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Cerebrum Illuminans: Mass Spectrometric Analysis of Protein and Peptide Dynamics in Neurological Diseases
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
2010 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The human brain (lat. cerebrum) is the most complex and heterogeneous organ in the human body. It is involved in a great number of body functions like movement, touch sensing, vision, hearing, smelling, hormone regulation and many more. In no other organ, the molecular communication mechanisms between different cells are so poorly understood. Due to the extensive diversity of processes that are controlled by the brain, diseases and injuries of the nervous system affect the human body significantly. Because of the immense complexity of the brain, the molecular mechanisms underlying the pathology of the diseases remain largely unknown.

Hence, there is an urgent need for the development of new analytical strategies in order to investigate these conditions on a molecular level. Here, a central focus lies in the study of protein and peptide expression profiles, which can provide an insight in ongoing molecular mechanisms underlying the pathophysiology of the diseases. A powerful approach for studying proteins and peptide dynamics is mass spectrometry based proteomics, which is defined as the comprehensive study of all proteins expressed in a biological matrix at a certain point of time.

The central objective of this thesis was to develop and employ different mass spectrometric techniques to study protein and peptide dynamics in the central nervous system in different neurological diseases. The individual studies comprise different aspects of proteome research. The first two studies included clinical proteomic applications for investigating protein dynamics in traumatic brain injury and amyotrophic lateral sclerosis. A further study was focused on method development for MS analysis of intact neural cells. The final three projects described in this thesis comprised MS based protein and peptide imaging in brain and spinal cord tissue samples. Here, the aim was to elucidate topological changes in protein expression in ALS as well as neuropeptide alterations in distinct brain structures in L-DOPA induced dyskinesia (LID) in Parkinson’s disease.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis , 2010. , p. 87
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 783
Keywords [en]
mass spectrometry, central nervous system, neurological disease, proteomics, MALDI imaging
National Category
Analytical Chemistry
Research subject
Chemistry with specialization in Analytical Chemistry
Identifiers
URN: urn:nbn:se:uu:diva-132908ISBN: 978-91-554-7937-4 (print)OAI: oai:DiVA.org:uu-132908DiVA, id: diva2:359600
Public defence
2010-12-10, B41, Uppsala Biomedical Center, Husargatan 3, Uppsala, 10:15 (English)
Opponent
Supervisors
Note
Felaktigt tryckt som Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology 713Available from: 2010-11-19 Created: 2010-10-28 Last updated: 2011-07-13
List of papers
1. Temporally resolved differential proteomic analysis of human ventricular CSF for monitoring traumatic brain injury biomarker candidates.
Open this publication in new window or tab >>Temporally resolved differential proteomic analysis of human ventricular CSF for monitoring traumatic brain injury biomarker candidates.
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2009 (English)In: Journal of Neuroscience Methods, ISSN 0165-0270, E-ISSN 1872-678X, Vol. 177, no 2, p. 469-478Article in journal (Refereed) Published
Abstract [en]

A shotgun proteomic approach based on nanoflow liquid chromatography (nanoLC) in conjunction with matrix assisted laser desorption/ionization time of flight tandem mass spectrometry (MALDI TOF MS/MS) was utilized to quantitatively analyze the protein content of consecutive ventricular cerebrospinal fluid (CSF) samples of severe traumatic brain injury (TBI) patients on an individual basis. CSF was acquired from the lateral ventricle 1–9 days after the TBI incident by canula drain to investigate temporally resolved protein changes in three patients that required intracranial pressure monitoring during neurointensive care. The samples were subjected to at once tryptic digestion followed by isobaric tag labeling before multiplexed peptide separation and MS analysis. By using this approach, we were able to follow characteristic changes in protein concentrations over time allowing new conclusions to be drawn about ongoing pathological processes during TBI. Certain suggested protein-biomarker candidates for TBI, like acute phase reactants (APRs), fibrinogens (FIB), cystatin C (CC) or more brain specific proteins like glial fibrillary acid protein (GFAP) and neuron-specific enolase (NSE) were found to be significantly up-regulated which is in strong consistence with previously reported results. This methodology appears to be a promising tool for studying candidate biomarkers of neurovascular and traumatic brain injuries in the neurointensive care setting.

Keywords
Traumatic brain injury (TBI), ventricular cerebrospinal fluid (CSF), shotgun proteomics, protein quantification, isobaric tag labeling, matrix assisted laser desorption/ionization, time of flight tandem mass spectrometry (MALDI TOF MS/MS)
National Category
Analytical Chemistry
Research subject
Analytical Chemistry
Identifiers
urn:nbn:se:uu:diva-88519 (URN)10.1016/j.jneumeth.2008.10.038 (DOI)000263393300027 ()19263575 (PubMedID)
Available from: 2009-02-03 Created: 2009-02-03 Last updated: 2017-12-14Bibliographically approved
2. Focused proteomics in post-mortem human spinal cord
Open this publication in new window or tab >>Focused proteomics in post-mortem human spinal cord
2006 (English)In: Journal of Proteome Research, ISSN 1535-3893, E-ISSN 1535-3907, Vol. 5, no 9, p. 2364-2371Article in journal (Refereed) Published
Abstract [en]

With a highly sensitive electrospray ionization-Fourier transform ion cyclotron resonance mass spectrometry (ESI-FTICR MS) system, proteins were identified in minimal amounts of spinal cord from patients with the neurodegenerative disease amyotrophic lateral sclerosis (ALS) and compared to proteins in spinal cord from control subjects. The results show 18 versus 16 significantly identified ( p < 0.05) proteins, respectively, all known to be found in the central nervous system. The most abundant protein in both groups was the glial fibrillary acidic protein, GFAP. Other proteins were, for example, hemoglobin alpha- and, chain, myelin basic protein, thioredoxin, R enolase, and cholin acetyltransferase. This study also includes the technique of laser microdissection in combination with pressure catapulting (LMPC) for the dissection of samples and specific neurons. Furthermore, complementary experiments with nanoLC-matrix assisted laser desorption ionization time-of-flight tandem mass spectrometry (MALDI-TOF-TOF MS) confirmed the results of the ESI-FTICR MS screening and provided additional results of further identified proteins.

Keywords
proteomics, electrospray ionization - Fourier transform ion cyclotron resonance mass spectrometry (ESI-FTICR MS), matrix-assisted laser desorption ionization time-of-flight tandem mass spectrometry (MALDI-TOF-TOF-MS), neurodegeneration, amyotrophic lateral sclerosis, spinal cord, laser microdissection with pressure catapulting (LMPC)
National Category
Chemical Sciences
Identifiers
urn:nbn:se:uu:diva-82961 (URN)10.1021/pr060237f (DOI)000240200700033 ()16944948 (PubMedID)
Available from: 2006-10-25 Created: 2006-10-25 Last updated: 2017-12-14Bibliographically approved
3. MALDI mass spectrometry based molecular phenotyping of CNS glial cells for prediction in mammalian brain tissue
Open this publication in new window or tab >>MALDI mass spectrometry based molecular phenotyping of CNS glial cells for prediction in mammalian brain tissue
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2011 (English)In: Analytical and Bioanalytical Chemistry, ISSN 1618-2642, E-ISSN 1618-2650, Vol. 401, no 1, p. 135-147Article in journal (Refereed) Published
Abstract [en]

The development of powerful analytical techniques for specific molecular characterization of neural cell types is of central relevance in neuroscience research for elucidating cellular functions in the central nervous system (CNS). This study examines the use of differential protein expression profiling of mammalian neural cells using direct analysis by means of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). MALDI-MS analysis is rapid, sensitive, robust, and specific for large biomolecules in complex matrices. Here, we describe a newly developed and straightforward methodology for direct characterization of rodent CNS glial cells using MALDI-MS-based intact cell mass spectrometry (ICMS). This molecular phenotyping approach enables monitoring of cell growth stages, (stem) cell differentiation, as well as probing cellular responses towards different stimulations. Glial cells were separated into pure astroglial, microglial, and oligodendroglial cell cultures. The intact cell suspensions were then analyzed directly by MALDI-TOF-MS, resulting in characteristic mass spectra profiles that discriminated glial cell types using principal component analysis. Complementary proteomic experiments revealed the identity of these signature proteins that were predominantly expressed in the different glial cell types, including histone H4 for oligodendrocytes and S100-A10 for astrocytes. MALDI imaging MS was performed, and signature masses were employed as molecular tracers for prediction of oligodendroglial and astroglial localization in brain tissue. The different cell type specific protein distributions in tissue were validated using immunohistochemistry. ICMS of intact neuroglia is a simple and straightforward approach for characterization and discrimination of different cell types with molecular specificity.

Keywords
Intact cell mass spectrometry (ICMS), MALDITOF- MS, Imaging mass spectrometry (IMS), Glial cells
National Category
Analytical Chemistry
Identifiers
urn:nbn:se:uu:diva-156047 (URN)10.1007/s00216-011-5043-y (DOI)000292156900012 ()21553124 (PubMedID)
Available from: 2011-07-08 Created: 2011-07-08 Last updated: 2017-12-11Bibliographically approved
4. MALDI Imaging of Post Mortem Spjnal Cord in Amyotrophic Lateral Sclerosis
Open this publication in new window or tab >>MALDI Imaging of Post Mortem Spjnal Cord in Amyotrophic Lateral Sclerosis
(English)Manuscript (preprint) (Other academic)
Identifiers
urn:nbn:se:uu:diva-132440 (URN)
Available from: 2010-10-20 Created: 2010-10-20 Last updated: 2012-10-18
5. Imaging mass spectrometry reveals association of nigral levels of dynorphin peptides and L-DOPA-induced dyskinesia in Parkinson’s disease
Open this publication in new window or tab >>Imaging mass spectrometry reveals association of nigral levels of dynorphin peptides and L-DOPA-induced dyskinesia in Parkinson’s disease
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(English)Article in journal (Refereed) Submitted
Identifiers
urn:nbn:se:uu:diva-132438 (URN)
Available from: 2010-10-20 Created: 2010-10-20 Last updated: 2011-08-22Bibliographically approved
6. Alterations of striatal dynorphin peptide levels associated with L-DOPA-induced dyskinesia elucidated by imaging mass spectrometry
Open this publication in new window or tab >>Alterations of striatal dynorphin peptide levels associated with L-DOPA-induced dyskinesia elucidated by imaging mass spectrometry
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2010 (English)Manuscript (preprint) (Other academic)
Publisher
p. 96
Identifiers
urn:nbn:se:uu:diva-132441 (URN)
Available from: 2010-10-28 Created: 2010-10-20 Last updated: 2013-06-12

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