Aβ Peptides In Cerebrospinal Fluid Research Articles

Fiber-in-tube SPME-CapLC-MS/MS method to determine Aβ peptides in cerebrospinal fluid obtained from Alzheimer's patients

Mass spectrometry is characterized by its high sensitivity, ability to measure very low analyte concentrations, specificity to distinguish between closely related compounds, availability to generate high-throughput methods for screening, and high multiplexing capacity. This technique has been used as a platform to analyze fluid biomarkers for Alzheimer's disease. However, more effective sample preparation procedures, preferably antibody-independent, and more automated mass spectrometry platforms with improved sensitivity, chromatographic separation, and high throughput are needed for this purpose. This short communication discusses the development of a fiber-in-tube SPME-CapLC-MS/MS method to determine Aβ peptides in cerebrospinal fluid obtained from Alzheimer's disease patients. To obtain the fiber-in-tube SPME capillary, we longitudinally packed 22 nitinol fibers coated with a zwitterionic polymeric ionic liquid into the same length of the PEEK tube. In addition, this communication compares this fiber-in-tube SPME method with the conventional HPLC scale (HPLC-MS/MS) and when directly coupled to CapESI-MS/MS without chromatographic separation, and, as a case study, discusses the benefits and challenges inherent in miniaturizing the flow scale of the sample preparation technique (fiber-in-tube SPME) to the CapLC-MS/MS system. Fiber-in-tube SPME-CapLC-MS/MS provided LLOQ ranging from 0.09 to 0.10 ng mL−1, accuracy ranging from 91 to 117 % (recovery), and reproducibility of less than 18 % (RSD). Analysis of the cerebrospinal fluid samples obtained from Alzheimer's disease patients evidenced that the method is robust. At the capillary scale (10 µL min−1), this innovative method presented higher analytical sensitivity than the conventional HPLC-MS/MS scale. Although fiber-in-tube SPME directly coupled to CapESI-MS/MS offers advantages in terms of high throughput, the sample was dispersed and non-quantitatively desorbed from the capillary at low flow rate. These results highlighted that chromatographic separation is important to decrease the matrix effect and to achieve higher detectability, which is indispensable for bioanalysis.

Profiling amyloid-β peptides as biomarkers for cerebral amyloid angiopathy.

Brain amyloid-β (Aβ) deposits are key pathological hallmarks of both cerebral amyloid angiopathy (CAA) and Alzheimer's disease (AD). Microvascular deposits in CAA mainly consist of the Aβ40 peptide, whereas Aβ42 is the predominant variant in parenchymal plaques in AD. The relevance in pathogenesis and diagnostic accuracy of various other Aβ isoforms in CAA remain understudied. We aimed to investigate the biomarker potential of various Aβ isoforms in cerebrospinal fluid (CSF) to differentiate CAA from AD pathology. We included 25 patients with probable CAA, 50 subjects with a CSF profile indicative of AD pathology (AD-like), and 23 age- and sex-matched controls. CSF levels of Aβ1-34 , Aβ1-37 , Aβ1-38 , Aβ1-39 , Aβ1-40 , and Aβ1-42 were quantified by liquid chromatography mass spectrometry. Lower CSF levels of all six Aβ peptides were observed in CAA patients compared with controls (p = 0.0005-0.03). Except for Aβ1-42 (p = 1.0), all peptides were decreased in CAA compared with AD-like subjects (p = 0.007-0.03). Besides Aβ1-42 , none of the Aβ peptides were decreased in AD-like subjects compared with controls. All Aβ peptides combined differentiated CAA from AD-like subjects better (area under the curve [AUC] 0.84) than individual peptide levels (AUC 0.51-0.75). Without Aβ1-42 in the model (since decreased Aβ1-42 served as AD-like selection criterion), the AUC was 0.78 for distinguishing CAA from AD-like subjects. CAA patients and AD-like subjects showed distinct disease-specific CSF Aβ profiles. Peptides shorter than Aβ1-42 were decreased in CAA patients, but not AD-like subjects, which could suggest different pathological mechanisms between vascular and parenchymal Aβ accumulation. This study supports the potential use of this panel of CSF Aβ peptides to indicate presence of CAA pathology with high accuracy.

Recent Electrokinetic and Microfluidic Strategies for Detection of Amyloid Beta Peptide Biomarkers: Towards Molecular Diagnosis of Alzheimer's Disease.

Among all neurodegenerative diseases, Alzheimer's Disease (AD) is the most prevalent worldwide, with a huge burden to the society and no efficient AD treatment so far. Continued efforts have been being made towards early and powerful diagnosis of AD, in the hope for a successful set of clinical trials and subsequently AD curative treatment. Towards this aim, detection and quantification of amyloid beta (Aβ) peptides in cerebrospinal fluid (CSF) and other biofluids, which are established and validated biomarkers for AD, have drawn attention of the scientific community and industry over almost two decades. In this work, an overview on our major contributions over 15 years to develop different electrokinetic and microfluidic strategies for Aβ peptides detection and quantification is reported. Accordingly, discussions and viewpoints on instrumental and methodological developments for microscale electrophoresis, microfluidic designs and immuno-enrichment / assays on magnetic beads in microchannels for tracing Aβ peptides in CSF are given in this review.

Amyloid-\u03b2 peptides in cerebrospinal fluid of patients with dementia with Lewy bodies

BackgroundOne of the major challenges in diagnosing dementia with Lewy bodies (DLB) is the common co-morbid presence of amyloid pathology. To understand the putative role of altered amyloid-β (Aβ) metabolism in dementia with DLB, we analyzed levels of different cerebrospinal fluid (CSF) Aβ peptides (Aβ38, Aβ40, Aβ42) in DLB, Alzheimer’s disease (AD), and cognitively normal controls.MethodsCSF from patients with DLB (n = 72; age 68 ± 6 years; 10%F; Mini-mental State examination (MMSE) 23 ± 4), AD (n = 38; age 68 ± 6 years; 8%F; MMSE 22 ± 5), and cognitively normal controls (n = 38; age 67 ± 7 years; 13%F; MMSE 29 ± 2) was analyzed using the Meso Scale Discovery assay for human Aβ peptides. We performed general linear models to compare CSF Aβ peptide levels between groups. Associations between CSF Aβ peptides and MMSE score at baseline and longitudinal changes over time were assessed with linear mixed models.ResultsFor all three CSF Aβ peptides and compared to controls (Aβ38 2676 ± 703 pg/ml, Aβ40 6243 ± 1500 pg/ml, and Aβ42 692 ± 205 pg/ml), we observed lower levels in DLB (Aβ38 2247 ± 638, Aβ40 5432 ± 1340, and Aβ42 441 ± 185, p < 0.05), whereas AD patients showed only lower Aβ42 levels (304 ± 71, p < 0.001). The observed differences in Aβ38 and Aβ40 were independent of co-morbid AD pathology (CSF tau/Aβ42 > 0.52) and APOE genotype. Finally, lower Aβ peptide levels were associated with lower MMSE score (β = 1.02–1.11, p < 0.05).ConclusionWe demonstrated different profiles of CSF Aβ reduction in DLB and AD. In particular, while AD is characterized by an isolated drop in Aβ42, DLB comes with reductions in Aβ38, Aβ40, and Aβ42. This suggests that amyloid metabolism is affected in DLB, even in the absence of co-morbid AD pathology.

PHARMACOKINETIC/PHARMACODYNAMIC (PK/PD) EFFECTS OF PF-06648671, A NOVEL GAMMA SECRETASE MODULATOR (GSM), FOLLOWING SINGLE AND MULTIPLE DOSE ADMINISTRATION IN HEALTHY VOLUNTEERS

Unlike gamma secretase inhibitors, GSMs shift the cleavage of amyloid-β (Aβ) peptides in favor of shorter forms without inhibiting overall Aβ precursor protein processing, thus preferentially lower the production of longer, amyloidogenic forms. This study aimed to characterize PF-06648671 plasma exposure and cerebrospinal fluid (CSF) Aβ (Aβx-42, Aβx-40, Aβx-38 and Aβx-37) response relationship in healthy subjects. Clinical data were obtained from three Phase 1 studies: first-in-human single ascending dose (PK), single dose CSF biomarker (PK/PD; 150, 300 mg, and placebo), and multiple dose (PK/PD; 40, 100, 200, 360 mg and placebo) studies. Drifts in baseline (a rising tendency) were observed in placebo subjects after serial CSF sampling over 36 hours, which was taken into account in the model. Indirect response model was applied in a way that PF-06648671 plasma concentrations change the production rate of CSF Aβ peptides (decreased Aβ42 and Aβ40; increased Aβ37 and Aβ38). Population PK/PD modeling was performed using NONMEM 7.3. The inhibitory effect on CSF Aβ42 was greater than on Aβ40, whereas the stimulatory effect on Aβ37 was greater than on Aβ38. Preliminary results suggested CSF Aβ42 reductions after 100 and 350 mg q.d. administrations at steady state are approximately 50% and 66% on average, and about 200% and 400% increase in Aβ37, respectively. Indirect response model was developed to characterize exposure-dependent and differential effects of PF-06648671 on CSF Aβ's in healthy subjects. PK/PD relationship of PF-06648671 in human demonstrated a good translatability from preclinical model. The clinical PK/PD model has been used to predict the drug effect on individual Aβ peptides as well as the ratio of these Aβ peptides in CSF across a range of PF-06648671 doses. These results will guide dose selection in future Phase 2/3 trials.

Cerebrospinal Fluid Levels of Amyloid Beta 1-43 Mirror 1-42 in Relation to Imaging Biomarkers of Alzheimer’s Disease

Introduction: Amyloid beta 1-43 (Aβ43), with its additional C-terminal threonine residue, is hypothesized to play a role in early Alzheimer’s disease pathology possibly different from that of amyloid beta 1-42 (Aβ42). Cerebrospinal fluid (CSF) Aβ43 has been suggested as a potential novel biomarker for predicting conversion from mild cognitive impairment (MCI) to dementia in Alzheimer’s disease. However, the relationship between CSF Aβ43 and established imaging biomarkers of Alzheimer’s disease has never been assessed.Materials and Methods: In this observational study, CSF Aβ43 was measured with ELISA in 89 subjects; 34 with subjective cognitive decline (SCD), 51 with MCI, and four with resolution of previous cognitive complaints. All subjects underwent structural MRI; 40 subjects on a 3T and 50 on a 1.5T scanner. Forty subjects, including 24 with SCD and 12 with MCI, underwent 18F-Flutemetamol PET. Seventy-eight subjects were assessed with 18F-fluorodeoxyglucose PET (21 SCD/7 MCI and 11 SCD/39 MCI on two different scanners). Ten subjects with SCD and 39 with MCI also underwent diffusion tensor imaging.Results: Cerebrospinal fluid Aβ43 was both alone and together with p-tau a significant predictor of the distinction between SCD and MCI. There was a marked difference in CSF Aβ43 between subjects with 18F-Flutemetamol PET scans visually interpreted as negative (37 pg/ml, n = 27) and positive (15 pg/ml, n = 9), p < 0.001. Both CSF Aβ43 and Aβ42 were negatively correlated with standardized uptake value ratios for all analyzed regions; CSF Aβ43 average rho -0.73, Aβ42 -0.74. Both CSF Aβ peptides correlated significantly with hippocampal volume, inferior parietal and frontal cortical thickness and axial diffusivity in the corticospinal tract. There was a trend toward CSF Aβ42 being better correlated with cortical glucose metabolism. None of the studied correlations between CSF Aβ43/42 and imaging biomarkers were significantly different for the two Aβ peptides when controlling for multiple testing.Conclusion: Cerebrospinal fluid Aβ43 appears to be strongly correlated with cerebral amyloid deposits in the same way as Aβ42, even in non-demented patients with only subjective cognitive complaints. Regarding imaging biomarkers, there is no evidence from the present study that CSF Aβ43 performs better than the classical CSF biomarker Aβ42 for distinguishing SCD and MCI.

Isobaric Quantification of Cerebrospinal Fluid Amyloid-β Peptides in Alzheimer's Disease: C-Terminal Truncation Relates to Early Measures of Neurodegeneration.

The amyloid beta (Aβ) peptide is the main constituent of the plaques characteristic of Alzheimer's disease (AD). Measurement of Aβ1-42 in cerebrospinal fluid (CSF) is a valuable marker in AD research, where low levels indicate AD. Although the use of immunoassays measuring Aβ1-38 and Aβ1-40 in addition to Aβ1-42 has increased, quantitative assays of other Aβ peptides remain rarely explored. We recently discovered novel Aβ peptides in CSF using antibodies recognizing the Aβ mid-domain region. Here we have developed a method using both Aβ N-terminal and mid-domain antibodies for immunoprecipitation in combination with isobaric labeling and liquid chromatography-tandem mass spectrometry (LC-MS/MS) for relative quantification of endogenous Aβ peptides in CSF. The developed method was used in a pilot study to produce Aβ peptide profiles from 38 CSF samples. Statistical comparison between CSF samples from 19 AD patients and 19 cognitively healthy controls revealed no significant differences at group level. A significant correlation was found between several larger C-terminally truncated Aβ peptides and protein biomarkers for neuronal damage, particularly prominent in the control group. Comparison of the isobaric quantification with immunoassays measuring Aβ1-38 or Aβ1-40 showed good correlation (r(2) = 0.84 and 0.85, respectively) between the two analysis methods. The developed method could be used to assess disease-modifying therapies directed at Aβ production or degradation.

Cerebrospinal fluid Aβ40 is similarly reduced in patients with Frontotemporal Lobar Degeneration and Alzheimer's Disease

Cerebrospinal fluid (CSF) biomarkers have been increasingly studied for dementia diagnosis, however the accuracy to distinguish between different forms of dementia is still unsatisfactory. In this study, the added value of another CSF Aβ-peptide (Aβ40), along with the core CSF markers t-Tau, p-Tau, and Aβ42, in the discrimination between two large dementia groups of Frontotemporal Lobar Degeneration (FTLD; n=107), Alzheimer's Disease (AD; n=107) and non-demented subjects (n=33) was evaluated.In FTLD, t-Tau and p-Tau were significantly increased in relation to controls, but lower than in AD, while Aβ42 was similar in FTLD and controls, but higher than in AD. Equally reduced Aβ40 levels were seen in both dementia groups, and therefore the combination of Aβ40 with core CSF biomarkers optimally discriminated FTLD and AD patients from controls. Aβ42 and t-Tau were selected as the best biomarker subset to differentiate FTLD from AD, with no added value of Aβ40 to the model. Diagnostic accuracy between FTLD and AD was still sub-optimal, with a significant percentage (23%) of FTLD patients, in particularly women, carrying an ApoE-ε4 allele, showing a CSF-AD biomarkers profile.Although CSF Aβ40 does not appear to have an additional value in the distinction between FTLD and AD, it increases the discrimination between subjects with dementia from controls. A CSF-AD biomarker profile can be seen in patients with a clinical phenotype of FTLD, reinforcing the need for autopsy confirmation.

Soluble BACE-1 Activity and sAβPPβ Concentrations in Alzheimer's Disease and Age-Matched Healthy Control Cerebrospinal Fluid from the Alzheimer's Disease Neuroimaging Initiative-1 Baseline Cohort.

β-site amyloid precursor protein-cleaving enzyme 1 (BACE1) plays an important role in the development of Alzheimer's disease (AD), freeing the amyloid-β (Aβ) N-terminus from the amyloid-β protein precursor (AβPP), the first step in Aβ formation. Increased BACE1 activity in AD brain or cerebrospinal fluid (CSF) has been reported. Other studies, however, found either no change or a decrease with AD diagnosis in either BACE1 activity or sAβPPβ, the N-terminal secreted product of BACE1 (sBACE1) activity on AβPP. Here, sBACE1 enzymatic activity and secreted AβPPβ (sAβPPβ) were measured in Alzheimer's Disease Neuroimaging Initiative-1 (ADNI-1) baseline CSF samples and no statistically significant changes were found in either measure comparing healthy control, mild cognitively impaired, or AD individual samples. While CSF sBACE1 activity and sAβPPβ demonstrated a moderate yet significant degree of correlation with each other, there was no correlation of either analyte to CSF Aβ peptide ending at residue 42. Surprisingly, a stronger correlation was demonstrated between CSF sBACE1 activity and tau, which was comparable to that between CSF Aβ₄₂ and tau. Unlike for these latter two analytes, receiver-operator characteristic curves demonstrate that neither CSF sBACE1 activity nor sAβPPβ concentrations can be used to differentiate between healthy elderly and AD individuals.

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.

Characterization of the Effect of a Novel &amp;#947;-secretase Modulator on A&amp;#946;: A Clinically Translatable Model

Alzheimer's disease (AD) is a slowly progressing disease and the evaluation of clinical effects of candidate drugs requires large clinical cohorts as well as long treatment trials. There is a great need for central biomarkers and translatable pre-clinical models to provide early indication of treatment effects. We set out to evaluate the guinea pig as a clinically translatable model looking at Aβ peptides. Our data demonstrate homology between β-amyloid (Aβ) peptide pattern in cerebrospinal fluid (CSF) from human and guinea pig. To further evaluate the model a novel γ-secretase modulator was used. Dose and time response studies confirm the modulatory properties with a statistically significant decrease in relative levels of Aβ1-40, Aβ1-42 and increase in Aβ1-37 already one hour after administration. We suggest that the guinea pig is a compelling pre-clinical model for evaluating and translating central effects on Aβ peptides in CSF after treatment. Further quantitative data are needed to confirm our data together with data from clinical trials in order to back translate and validate our findings.

Altered cerebrospinal fluid levels of amyloid β and amyloid precursor-like protein 1 peptides in Down's syndrome.

Down's syndrome (DS) patients develop early Alzheimer's disease pathology with abundant cortical amyloid plaques, likely due to overproduction of the amyloid precursor protein (APP), which subsequently leads to amyloid β (Aβ) aggregation. This is reflected in cerebrospinal fluid (CSF) levels of the 42-amino acid long Aβ peptide (Aβ1-42), which are increased in young DS patients and decreases with age. However, it is unclear whether DS also affects other aspects of Aβ metabolism, including production of shorter C- and N-terminal truncated Aβ peptides, and production of peptides from the amyloid precursor-like protein 1 (APLP1), which is related to APP, and cleaved by the same enzymatic processing machinery. APLP1-derived peptides may be surrogate markers for Aβ1-42 production in the brain. Here, we used hybrid immunoaffinity-mass spectrometry and enzyme-linked immunosorbent assays to monitor several Aβ and APLP1 peptides in CSF from DS patients (n = 12) and healthy controls (n = 20). CSF levels of Aβ1-42 and three endogenous peptides derived from APLP1 (APL1β25, APL1β27 and APL1β28) were decreased in DS compared with controls, while a specific Aβ peptide, Aβ1-28, was increased in a majority of the DS individuals. This study indicates that DS causes previously unknown specific alterations of APP and APLP1 metabolism.

Cerebrospinal fluid biomarkers of β-amyloid metabolism in multiple sclerosis

Background: Amyloid precursor protein (APP) and amyloid β (Aβ) peptides are intensely studied in neuroscience and their cerebrospinal fluid (CSF) measurements may be used to track the metabolic pathways of APP in vivo. Reduced CSF levels of Aβ and soluble APP (sAPP) fragments are reported in inflammatory diseases, including multiple sclerosis (MS); but in MS, the precise pathway of APP metabolism and whether it can be affected by disease-modifying treatments remains unclear. Objective: To characterize the CSF biomarkers of APP degradation in MS, including the effects of disease-modifying therapy. Methods: CSF samples from 87 MS patients (54 relapsing–remitting (RR) MS; 33 secondary progressive (SP) MS and 28 controls were analyzed for sAPP and Aβ peptides by immunoassays, plus a subset of samples was analyzed by immunoprecipitation and mass spectrometry (IP-MS). Patients treated with natalizumab or mitoxantrone were examined at baseline, and after 1–2 years of treatment. Results: CSF sAPP and Aβ peptide levels were reduced in MS patients; but they increased again towards normal, after natalizumab treatment. A multivariate model of IP-MS-measured Aβ species separated the SPMS patients from controls, with RRMS patients having intermediate levels. Conclusions: We confirmed and extended our previous observations of altered CSF sAPP and Aβ peptide levels in MS patients. We found that natalizumab therapy may be able to counteract the altered APP metabolism in MS. The CSF Aβ isoform distribution was found to be distinct in SPMS patients, as compared to the controls.

F2‐03‐04: Circadian patterns of beta‐amyloid in human cerebrospinal fluid and plasma

The amyloid hypothesis predicts that increased production or decreased clearance of amyloid-beta (Aβ) leads to amyloidosis, which ultimately culminates in Alzheimer's disease (AD). Changes in amyloid proteins, such as decreased cerebrospinal fluid (CSF) Aβ42 have been recognized as a biomarker for AD. However, the dynamics of Aβ concentrations over time are not well understood. Hourly CSF and plasma samples were obtained over a 36-hour study period in three groups of human volunteers: a dementia group with amyloid deposition, an age-matched control group, and a younger control group. We investigated dynamic patterns of Aβ peptides (Aβ40 and Aβ42 in CSF and plasma using bead-based Luminex XMAP assays), and Amyloid Precursor Protein (APP) (APP-α and APP-β in CSF using ELISA assays). We then analyzed the effects of aging and amyloidosis on circadian patterns and correlations between CSF and plasma Aβ peptides. Circadian patterns were observed in CSF and plasma Aβ, with decreased amplitudes with aging. Further, CSF APP also demonstrated a circadian pattern. No significant correlations were found between plasma and CSF Aβ levels on an hourly or individual basis. We also found that CSF APPα, APPβ, Aβ40, and Aβ42 are highly positively correlated in all participants without amyloidosis. However, in participants with amyloidosis, there is no correlation of Aβ42 to the other APP metabolites, suggesting that normal physiologic regulation of CSF Aβ42 is impaired in the presence of amyloidosis. This study suggests amyloid proteins in both central and peripheral compartments are dynamic, and regulated in a circadian pattern that is part of the normal dynamic physiologic control of Aβ concentrations. Regulatory mechanisms of these proteins may be altered with aging and amyloidosis. The lack of correlation of plasma and CSF Aβ levels suggest plasma Aβ concentrations can not be used for surrogates of central Aβ concentrations. These findings support an active circadian regulation of Aβ and may provide insight into the pathophysiological changes in AD.

Characterization of cerebrospinal fluid aminoterminally truncated and oxidized amyloid‐β peptides

Carboxyterminally elongated and aminoterminally truncated Aβ peptides as well as their pyroglutamate and oxidized derivates are major constituents of human amyloid plaques. The objective of the present study was to characterize aminoterminally truncated or oxidized Aβ38, Aβ40, and Aβ42 peptide species in immunoprecipitated human cerebrospinal fluid (CSF). We invented a novel sequential aminoterminally and carboxyterminally specific immunoprecipitation protocol and used the Aβ-SDS-PAGE/immunoblot for subsequent analysis of CSF Aβ peptide patterns. In the present study, we identified the aminoterminally truncated Aβ peptides 2-40 and 2-42 as well as oxidized forms of Aβ1-38 and Aβ1-42 in CSF. Our protocol allowed the quantification of a pattern of Aβ peptides 1-38(ox), 2-40, and 2-42 in addition to the well known panel of Aβ 1-37, 1-38, 1-39, 1-40, 1-40(ox), and 1-42 in a group of seven patients with peripheral polyneuropathy. In the present approach, we could broaden the range of quantifiable Aβ peptides described in previous studies (i.e., 1-37, 1-38, 1-39, 1-40, 1-40(ox), and 1-42) by Aβ 1-38(ox), 2-40, and 2-42. An exact analysis of CSF Aβ peptides regarding their carboxy- and aminoterminus as well as posttranslational modification seems promising with respect to diagnostic and pathogenic aspects.

Site-specific characterization of threonine, serine, and tyrosine glycosylations of amyloid precursor protein/amyloid β-peptides in human cerebrospinal fluid

The proteolytic processing of human amyloid precursor protein (APP) into shorter aggregating amyloid β (Aβ)-peptides, e.g., Aβ1-42, is considered a critical step in the pathogenesis of Alzheimer's disease (AD). Although APP is a well-known membrane glycoprotein carrying both N- and O-glycans, nothing is known about the occurrence of released APP/Aβ glycopeptides in cerebrospinal fluid (CSF). We used the 6E10 antibody and immunopurified Aβ peptides and glycopeptides from CSF samples and then liquid chromatography-tandem mass spectrometry for structural analysis using collision-induced dissociation and electron capture dissociation. In addition to 33 unglycosylated APP/Aβ peptides, we identified 37 APP/Aβ glycopeptides with sialylated core 1 like O-glycans attached to Thr(-39, -21, -20, and -13), in a series of APP/AβX-15 glycopeptides, where X was -63, -57, -52, and -45, in relation to Asp1 of the Aβ sequence. Unexpectedly, we also identified a series of 27 glycopeptides, the Aβ1-X series, where X was 20 (DAEFRHDSGYEVHHQKLVFF), 19, 18, 17, 16, and 15, which were all uniquely glycosylated on Tyr10. The Tyr10 linked O-glycans were (Neu5Ac)(1-2)Hex(Neu5Ac)HexNAc-O- structures with the disialylated terminals occasionally O-acetylated or lactonized, indicating a terminal Neu5Acα2,8Neu5Ac linkage. We could not detect any glycosylation of the Aβ1-38/40/42 isoforms. We observed an increase of up to 2.5 times of Tyr10 glycosylated Aβ peptides in CSF in six AD patients compared to seven non-AD patients. APP/Aβ sialylated O-glycans, including that of a Tyr residue, the first in a mammalian protein, may modulate APP processing, inhibiting the amyloidogenic pathway associated with AD.

A Window into the Heterogeneity of Human Cerebrospinal Fluid Aβ Peptides

The initiating event in Alzheimer's disease (AD) is an imbalance in the production and clearance of amyloid beta (Aβ) peptides leading to the formation of neurotoxic brain Aβ assemblies. Cerebrospinal Fluid (CSF), which is a continuum of the brain, is an obvious source of markers reflecting central neuropathologic features of brain diseases. In this review, we provide an overview and update on our current understanding of the pathobiology of human CSF Aβ peptides. Specifically, we focused our attention on the heterogeneity of the CSF Aβ world discussing (1) basic research studies and what has been translated to clinical practice, (2) monomers and other soluble circulating Aβ assemblies, and (3) communication modes for Aβ peptides and their microenvironment targets. Finally, we suggest that Aβ peptides as well as other key signals in the central nervous system (CNS), mainly involved in learning and hence plasticity, may have a double-edged sword action on neuron survival and function.

Microchip Electrophoresis Profiling of Aβ Peptides in the Cerebrospinal Fluid of Patients with Alzheimer’s Disease

The preferential aggregation of Aβ1-42 in amyloid plaques is one of the major neuropathological events in Alzheimer's disease. This is accompanied by a relative reduction of the concentration of Aβ1-42 in the cerebrospinal fluid (CSF) of patients developing the signs of Alzheimer's disease. Here, we describe a microchip gel electrophoresis method in polydimethylsiloxane (PDMS) chip that enables rapid profiling of major Aβ peptides in cerebrospinal fluid. To control the electroosmotic flow (EOF) in the PDMS channel and also to reduce the adsorption of the peptides to the surface of the channel, a new double coating using poly(dimethylacrylamide-co-allyl glycidyl ether) (PDMA-AGE) and methylcellulose-Tween-20 was developed. With this method, separation of five synthetic Aβ peptides (Aβ1-37, Aβ1-38, Aβ1-39, Aβ1-40, and Aβ1-42) was achieved, and relative abundance of Aβ1-42 to Aβ1-37 could be calculated in different standard mixtures. We applied our method for profiling of Aβ peptides in CSF samples from nonAlzheimer patients and patients with Alzheimer's disease. Aβ peptides in the CSF samples were captured and concentrated using a microfluidic system in which magnetic beads coated with anti-Aβ were self-organized into an affinity microcolumn under the a permanent magnetic field. Finally, we could detect two Aβ peptides (Aβ1-40 and Aβ1-42) in the CSF samples.

CSF amyloid-β and tau proteins, and cognitive performance, in early and untreated Parkinson's Disease: the Norwegian ParkWest study

BackgroundAlzheimer's disease (AD) pathology is found in a considerable portion of patients with Parkinson's disease (PD), particularly those with early dementia (PDD). Altered cerebrospinal fluid (CSF) levels of amyloid-β (Aβ)...

Distinct cerebrospinal fluid amyloid β peptide signatures in sporadic and PSEN1A431E-associated familial Alzheimer's disease

BackgroundAlzheimer's disease (AD) is associated with deposition of amyloid β (Aβ) in the brain, which is reflected by low concentration of the Aβ1-42 peptide in the cerebrospinal fluid (CSF). There are at least 15 additional Aβ peptides in human CSF and their relative abundance pattern is thought to reflect the production and degradation of Aβ. Here, we test the hypothesis that AD is characterized by a specific CSF Aβ isoform pattern that is distinct when comparing sporadic AD (SAD) and familial AD (FAD) due to different mechanisms underlying brain amyloid pathology in the two disease groups.ResultsWe measured Aβ isoform concentrations in CSF from 18 patients with SAD, 7 carriers of the FAD-associated presenilin 1 (PSEN1) A431E mutation, 17 healthy controls and 6 patients with depression using immunoprecipitation-mass spectrometry. Low CSF levels of Aβ1-42 and high levels of Aβ1-16 distinguished SAD patients and FAD mutation carriers from healthy controls and depressed patients. SAD and FAD were characterized by similar changes in Aβ1-42 and Aβ1-16, but FAD mutation carriers exhibited very low levels of Aβ1-37, Aβ1-38 and Aβ1-39.ConclusionSAD patients and PSEN1 A431E mutation carriers are characterized by aberrant CSF Aβ isoform patterns that hold clinically relevant diagnostic information. PSEN1 A431E mutation carriers exhibit low levels of Aβ1-37, Aβ1-38 and Aβ1-39; fragments that are normally produced by γ-secretase, suggesting that the PSEN1 A431E mutation modulates γ-secretase cleavage site preference in a disease-promoting manner.