A combined dataset of human cerebrospinal fluid proteins identified by multi-dimensional chromatography and tandem mass spectrometry

Human cerebrospinal fluid (CSF) is an important source for studying protein biomarkers of age-related neurodegenerative diseases. Before characterizing biomarkers unique to each disease, it is necessary to categorize CSF proteins systematically and extensively. However, the enormous complexity, great dynamic range of protein concentrations, and tremendous protein heterogeneity due to post-translational modification of CSF create significant challenges to the existing proteomics technologies for an in-depth, nonbiased profiling of the human CSF proteome. To circumvent these difficulties, in the last few years, we have utilized several different separation methodologies and mass spectrometric platforms that greatly enhanced the identification coverage and the depth of protein profiling of CSF to characterize CSF proteome. In total, 2594 proteins were identified in well-characterized pooled human CSF samples using stringent proteomics criteria. This report summarizes our efforts to comprehensively characterize the human CSF proteome to date.     

Expanding the cerebrospinal fluid endopeptidome

Biomarkers of neurodegenerative disorders are needed to assist in diagnosis, to monitor disease progression and therapeutic interventions, and to provide insight into disease mechanisms. One route to identify such biomarkers is by proteomic and peptidomic analysis of cerebrospinal fluid (CSF). In the current study, we performed an in‐depth analysis of the human CSF endopeptidome to establish an inventory that may serve as a basis for future targeted biomarker studies. High‐pH RP HPLC was employed for off‐line sample prefractionation followed by low‐pH nano‐LC‐MS analysis. Different software programs and scoring algorithms for peptide identification were employed and compared. A total of 18 031 endogenous peptides were identified at a FDR of 1%, increasing the number of known endogenous CSF peptides 10‐fold compared to previous studies. The peptides were derived from 2 053 proteins of which more than 60 have been linked to neurodegeneration. Notably, among the findings were six peptides derived from microtubule‐associated protein tau, three of which span the diagnostically interesting threonine‐181 (Tau‐F isoform). Also, 213 peptides from amyloid precursor protein were identified, 58 of which were partially or completely within the sequence of amyloid β 1–40/42, as well as 109 peptides from apolipoprotein E, spanning sequences that discriminate between the E2/E3/E4 isoforms of the protein.     

Understanding the importance of autophagy in human diseases using Drosophila

Autophagy is a lysosome-dependent intracellular degradation pathway that has been implicated in the pathogenesis of various human diseases, either positively or negatively impacting disease outcomes depending on the specific context. The majority of medical conditions including cancer, neurodegenerative diseases, infections and immune system disorders and inflammatory bowel disease could probably benefit from therapeutic modulation of the autophagy machinery. Drosophila represents an excellent model animal to study disease mechanisms thanks to its sophisticated genetic toolkit, and the conservation of human disease genes and autophagic processes. Here, we provide an overview of the various autophagy pathways observed both in flies and human cells(macroautophagy, microautophagy and chaperone-mediated autophagy), and discuss Drosophila models of the above-mentioned diseases where fly research has already helped to understand how defects in autophagy genes and pathways contribute to the relevant pathomechanisms.     

Associations between liver function and cerebrospinal fluid biomarkers of Alzheimer's disease pathology in non-demented adults: The CABLE study

Liver function has been suggested as a possible factor in the progression of Alzheimer's disease (AD) development. However, the association between liver function and cerebrospinal fluid (CSF) levels of AD biomarkers remains unclear. In this study, we analyzed the data from 1687 adults without dementia from the Chinese Alzheimer's Biomarker and LifestylE study to investigate differences in liver function between pathological and clinical AD groups, as defined by the 2018 National Institute on Aging-Alzheimer's Association Research Framework. We also examined the linear relationship between liver function, CSF AD biomarkers, and cognition using linear regression models. Furthermore, mediation analyses were applied to explore the potential mediation effects of AD pathological biomarkers on cognition. Our findings indicated that, with AD pathological and clinical progression, the concentrations of total protein (TP), globulin (GLO), and aspartate aminotransferase/alanine transaminase (ALT) increased, while albumin/globulin (A/G), adenosine deaminase, alpha-L-fucosidase, albumin, prealbumin, ALT, and glutamate dehydrogenase (GLDH) concentrations decreased. Furthermore, we also identified significant relationships between TP (β = −0.115, p_FDR < 0.001), GLO (β = −0.184, p_FDR < 0.001), and A/G (β = 0.182, p_FDR < 0.001) and CSF β-amyloid_1–42 (Aβ_1–42) (and its related CSF AD biomarkers). Moreover, after 10 000 bootstrapped iterations, we identified a potential mechanism by which TP and GLDH may affect cognition by mediating CSF AD biomarkers, with mediation effect sizes ranging from 3.91% to 16.44%. Overall, our results suggested that abnormal liver function might be involved in the clinical and pathological progression of AD. Amyloid and tau pathologies also might partially mediate the relationship between liver function and cognition. Future research is needed to fully understand the underlying mechanisms and causality to develop an approach to AD prevention and treatment approach.     

8-plex quantitation of changes in cerebrospinal fluid protein expression in subjects undergoing intravenous immunoglobulin treatment for Alzheimer's disease

An 8-plex version of an isobaric reagent for the quantitation of proteins using shotgun methods is presented. The 8-plex version of the reagent relies on amine-labeling chemistry of peptides similar to 4-plex reagents. MS/MS reporter ions at 113, 114, 115, 116, 117, 118, 119, and 121 m/z are used to quantify protein expression. This technology which was first applied to a test mixture consisting of eight proteins and resulted in accurate quantitation, has the potential to increase throughput of analysis for quantitative shotgun proteomics experiments when compared to 2- and 4-plex methods. The technology was subsequently applied to a longitudinal study of cerebrospinal fluid (CSF) proteins from subjects undergoing intravenous Ig treatment for Alzheimer's disease. Results from this study identify a number of protein expression changes that occur in CSF after 3 and 6 months of treatment compared to a baseline and compared to a drug washout period. A visualization tool was developed for this dataset and is presented. The tool can aid in the identification of key peptides and measurements. One conclusion aided by the visualization tool is that there are differences in considering peptide-based observations versus protein-based observations from quantitative shotgun proteomics studies.     

Rapid analysis of tryptically digested cerebrospinal fluid using capillary electrophoresis-electrospray ionization-Fourier transform ion cyclotron resonance-mass spectrometry

Mass spectrometry has in recent years been established as the method of choice for protein identification and characterization in proteomics. Capillary electrophoresis (CE) is a fast and efficient method for the separation of peptides and proteins. The on-line combination of CE with Fourier transform ion cyclotron resonance (FTICR) mass spectrometry (MS) has been shown to be a powerful tool in the analysis of complex mixtures of proteins. This paper presents the first results from a proteomic analysis of human cerebrospinal fluid proteins by tryptic digestion and CE-FTICR-MS, where 30 proteins could be identified on a 95% confidence level with mass measurement errors less than 5 ppm.     

Quantitative analysis of amyloid-beta peptides in cerebrospinal fluid using immunoprecipitation and MALDI-Tof mass spectrometry.

Immunoprecipitation (IP) combined with matrix-assisted laser desorption ionization (MALDI) time of flight (Tof) mass spectrometry has been used to develop quantitative assays for amyloid-beta (Abeta) peptides in cerebrospinal fluid (CSF). Inclusion of (15)N labelled standard peptides allows for absolute quantification of multiple Abeta isoforms in individual samples. Characterization of variability associated with all steps of the assay indicated that the IP step is the single largest contributor to overall variability. Optimization of the assay resulted in overall coefficient of variation 相似文献   

Protein interaction network for Alzheimer's disease using computational approach

Alzheimer''s disease (AD) is the most common form of dementia. It is the sixth leading cause of death in old age people. Despiterecent advances in the field of drug design, the medical treatment for the disease is purely symptomatic and hardly effective. Thusthere is a need to understand the molecular mechanism behind the disease in order to improve the drug aspects of the disease. Weprovided two contributions in the field of proteomics in drug design. First, we have constructed a protein-protein interactionnetwork for Alzheimer''s disease reviewed proteins with 1412 interactions predicted among 969 proteins. Second, the diseaseproteins were given confidence scores to prioritize and then analyzed for their homology nature with respect to paralogs andhomologs. The homology persisted with the mouse giving a basis for drug design phase. The method will create a new drug designtechnique in the field of bioinformatics by linking drug design process with protein-protein interactions via signal pathways. Thismethod can be improvised for other diseases in future.     

Wheat germ agglutinin-binding glycoproteins are decreased in Alzheimer's disease cerebrospinal fluid

A number of biomarkers (e.g. Abeta, tau) has been identified in Alzheimer's disease CSF. However, none fulfils the criteria of sensitivity and specificity (> 80%) needed for the development of an accurate diagnostic test. The lack of a suitable marker has prompted the search for new CSF biomarkers. In this study, the glycosylation of CSF proteins was examined using lectin blotting. Lumbar CSF was collected ante mortem from 22 non-Alzheimer's disease and 12 probable Alzheimer's disease cases and ventricular CSF collected post mortem from 7 non-Alzheimer's disease and 16 Alzheimer's disease cases confirmed by pathologic examination. When CSF glycoproteins were stained with wheat germ agglutinin (WGA), the staining intensity was found to be significantly lower in the Alzheimer's disease group. No difference in staining was found using other lectins (Canavalia ensiformis agglutinin, Ricinus communis agglutinin, Lens culinaris agglutinin). The measurement of WGA-reactive glycoproteins in CSF may be a useful biomarker for diagnosis of Alzheimer's disease.     

Characterization of the glycated human cerebrospinal fluid proteome

Protein glycation is a nonenzymatic modification that involves pathological functions in neurological diseases. Despite the high number of studies showing accumulation of advanced end glycation products (AGEs) at clinical stage, there is a lack of knowledge about which proteins are modified, where those modifications occur, and to what extent. The goal of this study was to achieve a comprehensive characterization of proteins modified by early glycation in human cerebrospinal fluid (CSF). Approaches based on glucose diferential labeling and mass spectrometry have been applied to evaluate the glycated CSF proteome at two physiological conditions: native glucose level and in vitro high glucose content. For both purposes, detection of glycated proteins was carried out by HCD-MS2 and CID-MS3 modes after endoproteinase Glu-C digestion and boronate affinity chromatography. The abundance of glycation was assessed by protein labeling with (13)C(6)-glucose incubation. The analysis of native glycated CSF identified 111 glycation sites corresponding to 48 glycated proteins. Additionally, the in vitro high glucose level approach detected 265 glycation sites and 101 glycated proteins. The comparison of glycation levels under native and 15 mM glucose conditions showed relative concentration increases up to ten folds for some glycated proteins. This report revealed for the first time a number of key glycated CSF proteins known to be involved in neuroinflammation and neurodegenerative disorders. Altogether, the present study contains valuable and unique information, which should further help to clarify the pathological role of glycation in central nervous system pathologies. This article is part of a Special Issue entitled: Translational Proteomics.     

Molecular docking analysis of hyperphosphorylated tau protein with compounds derived from Bacopa monnieri and Withania somnifera

Tau protein, the major player in Alzheimer’s disease forms neurofibrillary tangles in elderly people. Bramhi (Baccopa Monniera) is often used as an ayurvedic treatment for Alzheimer''s disease. Therefore it is of interest to study the interaction of compounds derived from Baccopa with the Tau protein involved in tangle formation. We show that compounds such as bacopaside II, bacopaside XII, and nicotine showed optimal binding features with the R2 repeat domain of hyperphosphorylated tau protein for further consideration in the context of Alzheimer''s disease (AD).     

Glycomics of pediatric and adulthood diseases of the central nervous system

Glycosylation consists in the covalent linkage of a carbohydrate structure to membrane bound and secreted glycoconjugates. It is a common post-translational modification that serves multiple functions in cell differentiation, signaling and intercellular communication. Unlike DNA/RNA/protein, the addition of complex carbohydrates is not-template driven and it is conceivable that both genetics and environmental factors might interact to influence glycosylation machinery in several pathological processes. Over the last few decades, the recognition of Congenital Disorders of Glycosylation (CDG) as an increasing number of genetic diseases of glycosylation with almost constant nervous system involvement, dramatically illustrated the consequences of abnormal glycosylation as improper CNS development and function. In addition, CDG recognition contributed to postulate that aberrant glycosylation processes might play a role in multifactorial, complex CNS diseases. On this context, CNS glycomics explores the effects of possible aberrant glycosylation to identify potential glyco-biomarkers useful for the diagnosis and ultimately for potential intervention strategies in neurological diseases. Up to date, CNS glycomics is an emerging, still uncharted area because of the specificity of CNS glycosylation, the complexity of the neurological disorders and for the inaccessibility and invasiveness of disease relevant samples. Here we review current knowledge on clinical glycomics of nervous system diseases, starting with CDG to include those pediatric and adulthood neuropsychiatric diseases where some evidences suggest that multifactor determinants converge to dysregulate glycosylation. Conventional and mass spectrometry-based high throughput technology for glyco-biomarker detection in CNS diseases is reported.