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  • 201.
    Kask, Lena
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Coagulation and inflammation science. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Jorsback, Anneli
    Winkvist, Maria
    Alfredsson, Jenny
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Coagulation and inflammation science.
    Ek, Bo
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Siegbahn, Agneta
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Coagulation and inflammation science.
    Identification of Novel Downstream Molecules of Tissue Factor Activation by Comparative Proteomic Analysis2014In: Journal of Proteome Research, ISSN 1535-3893, E-ISSN 1535-3907, Vol. 13, no 2, p. 477-488Article in journal (Refereed)
    Abstract [en]

    Tissue factor (TF) is both an initiator of blood coagulation and a signaling receptor. Using a proteomic approach, we investigated the role of TF in cell signaling when stimulated by its ligand, activated factor VII (FVIIa). From a 2-D difference gel electrophoresis (DIGE) study we found forty one spots that were differentially regulated over time in FVIIa stimulated cells or in comparison to nonstimulated cells. Mass spectrometry identifies 23 out of these as 13 different proteins. One of them, elongation factor 2 (EF-2), was investigated in greater detail by Western blot, a protein synthesis assay and cell cycle analysis. When tissue factor was stimulated by FVIIa, the phosphorylation of EF-2 increased which inactivates this protein. Analyzing the effect using site inactivated FVIIa (FVIIai), as well as the protease activated receptor 2 (PAR-2) agonist SLIGKV, indicated that the inactivation was not PAR-2 dependent. A panel of tissue factor mutants was analyzed further to try to pinpoint what part of the cytoplasmic domain that is needed for this effect. Performing a protein synthesis assay in two different cell lines we could confirm that protein synthesis decreased upon stimulation by FVIIa. Cell cycle analysis showed that FVIIa also promotes a higher degree of cell proliferation.

  • 202.
    Kiselova, Nadezda
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Dierker, Tabea
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Spillmann, Dorothe
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Ramström, Margareta
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Uppsala University, Science for Life Laboratory, SciLifeLab.
    An automated mass spectrometry-based screening method for analysis of sulfated glycosaminoglycans2014In: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 450, no 1, p. 598-603Article in journal (Refereed)
    Abstract [en]

    Glycosaminoglycans (GAGs) are linear polysaccharides, consisting of repeated disaccharide units, attached to core proteins in all multicellular organisms. Chondroitin sulfate (CS) and dermatan sulfate (DS) constitute a subgroup of sulfated GAGs for which the degree of sulfation varies between species and tissues. One major goal in GAG characterization is to correlate structure to function. A common approach is to exhaustively degrade the GAG chains and thereafter determine the amount of component disaccharide units. In large-scale studies, there is a need for high-throughput screening methods since existing methods are either very time- or samples consuming. Here, we present a new strategy applying MALDI-TOF MS in positive ion mode for semi-qualitative and quantitative analysis of CS/DS derived disaccharide units. Only a few picomoles of sample are required per analysis and 10 samples can be analyzed in 25 min, which makes this approach an attractive alternative to many established assay methods. The total CS/DS concentration in 19 samples derived from Caenorhabditis elegans and mammalian tissues and cells was determined. The obtained results were well in accordance with concentrations determined by a standard liquid chromatography-based method, demonstrating the applicability of the method for samples from various biological matrices containing CS/DS of different sulfation degrees.

  • 203.
    Kjellander, Marcus
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Biochemistry.
    Billinger, Erika
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Biochemistry.
    Ramachandraiah, Harisha
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC.
    Boman, Mats
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström, Inorganic Chemistry.
    Bergström Lind, Sara
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Johansson, Gunnar
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Biochemistry.
    A flow-through nanoporous alumina trypsin bioreactor for mass spectrometry peptide fingerprinting2018In: Journal of Proteomics, ISSN 1874-3919, E-ISSN 1876-7737, Vol. 172, p. 165-172Article in journal (Refereed)
    Abstract [en]

    Mass spectrometry-based proteomics benefits from efficient digestion of protein samples. In this study, trypsinwas immobilized on nanoporous anodized alumina membranes to create an enzyme reactor suitable for peptidemassfingerprinting. The membranes were derivatized with 3-aminopropyltriethoxysilane and the amino groupswere activated with carbonyldiimidazole to allow coupling of porcine trypsin viaε-amino groups. The functionwas assessed using the artificial substrate Nα-Benzoyl-L-arginine 4-nitroanilide hydrochloride, bovine ribonu-clease A and a human plasma sample. A 10-membraneflow-through reactor was used for fragmentation and MSanalysis after a single pass of substrate both by collection of product and subsequent off-line analysis, and bycoupling on-line to the instrument. The peptide pattern allowed correct identification of the single target proteinin both cases, and of > 70 plasma proteins in single pass mode followed by LC-MS analysis. The reactor retained76% of the initial activity after 14 days of storage and repeated use at room temperature.

    Significance:This manuscript describes the design of a stable enzyme reactor that allows efficient and fast di-gestion with negligible leakage of enzyme and enzyme fragments. The high stability facilitates the use in anonline-setup with MS detection since it allows the processing of multiple samples within an extended period of time without replacement.

  • 204.
    Klar, Jessica K.
    et al.
    Univ Southampton, Ocean & Earth Sci, Natl Oceanog Ctr Southampton, Waterfront Campus,European Way, Southampton SO14 3ZH, Hants, England.;Univ Toulouse, LEGOS, IRD, CNES,CNRS,UPS, 18 Ave Edouard Belin, F-31401 Toulouse, France..
    James, Rachael H.
    Univ Southampton, Ocean & Earth Sci, Natl Oceanog Ctr Southampton, Waterfront Campus,European Way, Southampton SO14 3ZH, Hants, England..
    Gibbs, Dakota
    Natl Oceanog Ctr Southampton, Marine Geosci, European Way, Southampton SO14 3ZH, Hants, England.;Southern Cross Geosci, Mil Rd, Lismore, NSW 2480, Australia..
    Lough, Alastair
    Univ Southampton, Ocean & Earth Sci, Natl Oceanog Ctr Southampton, Waterfront Campus,European Way, Southampton SO14 3ZH, Hants, England..
    Parkinson, Ian
    Univ Bristol, Sch Earth Sci, Queens Rd, Bristol BS8 1RJ, Avon, England..
    Milton, J. Andrew
    Univ Southampton, Ocean & Earth Sci, Natl Oceanog Ctr Southampton, Waterfront Campus,European Way, Southampton SO14 3ZH, Hants, England..
    Hawkes, Jeffrey A.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Connelly, Douglas P.
    Natl Oceanog Ctr Southampton, Marine Geosci, European Way, Southampton SO14 3ZH, Hants, England..
    Isotopic signature of dissolved iron delivered to the Southern Ocean from hydrothermal vents in the East Scotia Sea2017In: Geology, ISSN 0091-7613, E-ISSN 1943-2682, Vol. 45, no 4, p. 351-354Article in journal (Refereed)
    Abstract [en]

    It has recently been demonstrated that hydrothermal vents are an important source of dissolved Fe (dFe) to the Southern Ocean. The isotopic composition (delta Fe-56) of dFe in vent fluids appears to be distinct from other sources of dFe to the deep ocean, but the evolution of delta Fe-56 during mixing between vent fluids and seawater is poorly constrained. Here we present the evolution of delta Fe-56 for dFe in hydrothermal fluids and dispersing plumes from two sites in the East Scotia Sea. We show that delta Fe-56 values in the buoyant plume are distinctly lower (as low as -1.19 parts per thousand) than the hydrothermal fluids (-0.29 parts per thousand), attributed to (1) precipitation of Fe sulfides in the early stages of mixing, and (2) partial oxidation of Fe(II) to Fe(III), >55% of which subsequently precipitates as Fe oxyhydroxides. By contrast, the delta Fe-56 signature of stabilized dFe in the neutrally buoyant plume is -0.3 parts per thousand to -0.5 parts per thousand. This cannot be explained by continued dilution of the buoyant plume with background seawater; rather, we suggest that isotope fractionation of dFe occurs during plume dilution due to Fe ligand complexation and exchange with labile particulate Fe. The delta Fe-56 signature of stabilized hydrothermal dFe in the East Scotia Sea is distinct from background seawater and may be used to quantify the hydrothermal dFe input to the ocean interior.

  • 205.
    Kleint, Charlotte
    et al.
    Jacobs Univ Bremen, Sch Engn & Sci, Dept Phys & Earth Sci, Bremen, Germany.
    Hawkes, Jeffrey A.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Sander, Sylvia G.
    Univ Otago, NIWA Univ Otago Res Ctr Oceanog, Marine & Freshwater Chem, Dept Chem, Dunedin, New Zealand.
    Koschinsky, Andrea
    Jacobs Univ Bremen, Sch Engn & Sci, Dept Phys & Earth Sci, Bremen, Germany.
    Voltammetric Investigation of Hydrothermal Iron Speciation2016In: Frontiers in Marine Science, E-ISSN 2296-7745, Vol. 3, article id 75Article in journal (Refereed)
    Abstract [en]

    Hydrothermal vent fluids are highly enriched in iron (Fe) compared to ambient seawater, and organic ligands may play a role in facilitating the transport of some hydrothermal Fe into the open ocean. This is important since Fe is a limiting micronutrient for primary production in large parts of the world's surface ocean. We have investigated the concentration and speciation of Fe in several vent fluid and plume samples from the Nifonea vent field, Coriolis Troughs, New Hebrides Island Arc, South Pacific Ocean using competitive ligand exchange-adsorptive cathodic stripping voltammetry (CLE-AdCSV) with salicylaldoxime (SA) as the artificial ligand. Our results for total dissolved Fe (dFe) in the buoyant hydrothermal plume samples showed concentrations up to 3.86 mu M dFe with only a small fraction between 1.1 and 11.8% being chemically labile. Iron binding ligand concentrations (M) were found in mu M level with strong conditional stability constants up to logK(FeL,Fe)(3+) of 22.9. Within the non-buoyant hydrothermal plume above the Nifonea vent field, up to 84.7% of the available Fe is chemically labile and [L] concentrations up to 97 nM were measured. [L] was consistently in excess of Fe-lab, indicating that all available Fe is being complexed, which in combination with high F-lab values in the non-buoyant plume, signifies that a high fraction of hydrothermal dFe is potentially being transported away from the plume into the surrounding waters, contributing to the global oceanic Fe budget.

  • 206.
    Korvela, Marcus
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Method development for the analysis of complex samples with inductively coupled plasma mass spectrometry2019Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    In this thesis the development of methods for handling the problems associated with analyzing trace elements in complex matrixes using inductively coupled plasma mass spectrometry is presented. Trace elements such as Cu, Fe, Se, and Zn, to name a few, do play important roles in different organisms. Therefore it can be of importance to study trace elements in different samples of biological origin. As trace elements are low in abundance, sensitive instrumental techniques such as inductively coupled plasma mass spectrometry (ICP-MS) are required for accurate determination. Due to the complexity of samples with biological origin, careful method development, both regarding the sample preparation and instrumental analysis has to be performed to minimize negative effects on the instrument signal and introduction of interferences.

    For example the metal contents of mink livers were analyzed, after bomb digestion to investigate if the metal concentration could be linked to changes in the organ morphology as well as the minks’ environment. Morphological changes and capture locations could be linked to the metals investigated. The investigation of the elemental composition of cerebrospinal fluid from chronic pain patients using spinal cord stimulation electrode treatment on the other hand required less harsh sample treatment. No correlation between the spinal cord stimulation and element concentration could be found, but differences between patients and the control group were presented hinting that chronic pain intrinsically could affect the cerebrospinal fluid metal concentration. Another bodily fluid of interest is saliva and the use of paperpoint sticks as a sampling technique for Ti in saliva was investigated. As Ti is interfered by several components expected to be found in saliva, the use of reaction or collision gas was also investigated to reduce the effects of interferences. Simple leaching of the paperpoint sticks together with complexing the Ti with NH3 as reaction gas was shown to be optimal. Finally, how the selection of internal standard would be affected by the use of reaction and collision gases was also investigated. With collision gas most internal standards worked fine, while for reaction gas internal standard selection was harder. For elements with high ionization energy such as As, Se and Zn the choice of internal standard was very dependent on matrix. While ICP-MS suffers from problems when analyzing samples with complex matrixes many of them can be minimized by proper method development as shown in this thesis.

  • 207.
    Korvela, Marcus
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Andersson, Marit
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Pettersson, Jean
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Internal standards in inductively coupled plasma mass spectrometry using kinetic energy discrimination and dynamic reaction cells2018In: Journal of Analytical Atomic Spectrometry, ISSN 0267-9477, E-ISSN 1364-5544, Vol. 33, no 10, p. 1770-1776Article in journal (Refereed)
    Abstract [en]

    ICP-MS is a sensitive element analysis technique used for analyzing several different sample types. This can result in difficult matrixes which can affect both physical parameters and create overlaps of analyte elements. Some of the possible overlaps can be reduced by the use of reaction and/or collision cells, while the use of internal standards can help with reducing the physical interferences caused by a matrix. While both internal standardization and the use of cells have been studied separately, their effects on each other have not been investigated earlier. In this study ICP-MS was used to analyze Mg-24, Al-27, Ti-47, Ti-49, V-51, Cr-52, Cr-53, Mn-55, Fe-57, Co-59, Ni-60, Ni-61, Ni-62, Cu-63, Cu-65, Zn-66, Zn-67, As-75, Se-78, Se-82, Cd-111, and Pb-208 with Be-9, Y-89, Ga-69, Rh-103, In-115, Ir-193, and Tl-205 as internal standards with high concentrations of either HNO3, PBS-buffer, or Triton X-100 as the matrix, in reaction-, collision- and standard-cell modes. This was done to investigate which internal standards would compensate matrix effects in different cell modes. All internal standards, except Be, compensated fairly well (relative sensitivity RSD < 10%) even for severe matrix effects for most elements regardless of similarity in mass in the different cell modes. For Zn, As and Se no proper internal standard could be found, of the ones investigated.

  • 208.
    Korvela, Marcus
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Andersson, Marit
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Pettersson, Jean
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Pettersson, Mattias
    Division of Prosthetic Dentistry, Department of Odontology, Umeå University.
    Development of a method using paperpoint stick sampling to determine Ti concentration in saliva with ICP-MSManuscript (preprint) (Other academic)
    Abstract [en]

    Peri-implantitis is a disease that causes an inflammatory process in soft tissues as well as bone loss around dental implants integrated into bone. The relationship between peri-implantitis and titanium presence has previously been established and also a correlation between the inflammation severity and Ti concentration in biopsies from patients with dental implants. Biopsy can be considered as an invasive sampling procedure and therefore there is an interest to study alternative sampling methods, such as collecting saliva with paperpoint sticks. The small volume absorbed by the paperpoint sticks requires a sensitive detection method for Ti, therefore ICP-MS was used. To minimize the effect of common polyatomic interferences on Ti, the use of He as a collision gas and complexation using NH3 as reaction gas were investigated and both methods were compared to each other. Three sample preparation techniques were investigated: leaching, ultrasonic extraction and microwave bomb digestion. The use of paperpoint sticks for sampling was shown to be a valid approach with simple leaching in combination with ICP-MS using either He- or NH3-gas. The use of NH3-gas to create a 131Ti-complex, required more technical expertise, but gave a 2 times lower detection limit compared to using He as a collision gas.

  • 209.
    Korvela, Marcus
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Lind, Anne-Li
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Anaesthesiology and Intensive Care.
    Wetterhall, Magnus
    GE Healthcare Biosci, S-75148 Uppsala, Sweden..
    Gordh, Torsten
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Anaesthesiology and Intensive Care.
    Andersson, Marit
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Pettersson, Jean
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Quantification of 10 elements in human cerebrospinal fluid from chronic pain patients with and without spinal cord stimulation2016In: Journal of Trace Elements in Medicine and Biology, ISSN 0946-672X, E-ISSN 1878-3252, Vol. 37, p. 1-7Article in journal (Refereed)
    Abstract [en]

    Neuropathic pain affects 1-10% of the general population and is caused by a lesion or disease of the somatosensory nervous system. Spinal cord stimulation (SCS), a method where implanted electrodes stimulate the spinal cord, has been successfully used to treat drug-resistant neuropathic pain, but the mechanism of action is largely unknown. Studies show that SCS changes the protein levels in CSF (cerebrospinal fluid) of pain patients. Several neurological conditions have been shown to alter the elemental composition of CSF. Therefore changes in the levels of ions and trace elements in the CSF may correspond to SCS use. This study used ICP-MS (Inductively coupled plasma mass spectrometry) and ICP-AES (Inductively coupled plasma atomic emission spectroscopy) to quantify 10 elements in CSF from chronic neuropathic pain patients using SCS. The element concentrations in CSF from patients with SCS treatment on/off, were measured. No effect on the element concentrations in CSF from treatment with SCS could be detected. Also, the elemental concentrations in pooled CSF from patients without chronic neuropathic pain was determined and compared to the patients using SCS. The concentration of the elements Ca, Sr, Na, K, P, Mg and Ti were, significantly higher in patients compared to the CSF-control.

  • 210.
    Kovachev, Petar Stefanov
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology.
    Banerjee, Debapriya
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology.
    Rangel, Luciana Pereira
    Univ Fed Rio de Janeiro, Fac Farm, BR-21941902 Rio De Janeiro, Brazil..
    Eriksson, Jonny
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Pedrote, Murilo M
    Univ Fed Rio de Janeiro, Inst Bioquim Med Leopoldo de Meis, Inst Nacl Ciencia Tecnol Biol Estrutural & Bioima, BR-21941902 Rio De Janeiro, Brazil..
    Martins-Dinis, Mafalda Maria D C
    Univ Fed Rio de Janeiro, Inst Bioquim Med Leopoldo de Meis, Inst Nacl Ciencia Tecnol Biol Estrutural & Bioima, BR-21941902 Rio De Janeiro, Brazil..
    Edwards, Katarina
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Cordeiro, Yraima
    Univ Fed Rio de Janeiro, Fac Farm, BR-21941902 Rio De Janeiro, Brazil..
    Silva, Jerson L
    Univ Fed Rio de Janeiro, Inst Bioquim Med Leopoldo de Meis, Inst Nacl Ciencia Tecnol Biol Estrutural & Bioima, BR-21941902 Rio De Janeiro, Brazil..
    Sanyal, Suparna
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology.
    Distinct modulatory role of RNA in the aggregation of the tumor suppressor protein p53 core domain.2017In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 292, no 22, p. 9345-9357Article in journal (Refereed)
    Abstract [en]

    Inactivation of the tumor suppressor protein p53 by mutagenesis, chemical modification, protein-protein interaction, or aggregation has been associated with different human cancers. Although DNA is the typical substrate of p53, numerous studies have reported p53 interactions with RNA. Here, we have examined the effects of RNA of varied sequence, length, and origin on the mechanism of aggregation of the core domain of p53 (p53C) using light scattering, intrinsic fluorescence, transmission electron microscopy, thioflavin-T binding, seeding, and immunoblot assays. Our results are the first to demonstrate that RNA can modulate the aggregation of p53C and full-length p53. We found bimodal behavior of RNA in p53C aggregation. A low RNA:protein ratio (∼1:50) facilitates the accumulation of large amorphous aggregates of p53C. By contrast, at a high RNA:protein ratio (≥1:8), the amorphous aggregation of p53C is clearly suppressed. Instead, amyloid p53C oligomers are formed that can act as seeds nucleating de novo aggregation of p53C. We propose that structured RNAs prevent p53C aggregation through surface interaction and play a significant role in the regulation of the tumor suppressor protein.

  • 211.
    Kronschläger, Martin
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Ophthalmology.
    Forsman, Erik
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Yu, Zhaohua
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Ophthalmology.
    Talebizadeh, Nooshin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Ophthalmology.
    Löfgren, Stefan
    Meyer, Linda M
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Söderberg, Per
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Ophthalmology.
    Pharmacokinetics for topically applied caffeine in the rat2014In: Experimental Eye Research, ISSN 0014-4835, E-ISSN 1096-0007, Vol. 122, p. 94-101Article in journal (Refereed)
    Abstract [en]

    Topically applied caffeine was recently identified as a promising candidate molecule for cataract prevention. Little is known about the pharmacokinetics for topically applied caffeine. Potential toxicity of 72 mM caffeine on the ocular surface and the lens was qualitatively monitored and no toxic effects were observed. The concentration of caffeine was measured in the lens and the blood after topical application of 72 mM caffeine to groups of 10 animals sacrificed at 30, 60, 90 and 120 min after topical application. The lens concentration decreased throughout the observation period while the blood concentration increased up to 120 min. Further, the concentration of caffeine in the lens and blood was measured 30 min after topical application of caffeine, the concentration of caffeine being 0.72, 3.34, 15.51 and 72 mM depending on group belonging, in groups of 10 animals. The caffeine concentration in lens and blood, respectively, increased proportionally to the caffeine concentration topically applied. The rat blood concentrations achieved were far below the equivalent threshold dose of FDA recommended daily dose for humans. This information is important for further development of caffeine eye drops for cataract prevention.

  • 212. Kumar, Abhinav
    et al.
    Bicer, Elif Melis
    Morgan, Anna Babin
    Pfeffer, Paul E
    Monopoli, Marco
    Dawson, Kenneth A
    Eriksson, Jonny
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Edwards, Katarina
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Lynham, Steven
    Arno, Matthew
    Behndig, Annelie F
    Blomberg, Anders
    Somers, Graham
    Hassall, Dave
    Dailey, Lea Ann
    Forbes, Ben
    Mudway, Ian S
    Enrichment of immunoregulatory proteins in the biomolecular corona of nanoparticles within human respiratory tract lining fluid2016In: Nanomedicine: Nanotechnology, Biology and Medicine, ISSN 1549-9634, E-ISSN 1549-9642, Vol. 12, no 4, p. 1033-1043Article in journal (Refereed)
    Abstract [en]

    UNLABELLED: When inhaled nanoparticles deposit in the lungs, they transit through respiratory tract lining fluid (RTLF) acquiring a biomolecular corona reflecting the interaction of the RTLF with the nanomaterial surface. Label-free snapshot proteomics was used to generate semi-quantitative profiles of corona proteins formed around silica (SiO2) and poly(vinyl) acetate (PVAc) nanoparticles in RTLF, the latter employed as an archetype drug delivery vehicle. The evolved PVAc corona was significantly enriched compared to that observed on SiO2 nanoparticles (698 vs. 429 proteins identified); however both coronas contained a substantial contribution from innate immunity proteins, including surfactant protein A, napsin A and complement (C1q and C3) proteins. Functional protein classification supports the hypothesis that corona formation in RTLF constitutes opsonisation, preparing particles for phagocytosis and clearance from the lungs. These data highlight how an understanding of the evolved corona is necessary for the design of inhaled nanomedicines with acceptable safety and tailored clearance profiles.; FROM THE CLINICAL EDITOR: Inhaled nanoparticles often acquire a layer of protein corona while they go through the respiratory tract. Here, the authors investigated the identity of these proteins. The proper identification would improve the understanding of the use of inhaled nanoparticles in future therapeutics.

  • 213.
    Kumar, Abhinav
    et al.
    Institute of Pharmaceutical Science, Faculty of Life Sciences and Medicine, King's College, London, UK.
    Bicer, Elif Melis
    MRC-PHE Centre for Environment and Health, Analytical & Environmental Sciences Division, Faculty of Life Sciences and Medicine, King's College, London, UK.
    Pfeffer, Paul
    MRC-PHE Centre for Environment and Health, Analytical & Environmental Sciences Division, Faculty of Life Sciences and Medicine, King's College, London, UK.
    Monopoli, Marco P
    Pharmaceutical and Medical Chemistry, Royal College of Surgeons in Ireland, Dublin 2, Ireland.; Centre for BioNano Interactions, University College Dublin, Dublin 4, Ireland.
    Dawson, Kenneth A
    Centre for BioNano Interactions, University College Dublin, Dublin 4, Ireland.
    Eriksson, Jonny
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Edwards, Katarina
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Lynham, Steven
    Institute of Psychiatry, Psychology and Neuroscience, Faculty of Life Sciences and Medicine, King's College, London, UK.
    Arno, Matthew
    Genomics Centre, Faculty of Life Sciences and Medicine, King's College, London.
    Behndig, Annelie F
    Department of Public Health and Clinical Medicine, Division of Medicine/Respiratory Medicine, Umeå University, Umeå, Sweden.
    Blomberg, Anders
    Department of Public Health and Clinical Medicine, Division of Medicine/Respiratory Medicine, Umeå University, Umeå, Sweden.
    Somers, Graham
    GSK Medicines Research Centre, Gunnels Wood Road, Stevenage, Hertfordshire, UK.
    Hassall, Dave
    GSK Medicines Research Centre, Gunnels Wood Road, Stevenage, Hertfordshire, UK.
    Dailey, Lea Ann
    Institute of Pharmaceutical Science, Faculty of Life Sciences and Medicine, King's College, London, UK.
    Forbes, Ben
    Institute of Pharmaceutical Science, Faculty of Life Sciences and Medicine, King's College, London, UK.
    Mudway, Ian
    MRC-PHE Centre for Environment and Health, Analytical & Environmental Sciences Division, Faculty of Life Sciences and Medicine, King's College, London, UK.
    Differences in the coronal proteome acquired by particles depositing in the lungs of asthmatic versus healthy humans2017In: Nanomedicine: Nanotechnology, Biology and Medicine, ISSN 1549-9634, E-ISSN 1549-9642, Vol. 13, no 8, p. 2517-2521, article id S1549-9634(17)30115-6Article in journal (Refereed)
    Abstract [en]

    Most inhaled nanomedicines in development are for the treatment of lung disease, yet little is known about their interaction with the respiratory tract lining fluids (RTLFs). Here we combined the use of nano-silica, as a protein concentrator, with label-free snapshot proteomics (LC-MS/MS; key findings confirmed by ELISA) to generate a quantitative profile of the RTLF proteome and provided insight into the evolved corona; information that may be used in future to improve drug targeting to the lungs by inhaled medicines. The asthmatic coronal proteome displayed a reduced contribution of surfactant proteins (SP-A and B) and a higher contribution of α1-antitrypsin. Pathway analysis suggested that asthmatic RTLFs may also be deficient in proteins related to metal handling (e.g. lactoferrin). This study demonstrates how the composition of the corona acquired by inhaled nanoparticles is modified in asthma and suggests depressed mucosal immunity even in mild airway disease.

  • 214. Kumar, Abhinav
    et al.
    Terakosolphan, Wachirun
    Hassoun, Mireille
    Vandera, Kalliopi-Kelli
    Novicky, Astrid
    Harvey, Richard
    Royall, Paul G
    Bicer, Elif Melis
    Eriksson, Jonny
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Edwards, Katarina
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Valkenborg, Dirk
    Nelissen, Inge
    Hassall, Dave
    Mudway, Ian S
    Forbes, Ben
    A Biocompatible Synthetic Lung Fluid Based on Human Respiratory Tract Lining Fluid Composition.2017In: Pharmaceutical research, ISSN 0724-8741, E-ISSN 1573-904X, Vol. 34, no 12, p. 2454-2465Article in journal (Refereed)
    Abstract [en]

    PURPOSE: To characterise a biorelevant simulated lung fluid (SLF) based on the composition of human respiratory tract lining fluid. SLF was compared to other media which have been utilized as lung fluid simulants in terms of fluid structure, biocompatibility and performance in inhalation biopharmaceutical assays.

    METHODS: The structure of SLF was investigated using cryo-transmission electron microscopy, photon correlation spectroscopy and Langmuir isotherms. Biocompatibility with A549 alveolar epithelial cells was determined by MTT assay, morphometric observations and transcriptomic analysis. Biopharmaceutical applicability was evaluated by measuring the solubility and dissolution of beclomethasone dipropionate (BDP) and fluticasone propionate (FP), in SLF.

    RESULTS: SLF exhibited a colloidal structure, possessing vesicles similar in nature to those found in lung fluid extracts. No adverse effect on A549 cells was apparent after exposure to the SLF for 24 h, although some metabolic changes were identified consistent with the change of culture medium to a more lung-like composition. The solubility and dissolution of BDP and FP in SLF were enhanced compared to Gamble's solution.

    CONCLUSION: The SLF reported herein constitutes a biorelevant synthetic simulant which is suitable to study biopharmaceutical properties of inhalation medicines such as those being proposed for an inhaled biopharmaceutics classification system.

  • 215.
    Kushnir, Mark M.
    et al.
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Naessén, Tord
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Women's and Children's Health, Obstetrics and Gynaecology.
    Wånggren, Kjell
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Women's and Children's Health, Obstetrics and Gynaecology.
    Rockwood, Alan L.
    Crockett, David K.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Protein and Steroid Profiles in Follicular Fluid after Ovarian Hyperstimulation as Potential Biomarkers of IVF Outcome2012In: Journal of Proteome Research, ISSN 1535-3893, E-ISSN 1535-3907, Vol. 11, no 10, p. 5090-5100Article in journal (Refereed)
    Abstract [en]

    Controlled ovarian hyperstimulation is performed to assist with generation of multiple mature oocytes for use in in vitro fertilization (IVF). The goal of our study was to evaluate differences in protein and steroid profiles in ovarian follicular fluid (hFF) samples obtained during oocy-te retrieval from women undergoing IVF treatment and to identify physiological pathways associated with the proteins. The hFF samples were depleted of abundant proteins, fractionated by ultrafiltration, digested, and analyzed by nano-LC QTOF. Concentrations of 15 endogenous steroids were determined in the samples using LC-MS/MS methods. The total number of proteins identified in the samples was 75, of which 4, 7, and 2 were unique to the samples from women with viable pregnancy, miscarriage, and no pregnancy, respectively. Identified proteins were associated with the acute response signaling, coagulation system, intrinsic and extrinsic prothrombin activation, complement system, neuroprotective role of THOP1, FXR/RXR activation, role of tissue factor, and growth hormone pathways. A greater number of proteins associated with biosynthesis was found in hFF samples corresponding to the oocytes resulting in pregnancy. The abundance of seven proteins was found to be associated with steroidogenesis. The obtained data will contribute to better understanding of the pathogenesis and development of noninvasive markers for assessment of oocytes viability.

  • 216.
    Kushnir, Mark M.
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Rockwood, A. L.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    LC-MS/MS in clinical laboratories2013In: Bioanalysis, ISSN 1757-6180, E-ISSN 1757-6199, Vol. 5, no 1, p. 5-6Article in journal (Refereed)
  • 217.
    Källsten, Malin
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Zhao, Hongxing
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Konzer, Anne
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Lind, Sara Bergström
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Uppsala University, Science for Life Laboratory, SciLifeLab.
    A comparative study of phosphopeptide-selective techniques for a sub-proteome of a complex biological sample.2016In: Analytical and Bioanalytical Chemistry, ISSN 1618-2642, E-ISSN 1618-2650, Vol. 408, no 9, p. 2347-2356Article in journal (Refereed)
    Abstract [en]

    Phosphorylation of proteins is important for controlling cellular signaling and cell cycle regulatory events. The process is reversible and phosphoproteins normally constitute a minor part of the global proteome in a cell. Thus, sample preparation techniques tailored for phosphoproteome studies are continuously invented and evaluated. This paper aims at evaluating the performances of the most popular techniques for phospho-enrichments in sub-proteome analysis, such as viral proteomes expressed in human cells during infection. A two-species sample of Adenovirus type 2 infected human cells was used, and in-solution digestion, strong cation exchange (SCX), and electrostatic repulsion hydrophilic interaction chromatography (ERLIC) fractionation, and subsequent enrichment by TiO2, were compared with SDS-PAGE fractionation and in-gel digestion. Evaluation was focused on phosphopeptide detection in the sub-proteome. The results showed that the SCX+TiO2 or ERLIC+TiO2 combinations had the highest enrichment efficiencies, but SDS-PAGE fractionation and in-gel digestion resulted in the highest number of identified proteins and phosphopeptides. Furthermore, the study demonstrates the usefulness of applying as many orthogonal techniques as possible in deep phosphoproteome analysis, since the overlap between approaches was low.

  • 218.
    Källsten, Malin
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Ghorasaini, Mohan
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Hartmann, Rafael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Preparative Medicinal Chemistry.
    Lehmann, Fredrik
    Oncopeptides AB.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Kovac, Lucia
    Recipharm OT Chemistry AB.
    Lind, Sara
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Magnetic beads for desalting of monoclonal antibodies and antibody-drug conjugatesManuscript (preprint) (Other academic)
  • 219.
    Källsten, Malin
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Gromova, Arina
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Zhao, Hongxing
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology. Beijer Lab, Rudbeck Lab.
    Valdés, Alberto
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Konzer, Anne
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Pettersson, Ulf
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Medicinsk genetik och genomik. Uppsala University, Science for Life Laboratory, SciLifeLab. Beijer Lab, Rudbeck Lab.
    Lind, Sara Bergstrom
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Uppsala Universitet.
    Temporal characterization of the non-structural Adenovirus type 2 proteome and phosphoproteome using high-resolving mass spectrometry2017In: Virology, ISSN 0042-6822, E-ISSN 1096-0341, Vol. 511, p. 240-248Article in journal (Refereed)
    Abstract [en]

    The proteome and phosphoproteome of non-structural proteins of Adenovirus type 2 (Ad2) were time resolved using a developed mass spectrometry approach. These proteins are expressed by the viral genome and important for the infection process, but not part of the virus particle. We unambiguously confirm the existence of 95% of the viral proteins predicted to be encoded by the viral genome. Most non-structural proteins peaked in expression at late time post infection. We identified 27 non-redundant sites of phosphorylation on seven different non-structural proteins. The most heavily phosphorylated protein was the DNA binding protein (DBP) with 15 different sites. The phosphorylation occupancy rate could be calculated and monitored with time post infection for 15 phosphorylated sites on various proteins. In the DBP, phosphorylations with time-dependent relation were observed. The findings show the complexity of the Ad2 non-structural proteins and opens up a discussion for potential new drug targets.

  • 220.
    Källsten, Malin
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Recipharm OT Chem AB, Uppsala, Sweden.
    Hartmann, Rafael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Preparative Medicinal Chemistry.
    Artemenko, Konstantin
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Bergström Lind, Sara
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Lehmann, Fredrik
    Recipharm OT Chem AB, Uppsala, Sweden.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Qualitative analysis of antibody-drug conjugates (ADCs): an experimental comparison of analytical techniques of cysteine-linked ADCs.2018In: The Analyst, ISSN 0003-2654, E-ISSN 1364-5528, Vol. 143, no 22, p. 5487-5496Article in journal (Refereed)
    Abstract [en]

    Antibody-drug conjugates (ADCs) are an emerging type of biotherapeutics that utilize multiple tissue-specific antibodies combined with a range of linker designs to enable the transportation and selective release of cytotoxic drugs in close proximity to tumours. Consisting of antibodies conjugated to small drug molecules through a variety of linkers, ADCs are chemically complex analytes. Here we present a unique experimental comparison of four techniques for ADC analysis: hydrophobic interaction chromatography (HIC-UV/Vis), reversed phase liquid chromatography mass spectrometry (RPLC-MS), using either a QToF or an Orbitrap analyser, and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Four different ADCs consisting of Trastuzumab, monomethyl auristatin E (MMAE) and a peptidic linker moiety differing in their respective stoichiometric ratios in regard to drug-to-antibody ratio (DAR) were used for the comparison. We found that the determined DAR from all techniques was comparable, while the accuracy of the molecular weights for the conjugated light and heavy chain differed more extensively. This indicates that the choice of a mass analyser is more crucial for determining the accurate weights of the light and heavy chains than to evaluate the DAR of a given batch. However, ambiguous DAR assignment in HIC-UV/Vis or bias for either the light or heavy chain fragments in the mass spectrometry-based techniques can influence the obtained average DAR value and the use of complementary techniques is advisable. Out of the four techniques evaluated, HIC-UV/Vis and MALDI required less time to obtain an average DAR value and would therefore be good for initial screenings in the early stages of the discovery phase of new ADCs.

  • 221.
    Källsten, Malin
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Hartmann, Rafael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Preparative Medicinal Chemistry.
    Kovac, Lucia
    Recipharm OT Chemistry AB.
    Lehmann, Fredrik
    Oncopeptides AB.
    Lind, Sara
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Investigating the Impact of Sample Preparation on Mass Spectrometry-Based Drug-To-Antibody Ratio Determination for Cysteine- and Lysine-Linked Antibody-Drug ConjugatesManuscript (preprint) (Other academic)
  • 222.
    Källsten, Malin
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Recipharm OT Chem AB, Virdings Alle 32b, S-75450 Uppsala, Sweden.
    Pijnappel, Matthijs
    Recipharm OT Chem AB, Virdings Alle 32b, S-75450 Uppsala, Sweden.
    Hartmann, Rafael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Preparative Medicinal Chemistry.
    Lehmann, Fredrik
    Oncopeptides AB, Luntmakargatan 46, SE-11137 Stockholm, Sweden.
    Kovac, Lucia
    Recipharm OT Chem AB, Virdings Alle 32b, S-75450 Uppsala, Sweden.
    Lind, Sara
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Application of triple quadrupole mass spectrometry for the characterization of antibody-drug conjugates2019In: Analytical and Bioanalytical Chemistry, ISSN 1618-2642, E-ISSN 1618-2650, Vol. 411, no 12, p. 2569-2576Article in journal (Refereed)
    Abstract [en]

    Antibody-drug conjugates (ADCs) are an inherently heterogeneous class of biotherapeutics, the development of which requires extensive characterization throughout. During the earliest phases of preclinical development, when synthetic routes towards the desired conjugate are being assessed, the main interest lies in the determination of the average drug-to-antibody ratio (DAR) of a given batch as well as information about different conjugation species. There has been a trend in mass spectrometry (MS)-based characterization of ADCs towards the use of high-resolving mass spectrometry for many of these analyses. Considering the high cost for such an instrument, the evaluation of cheaper and more accessible alternatives is highly motivated. We have therefore tested the applicability of a quadrupole mass analyzer for the aforementioned characterizations. Eight ADCs consisting of trastuzumab and varying stoichiometries of Mc-Val-Cit-PABC-monomethyl auristatin E conjugated to native cysteines were synthesized and served as test analytes. The average DAR value and molecular weights (Mw) of all detected chains from the quadrupole mass analyzer showed surprisingly high agreement with results obtained from a time-of-flight (TOF) mass analyzer and hydrophobic interaction chromatography (HIC)-derived values for all investigated ADC batches. Acquired Mw were within 80ppm of TOF-derived values, and DAR was on average within 0.32 DAR units of HIC-derived values. Quadrupole mass spectrometers therefore represent a viable alternative for the characterization of ADC in early-stage development.

  • 223.
    Källsten, Malin
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Visanu, Diana
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Pijnappel, Matthijs
    Recipharm OT Chemistry AB.
    Lehmann, Fredrik
    Oncopeptides AB.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Lind, Sara
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Kovac, Lucia
    Recipharm OT Chemistry AB.
    The potential use of supercharging agents for improved mass spectrometric analysis of monoclonal antibodies and antibody-drug conjugatesManuscript (preprint) (Other academic)
  • 224.
    Lanekoff, Ingela
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Pacific Northwest Natl Lab, Div Phys Sci, Richland, WA 99352 USA..
    Cha, Jeeyeon
    Cincinnati Childrens Hosp Med Ctr, Div Reprod Sci, Cincinnati, OH 45229 USA.;Vanderbilt Univ, Dept Med, 221 Kirkland Hall, Nashville, TN 37235 USA..
    Kyle, Jennifer E.
    Pacific Northwest Natl Lab, Biol Sci Div, Richland, WA 99352 USA..
    Dey, Sudhansu K.
    Cincinnati Childrens Hosp Med Ctr, Div Reprod Sci, Cincinnati, OH 45229 USA..
    Laskin, Julia
    Pacific Northwest Natl Lab, Div Phys Sci, Richland, WA 99352 USA..
    Burnum-Johnson, Kristin E.
    Pacific Northwest Natl Lab, Biol Sci Div, Richland, WA 99352 USA..
    Trp53 deficient mice predisposed to preterm birth display region-specific lipid alterations at the embryo implantation site2016In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 6, article id 33023Article in journal (Refereed)
    Abstract [en]

    Here we demonstrate that conditional deletion of mouse uterine Trp53 (p53(d/d)), molecularly linked to mTORC1 activation and causally linked to premature uterine senescence and preterm birth, results in aberrant lipid signatures within the heterogeneous cell types of embryo implantation sites on day 8 of pregnancy. In situ nanospray desorption electrospray ionization mass spectrometry imaging (nano-DESI MSI) was used to characterize the molecular speciation of free fatty acids, monoacylglycerol species, unmodified and oxidized phosphatidylcholine (PC/Ox-PC), and diacylglycerol (DG) species within implantation sites of p53(d/d) mice and floxed littermates. Implantation sites from p53(d/d) mice exhibited distinct spatially resolved changes demonstrating accumulation of DG species, depletion of Ox-PC species, and increase in species with more unsaturated acyl chains, including arachidonic and docosahexaenoic acid. Understanding abnormal changes in the abundance and localization of individual lipid species early in the progression to premature birth is an important step toward discovering novel targets for treatments and diagnosis.

  • 225.
    Lanekoff, Ingela
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Laskin, Julia
    Imaging of lipids and metabolites using nanospray desorption electrospray ionization mass spectrometry.2015In: Methods in molecular biology, Springer, 2015, Vol. 1203, p. 99-106Chapter in book (Refereed)
    Abstract [en]

    Nanospray desorption electrospray ionization (nano-DESI) is an ambient ionization technique that uses localized liquid extraction for mass spectrometry imaging of molecules on surfaces. Nano-DESI enables imaging of ionizable molecules from a sample in its native state without any special sample pretreatment. In this chapter we describe the protocol for nano-DESI imaging of thin tissue sections.

  • 226.
    Lanekoff, Ingela
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Laskin, Julia
    Physical Sciences Division, Pacific Northwest National Laboratory, Richland, USA.
    Quantitative Mass Spectrometry Imaging of Molecules in Biological Systems2018In: Advances In Chromatography, Vol 54 / [ed] Grushka, E; Grinberg, N, Boca Raton: CRC Press, 2018, p. 43-72Chapter in book (Other academic)
  • 227.
    Laskin, Julia
    et al.
    Pacific Northwest National Laboratory.
    Andersson, Ingela
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Ambient Mass Spectrometry Imaging Using Direct Liquid Extraction Techniques2016In: Analytical Chemistry, ISSN 0003-2700, E-ISSN 1520-6882, Vol. 88, no 1, p. 52-73Article, review/survey (Refereed)
  • 228.
    Lee, Jae Kyoo
    et al.
    Stanford Univ, Dept Chem, 333 Campus Dr, Stanford, CA 94305 USA..
    Jansson, Erik T.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Stanford Univ, Dept Chem, 333 Campus Dr, Stanford, CA 94305 USA..
    Nam, Hong Gil
    Inst Basic Sci, Ctr Plant Aging Res, Daegu 42988, South Korea.;DGIST, Dept New Biol, Daegu 42988, South Korea..
    Zare, Richard N.
    Stanford Univ, Dept Chem, 333 Campus Dr, Stanford, CA 94305 USA..
    High-Resolution Live-Cell Imaging and Analysis by Laser Desorption/Ionization Droplet Delivery Mass Spectrometry2016In: Analytical Chemistry, ISSN 0003-2700, E-ISSN 1520-6882, Vol. 88, no 10, p. 5453-5461Article in journal (Refereed)
    Abstract [en]

    We have developed a new ambient-ionization mass spectrometric technique named laser desorption/ionization droplet delivery mass spectrometry (LDIDD-MS). LDIDD-MS permits high-resolution, high-sensitivity imaging of tissue samples as well as measurements of both single-cell apoptosis and live-cell exocytosis. A pulsed Hz) UV laser beam (266 nm) is focused on a surface covered with target analytes to trigger their desorption and ionization. A spray of liquid droplets is simultaneously directed onto the laser-focused surface region to capture the ionized analytes and deliver them to a mass spectrometer. The approach of rapid and effective capturing of molecules after laser desorption/ionization allows the limit of detection for the amino acid lysine to be as low as 2 amol under ambient ionization conditions. Two-dimensional maps of the desorbed/ionized species are recorded by moving the sample on an XY translational stage. The spatial resolution for imaging with LDIDD-MS was determined to be 2.4 mu m far an ink-printed pattern and 3 mu m for mouse brain tissue. We applied LDIDD-MS to single-cell analysis of apoptotic HEK cells. Differences were observed in the profiles of fatty acids and lipids between healthy HEK cells and those undergoing apoptosis. We observed upregulation of phosphatidylcholine (PC) with a relatively shorter carbon chain length and downregulation of PC with a relatively longer carbon chain length. We also applied LDIDD-MS for a real-time direct measurements of live-cell exocytosis. The catecholamine dopamine and trace amines (phenethylamine and tyramine) were detected from live PC12 cells without damaging them.

  • 229.
    Leito, Ivo
    et al.
    Univ Tartu, Inst Chem, Ravila 14a, EE-50411 Tartu, Estonia.
    Teearu, Anu
    Univ Tartu, Inst Chem, Ravila 14a, EE-50411 Tartu, Estonia.
    Bobacka, Johan
    Abo Akad Univ, Analyt Chem Lab, Johan Gadolin Proc Chem Ctr, Biskopsgatan 8, SF-20500 Turku, Finland.
    Randon, Jerome
    Univ Claude Bernard Lyon 1, CNRS, UMR 5280, Univ Lyon,Inst Sci Analyt, 5 Rue Doua, F-69100 Villeurbanne, France.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    EACH (Excellence in Analytical Chemistry), an Erasmus Mundus Joint Programme: progress and success2019In: Analytical and Bioanalytical Chemistry, ISSN 1618-2642, E-ISSN 1618-2650, Vol. 411, no 23, p. 5913-5921Article in journal (Other academic)
  • 230.
    Lennartsson, Anna-Karin
    et al.
    The Institute of Stress Medicine, Gothenburg AND The Department of Neuroscience and Physiology, Sahlgrenska Academy at the University of Gothenburg, Sweden.
    Kushnir, Mark M.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Billig, Håkan
    The Department of Neuroscience and Physiology, Sahlgrenska Academy at the University of Gothenburg, Sweden.
    Jonsdottir, Ingibjörg H
    The Institute of Stress Medicine, Gothenburg AND The Department of Neuroscience and Physiology, Sahlgrenska Academy at the University of Gothenburg, Sweden.
    Sex steroid levels temporarily increase in response to acute psychosocial stress in healthy men and women2012In: International Journal of Psychophysiology, ISSN 0167-8760, E-ISSN 1872-7697, Vol. 84, no 3, p. 246-253Article in journal (Refereed)
    Abstract [en]

    It is well known that acute psychosocial stress activates the hypothalamic-pituitary-adrenal (HPA) axis and the sympathetic nervous system (SNS). However, the effect of acute psychosocial stress on the hypothalamic-pituitary-gonadal (HPG) axis and levels of sex steroids are less known. The aim of this study was to investigate the effect of acute psychosocial stress on serum concentrations of sex steroids in healthy men and women. Twenty men and 19 women (age 30-50years) underwent Trier Social Stress Test (TSST), a tool for investigating psychobiological stress responses in a laboratory setting. Blood samples were collected before, directly after the stress test, and after 30 minutes of the recovery. Concentrations of androgens were measured with high specificity LC-MS/MS method; concentrations of cortisol, estradiol and sex hormone-binding globulin where determined using immunoassays. In both men and women we observed significantly elevated levels of testosterone, estradiol, androstenedione and sex hormone binding globulin along with significantly increased adrenocorticotropic hormone (ACTH), serum cortisol, heart rate, systolic blood pressure (SBP), and diastolic blood pressure (DBP) as a response to the stressor. Thus, even though the HPG axis and the production of sex steroids may be inhibited during prolonged periods of stress, the sex steroid levels may increase in the initial phase of the acute psychosocial stress.

  • 231. Lennartsson, Anna-Karin
    et al.
    Kushnir, Mark M.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Jonsdottir, Ingibjorg H.
    DHEA and DHEA-S response to acute psychosocial stress in healthy men and women2012In: Biological Psychology, ISSN 0301-0511, E-ISSN 1873-6246, Vol. 90, no 2, p. 143-149Article in journal (Refereed)
    Abstract [en]

    This study investigates the effect of acute psychosocial stress on serum concentrations of DHEA and DHEA-S in healthy men and women. Twenty men and 19 women (age 30-50 years) underwent Trier Social Stress Test (TSST). Physiological measurements were performed before, directly after the stress test and after 30 mins of recovery. In both men and women, significantly elevated DHEA and DHEA-S levels were observed in response to the stressor. There was a large inter-individual variation in the magnitude of the response, especially for DHEA but no statistical difference between men and women. Magnitude of the change in the levels of DHEA was found to be positively associated with the magnitude of the changes in ACTH, cortisol and heart rate. Furthermore, the results of this study suggest that the capacity to secrete DHEA and DHEA-S during acute psychosocial stress declines with age.

  • 232. Lennartsson, Anna-Karin
    et al.
    Theorell, Tores
    Kushnir, Mark M.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Bergquist, Jonas
    Jonsdottir, Ingibjorg H.
    Perceived stress at work is associated with attenuated DHEA-S response during acute psychosocial stress2013In: Psychoneuroendocrinology, ISSN 0306-4530, E-ISSN 1873-3360, Vol. 38, no 9, p. 1650-1657Article in journal (Refereed)
    Abstract [en]

    Background: Dehydroepiandrosterone (DHEA) and dehydroepiandrosterone sulfate (DHEA-S) have been suggested to play a protective role during acute psychosocial stress, because they act as antagonists to the effects of the stress hormone cortisol. This study aims to investigate whether prolonged psychosocial stress, measured as perceived stress at work during the past week, is related to the capacity to produce DHEA and DHEA-S during acute psychosocial stress. It also aims to investigate whether prolonged perceived stress affects the balance between production of cortisol and DHEA-S during acute psychosocial stress. Method: Thirty-six healthy subjects (19 men and 17 women, mean age 37 years, SD 5 years), were included. Perceived stress at work during the past week was measured by using the Stress-Energy (SE) Questionnaire. The participants were divided into three groups based on their mean scores; Low stress, Medium stress and High stress. The participants underwent the Trier Social Stress Test (TSST) and blood samples were collected before, directly after the stress test, and after 30 min of recovery. General Linear Models were used to investigate if the Medium stress group and the High stress group differ regarding stress response compared to the Low stress group. Results: Higher perceived stress at work was associated with attenuated DHEA-S response during acute psychosocial stress. Furthermore, the ratio between the cortisol production and the DHEA-S production during the acute stress test were higher in individuals reporting higher perceived stress at work compared to individuals reporting low perceived stress at work. There was no statistical difference in DHEA response between the groups. Conclusions: This study shows that prolonged stress, measured as perceived stress at work during the past week, seems to negatively affect the capacity to produce DHEA-S during acute stress. Given the protective functions of DHEA-S, attenuated DHEA-S production during acute stress may Lead to higher risk for adverse effects on psychological and physiological health, particularly if stress exposure continues.

  • 233.
    Li, Meishan
    et al.
    Department of Physiology and Pharmacology AND Department of Clinical Neuroscience, Clinical Neurophysiology, Karolinska Institutet, Sweden.
    Ogilvie, Hannah
    Department of Physiology and Pharmacology AND Department of Clinical Neuroscience, Clinical Neurophysiology, Karolinska Institutet, Sweden.
    Ochala, Julien
    Centre of Human and Aerospace Physiological Sciences, Faculty of Life Sciences and Medicine, King’s College London, UK.
    Artemenko, Konstantin A
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Iwamoto, Hiroyuki
    Japan Synchrotron Radiation Research Institute, Japan.
    Yagi, Naoto
    Japan Synchrotron Radiation Research Institute, Japan.
    Bergquist, Jonas
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Larsson, Lars
    Department of Physiology and Pharmacology AND Department of Clinical Neuroscience, Clinical Neurophysiology, Karolinska Institutet, Sweden.
    Aberrant post-translational modifications compromise human myosin motor function in old age2015In: Aging Cell, ISSN 1474-9718, E-ISSN 1474-9726, Vol. 14, no 2, p. 228-235Article in journal (Refereed)
    Abstract [en]

    Novel experimental methods, including a modified single fiber in vitro motility assay, X-ray diffraction experiments, and mass spectrometry analyses, have been performed to unravel the molecular events underlying the aging-related impairment in human skeletal muscle function at the motor protein level. The effects of old age on the function of specific myosin isoforms extracted from single human muscle fiber segments, demonstrated a significant slowing of motility speed (P < 0.001) in old age in both type I and IIa myosin heavy chain (MyHC) isoforms. The force-generating capacity of the type I and IIa MyHC isoforms was, on the other hand, not affected by old age. Similar effects were also observed when the myosin molecules extracted from muscle fibers were exposed to oxidative stress. X-ray diffraction experiments did not show any myofilament lattice spacing changes, but unraveled a more disordered filament organization in old age as shown by the greater widths of the 1, 0 equatorial reflections. Mass spectrometry (MS) analyses revealed eight age-specific myosin post-translational modifications (PTMs), in which two were located in the motor domain (carbonylation of Pro79 and Asn81) and six in the tail region (carbonylation of Asp900, Asp904, and Arg908; methylation of Glu1166; deamidation of Gln1164 and Asn1168). However, PTMs in the motor domain were only observed in the IIx MyHC isoform, suggesting PTMs in the rod region contributed to the observed disordering of myosin filaments and the slowing of motility speed. Hence, interventions that would specifically target these PTMs are warranted to reverse myosin dysfunction in old age.

  • 234.
    Lillja, Johan
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    MSn Structural Determination of Fatty AcidIsomers in Mixtures using Silver Cationization and Articial Neural Networks2018Independent thesis Advanced level (degree of Master (Two Years)), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    Fatty acid double bond positional isomers are expressed differently in healthy and diseased tissue. To utilize a mass spectrometry imaging approach to spatially map the distribution of fatty acids in tissue, a method needs to be developed to separate fatty acid isomers by the distribution of fragment ions. In this thesis, fatty acid isomers were cationized as silver ion adducts and subjected to MSn. The distribution of ions at MS3 was structurally informative, but no diagnostic ionswere present. An articial neural network was constructed that used the fragments of MS3 as inputs, to predict composite spectra. The network could classify fatty acid isomer standards in mixtures.

    The full text will be freely available from 2020-08-06 17:15
  • 235.
    Lind, Anne-Li
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Anaesthesiology and Intensive Care.
    Emami Khoonsari, Payam
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Cancer Pharmacology and Computational Medicine.
    Sjödin, Marcus
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Katila, Lenka
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Anaesthesiology and Intensive Care.
    Wetterhall, Magnus
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Gordh, Torsten
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Anaesthesiology and Intensive Care.
    Kultima, Kim
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Cancer Pharmacology and Computational Medicine.
    Spinal Cord Stimulation Alters Protein Levels in the Cerebrospinal Fluid of Neuropathic Pain Patients: A Proteomic Mass Spectrometric Analysis2016In: Neuromodulation (Malden, Mass.), ISSN 1094-7159, E-ISSN 1525-1403, Vol. 19, no 6, p. 549-562Article in journal (Refereed)
    Abstract [en]

    ObjectivesElectrical neuromodulation by spinal cord stimulation (SCS) is a well-established method for treatment of neuropathic pain. However, the mechanism behind the pain relieving effect in patients remains largely unknown. In this study, we target the human cerebrospinal fluid (CSF) proteome, a little investigated aspect of SCS mechanism of action. MethodsTwo different proteomic mass spectrometry protocols were used to analyze the CSF of 14 SCS responsive neuropathic pain patients. Each patient acted as his or her own control and protein content was compared when the stimulator was turned off for 48 hours, and after the stimulator had been used as normal for three weeks. ResultsEighty-six proteins were statistically significantly altered in the CSF of neuropathic pain patients using SCS, when comparing the stimulator off condition to the stimulator on condition. The top 12 of the altered proteins are involved in neuroprotection (clusterin, gelsolin, mimecan, angiotensinogen, secretogranin-1, amyloid beta A4 protein), synaptic plasticity/learning/memory (gelsolin, apolipoprotein C1, apolipoprotein E, contactin-1, neural cell adhesion molecule L1-like protein), nociceptive signaling (neurosecretory protein VGF), and immune regulation (dickkopf-related protein 3). ConclusionPreviously unknown effects of SCS on levels of proteins involved in neuroprotection, nociceptive signaling, immune regulation, and synaptic plasticity are demonstrated. These findings, in the CSF of neuropathic pain patients, expand the picture of SCS effects on the neurochemical environment of the human spinal cord. An improved understanding of SCS mechanism may lead to new tracks of investigation and improved treatment strategies for neuropathic pain.

  • 236.
    Lind, Anne-Li
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Anaesthesiology and Intensive Care.
    Yu, Di
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Freyhult, Eva
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Cancer Pharmacology and Computational Medicine.
    Bodolea, Constantin
    Department of Anaesthesia and Intensive Care, University of Medicine and Pharmacy, Cluj, Napoca, Romania..
    Ekegren, Titti
    Uppsala University, Disciplinary Domain of Science and Technology, Technology, Department of Engineering Sciences, Applied Materials Sciences.
    Larsson, Anders
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Biochemial structure and function.
    Gustafsson, Mats G
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Cancer Pharmacology and Computational Medicine.
    Katila, Lenka
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Anaesthesiology and Intensive Care.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Gordh, Torsten
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Anaesthesiology and Intensive Care.
    Landegren, Ulf
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Kamali-Moghaddam, Masood
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    A Multiplex Protein Panel Applied to Cerebrospinal Fluid Reveals Three New Biomarker Candidates in ALS but None in Neuropathic Pain Patients2016In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 11, no 2, article id e0149821Article in journal (Refereed)
    Abstract [en]

    The objective of this study was to develop and apply a novel multiplex panel of solid-phase proximity ligation assays (SP-PLA) requiring only 20 μL of samples, as a tool for discovering protein biomarkers for neurological disease and treatment thereof in cerebrospinal fluid (CSF). We applied the SP-PLA to samples from two sets of patients with poorly understood nervous system pathologies amyotrophic lateral sclerosis (ALS) and neuropathic pain, where patients were treated with spinal cord stimulation (SCS). Forty-seven inflammatory and neurotrophic proteins were measured in samples from 20 ALS patients and 15 neuropathic pain patients, and compared to normal concentrations in CSF from control individuals. Nineteen of the 47 proteins were detectable in more than 95% of the 72 controls. None of the 21 proteins detectable in CSF from neuropathic pain patients were significantly altered by SCS. The levels of the three proteins, follistatin, interleukin-1 alpha, and kallikrein-5 were all significantly reduced in the ALS group compared to age-matched controls. These results demonstrate the utility of purpose designed multiplex SP-PLA panels in CSF biomarker research for understanding neuropathological and neurotherapeutic mechanisms. The protein changes found in the CSF of ALS patients may be of diagnostic interest.

  • 237.
    Lind, Sara Bergström
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Artemenko, Konstantin A.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Elfineh, Lioudmila
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Cancer and Vascular Biology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Zhao, Yanhong
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Pettersson, Ulf
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Genomics. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Post translational modifications in adenovirus type 22013In: Virology, ISSN 0042-6822, E-ISSN 1096-0341, Vol. 447, no 1-2, p. 104-111Article in journal (Refereed)
    Abstract [en]

    We have combined 2-D SOS-PAGE with liquid chromatography-high resolving mass spectrometry (LC-MS) to explore the proteome of the adenovirus type 2 (Ad2) at the level of post translational modifications (PTMs). The experimental design included in-solution digestion, followed by titanium dioxide enrichment, as well as in-gel digestion of polypeptides after separation of Ad2 capsid proteins by 1-D and 2-D SOS-PAGE. All samples were analyzed using LC-MS with subsequent manual verification of PTM positions. The results revealed new phosphorylation sites that can explain the observed trains of protein spots observed for the pIII, pIIIa and ply proteins. The pin protein was found to be the most highly modified protein with now 18 verified sites of phosphorylation, three sites of nitrated tyrosine and one sulfated tyrosine. Nitrated tyrosines were also identified in pII. Lysine acetylations were detected in pII and pVI. The findings make the Ad2 virion much more complex than hitherto believed. 

  • 238.
    Lind, Sara Bergström
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Artemenko, Konstantin A.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Pettersson, Ulf
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Proteome Analysis of Adenovirus Using Mass Spectrometry2014In: Adenovirus: Methods and Protocols / [ed] Miguel Chillón, Assumpció Bosch, Springer Science+Business Media B.V., 2014, Vol. 1089, p. 25-44Chapter in book (Refereed)
    Abstract [en]

    Analysis of proteins and their posttranslational modifications is important for understanding different biological events. For analysis of viral proteomes, an optimal protocol includes production of a highly purified virus that can be investigated with a high-resolving analytical method. In this Methods in Molecular Biology paper we describe a working strategy for how structural proteins in the Adenovirus particle can be studied using liquid chromatography–high-resolving mass spectrometry. This method provides information on the chemical composition of the virus particle. Further, knowledge about amino acids carrying modifications that could be essential for any part of the virus life cycle is collected. We describe in detail alternatives available for preparation of virus for proteome analysis as well as choice of mass spectrometric instrumentation suitable for this kind of analysis.

  • 239.
    Lindegårdh, Niklas
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Dalarna University College, 781 88 Borlänge; and Uppsala University, Department of Analytical Chemistry, 751 21 Uppsala, Sweden.
    Blessborn, Daniel
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Dalarna University College, 781 88 Borlänge; and Uppsala University, Department of Analytical Chemistry, 751 21 Uppsala, Sweden.
    Bergqvist, Yngve
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Dalarna University College, 781 88 Borlänge; and Uppsala University, Department of Analytical Chemistry, 751 21 Uppsala, Sweden.
    Simultaneous quantitation of the highly lipophilic atovaquone and hydrophilic strong basic proguanil and its metabolites using a new mixed-mode SPE approach and steep-gradient LC2005In: Journal of Chromatographic Science, ISSN 0021-9665, E-ISSN 1945-239X, Vol. 43, no 5, p. 259-266Article in journal (Refereed)
    Abstract [en]

    A bioanalytical method is described for the simultaneous quantitative analysis of the highly lipophilic atovaquone and the strong basic proguanil with metabolites in plasma. The drugs are extracted from protein precipitated plasma samples on a novel mixed-mode solid-phase extraction (SPE) column containing carboxypropyl and octyl silica as functional groups. The analytes are further separated and quantitated using a steep-gradient liquid chromatographic method on a Zorbax SB-CN column with UV detection at 245 nm. Two different internal standards (IS) are used in the method to compensate for both types of analytes. A structurally similar IS to atovaquone is added with acetonitrile to precipitate proteins from plasma. A structurally similar IS to proguanil and its metabolites is added with phosphate buffer before samples are loaded onto the SPE columns. A single elution step is sufficient to elute all analytes. The method is validated according to published guidelines and shows excellent performance. The within-day precisions, expressed as relative standard deviation, are lower than 5% for all analytes at three tested concentrations within the calibration range. The between-day precisions are lower than 13% for all analytes at the same tested concentrations. The limit of quantitation is 25 nM for the basic substances and 50 nM for atovaquone. Several considerations regarding development and optimization of a method for determination of analytes with such a difference in physiochemical properties are discussed.

  • 240.
    Lindersson, Sebastian
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Method Development in Mass Spectrometry-based Proteomics for Determination of Early Pregnancy in Dogs2016Independent thesis Advanced level (degree of Master (Two Years)), 30 credits / 45 HE creditsStudent thesis
    Abstract [en]

    This project is concerned with method development in mass spectrometry (MS)-based proteomics in order to find putative biomarkers for early pregnancy ofdomesticated dogs. It is of importance for dog breeders to know whether the dogsbecome pregnant post-mating. Unlike humans, dogs are not known to possess aspecific hormone that indicates fetal development; therefore other biomarkers mustbe investigated. The approach of choice in this project was to look at proteins throughMS-based proteomics. For this purpose, serum samples from 11 pregnant dogs (case,different breeds) and 7 non-pregnant dogs (control, all beagle dogs) were sampledbefore-hand at the Swedish University of Agricultural Sciences. Each dog wassampled Day 1, Day 8, Day 15, Day 22 and Day 29 after optimal mating. Twodifferent proteomics approaches were conducted: Bottom-up (“Shotgun”) proteomicsand targeted proteomics (“targeted analysis”). In this study, Label-free Quantification(LFQ) was employed, which is a relative quantitative technique. The massspectrometer of choice was the Quadrupole-Orbitrap QExactive plus massspectrometer coupled to a nano-Ultra Performance Liquid Chromatography (UPLC).Method optimization was done with respect to concentration of samples prior to MSanalysis, as well as different LC-gradients. From shotgun screening experiments, itwas possible to identify 252 proteins. Ultimately, 9 proteins were investigated usingtargeted final analysis: CRP, SERPINC1, CP, PROS1, SERPING1, A2M, AGP,SERPINA1 and HP. For targeted final analysis, 21 peptides were considered.Calibration curves were constructed using 8 of the 21 targeted peptides; 1 peptide perprotein, except for HP which had 2 peptides per protein. The SERPINA1 and CPproteins had no appropriate peptides for targeted final analysis and were thusexcluded. It was confirmed that CRP was up-regulated in case dogs compared tocontrol dogs. The other investigatedproteins showed no significant signs of regulation. In order to improve the results; itwould be desirable to include more dogs in the study which would benefit thestatistics of protein regulation. However, the use of isotopic labeled standards andemployment of a Parallel Reaction Monitoring (PRM) method should be prioritizedfor obtaining absolute quantitative data.

  • 241.
    Lindgren, Fredrik
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström.
    Rehnlund, David
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström, Inorganic Chemistry. Karlsruhe Inst Technol, Dept Appl Biol, Inst Appl Biosci, Kaiserstr 12, D-76131 Karlsruhe, Germany.
    Pan, Ruijun
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström.
    Pettersson, Jean
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Younesi, Reza
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström, Structural Chemistry.
    Xu, Chao
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström, Structural Chemistry. Univ Cambridge, Dept Chem, Cambridge CB2 1EW, England.
    Gustafsson, Torbjörn
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström, Structural Chemistry.
    Edström, Kristina
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström, Structural Chemistry.
    Nyholm, Leif
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström, Inorganic Chemistry.
    On the Capacity Losses Seen for Optimized Nano-Si Composite Electrodes in Li-Metal Half-Cells2019In: Advanced Energy Materials, ISSN 1614-6832, Vol. 9, no 33, article id 1901608Article in journal (Refereed)
    Abstract [en]

    While the use of silicon‐based electrodes can increase the capacity of Li‐ion batteries considerably, their application is associated with significant capacity losses. In this work, the influences of solid electrolyte interphase (SEI) formation, volume expansion, and lithium trapping are evaluated for two different electrochemical cycling schemes using lithium‐metal half‐cells containing silicon nanoparticle–based composite electrodes. Lithium trapping, caused by incomplete delithiation, is demonstrated to be the main reason for the capacity loss while SEI formation and dissolution affect the accumulated capacity loss due to a decreased coulombic efficiency. The capacity losses can be explained by the increasing lithium concentration in the electrode causing a decreasing lithiation potential and the lithiation cut‐off limit being reached faster. A lithium‐to‐silicon atomic ratio of 3.28 is found for a silicon electrode after 650 cycles using 1200 mAhg−1 capacity limited cycling. The results further show that the lithiation step is the capacity‐limiting step and that the capacity losses can be minimized by increasing the efficiency of the delithiation step via the inclusion of constant voltage delithiation steps. Lithium trapping due to incomplete delithiation consequently constitutes a very important capacity loss phenomenon for silicon composite electrodes.

  • 242.
    Lindén, Mårten
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Urology.
    Segersten, Ulrika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Urology.
    Runeson, Marcus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Molecular and Morphological Pathology.
    Wester, Kenneth
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Busch, Christer
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Pettersson, Ulf
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Genomics.
    Bergström Lind, Sara
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Malmström, Per-Uno
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Urology.
    Tumour expression of bladder cancer-associated urinary proteins2013In: BJU International, ISSN 1464-4096, E-ISSN 1464-410X, Vol. 112, no 3, p. 407-415Article in journal (Refereed)
    Abstract [en]

    WHAT'S KNOWN ON THE SUBJECT? AND WHAT DOES THE STUDY ADD?:

    • The current basis for diagnosis and prognosis in urinary bladder cancer is based on the pathologists' assessment of a biopsy of the tumour. Urinary biomarkers are preferable as they can be non-invasively sampled. Urinary cytology is the only test with widespread use but is hampered by poor reproducibility and low sensitivity.
    • By studying the protein expression in bladder tumour tissue samples of proteins previously found in elevated levels in the urine of patients with bladder cancer, we have been able to show that these proteins originate from the tumour. The immunoreactivity of three of the investigated proteins increased with higher stage. Also a serine peptidase inhibitor was found to be predictive of progression from non-muscle-invasive to muscle-invasive tumours.

    OBJECTIVES:

    • To analyse the expression of five bladder cancer-associated urinary proteins and investigate if expression is related to the malignant phenotype of the tumour.
    • To explore the possible prognostic value of these proteins.

    PATIENTS AND METHODS:

    • Urine samples, 16 from patients with bladder cancer and 26 from controls, were used in Western Blotting experiments.
    • Tissue microarrays with bladder tissue from 344 patients diagnosed with bladder cancer between 1984 and 2005 was used in immunohistochemistry experiments.
    • The proteins apolipoprotein E (APOE), fibrinogen β chain precursor (FGB), leucine-rich α2-glycoprotein (LRG1), polymerase (RNA) I polypeptide E (POLR1E), α1-antitrypsin (SERPINA1) and topoisomerase 2A (TOP2A) were probed with antibodies validated by the Human Protein Atlas.

    RESULTS:

    • Increased expressions of APOE, FGB and POLR1E were correlated with increased tumour stage (P < 0.001).
    • Expression of SERPINA1 in Ta and T1 tumours was found to increase the risk of tumour progression (hazard ratio 2.57, 95% confidence interval 1.13-5.87; P = 0.025)

    CONCLUSIONS:

    • All proteins previously detected in urine from patients with bladder cancer were also expressed in bladder cancer tissue.
    • The expression of APOE, FGB and POLR1E increased with stage and they are potential diagnostic markers.
    • SERPINA1 was identified as a prognostic marker candidate.
  • 243. Liu, Dan
    et al.
    Ma, Yan
    Gu, Ming
    Janson, Jan-Christer
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Wang, Changhai
    Xiao, Hongbin
    Liquid-liquid/solid three-phase high-speed counter-current chromatography, a new technique for separation of polyphenols from Geranium wilfordii Maxim2012In: Journal of Separation Science, ISSN 1615-9306, E-ISSN 1615-9314, Vol. 35, no 16, p. 2146-2151Article in journal (Refereed)
    Abstract [en]

    High-speed counter-current chromatography using a new liquidliquid/solid three-phase system was used for the separation of the polyphenols corilagin and geraniin from a crude extract of Geranium wilfordii Maxim in one step. The optimized three-phase system was composed of n-hexane/ethyl acetate/methanol/acetic acid/water and to which was added 10-mu m average diameter microspheres of cross-linked 12% agarose at the ratio of 0.2:10:2:1:5 and 0.1 g/mL, respectively. The purities of geraniin and corilagin were 82 and 90%, which were determined by HPLC at 280 nm. A 14.5 and 7 mg of geraniin and corilagin were purified from 160 mg crude extract with the yields of 70 and 78%, respectively.

  • 244.
    Liu, Fang
    et al.
    Binzhou Med Univ, Med & Pharm Res Ctr, Yantai 264003, Shandong, Peoples R China.;Binzhou Med Univ, Dept Radiol, Affiliated Hosp, Binzhou 256603, Shandong, Peoples R China..
    Zhang, Yuan
    Binzhou Med Univ, Med & Pharm Res Ctr, Yantai 264003, Shandong, Peoples R China..
    Men, Tingting
    Binzhou Med Univ, Med & Pharm Res Ctr, Yantai 264003, Shandong, Peoples R China..
    Jiang, Xingyue
    Binzhou Med Univ, Dept Radiol, Affiliated Hosp, Binzhou 256603, Shandong, Peoples R China..
    Yang, Chunhua
    Binzhou Med Univ, Med & Pharm Res Ctr, Yantai 264003, Shandong, Peoples R China..
    Li, He
    Binzhou Med Univ, Yantai Affiliated Hosp, Dept Gastr & Intestine, Yantai 264003, Shandong, Peoples R China..
    Wei, Xiaodan
    Binzhou Med Univ, Med & Pharm Res Ctr, Yantai 264003, Shandong, Peoples R China..
    Yan, Dong
    Binzhou Med Univ, Med & Pharm Res Ctr, Yantai 264003, Shandong, Peoples R China..
    Feng, Gangming
    China Agr Univ, Yantai Inst, Yantai 264670, Shandong, Peoples R China..
    Yang, Jianke
    Binzhou Med Univ, Med & Pharm Res Ctr, Yantai 264003, Shandong, Peoples R China..
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Wang, Bin
    Binzhou Med Univ, Dept Radiol, Affiliated Hosp, Binzhou 256603, Shandong, Peoples R China..
    Jiang, Wenguo
    Binzhou Med Univ, Med & Pharm Res Ctr, Yantai 264003, Shandong, Peoples R China..
    Mi, Jia
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Binzhou Med Univ, Med & Pharm Res Ctr, Yantai 264003, Shandong, Peoples R China..
    Tian, Geng
    Binzhou Med Univ, Med & Pharm Res Ctr, Yantai 264003, Shandong, Peoples R China..
    Quantitative proteomic analysis of gastric cancer tissue reveals novel proteins in platelet-derived growth factor B signaling pathway2017In: OncoTarget, ISSN 1949-2553, E-ISSN 1949-2553, Vol. 8, no 13, p. 22059-22075Article in journal (Refereed)
    Abstract [en]

    Gastric cancer is one of the most common cancers in Asian countries. Searching for reliable biomarkers involving the development of gastric cancer is important for clinical practice. Quantitative proteomics has become an important method contributed to the discovery of novel diagnostic or therapeutic targets for the management of cancer. Here, we identified differently expressed proteins in gastric cancer and normal gastric tissues by using the high resolution mass spectrometer. Among the total of 2280 identified proteins, 87 were differentially expressed between gastric cancer and normal gastric tissues. Notably, several significant proteins are in the PDGF-B signaling pathway, including peroxiredoxin5 (PRDX5), S100A6, calreticulin (CALR) and cathepsin D (CTSD), which were validated by western blot. Furthermore, upstream regulators including PDGF-B, PDGFR-beta, Akt, eIF4E and p70s6K were found significantly increased in the gastric cancer tissues. In addition, silencing of PRDX5 and PDGF-B suppressed the proliferation of gastric cancer cells in vitro. The administration of exogenous PDGF-BB recovered the reduced expression of PDGF-B signaling pathway in PDGF-B knockdown cells. Taken together, our findings suggested that PDGF-B signaling pathway plays an important role in the regulation of gastric cancer proliferation and the inhibition of this pathway may be a potential approach for treatment of gastric cancer.

  • 245. Liu, Jiayin
    et al.
    Sandahl, Margareta
    Sjöberg, Per J. R.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Turner, Charlotta
    Pressurised hot water extraction in continuous flow mode for thermolabile compounds: extraction of polyphenols in red onions2014In: Analytical and Bioanalytical Chemistry, ISSN 1618-2642, E-ISSN 1618-2650, Vol. 406, no 2, p. 441-445Article in journal (Refereed)
    Abstract [en]

    Extraction and analysis of labile compounds in complex sample matrices, such as plants, is often a big analytical challenge. In this work, the use of a "green and clean" pressurised hot water extraction (PHWE) approach performed in continuous flow mode is explored. Experimental data for extraction and degradation kinetics of selected compounds were utilised to develop a continuous flow extraction (CFE) method targeting thermolabile polyphenols in red onions, with detection by high-performance liquid chromatography (HPLC)-diode array detection (DAD)-mass spectrometry (MS). Water containing ethanol and formic acid was used as extraction solvent. Method performance was focused on extraction yield with minimal analyte degradation. By adjusting the flow rate of the extraction solvent, degradation effects were minimised, and complete extraction could be achieved within 60 min. The CFE extraction yields of the polyphenols investigated were 80-90 % of the theoretically calculated quantitative yields and were significantly higher than the yields obtained by conventional methanol extraction and static batch extraction (70-79 and 58-67% of the theoretical yields, respectively). The precision of the developed method was lower than 8 % expressed as relative standard deviation.

  • 246.
    Ljungvall, Karl
    et al.
    Swedish Univ Agr Sci, Div Reprod, Dept Clin Sci, Uppsala, Sweden..
    Magnusson, Ulf
    Swedish Univ Agr Sci, Div Reprod, Dept Clin Sci, Uppsala, Sweden..
    Korvela, Marcus
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Norrby, Mattias
    Swedish Univ Agr Sci, Div Reprod, Dept Clin Sci, Uppsala, Sweden..
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Persson, Sara
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Heavy metal concentrations in female wild mink (Neovison vison) in Sweden: Sources of variation and associations with internal organ weights2017In: Environmental Toxicology and Chemistry, ISSN 0730-7268, E-ISSN 1552-8618, Vol. 36, no 8, p. 2030-2035Article in journal (Refereed)
    Abstract [en]

    The American mink is an invasive species in Sweden, and it is legally hunted all year. Therefore, the mink is well suited as a sentinel species for environmental monitoring. In the present study female mink (n = 91) from 6 different areas in Sweden were analyzed for the concentrations of silver, cadmium, mercury and lead in liver tissue using inductively coupled plasma mass spectrometry. The wet concentrations in liver tissue were 42.6 +/- 52.7 ng/g for silver, 99.5 +/- 100 ng/g for cadmium, 652 +/- 537 ng/g formercury, and 196 +/- 401 ng/g for lead (expressed as mean + standard deviation). There were associations between the sample area and the concentrations of silver, lead, and mercury. The concentrations of lead and cadmium varied with season of capture and lead, cadmium, and mercury were positively associated with increasing age. Relative liver weight was positively associated with concentrations of mercury and negatively associated with lead and cadmium. Relative kidney weight was negatively associated with lead concentrations. In summary, it is of importance to take age and season of capture into account when assessing levels of heavy metals in wild mink. Also, liver and kidneys seem to be potential targets for heavy metal toxicity in wild female mink in Sweden.

  • 247.
    Llano-Diez, Monica
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Clinical Neurophysiology.
    Renaud, Guillaume
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Clinical Neurophysiology.
    Andersson, Magnus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurosurgery.
    Gonzales Marrero, Humberto
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Clinical Neurophysiology.
    Cacciani, Nicola
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Clinical Neurophysiology.
    Engquist, Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurosurgery.
    Corpeno, Rebeca
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Clinical Neurophysiology.
    Artemenko, Konstantin
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Larsson, Lars
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Clinical Neurophysiology.
    Mechanisms underlying intensive care unit muscle wasting and effects of passive mechanical loading2012In: Critical Care, ISSN 1364-8535, E-ISSN 1466-609X, Vol. 16, no 5, p. R209-Article in journal (Refereed)
    Abstract [en]

    ABSTRACT: INTRODUCTION: Critical ill intensive care unit (ICU) patients commonly develop severe muscle wasting and impaired muscle function, leading to delayed recovery, with subsequent increased morbidity and financial costs, and decreased quality of life of survivors. Critical illness myopathy (CIM) is a frequently observed neuromuscular disorder in ICU patients. Sepsis, systemic corticosteroid hormone treatment and post-synaptic neuromuscular blockade have been forwarded as the dominating triggering factors. Recent experimental results from our group using a unique experimental rat ICU model have shown that the "mechanical silencing" associated with the ICU condition is the primary triggering factor. This study aims at (1) unraveling the mechanisms underlying CIM, and (2) evaluating the effects of a specific intervention aiming at reducing the mechanical silencing in sedated and mechanically ventilated ICU patients. METHODS: Muscle gene/protein expression, post-translational modifications (PTMs), muscle membrane excitability, muscle mass measurements, and contractile properties at the single muscle fiber level were explored in seven deeply sedated and mechanically ventilated ICU patients (not exposed to systemic corticosteroid hormone treatment, post-synaptic neuromuscular blockade or sepsis) subjected to unilateral passive mechanical loading 10 hours per day (2.5 hours, 4 times) for 9 +/- 1 days. RESULTS: These patients developed a phenotype considered pathognomonic of CIM, i.e., severe muscle wasting and a preferential myosin loss (P<0.001). In addition, myosin PTMs specific to the ICU condition were observed in parallel with an increased sarcolemmal expression and cytoplasmic translocation of nNOS. Passive mechanical loading for 9 +/- 1 resulted in a 35% higher specific force (P<0.001) compared with the unloaded leg, although it was not sufficient to prevent the loss of muscle mass. CONCLUSIONS: Mechanical silencing is suggested to be a primary mechanism underlying CIM, i.e., triggering the myosin loss, muscle wasting and myosin PTMs. The higher nNOS expression found in the ICU patients and its cytoplasmic translocation are forwarded as a probable mechanism underlying these modifications. The positive effect of passive loading on muscle fiber function strongly supports the importance of early physical therapy and mobilization in deeply sedated and mechanically ventilated ICU patients.

  • 248.
    Loeffler, Christopher R.
    et al.
    US FDA, Div Seafood Sci & Technol, Dauphin Isl, AL 36528 USA;German Fed Inst Risk Assessment, Contaminants, Max Dohrn Str 8-10, D-10589 Berlin, Germany;Univ Napoli Federico II, Sch Med & Surg, Dept Pharm, Via D Montesano 49, I-80131 Naples, Italy.
    Handy, Sara M.
    US FDA, Off Regulatory Sci, College Pk, MD 20740 USA.
    Quintana, Harold A. Flores
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. US FDA, Div Seafood Sci & Technol, Dauphin Isl, AL 36528 USA.
    Deeds, Jonathan R.
    US FDA, Off Regulatory Sci, College Pk, MD 20740 USA.
    Fish Hybridization Leads to Uncertainty Regarding Ciguatera Fish Poisoning Risk; Confirmation of Hybridization and Ciguatoxin Accumulation with Implications for Stakeholders2019In: Journal of Marine Science and Engineering, E-ISSN 2077-1312, Vol. 7, no 4, article id 105Article in journal (Refereed)
    Abstract [en]

    Globally, ciguatera fish poisoning (CFP) avoidance efforts rely primarily on local knowledge of the fish being consumed, its collection location, and association with illnesses. In 2016, several fish that appeared to be hybrids between a local commercially prized species, Ocyurus chrysurus, and a regionally prohibited species Lutjanus apodus (due to CFP concerns), were caught nearshore in United States Virgin Islands waters, leading to confusion regarding the safety of consuming the fish. The hybrid status of the fish was verified as O. chrysurus (male) x L. apodus (female) by comparing two sets of gene sequences (mitochondrial CO1 and nuclear S7). Using an in vitro mouse neuroblastoma (N2a) assay, one of the hybrid fish exhibited a composite cytotoxicity of 0.038 ppb Caribbean ciguatoxin-1 (C-CTX-1) equivalents (Eq.); a concentration below the US Food and Drug Administration (FDA) guidance level for safety in fish products for CFP (0.1 ppb C-CTX-1 Eq.) but approximately 2x above the maximum described in the commercially prized parent species (0.019 ppb C-CTX-1 Eq./g). C-CTX-1 was confirmed in the hybrid sample by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The second hybrid fish tested negative for CTXs. This research confirms hybridization between two species with contrasting commercial statuses, discusses CTX accumulation implications for hybridization, and provides a methodology for future studies into novel CFP vectors, with the goal of providing critical information for fishermen and consumers regarding CFP risk management.

  • 249.
    Loov, Camilla
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurosurgery.
    Shevchenko, Ganna
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Nadadhur, Aishwarya Geeyarpuram
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurosurgery.
    Clausen, Fredrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurosurgery.
    Hillered, Lars
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurosurgery.
    Wetterhall, Magnus
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Erlandsson, Anna
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurosurgery.
    Identification of Injury Specific Proteins in a Cell Culture Model of Traumatic Brain Injury2013In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 8, no 2, p. e55983-Article in journal (Refereed)
    Abstract [en]

    The complicated secondary molecular and cellular mechanisms following traumatic brain injury (TBI) are still not fully understood. In the present study, we have used mass spectrometry to identify injury specific proteins in an in vitro model of TBI. A standardized injury was induced by scalpel cuts through a mixed cell culture of astrocytes, oligodendrocytes and neurons. Twenty-four hours after the injury, cell culture medium and whole-cell fractions were collected for analysis. We found 53 medium proteins and 46 cell fraction proteins that were specifically expressed after injury and the known function of these proteins was elucidated by an extensive literature survey. By using time-lapse microscopy and immunostainings we could link a large proportion of the proteins to specific cellular processes that occur in response to trauma; including cell death, proliferation, lamellipodia formation, axonal regeneration, actin remodeling, migration and inflammation. A high percentage of the proteins uniquely expressed in the medium after injury were actin-related proteins, which normally are situated intracellularly. We show that two of these, ezrin and moesin, are expressed by astrocytes both in the cell culture model and in mouse brain subjected to experimental TBI. Interestingly, we found many inflammation-related proteins, despite the fact that cells were present in the culture. This study contributes with important knowledge about the cellular responses after trauma and identifies several potential cell-specific biomarkers.

  • 250.
    Lough, Alastair J. M.
    et al.
    Univ Southampton, Natl Oceanog Ctr Southampton, Ocean & Earth Sci, Southampton, Hants, England;Natl Oceanog Ctr, European Way, Southampton, Hants, England.
    Connelly, Douglas P.
    Natl Oceanog Ctr, European Way, Southampton, Hants, England.
    Homoky, William B.
    Univ Oxford, Dept Earth Sci, Oxford, England.
    Hawkes, Jeffrey A.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Univ Southampton, Natl Oceanog Ctr Southampton, Ocean & Earth Sci, Southampton, Hants, England.
    Chavagnac, Valerie
    Univ Toulouse, GET CNRS UMR5563, Geosci Environement Toulouse, Toulouse, France.
    Castillo, Alain
    Univ Toulouse, GET CNRS UMR5563, Geosci Environement Toulouse, Toulouse, France.
    Kazemian, Majid
    Diamond Light Source Ltd, Didcot, Oxon, England.
    Nakamura, Ko-ichi
    Natl Inst Adv Ind Sci & Technol, Tsukuba, Ibaraki, Japan.
    Araki, Tohru
    Diamond Light Source Ltd, Didcot, Oxon, England.
    Kaulich, Burkhard
    Diamond Light Source Ltd, Didcot, Oxon, England.
    Mills, Rachel A.
    Univ Southampton, Natl Oceanog Ctr Southampton, Ocean & Earth Sci, Southampton, Hants, England.
    Diffuse Hydrothermal Venting: A Hidden Source of Iron to the Oceans2019In: Frontiers in Marine Science, E-ISSN 2296-7745, Vol. 6, article id 329Article in journal (Refereed)
    Abstract [en]

    Iron (Fe) limits primary productivity and nitrogen fixation in large regions of the world's oceans. Hydrothermal supply of Fe to the global deep ocean is extensive; however, most of the previous work has focused on examining high temperature, acidic, focused flow on ridge axes that create "black smoker" plumes. The contribution of other types of venting to the global ocean Fe cycle has received little attention. To thoroughly understand hydrothermal Fe sources to the ocean, different types of vent site must be compared. To examine the role of more diffuse, higher pH sources of venting, a hydrothermal plume above the Von Damm vent field (VDVF) was sampled for Total dissolvable Fe (unfiltered, TDFe), dissolved Fe (< 0.2 mu m, dFe) and soluble Fe (< 0.02 mu m, sFe). Plume particles sampled in situ were characterized using scanning electron microscopy and soft X-ray spectromicroscopy. The VDVF vents emit visibly clear fluids with particulate Fe (TDFe-dFe, > 0.2 mu m) concentrations up to 196 nmol kg(-1) comparable to concentrations measured in black smoker plumes on the Mid-Atlantic Ridge. Colloidal Fe (cFe) and sFe increased as a fraction of TDFe with decreasing TDFe concentration. This increase in the percentage of sFe and cFe within the plume cannot be explained by settling of particulates or mixing with background seawater. The creation of new cFe and sFe within the plume from the breakdown of pFe is required to close the Fe budget. We suggest that the proportional increase in cFe and sFe reflects the entrainment, breakdown and recycling of Fe bearing organic particulates near the vents. Fe plume profiles from the VDVF differ significantly from previous studies of "black smoker" vents where formation of new pFe in the plume decreases the amount of cFe. Formation and removal of Fe-rich colloids and particles will control the amount and physico-chemical composition of dFe supplied to the deep ocean from hydrothermal systems. This study highlights the differences in the stabilization of hydrothermal Fe from an off-axis diffuse source compared to black smokers. Off-axis diffuse venting represent a potentially significant and previously overlooked Fe source to the ocean due to the difficulties in detecting and locating such sites.

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