with UV light, presenting a new and convenient dietary source of Vitamin D₂. We hypothesised that Vitamin D₂-enriched mushrooms (VDM) could prevent the cognitive and pathological abnormalities associated with dementia. Two month old wild type (B6C3) and AD transgenic (APP_Swe/PS1dE9) mice were fed a diet either deficient in Vitamin D₂ or a diet which was supplemented with VDM, containing 1±0.2 µg/kg (∼54 IU/kg) vitamin D₂, for 7 months. Effects of the dietary intervention on memory were assessed pre- and post-feeding. Brain sections were evaluated for amyloid β (Aβ) plaque loads and inflammation biomarkers using immuno-histochemical methods. Plasma vitamin D metabolites, Aβ40, Aβ42, calcium, protein and cholesterol were measured using biochemical assays. Compared with mice on the control diet, VDM-fed wild type and AD transgenic mice displayed improved learning and memory, had significantly reduced amyloid plaque load and glial fibrillary acidic protein, and elevated interleukin-10 in the brain. The results suggest that VDM might provide a dietary source of Vitamin D₂ and other bioactives for preventing memory-impairment in dementia. This study supports the need for a randomised clinical trial to determine whether or not VDM consumption can benefit cognitive performance in the wider population.     

Increased CSF Levels of Phosphorylated Neurofilament Heavy Protein following Bout in Amateur Boxers

Introduction

Diagnosis of mild TBI is hampered by the lack of imaging or biochemical measurements for identifying or quantifying mild TBI in a clinical setting. We have previously shown increased biomarker levels of protein reflecting axonal (neurofilament light protein and tau) and glial (GFAP and S-100B) damage in cerebrospinal fluid (CSF) after a boxing bout. The aims of this study were to find other biomarkers of mild TBI, which may help clinicians diagnose and monitor mild TBI, and to calculate the role of APOE ε4 allele genotype which has been associated with poor outcome after TBI.

Materials and Methods

Thirty amateur boxers with a minimum of 45 bouts and 25 non-boxing matched controls were included in a prospective cohort study. CSF and blood were collected at one occasion between 1 and 6 days after a bout, and after a rest period for at least 14 days (follow up). The controls were tested once. CSF levels of neurofilament heavy (pNFH), amyloid precursor proteins (sAPPα and sAPPβ), ApoE and ApoA1 were analyzed. In blood, plasma levels of Aβ42 and ApoE genotype were analyzed.

Results

CSF levels of pNFH were significantly increased between 1 and 6 days after boxing as compared with controls (p<0.001). The concentrations decreased at follow up but were still significantly increased compared to controls (p = 0.018). CSF pNFH concentrations correlated with NFL (r =  0.57 after bout and 0.64 at follow up, p<0.001). No significant change was found in the other biomarkers, as compared to controls. Boxers carrying the APOE ε4 allele had similar biomarker concentrations as non-carriers.

Conclusions

Subconcussive repetitive trauma in amateur boxing causes a mild TBI that may be diagnosed by CSF analysis of pNFH, even without unconsciousness or concussion symptoms. Possession of the APOE ε4 allele was not found to influence biomarker levels after acute TBI.     

Enhanced Aβ1–40 Production in Endothelial Cells Stimulated with Fibrillar Aβ1–42

Amyloid accumulation in the brain of Alzheimer’s patients results from altered processing of the 39- to 43-amino acid amyloid β protein (Aβ). The mechanisms for the elevated amyloid (Aβ_1–42) are considered to be over-expression of the amyloid precursor protein (APP), enhanced cleavage of APP to Aβ, and decreased clearance of Aβ from the central nervous system (CNS). We report herein studies of Aβ stimulated effects on endothelial cells. We observe an interesting and as yet unprecedented feedback effect involving Aβ_1–42 fibril-induced synthesis of APP by Western blot analysis in the endothelial cell line Hep-1. We further observe an increase in the expression of Aβ_1–40 by flow cytometry and fluorescence microscopy. This phenomenon is reproducible for cultures grown both in the presence and absence of serum. In the former case, flow cytometry reveals that Aβ_1–40 accumulation is less pronounced than under serum-free conditions. Immunofluorescence staining further corroborates these observations. Cellular responses to fibrillar Aβ_1–42 treatment involving eNOS upregulation and increased autophagy are also reported.     

Feasibility of β-Sheet Breaker Peptide-H102 Treatment for Alzheimer's Disease Based on β-Amyloid Hypothesis

β-amyloid hypothesis is the predominant hypothesis in the study of pathogenesis of Alzheimer''s disease. This hypothesis claims that aggregation and neurotoxic effects of amyloid β (Aβ) is the common pathway in a variety of etiological factors for Alzheimer''s disease. Aβ peptide derives from amyloid precursor protein (APP). β-sheet breaker peptides can directly prevent and reverse protein misfolding and aggregation in conformational disorders. Based on the stereochemical structure of Aβ1-42 and aggregation character, we had designed a series of β-sheet breaker peptides in our previous work and screened out a 10-residue peptide β-sheet breaker peptide, H102. We evaluated the effects of H102 on expression of P-tau, several associated proteins, inflammatory factors and apoptosis factors, and examined the cognitive ability of APP transgenic mice by behavioral test. This study aims to validate the β-amyloid hypothesis and provide an experimental evidence for the feasibility of H102 treatment for Alzheimer''s disease.