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Increased levels of extracellular microvesicle markers and decreased levels of endocytic/exocytic proteins in the Alzheimer’s disease brain
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Cancer Pharmacology and Computational Medicine.
KTH Royal Inst Technol, Sch Biotechnol, Sci Life Lab, Affin Prote, Stockholm, Sweden.
GE Healthcare Life Sci, Uppsala, Sweden.
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2016 (English)In: Journal of Alzheimer's Disease, ISSN 1387-2877, E-ISSN 1875-8908, Vol. 54, no 4, 71 p.1671-1686 p.Article in journal (Refereed) Published
Abstract [en]

Background: Alzheimer's disease (AD) is a chronic neurodegenerative disorder accounting for more than 50% of all dementia cases. AD neuropathology is characterized by the formation of extracellular plaques and intracellular neurofibrillary tangles consisting of aggregated amyloid-beta and tau, respectively. The disease mechanism has only been partially elucidated and is believed to also involve many other proteins.

Objective: This study intended to perform a proteomic profiling of post mortem AD brains and compare it with control brains as well as brains from other neurological diseases to gain insight into the disease pathology.

Methods: Here we used label-free shotgun mass spectrometry to analyze temporal neocortex samples from AD, other neurological disorders, and non-demented controls, in order to identify additional proteins that are altered in AD. The mass spectrometry results were verified by antibody suspension bead arrays.

Results: We found 50 proteins with altered levels between AD and control brains. The majority of these proteins were found at lower levels in AD. Pathway analyses revealed that several of the decreased proteins play a role in exocytic and endocytic pathways, whereas several of the increased proteins are related to extracellular vesicles. Using antibody-based analysis, we verified the mass spectrometry results for five representative proteins from this group of proteins (CD9, HSP72, PI42A, TALDO, and VAMP2) and GFAP, a marker for neuroinflammation.

Conclusions: Several proteins involved in exo-endocytic pathways and extracellular vesicle functions display altered levels in the AD brain. We hypothesize that such changes may result in disturbed cellular clearance and a perturbed cell-to-cell communication that may contribute to neuronal dysfunction and cell death in AD.

Place, publisher, year, edition, pages
2016. Vol. 54, no 4, 71 p.1671-1686 p.
Keyword [en]
Brain, Proteomics, Mass spectrometry, Alzheimer's disease
National Category
Analytical Chemistry Geriatrics Neurosciences
Research subject
Chemistry with specialization in Analytical Chemistry
Identifiers
URN: urn:nbn:se:uu:diva-277617DOI: 10.3233/JAD-160271ISI: 000386749900034PubMedID: 27636840OAI: oai:DiVA.org:uu-277617DiVA: diva2:905201
Funder
VINNOVALars Hierta Memorial FoundationSwedish Research Council, P29797-1; 621-2011-4423Knut and Alice Wallenberg FoundationStiftelsen Gamla Tjänarinnor
Available from: 2016-02-22 Created: 2016-02-22 Last updated: 2017-10-17Bibliographically approved
In thesis
1. Mass Spectrometry-based Neuroproteomics: Deciphering the Human Brain Proteome
Open this publication in new window or tab >>Mass Spectrometry-based Neuroproteomics: Deciphering the Human Brain Proteome
2016 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Mammalian brain is challenging to study due to its heterogeneity and complexity. However, recent advances in molecular imaging, genomics and proteomics have contributed significantly to achieve insights into molecular basis of brain function and pathogenesis of neurological disorders. Efficient sample preparation is an integral part of a successful mass spectrometry (MS)-based proteomics. Apart from the identification, quantification of proteins is needed to investigate the alterations between proteome profiles from different sample sets. Therefore, this thesis investigates optimizing and application of the MS compatible sample preparation techniques for the identification and quantification of proteins from brain tissue.

The central objective of this thesis was (i) to improve the extraction of proteins as well as membrane proteins (MPs) from the brain tissue and (ii) to apply the optimized method along with the stable isotope dimethyl labeling (DML) and label-free (LF) MS approaches for the relative quantification of the brain proteome profiles during neurological conditions such as Alzheimer’s disease (AD) and traumatic brain injury (TBI).  First study described in this thesis is focused on the qualitative aspects for the brain tissue sample preparation. The optimized extraction buffers from first study containing n-octyl-β-glucopyranside or triton X-114 were used in the further quantitative studies to extract the proteins from patient (AD or TBI) and control human brain samples. Triton X-114 has additional advantage of separating MPs into a micellar phase. Therefore we also investigated the possibility to apply this in combination with DML quantitation approach for enrichment of low abundant MPs from AD brains.

AD and TBI causes severe socio-economic burden on the society and therefore there is a need to develop diagnostic markers to detect the early changes in the pathology of the disease. Analytical tools and techniques applied and discussed in this thesis for neuroproteomics applications proved to be powerful and reliable for analyzing complex biological samples to generate high-throughput screening and unbiased identification and quantitation of disease-specific proteins that are of great importance in understanding the disease pathology. 

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2016. 70 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 1347
Keyword
Brain, Proteomics, Mass spectrometry, Alzheimer's disease, Traumatic brain injury, Membrane proteins, Sample preparation
National Category
Analytical Chemistry
Research subject
Chemistry with specialization in Analytical Chemistry
Identifiers
urn:nbn:se:uu:diva-277613 (URN)978-91-554-9485-8 (ISBN)
Public defence
2016-04-08, B42, BMC, Husargatan 3, Uppsala, 10:15 (English)
Opponent
Supervisors
Available from: 2016-03-18 Created: 2016-02-22 Last updated: 2016-04-04
2. Proteomics Studies of Subjects with Alzheimer’s Disease and Chronic Pain
Open this publication in new window or tab >>Proteomics Studies of Subjects with Alzheimer’s Disease and Chronic Pain
2017 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Alzheimer’s disease (AD) is a neurodegenerative disease and the major cause of dementia, affecting more than 50 million people worldwide. Chronic pain is long-lasting, persistent pain that affects more than 1.5 billion of the world population. Overlapping and heterogenous symptoms of AD and chronic pain conditions complicate their diagnosis, emphasizing the need for more specific biomarkers to improve the diagnosis and understand the disease mechanisms.

To characterize disease pathology of AD, we measured the protein changes in the temporal neocortex region of the brain of AD subjects using mass spectrometry (MS). We found proteins involved in exo-endocytic and extracellular vesicle functions displaying altered levels in the AD brain, potentially resulting in neuronal dysfunction and cell death in AD.

To detect novel biomarkers for AD, we used MS to analyze cerebrospinal fluid (CSF) of AD patients and found decreased levels of eight proteins compared to controls, potentially indicating abnormal activity of complement system in AD.

By integrating new proteomics markers with absolute levels of Aβ42, total tau (t-tau) and p-tau in CSF, we improved the prediction accuracy from 83% to 92% of early diagnosis of AD. We found increased levels of chitinase-3-like protein 1 (CH3L1) and decreased levels of neurosecretory protein VGF (VGF) in AD compared to controls.

By exploring the CSF proteome of neuropathic pain patients before and after successful spinal cord stimulation (SCS) treatment, we found altered levels of twelve proteins, involved in neuroprotection, synaptic plasticity, nociceptive signaling and immune regulation.

To detect biomarkers for diagnosing a chronic pain state known as fibromyalgia (FM), we analyzed the CSF of FM patients using MS. We found altered levels of four proteins, representing novel biomarkers for diagnosing FM. These proteins are involved in inflammatory mechanisms, energy metabolism and neuropeptide signaling.

Finally, to facilitate fast and robust large-scale omics data handling, we developed an e-infrastructure. We demonstrated that the e-infrastructure provides high scalability, flexibility and it can be applied in virtually any fields including proteomics. This thesis demonstrates that proteomics is a promising approach for gaining deeper insight into mechanisms of nervous system disorders and find biomarkers for diagnosis of such diseases.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2017. 82 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1385
Keyword
Bioinformatics, microservices, biomarkers, Alzheimer's disease, chronic pain, fibromyalgia, neuropathic pain, spinal cord stimulation, cloud computing, proteomics, metabolomics, software, workflows, data analysis, mass spectrometry
National Category
Geriatrics Neurology Neurosciences
Research subject
Bioinformatics; Neurology; Geriatrics
Identifiers
urn:nbn:se:uu:diva-331748 (URN)978-91-513-0111-2 (ISBN)
Public defence
2017-12-05, Rosénsalen, Akademiska sjukhuset, Ing 95/96, nbv, Uppsala, 09:00 (English)
Opponent
Supervisors
Available from: 2017-11-14 Created: 2017-10-17 Last updated: 2017-11-14

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Sravani, MusunuriEmami Khoonsari, Payam

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