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Applications of nanospray desorption electrospray ionization mass spectrometry: In situ lipid and metabolite analysis from cells to tissue
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
2018 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Ambient mass spectrometry (MS) has proved to be an important addition to the bioanalytical toolbox. These methods perform analyte sampling and ionization under atmospheric pressure, and require very little sample preparation other than the sampling process in front of the machine. Nanospray desorption electrospray ionization (nano-DESI) is an ambient MS technique developed in 2010 that utilizes localized liquid extraction for surface sampling. The aim of this thesis was to explore the possibilities of this technique, and identify areas in which nano-DESI MS could further contribute to the community of MS-based surface analysis.

One such area was found to be mass spectrometry imaging (MSI) of small-molecule neurotransmitters. By the use of deuterated standards of acetylcholine, γ-aminobutyric acid and glutamate, the respective endogenous compounds were successfully imaged in coronal sections of rat brain. The use of internal standards was shown to be essential to compensatee for matrix effects in different regions of the brain. In a second imaging study, nano-DESI MSI was used to compare the chemical profiles of diabetic rat kidney tissue and control. Analysis was performed on kidney two weeks after diabetic onset, before any pathohistological changes relating to diabetic nephropathy can be seen in a microscope. In our study, it was shown that a large number of chemical species related to energy metabolism were detected with altered signal intensity in diabetic kidney tissue.

To push the limits of nano-DESI analysis, its use for single-cell analysis was evaluated. By placing buccal epithelial cells in contact with the nano-DESI probe, it was possible to identify 46 endogenous compounds and detect differences between cells from three human donors. In addition, it was shown that molecules from single cells on a surface could be detected by scanning the surface with the nano-DESI probe, which opens up for development of an automated analysis with higher throughput.

The last study in this thesis was concerned with method development rather than application, as it presented a setup for pneumatically assisted nano-DESI. Evaluation showed that the setup provided improved sensitivity in the analysis of small metabolites, and provided the possibility of using pure water as nano-DESI solvent.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2018. , p. 78
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 1660
Keywords [en]
Mass spectrometry, mass spectrometry imaging (MSI), nanospray desorption electrospray ionization (nano-DESI), single-cell analysis, neurotransmitter imaging, diabetic nephropathy, pneumatic nebulization, lipidomics, metabolomics
National Category
Chemical Sciences
Research subject
Chemistry with specialization in Analytical Chemistry
Identifiers
URN: urn:nbn:se:uu:diva-347674ISBN: 978-91-513-0307-9 (print)OAI: oai:DiVA.org:uu-347674DiVA, id: diva2:1195599
Public defence
2018-05-25, A1:107a, BMC, Husargatan 3, Uppsala, 13:00 (English)
Opponent
Supervisors
Available from: 2018-05-03 Created: 2018-04-05 Last updated: 2018-10-08
List of papers
1. Quantitative mass spectrometry imaging of small-molecule neurotransmitters in rat brain tissue sections using nanospray desorption electrospray ionization
Open this publication in new window or tab >>Quantitative mass spectrometry imaging of small-molecule neurotransmitters in rat brain tissue sections using nanospray desorption electrospray ionization
2016 (English)In: The Analyst, ISSN 0003-2654, E-ISSN 1364-5528, Vol. 141, no 12, p. 3686-3695Article in journal (Refereed) Published
Abstract [en]

Small molecule neurotransmitters are essential for the function of the nervous system, and neurotransmitter imbalances are often connected to neurological disorders. The ability to quantify such imbalances is important to provide insights into the biochemical mechanisms underlying the disorder. This proof-of-principle study presents online quantification of small molecule neurotransmitters, specifically acetylcholine, γ-aminobutyric acid (GABA) and glutamate, in rat brain tissue sections using nanospray desorption electrospray ionization (nano-DESI) mass spectrometry imaging. By incorporating deuterated internal standards in the nano-DESI solvent we show identification, accurate mapping, and quantification of these small neurotransmitters in rat brain tissue without introducing any additional sample preparation steps. We find that GABA is about twice as abundant in the medial septum-diagonal band complex (MSDB) as in the cortex, while glutamate is about twice as abundant in the cortex as compared to the MSDB. The study shows that nano-DESI is well suited for imaging of small molecule neurotransmitters in health and disease.

National Category
Analytical Chemistry
Identifiers
urn:nbn:se:uu:diva-281314 (URN)10.1039/c5an02620b (DOI)000378942900021 ()26859000 (PubMedID)
Funder
Swedish Research Council, VR 621-2013-4231Swedish Foundation for Strategic Research , SSF ICA-6
Available from: 2016-03-22 Created: 2016-03-22 Last updated: 2018-11-29
2. Metabolite aberrations in early diabetes detected in rat kidney using mass spectrometry imaging
Open this publication in new window or tab >>Metabolite aberrations in early diabetes detected in rat kidney using mass spectrometry imaging
Show others...
2019 (English)In: Analytical and Bioanalytical Chemistry, ISSN 1618-2642, E-ISSN 1618-2650, Vol. 411, no 13, p. 2809-2816Article in journal (Refereed) Published
Abstract [en]

Diabetic kidney disease is a serious complication of diabetes that can ultimately lead to end-stage renal disease. The pathogenesis of diabetic kidney disease is complex, and fundamental research is still required to provide a better understanding of the driving forces behind it. We report regional metabolic aberrations from an untargeted mass spectrometry imaging study of kidney tissue using an insulinopenic rat model of diabetes. Diabetes was induced by intravenous injection of streptozotocin, and kidneys were harvested 2weeks thereafter. Imaging was performed using nanospray desorption electrospray ionization connected to a high-mass-resolving mass spectrometer. No histopathological changes were observed in the kidney sections; however, mass spectrometry imaging revealed a significant increase in several 18-carbon unsaturated non-esterified fatty acid species and monoacylglycerols. Notably, these 18-carbon acyl chains were also constituents of several increased diacylglycerol species. In addition, a number of short- and long-chain acylcarnitines were found to be accumulated while several amino acids were depleted. This study presents unique regional metabolic data indicating a dysregulated energy metabolism in renal mitochondria as an early response to streptozotocin-induced type I diabetes.

National Category
Analytical Chemistry
Research subject
Chemistry with specialization in Analytical Chemistry
Identifiers
urn:nbn:se:uu:diva-347672 (URN)10.1007/s00216-019-01721-5 (DOI)000468133600008 ()30895347 (PubMedID)
Funder
Swedish Foundation for Strategic Research Swedish Research CouncilSwedish Diabetes AssociationAstraZeneca
Note

Title in dissertation list of papers: Metabolite aberrations at early onset of diabetes detected in rat kidney using mass spectrometry imaging

Available from: 2018-04-05 Created: 2018-04-05 Last updated: 2020-03-24Bibliographically approved
3. Profiling and quantifying endogenous molecules in single cells using nano-DESI MS
Open this publication in new window or tab >>Profiling and quantifying endogenous molecules in single cells using nano-DESI MS
2017 (English)In: The Analyst, ISSN 0003-2654, E-ISSN 1364-5528, Vol. 142, no 19, p. 3639-3647Article in journal (Refereed) Published
Abstract [en]

Molecular profiling of single cells has the potential to significantly advance our understanding of cell function and cellular processes of importance to health and disease. In particular, small molecules with rapid turn-over rates can reveal activated metabolic pathways resulting from an altered chemical environment or cellular events such as differentiation. Consequently, techniques for quantitative metabolite detection acquired in a higher throughput manner are needed to characterize the biological variability between seemingly homogenous cells. Here, we show that nanospray desorption electrospray ionization (nano-DESI) mass spectrometry ( MS) enables sensitive molecular profiling and quantification of endogenous species in single cells in a higher throughput manner. Specifically, we show a large number of detected amino acids and phospholipids, including plasmalogens, readily detected from single cheek cells. Further, by incorporating a phosphatidylcholine ( PC) internal standard into the nano-DESI solvent, we determined the total amount of PC in one cell to be 1.2 pmoles. Finally, we describe a higher throughput approach where molecules in single cells are automatically profiled. These developments in single cell analysis provide a basis for future studies to understand cellular processes related to drug effects, cell differentiation and altered chemical microenvironments.

Place, publisher, year, edition, pages
ROYAL SOC CHEMISTRY, 2017
National Category
Analytical Chemistry
Research subject
Chemistry with specialization in Analytical Chemistry
Identifiers
urn:nbn:se:uu:diva-336430 (URN)10.1039/c7an00885f (DOI)000411703800013 ()28835951 (PubMedID)
Funder
Swedish Research Council, VR 621-2013-4231Swedish Foundation for Strategic Research , SSF ICA-6
Available from: 2017-12-15 Created: 2017-12-15 Last updated: 2018-11-29
4. A pneumatically assisted nanospray desorption electrospray ionization source for increased solvent versatility and enhanced metabolite detection from tissue
Open this publication in new window or tab >>A pneumatically assisted nanospray desorption electrospray ionization source for increased solvent versatility and enhanced metabolite detection from tissue
2017 (English)In: The Analyst, ISSN 0003-2654, E-ISSN 1364-5528, Vol. 142, no 18, p. 3424-3431Article in journal (Refereed) Published
Abstract [en]

Nanospray desorption electrospray ionization (nano-DESI) has been established as a powerful technique for mass spectrometry imaging (MSI) of biomolecules from tissue samples. The direct liquid extraction of analytes from a surface at ambient pressure negates the need for significant sample preparation or matrix application. Although many recent studies have applied nano-DESI to new and exciting applications, there has not been much work in the development and improvement of the nano-DESI source. Here, we incorporate a nebulizer to replace the self-aspirating secondary capillary in the conventional nano-DESI setup, and characterize the device by use of rat kidney tissue sections. We find that the pneumatically assisted nano-DESI device offers improved sensitivity for metabolite species by 1-3 orders of magnitude through more complete desolvation and reduced ionization suppression. Further, the pneumatically assisted nano-DESI device reduces the dependence on probe-to-surface distance and enables sampling and imaging using pure water as the nano-DESI solvent. This provides exclusive detection and imaging of many highly polar endogenous species. Overall, the developed pneumatically assisted nano-DESI device provides more versatile solvent selection and an increased sensitivity for metabolites, which generates ion images of higher contrast - allowing for more intricate studies of metabolite distribution.

Place, publisher, year, edition, pages
ROYAL SOC CHEMISTRY, 2017
National Category
Chemical Sciences
Identifiers
urn:nbn:se:uu:diva-334756 (URN)10.1039/c7an00901a (DOI)000409919200016 ()28828451 (PubMedID)
Funder
Swedish Foundation for Strategic Research , SSF ICA-6Swedish Research Council, VR 621-2013-4231
Available from: 2017-11-27 Created: 2017-11-27 Last updated: 2018-11-29

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