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  • 351.
    Frisk, Jun Mei Hu
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Kjellén, Lena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Melo, Fabio R.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Öhrvik, Helena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anat Physiol & Biochem, Uppsala, Sweden.
    Mitogen-Activated Protein Kinase Signaling Regulates Proteoglycan Composition of Mast Cell Secretory Granules2018In: Frontiers in Immunology, ISSN 1664-3224, E-ISSN 1664-3224, Vol. 9, article id 1670Article in journal (Refereed)
    Abstract [en]

    Mast cells (MCs) are characterized by an abundance of lysosome-like secretory granules filled with immunomodulatory compounds including histamine, cytokines, lysosomal hydrolases, MC-restricted proteases, and serglycin proteoglycans. The latter are essential for promoting the storage of other granule compounds and are built up of the serglycin core protein to which highly sulfated and thereby negatively charged glycosaminoglycan (GAG) side chains of heparin or chondroitin sulfate type are attached. In the search for mechanisms operating in regulating MC granule homeostasis, we here investigated the role of mitogen-activated protein kinase (MAPK) signaling. We show that inhibition of MEK1/2 (a MAPK kinase) leads to increased metachromatic staining of MC granules, indicative of increased proteoglycan content. Indeed, MEK1/2 inhibition caused a profound increase in the expression of the gene coding for the serglycin core protein and of genes coding for various enzymes involved in the biosynthesis/sulfation of the GAGs attached to the serglycin core protein. This was accompanied by corresponding increases in the levels of the respective GAGs. Deletion of the serglycin core protein abrogated the induction of enzymes operative in proteoglycan synthesis, indicating that availability of the serglycin proteoglycan core protein has a regulatory function impacting on the expression of the various serglycin-modifying enzymes. MEK1/2 inhibition also caused a substantial increase in the expression of granule-localized, proteoglycan-binding proteases. Altogether, this study identifies a novel role for MAPK signaling in regulating the content of secretory granules in MCs.

  • 352.
    Fritsch Fredin, Maria
    et al.
    Astra-Zeneca, Mölndal.
    Hultin, Leif
    Astra-Zeneca, Mölndal.
    Hyberg, Gina
    Astra-Zeneca, Mölndal.
    Rehnström, Erika
    Astra-Zeneca, Mölndal.
    Hultgren Hörnquist, Elisabeth
    Örebro University, School of Health and Medical Sciences.
    Melgar, Silvia
    Astra-Zeneca, Mölndal.
    Jansson, Liselotte
    Astra-Zeneca, Mölndal.
    Predicting and monitoring colitis development in mice by micro-computed tomography2008In: Inflammatory Bowel Diseases, ISSN 1078-0998, E-ISSN 1536-4844, Vol. 14, no 4, p. 491-499Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Computed tomography (CT) has been developed as a tool for monitoring human inflammatory bowel disease (IBD). The aim of this study was to evaluate colon wall thickness as a noninvasive marker in the dextran sodium sulfate (DSS) mouse model of colitis using micro-CT. METHODS: Mice were examined by micro-CT 1, 2, or 4 times between day 0 (d0) and d26 after induction of colitis to document the kinetics of changes in colon wall thickness and its relation to colitis development. RESULTS: DSS-treated mice displayed a significantly thicker colon wall at all timepoints (days 5, 8, 12, 19, and 26) investigated compared to healthy controls. Colon wall thickness showed a good correlation to the macroscopic grading of colitis (r = 0.81). The increase in colon wall thickness occurred mainly during the acute phase of colitis (between days 5 and 12) and did not progress much further in the chronic phase of colitis (d26). Colon wall thickness at d26 was thereby predicted by measurements at d12. All mice did not respond equally to DSS and this difference was manifest during the first 2 weeks of colitis, providing an important tool in stratifying responders from nonresponders. CONCLUSIONS: While the potential impact of handling and anesthesia should be considered on repeated micro-CT, irradiation exposure during repeated micro-CT did not affect the development of colitis. Thus, the results suggest that micro-CT can be used for monitoring and prediction of the inflammatory response in mouse colitis in future therapeutic studies.

  • 353.
    Fritz, Michael
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Center for Social and Affective Neuroscience. Linköping University, Faculty of Medicine and Health Sciences.
    Klawonn, Anna
    Linköping University, Department of Clinical and Experimental Medicine, Center for Social and Affective Neuroscience. Linköping University, Faculty of Medicine and Health Sciences.
    Jaarola, Maarit
    Linköping University, Department of Clinical and Experimental Medicine, Center for Social and Affective Neuroscience. Linköping University, Faculty of Medicine and Health Sciences.
    Engblom, David
    Linköping University, Faculty of Medicine and Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Center for Social and Affective Neuroscience.
    Interferon-ɣ mediated signaling in the brain endothelium is critical for inflammation-induced aversion2018In: Brain, behavior, and immunity, ISSN 0889-1591, E-ISSN 1090-2139, Vol. 67, p. 54-58Article in journal (Refereed)
    Abstract [en]

    Systemic inflammation elicits malaise and a negative affective state. The mechanism underpinning the aversive component of inflammation include cerebral prostaglandin synthesis and modulation of dopaminergic reward circuits, but the messengers that mediate the signaling between the peripheral inflammation and the brain have not been sufficiently characterized. Here we investigated the role of interferon-ɣ (IFN-ɣ) in the aversive response to systemic inflammation induced by a low dose (10μg/kg) of lipopolysaccharide (LPS) in mice. LPS induced IFN-ɣ expression in the blood and deletion of IFN-ɣ or its receptor prevented the development of conditioned place aversion to LPS. LPS induced expression of the chemokine Cxcl10 in the striatum of normal mice, but this induction was absent in mice lacking IFN-ɣ receptors or Myd88 in blood brain barrier endothelial cells. Furthermore, inflammation-induced aversion was blocked in mice lacking Cxcl10 or its receptor Cxcr3. Finally, mice with a selective deletion of the IFN-ɣ receptor in brain endothelial cells did not develop inflammation-induced aversion, demonstrating that the brain endothelium is the critical site of IFN-ɣ action. Collectively, these findings show that circulating IFN-ɣ that binds to receptors on brain endothelial cells and induces Cxcl10, is a central link in the signaling chain eliciting inflammation-induced aversion.

  • 354.
    Frodlund, M.
    et al.
    Linkoping Univ, Div Neuro & Inflammat Sci, Dept Clin & Expt Med, Linkoping, Sweden.
    Vikerfors, A.
    Swedish Med Prod Agcy, SE-75103 Uppsala, Sweden;Karolinska Univ Hosp, Karolinska Inst, Dept Med Solna, Unit Rheumatol, Stockholm, Sweden.
    Grosso, G.
    Karolinska Univ Hosp, Karolinska Inst, Dept Med Solna, Unit Rheumatol, Stockholm, Sweden.
    Skogh, T.
    Linkoping Univ, Div Neuro & Inflammat Sci, Dept Clin & Expt Med, Linkoping, Sweden.
    Wetterö, J.
    Linkoping Univ, Div Neuro & Inflammat Sci, Dept Clin & Expt Med, Linkoping, Sweden.
    Elvin, K.
    Karolinska Inst, Dept Clin Immunol & Transfus Med, Unit Clin Immunol, Stockholm, Sweden.
    Gunnarsson, I.
    Karolinska Univ Hosp, Karolinska Inst, Dept Med Solna, Unit Rheumatol, Stockholm, Sweden.
    Kastbom, A.
    Linkoping Univ, Div Neuro & Inflammat Sci, Dept Clin & Expt Med, Linkoping, Sweden.
    Dahlström, Ö.
    Linkoping Univ, Swedish Inst Disabil Res, Dept Behav Sci & Learning, Linkoping, Sweden.
    Rönnelid, Johan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Svenungsson, E.
    Karolinska Univ Hosp, Karolinska Inst, Dept Med Solna, Unit Rheumatol, Stockholm, Sweden.
    Sjöwall, C.
    Linkoping Univ, Div Neuro & Inflammat Sci, Dept Clin & Expt Med, Linkoping, Sweden.
    Immunoglobulin A anti-phospholipid antibodies in Swedish cases of systemic lupus erythematosus: associations with disease phenotypes, vascular events and damage accrual2018In: Clinical and Experimental Immunology, ISSN 0009-9104, E-ISSN 1365-2249, Vol. 194, no 1, p. 27-38Article in journal (Refereed)
    Abstract [en]

    Immunoglobulin (Ig) G- and IgM-class anti-cardiolipin antibodies (aCL) and lupus anti-coagulant (LA) are included in the 1997 update of the American College of Rheumatology (ACR-97) systemic lupus erythematosus (SLE) criteria. Despite limited evidence, IgA-aCL and IgA anti-(2)-glycoprotein-I (anti-(2)GPI) were included in the 2012 Systemic Lupus International Collaborating Clinics criteria. The present study aimed to evaluate IgG-/IgA-/IgM-aCL and anti-(2)GPI occurrence in relation to disease phenotype, smoking habits, pharmacotherapy, anti-phospholipid syndrome (APS) and organ damage among 526 Swedish SLE patients meeting ACR-97. Patients with rheumatoid arthritis (n=100), primary Sjogren's syndrome (n=50) and blood donors (n=507) served as controls. Anti-phospholipid antibodies (aPL) were analysed by fluoroenzyme-immunoassays detecting aCL/anti-(2)GPI. Seventy-six (14%) SLE cases fulfilled the Sydney APS-criteria, and 1 aCL/anti-(2)GPI isotype (IgG/IgA/IgM) occurred in 138 SLE patients (26%). Forty-five (9%) of the SLE cases had IgA-aCL, 20 of whom (4%) lacked IgG-/IgM-aCL. Seventy-four (14%) tested positive for IgA anti-(2)GPI, 34 (6%) being seronegative regarding IgG/IgM anti-(2)GPI. Six (1%) had APS manifestations but were seropositive regarding IgA-aCL and/or IgA anti-(2)GPI in the absence of IgG/IgM-aPL and LA. Positive LA and IgG-aPL tests were associated with most APS-related events and organ damage. Exclusive IgA anti-(2)GPI occurrence associated inversely with Caucasian ethnicity [odds ratio (OR)=021, 95% confidence interval (CI)=006-072) and photosensitivity (OR=019, 95% CI=005-072). Nephritis, smoking, LA-positivity and statin/corticosteroid-medication associated strongly with organ damage, whereas hydroxychloroquine-medication was protective. In conclusion, IgA-aPL is not rare in SLE (16%) and IgA-aPL analysis may have additional value among SLE cases with suspected APS testing negative for other isotypes of aPL and LA.

  • 355.
    Fromell, Karin
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Dührkop, Claudia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Johansson, Ulrika
    Linnaeus Univ, Linnaeus Ctr Biomat Chem, Vaxjo, Sweden..
    Nilsson Ekdahl, Kristina
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. Linnaeus Univ, Linnaeus Ctr Biomat Chem, Vaxjo, Sweden..
    Nilsson, Bo
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Forms of contact-activated C3 associated with AP convertase formation2017In: Molecular Immunology, ISSN 0161-5890, E-ISSN 1872-9142, Vol. 89, p. 141-141Article in journal (Other academic)
  • 356.
    Fromell, Karin
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Dührkop, Claudia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Kozarcanin, Huda
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Johansson, Ulrika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Skjoedt, Mikkel-Ole
    Rigshosp, Univ Hosp, Clin Immunol, Copenhagen, Sweden..
    Garred, Peter
    Rigshosp, Univ Hosp, Clin Immunol, Copenhagen, Sweden..
    Nilsson Ekdahl, Kristina
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. Linneus Univ, Biomat Chem, Kalmar, Sweden..
    Nilsson, Bo
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    The lectin pathway of complement and the contact/kallikrein system are integrated2018In: Molecular Immunology, ISSN 0161-5890, E-ISSN 1872-9142, Vol. 102, p. 151-152Article in journal (Other academic)
  • 357.
    Fu, Zhirong
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Microbiology.
    Thorpe, Michael
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Microbiology.
    Akula, Srinivas
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Microbiology.
    Chahal, Gurdeep
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Microbiology.
    Hellman, Lars
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Microbiology.
    Extended Cleavage Specificity of Human Neutrophil Elastase, Human Proteinase 3, and Their Distant Ortholog Clawed Frog PR3-Three Elastases With Similar Primary but Different Extended Specificities and Stability2018In: Frontiers in Immunology, ISSN 1664-3224, E-ISSN 1664-3224, Vol. 9, article id 2387Article in journal (Refereed)
    Abstract [en]

    Serine proteases are major granule constituents of several of the human hematopoietic cell lineages. Four proteolytically active such proteases have been identified in human neutrophils: cathepsin G (hCG), N-elastase (hNE), proteinase 3 (hPR-3), and neutrophil serine protease 4 (hNSP-4). Here we present the extended cleavage specificity of two of the most potent and most abundant of these enzymes, hNE and hPR-3. Their extended specificities were determined by phage display and by the analysis of a panel of chromogenic and recombinant substrates. hNE is an elastase with a relatively broad specificity showing a preference for regions containing several aliphatic amino acids. The protease shows self-cleaving activity, which results in the loss of activity during storage even at +4 degrees C. Here we also present the extended cleavage specificity of hPR-3. Compared with hNE, it shows considerably lower proteolytic activity. However, it is very stable, shows no self-cleaving activity and is actually more active in the presence of SDS, possibly by enhancing the accessibility of the target substrate. This enables specific analysis of hPR-3 activity even in the presence of all the other neutrophil enzymes with addition of 1% SDS. Neutrophils are the most abundant white blood cell in humans and one of the key players in our innate immune defense. The neutrophil serine proteases are very important for the function of the neutrophils and therefore also interesting from an evolutionary perspective. In order to study the origin and functional conservation of these neutrophil proteases we have identified and cloned an amphibian ortholog, Xenopus PR-3 (xPR-3). This enzyme was found to have a specificity very similar to hPR-3 but did not show the high stability in the presence of SDS. The presence of an elastase in Xenopus closely related to hPR-3 indicates a relatively early appearance of these enzymes during vertebrate evolution.

  • 358. Fuks, Jonas M
    et al.
    Arrighi, Romanico B G
    Weidner, Jessica M
    Kumar Mendu, Suresh
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
    Jin, Zhe
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Physiology.
    Wallin, Robert P A
    Rethi, Bence
    Birnir, Bryndis
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Physiology.
    Barragan, Antonio
    GABAergic Signaling Is Linked to a Hypermigratory Phenotype in Dendritic Cells Infected by Toxoplasma gondii2012In: PLoS pathogens, ISSN 1553-7374, Vol. 8, no 12, p. e1003051-Article in journal (Refereed)
    Abstract [en]

    During acute infection in human and animal hosts, the obligate intracellular protozoan Toxoplasma gondii infects a variety of cell types, including leukocytes. Poised to respond to invading pathogens, dendritic cells (DC) may also be exploited by T. gondii for spread in the infected host. Here, we report that human and mouse myeloid DC possess functional γ-aminobutyric acid (GABA) receptors and the machinery for GABA biosynthesis and secretion. Shortly after T. gondii infection (genotypes I, II and III), DC responded with enhanced GABA secretion in vitro. We demonstrate that GABA activates GABA(A) receptor-mediated currents in T. gondii-infected DC, which exhibit a hypermigratory phenotype. Inhibition of GABA synthesis, transportation or GABA(A) receptor blockade in T. gondii-infected DC resulted in impaired transmigration capacity, motility and chemotactic response to CCL19 in vitro. Moreover, exogenous GABA or supernatant from infected DC restored the migration of infected DC in vitro. In a mouse model of toxoplasmosis, adoptive transfer of infected DC pre-treated with GABAergic inhibitors reduced parasite dissemination and parasite loads in target organs, e.g. the central nervous system. Altogether, we provide evidence that GABAergic signaling modulates the migratory properties of DC and that T. gondii likely makes use of this pathway for dissemination. The findings unveil that GABA, the principal inhibitory neurotransmitter in the brain, has activation functions in the immune system that may be hijacked by intracellular pathogens.

  • 359.
    Galant, Natalie J.
    et al.
    Univ Toronto, Dept Med Biophys, Univ Hlth Network, Princess Margaret Canc Ctr, TMDT 4-305,101 Coll St, Toronto, ON M5G 1L7, Canada.;Univ Toronto, Dept Biochem, Univ Hlth Network, Princess Margaret Canc Ctr, TMDT 4-305,101 Coll St, Toronto, ON M5G 1L7, Canada..
    Bugyei-Twum, Antoinette
    Univ Toronto, Dept Med Biophys, Univ Hlth Network, Princess Margaret Canc Ctr, TMDT 4-305,101 Coll St, Toronto, ON M5G 1L7, Canada.;Univ Toronto, Dept Biochem, Univ Hlth Network, Princess Margaret Canc Ctr, TMDT 4-305,101 Coll St, Toronto, ON M5G 1L7, Canada..
    Rakhit, Rishi
    Stanford Univ, Dept Chem & Syst Biol, Stanford, CA 94305 USA..
    Walsh, Patrick
    Univ Toronto, Dept Biochem, Hosp Sick Children, Mol Struct & Funct Program, 1 Kings Coll Circle, Toronto, ON M5S 1A8, Canada..
    Sharpe, Simon
    Univ Toronto, Dept Biochem, Hosp Sick Children, Mol Struct & Funct Program, 1 Kings Coll Circle, Toronto, ON M5S 1A8, Canada..
    Arslan, Pharhad Eli
    Univ Toronto, Dept Med Biophys, Univ Hlth Network, Princess Margaret Canc Ctr, TMDT 4-305,101 Coll St, Toronto, ON M5G 1L7, Canada.;Univ Toronto, Dept Biochem, Univ Hlth Network, Princess Margaret Canc Ctr, TMDT 4-305,101 Coll St, Toronto, ON M5G 1L7, Canada..
    Westermark, Per
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical and experimental pathology.
    Higaki, Jeffrey N.
    Prothena Biosci Inc, Dept Biochem, San Francisco, CA 94080 USA.;Prothena Biosci Inc, Dept Histopathol, San Francisco, CA 94080 USA..
    Torres, Ronald
    Prothena Biosci Inc, Dept Biochem, San Francisco, CA 94080 USA.;Prothena Biosci Inc, Dept Histopathol, San Francisco, CA 94080 USA..
    Tapia, Jose
    Prothena Biosci Inc, Dept Biochem, San Francisco, CA 94080 USA.;Prothena Biosci Inc, Dept Histopathol, San Francisco, CA 94080 USA..
    Chakrabartty, Avijit
    Univ Toronto, Dept Med Biophys, Univ Hlth Network, Princess Margaret Canc Ctr, TMDT 4-305,101 Coll St, Toronto, ON M5G 1L7, Canada.;Univ Toronto, Dept Biochem, Univ Hlth Network, Princess Margaret Canc Ctr, TMDT 4-305,101 Coll St, Toronto, ON M5G 1L7, Canada..
    Substoichiometric inhibition of transthyretin misfolding by immune-targeting sparsely populated misfolding intermediates: a potential diagnostic and therapeutic for TTR amyloidoses2016In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 6, article id 25080Article in journal (Refereed)
    Abstract [en]

    Wild-type and mutant transthyretin (TTR) can misfold and deposit in the heart, peripheral nerves, and other sites causing amyloid disease. Pharmacological chaperones, Tafamidis (R) and diflunisal, inhibit TTR misfolding by stabilizing native tetrameric TTR; however, their minimal effective concentration is in the micromolar range. By immune-targeting sparsely populated TTR misfolding intermediates (i.e. monomers), we achieved fibril inhibition at substoichiometric concentrations. We developed an antibody (misTTR) that targets TTR residues 89-97, an epitope buried in the tetramer but exposed in the monomer. Nanomolar misTTR inhibits fibrillogenesis of misfolded TTR under micromolar concentrations. Pan-specific TTR antibodies do not possess such fibril inhibiting properties. We show that selective targeting of misfolding intermediates is an alternative to native state stabilization and requires substoichiometric concentrations. MisTTR or its derivative may have both diagnostic and therapeutic potential.

  • 360.
    Galindo-Feria, Angeles Shunashy
    et al.
    Karolinska Inst, Karolinska Univ Hosp, Stockholm, Sweden..
    Albrecht, Inka
    Karolinska Inst, Karolinska Univ Hosp, Stockholm, Sweden..
    Notarnicola, Antonella
    Karolinska Inst, Karolinska Univ Hosp, Stockholm, Sweden..
    Dastmalchi, Maryam
    Karolinska Inst, Karolinska Univ Hosp, Stockholm, Sweden..
    Dubnovitsky, Anatoly
    Sci Life Labs, Stockholm, Sweden..
    Sandalova, Tatiana
    Sci Life Labs, Stockholm, Sweden..
    Kozhukh, Genadiy
    Sci Life Labs, Stockholm, Sweden..
    Rönnblom, Lars
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Rheumatology.
    Achour, Adnane
    Sci Life Labs, Stockholm, Sweden..
    Malmstrom, Vivianne
    Karolinska Inst, Karolinska Univ Hosp, Stockholm, Sweden..
    Lundberg, Ingrid
    Karolinska Inst, Karolinska Univ Hosp, Stockholm, Sweden..
    Identification of a novel pro-inflammatory T cell epitope from his-tRNA-synthetase associated with interstitial lung disease in anti-Jo-1 positive patients2017In: Scandinavian Journal of Immunology, ISSN 0300-9475, E-ISSN 1365-3083, Vol. 86, no 4, p. 317-317Article in journal (Other academic)
  • 361.
    Gallini, Radiosa
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Molecular tools. Karolinska Inst, Dept Med Biochem & Biophys, Stockholm, Sweden..
    Huusko, Jenni
    Univ Kuopio, Dept Biotechnol & Mol Med, Al Virtanen Inst Mol Sci, Kuopio, Finland..
    Yla-Herttuala, Seppo
    Univ Kuopio, Dept Biotechnol & Mol Med, Al Virtanen Inst Mol Sci, Kuopio, Finland..
    Betsholtz, Christer
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Vascular Biology. Karolinska Inst, Dept Med Biochem & Biophys, Stockholm, Sweden..
    Andrae, Johanna
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Vascular Biology.
    Isoform-Specific Modulation of Inflammation Induced by Adenoviral Mediated Delivery of Platelet-Derived Growth Factors in the Adult Mouse Heart2016In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 11, no 8, article id e0160930Article in journal (Refereed)
    Abstract [en]

    Platelet-derived growth factors (PDGFs) are key regulators of mesenchymal cells in vertebrate development. To what extent PDGFs also exert beneficial homeostatic or reparative roles in adult organs, as opposed to adverse fibrogenic responses in pathology, are unclear. PDGF signaling plays critical roles during heart development, during which forced overexpression of PDGFs induces detrimental cardiac fibrosis; other studies have implicated PDGF signaling in post-infarct myocardial repair. Different PDGFs may exert different effects mediated through the two PDGF receptors (PDGFR alpha and PDGFR beta) in different cell types. Here, we assessed responses induced by five known PDGF isoforms in the adult mouse heart in the context of adenovirus vector-mediated inflammation. Our results show that different PDGFs have different, in some cases even opposing, effects. Strikingly, whereas the major PDGFRa agonists (PDGF-A and -C) decreased the amount of scar tissue and increased the numbers of PDGFR alpha-positive fibroblasts, PDGFR beta agonists either induced large scars with extensive inflammation (PDGF-B) or dampened the adenovirusinduced inflammation and produced a small and dense scar (PDGF-D). These results provide evidence for PDGF isoform-specific inflammation-modulating functions that may have therapeutic implications. They also illustrate a surprising complexity in the PDGF-mediated pathophysiological responses.

  • 362.
    Gallwitz, Maike
    et al.
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Immunology. Molekylär immunologi.
    Hellman, Lars
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Immunology. Molekylär immunologi.
    Rapid species-specific diversification of the mast cell chymase locus during mammalian evolution.2006In: Immunogenetics, Vol. 58, p. 641-654Article in journal (Refereed)
  • 363. Garcia-Vilas, Javier A.
    et al.
    Medina, Miguel A.
    Melo, Fabio R.
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Garcia-Faroldi, Gianni
    Damnacanthal inhibits IgE receptor-mediated activation of mast cells2015In: Molecular Immunology, ISSN 0161-5890, E-ISSN 1872-9142, Vol. 65, no 1, p. 86-93Article in journal (Refereed)
    Abstract [en]

    Damnacanthal, an anthraquinone obtained from the noni fruit (Morinda citrifolia L), has been described to possess anti-cancer and anti-inflammatory properties. Since mast cells are key players in various inflammatory conditions as well as in cancer, we considered the possibility that the biological actions of damnacanthal, at least partly, could be due to effects on mast cells. Many of the biological activities of mast cells are mediated by IgE receptor cross-linking, which results in degranulation with release of preformed granule mediators, as well as de nova synthesis and release of additional compounds. Here we show that damnacanthal has profound inhibitory activity on mast cell activation through this pathway. The release of the granule compounds beta-hexosaminidase and tryptase release was completely abrogated by damnacanthal at doses that were non-toxic to mast cells. In addition, damnacanthal inhibited activation-dependent pro-inflammatory gene induction, as well as cytokine/chemokine release in response to mast cell stimulation. The mechanism underlying damnacanthal inhibition was linked to impaired phosphorylation of Syk and Akt. Furthermore, damnacanthal inhibited mast cell activation in response to calcium ionophore A23187. Altogether, the data presented here demonstrate that damnacanthal inhibits mast cell activation induced by different stimuli and open a new window for the use of this compound as a mast cell stabilizer. 

  • 364.
    Garousi, Javad
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Andersson, Ken G.
    KTH Royal Inst Technol, Dept Prot Technol.
    Dam, Johan H.
    Odense Univ Hosp, Dept Nucl Med.
    Olsen, Birgitte B.
    Odense Univ Hosp, Dept Nucl Med..
    Mitran, Bogdan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Division of Molecular Imaging.
    Orlova, Anna
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Division of Molecular Imaging.
    Buijs, Jos
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Medical Radiation Science.
    Ståhl, Stefan
    KTH Royal Inst Technol, Dept Prot Technol.
    Löfblom, John
    KTH Royal Inst Technol, Dept Prot Technol.
    Thisgaard, Helge
    Odense Univ Hosp, Dept Nucl Med.
    Tolmachev, Vladimir
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    The use of radiocobalt as a label improves imaging of EGFR using DOTA-conjugated Affibody molecule2017In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 7, article id 5961Article in journal (Refereed)
    Abstract [en]

    Several anti-cancer therapies target the epidermal growth factor receptor (EGFR). Radionuclide imaging of EGFR expression in tumours may aid in selection of optimal cancer therapy. The In-111-labelled DOTA-conjugated Z(EGFR:2377) Affibody molecule was successfully used for imaging of EGFR-expressing xenografts in mice. An optimal combination of radionuclide, chelator and targeting protein may further improve the contrast of radionuclide imaging. The aim of this study was to evaluate the targeting properties of radiocobalt-labelled DOTA-Z(EGFR:2377). DOTA-Z(EGFR:2377) was labelled with Co-57 (T-1/2 = 271.8 d), Co-55 (T-1/2 = 17.5 h), and, for comparison, with the positron-emitting radionuclide Ga-68 (T-1/2 = 67.6 min) with preserved specificity of binding to EGFR-expressing A431 cells. The long-lived cobalt radioisotope Co-57 was used in animal studies. Both Co-57-DOTA-Z(EGFR:2377) and Ga-68-DOTA-Z(EGFR:2377) demonstrated EGFR-specific accumulation in A431 xenografts and EGFR-expressing tissues in mice. Tumour-to-organ ratios for the radiocobalt-labelled DOTA-Z(EGFR:2377) were significantly higher than for the gallium-labelled counterpart already at 3 h after injection. Importantly, Co-57-DOTA-Z(EGFR:2377) demonstrated a tumour-to-liver ratio of 3, which is 7-fold higher than the tumour-to-liver ratio for (68)GaDOTA-Z(EGFR:2377). The results of this study suggest that the positron-emitting cobalt isotope 55Co would be an optimal label for DOTA-Z(EGFR:2377) and further development should concentrate on this radionuclide as a label.

  • 365.
    Garraud, O.
    et al.
    Univ Lyon, Fac Med St Etienne, EA3064, F-42023 St Etienne 02, France;Inst Natl Transfus Sanguine, 6 Rue Alexandre Cabanel, F-7539 Paris, France.
    Martin, C.
    Univ Lyon, Fac Med St Etienne, EA3064, F-42023 St Etienne 02, France;Univ Hosp St Etienne, F-42023 St Etienne, France.
    Haddad, A.
    Univ Lyon, Fac Med St Etienne, EA3064, F-42023 St Etienne 02, France;Lebanese Univ, Sacre Coeur Hosp, Dept Clin Pathol & Blood Banking, Beirut, Lebanon.
    Hansen, M. Bagge
    Rigshosp, Copenhagen, Denmark.
    Rautmann, G.
    Council Europe, European Directorate Qual Med, F-75185 Strasbourg, France.
    MacLennan, S.
    NHS Blood & Transplant, Leeds, W Yorkshire, England.
    Norda, Rut
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    A European survey on donor deferral for allergy: Rationale and initial results of a survey in 35 countries2017In: Transfusion Clinique et Biologique, ISSN 1246-7820, E-ISSN 1953-8022, Vol. 24, no 1, p. 34-35Article in journal (Refereed)
    Abstract [en]

    Allergy accounts to near 0.5% of all reported transfusion adverse events. The responsibility of blood components themselves and - therefore - of blood donors is still questioned. The European Community undertook a large international survey to address the consistency and homogeneity of medical selection of blood donors with regard to the risk of allergy, and especially of transferring allergy to recipients. This short report presents the salient points of the survey, stressing that there is inconsistency in addressing the allergy question within countries or systems, with paths of improvement.

  • 366.
    Gasperov, Nina Milutin
    et al.
    Division of Molecular Medicine, Rudjer Boskovic Institute, Zagreb, Croatia.
    Farkas, Sanja A.
    Örebro University Hospital. Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Nilsson, Torbjörn K.
    Department of Medical Biosciences, Unit of Clinical Chemistry, Umeå University, Umeå, Sweden.
    Grce, Magdalena
    Division of Molecular Medicine, Rudjer Boskovic Institute, Zagreb, Croatia.
    Epigenetic activation of immune genes in cervical cancer2014In: Immunology Letters, ISSN 0165-2478, E-ISSN 1879-0542, Vol. 162, no 2, p. 256-257Article in journal (Refereed)
    Abstract [en]

    Immune system provides us protection from infectious pathogens and tumors formation during lifetime. Cervical cancer (CC), and its cause, human papillomavirus (HPV) are both challenges for the immune system. We present here evidence of epigenetic activation of immune system genes in CC. Illumina Infinium Human Methylation 450 K BeadChip identified genes, which were all significantly hypomethylated in CC tissue versus normal tissue. The GeneMANIA computer program identified a tight network between those genes. The most strongly correlated genes based on their function are immune effectors' process (AIM2, BST2, BTN3A3, and IL12RB1) and response to virus related genes (AIM2, BST2, and IL12RB1). Thus, activation of those genes through demethylation is probably triggered by HPV oncogenes. In conclusion, the immune system of women who do not develop CC is probably activated earlier through DNA demethylation.

  • 367.
    Ge, Changrong
    et al.
    Karolinska Inst, Stockholm, Sweden..
    Xu, Bingze
    Karolinska Inst, Stockholm, Sweden..
    Liang, Bibo
    Karolinska Inst, Stockholm, Sweden.;Southern Med Univ, Guangzhou, Guangdong, Peoples R China..
    Lonnblom, Erik
    Karolinska Inst, Stockholm, Sweden..
    Lundstrom, Susanna L.
    Karolinska Inst, Stockholm, Sweden..
    Zubarev, Roman A.
    Karolinska Inst, Stockholm, Sweden..
    Ayoglu, Burcu
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Nilsson, Peter
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Skogh, Thomas
    Linkoping Univ, Linkoping, Sweden..
    Kastbom, Alf
    Linkoping Univ, Linkoping, Sweden..
    Malmstrom, Vivianne
    Karolinska Inst, Stockholm, Sweden.;Karolinska Univ Hosp, Stockholm, Sweden..
    Klareskog, Lars
    Karolinska Inst, Stockholm, Sweden.;Karolinska Univ Hosp, Stockholm, Sweden..
    Toes, Rene E. M.
    Leiden Univ, Med Ctr, Leiden, Netherlands..
    Rispens, Theo
    Univ Amsterdam, Amsterdam, Netherlands..
    Dobritzsch, Doreen
    Uppsala Univ, Uppsala, Sweden..
    Holmdahl, Rikard
    Karolinska Inst, Stockholm, Sweden.;Southern Med Univ, Guangzhou, Guangdong, Peoples R China..
    Structural Basis of Cross-Reactivity of Anti-Citrullinated Protein Antibodies2019In: Arthritis & Rheumatology, ISSN 2326-5191, E-ISSN 2326-5205, Vol. 71, no 2, p. 210-221Article in journal (Refereed)
    Abstract [en]

    Objective Anti-citrullinated protein antibodies (ACPAs) develop many years before the clinical onset of rheumatoid arthritis (RA). This study was undertaken to address the molecular basis of the specificity and cross-reactivity of ACPAs from patients with RA. Methods Antibodies isolated from RA patients were expressed as monoclonal chimeric antibodies with mouse Fc. These antibodies were characterized for glycosylation using mass spectrometry, and their cross-reactivity was assessed using Biacore and Luminex immunoassays. The crystal structures of the antigen-binding fragment (Fab) of the monoclonal ACPA E4 in complex with 3 different citrullinated peptides were determined using x-ray crystallography. The prevalence of autoantibodies reactive against 3 of the citrullinated peptides that also interacted with E4 was investigated by Luminex immunoassay in 2 Swedish cohorts of RA patients. Results Analysis of the crystal structures of a monoclonal ACPA from human RA serum in complex with citrullinated peptides revealed key residues of several complementarity-determining regions that recognized the citrulline as well as the neighboring peptide backbone, but with limited contact with the side chains of the peptides. The same citrullinated peptides were recognized by high titers of serum autoantibodies in 2 large cohorts of RA patients. Conclusion These data show, for the first time, how ACPAs derived from human RA serum recognize citrulline. The specific citrulline recognition and backbone-mediated interactions provide a structural explanation for the promiscuous recognition of citrullinated peptides by RA-specific ACPAs.

  • 368.
    Gekara, Nelson O
    et al.
    Molecular Immunology, Helmholtz Center for Infection Research, Braunschweig, Germany.
    Zietara, Natalia
    Molecular Immunology and 2Department of Cell Biology, Helmholtz Center for Infection Research, Braunschweig, Germany.
    Geffers, Robert
    Molecular Immunology and 2Department of Cell Biology, Helmholtz Center for Infection Research, Braunschweig, Germany.
    Weiss, Siegfried
    Molecular Immunology and 2Department of Cell Biology, Helmholtz Center for Infection Research, Braunschweig, Germany.
    Listeria monocytogenes induces T cell receptor unresponsiveness through pore-forming toxin listeriolysin O2010In: Journal of Infectious Diseases, ISSN 0022-1899, E-ISSN 1537-6613, Vol. 202, no 11, p. 1698-1707Article in journal (Refereed)
    Abstract [en]

    Background.  The success of many pathogens relies on their ability to circumvent the innate and adaptive immune defenses. How bacterial pathogens subvert adaptive immune defenses is not clear. Cholesterol-dependent cytolysins (CDCs) represent an expansive family of homologous pore-forming toxins that are produced by more than 20 gram-positive bacterial species. Listeriolysin O (LLO), a prototype CDC, is the main virulence factor of Listeria monocytogenes. Methods.  We employed flow cytometric and microarray techniques to analyze the effect of LLO on T cell activation in vitro and in vivo. Results.  In vivo and in vitro proliferation of CD4(+) T cells upon T cell receptor (TCR) activation was highly diminished in the presence of LLO or wild-type L. monocytogenes but not in the presence of LLO-deficient L. monocytogenes. This block in T cell proliferation was specific to T cell activation via the TCR and not by phorbol 12-myristate 13-acetate-ionomycin, which bypasses the proximal TCR signaling event. The results of microarray analysis suggest that LLO-induced T cell unresponsiveness is due to the induction of a calcium-nuclear factor of activated T cells-dependent transcriptional program that drives the expression of negative regulators of TCR signaling. Conclusion. These findings provide important insights into how bacterial toxins silence adaptive immune responses and thus enable prolonged survival of the pathogen in the host.

  • 369.
    Gellerstedt, Martin
    et al.
    University West, Department of Economics and IT, Division of Computer Science and Informatics.
    Bengtsson, U
    The Asthma and Allergy Research Group, Department of Respiratory Medicine and Allergy,Sahlgrenska University Hospital, Gothenburg, Sweden.
    Niggemann, B
    Department of Pneumology and Immunology, University Children’s Hospital Charité, Berlin, Germany.
    Methodological issues in the methodological issues in the diagnostic work-up of food allergy: a real challenge2007In: Journal of investigational allergology & clinical immunology, ISSN 1018-9068, E-ISSN 1698-0808, Vol. 17, no 6, p. 350-356Article in journal (Refereed)
    Abstract [en]

    The standard of reporting in diagnostic studies has generally been low. Fortunately, this issue has begun to be addressed in recent years through the discussion of important methodological issues in educational series, textbooks, and checklists. Double-blind, placebocontrolled, oral food challenges (DBPCFC) are considered to be the gold standard for diagnosis of food allergy. However, there is no consensus regarding how to interpret the outcome and how to defi ne positive and negative provocations in DBPCFC. Furthermore, since most theories on the diagnosis of food allergy rely on the assumption that the DBPCFC has a high accuracy, this accuracy must be formally statistically evaluated. In this review, we discuss essential methodological issues for diagnostic accuracy studies in general and for oral food challenges in particular and discuss the importance of methodological issues as a guide for forthcoming studies of diagnostic procedures.

  • 370. Gendrin, Claire
    et al.
    Shubin, Nicholas J
    Boldenow, Erica
    Merillat, Sean
    Clauson, Morgan
    Power, Danial
    Doran, Kelly S
    Abrink, Magnus
    Pejler, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Department of Anatomy, Physiology and Biochemistry, the Swedish University of Agricultural Sciences, Uppsala, Sweden.
    Rajagopal, Lakshmi
    Piliponsky, Adrian M
    Mast cell chymase decreases the severity of group B Streptococcus infections2018In: Journal of Allergy and Clinical Immunology, ISSN 0091-6749, E-ISSN 1097-6825, Vol. 142, no 1, p. 120-129Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Group B Streptococcus (GBS) or Streptococcus agalactiae are β-hemolytic gram-positive bacteria that colonize the lower genital tracts of women and are frequently associated with infections during pregnancy. Innate immune defenses are critical for controlling GBS dissemination and systemic infection. Mast cells are resident sentinel cells that come into contact with pathogens early during colonization and infection.

    OBJECTIVE: We aimed to investigate the contribution of chymase to systemic GBS infection and rates of preterm birth.

    METHODS: Pharmacologic and genetic approaches using mice deficient in mast cell protease (MCPT) 4, the mouse functional homologue of human chymase, were used.

    RESULTS: Our studies show that mast cells release a protease with chymotrypsin-like cleavage specificity in response to GBS. Additionally, increased GBS systemic infection and preterm births were observed in MCPT4-deficient mice versus MCPT4-sufficient mice. Furthermore, we observed that proteolytic cleavage of the host extracellular matrix protein fibronectin by peritoneal cell-derived mast cell lysates diminished GBS adherence. Consistent with this observation, the increase in GBS dissemination and preterm births observed in MCPT4-deficient mice was abolished when GBS was deficient in expression of the fibronectin-binding protein SfbA.

    CONCLUSIONS: Taken together, our results suggest that the protective effect of MCPT4 against GBS dissemination and preterm labor can be attributed in part to MCPT4-mediated proteolysis of fibronectin. Our studies reveal a novel role of mast cells in defense against bacterial infections.

  • 371.
    Georganaki, Maria
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Vascular Biology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    van Hooren, Luuk
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Vascular Biology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Dimberg, Anna
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Vascular Biology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Vascular Targeting to Increase the Efficiency of Immune Checkpoint Blockade in Cancer2018In: Frontiers in Immunology, ISSN 1664-3224, E-ISSN 1664-3224, Vol. 9, article id 3081Article, review/survey (Refereed)
    Abstract [en]

    Boosting natural immunity against malignant cells has had a major breakthrough in clinical cancer therapy. This is mainly due to the successful development of immune checkpoint blocking antibodies, which release a break on cytolytic anti-tumor-directed T-lymphocytes. However, immune checkpoint blockade is only effective for a proportion of cancer patients, and a major challenge in the field is to understand and overcome treatment resistance. Immune checkpoint blockade relies on successful trafficking of tumor-targeted T-lymphocytes from the secondary lymphoid organs, through the blood stream and into the tumor tissue. Resistance to therapy is often associated with a low density of T-lymphocytes residing within the tumor tissue prior to treatment. The recruitment of leukocytes to the tumor tissue relies on up-regulation of adhesion molecules and chemokines by the tumor vasculature, which is denoted as endothelial activation. Tumor vessels are often poorly activated due to constitutive pro-angiogenic signaling in the tumor microenvironment, and therefore constitute barriers to efficient leukocyte recruitment. An emerging possibility to enhance the efficiency of cancer immunotherapy is to combine pro-inflammatory drugs with anti-angiogenic therapy, which can enable tumor-targeted T-lymphocytes to access the tumor tissue by relieving endothelial anergy and increasing adhesion molecule expression. This would pave the way for efficient immune checkpoint blockade. Here, we review the current understanding of the biological basis of endothelial anergy within the tumor microenvironment, and discuss the challenges and opportunities of combining vascular targeting with immunotherapeutic drugs as suggested by data from key pre-clinical and clinical studies.

  • 372.
    Gerasimcik, Natalija
    et al.
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute. University of Gothenburg, Sweden.
    He, Minghui
    Dahlberg, Carin I. M.
    Kuznetsov, Nikolai V.
    Severinson, Eva
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Westerberg, Lisa S.
    The small rho GTPases Rac1 and Rac2 are important for T-cell independent antigen responses and for suppressing switching to IgG2b in Mice2017In: Frontiers in Immunology, ISSN 1664-3224, E-ISSN 1664-3224, Vol. 8, article id 1264Article in journal (Refereed)
    Abstract [en]

    The Rho GTPases Cdc42, Rac1, and Rac2 coordinate receptor signaling to cell adhesion, migration, and proliferation. Deletion of Rac1 and Rac2 early during B cell development leads to failure in B cell entry into the splenic white pulp. Here, we sought to understand the role of Rac1 and Rac2 in B cell functionality and during the humoral antibody response. To circumvent the migratory deficiency of B cells lacking both Rac1 and Rac2, we took the approach to inducibly delete Rac1 in Rac2(-/-) B cells in the spleen (Rac1(B)Rac2(-/-) B cells). Rac1(B)Rac2(-/-) mice had normal differentiation of splenic B cell populations, except for a reduction in marginal zone B cells. Rac1(B)Rac2(-/-) B cells showed normal spreading response on antibody-coated layers, while both Rac2(-/-) and Rac1(B)Rac2(-/-) B cells had reduced homotypic adhesion and decreased proliferative response when compared to wild-type B cells. Upon challenge with the T-cell-independent antigen TNP-conjugated lipopolysaccharide, Rac1(B)Rac2(-/-) mice showed reduced antibody response. In contrast, in response to the T-cell-dependent antigen sheep red blood cells, Rac1(B)Rac2(-/-) mice had increased serum titers of IgG1 and IgG2b. During in vitro Ig class switching, Rac1(B)Rac2(-/-) B cells had elevated germline gamma 2b transcripts leading to increased Ig class switching to IgG2b. Our data suggest that Rac1 and Rac2 serve an important role in regulation of the B cell humoral immune response and in suppressing Ig class switching to IgG2b.

  • 373. Gerold, Gisa
    et al.
    Meissner, Felix
    Bruening, Janina
    Welsch, Kathrin
    Perin, Paula M
    Baumert, Thomas F
    Vondran, Florian W
    Kaderali, Lars
    Marcotrigiano, Joseph
    Khan, Abdul G
    Mann, Matthias
    Rice, Charles M
    Pietschmann, Thomas
    Quantitative Proteomics Identifies Serum Response Factor Binding Protein 1 as a Host Factor for Hepatitis C Virus Entry2015In: Cell reports, ISSN 2211-1247, E-ISSN 2211-1247, Vol. 12, no 5, p. 864-878, article id S2211-1247(15)00689-0Article in journal (Refereed)
    Abstract [en]

    Hepatitis C virus (HCV) enters human hepatocytes through a multistep mechanism involving, among other host proteins, the virus receptor CD81. How CD81 governs HCV entry is poorly characterized, and CD81 protein interactions after virus binding remain elusive. We have developed a quantitative proteomics protocol to identify HCV-triggered CD81 interactions and found 26 dynamic binding partners. At least six of these proteins promote HCV infection, as indicated by RNAi. We further characterized serum response factor binding protein 1 (SRFBP1), which is recruited to CD81 during HCV uptake and supports HCV infection in hepatoma cells and primary human hepatocytes. SRFBP1 facilitates host cell penetration by all seven HCV genotypes, but not of vesicular stomatitis virus and human coronavirus. Thus, SRFBP1 is an HCV-specific, pan-genotypic host entry factor. These results demonstrate the use of quantitative proteomics to elucidate pathogen entry and underscore the importance of host protein-protein interactions during HCV invasion.

  • 374.
    Gerstner, Christina
    et al.
    Karolinska Univ Hosp, Karolinska Inst, Ctr Mol Med, Rheumatol Unit,Dept Med Solna, Stockholm, Sweden..
    Dubnovitsky, Anatoly
    Karolinska Inst, Ctr Mol Med, Dept Clin Neurosci, Neuroimmunol Unit, Stockholm, Sweden.;Karolinska Inst, Dept Med Solna, Sci Life Lab, Stockholm, Sweden..
    Sandin, Charlotta
    Karolinska Univ Hosp, Karolinska Inst, Ctr Mol Med, Rheumatol Unit,Dept Med Solna, Stockholm, Sweden..
    Kozhukh, Genadiy
    Karolinska Univ Hosp, Karolinska Inst, Ctr Mol Med, Rheumatol Unit,Dept Med Solna, Stockholm, Sweden.;Karolinska Inst, Dept Med Solna, Sci Life Lab, Stockholm, Sweden..
    Uchtenhagen, Hannes
    Karolinska Univ Hosp, Karolinska Inst, Ctr Mol Med, Rheumatol Unit,Dept Med Solna, Stockholm, Sweden.;Virginia Mason, BRI, Translat Res Program, Seattle, WA USA..
    James, Eddie A.
    Virginia Mason, BRI, Tetramer Core, Seattle, WA USA..
    Rönnelid, Johan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Ytterberg, Anders Jimmy
    Karolinska Univ Hosp, Karolinska Inst, Ctr Mol Med, Rheumatol Unit,Dept Med Solna, Stockholm, Sweden.;Karolinska Inst, Dept Med Biochem & Biophys, Stockholm, Sweden..
    Pieper, Jennifer
    Karolinska Univ Hosp, Karolinska Inst, Ctr Mol Med, Rheumatol Unit,Dept Med Solna, Stockholm, Sweden..
    Reed, Evan
    Karolinska Univ Hosp, Karolinska Inst, Ctr Mol Med, Rheumatol Unit,Dept Med Solna, Stockholm, Sweden..
    Tandre, Carolina
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Rheumatology.
    Rieck, Mary
    Virginia Mason, BRI, Translat Res Program, Seattle, WA USA..
    Zubarev, Roman A.
    Karolinska Inst, Dept Med Biochem & Biophys, Stockholm, Sweden..
    Ronnblom, Lars
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Rheumatology. Uppsala Univ, Dept Med Sci, Sci Life Lab, Rheumatol, Uppsala, Sweden..
    Sandalova, Tatyana
    Karolinska Inst, Dept Med Solna, Sci Life Lab, Stockholm, Sweden.;Karolinska Univ Hosp Solna, Dept Infect Dis, Stockholm, Sweden..
    Buckner, Jane H.
    Virginia Mason, BRI, Translat Res Program, Seattle, WA USA..
    Achour, Adnane
    Karolinska Inst, Dept Med Solna, Sci Life Lab, Stockholm, Sweden.;Karolinska Univ Hosp Solna, Dept Infect Dis, Stockholm, Sweden..
    Malmstrom, Vivianne
    Karolinska Univ Hosp, Karolinska Inst, Ctr Mol Med, Rheumatol Unit,Dept Med Solna, Stockholm, Sweden..
    Functional and Structural Characterization of a Novel HLA-DRB1*04:01-Restricted alpha-Enolase T Cell Epitope in Rheumatoid Arthritis2016In: Frontiers in Immunology, ISSN 1664-3224, E-ISSN 1664-3224, Vol. 7, article id 494Article in journal (Refereed)
    Abstract [en]

    Antibodies to citrullinated proteins, common in rheumatoid arthritis (RA) patients, are strongly associated to a specific set of HLA-DR alleles including HLA-DRB1*04:01, *04:04, and *01:01. Here, we first demonstrate that autoantibody levels toward the dominant citrullinated B cell epitope from alpha-enolase are significantly elevated in HLA-DRB1*04:01-positive RA patients. Furthermore, we identified alpha-enolase-derived T cell epitopes and demonstrated that native and citrullinated versions of several peptides bind with different affinities to HLA-DRB1*04:01, *04:04, and *01:01. The citrulline residues in the eight identified peptides are distributed throughout the entire length of the presented epitopes and more specifically, localized at peptide positions p-2, p2, p4, p6, p7, p10, and p11. Importantly, in contrast to its native version peptide 26 (TSKGLFRAAVPSGAS), the HLA-DRB1*04:01-restricted citrullinated peptide Cit26 (TSKGLFCitAAVPSGAS) elicited significant functional T cell responses in primary cells from RA patients. Comparative analysis of the crystal structures of HLA-DRB1*04:01 in complex with peptide 26 or Cit26 demonstrated that the posttranslational modification did not alter the conformation of the peptide. And since citrullination is the only structural difference between the two complexes, this indicates that the neo-antigen Cit26 is recognized by T cells with high specificity to the citrulline residue.

  • 375.
    Getahun, Andrew
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Genetics and Pathology.
    Antibody Feedback Regulation: From Epitope Masking to T Helper Cell Activation2004Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Antibodies have the ability to influence the antibody response against the very antigen they are specific for, in a process called antibody feedback regulation. Depending on the nature of the antigen, the antibody response can be either enhanced or almost completely inhibited. This thesis focuses on the underlying mechanisms of antibody feedback regulation in vivo.

    Antigen-specific IgG can inhibit the antibody response to a particulate antigen. Based on its ability to inhibit B cell activation, the inhibitory FcγRIIB (low affinity receptor for IgG) has been suggested to be involved. Here we show that although FcγRIIB is required for efficient suppression in vitro, it is not required in vivo. Therefore, even though FcγRIIB can inhibit antibody responses, other mechanisms (such as epitope masking and enhanced antigen clearance) play a more dominant role in vivo.

    The antibody response to soluble antigen is greatly enhanced when it is introduced to the immune system in complex with antigen-specific IgG or IgE. We found that FcγRIIB attenuates the magnitude of IgG-mediated enhancement. In mice lacking FcγRIIB, IgG enhanced the antibody response much more efficiently than in normal mice.

    Since B cells require CD4+ T cell help in order to become antibody-producing cells, we examined the CD4+ T cell response to immune complexes in vivo. Using an adoptive transfer strategy with transgenic ovalbumin (OVA)-specific CD4+T cells, we could show that the enhanced OVA-specific IgG response to IgG2a/OVA and IgE/OVA complexes was preceded by a potent OVA-specific CD4+ T cell response. IgG2a-mediated enhancement was dependent on activating Fcγ receptors, whereas IgE-mediated enhancement was dependent on CD23, the low affinity receptor for IgE. We identified CD23+ B cells as the responsible effector cells for IgE-mediated enhancement in vivo. Taken together, these results show that Fc receptor-mediated antigen presentation is a major mechanism underlying antibody feedback enhancement.

  • 376. Gibbs, Anna
    et al.
    Buggert, Marcus
    Edfeldt, Gabriella
    Ranefall, Petter
    Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Division of Visual Information and Interaction. Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Computerized Image Analysis and Human-Computer Interaction.
    Introini, Andrea
    Cheuk, Stanley
    Martini, Elisa
    Eidsmo, Liv
    Hirbod, Taha
    Ball, Terry B.
    Kimani, Joshua
    Kaul, Rupert
    Karlsson, Annika C.
    Wählby, Carolina
    Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Division of Visual Information and Interaction. Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Computerized Image Analysis and Human-Computer Interaction.
    Broliden, Kristina
    Tjernlund, Annelie
    Increased numbers of CD103-CD8+ TRM cells in the cervical mucosa of HIV-infected women2017In: Scandinavian Journal of Immunology, ISSN 0300-9475, E-ISSN 1365-3083, Vol. 86, no 4, p. 288-289Article in journal (Other academic)
  • 377. Giha, Hayder A
    et al.
    Nasr, Amre
    Stockholm University, Faculty of Science, The Wenner-Gren Institute , Immunology.
    Iriemenam, Nnaemeka C
    Stockholm University, Faculty of Science, The Wenner-Gren Institute , Immunology.
    Arnot, David
    Troye-Blomberg, Marita
    Stockholm University, Faculty of Science, The Wenner-Gren Institute , Immunology.
    Theander, Thor G
    Berzins, Klavs
    Stockholm University, Faculty of Science, The Wenner-Gren Institute , Immunology.
    Elghazali, Gehad
    Pandey, Janardan P
    Antigen-specific influence of GM/KM allotypes on IgG isotypes and association of GM allotypes with susceptibility to Plasmodium falciparum malaria.2009In: Malaria journal, ISSN 1475-2875, Vol. 8, no 1, p. 306-Article in journal (Refereed)
    Abstract [en]

    ABSTRACT: BACKGROUND: Plasmodium falciparum malaria is a complex disease in which genetic and environmental factors influence susceptibility. IgG isotypes are in part genetically controlled, and GM/KM allotypes are believed to be involved in this control. METHODS: In this study, 216 individuals from Daraweesh, an area of seasonal malaria transmission in Sudan, were followed for nine years for malaria infection. Total IgG and IgG isotypes against four malaria antigens, MSP2-3D7, MSP2-FC27, AMA1, and Pf332-C231 were measured in plasma obtained from the cohort at the end of the study, during the dry malaria-free period. The GM/KM allotypes of the donors were determined. RESULTS: The GM 1,17 5,13,14,6 phenotype was associated with a higher incidence of malaria compared with the non-1,17 5,13,14,6 phenotypes (P = 0.037). Paradoxically, the carriers of the GM 1,17 5,13,14,6 phenotype had significantly higher baseline levels of total IgG and non-cytophilic IgG isotypes as compared to non-carriers. The KM allotypes influence on IgG isotypes level was limited. Finally, the differences in the baseline concentrations of total IgG and IgG isotypes between the different GK/KM phenotype carriers were antigen-dependent. DISCUSSION: The results show that GM but not KM allotypes appeared to influence host susceptibility to uncomplicated malaria as well as the antibody profile of the donors, and the carriers of the GM 1,17 5,13,14,6 phenotype were the most susceptible CONCLUSIONS: The GM allotypes have significant influence on susceptibility to uncomplicated P. falciparum malaria and antigen-dependent influence on total IgG and IgG subclasses.

  • 378.
    Giusti, Pablo
    Stockholm University, Faculty of Science, The Wenner-Gren Institute , Immunology.
    Dendritic cells and Plasmodium falciparum: studies in vitro and in the human host2009Licentiate thesis, comprehensive summary (Other academic)
    Abstract [en]

    Malaria is one of the world’s most threatening diseases. About half the world’s population is at risk of infection and the infection claims a million lives each year. A vast majority of the deaths occur in children below the age of 5 in sub-Saharan Africa. Survivors typically acquire immunity only after long time of repeated exposure and immunity is rapidly lost. Immunity is created by the activation of naive T cells and their differentiation into effector cells. The most potent activators of naive T cells are dendritic cells (DCs). The life cycle of DCs is adapted to find and process microbes in order to be able to present their antigens to T cells and thereby activate them. Antigen presentation typically takes place in the lymph nodes and that is why migration to these areas is an essential part of the DC life cycle. Various studies have shown that DC function may be hampered by the malaria parasite or its components.

    We have investigated activation and migratory capacities of DCs upon in vitro exposure of the malarial pigment hemozoin and Plasmodium falciparum infected red blood cells. Furthermore, we have assessed the activation status of blood DCs in the Fulani, a traditionally nomadic population that respond better to malaria infection and exhibit less clinical symptoms than other ethnicities living under similar conditions, and a neighbouring ethnic group, the Dogon, in Mali.

    Our results indicate that DCs are semi-activated upon malaria exposure in vitro, including enhanced migratory capacity, partial up-regulation of co-stimulatory markers and no IL-12, which may lead to inappropriate T-cell priming. We also observed that DCs from the Fulani have a higher degree of activation than DCs from the Dogon upon malaria exposure in vivo.

    We hypothesize that this increased DC activation may be the reason for the relatively increased protection against malaria.

    Taken together, our findings suggest that improper DC activation may contribute to poor immunity in Malaria.

  • 379.
    Giusti, Pablo
    Stockholm University, Faculty of Science, The Wenner-Gren Institute , Immunology.
    The novel anti-rheumatic compound Rabeximod impairs differentiation and function of human pro-inflammatory dendritic cells and macrophages2011In: Immunobiology, ISSN 0171-2985, E-ISSN 1878-3279, Vol. 216, no 1-2, p. 243-250Article in journal (Refereed)
    Abstract [en]

    Rabeximod (9-chloro-2,3-dimethyl-6-(N,N-dimethylaminoethylamino-2-oxoethyl)-6H-indolo[2,3-b]quinoxaline) is a synthetic compound that is currently being developed for the treatment of rheumatoid arthritis (RA). Here, we investigated the effects of Rabeximod on the functionality of human antigen-presenting cells (APCs) of myeloid origin. Different subsets of professional APCs were generated from human monocytes in vitro and simultaneously treated with different doses of Rabeximod. Although Rabeximod had no effect on the differentiation of monocytes into anti-inflammatory macrophages (AI-Ms), this compound impaired monocyte differentiation into monocyte-derived dendritic cells (MDCs) and pro-inflammatory allostimulated macrophages (Allo-Ms). MDCs that were treated with Rabeximod resulted in a significant decrease in their ability to pinocytose antigens, while no effect was exerted by the drug on the ability of Allo-Ms and AI-Ms to phagocytose. Furthermore, we observed a significant reduction in the allostimulatory ability of MDCs and Allo-Ms after treatment with Rabeximod, although this compound did not affect the low immunostimulatory capacity of AI-Ms. Conversely, the effect of Rabeximod in influencing cytokine secretion by APCs appeared to be limited. In conclusion, Rabeximod impairs differentiation of monocytes into different pro-inflammatory APCs, leading to impaired immunostimulatory abilities of these cells. Our observations shed light on the cellular mode of action and the immunomodulatory effect of Rabeximod.

  • 380.
    Giusti, Pablo
    et al.
    Stockholm University, Faculty of Science, The Wenner-Gren Institute , Immunology.
    Troye Blomberg, Marita
    Stockholm University, Faculty of Science, The Wenner-Gren Institute , Immunology.
    Varani, Stefania
    Stockholm University, Faculty of Science, The Wenner-Gren Institute , Immunology.
    Plasmodium falciparum-Infected Erythrocytes and beta-Hematin Induce Partial Maturation of Human Dendritic Cells and Increase Their Migratory Ability in Response to Lymphoid Chemokines2011In: Infection and Immunity, ISSN 0019-9567, E-ISSN 1098-5522, Vol. 79, no 7, p. 2727-2736Article in journal (Refereed)
    Abstract [en]

    Acute and chronic Plasmodium falciparum infections alter theimmune competence of the host possibly through changes in dendriticcell (DC) functionality. DCs are the most potent activatorsof T cells, and migration is integral to their function. MatureDCs express lymphoid chemokine receptors (CCRs), expressionof which enables them to migrate to the lymph nodes, where theyencounter naïve T cells. The present study aimed to investigatethe impact of the synthetic analog to malaria parasite pigmenthemozoin, i.e., β-hematin, or infected erythrocytes (iRBCs)on the activation status of human monocyte-derived DCs and ontheir expression of CCRs. Human monocyte-derived DCs partiallymatured upon incubation with β-hematin as indicated byan increased expression of CD80 and CD83. Both β-hematinand iRBCs provoked the release of proinflammatory and anti-inflammatorycytokines, such as interleukin-6 (IL-6), IL-10, and tumor necrosisfactor alpha, but not IL-12, and induced upregulation of thelymphoid chemokine receptor CXCR4, which was coupled to an increasedmigration to lymphoid ligands. Taken together, these resultssuggest that the partial and transient maturation of human myeloidDCs upon stimulation with malaria parasite-derived productsand the increased IL-10 but lack of IL-12 secretion may leadto suboptimal activation of T cells. This may in turn lead toimpaired adaptive immune responses and therefore insufficientclearance of the parasites.

  • 381. Gnann, John W, Jr
    et al.
    Sköldenberg, Birgit
    Hart, John
    Aurelius, Elisabeth
    Schliamser, Silvia
    Studahl, Marie
    Eriksson, Britt-Marie
    Hanley, Daniel
    Aoki, Fred
    Jackson, Alan C
    Griffiths, Paul
    Miedzinski, Lil
    Hanfelt-Goade, Diane
    Hinthorn, Daniel
    Ahlm, Clas
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology.
    Aksamit, Allen
    Cruz-Flores, Salvador
    Dale, Ilet
    Cloud, Gretchen
    Jester, Penelope
    Whitley, Richard J
    Herpes Simplex Encephalitis: Lack of Clinical Benefit of Long-term Valacyclovir Therapy2015In: Clinical Infectious Diseases, ISSN 1058-4838, E-ISSN 1537-6591, Vol. 61, no 5, p. 683-691Article in journal (Refereed)
    Abstract [en]

    Background: Despite the proven efficacy of acyclovir (ACV) therapy, herpes simplex encephalitis (HSE) continues to cause substantial morbidity and mortality. Among patients with HSE treated with ACV, the mortality rate is approximately 14%-19%. Among survivors, 45%-60% have neuropsychological sequelae at 1 year. Thus, improving therapeutic approaches to HSE remains a high priority. Methods: Following completion of a standard course of intravenous ACV, 87 adult patients with HSE (confirmed by positive polymerase chain reaction [PCR] for herpes simplex virus DNA in cerebrospinal fluid) were randomized to receive either valacyclovir (VACV) 2 g thrice daily (n = 40) or placebo tablets (n = 47) for 90 days (12 tablets of study medication daily). The primary endpoint was survival with no or mild neuropsychological impairment at 12 months, as measured by the Mattis Dementia Rating Scale (MDRS). Logistic regression was utilized to assess factors related to the primary endpoint. Results: The demographic characteristics of the 2 randomization groups were statistically similar with no significant differences in age, sex, or race. At 12 months, there was no significant difference in the MDRS scoring for VACV-treated vs placebo recipients, with 85.7% and 90.2%, respectively, of patients demonstrating no or mild neuropsychological impairment (P = .72). No significant study-related adverse events were encountered in either treatment group. Conclusions: Following standard treatment with intravenous ACV for PCR-confirmed HSE, an additional 3-month course of oral VACV therapy did not provide added benefit as measured by neuropsychological testing 12 months later in a population of relatively high-functioning survivors.

  • 382. Goeijenbier, M.
    et al.
    Hartskeerl, R. A.
    Reimerink, J.
    Verner-Carlsson, J.
    Wagenaar, J. F.
    Goris, M. G.
    Martina, B. E.
    Lundkvist, Åke
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Koopmans, M.
    Osterhaus, A. D.
    van Gorp, E. C.
    Reusken, C. B.
    The hanta hunting study: underdiagnosis of Puumala hantavirus infections in symptomatic non-travelling leptospirosis-suspected patients in the Netherlands, in 2010 and April to November 20112014In: Eurosurveillance, ISSN 1025-496X, E-ISSN 1560-7917, Vol. 19, no 32, p. 27-36Article in journal (Refereed)
    Abstract [en]

    Leptospirosis and haemorrhagic fever with renal syndrome (HFRS) are hard to distinguish clinically since these two important rodent-borne zoonoses share hallmark symptoms such as renal failure and haemorrhage. Leptospirosis is caused by infection with a spirochete while HFRS is the result of an infection with certain hantaviruses. Both diseases are relatively rare in the Netherlands. Increased incidence of HFRS has been observed since 2007 in countries that border the Netherlands. Since a similar rise in incidence has not been registered in the Netherlands, we hypothesise that due to overlapping clinical manifestations, hantavirus infections may be confused with leptospirosis, leading to underdiagnosis. Therefore, we tested a cohort of non-travelling Dutch patients with symptoms compatible with leptospirosis, but with a negative diagnosis, during 2010 and from April to November 2011. Sera were screened with pan-hantavirus IgG and IgM enzyme-linked immunosorbent assays (ELISAs). Sera with IgM reactivity were tested by immunofluorescence assay (IFA). ELISA (IgM positive) and IFA results were confirmed using focus reduction neutralisation tests (FRNTs). We found hantavirus-specific IgG and/or IgM antibodies in 4.3% (11/255) of samples taken in 2010 and in 4.1% (6/146) of the samples during the 2011 period. After FRNT confirmation, seven patients were classed as having acute Puumala virus infections. A review of hantavirus diagnostic requests revealed that at least three of the seven confirmed acute cases as well as seven probable acute cases of hantavirus infection were missed in the Netherlands during the study period.

  • 383.
    Golparian, Daniel
    et al.
    WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Faculty of Medicine and Health, Örebro University, Örebro, Sweden.
    Hellmark, Bengt
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Faculty of Medicine and Health, Örebro University, Örebro, Sweden.
    Unemo, Magnus
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Faculty of Medicine and Health, Örebro University, Örebro, Sweden.
    Analytical specificity and sensitivity of the novel dual-target GeneProof Neisseria gonorrhoeae PCR kit for detection of N-gonorrhoeae2015In: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS), ISSN 0903-4641, E-ISSN 1600-0463, Vol. 123, no 11, p. 955-958Article in journal (Refereed)
    Abstract [en]

    Detection of Neisseria gonorrhoeae relies increasingly on nucleic acid amplification tests (NAATs). The specificity of many gonococcal NAATs has been suboptimal and supplementary testing remains recommended in Europe and several additional countries. The novel dual-target GeneProofNeisseria gonorrhoeae PCR kit, targeting porA pseudogene and 16S rRNA gene, showed a high specificity and sensitivity when isolates of non-gonococcal Neisseria and related species (n=144), and gonococci (n=104) were tested. However, rare gonococcal porA mutants were only detected in the 16S rRNA gene target and two non-gonococcal isolates showed a low-level cross-reactivity in the 16S rRNA gene target. The detection limit for both targets was 1.5 copies per reaction.

  • 384.
    Goto, Masafumi
    et al.
    Tohoku Univ, New Ind Creat Hatchery Ctr, Sendai, Miyagi 980, Japan.;Tohoku Univ, Div Adv Surg Sci & Technol, Sendai, Miyagi 980, Japan..
    Friberg, Andrew
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Ståhle, Magnus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Imura, Takehiro
    Tohoku Univ, New Ind Creat Hatchery Ctr, Sendai, Miyagi 980, Japan..
    Yamagata, Youhei
    Tokyo Univ Agr & Technol, Dept Appl Biol Chem, Tokyo, Japan..
    Watanabe, Kimiko
    Tohoku Univ, New Ind Creat Hatchery Ctr, Sendai, Miyagi 980, Japan..
    Murayama, Kazutaka
    Tohoku Univ, Div Biomed Measurements & Diagnost, Sendai, Miyagi 980, Japan..
    Inagaki, Akiko
    Tohoku Univ, New Ind Creat Hatchery Ctr, Sendai, Miyagi 980, Japan..
    Igarashi, Yasuhiro
    Tohoku Univ, New Ind Creat Hatchery Ctr, Sendai, Miyagi 980, Japan..
    Satomi, Susumu
    Tohoku Univ, Div Adv Surg Sci & Technol, Sendai, Miyagi 980, Japan..
    Korsgren, Olle
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Proof Of Concept For The Clinical Application Of Animal Component Free Recombinant Collagenase For Isolating Pancreatic Islets2015In: Transplantation, ISSN 0041-1337, E-ISSN 1534-6080, Vol. 99, no 11, p. S80-S80Article in journal (Other academic)
  • 385.
    Grapensparr, Liza
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology, Integrative Physiology.
    Olerud, Johan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Vasylovska, Svitlana
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neuroanatomy.
    Carlsson, Per-Ola
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    The therapeutic role of endothelial progenitor cells in Type 1 diabetes mellitus2011In: Regenerative Medicine, ISSN 1746-0751, E-ISSN 1746-076X, Vol. 6, no 5, p. 599-605Article, review/survey (Refereed)
    Abstract [en]

    Pancreatic beta-cells sense and adjust the blood glucose level by secretion of insulin. In Type 1 diabetes mellitus, these insulin-producing cells are destroyed, leaving the patients incapable of regulating blood glucose homeostasis. At the time of diagnosis, most patients still have 20-30% of their original beta-cell mass remaining. These residual beta-cells are targets for intervention therapies aimed at preventing further autoimmune destruction, in addition to increasing the number of existing beta-cells. Such a therapeutic option is highly desirable since it may lead to a full recovery of newly diagnosed patients, with no need for further treatment with immunosuppressant drugs or exogenous insulin administration. In this article, we propose that endothelial progenitor cells, a cell type known to promote and support neovascularization following endothelial injury, may be used as part of a combinational stem cell therapy aimed to improve the vascularization, survival and proliferation of beta-cells.

  • 386.
    Grero, Dhanya
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Cytotoxicity of Vγ9Vδ2 T cells towards Colon Cancer Cells2014Independent thesis Advanced level (degree of Master (One Year)), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    Immunotherapies for cancer are widely studied at present. We are currently studying a specific form of “Vγ9Vδ2 T cells” found in the peripheral blood of healthy donors that can be used for the killing of HT-29 colon cancer cells. In order to determine the cytotoxicity of effectors, Vγ9Vδ2 T cells towards target cells, HT-29, it is important to first evaluate the absolute number of Vγ9Vδ2 T cells in a mixed cell population, and next to determine the phenotypic characterization, their activity and cytotoxicity in the presence of target cells. A flow cytometry and bead based assay was developed to evaluate the absolute number of Vγ9Vδ2 T cells in a mixed cell population. Peripheral Blood Mononuclear Cells (PBMCs) were surface stained with monoclonal antibodies (MoAbs) conjugated to fluorochromes that are cross reactive to cell surface markers such as CD3 (T Lymphocytes), γδ2 and were mixed with fluorophore beads. In these assays, no washes and centrifugation steps were performed after the cell surface staining and bead addition. The absolute cell counts were evaluated based on referencing a known concentration of beads. In addition, quantification assays were also performed to measure the cell and bead loss on surface staining that included washes and centrifugation steps and thus found a higher percentage loss of cells than beads. Immunophenotyping assays with four color staining were performed to monitor the phenotypic differentiation of effector cells based on cell surface markers CD27 and CD45RA. Only the naïve (CD27+CDRA+) and terminally differentiated effector memory (CD27-CD45RA+) were identified on the assays performed using Vγ9Vδ2 T cells of different donors. A flow cytometry based cytotoxicity (FCC) assay was completed to monitor the effector cell activity (CD69+) in the presence and absence of target cells and also the cytotoxicity was measured based on % specific lysis of target cells at four different effector to target (E:T) cell ratios. Only preliminary data were obtained for the FCC assay and the development is still in progress.  

  • 387. Gruden, Marina A.
    et al.
    Sewell, Robert D. E.
    Yanamandra, Kiran
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Davidova, Tatyana V.
    Kucheryanu, Valery G.
    Bocharov, Evgeny V.
    Bocharova, Olga A.
    Poleschuk, Vsevolod V.
    Sherstnev, Vladimir V.
    Morozova-Roche, Ludmilla A.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Immunoprotection against toxic biomarkers is retained during Parkinson's disease progression (vol 233, pg 221, 2011)2014In: Journal of Neuroimmunology, ISSN 0165-5728, E-ISSN 1872-8421, Vol. 268, no 1-2, p. 99-99Article in journal (Refereed)
  • 388. Gruden, Marina A.
    et al.
    Yanamandra, Kiran
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Kucheryanu, Valery G.
    Bocharova, Olga R.
    Sherstnev, Vladimir V.
    Morozova-Roche, Ludmilla A.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Sewell, Robert D. E.
    Correlation between Protective Immunity to alpha-Synuclein Aggregates, Oxidative Stress and Inflammation2012In: Neuroimmunomodulation, ISSN 1021-7401, E-ISSN 1423-0216, Vol. 19, no 6, p. 334-342Article in journal (Refereed)
    Abstract [en]

    Objective: Protein aggregation leading to central amyloid deposition is implicated in Parkinson's disease (PD). During disease progression, inflammation and oxidative stress may well invoke humoral immunity against pathological aggregates of PD-associated alpha-synuclein. The aim was to investigate any possible concurrence between autoimnnune responses to alpha-synuclein monomers, oligomers or fibrils with oxidative stress and inflammation.

    Methods: The formation of alpha-synuclein amyloid species was assessed by thioflavin-T assay and atomic force microscopy was employed to confirm their morphology. Serum autoantibody titers to alpha-synuclein conformations were determined by ELISA. Enzyme activity and concentrations of oxidative stress/inflammatory indicators were evaluated by enzyme and ELISA protocols.

    Results: In PD patient sera, a differential increase in autoantibody titers to alpha-synuclein monomers, toxic oligomers or fibrils was associated with boosted levels of the pro-inflammatory cytokine interleukin-6 and tumour necrosis factor-alpha, but a decrease in interferon-gamma concentration. In addition, levels of malondialdehyde were elevated whilst those of glutathione were reduced along with decrements in the activity of the antioxidants: superoxide dismutase, catalase and glutathione transferase.

    Conclusions: It is hypothesized that the generation of alpha-synuclein amyloid aggregates allied with oxidative stress and inflammatory reactions may invoke humoral immunity protecting against dopaminergic neuronal death. Hence, humoral immunity is a common integrative factor throughout PD progression which is directed towards prevention of further neurodegeneration, so potential treatment strategies should attempt to maintain PD patient immune status. Copyright (c) 2012 S. Karger AG, Basel

  • 389.
    Grundström, Christine
    et al.
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Kumar, Anjani
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Priya, Anshu
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Negi, Neema
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Grundström, Thomas
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    ETS1 and PAX5 transcription factors recruit AID to Igh DNA2018In: European Journal of Immunology, ISSN 0014-2980, E-ISSN 1521-4141, Vol. 48, no 10, p. 1687-1697Article in journal (Refereed)
    Abstract [en]

    B lymphocytes optimize antibody responses by class switch recombination (CSR), which changes the expressed constant region exon of the immunoglobulin heavy chain (IgH), and by somatic hypermutation (SH) that introduces point mutations in the variable regions of the antibody genes. Activation-induced cytidine deaminase (AID) is the key mutagenic enzyme that initiates both these antibody diversification processes by deaminating cytosine to uracil. Here we asked the question if transcription factors can mediate the specific targeting of the antibody diversification by recruiting AID. We have recently reported that AID is together with the transcription factors E2A, PAX5 and IRF4 in a complex on key sequences of the Igh locus. Here we report that also ETS1 is together with AID in this complex on key sequences of the Igh locus in splenic B cells of mice. Furthermore, we show that both ETS1 and PAX5 can directly recruit AID to DNA sequences from the Igh locus with the specific binding site for the transcription factor. Taken together, our findings support the notion of a targeting mechanism for the selective diversification of antibody genes with limited genome wide mutagenesis by recruitment of AID by PAX5 and ETS1 in a transcription factor complex.

  • 390. Grunewald, Johan
    et al.
    Kaiser, Ylva
    Ostadkarampour, Mahyar
    Rivera, Natalia V.
    Vezzi, Francesco
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Lötstedt, Britta
    Olsen, Remi-André
    Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
    Sylwan, Lina
    Lundin, Sverker
    Käller, Max
    Sandalova, Tatiana
    Ahlgren, Kerstin M.
    Wahlström, Jan
    Achour, Adnane
    Ronninger, Marcus
    Eklund, Anders
    T-cell receptor-HLA-DRB1 associations suggest specific antigens in pulmonary sarcoidosis2016In: European Respiratory Journal, ISSN 0903-1936, E-ISSN 1399-3003, Vol. 47, no 3, p. 898-909Article in journal (Refereed)
    Abstract [en]

    In pulmonary sarcoidosis, CD4(+) T-cells expressing T-cell receptor V alpha 2.3 accumulate in the lungs of HLA-DRB1*03(+) patients. To investigate T-cell receptor-HLA-DRB1*03 interactions underlying recognition of hitherto unknown antigens, we performed detailed analyses of T-cell receptor expression on bronchoalveolar lavage fluid CD4(+) T-cells from sarcoidosis patients. Pulmonary sarcoidosis patients (n=43) underwent bronchoscopy with bronchoalveolar lavage. T-cell receptor alpha and beta chains of CD4(+) T-cells were analysed by flow cytometry, DNA-sequenced, and three-dimensional molecular models of T-cell receptor-HLA-DRB1*03 complexes generated. Simultaneous expression of V alpha 2.3 with the V beta 22 chain was identified in the lungs of all HLA-DRB1*03(+) patients. Accumulated V alpha 2.3/V beta 22-expressing T-cells were highly clonal, with identical or near-identical V alpha 2.3 chain sequences and inter-patient similarities in V beta 22 chain amino acid distribution. Molecular modelling revealed specific T-cell receptor-HLA-DRB1*03-peptide interactions, with a previously identified, sarcoidosis-associated vimentin peptide, (Vim)(429-443) DSLPLVDTHSKRTLL, matching both the HLA peptide-binding cleft and distinct T-cell receptor features perfectly. We demonstrate, for the first time, the accumulation of large clonal populations of specific V alpha 2.3/V beta 22 T-cell receptor-expressing CD4(+) T-cells in the lungs of HLA-DRB1*03(+) sarcoidosis patients. Several distinct contact points between V alpha 2.3/V beta 22 receptors and HLA-DRB1*03 molecules suggest presentation of prototypic vimentin-derived peptides.

  • 391.
    Gräslund, Susanne
    et al.
    KTH, Superseded Departments, Biotechnology.
    Eklund, Malin
    KTH, Superseded Departments, Biotechnology.
    Falk, Ronny
    KTH, Superseded Departments, Biotechnology.
    Uhlén, Mathias
    KTH, Superseded Departments, Biotechnology.
    Nygen, Per-Åke
    KTH, Superseded Departments, Biotechnology.
    Ståhl, Stefan
    KTH, Superseded Departments, Biotechnology.
    A novel affinity gene fusion system allowing protein A-based recovery of non-immunoglobulin gene products2002In: Journal of Biotechnology, ISSN 0168-1656, E-ISSN 1873-4863, Journal of biotechnology, Vol. 99, no 1, p. 41-50Article in journal (Refereed)
    Abstract [en]

    An expression vector system has been developed, taking advantage of a novel, Staphylococcus aureus protein A (SPA)-binding affinity tag ZSPA-1, enabling straightforward affinity blotting procedures and efficient recovery by affinity purification of expressed gene products on readily available reagents and chromatography media. The 58 amino acid SPA-binding affinity tag ZSPA-1, was previously selected from a library constructed by combinatorial mutagenesis of a protein domain from SPA. An Escherichia coli expression vector for intracellular T7 promoter (PT7) driven production was constructed with an N-terminal dual affinity tag, consisting of a hexahistidyl (His6) tag in frame with the ZSPA-1 tag, thus allowing alternative affinity recovery methods. To evaluate the system, five cDNA clones from a mouse testis cDNA library were expressed, and two alternative blotting procedures were developed for convenient screening of expression efficiencies. The five produced fusion proteins were recovered on both immobilized metal-ion affinity chromatography (IMAC) columns and on Protein A-based chromatography media, to allow comparative studies. It was found that the Protein A-based recovery resulted in the highest degree of purity, and furthermore, gene products that were produced as inclusion bodies could after denaturation be efficiently affinity purified on Protein A-Sepharose in the presence of 0.5 M guanidine hydrochloride. The convenience and robustness of the presented expression system should make it highly suitable for various high-throughput protein expression efforts.

  • 392. Guenot, Marianne
    et al.
    Loizon, Séverine
    Howard, Jennifer
    Costa, Giulia
    Baker, David A.
    Mohabeer, Shaneel Y.
    Troye-Blomberg, Marita
    Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
    Moreau, Jean-Francois
    Dechanet-Merville, Julie
    Mercereau-Puijalon, Odile
    Mamani-Matsuda, Maria
    Behr, Charlotte
    Phosphoantigen Burst upon Plasmodium falciparum Schizont Rupture Can Distantly Activate V gamma 9V delta 2 T Cells2015In: Infection and Immunity, ISSN 0019-9567, E-ISSN 1098-5522, Vol. 83, no 10, p. 3816-3824Article in journal (Refereed)
    Abstract [en]

    Malaria induces potent activation and expansion of the V gamma 9V delta 2 subpopulation of gamma delta T cells, which inhibit the Plasmodium falciparum blood cycle through soluble cytotoxic mediators, abrogating merozoite invasion capacity. Intraerythrocytic stages efficiently trigger V gamma 9V delta 2 T-cell activation and degranulation through poorly understood mechanisms. P. falciparum blood-stage extracts are known to contain phosphoantigens able to stimulate V gamma 9V delta 2 T cells, but how these are presented by intact infected red blood cells (iRBCs) remains elusive. Here we show that, unlike activation by phosphoantigen-expressing cells, V gamma 9V delta 2 T-cell activation by intact iRBCs is independent of butyrophilin expression by the iRBC, and contact with an intact iRBC is not required. Moreover, blood-stage culture supernatants proved to be as potent activators of V gamma 9V delta 2 T cells as iRBCs. Bioactivity in the microenvironment is attributable to phosphoantigens, as it is dependent on the parasite DOXP pathway, on V gamma 9V delta 2 TCR signaling, and on butyrophilin expression by V gamma 9V delta 2 T cells. Kinetic studies showed that the phosphoantigens were released at the end of the intraerythrocytic cycle at the time of parasite egress. We document exquisite sensitivity of V gamma 9V delta 2 T cells, which respond to a few thousand parasites. These data unravel a novel framework, whereby release of phosphoantigens into the extracellular milieu by sequestered parasites likely promotes activation of distant V gamma 9V delta 2 T cells that in turn exert remote antiparasitic functions.

  • 393.
    Guerrerio, Pamela A.
    et al.
    NIAID, LAD, NIH, 9000 Rockville Pike, Bethesda, MD 20892 USA.
    Rasooly, Marjohn
    NIAID, LAD, NIH, 9000 Rockville Pike, Bethesda, MD 20892 USA.
    Gu, Wenjuan
    Leidos Biomed Res Inc, bClin Res Directorate, Clin Monitoring Res Program, NCI Campus, Frederick, MD USA.
    Levin, Samara
    NIAID, LAD, NIH, 9000 Rockville Pike, Bethesda, MD 20892 USA.
    Jhamnani, Rekha
    NIAID, LAD, NIH, 9000 Rockville Pike, Bethesda, MD 20892 USA.
    Milner, Joshua D.
    NIAID, LAD, NIH, 9000 Rockville Pike, Bethesda, MD 20892 USA.
    Stone, Kelly D.
    NIAID, NIH, Lab Allerg Dis, 9000 Rockville Pike, Bethesda, MD 20892 USA.
    Guerrerio, Anthony L.
    Johns Hopkins Univ, Baltimore, MD USA.
    Jones, Joseph
    Phadia US Inc, dImmuno Diagnost Branch, Thermo Fisher Sci, Portage, MI USA.
    Borres, Magnus P
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Women's and Children's Health, Research group (Dept. of women´s and children´s health), Paediatric Inflammation Research. ThermoFisher Sci, Uppsala, Sweden.
    Brittain, Erica
    NIH, Biostat Res Branch, DCR, Bldg 10, Bethesda, MD 20892 USA.
    IgE Testing Can Predict Food Allergy Status in Patients with Moderate-Severe Atopic Dermatitis2019In: Journal of Allergy and Clinical Immunology, ISSN 0091-6749, E-ISSN 1097-6825, Vol. 143, no 2, p. AB130-AB130, article id 392Article in journal (Other academic)
  • 394.
    Guldevall, Karolin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Single Cell Investigations of the Functional Heterogeneity Within Immune Cell Populations: a Microchip-based Study2014Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Immune cell populations are constantly divided into smaller and smaller subsets defined by newly emerging cellular markers. However, there is a growing awareness of the functional heterogeneities in between cells even within small populations, in addition to the heterogeneity over time. One may ask whether a population is correctly defined only by cellular markers or if the functionality should be regarded as well? Many of today’s techniques only measure at the population level, giving an average estimate of the behavior of that pool of cells, but failing to detect rare possibly important events. Thus, high-throughput experimental approaches to analyze single cells over time are required to address cellular heterogeneity.

    Progress in the fields of microfabrication, microscopy and computing have paved the way for increasingly efficient tools for studies on the single cell level, and a variety of devices have been described by others. However, few of them are suitable for long-term imaging of dynamic events such as cell-cell interactions or migration. In addition, for efficient recording of many individual events it is desirable to scale down the cells’ interaction volume; not only to shorten the time to interaction, but also to increase the number of individual events in a given area; thereby pushing a screening approach.

    To address these questions, a complete microwell array system for imaging of immune cell responses with single-cell resolution was designed. The platform consists of a range of silicon-glass microchips with arrays of miniature wells for incubation of cells and a custom made holder that fits conventional microscopes. The device has been designed to allow cells to be kept viable for several days in the wells, to be easy to use and to allow high-resolution imaging. Five different designs were fabricated; all with a specific type of assay in mind, and were evaluated regarding biocompatibility and functionality. Here, the design aimed for screening applications is the main focus. In this approach a large amount, tens of thousands, of small wells are imaged two to three times: first directly post-seeding of effector and target cells to register the well’s content, and second after some time has passed to allow for cell-cell interactions. The final read-out is the number of killed target cells in each well, making an automatic cell counting protocol necessary in order to analyze the massive amount of data generated.

    We here show that our silicon microwell platform allows long-term studies with the possibility of both time-lapse and high-resolution imaging of a variety of immune cell behavior. Using both time-lapse imaging and the screening approach we confirmed and investigated immune cell heterogeneity within NK cell populations in regards to both cytotoxicity and migrational behavior. In addition, two different types of cytolytic behavior in NK cells, termed fast and slow killing, were described and evaluated in regards to dynamic parameters; like conjugation and attachment time. We could also quantify the type of cytolytic response in relation to serial killing NK cells, and saw that serial killing NK cells more often induced fast target cell death. Further investigations using the screening approach have shown that serial killing NK cells also differ from other NK cells in their morphology, being both larger and with a more elongated shape. So far the platform has been used to gain better understanding of some aspects of NK cell biology, but there is still much left to explore. With the addition of an automatic counting program, the large numbers of wells that can be simultaneously imaged will provide new statistical information and enable higher throughput.

    Altogether, our family of techniques enables novel types of cellular imaging assays allowing data collection at a level of resolution not previously obtained – this was shown to be important for performing basic cell biological studies, but may also prove valuable in the proposed future medical applications such as adoptive cell therapy and stem cell transplantation.

  • 395.
    Guldevall, Karolin
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Gustafsson, Karin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Forslund, Elin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Frisk, Thomas
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Manneberg, Otto
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Olofsson, Per E.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Tauriainen, Johanna
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Stikvoort, Arwen
    Karolinska Institute.
    Vanherberghen, Bruno
    KTH, School of Engineering Sciences (SCI), Applied Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Brismar, Hjalmar
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Mattsson, Jonas
    Karolinska Institute.
    Kärre, Klas
    Karolinska Institute.
    Uhlin, Michael
    Karolinska Institute.
    Önfelt, Bjorn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Microchip screening platform for assessment of cytotoxic effector cellsManuscript (preprint) (Other academic)
    Abstract [en]

    Here we report a screening platform for assessment of the cytotoxic potential of individual natural killer (NK) or T cells within larger populations. Human primary NK cells or human Epstein-Barr virus (EBV)- specific T cells were distributed across a silicon-glass microchip containing 32 400 individual microwells loaded with target cells. Through fluorescence screening and automated image analysis the numbers of effector and live or dead target cells in each well could be assessed at different time-points after initial mixing. Cytotoxicity was also studied by time-lapse live-cell imaging in microwells quantifying the killing potential of individual NK cells. Although most resting NK cells (≈75%) were non-cytotoxic to the leukemia cell line K562, some NK cells were able to kill several (≥3) target cells within the 12 hours long experiment. We demonstrate that this assay can be used to enumerate and characterize cytotoxic cells, something that could find clinical applications, e.g. in the selection of donors for stem cell transplantation or generation of highly specific and cytotoxic cells for adoptive immunotherapy.

  • 396.
    Gullberg, Martin
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Immunology.
    Recognition requirements and regulatory events directing T cell responses1983Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    The present study has considered cellular and molecular requirements in T cell responses. The central role of T cell growth factors (TCGF) in T cell responses prompted us to study the regulatory events directing TCGF production in lectin stimulated cultures. It was found that normal spleen cells, activated with Concanavalin A for 24 h, develop suppressive cells that block de novo TCGF production by fresh spleen cells. The induction time for effector suppressor cells (nonadherent, Lyt-2-positive T cells) was found to be 18 h and to parallel the termination of TCGF production in situ. The suppressive mechanism is neither iji situ absorption of TCGF produced at control rates nor killing of TCGF producing cells. These results suggest that suppression of TCGF production is an active process which directly and reversibly blocks TCGF-producing cells.

    This study also indicated that ConA induced a very limited proliferation of Lyt-2- T helper cells (TH) in unselected T cell populations. The activation and growth requirements of Lyt-1+ TH cells were directly investigated and compared with those of Lyt-2+ cytotoxic T lymphocytes (CTL), as defined by the selective expression of Lyt differentiation antigens and functional activities. This analysis revealed a profound difference in activation and growth requirements between these T cell subsets. Thus, while Lyt-2+ CTL precursors can be induced to TCGF reactivity by soluble lectins, in the absence of specialized accessory cells,; Lyt-2" TH cell precursors show a strict accessory cell requirement both for activation and proliteration. Finally, the low level of TH cell effector function, detected in a primary responses to allo-MHC-antigens or lectins, appears to be due to the development of suppressive Lyt2+ T cells.

    The functional relevance of Lyt-2 antigens expressed on CTL membranes was further assessed in the last part of this study. Two distinct activation systems were used, namely MHC-antigens, provided as UV-irradiated stimulator cells or polyclonal induction by a 4 h pulse, with lectins. Both procedures were shown to selectively induce Lyt-2+ CTL precursors into TCGF reactivity without leading to mitosis, unless TCGF was added. In both cases it was found that monoclonal anti-Lyt-2 antibodies inhibited the two antigen- dependent phases of CTL responses namely, the initial induction step and target cytolysis. The analogy observed between antigen specific and lectin mediated indueton and target cytolysis, with regard to the susceptibility of inhibition by anti-Lyt-2 antibodies has lead to a general hypothesis on CTL activation.

  • 397.
    Gunaltay, Sezin
    et al.
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden.
    Ghiboub, M.
    Frenche Comte Univ, Besancon, France.
    Hultgren, Olof
    Örebro University, School of Medicine, Örebro University, Sweden.
    Hultgren-Hörnquist, Elisabeth
    Örebro University, School of Medicine, Örebro University, Sweden.
    The role of IL-37 on cytokine responses downstream of TLR4 and TLR5 signalling in intestinal epithelial cells2014In: Immunology, ISSN 0019-2805, E-ISSN 1365-2567, Vol. 143, p. 94-95Article in journal (Other academic)
  • 398.
    Gunaltay, Sezin
    et al.
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden.
    Kumawat, Ashok Kumar
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. Institute of Infection, College of Medical, Veterinary & Life Sciences, University of Glasgow, Glasgow, United Kingdom.
    Nyhlin, Nils
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. Örebro University Hospital.
    Bohr, Johan
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. Örebro University Hospital.
    Tysk, Curt
    Örebro University Hospital. Division of Gastroenterology, Department of Medicine, Örebro University, Örebro, Sweden.
    Hultgren, Olof
    Örebro University Hospital.
    Hultgren-Hörnquist, Elisabeth
    Örebro University, School of Medicine, Örebro University, Sweden.
    Enhanced levels of chemokines and their receptors in the colon of microscopic colitis patients indicate mixed immune cell recruitment2015In: Mediators of Inflammation, ISSN 0962-9351, E-ISSN 1466-1861, article id 132458Article in journal (Refereed)
    Abstract [en]

    Microscopic colitis (MC), comprising collagenous colitis (CC) and lymphocytic colitis (LC), is a common cause of chronic diarrhea. Various immune cell infiltrations in the epithelium and lamina propria are seen in MC immunopathology. We compared gene and protein expressions of different immune cell attracting chemokines and their receptors in colon biopsies from MC patients in active disease or histopathological remission (CC/LC-HR) with controls, using qRT-PCR and Luminex, respectively. CC and LC patients with active disease demonstrated a mixed chemokine profile with significantly enhanced gene and/or protein expressions of the chemokines CCL2, CCL3, CCL4, CCL5, CCL7, CCL22, CXCL8, CXCL9, CXCL10, CXCL11, and CX(3)CL1 and the receptors CCR2, CCR3, CCR4, CXCR1, CXCR2, and CX(3)CR1. Enhanced chemokine/chemokine receptor gene and protein levels in LC-HR patients were similar to LC patients, whereas CC-HR patients demonstrated almost normalized levels. These findings expand the current understanding of the involvement of various immune cells in MC immunopathology and endorse chemokines as potential diagnostic markers as well as therapeutic candidates. Moreover, this study further supports the hypothesis that CC and LC are two different entities due to differences in their immunoregulatory responses.

  • 399.
    Gunnar, Erika
    Linköping University, Department of Clinical and Experimental Medicine, Division of Microbiology and Molecular Medicine. Linköping University, Faculty of Medicine and Health Sciences.
    Regulatory programs controlling profileration during Drosophila nervous system development2017Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    The central nervous system (CNS) is the most complex organ in the body, responsible for complex functions, including thinking, reasoning and memory. The CNS contains cells of many different types, often generated in vast numbers. Hence, CNS development requires precise genetic control of both cell fate and of cell proliferation, to generate the right number of cells, with the proper identity, and in the proper location. The cells also need to make connections with each other for correct signaling and function. This complexity evokes the question of how this is regulated. How does the stem cells, responsible for building the CNS, know how many times to divide, and how does the daughters know which identity to acquire and in which location they shall end up? During Drosophila melanogaster development, the neuroblasts (NBs) are responsible for generating the CNS. In each hemisegment, every NB is unique in identity, and generates a predetermined number of daughters with specific identities. The lineages of different NBs vary in size, but are always the same for each specific NB, and the division modes of each NBs is hence stereotyped. Most NBs start dividing by renewing themselves while generating daughters that will in turn divide once to generate two neurons and/or glia (denoted type I mode). Many, maybe all, NBs later switch to generating daughters that will differentiate directly into a neuron or glia (denoted type 0 mode). This type I>0 switch occurs at different time-points during lineage progression, and influences the total numbers of cells generated from a single NB.

    The work presented in this thesis aimed at investigating the genetic regulation of proliferation, with particular focus on the type I>0 switch. In the first project, the implication of the Notch pathway on the type I>0 switch was studied. Mutants of the Notch pathway do not switch, and the results show that the Notch pathway regulates the switch by activation of several target genes, both regulators and cell cycle genes. One of the target genes, the E(spl)-C genes, have been difficult to study due to functional redundancy. This study reveals that even though they can functionally compensate for each other, they have individual functions in different lineages. Regarding cell cycle genes, both Notch and E(spl)-C regulate several key cell cycle genes, and molecular analysis indicated that this regulation is direct. In the second project we studied the seq gene, previously identified in a genetic screen. We found that seq controls the type I>0 switch by regulating the key cell cycle genes, but also through interplay with the Notch pathway. Notch and seq stop proliferation, and in the third project we wanted to identify genes that drive proliferation. We found that there is battery of early NB genes, socalled early factors, which activate the cell cycle, and drive NB and daughter proliferation. These are gradually replaced by late regulators, and the interplay between early and late factors acts to achieve precise control of lineage progression.

    The work presented here increases our understanding of how regulatory programs act to control the development of the CNS; to generate the right number of cells of different identities. These results demonstrate the importance of correct regulation of proliferation in both stem cells and daughters. Problems in this control can result in either an underdeveloped CNS or loss of control such as in cancer. Knowledge about these regulatory programs can contribute to the development of therapeutics against these diseases.

  • 400.
    Gustafson, Elisabet
    et al.
    Uppsala University Hospital, Sweden.
    Asif, Sana
    Uppsala University, Sweden.
    Kozarcanin, Huda
    Uppsala University, Sweden.
    Elgue, Graciela
    Uppsala University, Sweden.
    Meurling, Staffan
    Uppsala University Hospital, Sweden.
    Nilsson Ekdahl, Kristina
    Linnaeus University, Faculty of Health and Life Sciences, Department of Chemistry and Biomedical Sciences.
    Nilsson, Bo
    Uppsala University, Sweden.
    Control of IBMIR Induced by Fresh and Cryopreserved Hepatocytes by Low Molecular Weight Dextran Sulfate Versus Heparin2017In: Cell Transplantation, ISSN 0963-6897, E-ISSN 1555-3892, Vol. 26, no 1, p. 71-81Article in journal (Refereed)
    Abstract [en]

    Rapid destruction of hepatocytes after hepatocyte transplantation has hampered the application of this procedure clinically. The instant blood-mediated inflammatory reaction (IBMIR) is a plausible underlying cause for this cell loss. The present study was designed to evaluate the capacity of low molecular weight dextran sulfate (LMW-DS) to control these initial reactions from the innate immune system. Fresh and cryopreserved hepatocytes were tested in an in vitro whole-blood model using ABO-compatible blood. The ability to elicit IBMIR and the capacity of LMW-DS (100 mu g/ml) to attenuate the degree of activation of the cascade systems were monitored. The effect was also compared to conventional anticoagulant therapy using unfractionated heparin (1 IU/ml). Both fresh and freeze thawed hepatocytes elicited IBMIR to the same extent. LMW-DS reduced the platelet loss and maintained the cell counts at the same degree as unfractionated heparin, but controlled the coagulation and complement systems significantly more efficiently than heparin. LMW-DS also attenuated the IBMIR elicited by freeze thawed cells. Therefore, LMW-DS inhibits the cascade systems and maintains the cell counts in blood triggered by both fresh and cryopreserved hepatocytes in direct contact with ABO-matched blood. LMW-DS at a previously used and clinically applicable concentration (100 mu g/ml) inhibits IBMIR in vitro and is therefore a potential IBMIR inhibitor in hepatocyte transplantation.

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