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Increased striatal mRNA transcription and active protein expression of the immunophilin FKBP-12 in experimental Parkinson’s disease models
Uppsala University, Medicinska vetenskapsområdet, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
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Identifiers
URN: urn:nbn:se:uu:diva-92420OAI: oai:DiVA.org:uu-92420DiVA: diva2:165488
Available from: 2004-11-09 Created: 2004-11-09 Last updated: 2010-01-13Bibliographically approved
In thesis
1. Development of Methods for Protein and Peptide Analysis Applied in Neuroscience Utilizing Mass Spectrometry
Open this publication in new window or tab >>Development of Methods for Protein and Peptide Analysis Applied in Neuroscience Utilizing Mass Spectrometry
2004 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

This thesis describes the utilization of the matrix-assisted laser desorption ionization mass spectrometry (MALDI MS) and electrospray ionization (ESI) MS techniques for analysis of complex brain tissue samples.

Direct molecular profiling of biological samples using MALDI MS is a powerful tool for identifying phenotypic markers. MALDI MS-profiling of proteins and peptides directly on brain tissue sections was used for the first time to study experimental models of Parkinson’s disease (PD). The mass spectrometer was used to map the peptide and protein expression directly on 12 µm tissue sections in mass-to-charge (m/z) values, providing the capability of mapping specific molecules of the original sample, that is, localization, intensity and m/z ratio. Several protein and peptide expression profile differences were found in the dopamine denervated brains when compared to the corresponding controls, for example, calmodulin, cytochrome c, cytochrome c oxidase, and the neuroimmunophilin protein FKBP-12. The increased expression of FKBP-12 from the profiling experiments was supported by mRNA expression analysis and two-dimensional gel electrophoresis separation analysis. Multiple genetic deficits have linked impaired ubiquitin-conjugation pathways to various forms of familiar PD. This study showed for the first time an increased level of unconjugated ubiquitin specifically in the dorsal striatum of the dopamine depleted PD brain. The strength of the MALDI MS-profiling technique is that a minimum of sample handling and manipulation is necessary pre-analysis. This ensures preservation of the spatial localization of the biomolecules in the tissue section.

Biological liquid samples often contain high amounts of salt that is non-compatible with the ESI MS technique. A nano-flow capillary liquid chromatography (nanoLC) system coupled on-line with ESI-MS was used to study the metabolism of the peptide LVV-hemorphin-7 in the brain and blood using in vivo microdialysis. The microdialysis technique provides capabilities for very precise sampling in specific brain regions. The combination of on-line desalting and pre-concentration by nanoLC with ESI MS is a powerful tool to detect minute concentration of metabolic fragments and endogenous biomolecules.

The utilization of mass spectrometry in neuroscience applications provides a uniquely advantageous tool for the analysis of complex biochemical events that underlie the pathological symptoms expressed in different disease states. Furthermore, the MALDI-MS profiling technique shows great potential for the future with regards to proteome analysis and drug discovery.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2004. 41 p.
Series
Comprehensive Summaries of Uppsala Dissertations from the Faculty of Pharmacy, ISSN 0282-7484 ; 318
Keyword
Analytical chemistry, Mass spectrometry, Parkinson’s disease, Metabolism, Profiling Mass Spectrometry, Proteomics, Analytisk kemi
National Category
Analytical Chemistry
Identifiers
urn:nbn:se:uu:diva-4685 (URN)91-554-6104-2 (ISBN)
Public defence
2004-12-01, B:41, BMC, 13:15
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Available from: 2004-11-09 Created: 2004-11-09Bibliographically approved

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