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Träfflista för sökning "WFRF:(Andren Per E.) "

Sökning: WFRF:(Andren Per E.)

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1.
  • Becker, K., et al. (författare)
  • Antibacterial activity of apramycin at acidic pH warrants wide therapeutic window in the treatment of complicated urinary tract infections and acute pyelonephritis
  • 2021
  • Ingår i: EBioMedicine. - : Elsevier B.V.. - 2352-3964. ; 73
  • Tidskriftsartikel (refereegranskat)abstract
    • Background: The clinical-stage drug candidate EBL-1003 (apramycin) represents a distinct new subclass of aminoglycoside antibiotics for the treatment of drug-resistant infections. It has demonstrated best-in-class coverage of resistant isolates, and preclinical efficacy in lung infection models. However, preclinical evidence for its utility in other disease indications has yet to be provided. Here we studied the therapeutic potential of EBL-1003 in the treatment of complicated urinary tract infection and acute pyelonephritis (cUTI/AP). Methods: A combination of data-base mining, antimicrobial susceptibility testing, time-kill experiments, and four murine infection models was used in a comprehensive assessment of the microbiological coverage and efficacy of EBL-1003 against Gram-negative uropathogens. The pharmacokinetics and renal toxicology of EBL-1003 in rats was studied to assess the therapeutic window of EBL-1003 in the treatment of cUTI/AP. Findings: EBL-1003 demonstrated broad-spectrum activity and rapid multi-log CFU reduction against a phenotypic variety of bacterial uropathogens including aminoglycoside-resistant clinical isolates. The basicity of amines in the apramycin molecule suggested a higher increase in positive charge at urinary pH when compared to gentamicin or amikacin, resulting in sustained drug uptake and bactericidal activity, and consequently in potent efficacy in mouse infection models. Renal pharmacokinetics, biomarkers for toxicity, and kidney histopathology in adult rats all indicated a significantly lower nephrotoxicity of EBL-1003 than of gentamicin. Interpretation: This study provides preclinical proof-of-concept for the efficacy of EBL-1003 in cUTI/AP. Similar efficacy but lower nephrotoxicity of EBL-1003 in comparison to gentamicin may thus translate into a higher safety margin and a wider therapeutic window in the treatment of cUTI/API. Funding: A full list of funding bodies that contributed to this study can be found in the Acknowledgements section. © 2021 The Author(s)
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2.
  • Becker, K., et al. (författare)
  • Efficacy of EBL-1003 (apramycin) against Acinetobacter baumannii lung infections in mice
  • 2021
  • Ingår i: Clinical Microbiology and Infection. - : Elsevier B.V.. - 1198-743X .- 1469-0691. ; 27:9, s. 1315-
  • Tidskriftsartikel (refereegranskat)abstract
    • Objectives: Novel therapeutics are urgently required for the treatment of carbapenem-resistant Acinetobacter baumannii (CRAB) causing critical infections with high mortality. Here we assessed the therapeutic potential of the clinical-stage drug candidate EBL-1003 (crystalline free base of apramycin) in the treatment of CRAB lung infections. Methods: The genotypic and phenotypic susceptibility of CRAB clinical isolates to aminoglycosides and colistin was assessed by database mining and broth microdilution. The therapeutic potential was assessed by target attainment simulations on the basis of time–kill kinetics, a murine lung infection model, comparative pharmacokinetic analysis in plasma, epithelial lining fluid (ELF) and lung tissue, and pharmacokinetic/pharmacodynamic (PKPD) modelling. Results: Resistance gene annotations of 5451 CRAB genomes deposited in the National Database of Antibiotic Resistant Organisms (NDARO) suggested >99.9% of genotypic susceptibility to apramycin. Low susceptibility to standard-of-care aminoglycosides and high susceptibility to EBL-1003 were confirmed by antimicrobial susceptibility testing of 100 A. baumannii isolates. Time–kill experiments and a mouse lung infection model with the extremely drug-resistant CRAB strain AR Bank #0282 resulted in rapid 4-log CFU reduction both in vitro and in vivo. A single dose of 125 mg/kg EBL-1003 in CRAB-infected mice resulted in an AUC of 339 h × μg/mL in plasma and 299 h × μg/mL in ELF, suggesting a lung penetration of 88%. PKPD simulations suggested a previously predicted dose of 30 mg/kg in patients (creatinine clearance (CLCr) = 80 mL/min) to result in >99% probability of –2 log target attainment for MICs up to 16 μg/mL. Conclusions: This study provides proof of concept for the efficacy of EBL-1003 in the treatment of CRAB lung infections. Broad in vitro coverage, rapid killing, potent in vivo efficacy, and a high probability of target attainment render EBL-1003 a strong therapeutic candidate for a priority pathogen for which treatment options are very limited. © 2020 The Author(s)
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3.
  • Nilsson, C. L., et al. (författare)
  • Use of ENCODE Resources to Characterize Novel Proteoforms and Missing Proteins in the Human Proteome
  • 2015
  • Ingår i: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 14:2, s. 603-608
  • Tidskriftsartikel (refereegranskat)abstract
    • We describe the utility of integrated strategies that employ both translation of ENCODE data and major proteomic technology pillars to improve the identification of the "missing proteins", novel proteoforms, and PTMs. On one hand, databases in combination with bioinformatic tools are efficiently utilized to establish microarray-based transcript analysis and supply rapid protein identifications in clinical samples. On the other hand, sequence libraries are the foundation of targeted protein identification and quantification using mass spectrometric and immunoaffinity techniques. The results from combining proteoENCODEdb searches with experimental mass spectral data indicate that some alternative splicing forms detected at the transcript level are in fact translated to proteins. Our results provide a step toward the directives of the C-HPP initiative and related biomedical research.
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4.
  • Dahlin, Andreas P., et al. (författare)
  • Poly(dimethylsiloxane)-Based Microchip for Two-Dimensional Solid-Phase Extraction-Capillary Electrophoresis with an Integrated Electrospray Emitter Tip
  • 2005
  • Ingår i: Analytical Chemistry. - : American Chemical Society (ACS). - 0003-2700 .- 1520-6882. ; 77:16, s. 5356-5363
  • Tidskriftsartikel (refereegranskat)abstract
    • A microchip in poly(dimethylsiloxane) (PDMS) for in-line solid-phase extraction-capillary electrophoresis-electrospray ionization-time-of-flight mass spectrometry (SPE-CE-ESI-TOF-MS) has been developed and evaluated. The chip was fabricated in a novel one-step procedure where mixed PDMS was cast over steel wires in a mold. The removed wires defined 50-um cylindrical channels. Fused-silica capillaries were inserted into the structure in a tight fit connection. The inner walls of the inserted fused-silica capillaries and the PDMS microchip channels were modified with a positively charged polymer, PolyE-323. The chip was fabricated in a two-level cross design. The channel at the lower level was packed with 5-um hyper-cross-linked polystyrene beads acting as a SPE medium used for desalting. The upper level channel acted as a CE channel and ended in an integrated emitter tip coated with conducting graphite powder to facilitate the electrical contact for sheathless ESI. An overpressure continuously provided fresh CE electrolyte independently of the flows in the different levels. Further studies were carried out in order to investigate the electrophoretic and flow rate properties of the chip. Finally, six-peptide mixtures, in different concentrations, dissolved in physiological salt solution was injected, desalted, separated, and sprayed into the mass spectrometer for analysis with a limit of detection in femtomole levels.
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5.
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6.
  • Fridjonsdottir, Elva, et al. (författare)
  • Mass spectrometry imaging identifies abnormally elevated brain L-DOPA levels and extrastriatal monoaminergic dysregulation in L-DOPA-induced dyskinesia
  • 2021
  • Ingår i: Science Advances. - : American Association for the Advancement of Science (AAAS). - 2375-2548. ; 7:2
  • Tidskriftsartikel (refereegranskat)abstract
    • L-DOPA treatment for Parkinson's disease frequently leads to dyskinesias, the pathophysiology of which is poorly understood. We used MALDI-MSI to map the distribution of L-DOPA and monoaminergic pathways in brains of dyskinetic and nondyskinetic primates. We report elevated levels of L-DOPA, and its metabolite 3-O-methyldopa, in all measured brain regions of dyskinetic animals and increases in dopamine and metabolites in all regions analyzed except the striatum. In dyskinesia, dopamine levels correlated well with L-DOPA levels in extrastriatal regions, such as hippocampus, amygdala, bed nucleus of the stria terminalis, and cortical areas, but not in the striatum. Our results demonstrate that L-DOPA-induced dyskinesia is linked to a dysregulation of L-DOPA metabolism throughout the brain. The inability of extrastriatal brain areas to regulate the formation of dopamine during L-DOPA treatment introduces the potential of dopamine or even L-DOPA itself to modulate neuronal signaling widely across the brain, resulting in unwanted side effects.
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7.
  • Fridjonsdottir, Elva, et al. (författare)
  • Mass spectrometry imaging reveals brain-region specific changes in metabolism and acetylcholine levels in experimental Parkinson’s disease and L-DOPA-induced dyskinesia
  • Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
    • There is evidence that cholinergic alterations are linked to various motor and non-motor symptoms of Parkinson’s disease. We therefore used mass spectrometry imaging to investigate regional changes in acetylcholine abundance in the brain of a non-human primate model of Parkinson’s disease (PD) and L-DOPA-induced dyskinesia (LID). We also present an experimental design for performing untargeted analysis using MALDI-MSI with multiple experiments incorporating quality control samples to monitor experimental variability. We observed that MPTP treatment (i) led to reductions in putaminal acetylcholine levels that persisted after L-DOPA treatment and (ii) appeared to induce a shift of choline metabolism from α-glycerophosphocholine towards betaine. LID animals exhibited reduced levels of various metabolites important for brain homeostasis including S-adenosylmethionine, glutathione, adenosine monophosphate, and acylcarnitines. The vasculature marker heme B was upregulated in the putamen of LID animals, suggesting increased blood-flow in the dyskinetic putamen. These results provide new insights into pathological choline-related metabolic changes in PD and LID.  
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8.
  • Fridjonsdottir, Elva, et al. (författare)
  • Region-Specific and Age-Dependent Multitarget Effects of Acetylcholinesterase Inhibitor Tacrine on Comprehensive Neurotransmitter Systems
  • 2022
  • Ingår i: ACS Chemical Biology. - : American Chemical Society (ACS). - 1554-8929 .- 1554-8937. ; 17:1, s. 147-158
  • Tidskriftsartikel (refereegranskat)abstract
    • Regional brain distribution and metabolism of neurotransmitters and their response to drug treatment are fundamentally important for understanding the central effects of neuroactive substances. We used matrix-assisted laser desorption/ionization mass spectrometry imaging in combination with multivariate analysis to visualize in anatomical detail metabolic effects of aging and tacrine-mediated acetylcholinesterase inhibition on comprehensive neurotransmitter systems in multiple mouse brain regions of 12-week-old and 14-month-old mice. We detected age-related increases in 3,4-dihydroxyphenylacetaldehyde and histamine, indicating oxidative stress and aging deficits in astrocytes. Tacrine had a significant impact on the metabolism of neurotransmitters in both age groups; predominantly, there was an increased norepinephrine turnover throughout the brain and decreased 3-methoxy tyramine, a marker for dopamine release, in the striatum. The striatal levels of histamine were only elevated after tacrine administration in the older animals. Our results demonstrated that tacrine is a multitarget and region-specific neuroactive agent, inducing age-specific responses. Although well-studied, the complete mechanisms of the action of tacrine are not fully understood, and the current findings reveal features that may help explain its treatment-related effectiveness and central side effects.
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9.
  • He, Yachao, et al. (författare)
  • Prosaposin maintains lipid homeostasis in dopamine neurons and counteracts experimental parkinsonism in rodents
  • 2023
  • Ingår i: Nature Communications. - : Springer Nature. - 2041-1723. ; 14
  • Tidskriftsartikel (refereegranskat)abstract
    • Prosaposin (PSAP) modulates glycosphingolipid metabolism and variants have been linked to Parkinson's disease (PD). Here, we find altered PSAP levels in the plasma, CSF and post-mortem brain of PD patients. Altered plasma and CSF PSAP levels correlatewith PD-relatedmotor impairments. Dopaminergic PSAP-deficient (cPSAP(DAT)) mice display hypolocomotion and depression/anxiety-like symptoms with mildly impaired dopaminergic neurotransmission, while serotonergic PSAP-deficient (cPSAP(SERT)) mice behave normally. Spatial lipidomics revealed an accumulation of highly unsaturated and shortened lipids and reduction of sphingolipids throughout the brains of cPSAP(DAT) mice. The overexpression of alpha-synuclein via AAV lead to more severe dopaminergic degeneration and higher p-Ser129 alpha-synuclein levels in cPSAP(DAT) mice compared to WT mice. Overexpression of PSAP via AAV and encapsulated cell biodelivery protected against 6-OHDA and alpha-synuclein toxicity in wild-type rodents. Thus, these findings suggest PSAP may maintain dopaminergic lipid homeostasis, which is dysregulated in PD, and counteract experimental parkinsonism.
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10.
  • Hulme, Heather E., et al. (författare)
  • Mass spectrometry imaging identifies palmitoylcarnitine as an immunological mediator during Salmonella Typhimurium infection
  • 2017
  • Ingår i: Scientific Reports. - : Nature Publishing Group. - 2045-2322. ; 7
  • Tidskriftsartikel (refereegranskat)abstract
    • Salmonella Typhimurium causes a self-limiting gastroenteritis that may lead to systemic disease. Bacteria invade the small intestine, crossing the intestinal epithelium from where they are transported to the mesenteric lymph nodes (MLNs) within migrating immune cells. MLNs are an important site at which the innate and adaptive immune responses converge but their architecture and function is severely disrupted during S. Typhimurium infection. To further understand host-pathogen interactions at this site, we used mass spectrometry imaging (MSI) to analyse MLN tissue from a murine model of S. Typhimurium infection. A molecule, identified as palmitoylcarnitine (PalC), was of particular interest due to its high abundance at loci of S. Typhimurium infection and MLN disruption. High levels of PalC localised to sites within the MLNs where B and T cells were absent and where the perimeter of CD169(+) sub capsular sinus macrophages was disrupted. MLN cells cultured ex vivo and treated with PalC had reduced CD4(+) CD25(+) T cells and an increased number of B220(+) CD19(+) B cells. The reduction in CD4(+) CD25(+) T cells was likely due to apoptosis driven by increased caspase-3/7 activity. These data indicate that PalC significantly alters the host response in the MLNs, acting as a decisive factor in infection outcome.
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11.
  • Hulme, Heather, et al. (författare)
  • Simultaneous mass spectrometry imaging of multiple neuropeptides in the brain and alterations induced by experimental parkinsonism and L-DOPA therapy
  • 2020
  • Ingår i: Neurobiology of Disease. - : ACADEMIC PRESS INC ELSEVIER SCIENCE. - 0969-9961 .- 1095-953X. ; 137
  • Tidskriftsartikel (refereegranskat)abstract
    • Neuropeptides are important signalling molecules in the brain and alterations in their expression levels have been linked to neurological disorders such as Parkinson's disease. It is challenging to map neuropeptide changes across and within brain regions because of their low in vivo concentrations and complex post-translational processing. Consequently, the role of neuropeptides in Parkinson's disease is not well understood. Thus, we have developed and evaluated a method to image multiple neuropeptides simultaneously in both rat and primate brain tissue sections by matrix-assisted laser desorption/ionisation mass spectrometry imaging at high lateral resolution. Using a unilateral 6-hydroxydopamine rat model of Parkinson's disease, we imaged changes in enkephalins, dynorphins, tachykinins and neurotensin associated with the dopaminergic denervation and L-DOPA treatment in multiple brain regions. L-DOPA administration significantly affected neuropeptides in the globus pallidus, while neuropeptides in the caudate-putamen were mostly affected by dopamine depletion. Using high lateral resolution imaging, we observed an increase of neurotensin in the dorsal sub-region of the globus pallidus after dopamine depletion. This study highlights the capacity of mass spectrometry imaging to elucidate the dynamics of neuropeptide signalling during Parkinson's disease and its treatment.
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12.
  • Kaplan, Anders, et al. (författare)
  • An Automated Method for Scanning LC−MS Data Sets for Significant Peptides and Proteins, Including Quantitative Profiling and Interactive Confirmation : An Automated Method for Scanning LC−MS Data Sets for Significant Peptides and Proteins, Including Quantitative Profiling and Interactive Confirmation
  • 2007
  • Ingår i: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 6:7, s. 2888-2895
  • Tidskriftsartikel (refereegranskat)abstract
    • Differential quantification of proteins and peptides by LC-MS is a promising method to acquire knowledge about biological processes, and for finding drug targets and biomarkers. However, differential protein analysis using LC-MS has been held back by the lack of suitable software tools. Large amounts of experimental data are easily generated in protein and peptide profiling experiments, but data analysis is time-consuming and labor-intensive. Here, we present a fully automated method for scanning LC-MS/MS data for biologically significant peptides and proteins, including support for interactive confirmation and further profiling. By studying peptide mixtures of known composition, we demonstrate that peptides present in different amounts in different groups of samples can be automatically screened for using statistical tests. A linear response can be obtained over almost 3 orders of magnitude, facilitating further profiling of peptides and proteins of interest. Furthermore, we apply the method to study the changes of endogenous peptide levels in mouse brain striatum after administration of reserpine, a classical model drug for inducing Parkinson disease symptoms.
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13.
  • Kaya, Ibrahim, et al. (författare)
  • On-Tissue Chemical Derivatization for Comprehensive Mapping of Brain Carboxyl and Aldehyde Metabolites by MALDI-MS Imaging
  • 2023
  • Ingår i: Journal of the American Society for Mass Spectrometry. - : American Chemical Society (ACS). - 1044-0305 .- 1879-1123. ; 34:5, s. 836-846
  • Tidskriftsartikel (refereegranskat)abstract
    • The visualization of small metabolites by MALDI mass spectrometry imaging in brain tissue sections is challenging due to low detection sensitivity and high background interference. We present an on-tissue chemical derivatization MALDI mass spectrometry imaging approach for the comprehensive mapping of carboxyls and aldehydes in brain tissue sections. In this approach, the AMPP (1-(4-(aminomethyl)phenyl)pyridin-1-ium chloride) derivatization reagent is used for the covalent charge-tagging of molecules containing carboxylic acid (in the presence of peptide coupling reagents) and aldehydes. This includes free fatty acids and the associated metabolites, fatty aldehydes, dipeptides, neurotoxic reactive aldehydes, amino acids, neurotransmitters and associated metabolites, as well as tricarboxylic acid cycle metabolites. We performed sensitive ultrahigh mass resolution MALDI-MS detection and imaging of various carboxyl-and aldehyde containing endogenous metabolites simultaneously in rodent brain tissue sections. We verified the AMPP-derivatized metabolites by tandem MS for structural elucidation. This approach allowed us to image numerous aldehydes and carboxyls, including certain metabolites which had been undetectable in brain tissue sections. We also demonstrated the application of on-tissue derivatization to carboxyls and aldehydes in coronal brain tissue sections of a nonhuman primate Parkinson's disease model. Our methodology provides a powerful tool for the sensitive, simultaneous spatial molecular imaging of numerous aldehydes and carboxylic acids during pathological states, including neurodegeneration, in brain tissue.
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14.
  • Kaya, Ibrahim, et al. (författare)
  • Spatial lipidomics reveals brain region-specific changes of sulfatides in an experimental MPTP Parkinson's disease primate model
  • 2023
  • Ingår i: NPJ PARKINSONS DISEASE. - : Springer Nature. - 2373-8057. ; 9:1
  • Tidskriftsartikel (refereegranskat)abstract
    • Metabolism of MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) to the neurotoxin MPP+ in the brain causes permanent Parkinson's disease-like symptoms by destroying dopaminergic neurons in the pars compacta of the substantia nigra in humans and non-human primates. However, the complete molecular pathology underlying MPTP-induced parkinsonism remains poorly understood. We used dual polarity matrix-assisted laser desorption/ionization mass spectrometry imaging to thoroughly image numerous glycerophospholipids and sphingolipids in coronal brain tissue sections of MPTP-lesioned and control non-human primate brains (Macaca mulatta). The results revealed specific distributions of several sulfatide lipid molecules based on chain-length, number of double bonds, and importantly, hydroxylation stage. More specifically, certain long-chain hydroxylated sulfatides with polyunsaturated chains in the molecular structure were depleted within motor-related brain regions in the MPTP-lesioned animals, e.g., external and internal segments of globus pallidus and substantia nigra pars reticulata. In contrast, certain long-chain non-hydroxylated sulfatides were found to be elevated within the same brain regions. These findings demonstrate region-specific dysregulation of sulfatide metabolism within the MPTP-lesioned macaque brain. The depletion of long-chain hydroxylated sulfatides in the MPTP-induced pathology indicates oxidative stress and oligodendrocyte/myelin damage within the pathologically relevant brain regions. Hence, the presented findings improve our current understanding of the molecular pathology of MPTP-induced parkinsonism within primate brains, and provide a basis for further research regarding the role of dysregulated sulfatide metabolism in PD.
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15.
  • Källback, Patrik, et al. (författare)
  • Cross-validated Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry Imaging Quantitation Protocol for a Pharmaceutical Drug and Its Drug-Target Effects in the Brain Using Time-of-Flight and Fourier Transform Ion Cyclotron Resonance Analyzers
  • 2020
  • Ingår i: Analytical Chemistry. - : American Chemical Society (ACS). - 0003-2700 .- 1520-6882. ; 92:21, s. 14676-14684
  • Tidskriftsartikel (refereegranskat)abstract
    • Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) is an established tool in drug development, which enables visualization of drugs and drug metabolites at spatial localizations in tissue sections from different organs. However, robust and accurate quantitation by MALDI-MSI still remains a challenge. We present a quantitative MALDI-MSI method using two instruments with different types of mass analyzers, i.e., time-of-flight (TOF) and Fourier transform ion cyclotron resonance (FTICR) MS, for mapping levels of the in vivo-administered drug citalopram, a selective serotonin reuptake inhibitor, in mouse brain tissue sections. Six different methods for applying calibration standards and an internal standard were evaluated. The optimized method was validated according to authorities' guidelines and requirements, including selectivity, accuracy, precision, recovery, calibration curve, sensitivity, reproducibility, and stability parameters. We showed that applying a dilution series of calibration standards followed by a homogeneously applied, stable, isotopically labeled standard for normalization and a matrix on top of the tissue section yielded similar results to those from the reference method using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The validation results were within specified limits and the brain concentrations for TOF MS (51.1 +/- 4.4 pmol/mg) and FTICR MS (56.9 +/- 6.0 pmol/mg) did not significantly differ from those of the cross-validated LC-MS/MS method (55.0 +/- 4.9 pmol/mg). The effect of in vivo citalopram administration on the serotonin neurotransmitter system was studied in the hippocampus, a brain region that is the principal target of the serotonergic afferents along with the limbic system, and it was shown that serotonin was significantly increased (2-fold), but its metabolite 5-hydroxyindoleacetic acid was not. This study makes a substantial step toward establishing MALDI-MSI as a fully quantitative validated method.
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16.
  • Källback, Patrik (författare)
  • Development and Application of Software Tools for Mass Spectrometry Imaging
  • 2017
  • Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
    • Mass spectrometry imaging (MSI) has been extensively used to produce qualitative maps of distributions of proteins, peptides, lipids, neurotransmitters, small molecule pharmaceuticals and their metabolites directly in biological tissue sections. Moreover, during the last 10 years, there has been growing demand to quantify target compounds in tissue sections of various organs. This thesis focuses on development and application of a novel instrument- and manufacturer-independent MSI software suite, msIQuant, in the open access format imzML, which has been developed specifically for quantitative analysis of MSI data. The functionality of msIQuant facilitates automatic generation of calibration curves from series of standards that can be used to determine concentrations of specific analytes. In addition, it provides many tools for image visualization, including modules enabling multiple interpolation, low intensity transparency display, and image fusion and sharpening. Moreover, algorithms and advanced data management modules in msIQuant facilitate management of the large datasets generated following rapid recent increases in the mass and spatial resolutions of MSI instruments, by using spectra transposition and data entropy reduction (at four selectable levels: coarse, medium, fine or superfine) before lossless compression of the data. As described in the thesis, implementation of msIQuant has been exemplified in both quantitative (relative or absolute) and qualitative analyses of distributions of neurotransmitters, endogenous substances and pharmaceutical drugs in brain tissue sections. Our laboratory have developed a molecular-specific approach for the simultaneous imaging and quantitation of multiple neurotransmitters, precursors, and metabolites, such as tyrosine, tryptamine, tyramine, phenethylamine, dopamine, 3-methoxytyramine, serotonin, gamma-aminobutyric acid (GABA), and acetylcholine, in histological tissue sections at high spatial resolution by matrix-assisted laser desorption ionization (MALDI) and desorption electrospray ionization (DESI) MSI. Chemical derivatization by charge-tagging primary amines of analytes significantly increased the sensitivity, enabling mapping of neurotransmitters that were not previously detectable by MSI. The two MSI approaches have been used to directly measure changes in neurotransmitter levels in specific brain structures in animal disease models, which facilitates understanding of biochemical mechanisms of drug treatments. In summary, msIQuant software has proven potency (particularly in combination with the reported derivatization technique) for both qualitative and quantitative analyses. Further developments will enable its implementation in multiple operating system platforms and use for statistical analysis.
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17.
  • Madeira, Alexandra, et al. (författare)
  • Caveolin-1 interacts with alpha-synuclein and mediates toxic actions of cellular alpha-synuclein overexpression
  • 2011
  • Ingår i: Neurochemistry International. - : Elsevier BV. - 0197-0186 .- 1872-9754. ; 59:2, s. 280-289
  • Tidskriftsartikel (refereegranskat)abstract
    • Caveolin-1 (Cav-1) is a transmembrane protein which clusters proteins and lipids at the cell membrane into a subclass of lipid rafts named caveolae. To increase our understanding about putative functions of Cav-1 in neuronal cells, we used mouse brain extracts and a novel technology coupling surface plasmon resonance to mass spectrometry to find binding partners to Cav-1. An interaction between Cav-1 and alpha-synclein was found and confirmed in reciprocal pulldown experiments. Genetic overexpression of alpha-synclein in mouse neuroblastoma Neuro2A cells (N2A) expectedly decreased cell survival, but also significantly increased the levels of Cav-1. Furthermore, si-RNA-mediated knockdown of Cav-1 counteracted cell death induced by overexpression of alpha-synuclein. We also used an inhibitor of proteasome (MG132) to induce cell death in a Parkinson's disease context. Cav-1 knockdown had no effect on cell death induced by MG132. Conversely, treating the cells with mevastatin, an inhibitor of cholesterol synthesis, inhibits cell death induced by MG132, but not by alpha synuclein overexpression. It can be concluded that Cav-1 may play a functional role in neuronal cells by virtue of its physical interaction with alpha-synuclein and regulate alpha synuclein-mediated actions on cell death, processes known to be involved in synucleinopathies including Parkinson's disease.
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18.
  • Madeira, Alexandra, et al. (författare)
  • Coupling surface plasmon resonance to mass spectrometry to discover novel protein-protein interactions
  • 2009
  • Ingår i: Nature Protocols. - : Springer Science and Business Media LLC. - 1754-2189 .- 1750-2799. ; 4:7, s. 1023-1037
  • Tidskriftsartikel (refereegranskat)abstract
    • The elucidation of protein-protein interaction networks is a crucial task in the postgenomic era. In this protocol, we describe our approach to discover protein-protein interactions using the surface plasmon resonance technique coupled to mass spectrometry ( MS). A peptide or a protein is immobilized on a sensor chip and then exposed to brain extracts injected through the surface of the chip by a microfluidic system. The interactions between the immobilized ligand and the extracts can be monitored in real time. Proteins interacting with the peptide/protein are recovered, trypsinated and identified using MS. The data obtained are searched against a sequence database using the Mascot 2.1 software. Control experiments using blank sensor chips and/or randomized peptides are carried out to exclude nonspecific interactors. The protocol can be carried out in < 3 days. Other methods, such as yeast two-hybrid systems or pull-down approaches followed by MS, are widely used to screen protein-protein interactions. However, as the yeast two-hybrid system requires protein interactions in the nucleus of yeast, proteins that are abundant in other compartments may not be detected. Pull-down approaches based on immunoprecipitation can be used to study endogenous proteins but they require specific antibodies. The protocol presented here does not require the specific labeling or modification of proteins.
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19.
  • Madeira, Alexandra, et al. (författare)
  • Identification of protein-protein interactions by surface plasmon resonance followed by mass spectrometry
  • 2011
  • Ingår i: Current protocols in protein science. - Hoboken, NJ, USA : John Wiley & Sons, Inc.. - 1934-3663. ; Suppl. 65, s. 19.21.1-19.21.9
  • Tidskriftsartikel (refereegranskat)abstract
    • Elucidation of the function and meaning of the protein networks can be useful in the understanding of many pathological processes and the identification of new therapeutic targets. This unit describes an approach to discover protein-protein interactions by coupling surface plasmon resonance to mass spectrometry. Briefly, a protein is covalently bound to a sensor chip, which is then exposed to brain extracts injected over the surface via a microfluidic system. This allows the monitoring in real-time of the interactions between the immobilized ligand and the extracts. Interacting proteins from the extracts are then recovered, trypsinized, and identified using mass spectrometry. The data obtained are searched against a sequence database using the Mascot software. To exclude nonspecific interactors, control experiments using blank sensor chips, and/or randomized peptides, are performed. The protocol presented here does not require specific labeling or modification of proteins and can be performed in <4 days.
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20.
  • Mantas, Ioannis, et al. (författare)
  • TAAR1-Dependent and-Independent Actions of Tyramine in Interaction With Glutamate Underlie Central Effects of Monoamine Oxidase Inhibition
  • 2021
  • Ingår i: Biological Psychiatry. - : Elsevier. - 0006-3223 .- 1873-2402. ; 90:1, s. 16-27
  • Tidskriftsartikel (refereegranskat)abstract
    • BACKGROUND: Monoamine oxidase inhibitors (MAOIs) exert therapeutic actions by elevating extracellular levels of monoamines in the brain. Irreversible MAOIs cause serious hypertensive crises owing to peripheral accumulation of tyramine, but the role of tyramine in the central effects of MAOIs remains elusive, an issue addressed herein. To achieve robust inhibition of MAOA/B, the clinically used antidepressant tranylcypromine (TCP) was employed. METHODS: Behavioral, histological, mass spectrometry imaging, and biosensor-mediated measures of glutamate were conducted with MAOIs in wild-type and TAAR1-knockout (KO) mice. RESULTS: Both antidepressant and locomotion responses to TCP were enhanced in TAAR1-KO mice. A recently developed fluoromethylpyridinium-based mass spectrometry imaging method revealed robust accumulation of striatal tyramine on TCP administration. Furthermore, tyramine accumulation was higher in TAAR1-KO versus wild type mice, suggesting a negative feedback mechanism for TAAR1 in sensing tyramine levels. Combined histoenzymological and immunohistological studies revealed hitherto unknown TAAR1 localization in brain areas projecting to the substantia nigra/ventral tegmental area. Using an enzyme-based biosensor technology, we found that both TCP and tyramine reduced glutamate release in the substantia nigra in wild-type but not in TAAR1-KO mice. Moreover, glutamate measures in freely moving animals treated with TCP demonstrated that TAAR1 prevents glutamate accumulation in the substantia nigra during hyperlocomotive states. CONCLUSIONS: These observations suggest that tyramine, in interaction with glutamate, is involved in centrally mediated behavioral, transcriptional, and neurochemical effects of MAOIs.
  •  
21.
  •  
22.
  • Nilsson, Anna, et al. (författare)
  • Fine Mapping the Spatial Distribution and Concentration of Unlabeled Drugs within Tissue Micro-Compartments Using Imaging Mass Spectrometry
  • 2010
  • Ingår i: PloS one. - : Public Library of Science (PLoS). - 1932-6203. ; 5:7, s. e11411-
  • Tidskriftsartikel (refereegranskat)abstract
    • Readouts that define the physiological distributions of drugs in tissues are an unmet challenge and at best imprecise, but are needed in order to understand both the pharmacokinetic and pharmacodynamic properties associated with efficacy. Here we demonstrate that it is feasible to follow the in vivo transport of unlabeled drugs within specific organ and tissue compartments on a platform that applies MALDI imaging mass spectrometry to tissue sections characterized with high definition histology. We have tracked and quantified the distribution of an inhaled reference compound, tiotropium, within the lungs of dosed rats, using systematic point by point MS and MS/MS sampling at 200 mu m intervals. By comparing drug ion distribution patterns in adjacent tissue sections, we observed that within 15 min following exposure, tiotropium parent MS ions (mass-to-charge; m/z 392.1) and fragmented daughter MS/MS ions (m/z 170.1 and 152.1) were dispersed in a concentration gradient (80 fmol-5 pmol) away from the central airways into the lung parenchyma and pleura. These drug levels agreed well with amounts detected in lung compartments by chemical extraction. Moreover, the simultaneous global definition of molecular ion signatures localized within 2-D tissue space provides accurate assignment of ion identities within histological landmarks, providing context to dynamic biological processes occurring at sites of drug presence. Our results highlight an important emerging technology allowing specific high resolution identification of unlabeled drugs at sites of in vivo uptake and retention.
  •  
23.
  • Nilsson, Anna, et al. (författare)
  • In Situ Mass Spectrometry Imaging and Ex Vivo Characterization of Renal Crystalline Deposits Induced in Multiple Preclinical Drug Toxicology Studies
  • 2012
  • Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 7:10, s. e47353-
  • Tidskriftsartikel (refereegranskat)abstract
    • Drug toxicity observed in animal studies during drug development accounts for the discontinuation of many drug candidates, with the kidney being a major site of tissue damage. Extensive investigations are often required to reveal the mechanisms underlying such toxicological events and in the case of crystalline deposits the chemical composition can be problematic to determine. In the present study, we have used mass spectrometry imaging combined with a set of advanced analytical techniques to characterize such crystalline deposits in situ. Two potential microsomal prostaglandin E synthase 1 inhibitors, with similar chemical structure, were administered to rats over a seven day period. This resulted in kidney damage with marked tubular degeneration/regeneration and crystal deposits within the tissue that was detected by histopathology. Results from direct tissue section analysis by matrix-assisted laser desorption ionization mass spectrometry imaging were combined with data obtained following manual crystal dissection analyzed by liquid chromatography mass spectrometry and nuclear magnetic resonance spectroscopy. The chemical composition of the crystal deposits was successfully identified as a common metabolite, bisulphonamide, of the two drug candidates. In addition, an un-targeted analysis revealed molecular changes in the kidney that were specifically associated with the area of the tissue defined as pathologically damaged. In the presented study, we show the usefulness of combining mass spectrometry imaging with an array of powerful analytical tools to solve complex toxicological problems occurring during drug development.
  •  
24.
  • Nilsson, Anna, et al. (författare)
  • Increased striatal mRNA and protein levels of the immunophilin FKBP-12 in experimental Parkinson’s Disease and identification of FKBP-12-binding proteins
  • 2007
  • Ingår i: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 6:10, s. 3952-3961
  • Tidskriftsartikel (refereegranskat)abstract
    • FKBP-12, a 12 kDa FK506-binding protein (neuroimmunophilin), acts as a receptor for the immunosuppressant drug FK506. Neuroimmunophilins, including FKBP-12, are abundant in the brain and have been shown to be involved in reversing neuronal degeneration and preventing cell death. In this report, we have utilized several analytical techniques, such as in situ hybridization, Western blotting, two-dimensional gel electrophoresis, and liquid chromatography electrospray tandem mass spectrometry to study the transcriptional expression as well as protein levels of FKBP-12 in the unilateral 6-hydroxydopamine (6-OHDA) rat model of Parkinson's disease. The FKBP-12 protein was also detected directly on brain tissue sections using mass spectrometry profiling. We found increased levels of FKBP-12 mRNA and protein in the dorsal and middle part of the 6-OHDA lesioned striatum. Thus, these studies clearly demonstrate that FKBP-12 is increased in the brain of a common animal model of Parkinson's disease (PD). Additionally, we have identified potential binding partners to FKBP-12 that may be implicated in the pathophysiology of Parkinson's disease, such as alpha-enolase, 14-3-3 zeta/delta, pyruvate kinase isozymes, and heat shock protein 70, using surface plasmon resonance sensor technology in combination with mass spectrometry. In conclusion, these data strongly suggests that FKBP-12 is altered in an experimental model of PD.
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25.
  • Nilsson, Anna, et al. (författare)
  • Neuropeptidomics of mouse hypothalamus after imipramine treatment reveal somatostatin as a potential mediator of antidepressant effects
  • 2012
  • Ingår i: Neuropharmacology. - : Elsevier BV. - 0028-3908 .- 1873-7064. ; 62:1, s. 347-357
  • Tidskriftsartikel (refereegranskat)abstract
    • Excessive activation of the hypothalamic pituitary adrenal (HPA) axis has been associated with numerous diseases, including depression, and the tricyclic antidepressant imipramine has been shown to suppress activity of the HPA axis. Central hypothalamic control of the HPA axis is complex and involves a number of neuropeptides released from multiple hypothalamic subnuclei. The present study was therefore designed to determine the effects of imipramine administration on the mouse hypothalamus using a peptidomics approach. Among the factors found to be downregulated after acute (one day) or chronic (21 days) imipramine administration were peptides derived from secretogranin 1 (chromogranin B) as well as peptides derived from cerebellin precursors. In contrast, peptides SRIF-14 and SRIF-28 (1-11) derived from somatostatin (SRIF, somatotropin release inhibiting factor) were significantly upregulated by imipramine in the hypothalamus. Because diminished SRIF levels have long been known to occur in depression, a second part of the study investigated the roles of individual SRIF receptors in mediating potential antidepressant effects. SRA880, an antagonist of the somatostatin-1 autoreceptor (sst1) which positively modulates release of endogenous SRIF, was found to synergize with imipramine in causing antidepressant-like effects in the tail suspension test. Furthermore, chronic co-administration of SRA880 and imipramine synergistically increased BDNF mRNA expression in the cerebral cortex. Application of SRIF or L054264, an sst2 receptor agonist, but not 1,803807, an sst4 receptor agonist, increased phosphorylation of CaMKII and GluR1 in cerebrocortical slices. Our present experiments thus provide evidence for antidepressant-induced upregulation of SRIF in the brain, and strengthen the notion that augmented SRIF expression and signaling may counter depressive-like symptoms.
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26.
  • Nilsson, Anna, et al. (författare)
  • Striatal alterations of secretogranin-1, somatostatin, prodynorphin and cholecystokinin peptides in an experimental mouse model of Parkinson’s disease
  • 2009
  • Ingår i: Molecular & Cellular Proteomics. - 1535-9476 .- 1535-9484. ; 8:5, s. 1094-1104
  • Tidskriftsartikel (refereegranskat)abstract
    • The principal causative pathology of Parkinson disease is the progressive degeneration of dopaminergic neurons in the substantia nigra pars compacta projecting to the striatum in the brain. The information regarding the expression of neuropeptides in parkinsonism is very limited. Here we have elucidated striatal neuropeptide mechanisms in experimental parkinsonism using the unilateral  6-hydroxydopamine model to degenerate dopamine neurons. A thoroughly controlled sample preparation technique together with a peptidomics approach and targeted neuropeptide sequence collections enabled sensitive detection, identification, and relative quantitation of a great number of endogenous neuropeptides. Previously not recognized alterations in neuropeptide levels were identified in the unilateral   lesioned mice with or without subchronic 3,4-dihydroxy-L-phenylalanine   administration, the conventional treatment of Parkinson disease. Several of these peptides originated from the same precursor such as secretogranin-1, somatostatin, prodynorphin, and cholecystokinin. Disease-related biotransformation of precursors into individual   peptides was observed in the experimental model of Parkinson disease. Several previously unreported potentially biologically active peptides were also identified from the striatal samples. This study provides further evidence that neuropeptides take part in mediating the central nervous system failure associated with Parkinson disease.
  •  
27.
  • Nilsson, C. L., et al. (författare)
  • Chromosome 19 Annotations with Disease Speciation: A First Report from the Global Research Consortium
  • 2013
  • Ingår i: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 12:1, s. 134-149
  • Tidskriftsartikel (refereegranskat)abstract
    • A first research development progress report of the Chromosome 19 Consortium with members from Sweden, Norway, Spain, United States, China and India, a part of the Chromosome-centric Human Proteome Project (C-HPP) global initiative, is presented (http://www.c-hpp.org). From the chromosome 19 peptide-targeted library constituting 6159 peptides, a pilot study was conducted using a subset with 125 isotope-labeled peptides. We applied an annotation strategy with triple quadrupole, ESI-Qtrap, and MALDI mass spectrometry platforms, comparing the quality of data within and in between these instrumental set-ups. LC–MS conditions were outlined by multiplex assay developments, followed by MRM assay developments. SRM was applied to biobank samples, quantifying kallikrein 3 (prostate specific antigen) in plasma from prostate cancer patients. The antibody production has been initiated for more than 1200 genes from the entire chromosome 19, and the progress developments are presented. We developed a dedicated transcript microarray to serve as the mRNA identifier by screening cancer cell lines. NAPPA protein arrays were built to align with the transcript data with the Chromosome 19 NAPPA chip, dedicated to 90 proteins, as the first development delivery. We have introduced an IT-infrastructure utilizing a LIMS system that serves as the key interface for the research teams to share and explore data generated within the project. The cross-site data repository will form the basis for sample processing, including biological samples as well as patient samples from national Biobanks.
  •  
28.
  • Paslawski, Wojciech, et al. (författare)
  • alpha-synuclein-lipoprotein interactions and elevated ApoE level in cerebrospinal fluid from Parkinson's disease patients
  • 2019
  • Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : NATL ACAD SCIENCES. - 0027-8424 .- 1091-6490. ; 116:30, s. 15226-15235
  • Tidskriftsartikel (refereegranskat)abstract
    • The progressive accumulation, aggregation, and spread of alpha-synuclein (alpha SN) are common hallmarks of Parkinson's disease (PD) pathology. Moreover, numerous proteins interact with alpha SN species, influencing its toxicity in the brain. In the present study, we extended analyses of alpha SN-interacting proteins to cerebrospinal fluid (CSF). Using coimmunoprecipitation, followed by mass spectrometry, we found that alpha SN colocalize with apolipoproteins on lipoprotein vesicles. We confirmed these interactions using several methods, including the enrichment of lipoproteins with a recombinant alpha SN, and the subsequent uptake of prepared vesicles by human dopaminergic neuronal-like cells. Further, we report an increased level of ApoE in CSF from early PD patients compared with matched controls in 3 independent cohorts. Moreover, in contrast to controls, we observed the presence of ApoE-positive neuromelanin-containing dopaminergic neurons in substantia nigra of PD patients. In conclusion, the cooccurrence of alpha SN on lipoprotein vesicles, and their uptake by dopaminergic neurons along with an increase of ApoE in early PD, proposes a mechanism(s) for alpha SN spreading in the extracellular milieu of PD.
  •  
29.
  • Pierson, Johan, et al. (författare)
  • Increased Levels of Ubiquitin in the 6-OHDA-Lesioned Striatum of Rats
  • 2005
  • Ingår i: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 4:2, s. 223-226
  • Tidskriftsartikel (refereegranskat)abstract
    • Multiple genetic deficits have linked impaired ubiquitin-conjugation pathways to various forms of familiar Parkinson's disease. We therefore examined the possible role of 6-hydroxydopamine, a dopaminergic neurotoxin used in Parkinson's disease experimental models, in causing protein degradation and its association with the ubiquitin proteasome system. Using unilaterally 6-hydroxydopamine-denervated rats and mass spectrometry profiling directly on brain tissue sections, we here report for the first time an increased level of unconjugated ubiquitin specifically in the dorsal striatum of the dopamine depleted hemisphere. No similar changes were found in the intact hemisphere or in the ventral striatum of the dopamine depleted hemisphere. The lesioning of the dopamine innervation to the striatum was confirmed by a strongly reduced dopamine transporter binding in the striatum, indicating an abundant loss of dopamine neurons. These results suggest that denervation of dopamine neurons per se is implicated in the regulation of ubiquitin pathways, at least in a classical animal model of Parkinson's disease. This study adds additional information regarding the involvement of the ubiquitin-proteasome system in Parkinson's disease.
  •  
30.
  •  
31.
  • Schintu, Nicoletta, et al. (författare)
  • Non-dopaminergic Alterations in Depression-Like FSL Rats in Experimental Parkinsonism and L-DOPA Responses
  • 2020
  • Ingår i: Frontiers in Pharmacology. - : FRONTIERS MEDIA SA. - 1663-9812. ; 11
  • Tidskriftsartikel (refereegranskat)abstract
    • Depression is a common comorbid condition in Parkinson's disease (PD). Patients with depression have a two-fold increased risk to develop PD. Further, depression symptoms often precede motor symptoms in PD and are frequent at all stages of the disease. However, the influence of a depressive state on the responses to antiparkinson treatments is largely unknown. In this study, the genetically inbred depression-like flinders sensitive line (FSL) rats and control flinders resistant line (FRL) rats were studied in models of experimental parkinsonism. FSL rats showed a potentiated tremorgenic response to tacrine, a cholinesterase inhibitor used experimentally to induce 6 Hz resting tremor reminiscent of parkinsonian tremor. We also studied rats lesioned with 6-OHDA to induce hemiparkinsonism. No baseline differences in dopaminergic response to acute apomorphine or L-DOPA was found. However, following chronic treatment with L-DOPA, FRL rats developed sensitization of turning and abnormal involuntary movements (AIMs); these effects were counteracted by the anti-dyskinetic 5-HT1A agonist/D-2 partial agonist sarizotan. In contrast, FSL rats did not develop sensitization of turning and only minor AIMs in response to L-DOPA treatment. The roles of several non-dopamine systems underlying this discrepancy were studied. Unexpectedly, no differences of opioid neuropeptides or serotonin markers were found between FRL and FSL rats. The marked behavioral difference between the FRL and FSL rats was paralleled with the striatal expression of the established marker, c-fos, but also the GABAergic transporter (vGAT), and a hitherto unknown marker, tamalin, that is known to regulate mGluR5 receptor function and postsynaptic organization. This study demonstrates that behavioral and transcriptional responses of non-dopaminergic systems to experimental parkinsonism and L-DOPA are modified in a genetic rat model of depression.
  •  
32.
  • Shariatgorji, Mohammadreza, et al. (författare)
  • Bromopyrylium Derivatization Facilitates Identification by Mass Spectrometry Imaging of Monoamine Neurotransmitters and Small Molecule Neuroactive Compounds
  • 2020
  • Ingår i: Journal of the American Society for Mass Spectrometry. - : AMER CHEMICAL SOC. - 1044-0305 .- 1879-1123. ; 31:12, s. 2553-2557
  • Tidskriftsartikel (refereegranskat)abstract
    • Mass spectrometry imaging using matrix-assisted laser desorption/ionization and desorption electrospray ionization has recently been employed to investigate the distribution of neurotransmitters, including biogenic amines and amino acids, directly in brain tissue sections. Ionization is facilitated by charge-tagging through pyrylium derivatization of primary amine containing neurotransmitters directly in tissue sections, significantly improving the limit of detection. Since the derivatization adds carbon and hydrogen to the target compounds, the resulting isotopic patterns of the products are not distinctive from those of the nonderivatized species. Here, we describe an approach for chemically modifying the reactive pyrylium ion to introduce the distinct isotopic signature of bromine in mass spectra of chemically derivatized substances in tissue sections. The method enables monoamine compounds to be distinguished directly in tissue sections, facilitating their identification.
  •  
33.
  • Shariatgorji, Mohammadreza, et al. (författare)
  • Comprehensive mapping of neurotransmitter networks by MALDI-MS imaging
  • 2019
  • Ingår i: Nature Methods. - : NATURE PUBLISHING GROUP. - 1548-7091 .- 1548-7105. ; 16:10, s. 1021-1028
  • Tidskriftsartikel (refereegranskat)abstract
    • We present a mass spectrometry imaging (MSI) approach for the comprehensive mapping of neurotransmitter networks in specific brain regions. Our fluoromethylpyridinium-based reactive matrices facilitate the covalent charge-tagging of molecules containing phenolic hydroxyl and/or primary or secondary amine groups, including dopaminergic and serotonergic neurotransmitters and their associated metabolites. These matrices improved the matrix-assisted laser desorption/ionization (MALDI)-MSI detection limit toward low-abundance neurotransmitters and facilitated the simultaneous imaging of neurotransmitters in fine structures of the brain at a lateral resolution of 10 mu m. We demonstrate strategies for the identification of unknown molecular species using the innate chemoselectivity of the reactive matrices and the unique isotopic pattern of a brominated reactive matrix. We illustrate the capabilities of the developed method on Parkinsonian brain samples from human post-mortem tissue and animal models. The direct imaging of neurotransmitter systems provides a method for exploring how various neurological diseases affect specific brain regions through neurotransmitter modulation.
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34.
  • Shariatgorji, Mohammadreza, et al. (författare)
  • Controlled-pH Tissue Cleanup Protocol for Signal Enhancement of Small Molecule Drugs Analyzed by MALDI-MS Imaging
  • 2012
  • Ingår i: Analytical Chemistry. - : American Chemical Society (ACS). - 1520-6882 .- 0003-2700. ; 84:10, s. 4603-4607
  • Tidskriftsartikel (refereegranskat)abstract
    • The limit of detection of low-molecular weight compounds in tissue sections, analyzed by matrix assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI), was significantly improved by employing sample washing using a pH-controlled buffer solution. The pH of the washing solutions were set at values whereby the target analytes would have low solubility. Washing the tissue sections in the buffered solution resulted in removal of endogenous soluble ionization-suppressing compounds and salts, while the target compound remained in situ with minor or no delocalization during the buffered washing procedure. Two pharmaceutical compounds (cimetidine and imipramine) and one new protease inhibitor compound were successfully used to evaluate the feasibility of the pH-controlled tissue washing protocol for MALDI-MSI. Enhancement in signal-to-noise ratio was achieved by a factor of up to 10.
  •  
35.
  • Shariatgorji, Mohammadreza, et al. (författare)
  • Deuterated Matrix-Assisted Laser Desorption Ionization Matrix Uncovers Masked Mass Spectrometry Imaging Signals of Small Molecules
  • 2012
  • Ingår i: Analytical Chemistry. - : American Chemical Society (ACS). - 0003-2700 .- 1520-6882. ; 84:16, s. 7152-7157
  • Tidskriftsartikel (refereegranskat)abstract
    • D-4-alpha-Cyano-4-hydroxycinnamic acid (D-4-CHCA) has been synthesized for use as a matrix for matrix-assisted laser desorption ionization-mass spectrometry (MALDI-MS) and MALDI-MS imaging (MSI) of small molecule drugs and endogenous compounds. MALDI-MS analysis of small molecules has historically been hindered by interference from matrix ion clusters and fragment peaks that mask signals of low molecular weight compounds of interest. By using D-4-CHCA, the cluster and fragment peaks of CHCA, the most common matrix for analysis of small molecules, are shifted by + 4, + 8 and + 12 Da, which expose signals across areas of the previously concealed low mass range. Here, obscured MALDI-MS signals of a synthetic small molecule pharmaceutical, a naturally occurring isoquinoline alkaloid, and endogenous compounds including the neurotransmitter acetylcholine have been unmasked and imaged directly from biological tissue sections.
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36.
  • Shariatgorji, Mohammadreza, et al. (författare)
  • Direct targeted quantitative molecular imaging of neurotransmitters in brain tissue sections
  • 2014
  • Ingår i: Neuron. - : Elsevier BV. - 0896-6273 .- 1097-4199. ; 84:4, s. 697-707
  • Tidskriftsartikel (refereegranskat)abstract
    • Current neuroimaging techniques have very limited abilities to directly identify and quantify neurotransmitters from brain sections. We have developed a molecular-specific approach for the simultaneous imaging and quantitation of multiple neurotransmitters, precursors, and metabolites, such as tyrosine, tryptamine, tyramine, phenethylamine, dopamine, 3-methoxytyramine, serotonin, GABA, glutamate, acetylcholine, and L-alpha-glycerylphosphorylcholine, in histological tissue sections at high spatial resolutions. The method is employed to directly measure changes in the absolute and relative levels ofneurotransmitters in specific brain structures in animal disease models and in response to drug treatments, demonstrating the power of mass spectrometry imaging in neuroscience.
  •  
37.
  • Shariatgorji, Mohammadreza, et al. (författare)
  • Mass Spectrometry Imaging, an Emerging Technology in Neuropsychopharmacology
  • 2014
  • Ingår i: Neuropsychopharmacology. - : Springer Science and Business Media LLC. - 0893-133X .- 1740-634X. ; 39:1, s. 34-49
  • Forskningsöversikt (refereegranskat)abstract
    • Mass spectrometry imaging is a powerful tool for directly determining the distribution of proteins, peptides, lipids, neurotransmitters, metabolites and drugs in neural tissue sections in situ. Molecule-specific imaging can be achieved using various ionization techniques that are suited to different applications but which all yield data with high mass accuracies and spatial resolutions. The ability to simultaneously obtain images showing the distributions of chemical species ranging from metal ions to macromolecules makes it possible to explore the chemical organization of a sample and to correlate the results obtained with specific anatomical features. The imaging of biomolecules has provided new insights into multiple neurological diseases, including Parkinson's and Alzheimer's disease. Mass spectrometry imaging can also be used in conjunction with other imaging techniques in order to identify correlations between changes in the distribution of important chemical species and other changes in the properties of the tissue. Here we review the applications of mass spectrometry imaging in neuroscience research and discuss its potential. The results presented demonstrate that mass spectrometry imaging is a useful experimental method with diverse applications in neuroscience.
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38.
  • Shariatgorji, Mohammadreza, et al. (författare)
  • Pyrylium Salts as Reactive Matrices for MALDI-MS Imaging of Biologically Active Primary Amines
  • 2015
  • Ingår i: Journal of the American Society for Mass Spectrometry. - : American Chemical Society (ACS). - 1044-0305 .- 1879-1123. ; 26:6, s. 934-939
  • Tidskriftsartikel (refereegranskat)abstract
    • Many neuroactive substances, including endogenous biomolecules, environmental compounds, and pharmaceuticals possess primary amine functional groups. Among these are catecholamine neurotransmitters (e.g., dopamine), many substituted phenethylamines (e.g., amphetamine), as well as amino acids and neuropeptides. In most cases, mass spectrometric (ESI and MALDI) analyses of trace amounts of such compounds are challenging because of their poor ionization properties. We present a method for chemical derivatization of primary amines by reaction with pyrylium salts that facilitates their detection by MALDI-MS and enables the imaging of primary amines in brain tissue sections. A screen of pyrylium salts revealed that the 2,4-diphenyl-pyranylium ion efficiently derivatizes primary amines and can be used as a reactive MALDI-MS matrix that induces both derivatization and desorption. MALDI-MS imaging with such matrix was used to map the localization of dopamine and amphetamine in brain tissue sections and to quantitatively map the distribution of the neurotoxin beta-N-methylamino-L-alanine.
  •  
39.
  • Shariatgorji, Mohammadreza, et al. (författare)
  • Simultaneous imaging of multiple neurotransmitters and neuroactive substances in the brain by desorption electrospray ionization mass spectrometry
  • 2016
  • Ingår i: NeuroImage. - : Elsevier BV. - 1053-8119 .- 1095-9572. ; 136, s. 129-138
  • Tidskriftsartikel (refereegranskat)abstract
    • With neurological processes involving multiple neurotransmitters and neuromodulators, it is important to have the ability to directly map and quantify multiple signaling molecules simultaneously in a single analysis. By utilizing a molecular-specific approach, namely desorption electrospray ionization mass spectrometry imaging (DESI-MSI), we demonstrated that the technique can be used to image multiple neurotransmitters and their metabolites (dopamine, dihydroxyphenylacetic acid, 3-methoxytyramine, serotonin, glutamate, glutamine, aspartate,gamma-aminobutyric acid, adenosine) as well as neuroactive drugs (amphetamine, sibutramine, fluvoxamine) and drug metabolites in situ directly in brain tissue sections. The use of both positive and negative ionization modes increased the number of identified molecular targets. Chemical derivatization by charge-tagging the primary amines of molecules significantly increased the sensitivity, enabling the detection of low abundant neurotransmitters and other neuroactive substances previously undetectable by MSI. The sensitivity of the imaging approach of neurochemicals has a great potential in many diverse applications in fields such as neuroscience, pharmacology, drug discovery, neurochemistry, and medicine.
  •  
40.
  • Shariatgorji, Reza, et al. (författare)
  • Spatial visualization of comprehensive brain neurotransmitter systems and neuroactive substances by selective in situ chemical derivatization mass spectrometry imaging
  • 2021
  • Ingår i: Nature Protocols. - : Springer Nature. - 1754-2189 .- 1750-2799. ; 16:7, s. 3298-3321
  • Tidskriftsartikel (refereegranskat)abstract
    • Molecule-specific techniques such as MALDI and desorption electrospray ionization mass spectrometry imaging enable direct and simultaneous mapping of biomolecules in tissue sections in a single experiment. However, neurotransmitter imaging in the complex environment of biological samples remains challenging. Our covalent charge-tagging approach using on-tissue chemical derivatization of primary and secondary amines and phenolic hydroxyls enables comprehensive mapping of neurotransmitter networks. Here, we present robust and easy-to-use chemical derivatization protocols that facilitate quantitative and simultaneous molecular imaging of complete neurotransmitter systems and drugs in diverse biological tissue sections with high lateral resolution. This is currently not possible with any other imaging technique. The protocol, using fluoromethylpyridinium and pyrylium reagents, describes all steps from tissue preparation (-1 h), chemical derivatization (1-2 h), data collection (timing depends on the number of samples and lateral resolution) and data analysis and interpretation. The specificity of the chemical reaction can also help users identify unknown chemical identities. Our protocol can reveal the cellular locations in which signaling molecules act and thus shed light on the complex responses that occur after the administration of drugs or during the course of a disease.
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41.
  • Sköld, Karl, 1974- (författare)
  • Neuropeptidomics – Methods and Applications
  • 2006
  • Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
    • The sequencing of genomes has caused a growing demand for functional analysis of gene products. This research field named proteomics is derived from the term proteome, which by analogy to genome is defined as all proteins expressed by a cell or a tissue. Proteomics is however methodologically restricted to the analysis of proteins with higher molecular weights. The development of a technology which includes peptides with low molecular weight and small proteins is needed, since peptides play a central role in many biological processes. To study endogenous peptides and hormones, the peptidome, an improved method comprising rapid deactivation in combination with nano-flow liquid chromatography (LC) and mass spectrometry (MS) was developed. The method has been used to investigate endogenous peptides in brains of mouse and rat. Several novel peptides have been discovered together with known neuropeptides. To elucidate the post mortem time influence on peptides and proteins, a time course study was performed using peptidomics and proteomics technologies. Already after three minutes a substantial amount of protein fragments emerged in the peptidomics study and some endogenous peptides were drastically reduced with increasing post mortem time. Of about 1500 proteins investigated, 53 were found to be significantly changed at 10 minutes post mortem as compared to control. Moreover, using western blot the level of MAPK phosphorylation was shown to decrease by 95% in the 10 minutes post mortem sample. A database, SwePep (a repository of endogenous peptides, hormones and small proteins), was constructed to facilitate identification using MS. The database also contains additional information concerning the peptides such as physical properties. A method for analysis of LC-MS data, including scanning for, and further profiling of, biologically significant peptides was developed. We show that peptides present in different amounts in groups of samples can be automatically detected.The peptidome approach was used to investigate levels of peptides in two animal models of Parkinson’s disease. PEP-19, was found to be significantly decreased in the striatum of MPTP lesioned parkinsonian mice. The localization and expression was further investigated by imaging MALDI MS and by in situ hybridization. The brain peptidome of reserpine treated mice was investigated and displayed a number of significantly altered peptides. This thesis demonstrates that the peptidomics approach allows for the study of complex biochemical processes.
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42.
  • Sköld, Karl, et al. (författare)
  • The significance of biochemical and molecular sample integrity in brain proteomics and peptidomics: Stathmin (2-20) and peptides as sample quality indicators.
  • 2007
  • Ingår i: Proteomics. - : Wiley. - 1615-9853 .- 1615-9861. ; 7:24, s. 4445-4456
  • Tidskriftsartikel (refereegranskat)abstract
    • Comparisons of transcriptional and translational expression in normal and abnormal states are important to reach an understanding of pathogenesis and pathophysiology. Maintaining the biochemical, molecular, and structural sample integrity is essential for correct sample comparisons. We demonstrate that both proteins and neuropeptides, including their PTMs, are subjected to massive degradation in the brain already 1 min postmortem. Further, markers for determining the integrity and status of a biological sample were identified. The protein fragment stathmin 2-20 correlated well with the general level of postmortem degradation and may serve as a sample quality indicator for future work, both in animal and human postmortem brains. Finally, a novel method for preventing degradation of proteins and peptides in postmortem tissue is presented using rapid and uniform conductive heat transfer on tissue prior to the actual sample preparation procedures, which enables the relatively low-abundant neuropeptides to remain intact, minimizes degradation of proteins by proteolysis, and conserves the PTMs of the neuropeptides.
  •  
43.
  • Svensson, Marcus, et al. (författare)
  • Heat stabilization of the tissue proteome : a new technology for improved proteomics
  • 2009
  • Ingår i: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 8:2, s. 974-981
  • Tidskriftsartikel (refereegranskat)abstract
    • After tissue or body fluid sampling, proteases and other protein-modifying enzymes can rapidly change composition of the proteome. As a direct consequence, analytical results will reflect a mix of in vivo proteome and ex vivo degradation products. Vital information about the presampling state may be destroyed or distorted, leading to variation between samples and incorrect conclusions. Sample stabilization and standardization of sample handling can reduce or eliminate this problem. Here, a novel tissue stabilization system which utilizes a combination of heat and pressure under vacuum was used to stop degradation in mouse brain tissue immediately after sampling. It was found by biochemical assays that enzymatic activity was reduced to background levels in stabilized samples. Western blot analysis confirmed that post-translational phosphorylations of analyzed proteins were stable and conserved for up to 2 h at room temperature and that peptide extracts were devoid of abundant protein degradation fragments. The combination of reduced complexity and proteolytic inactivation enabled mass spectrometric identification of several neuropeptides and endogenous peptides including modified species at higher levels compared to nonstabilized samples. The tissue stabilizing system ensures reproducible and rapid inactivation of enzymes. Therefore, the system provides a powerful improvement to proteomics by greatly reducing the complexity and dynamic range of the proteome in tissue samples and enables enhanced possibilities for discovery and analysis of clinically relevant protein/peptide biomarkers.
  •  
44.
  • Svensson, Marcus, et al. (författare)
  • Neuropeptidomics : MS applied to the discovery of novel peptides from the brain
  • 2007
  • Ingår i: Analytical Chemistry. - : American Chemical Society (ACS). - 0003-2700 .- 1520-6882. ; 79:1, s. 14-21
  • Tidskriftsartikel (refereegranskat)abstract
    • Peptidomics involves the comprehensive analysis of the peptide content of a certain cell, organ, body fluid, or organism. Per E. Andrén and colleagues at Uppsala University and the Karolinska Institutet (both in Sweden) describe neuropeptidomics approaches in brain lysates and tissue sections to study peptide expression in disease models and to identify potentially biologically active neuropeptides.
  •  
45.
  •  
46.
  • Swales, John G., et al. (författare)
  • Quantitation of Endogenous Metabolites in Mouse Tumors Using Mass-Spectrometry Imaging
  • 2018
  • Ingår i: Analytical Chemistry. - : AMER CHEMICAL SOC. - 0003-2700 .- 1520-6882. ; 90:10, s. 6051-6058
  • Tidskriftsartikel (refereegranskat)abstract
    • Described is a quantitative-mass-spectrometry-imaging (qMSI) methodology for the analysis of lactate and glutamate distributions in order to delineate heterogeneity among mouse tumor models used to support drug-discovery efficacy testing. We evaluate and report on preanalysis-stabilization methods aimed at improving the reproducibility and efficiency of quantitative assessments of endogenous molecules in tissues. Stability experiments demonstrate that optimum stabilization protocols consist of frozen-tissue embedding, post-tissue-sectioning desiccation, and storage at -80 degrees C of tissue sections sealed in vacuum-tight containers. Optimized stabilization protocols are used in combination with qMSI methodology for the absolute quantitation of lactate and glutamate in tumors, incorporating the use of two different stable-isotope-labeled versions of each analyte and spectral-clustering performed on each tissue section using k-means clustering to allow region-specific, pixel-by-pixel quantitation. Region-specific qMSI was used to screen different tumor models and identify a phenotype that has low lactate heterogeneity, which will enable accurate measurements of lactate modulation in future drug-discovery studies. We conclude that using optimized qMSI protocols, it is possible to quantify endogenous metabolites within tumors, and region-specific quantitation can provide valuable insight into tissue heterogeneity and the tumor microenvironment.
  •  
47.
  • Vallianatou, Theodosia, et al. (författare)
  • A mass spectrometry imaging approach for investigating how drug-drug interactions influence drug blood-brain barrier permeability
  • 2018
  • Ingår i: NeuroImage. - : Elsevier BV. - 1053-8119 .- 1095-9572. ; 172, s. 808-816
  • Tidskriftsartikel (refereegranskat)abstract
    • There is a high need to develop quantitative imaging methods capable of providing detailed brain localization information of several molecular species simultaneously. In addition, extensive information on the effect of the blood-brain barrier on the penetration, distribution and efficacy of neuroactive compounds is required. Thus, we have developed a mass spectrometry imaging method to visualize and quantify the brain distribution of drugs with varying blood-brain barrier permeability. With this approach, we were able to determine blood-brain barrier transport of different drugs and define the drug distribution in very small brain structures (e.g., choroid plexus) due to the high spatial resolution provided. Simultaneously, we investigated the effect of drug-drug interactions by inhibiting the membrane transporter multidrug resistance 1 protein. We propose that the described approach can serve as a valuable analytical tool during the development of neuroactive drugs, as it can provide physiologically relevant information often neglected by traditional imaging technologies.
  •  
48.
  •  
49.
  • Vallianatou, Theodosia, et al. (författare)
  • Integration of Mass Spectrometry Imaging and Machine Learning Visualizes Region-Specific Age-Induced and Drug-Target Metabolic Perturbations in the Brain
  • 2021
  • Ingår i: ACS Chemical Neuroscience. - : American Chemical Society (ACS). - 1948-7193. ; 12:10, s. 1811-1823
  • Tidskriftsartikel (refereegranskat)abstract
    • Detailed metabolic imaging of specific brain regions in early aging may expose pathophysiological mechanisms and indicate effective neuropharmacological targets in the onset of cognitive decline. Comprehensive imaging of brain aging and drug-target effects is restricted using conventional methodology. We simultaneously visualized multiple metabolic alterations induced by normal aging in specific regions of mouse brains by integrating Fourier-transform ion cyclotron resonance mass spectrometry imaging and combined supervised and unsupervised machine learning models. We examined the interplay between aging and the response to tacrine-induced acetylcholinesterase inhibition, a well-characterized therapeutic treatment against dementia. The dipeptide carnosine (β-alanyl-l-histidine) and the vitamin α-tocopherol were significantly elevated by aging in different brain regions. l-Carnitine and acetylcholine metabolism were found to be major pathways affected by aging and tacrine administration in a brain region-specific manner, indicating altered mitochondrial function and neurotransmission. The highly interconnected hippocampus and retrosplenial cortex displayed different age-induced alterations in lipids and acylcarnitines, reflecting diverse region-specific metabolic effects. The subregional differences observed in the hippocampal formation of several lipid metabolites demonstrate the unique potential of the technique compared to standard mass spectrometry approaches. An age-induced increase of endogenous antioxidants, such as α-tocopherol, in the hippocampus was detected, suggesting an augmentation of neuroprotective mechanisms in early aging. Our comprehensive imaging approach visualized heterogeneous age-induced metabolic perturbations in mitochondrial function, neurotransmission, and lipid signaling, not always attenuated by acetylcholinesterase inhibition.
  •  
50.
  • Vallianatou, Theodosia, et al. (författare)
  • Molecular imaging identifies age-related attenuation of acetylcholine in retrosplenial cortex in response to acetylcholinesterase inhibition
  • 2019
  • Ingår i: Neuropsychopharmacology. - : Springer Science and Business Media LLC. - 0893-133X .- 1740-634X. ; 44, s. 2091-2098
  • Tidskriftsartikel (refereegranskat)abstract
    • The neurotransmitter of the cholinergic system, acetylcholine plays a major role in the brain's cognitive function and is involved in neurodegenerative disorders. Here, we present age-related alterations of acetylcholine levels after administration of the acetylcholinesterase inhibitor drug tacrine in normal mice. Using a quantitative, robust and molecular-specific mass spectrometry imaging method we found that tacrine administration significantly raised acetylcholine levels in most areas of sectioned mice brains, inter alia the striatum, hippocampus and cortical areas. However, acetylcholine levels in retrosplenial cortex were significantly lower in 14-month-old than in 12-week-old animals following its administration, indicating that normal aging affects the cholinergic system's responsivity. This small brain region is interconnected with an array of brain networks and is involved in numerous cognitive tasks. Simultaneous visualization of distributions of tacrine and its hydroxylated metabolites in the brain revealed a significant decrease in levels of the metabolites in the 14-month-old mice. The results highlight strengths of the imaging technique to simultaneously investigate multiple molecular species and the drug-target effects in specific regions of the brain. The proposed approach has high potential in studies of neuropathological conditions and responses to neuroactive treatments.
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