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  • 201.
    Tafazoli, Farideh
    et al.
    Linköping University, Faculty of Arts and Sciences. Linköping University, Department of health and environment.
    Magnusson, Karl-Eric
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology .
    Ultrastructural characteristics of the perturbation of the barrier properties of epithelial cells by pathogenic Yersiniae pseudotuberculosis bacteria2000In: Molecular Biology of the Cell, ISSN 1059-1524, E-ISSN 1939-4586, Vol. 11, p. 2653-Conference paper (Other academic)
  • 202. Order onlineBuy this publication >>
    Tejle, Katarina
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Leishmania donovani Lipophosphoglycan: Modulation of Macrophage and Dendritic Cell Function2006Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Leishmania donovani is a blood-borne tropicial parasite, which infects humans through bites by Phlebotomus sandflies. The parasite survives and multiplies inside macrophages in inner organs, and causes the deadly disease visceral leishmaniasis (Kala-Azar).

    Macrophages and dendritic cells (DC) are professional antigen-presenting cells involved in the initiation of immune responses. Immature DC are present in all tissues where they internalise and process antigen, in response to which they migrate from tissue, into draining lymphoid organs, undergo maturation and present antigens to lymphocytes. Control measures for leishmaniasis include testing of new diagnostics and development of affordable and effective vaccines for humans.

    Lipophosphoglycan (LPG) is the major surface component of Leishmania donovani promastigotes. LPG comprises a membrane-anchoring lysophosphatidylinositol part and an extracellular chain of disaccharide phosphates. These repetitions are crucial for parasite survival inside macrophages following phagocytosis. LPG has several specific effects on the host cell including inhibition of protein kinase C (PKC) activity, and inhibition of phagosomal maturation, a process requiring depolymerization of periphagosomal F-actin.

    Confocal microscopy and image analysis were used to follow F-actin dynamics in single macrophages during phagocytosis of L. donovani promastigotes and LPG-coated particles. F-actin did not depolymerize, but instead progressively polymerized around phagosomes with LPG-containing prey. This correlated with reduced translocation of PKCα to the phagosome and blocked phagosomal maturation. LPG also inhibited cortical actin turnover, which could be the underlying cause of the reduced uptake of LPG-containing prey. Extracellular- and intracellular calcium was necessary for phagocytosis, periphagosomal F-actin breakdown and phagosomal maturation in macrophages interacting with unopsonized prey,and for the action of LPG.

    We also studied F-actin turnover in macrophages overexpressing dominant-negative (DN) PKCα. DN PKCα macrophages showed increased amounts of cortical F-actin, decreased phagocytic capacity, inhibition of periphagosomal F-actin breakdown and defective phagosomal maturation. When DN PKCα macrophages interacted with LPG-containing prey, phagocytosis was almost completely blocked.

    Moreover, we found that Leishmania promastigotes and particularly LPG inhibit DC maturation and detachment from distinct surfaces. Thus, LPG from Leishmania donovani could directly inhibit DC migration to lymphoid organs, antigen-presentation and development of immunity.

    List of papers
    1. Leishmania donovani lipophosphoglycan causes periphagosomal actin accumulation: correlation with impaired translocation of PKCα and defective phagosome maturation
    Open this publication in new window or tab >>Leishmania donovani lipophosphoglycan causes periphagosomal actin accumulation: correlation with impaired translocation of PKCα and defective phagosome maturation
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    2001 (English)In: Cellular Microbiology, ISSN 1462-5814, E-ISSN 1462-5822, Vol. 3, no 7, p. 439-447Article in journal (Refereed) Published
    Abstract [en]

    Lipophosphoglycan (LPG) is the major surface glycoconjugate of Leishmania donovani promastigotes. The repeating disaccharide–phosphate units of LPG are crucial for promastigote survival inside macrophages and establishment of infection. LPG has a number of effects on the host cell, including inhibition of PKC activity, inhibition of nitric oxide production and altered expression of cytokines. LPG also inhibits phagosomal maturation, a process requiring depolymerization of periphagosomal F-actin. In the present study, we have characterized the dynamics of F-actin during the phagocytosis of L. donovani promastigotes in J774 macrophages. We observed that F-actin accumulated progressively around phagosomes containing wild-type L. donovani promastigotes during the first hour of phagocytosis. Using LPG-defective mutants and yeast particles coated with purified LPG, we obtained evidence that this effect could be attributed to the repeating units of LPG. LPG also disturbed cortical actin turnover during phagocytosis. The LPG-dependent accumulation of periphagosomal F-actin correlated with an impaired recruitment of the lysosomal marker LAMP1 and PKCα to the phagosome. Accumulation of periphagosomal F-actin during phagocytosis of L. donovani promastigotes may contribute to the inhibition of phagosomal maturation by physically preventing vesicular trafficking to and from the phagosome.

    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:liu:diva-13848 (URN)10.1046/j.1462-5822.2001.00127.x (DOI)
    Available from: 2006-05-23 Created: 2006-05-23 Last updated: 2017-12-13Bibliographically approved
    2. Phagocytosis and phagosome maturation are regulated by calcium in J774 macrophages interacting with un-opsonized prey
    Open this publication in new window or tab >>Phagocytosis and phagosome maturation are regulated by calcium in J774 macrophages interacting with un-opsonized prey
    2002 (English)In: Bioscience Reports, ISSN 0144-8463, Vol. 22, no 5-6, p. 529-540Article in journal (Refereed) Published
    Abstract [en]

    Phagocytosis by neutrophils, macrophages, and other professional phagocytes requires rapid remodeling of actin. Early phagosomes are surrounded by a rim of F-actin that is disassembled during phagosomoal maturation. Breakdown of periphagosomal F-actin and phagolysosome fusion are calcium dependent processes in neutrophils interacting with serum-opsonized prey, but appears to be calcium independent in macrophages interacting with serum- or IgG-opsonized prey. In the present study, we found that calcium was necessary for phagocytosis, breakdown of periphagosomal F-actin, and phagosomal maturation in J774 macrophages interacting with unopsonized prey. We also observed that lipophosphoglycan (LPG) from Leishmania donovani promastigotes required calcium to exert its inhibitory effect on macrophage phagocytosis and periphagosomal F-actin breakdown. We conclude that calcium is essential for phagocytosis, depolymerization of periphagosomal F-actin, and phagosomal maturation in J774 macrophages interacting with unopsonized prey, as well as for proper functioning of LPG.

    Keywords
    Macrophage, phagocytosis, calcium, actin, phagosome maturation, lipophosphoglycan
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:liu:diva-13849 (URN)10.1023/A:1022025903688 (DOI)
    Available from: 2006-05-23 Created: 2006-05-23 Last updated: 2013-09-18
    3. Role of protein kinase C α for uptake of unopsonized prey and phagosomal maturation in macrophages
    Open this publication in new window or tab >>Role of protein kinase C α for uptake of unopsonized prey and phagosomal maturation in macrophages
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    2003 (English)In: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 302, no 4, p. 653-658Article in journal (Refereed) Published
    Abstract [en]

    Protein kinase C α (PKCα) participates in F-actin remodeling during phagocytosis and phagosomal maturation in macrophages. Leishmania donovani promastigotes, which inhibit phagosomal maturation, cause accumulation of periphagosomal F-actin instead of the dissassembly observed around other prey [Cell. Microbiol. 7 (2001) 439]. This accumulation is induced by promastigote lipophosphoglycan (LPG), which has several effects on macrophages including inhibition of PKCα. To investigate a possible connection between PKCα and LPG’s effects on actin dynamics, we utilized RAW264.7 macrophages overexpressing dominant-negative PKCα (DN PKCα). We found increased cortical F-actin and decreased phagocytic capacity, as well as defective periphagosomal F-actin breakdown and inhibited phagosomal maturation in the DN PKCα-overexpressing cells, effects similar to those seen in controls subjected to LPG-coated prey. The results indicate that PKCα is involved in F-actin turnover in macrophages and that PKCα-dependent breakdown of periphagosomal F-actin is required for phagosomal maturation, and endorse the hypothesis that intracellular survival of L. donovani involves inhibition of PKCα by LPG.

    Keywords
    PKCα, Actin, Phagocytosis, Macrophage, Lipophosphoglycan
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:liu:diva-13850 (URN)10.1016/S0006-291X(03)00231-6 (DOI)
    Available from: 2006-05-23 Created: 2006-05-23 Last updated: 2017-12-13Bibliographically approved
    4. Leishmania donovani promastigotes block maturation, increase integrin expression and inhibit detachment of human monocytederived dendritic cells – a role for lipophosphoglycan (LPG)
    Open this publication in new window or tab >>Leishmania donovani promastigotes block maturation, increase integrin expression and inhibit detachment of human monocytederived dendritic cells – a role for lipophosphoglycan (LPG)
    Manuscript (Other academic)
    Identifiers
    urn:nbn:se:liu:diva-13851 (URN)
    Available from: 2006-05-23 Created: 2006-05-23 Last updated: 2010-01-13
    Download full text (pdf)
    FULLTEXT01
  • 203.
    Tejle, Katarina
    et al.
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology .
    Lindroth, Margaretha
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology .
    Magnusson, Karl-Eric
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology .
    Rasmusson, Birgitta
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology .
    Wild-type Leishmania donovani promastigotes block maturation, increase integrin expression and inhibit detachment of human monocyte-derived dendritic cells - The influence of phosphoglycans2008In: FEMS Microbiology Letters, ISSN 0378-1097, E-ISSN 1574-6968, Vol. 279, no 1, p. 92-102Article in journal (Refereed)
    Abstract [en]

    The protective immune response against the parasite, including the role of dendritic cells (DC) in the course of infection, plays a fundamental role. This study shows that wild-type (WT) Leishmania promastigotes and specifically the phosphoglycans family of virulence-associated antigens inhibit human monocyte-derived dendritic cells (MoDC) maturation and detachment to distinct surfaces. Immature phagocytosis of Leishmania donovani promastigotes by immature MoDC results in the increased expression of CD11b and CD51, and inhibition of cell detachment to distinct surfaces, which was dependent on the presence of phosphoglycans. These findings demonstrate that phosphoglycans of WT L. donovani might also inhibit human DC migration to lymphoid organs. © 2007 Federation of European Microbiological Societies.

  • 204.
    Tejle, Katarina
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Magnusson, Karl-Eric
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Rasmusson, Birgitta
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Phagocytosis and phagosome maturation are regulated by calcium in J774 macrophages interacting with un-opsonized prey2002In: Bioscience Reports, ISSN 0144-8463, Vol. 22, no 5-6, p. 529-540Article in journal (Refereed)
    Abstract [en]

    Phagocytosis by neutrophils, macrophages, and other professional phagocytes requires rapid remodeling of actin. Early phagosomes are surrounded by a rim of F-actin that is disassembled during phagosomoal maturation. Breakdown of periphagosomal F-actin and phagolysosome fusion are calcium dependent processes in neutrophils interacting with serum-opsonized prey, but appears to be calcium independent in macrophages interacting with serum- or IgG-opsonized prey. In the present study, we found that calcium was necessary for phagocytosis, breakdown of periphagosomal F-actin, and phagosomal maturation in J774 macrophages interacting with unopsonized prey. We also observed that lipophosphoglycan (LPG) from Leishmania donovani promastigotes required calcium to exert its inhibitory effect on macrophage phagocytosis and periphagosomal F-actin breakdown. We conclude that calcium is essential for phagocytosis, depolymerization of periphagosomal F-actin, and phagosomal maturation in J774 macrophages interacting with unopsonized prey, as well as for proper functioning of LPG.

  • 205.
    Thiere, Geraldine
    et al.
    Halmstad University.
    Milenkovski, Susanne
    Linköping University, Department of Clinical and Experimental Medicine. Linköping University, Faculty of Health Sciences.
    Lindgren, Per-Eric
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Sahlen, Goran
    Halmstad University.
    Berglund, Olof
    Lund University.
    Weisner , Stefan E B
    Halmstad University.
    Wetland creation in agricultural landscapes: Biodiversity benefits on local and regional scales2009In: BIOLOGICAL CONSERVATION, ISSN 0006-3207 , Vol. 142, no 5, p. 964-973Article in journal (Refereed)
    Abstract [en]

    Wetland creation aiming at a simultaneous increase in nutrient retention and species diversity in agricultural landscapes has recently become applied as a catchment-scale compensation measure for past wetland losses. Here, we evaluate if, and to what extend, dual-purpose wetlands benefit local and regional diversity of agricultural landscapes. We analysed composition and alpha, beta, and gamma diversity of aquatic macroinvertebrate assemblages among dual-purpose wetlands in an agricultural region in southwest Sweden in relation to local (water quality, wetland morphology, succession stage, proximity to other aquatic habitats) and landscape parameters (regional connectivity, wetland density). Diversity of mature agricultural ponds was used as a standard to evaluate the value of dual-purpose wetlands. Dual-purpose wetlands sustained alpha, beta, and gamma diversity similar to that of natural lentic water bodies in agricultural landscapes in the region and elsewhere. Over 80% of the overall species richness was attributed to beta diversity, and each created wetland contributed to overall species accumulation. Ecosystem parameters explained 19% of the compositional variation among assemblages, but were only marginally related to diversity. Wetland density promoted alpha and gamma diversity, while spatial heterogeneity (beta) remained equally high, independent of wetland density. Our results indicate that catchment-scale wetland creation for simultaneous retention and diversity purposes benefits the biodiversity of agricultural landscapes, particularly if the density of aquatic habitats is increased by at least 30%.

  • 206.
    Tjellstrom, Bo
    et al.
    Karolinska Institute, Sweden Norrkoping Hospital, Sweden .
    Högberg, Lotta
    Linköping University, Department of Clinical and Experimental Medicine, Pediatrics. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre of Paediatrics and Gynecology and Obstetrics, Department of Paediatrics in Norrköping.
    Stenhammar, Lars
    Linköping University, Department of Clinical and Experimental Medicine, Pediatrics. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre of Paediatrics and Gynecology and Obstetrics, Department of Paediatrics in Norrköping.
    Magnusson, Karl-Eric
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Midtvedt, Tore
    Karolinska Institute, Sweden .
    Norin, Elisabeth
    Karolinska Institute, Sweden .
    Sundqvist, Tommy
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Effect of exclusive enteral nutrition on gut microflora function in children with Crohns disease2012In: Scandinavian Journal of Gastroenterology, ISSN 0036-5521, E-ISSN 1502-7708, Vol. 47, no 12, p. 1454-1459Article in journal (Refereed)
    Abstract [en]

    Objective. Exclusive enteral nutrition (EEN) is a first-line treatment in children with active Crohns disease (CD) but is seldom used in adults with active disease. The mode of action of EEN in suppressing mucosa] inflammation is not fully understood, but modulation of intestinal microflora activity is one possible explanation. The aim of this study was to investigate the effect of 6-week EEN in children with active CD, with special reference to intestinal microflora function. Materials and methods. Fecal samples from 18 children (11 boys, 7 girls; median age 13.5 years) with active CD (13 children with small bowel/colonic and 5 with perianal disease) were analyzed for short chain fatty acid (SCFA) pattern as marker of gut microflora function. The children were studied before and after EEN treatment. Results from 12 healthy teenagers were used for comparison. Results. Eleven (79%) of the children with small bowel/colonic CD responded clinically positively to EEN treatment showing decreased levels of pro-inflammatory acetic acid as well as increased concentrations of anti-inflammatory butyric acids and also of valeric acids, similar to the levels in healthy age-matched children. In children with active perianal CD, however, EEN had no positive effect on clinical status or inflammatory parameters. Conclusions. The authors present new data supporting the hypothesis that the well-documented anti-inflammatory effect of EEN in children with active small bowel/colonic CD is brought about by modulation of gut microflora activity, resulting in an anti-inflammatory SCFA pattern. By contrast, none of the children with perianal disease showed clinical or biochemical improvement after EEN treatment.

  • 207.
    Tjellström, Bo
    et al.
    Östergötlands Läns Landsting, Center of Paediatrics and Gynaecology and Obstetrics, Department of Paediatrics in Norrköping. Department of Microbiology, Tumor and Cell Biology, Karolinska Institute, Stockholm, Sweden.
    Högberg, Lotta
    Linköping University, Department of Clinical and Experimental Medicine, Pediatrics. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Center of Paediatrics and Gynaecology and Obstetrics, Department of Paediatrics in Norrköping.
    Stenhammar, Lars
    Linköping University, Department of Clinical and Experimental Medicine, Pediatrics. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Center of Paediatrics and Gynaecology and Obstetrics, Department of Paediatrics in Norrköping.
    Fälth-Magnusson, Karin
    Linköping University, Department of Clinical and Experimental Medicine, Pediatrics. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Center of Paediatrics and Gynaecology and Obstetrics, Department of Paediatrics in Linköping.
    Magnusson, Karl-Erik
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Norin, Elisabeth
    Department of Microbiology, Tumor and Cell Biology, Karolinska Institute, Stockholm, Sweden.
    Sundqvist, Tommy
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Midtvedt, Tore
    Department of Microbiology, Tumor and Cell Biology, Karolinska Institute, Stockholm, Sweden.
    Faecal short-chain fatty acid pattern in childhood coeliac disease is normalised after more than one year's gluten-free diet2013In: Microbiological Ecology in Health and Disease, ISSN 0891-060X, E-ISSN 1651-2235, Vol. 24Article in journal (Refereed)
    Abstract [en]

    OBJECTIVE: Recent work indicates that the gut microflora is altered in patients with coeliac disease (CD). Faecal short-chain fatty acids (SCFAs) are produced by the gut microflora. We have previously reported a high SCFA output in children with symptomatic and asymptomatic CD at presentation, as well as in CD children on a gluten-free diet (GFD) for less than 1 year, indicating deviant gut microfloral function. In this report, we focus on faecal SCFA production in coeliacs on GFD for more than 1 year.

    MATERIALS AND METHODS: Faecal samples were collected from 53 children with CD at presentation, 74 coeliac children on GFD for less than 1 year, and 25 individuals diagnosed with CD in childhood and on GFD for more than 1 year. The control group comprised 54 healthy children (HC). The faecal samples were analysed to show the SCFA pattern taken as a marker of gut microflora function. We applied a new fermentation index, reflecting the inflammatory activity of the SCFAs (amount of acetic acid minus propionic acid and n-butyric acid, together divided by the total amount of SCFAs).

    RESULTS: In coeliacs on GFD for more than 1 year, the individual SCFAs, total SCFA, and fermentation index did not differ significantly from the findings in controls. In contrast, the faecal SCFA level was clearly higher in coeliacs treated with GFD for less than 1 year compared to those more than 1 year.

    CONCLUSIONS: This is the first study on SCFA patterns in faecal samples from individuals with CD on GFD for more than 1 year. Our study indicates that the disturbed gut microflora function in children with CD at presentation and after less than 1 year of GFD, previously demonstrated by us, is normalised on GFD for more than 1 year.

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  • 208.
    Tjellström, Bo
    et al.
    Karolinska Institute.
    Stenhammar, Lars
    Linköping University, Department of Clinical and Experimental Medicine, Pediatrics . Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre of Paediatrics and Gynecology and Obstetrics, Department of Paediatrics in Norrköping.
    Högberg, Lotta
    Linköping University, Department of Clinical and Experimental Medicine, Pediatrics . Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre of Paediatrics and Gynecology and Obstetrics, Department of Paediatrics in Norrköping.
    Fälth-Magnusson, Karin
    Linköping University, Department of Clinical and Experimental Medicine, Pediatrics . Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre of Paediatrics and Gynecology and Obstetrics, Department of Paediatrics in Linköping.
    Magnusson, Karl-Eric
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Midtvedt, Tore
    Karolinska Institute.
    Sundqvist, Tommy
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Norin, Elisabeth
    Karolinska Institute.
    Screening-detected and symptomatic untreated celiac children show similar gut microflora-associated characteristics2010In: SCANDINAVIAN JOURNAL OF GASTROENTEROLOGY, ISSN 0036-5521, Vol. 45, no 9, p. 1059-1062Article in journal (Refereed)
    Abstract [en]

    Objective. The aim of this study was to investigate the metabolic function of intestinal microflora in children with screening-detected celiac disease (CD) to see if there is an aberrant gut flora in screening-detected CD similar to symptomatic CD and contrary to healthy controls. Materials and methods. As part of a Swedish multicenter screening for CD, 912 12-year-old children were screened with serum anti-human tissue transglutaminase-IgA. Small bowel biopsy specimens from children with positive serology revealed 17 individuals with CD. The functional status of the intestinal microflora was evaluated by gas liquid chromatography of short chain fatty acids (SCFAs) in fecal samples. Our previously published findings in children with symptomatic CD and healthy controls were used as comparison. Results. The children with screening-detected CD had a similar fecal SCFA profile to children with symptomatic CD, but differed significantly from that in healthy children. Conclusions. This is the first study on SCFA patterns in fecal samples from children with screening-detected CD. The similarity of the fecal SCFA profile in screening-detected and symptomatic CD indicates common pathogenic mechanisms. This could open the way for new therapeutic or prophylactic measures based on novel biological principles.

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  • 209.
    Tjernberg, Ivar
    et al.
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Infectious Diseases .
    Schön, Thomas
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology .
    Ernerudh, Jan
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Clinical Immunology . Östergötlands Läns Landsting, Centre for Laboratory Medicine, Department of Clinical Immunology and Transfusion Medicine.
    Carlsson-Wistedt, Annika
    Clinical Microbiology Kalmar County Hospital, Kalmar.
    Forsberg, Pia
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Infectious Diseases . Östergötlands Läns Landsting, Centre for Medicine, Department of Infectious Diseases in Östergötland.
    Eliasson, Ingvar
    Department of Clinical Microbiology and Immunology Lund University Hospital.
    C6-peptide serology as diagnostic tool in neuroborreliosis2008In: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS), ISSN 0903-4641, E-ISSN 1600-0463, Vol. 116, no 5, p. 393-399Article in journal (Refereed)
    Abstract [en]

    The aim of this study was to evaluate the usefulness of borrelia serology (Quick ELISA C6 Borrelia assay kit) as a diagnostic tool in cases of suspected neuroborreliosis. A retrospective patient material consisting of 124 paired serum and cerebrospinal fluid samples with a positive anti-borrelia antibody index (AI) using the IDEIA Lyme Neuroborreliosis test was compared with 124 AI-negative matched control subjects. The patients were divided into four groups based on presence of pleocytosis and age above or below 12 years. The presence of positive C6 serology in AI-positive patients with pleocytosis was 89% (83/93), significantly different (p<0.01) from in patients without pleocytosis (58%, 18/31). In AI-positive patients aged ≥12 years with pleocytosis, 94% (51/54) had a positive C6 serology. Of AI-positive patients with a symptom duration of more than 30 days, 93% (27/29) were positive by the C6 test. We conclude that the C6 serum test, together with clinical evaluation, is a powerful diagnostic tool in adult (≥12 years) European patients with suspected neuroborreliosis with a symptom duration of more than 30 days. Patients with suspected neuroborreliosis and positive C6 results should be further investigated by lumbar puncture for definite diagnosis. © The Authors 2008.

  • 210.
    Tjomsland, Vegard
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Molecular Virology . Linköping University, Faculty of Health Sciences.
    Sandström, Per
    Linköping University, Department of Clinical and Experimental Medicine, Surgery . Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre of Surgery and Oncology, Department of Surgery in Östergötland.
    Spangeus, Anna
    Linköping University, Department of Medicine and Health Sciences, Internal Medicine . Linköping University, Faculty of Health Sciences.
    Messmer, Davorka
    University of California.
    Emilsson, Johan
    Linköping University, Department of Clinical and Experimental Medicine. Linköping University, Faculty of Health Sciences.
    Falkmer, Ursula
    Jonköping Hospital.
    Falkmer, Sture
    Jonköping Hospital.
    Magnusson, Karl-Eric
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Borch, Kurt
    Linköping University, Department of Clinical and Experimental Medicine, Surgery . Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre of Surgery and Oncology, Department of Surgery in Östergötland.
    Larsson, Marie
    Linköping University, Department of Clinical and Experimental Medicine, Molecular Virology . Linköping University, Faculty of Health Sciences.
    Pancreatic adenocarcinoma exerts systemic effects on the peripheral blood myeloid and plasmacytoid dendritic cells: an indicator of disease severity?2010In: BMC CANCER, ISSN 1471-2407, Vol. 10, no 87Article in journal (Refereed)
    Abstract [en]

    Background: Dendritic cells (DCs) isolated from tumor bearing animals or from individuals with solid tumors display functional abnormalities and the DC impairment has emerged as one mechanism for tumor evasion from the control of the immune system. Ductal pancreatic adenocarcinoma (PDAC), the most common pancreatic cancer, is recognized as a very aggressive cancer type with a mortality that almost matches the rate of incidence. Methods: We examined the systemic influence ductal pancreatic adenocarcinoma ( PDAC) exerted on levels of peripheral blood DCs and inflammatory mediators in comparison to the effects exerted by other pancreatic tumors, chronic pancreatitis, and age-matched controls. Results: All groups examined, including PDAC, had decreased levels of myeloid DCs (MDC) and plasmacytoid DCs (PDC) and enhanced apoptosis in these cells as compared to controls. We found elevated levels of PGE2 and CXCL8 in subjects with PDAC, and chronic pancreatitis. Levels of these inflammatory factors were in part restored in PDAC after tumor resection, whereas the levels of DCs were impaired in the majority of these patients similar to 12 weeks after tumor removal. Our results prove that solid pancreatic tumors, including PDAC, systemically affect blood DCs. The impairments do not seem to be tumor-specific, since similar results were obtained in subjects with chronic pancreatitis. Furthermore, we found that PDAC patients with a survival over 2 years had significant higher levels of blood DCs compared to patients with less than one year survival. Conclusions: Our findings points to the involvement of inflammation in the destruction of the blood MDCs and PDCs. Furthermore, the preservation of the blood DCs compartment in PDAC patients seems to benefit their ability to control the disease and survival.

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  • 211.
    Tunaitis, Virginijus
    et al.
    State Research Institute Centre Innovat Med, Lithuania .
    Borutinskaite, Veronika
    Institute Biochem, Lithuania .
    Navakauskiene, Ruta
    Institute Biochem, Lithuania .
    Treigyte, Grazina
    Institute Biochem, Lithuania .
    Unguryte, Ausra
    State Research Institute Centre Innovat Med, Lithuania .
    Aldonyte, Ruta
    State Research Institute Centre Innovat Med, Lithuania.
    Magnusson, Karl-Eric
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Pivoriunas, Augustas
    State Research Institute Centre Innovat Med, Lithuania.
    Effects of different sera on adipose tissue-derived mesenchymal stromal cells2011In: JOURNAL OF TISSUE ENGINEERING AND REGENERATIVE MEDICINE, ISSN 1932-6254, Vol. 5, no 9, p. 733-746Article in journal (Refereed)
    Abstract [en]

    Current cell therapy protocols require considerable numbers of mesenchymal stromal cells (MSCs), which can be obtained only by in vitro expansion. The most important issue is a choice of optimal growth supplements for cell culture. Ideally, these should be of known composition, free of animal components and allow production of large homogenic populations of MSCs in a considerably short period of time. Since this standard has not been achieved to date, we aimed to assess the molecular responses of MSCs to different growth supplements commonly in use. MSCs were isolated from breast or abdominal adipose tissue and plated into DMEM supplemented with one of four different sera: fetal calf serum (FCS), pretested fetal calf serum (FCS-Sp), human allogeneic serum (HS) or artificial serum substitute (AS). MSCs cultivated with different serum supplements demonstrated distinct morphologies, high adipogenic and osteogenic differentiation potential and expressed characteristic antigens. Using real-time PCR, we found a large increase in PPAR gamma and Msx2 gene expression in both lines of proliferating MSCs cultivated with AS. We found that MSCs cultivated in the presence of different sera had similar global proteomic expression patterns, but comparisons of identified proteins revealed most differences in the MSCs cultivated with AS. Our results indicate that MSCs cultivated in the presence of FCS and HS display similar growth, differentiation, immunophenotypic and proteomic properties, while AS induces more profound changes in the physiology of MSCs, suggesting that further fundamental studies should be done before its introduction into clinical practice.

  • 212.
    Turina, Dean
    et al.
    Linköping University, Department of Medical and Health Sciences, Anesthesiology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Sinnescentrum, Department of Anaesthesiology and Surgery UHL.
    Björnström-Karlsson, Karin
    Linköping University, Department of Medical and Health Sciences, Anesthesiology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Sinnescentrum, Department of Anaesthesiology and Surgery UHL.
    Sundqvist, Tommy
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Eintrei, Christina
    Linköping University, Department of Medical and Health Sciences, Anesthesiology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Sinnescentrum, Department of Intensive Care UHL.
    Propofol alters vesicular transport in rat cortical neuronalcultures2011In: Journal of Physiology and Pharmacology, ISSN 0867-5910, E-ISSN 1899-1505, Vol. 62, no 1, p. 119-124Article in journal (Refereed)
    Abstract [en]

    Neuronal intracellular transport is performed by motor proteins, which deliver vesicles, organelles and proteins along cytoskeletal tracks inside the neuron. We have previously shown that the anesthetic propofol causes dose- and time-dependent, reversible retraction of neuronal neurites. We hypothesize that propofol alters the vesicular transport of cortical neurons due to this neurite retraction. Primary cultures of co-cultivated rat cortical neurons and glial cells were exposed to either 2 mu M propofol, control medium or the lipid vehicle, in time-response experiments. Reversibility was tested by washing propofol off the cells. The role of the GABA(A) receptor (GABA(A)R) was assessed with the GABA(A)R antagonist gabazine. Vesicles were tracked using differential interference contrast video microscopy. Propofol caused a retrograde movement in 83.4 +/- 5.2% (mean +/- S.E.M.) of vesicles, which accelerated over the observed time course (0.025 +/- 0.012 mu m.s(-1)). In control medium, vesicles moved predominantly anterograde (84.6 +/- 11.1%) with lower velocity (0.011 +/- 0.004 mu m.s(-1)). Cells exposed to the lipid vehicle showed the same dynamic characteristics as cells in control medium. The propofol-induced effect on vesicle transport was reversible and blocked by the GABA(A)R antagonist gabazine in low concentration. Our results show that propofol causes a reversible, accelerating vesicle movement toward the neuronal cell body that is mediated via synaptic GABA(A)R. We have previously reported that propofol initiates neurite retraction, and we propose that propofol causes vesicle movement by retrograde flow of cytoplasm from the narrowed neurite.

  • 213.
    Turina, Dean
    et al.
    Linköping University, Department of Medical and Health Sciences, Anesthesiology. Linköping University, Faculty of Health Sciences.
    Karin, Björnström Karlsson
    Linköping University, Department of Medical and Health Sciences, Anesthesiology. Linköping University, Faculty of Health Sciences.
    Sundqvist, Tommy
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Eintrei, Christina
    Linköping University, Department of Medical and Health Sciences, Anesthesiology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Sinnescentrum, Department of Intensive Care UHL.
    Orexin A inhibits propofol-induced neurite retraction by a PLD-dependent mechanism in neuronsManuscript (preprint) (Other academic)
    Abstract [en]

    Background: Propofol retracts neurites and reverses the transport of vesicles in rat cortical neurons in a γ-aminobutyric acid type A (GABAA) receptor dependent manner. Orexin A (OA) is an endogenous peptide regulating wakefulness, and is known to interact with anaesthetics. We aim to investigate whether OA inhibits propofol-induced neurite retraction and elucidate the intracellular signalling involved.

    Methods: In primary cortical cell cultures from newborn rat brains, live cell light microscopy was used to measure neurite retraction after propofol (2 μM) with or without OA (10 nM) application after preincubation with the Rhokinase inhibitor (HA-1077), phospholipase D (PLD) inhibitor [5-fluoro-2- indolyl des-chlorohalopemide (FIPI)], protein kinase C (PKC) inhibitor (staurosporine) or PKC activator phorbol 12-myristate 13-acetate (PMA).

    Results: The neurite retraction induced by propofol is blocked by HA-1077 and PMA. OA blocks neurite retraction induced by propofol, and this inhibitory effect could be prevented by FIPI, as well as staurosporine.

    Conclusions: Rho-kinase is essential for propofol-induced neurite retraction in cortical neuronal cells. Activation of PKC plays an inhibitive role during neurite retraction caused by propofol. OA blocks propofol-induced neurite retraction by a PLD/PKC-mediated pathway.

  • 214.
    Turina, Dean
    et al.
    Linköping University, Department of Medical and Health Sciences, Anesthesiology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Sinnescentrum, Department of Anaesthesiology and Surgery UHL.
    Loitto, Vesa
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Björnström, Karin
    Linköping University, Department of Medical and Health Sciences, Anesthesiology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Sinnescentrum, Department of Anaesthesiology and Surgery UHL.
    Sundqvist, Tommy
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Eintrei, Christina
    Linköping University, Department of Medical and Health Sciences, Anesthesiology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Sinnescentrum, Department of Anaesthesiology and Surgery UHL.
    Propofol causes neurite retraction in neurons2008Conference paper (Refereed)
  • 215.
    Turina, Dean
    et al.
    Linköping University, Department of Medical and Health Sciences, Anesthesiology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Sinnescentrum, Department of Anaesthesiology and Surgery UHL.
    Loitto, Vesa
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Björnström, Karin
    Linköping University, Department of Medical and Health Sciences, Anesthesiology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Sinnescentrum, Department of Anaesthesiology and Surgery UHL.
    Sundqvist, Tommy
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Eintrei, Christina
    Linköping University, Department of Medical and Health Sciences, Anesthesiology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Sinnescentrum, Department of Anaesthesiology and Surgery UHL.
    Propofol causes neurite retraction in neurons2008In: British Journal of Anaesthesia, ISSN 0007-0912, E-ISSN 1471-6771, Vol. 101, no 3, p. 374-379Article in journal (Refereed)
    Abstract [en]

    Background The mechanism by which anaesthetic agents produce general anaesthesia is not yet fully understood. Retraction of neurites is an important function of individual neurones and neural plexuses during normal and pathological conditions, and it has been shown that such a retraction pathway exists in developing and mature neurones. We hypothesized that propofol decreases neuronal activity by causing retraction of neuronal neurites.

    Methods Primary cultures of rat cortical neurones were exposed in concentration– and time–response experiments to 0.02, 0.2, 2, and 20 µM propofol or lipid vehicle. Neurones were pretreated with the GABAA receptor (GABAAR) antagonist, bicuculline, the myosin II ATPase activity inhibitor, blebbistatin, and the F-actin stabilizing agent, phalloidin, followed by administration of propofol (20 µM). Changes in neurite retraction were evaluated using time-lapse light microscopy.

    Results Propofol caused a concentration- and time-dependent reversible retraction of cultured cortical neurone neurites. Bicuculline, blebbistatin, and phalloidin completely inhibited propofol-induced neurite retraction. Images of retracted neurites were characterized by a retraction bulb and a thin trailing membrane remnant.

    Conclusions Cultured cortical rat neurones retract their neurites after exposure to propofol in a concentration- and time-dependent manner. This retraction is GABAAR mediated, reversible, and dependent on actin and myosin II. Furthermore, the concentrations and times to full retraction and recovery correspond to those observed during propofol anaesthesia.

  • 216.
    Turoverova, L.V.
    et al.
    Institute of Cytology RAS, St. Petersburg, Russian Federation.
    Khotin, M.G.
    Institute of Cytology RAS, St. Petersburg, Russian Federation.
    Yudintseva, N.M.
    Institute of Cytology RAS, St. Petersburg, Russian Federation.
    Magnusson, Karl-Eric
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Blinova, M.I.
    Institute of Cytology RAS, St. Petersburg, Russian Federation.
    Pinaev, G.P.
    Institute of Cytology RAS, St. Petersburg, Russian Federation.
    Tentler, D.G.
    Institute of Cytology RAS, St. Petersburg, Russian Federation.
    Analysis of extracellular matrix proteins produced by cultured cells2009In: Cell and Tissue Biology, ISSN 1990-519X, Vol. 3, no 5, p. 497-502Article in journal (Refereed)
    Abstract [en]

    The extracellular matrix (ECM) is a highly organized multimolecular structure essential for the vital functions of any organism. Although much of the data of extracellular matrix components has been accumulated, the isolation of an entire set of these proteins remains a complex procedure due to the high content of fibrillar proteins and proteoglycans, which form multidomain, netlike structures. In the study presented, we developed a method for isolating ECM proteins from cell cultures. Human epidermoid carcinoma cells A431 and fibroblasts obtained from normal and scar human skin were used. We showed that EDTA solution removed cells from culture plates without destroying the cell membranes. Subsequent treatment of remaining ECM proteins with acetic acid in order to dissociate collagen fibers significantly improved the fractioning of ECM proteins. The extraction of remaining proteins from the surface of the culture plate was preformed by a buffer developed based on Laemmli probe buffer. Using this method, we isolated ECM proteins synthesized by cultured cells, and the extracted proteins were suitable for future analysis by SDS PAGE and two-dimentional electrophoresis, as well as for identifying individual proteins by mass spectrometry. This study may allow us to compare assortments of ECM proteins isolated from different sources, and elucidate impact of various proteins on structure and property of extracellular matrix of investigated cells.

  • 217.
    Turoverova, L.V.
    et al.
    Institute of Cytology RAS, St. Petersburg, Russian Federation.
    Khotin, M.G.
    Institute of Cytology RAS, St. Petersburg, Russian Federation.
    Yudintseva, N.M.
    Institute of Cytology RAS, St. Petersburg, Russian Federation.
    Magnusson, Karl-Eric
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Blinova, M.I.
    Institute of Cytology RAS, St. Petersburg, Russian Federation.
    Pinaev, G.P.
    Institute of Cytology RAS, St. Petersburg, Russian Federation.
    Tentler, D.G.
    Institute of Cytology RAS, St. Petersburg, Russian Federation.
    Analysis of extracellular matrix proteins produced by cultured cells2009In: Tsitologiya, ISSN 0041-3771, Vol. 51, no 8, p. 691-696Article in journal (Refereed)
    Abstract [en]

    Extracellular matrix (ECM) is a highly organized multimolecular structure essential for vital function of any organism. Although a lot of data on the extracellular matrix components has been accumulated, an isolation of the entire set of these proteins still remains to be a complex procedure since ECM contains fibrillar proteins and proteoglycans, which form multidomain net-like structures. In the presented study, we developed a method for isolation of ECM proteins from cell cultures. Human epidermoid carcinoma cells A431 and fibroblasts obtained from normal and scar human skin were used. We showed that EDTA solution removed cells from culture plates without destroying the cell membrane. Following treatment of remaining ECM proteins with acetic acid in order to dissociate collagen fibrils significantly improved fractioning of ECM proteins. Extraction of the remained proteins from culture plate surface was preformed using a buffer developed on the basis of Laemmli probe buffer. With this method, we isolated ECM proteins synthesized by culturing cells and suitable for a future analysis by SDS PAGE and two-dimentional electrophoresis as well as for identification of individual proteins by mass-spectrometry. This study may allow comparing protein contents of ECMs isolated from different sources, and elucidate influences of various proteins on the protein and the properties of extracellular matrix of investigated cells.

  • 218.
    Verma, Deepti
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Cell Biology. Linköping University, Faculty of Health Sciences.
    Eriksson, Per
    Linköping University, Department of Clinical and Experimental Medicine, Rheumatology. Linköping University, Faculty of Health Sciences.
    Sahdo, Berolla
    Örebro universitet, Sweden.
    Persson, Alexander
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Ejdebäck, Mikael
    Skövde universitet, Sweden.
    Särndahl, Eva
    Örebro universitet.
    Söderkvist, Peter
    Linköping University, Department of Clinical and Experimental Medicine, Cell Biology. Linköping University, Faculty of Health Sciences.
    Two adult siblings with atypical cryopyrin-associated periodic syndrome due to a novel M299V mutation in NLRP32010In: Arthritis and Rheumatism, ISSN 0004-3591, E-ISSN 1529-0131, Vol. 62, no 7, p. 2138-2143Article in journal (Refereed)
    Abstract [en]

    OBJECTIVE: The NALP3 inflammasome is a multiprotein complex that triggers caspase 1-mediated interleukin-1beta (IL-1beta) release. Mutations in the gene encoding NALP3 (NLRP3) underlie the cryopyrin-associated periodic syndrome (CAPS). The aim of this study was to report a novel NLRP3 mutation in 2 siblings of Swedish descent in whom symptoms first presented in adulthood.

    METHODS: Mutation analysis of NLRP3 was performed on DNA from patients with CAPS and 100 control subjects. For assessment of caspase 1 and IL-1beta, blood was collected from patients and age- and sex-matched healthy control subjects. Genetic constructs containing mutant or wild-type NLRP3 were transduced into THP-1 cells, followed by assessment of IL-1beta levels in cell supernatant.

    RESULTS: Both siblings carried a novel M299V mutation in NLRP3, which was not present in the control population. The samples obtained from the patients displayed increased caspase 1 activity and elevated IL-1beta levels at basal conditions as compared with healthy control subjects. THP-1 cells expressing mutated M299V revealed almost 10-fold higher IL-1beta production compared with the wild-type construct.

    CONCLUSION: M299V is an activating mutation in NLRP3 resulting in elevated spontaneous caspase 1 activity and IL-1beta levels. The classic CAPS phenotype was lacking in these adult siblings. Whereas one sibling displayed a milder phenotype that has so far responded satisfactorily to oral nonsteroidal antiinflammatory drugs in combination with low-dose corticosteroids, the inflammatory symptoms in the sibling with the more severe case responded well to IL-1beta blockade. Understanding the pathogenic mechanism underlying such disorders can be helpful for the physician. Our study reinforces the importance of genetic testing and laboratory investigations in combination with careful phenotypic evaluation for the diagnosis of such patients.

  • 219.
    Verma, Deepti
    et al.
    Linköping University, Department of Clinical and Experimental Medicine. Linköping University, Faculty of Health Sciences.
    Lerm, Maria
    Linköping University, Department of Clinical and Experimental Medicine. Linköping University, Faculty of Health Sciences.
    Blomgran Julinder, Robert
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Eriksson, Per
    Linköping University, Department of Clinical and Experimental Medicine, Rheumatology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre for Medicine, Department of Rheumatology in Östergötland.
    Söderkvist, Peter
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Cell Biology.
    Särndahl, Eva
    Örebro Universitet, Örebro.
    Gene polymorphisms in the NALP3 inflammasome are associated with interleukin-1 production and severe inflammation: Relation to Common Inflammatory Diseases?2008In: Arthritis and Rheumatism, ISSN 0004-3591, E-ISSN 1529-0131, Vol. 58, no 3, p. 888-894Article in journal (Refereed)
    Abstract [en]

    Objective: NALP3, ASC, and TUCAN are components of the NALP3 inflammasome, which triggers caspase 1-mediated interleukin-1β (IL-1β) release. Activating mutations in the gene encoding NALP3 (NLRP3) have recently been linked to familial periodic fever syndromes. We undertook this study to determine whether a patient with arthritis and antibiotic-resistant fever carried mutations in the genes encoding the NALP3 inflammasome.

    Methods: Genetic analysis of NLRP3 and the gene encoding TUCAN (CARD-8) was performed on genomic DNA from the patient and from a population-based collection of DNA (806 subjects). For in vitro studies of IL-1β production and caspase 1 activity, blood was obtained from the patient at different time points after administration of anakinra, an IL-1 receptor antagonist, as well as from 5 healthy age- and sex-matched control subjects.

    Results: Mutation analysis of the patient's genes encoding NALP3, ASC, and TUCAN revealed variations in the NLRP3 (Q705K) and CARD-8 (C10X) genes. The allele frequencies of these single-nucleotide polymorphisms (SNPs) in the population were 6.5% and 34%, respectively. The elevated activity of caspase 1 and the high levels of IL-1β measured in samples from the patient returned to normal levels after treatment with anakinra.

    Conclusion: Our results indicate that the patient's symptoms were due to elevated levels of IL-1β, since treatment with anakinra effectively abolished the symptoms. The compound SNPs may explain the increased IL-1β levels and inflammatory symptoms observed, but further studies are needed to reveal a functional relationship. The prevalence of the polymorphisms (4% of the population carry both SNPs) in the general population may suggest a genetic predisposition for common inflammatory disorders.

  • 220.
    Vikström, Elena
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Bui, Lan
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Konradsson, Peter
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    Magnusson, Karl-Eric
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Role of calcium signalling and phosphorylations in disruption of the epithelial junctions by Pseudomonas aeruginosa quorum sensing molecule2010In: European Journal of Cell Biology, ISSN 0171-9335, E-ISSN 1618-1298, Vol. 89, no 8, p. 584-597Article in journal (Refereed)
    Abstract [en]

    In Pseudomonas aeruginosa. cell-cell communication based on acyl-homoserine lactone (HSL) quorum sensing molecules is known to coordinate the production of virulence factors and biofilms by the bacterium. Incidentally, these bacterial signals can also modulate mammalian cell behaviour. We demonstrate here that 3O-C-12-HSL can induce changes in calcium signalling through influx and release of calcium from thapsigargin-sensitive stores and delocalization of inositol 1,4,5-trisphosphate receptors (IP3R), but not of ryanodine receptors (RyR). In parallel, P. aeruginosa 3O-C-12-HSL disrupts junctions in human Caco-2 cells as evidenced by a reduction of the expression and distribution of ZO-3 and JAM-A. Using co-immunoprecipitation we also found an alteration in the binding of ZO-3 to JAM-A in protein complexes. Moreover, 3O-C-12-HSL-treatment resulted in tyrosine hyperphosphorylation of ZO-3 and JAM-A. On the contrary, serine and threonine residues of ZO-1 and JAM-A became less phosphorylated after exposition of 3O-C-12-HSL. The 3O-C-12-HSL-induced intracellular calcium signalling and alteration in the phosphorylation status of junction proteins furthermore correlated with changes in the association between JAM-A-ZO-3. The calcium inhibitors thapsigargin, xestospongin C. and dantrolene partly prevented the 3O-C-12-HSL-induced decreases in TER and increases in the paracellular flux of 10 kDa dextran. These findings clearly suggest that P. aeruginosa 3O-C-12-HSL can cause the loss of epithelial barrier function via calcium signalling and further alteration in the phosphorylation status of junction proteins; and that bacterial quorum sensing signals represent inter-kingdom signalling.

  • 221.
    Vikström, Elena
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Bui, Lan
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Konradsson, Peter
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Magnusson, Karl-Eric
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    The junctional integrity of epithelial cells is modulated by Pseudomonas aeruginosa quorum sensing molecule through phosphorylation-dependent mechanisms2009In: Experimental Cell Research, ISSN 0014-4827, E-ISSN 1090-2422, Vol. 315, no 2, p. 313-326Article in journal (Refereed)
    Abstract [en]

    In Pseudomonas aeruginosa, cell-cell Communication based on acyl-homoserine lactone (HSL) quorum sensing molecules is known to coordinate the production of virulence factors and biofilms by the bacterium. Incidentally, these bacterial signals can also modulate mammalian cell behaviour. We report that 3O-C-12-HSL can disrupt adherens junctions in human epithelial Caco-2 cells as evidenced by a reduction of the expression and distribution of E-cadherin and beta-catenin. Using co-immunoprecipitation we also found that P. aeruginosa 3O-C-12-HSL-treatment resulted in tyrosine hyperphosphorylation of E-cadherin, beta-catenin, occludin and ZO-1. Similarly, serine and threonine residues of E-cadherin and ZO-1 became more phosphorylated after 3O-C-12-HSL treatment. On the contrary, occludin and beta-catenin underwent dephosphorylation on serine and threonine residues after exposition of 3O-C-12-HSL. These changes in the phosphorylation state were paralleled by alteration in the Structure of junction complexes and increased paracellular permeability. Moreover, pre-treatment of the Caco-2 cells with protein phosphatase and kinase inhibitors prevented 3O-C-12-HSL-induced changes in paracellular permeability and interactions between occludin-ZO-1 and the E-cadherin-beta-catenin. These findings clearly suggest that an alteration in the phosphorylation status of junction proteins are involved in the changes in cell junction associations and enhanced paracellular permeability, and that bacterial signals are indeed sensed by the host cells.

  • 222.
    Viljanen, Johan
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    Larsson, Jenny
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Larsson (Kaiser), Andréas
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Broo, Kerstin S.
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    A Multipurpose Receptor Composed of Promiscuous Proteins. Analyte Detection through Pattern Recognition2007In: Bioconjugate Chemistry, ISSN 1043-1802, Vol. 18, no 6, p. 1935-1945Article in journal (Refereed)
    Abstract [en]

    A multipurpose receptor akin to the “electronic nose” was composed of coumarin-labeled mutants of human glutathione transferase A1. We have previously constructed a kit for site-specific modification of a lysine residue (A216K) using a thiol ester of glutathione (GSC-Coubio) as a modifying reagent. In the present investigation, we scrambled the hydrophobic binding site (H-site) of the protein scaffold through mutations at position M208 via random mutagenesis and isolated a representative library of 11 A216K/M208X mutants. All of the double mutants could be site-specifically labeled to form the K216Cou conjugates. The labeled proteins responded to the addition of different analytes with signature changes in their fluorescence spectra resulting in a matrix of 96 data points per analyte. Ligands as diverse as n-valeric acid, fumaric acid monoethyl ester, lithocholic acid, 1-chloro-2,4-dinitrobenzene (CDNB), glutathione (GSH), S-methyl-GSH, S-hexyl-GSH, and GS-DNB all gave rise to signals that potentially can be interpreted through pattern recognition. The measured Kd values range from low micromolar to low millimolar. The cysteine residue C112 was used to anchor the coumarin-labeled protein to a PEG-based hydrogel chip in order to develop surface-based biosensing systems. We have thus initiated the development of a multipurpose, artificial receptor composed of an array of promiscuous proteins where detection of the analyte occurs through pattern recognition of fluorescence signals. In this system, many relatively poor binders each contribute to detailed readout in a truly egalitarian fashion.

  • 223. Order onlineBuy this publication >>
    Welin, Amanda
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Survival strategies of Mycobacterium tuberculosis inside the human macrophage2011Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Mycobacterium tuberculosis (Mtb) is the bacterium responsible for tuberculosis (TB). For decades, it was believed that TB was a disease of the past, but the onset of the HIV epidemic resulting in a greatly increased number of TB cases, the emergence of antibiotic resistant Mtb strains, and the relative ineffectiveness of the BCG vaccine have put TB back on the agenda. With almost two million people being killed by TB each year, the World Health Organization has declared it a global emergency. TB is an especially big issue in low-income countries, where crowded living conditions accelerates spread of the disease, and where access to health care and medication is problematic. Mtb spreads by aerosol and infects its host through the airways. The bacterium is phagocytosed by resident macrophages in the lung, and when successful is able to replicate inside these cells, which are actually designed to kill invading microbes. Mtb is able to evade macrophage responses in part by inhibiting the fusion between the phagosome in which it resides and bactericidal lysosomes, as well as by dampening the acidification of the vacuole. The initial macrophage infection results in a pro-inflammatory response and the recruitment of other cells of the innate and adaptive immune systems, giving rise to the hallmark of Mtb infection – the granuloma. It is believed that in up to 50 % of exposed individuals, however, the infection is cleared without the involvement of the adaptive immune system, indicating that the innate immune system may be able to control or clear the infection if activated appropriately. This thesis focuses on the interaction between the host macrophage and Mtb, aiming to understand some of the mechanisms employed by the bacterium to evade macrophage responses to enable replication and spread to new host cells. Furthermore, mechanisms used by the macrophage to keep the infection under control were studied, and a method that could be used to measure the replication of the bacilli inside macrophages in vitro in an efficient way was developed. We found that a mycobacterial glycoprotein, mannose-capped lipoarabinomannan (ManLAM), which is shed from the bacilli during phagocytosis by macrophages, integrates into membrane raft domains of the host cell membrane via its GPI anchor. This integration leads to an inhibition of phagosomal maturation. Subsequently, we developed a luciferase-based method by which intracellular replication of Mtb as well as viability of the host macrophage could be measured in a rapid, inexpensive and quantitative way in a 96-well plate. This method could be used for drug screening as well as for studying the different host and bacterial factors that influence the growth of Mtb inside the host cell. Using this method, we discovered that infection of macrophages with Mtb at a low multiplicity of infection (MOI) led to effective control of bacterial growth by the cell, and that this was dependent on functional lysosomal proteases as well as phagosomal acidification. However, we found no correlation between controlled bacterial growth and the translocation of late endosomal membrane proteins to the phagosome, showing that these markers are poor indicators of phagosomal functionality. Furthermore, we discovered that infection of macrophages with Mtb at a higher MOI led to replication of the bacilli accompanied by host cell death within a few days. We characterized this cell death, and concluded that when replication of Mtb inside macrophages reaches a certain threshold and the bacteria secrete a protein termed ESAT-6, necrotic cell death of the host cell occurs. However, although the bacilli activated inflammasome complexes in the host cell and IL-1β was secreted during infection of macrophages, Mtb infection did not induce either of the recently characterized inflammasome-related cell death types pyroptosis or pyronecrosis. Thus, we have elucidated some of the strategies that Mtb uses to be able to survive and replicate inside the macrophage and spread to new cells, as well as studied the conditions under which the host cell is able to control infection. This knowledge could be used in the future for developing drugs that boost the innate immune system or targets bacterial virulence factors in the macrophage.

    List of papers
    1. Incorporation of Mycobacterium tuberculosis lipoarabinomannan into macrophage membrane rafts is a prerequisite for the phagosomal maturation block.
    Open this publication in new window or tab >>Incorporation of Mycobacterium tuberculosis lipoarabinomannan into macrophage membrane rafts is a prerequisite for the phagosomal maturation block.
    Show others...
    2008 (English)In: Infection and Immunity, ISSN 0019-9567, E-ISSN 1098-5522, Vol. 76, no 7, p. 2882-2887Article in journal (Refereed) Published
    Abstract [en]

    Lipoarabinomannan (LAM) is one of the key virulence factors for Mycobacterium tuberculosis, the etiological agent of tuberculosis. During uptake of mycobacteria, LAM interacts with the cell membrane of the host macrophage and can be detected throughout the cell upon infection. LAM can inhibit phagosomal maturation as well as induce a proinflammatory response in bystander cells. The aim of this study was to investigate how LAM exerts its action on human macrophages. We show that LAM is incorporated into membrane rafts of the macrophage cell membrane via its glycosylphosphatidylinositol anchor and that incorporation of mannose-capped LAM from M. tuberculosis results in reduced phagosomal maturation. This is dependent on successful insertion of the glycosylphosphatidylinositol anchor. LAM does not, however, induce the phagosomal maturation block through activation of p38 mitogen-activated protein kinase, contradicting some previous suggestions.

    National Category
    Microbiology in the medical area
    Identifiers
    urn:nbn:se:liu:diva-20816 (URN)10.1128/IAI.01549-07 (DOI)18426888 (PubMedID)
    Available from: 2009-09-22 Created: 2009-09-22 Last updated: 2018-01-13Bibliographically approved
    2. Validation of a Medium-Throughput Method for Evaluation of Intracellular Growth of Mycobacterium tuberculosis
    Open this publication in new window or tab >>Validation of a Medium-Throughput Method for Evaluation of Intracellular Growth of Mycobacterium tuberculosis
    Show others...
    2010 (English)In: Clinical and Vaccine Immunology, ISSN 1556-6811, E-ISSN 1556-679X, Vol. 17, no 4, p. 513-517Article in journal (Refereed) Published
    Abstract [en]

    Intracellular pathogens such as Mycobacterium tuberculosis have adapted to a life inside host cells, in which they utilize host nutrients to replicate and spread. Ineffective methods for the evaluation of growth of intracellular pathogens in their true environment pose an obstacle for basic research and drug screening. Here we present the validation of a luminometry-based method for the analysis of intramacrophage growth of M. tuberculosis. The method, which is performed in a medium-throughput format, can easily be adapted for studies of other intracellular pathogens and cell types. The use of host cells in drug-screening assays dedicated to find antimicrobials effective against intracellular pathogens permits the discovery of not only novel antibiotics but also compounds with immunomodulatory and virulence-impairing activities, which may be future alternatives or complements to antibiotics.

    Place, publisher, year, edition, pages
    American Society for Microbiology, 2010
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:liu:diva-54868 (URN)10.1128/CVI.00446-09 (DOI)000276170900004 ()
    Available from: 2010-04-16 Created: 2010-04-16 Last updated: 2017-10-31
    3. Importance of phagosomal functionality for growth restriction of Mycobacterium tuberculosis in primary human macrophages
    Open this publication in new window or tab >>Importance of phagosomal functionality for growth restriction of Mycobacterium tuberculosis in primary human macrophages
    Show others...
    2011 (English)In: Journal of Innate Immunity, ISSN 1662-811X, E-ISSN 1662-8128, Vol. 3, no 5, p. 508-518Article in journal (Refereed) Published
    Abstract [en]

    The best characterized survival mechanism of Mycobacterium tuberculosis inside the macrophage is the inhibition of phagosomal maturation. Phagosomal maturation involves several steps including fusion with lysosomes and acidification. However, it has not been elucidated which components of phagosomal maturation correlate with growth restriction of virulent mycobacteria in human macrophages, and we aimed to study this. We infected human monocyte-derived macrophages with M. tuberculosis and assessed bacterial replication, translocation of CD63 to the phagosome, and phagosomal acidification. We found that unstimulated macrophages were able to control infection with M. tuberculosis upon inoculation at a low, but not high, multiplicity of infection (MOI). H37Rv and H37Ra infection, at both high and low MOI, led to equally ineffective translocation of CD63 to the phagosome. This was true despite the impaired growth ability of H37Rv at the low MOI and of H37Ra even at the high MOI, indicating that inhibition of CD63 translocation was not sufficient for growth to occur. On the other hand, acidification of mycobacterial phagosomes was more efficient at a low MOI with both mycobacterial strains, consistent with a role for phagosomal acidification in restricting M. tuberculosis growth. Inhibition of the vacuolar H+-ATPase as well as of cathepsin D led to enhanced mycobacterial replication inside the macrophage. We conclude that acidification and related functional aspects of the mature phagosome are important factors for restriction of M. tuberculosis replication in human macrophages.

    Place, publisher, year, edition, pages
    S. Karger, 2011
    National Category
    Basic Medicine
    Identifiers
    urn:nbn:se:liu:diva-65447 (URN)10.1159/000325297 (DOI)000294572500008 ()21576918 (PubMedID)
    Note

    Funding Agencies|Swedish Research Council|529-2003-5994,2005-7046,2006-5968,2007-2673,2009-3821|Bill and Melinda Gates Foundation||SIDA/SAREC||Ekhaga Foundation||Carl Trygger Foundation||King Gustaf V 80-Year Memorial Foundation||County Council of Ostergotland||Swedish Heart Lung Foundation||Oskar II Jubilee Foundation||Clas Groschinsky Foundation||Soderbergs Foundation||Colorado State University, Fort Collins (NIH, NIAID)|HHSN26620040 0091C|

    Available from: 2011-02-08 Created: 2011-02-08 Last updated: 2018-01-12Bibliographically approved
    4. Human macrophages infected with virulent Mycobacterium tuberculosis undergo ESAT-6-dependent necrosis, but not pyroptosis or pyronecrosis
    Open this publication in new window or tab >>Human macrophages infected with virulent Mycobacterium tuberculosis undergo ESAT-6-dependent necrosis, but not pyroptosis or pyronecrosis
    (English)Manuscript (preprint) (Other academic)
    Abstract [en]

    Mycobacterium tuberculosis (Mtb) infects lung macrophages, which instead of killing the pathogen can be manipulated by the bacilli, creating an environment suitable for intracellular replication and spread to adjacent cells. The role of host cell death during Mtb infection is debated because the bacilli have been shown to be both anti-apoptotic, keeping the host cell alive to avoid the antimicrobial effects of apoptosis, and pro-necrotic, killing the host macrophage to allow infection of neighboring cells. Since mycobacteria are able to activate the NLRP3 inflammasome, we investigated whether Mtb could induce one of the recently described inflammasome-linked cell death modes pyroptosis and pyronecrosis, in human monocyte-derived macrophages. Cells were infected with virulent (H37Rv) Mtb at a multiplicity of infection (MOI) of 1 or 10. The higher MOI resulted in strongly enhanced release of IL-1β, while a low MOI gave no IL-1β response. The infected macrophages were collected and cell viability in terms of the integrity of DNA, mitochondria and the plasma membrane was determined. We found that infection with H37Rv at MOI 10, but not MOI 1, over two days led to extensive DNA fragmentation, loss of mitochondrial membrane potential and loss of plasma membrane integrity. Although we observed plasma membrane permeabilization and IL-1 β release from infected cells, the cell death induced by Mtb was not pyroptosis or pyronecrosis, as it was independent of caspase-1 and cathepsin B. Instead, we conclude that as virulent Mtb reaches a threshold number of bacilli inside the macrophage, ESAT-6-dependent necrosis occurs, activating caspase-1 in the process.

    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:liu:diva-65451 (URN)
    Note
    Submitted manuscriptAvailable from: 2011-02-08 Created: 2011-02-08 Last updated: 2011-02-22Bibliographically approved
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    Survival strategies of Mycobacterium tuberculosis inside the human macrophage
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  • 224.
    Welin, Amanda
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Eklund, Daniel
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology.
    Stendahl, Olle
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Lerm, Maria
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Human Macrophages Infected with a High Burden of ESAT-6-Expressing M. tuberculosis Undergo Caspase-1-and Cathepsin B-Independent Necrosis2011In: PLOS ONE, ISSN 1932-6203, Vol. 6, no 5Article in journal (Refereed)
    Abstract [en]

    Mycobacterium tuberculosis (Mtb) infects lung macrophages, which instead of killing the pathogen can be manipulated by the bacilli, creating an environment suitable for intracellular replication and spread to adjacent cells. The role of host cell death during Mtb infection is debated because the bacilli have been shown to be both anti-apoptotic, keeping the host cell alive to avoid the antimicrobial effects of apoptosis, and pro-necrotic, killing the host macrophage to allow infection of neighboring cells. Since mycobacteria activate the NLRP3 inflammasome in macrophages, we investigated whether Mtb could induce one of the recently described inflammasome-linked cell death modes pyroptosis and pyronecrosis. These are mediated through caspase-1 and cathepsin-B, respectively. Human monocyte-derived macrophages were infected with virulent (H37Rv) Mtb at a multiplicity of infection (MOI) of 1 or 10. The higher MOI resulted in strongly enhanced release of IL-1 beta, while a low MOI gave no IL-1 beta response. The infected macrophages were collected and cell viability in terms of the integrity of DNA, mitochondria and the plasma membrane was determined. We found that infection with H37Rv at MOI 10, but not MOI 1, over two days led to extensive DNA fragmentation, loss of mitochondrial membrane potential, loss of plasma membrane integrity, and HMGB1 release. Although we observed plasma membrane permeabilization and IL-1 beta release from infected cells, the cell death induced by Mtb was not dependent on caspase-1 or cathepsin B. It was, however, dependent on mycobacterial expression of ESAT-6. We conclude that as virulent Mtb reaches a threshold number of bacilli inside the human macrophage, ESAT-6-dependent necrosis occurs, activating caspase-1 in the process.

    Download full text (pdf)
    fulltext
  • 225.
    Welin, Amanda
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Eklund, Daniel
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Stendahl, Olle
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Lerm, Maria
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Human macrophages infected with virulent Mycobacterium tuberculosis undergo ESAT-6-dependent necrosis, but not pyroptosis or pyronecrosisManuscript (preprint) (Other academic)
    Abstract [en]

    Mycobacterium tuberculosis (Mtb) infects lung macrophages, which instead of killing the pathogen can be manipulated by the bacilli, creating an environment suitable for intracellular replication and spread to adjacent cells. The role of host cell death during Mtb infection is debated because the bacilli have been shown to be both anti-apoptotic, keeping the host cell alive to avoid the antimicrobial effects of apoptosis, and pro-necrotic, killing the host macrophage to allow infection of neighboring cells. Since mycobacteria are able to activate the NLRP3 inflammasome, we investigated whether Mtb could induce one of the recently described inflammasome-linked cell death modes pyroptosis and pyronecrosis, in human monocyte-derived macrophages. Cells were infected with virulent (H37Rv) Mtb at a multiplicity of infection (MOI) of 1 or 10. The higher MOI resulted in strongly enhanced release of IL-1β, while a low MOI gave no IL-1β response. The infected macrophages were collected and cell viability in terms of the integrity of DNA, mitochondria and the plasma membrane was determined. We found that infection with H37Rv at MOI 10, but not MOI 1, over two days led to extensive DNA fragmentation, loss of mitochondrial membrane potential and loss of plasma membrane integrity. Although we observed plasma membrane permeabilization and IL-1 β release from infected cells, the cell death induced by Mtb was not pyroptosis or pyronecrosis, as it was independent of caspase-1 and cathepsin B. Instead, we conclude that as virulent Mtb reaches a threshold number of bacilli inside the macrophage, ESAT-6-dependent necrosis occurs, activating caspase-1 in the process.

  • 226.
    Welin, Amanda
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Lerm, Maria
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Inside or outside the phagosome? The controversy of the intracellular localization of Mycobacterium tuberculosis2012In: Tuberculosis, ISSN 1472-9792, E-ISSN 1873-281X, Vol. 92, no 2, p. 113-120Article, review/survey (Refereed)
    Abstract [en]

    The localization of Mycobacterium tuberculosis (Mtb) inside the macrophage has been a matter of debate in recent years. Upon inhalation, the bacterium is taken up into macrophage phagosomes, which are manipulated by the bacterium. Subsequent translocation of the bacilli into the cytosol has been observed by several groups, while others fail to observe this phenomenon. Here, we review the available literature in favour of and against this idea, and scrutinize the existing data on how human macrophages control Mtb infection, relating this to the robustness of the host cell. We conclude that both phagosomal maturation inhibition and escape from the phagosome are part of the greater infection strategy of Mtb. The balance between the host cell and the infecting bacterium is an important factor in determining the outcome of infection as well as whether phagosomal escape occurs and can be captured.

  • 227.
    Welin, Amanda
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Raffetseder, Johanna
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Eklund, Daniel
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Stendahl, Olle
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Lerm, Maria
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Importance of phagosomal functionality for growth restriction of Mycobacterium tuberculosis in primary human macrophages2011In: Journal of Innate Immunity, ISSN 1662-811X, E-ISSN 1662-8128, Vol. 3, no 5, p. 508-518Article in journal (Refereed)
    Abstract [en]

    The best characterized survival mechanism of Mycobacterium tuberculosis inside the macrophage is the inhibition of phagosomal maturation. Phagosomal maturation involves several steps including fusion with lysosomes and acidification. However, it has not been elucidated which components of phagosomal maturation correlate with growth restriction of virulent mycobacteria in human macrophages, and we aimed to study this. We infected human monocyte-derived macrophages with M. tuberculosis and assessed bacterial replication, translocation of CD63 to the phagosome, and phagosomal acidification. We found that unstimulated macrophages were able to control infection with M. tuberculosis upon inoculation at a low, but not high, multiplicity of infection (MOI). H37Rv and H37Ra infection, at both high and low MOI, led to equally ineffective translocation of CD63 to the phagosome. This was true despite the impaired growth ability of H37Rv at the low MOI and of H37Ra even at the high MOI, indicating that inhibition of CD63 translocation was not sufficient for growth to occur. On the other hand, acidification of mycobacterial phagosomes was more efficient at a low MOI with both mycobacterial strains, consistent with a role for phagosomal acidification in restricting M. tuberculosis growth. Inhibition of the vacuolar H+-ATPase as well as of cathepsin D led to enhanced mycobacterial replication inside the macrophage. We conclude that acidification and related functional aspects of the mature phagosome are important factors for restriction of M. tuberculosis replication in human macrophages.

    Download full text (pdf)
    fulltext
  • 228.
    Welin, Amanda
    et al.
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Winberg Tinnerfelt, Martin
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Abdalla, Hana
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Särndahl Lindblom, Eva
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Rasmusson, Birgitta
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Stendahl, Olle
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Lerm, Maria
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Incorporation of Mycobacterium tuberculosis lipoarabinomannan into macrophage membrane rafts is a prerequisite for the phagosomal maturation block.2008In: Infection and Immunity, ISSN 0019-9567, E-ISSN 1098-5522, Vol. 76, no 7, p. 2882-2887Article in journal (Refereed)
    Abstract [en]

    Lipoarabinomannan (LAM) is one of the key virulence factors for Mycobacterium tuberculosis, the etiological agent of tuberculosis. During uptake of mycobacteria, LAM interacts with the cell membrane of the host macrophage and can be detected throughout the cell upon infection. LAM can inhibit phagosomal maturation as well as induce a proinflammatory response in bystander cells. The aim of this study was to investigate how LAM exerts its action on human macrophages. We show that LAM is incorporated into membrane rafts of the macrophage cell membrane via its glycosylphosphatidylinositol anchor and that incorporation of mannose-capped LAM from M. tuberculosis results in reduced phagosomal maturation. This is dependent on successful insertion of the glycosylphosphatidylinositol anchor. LAM does not, however, induce the phagosomal maturation block through activation of p38 mitogen-activated protein kinase, contradicting some previous suggestions.

  • 229.
    Wetterö, Jonas
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Rheumatology . Linköping University, Faculty of Health Sciences.
    Pettersson, Sofia
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Surgery .
    Holmgren Peterson, Kajsa
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    A cellular imaging CDIO project for 2nd semester students in engineering biology2006In: World Transactions on Engineering and Technology Education, ISSN 1446-2257, Vol. 5, no 2, p. 279-282Article in journal (Refereed)
    Abstract [en]

    The demand for exact engineering within the life sciences is growing and the Engineering Biology programme at Linköping University, Linköping, Sweden, prepares students for a career at this interface. Conceive – Design – Implement – Operate (CDIO) was recently pioneered in an introductory project course. Groups of six to seven students apply a LIPS scalable project model from traditional engineering educational environments on, for example, a cellular imaging task in a hospital setting, prior to taking courses in cell biology/optics. Besides facilitating the implementation of CDIO in higher courses, students gain early career insight and enhance their communication skills. A customer (senior teacher) needs to visualise structures in cells, and the student group is contracted to deliver an applied and optimised method to meet specified requirements. The customer reviews deliverables before the tollgates and communicates with the student project leader. Other students are responsible for documentation and subsystems. The project is allocated laboratory facilities and hardware, and two fictitious subcontractors supply samples and consumables. Extra teachers perform supervision and methodological consultation. In summary, CDIO is indeed applicable and rewarding in cellular imaging, yet is also challenging.

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  • 230.
    Wetterö, Jonas
    et al.
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Rheumatology.
    Hellerstedt, T.
    Nygren, Patrik
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics.
    Broo, K.
    Occupational and Environmental Medicine, Sahlgrenska University Hospital, Göteborg University, Göteborg, Sweden.
    Aili, Daniel
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics.
    Liedberg, Bo
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics.
    Magnusson, Karl-Eric
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology.
    Immobilized chemoattractant peptides mediate adhesion and distinct calcium-dependent cell signaling in human neutrophils2008In: Langmuir, ISSN 0743-7463, E-ISSN 1520-5827, Vol. 24, no 13, p. 6803-6811Article in journal (Refereed)
    Abstract [en]

    Chemotaxis is the stimulated directional migration of cells in response to chemotactic factors, manifested for instance during leukocyte interaction with chemoattractants in inflammation. The N-formyl-Met-Leu-Phe (fMLF) bacterial peptide family is particularly potent in attracting and activating neutrophilic granulocytes. To accomplish defined circumstances for recruitment and activation of cells, we fabricated semitransparent gold-coated glass coverslips functionalized with chemoattractant fMLF receptor peptide agonist analogues. Peptides based on a common leading four-amino-acid sequence Gly-Gly-Gly-Cys were thus coupled to two potent fMLF receptor agonists, N-formyl-Tyr-Nle-Phe-Leu- Nle-Gly-Gly-Gly-Cys and N-formyl-Met-Leu-Phe-Gly-Gly-Gly-Cys, and a formylated control peptide, N-formyl-Gly-Gly-Gly-Cys. They were anchored via the SH group of Cys either directly to the gold surface or a mixed self-assembled monolayer composed of maleimide- and hydroxyl-terminated oligo(ethylene glycol) alkyldisulfides. The overall peptide immobilization procedure was characterized with ellipsometry, contact angle measurement, and infrared spectroscopy. When exposed to granulocytes, the agonist surface rapidly recruited neutrophils and the cells responded with extensive spreading and intracellular calcium transients within minutes. The reference peptide generated no such activation, and the cells maintained a more spherical morphology, suggesting that we have been able to immobilize chemoattractant receptor agonist peptides with retained bioactivity. This is a crucial step in designing surfaces with specific effects on cellular behavior. © 2008 American Chemical Society.

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  • 231.
    Wilhelmsson, Peter
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Cell Biology. Linköping University, Faculty of Health Sciences.
    Fryland, Linda
    Linköping University, Department of Clinical and Experimental Medicine. Linköping University, Faculty of Health Sciences.
    Börjesson, Stefan
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Nordgren, Johan
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Bergström, Sven
    Ernerudh, Jan
    Linköping University, Department of Clinical and Experimental Medicine, Clinical Immunology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Center for Diagnostics, Department of Clinical Immunology and Transfusion Medicine.
    Forsberg, Pia
    Linköping University, Department of Clinical and Experimental Medicine, Infectious Diseases. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Heart and Medicine Center, Department of Infectious Diseases.
    Lindgren, Per-Eric
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Erratum: Prevalence and Diversity of Borrelia Species in Ticks That Have Bitten Humans in Sweden (vol 48, pg 4169, 2010)2011In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 49, no 1, p. 481-481Article in journal (Other academic)
    Abstract [en]

    n/a

  • 232.
    Wilhelmsson, Peter
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Cell Biology. Linköping University, Faculty of Health Sciences.
    Fryland, Linda
    Linköping University, Department of Clinical and Experimental Medicine. Linköping University, Faculty of Health Sciences.
    Börjesson, Stefan
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Nordgren, Johan
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Bergström, Sven
    Umeå University.
    Ernerudh, Jan
    Linköping University, Department of Clinical and Experimental Medicine, Clinical Immunology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre for Diagnostics, Department of Clinical Immunology and Transfusion Medicine.
    Forsberg, Pia
    Linköping University, Department of Clinical and Experimental Medicine, Infectious Diseases. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Heart and Medicine Centre, Department of Infectious Diseases in Östergötland.
    Lindgren, Per-Eric
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Prevalence and Diversity of Borrelia Species in Ticks That Have Bitten Humans in Sweden2010In: JOURNAL OF CLINICAL MICROBIOLOGY, ISSN 0095-1137, Vol. 48, no 11, p. 4169-4176Article in journal (Refereed)
    Abstract [en]

    Members of the genus Borrelia are among the most common infectious agents causing tick-borne disease in humans worldwide. Here, we developed a Light Upon eXtension (LUX) real-time PCR assay that can detect and quantify Borrelia species in ticks that have fed on humans, and we applied the assay to 399 such ticks. Borrelia PCR-positive ticks were identified to species level by sequencing the products of conventional PCR performed using Borrelia group-specific primers. There was a 19% prevalence of Borrelia spp. in the detached ticks, and the number of spirochetes per Borrelia PCR-positive tick ranged from 2.0 x 10(2) to 4.9 x 10(5), with a median of 7.8 x 10(3) spirochetes. Adult ticks had a significantly larger number of spirochetes, with a median of 8.4 x 10(4) compared to the median of nymphs of 4.4 x 10(4). Adult ticks also exhibited a higher prevalence of Borrelia (33%) than nymphs (14%). Among the identified species, Borrelia afzelii was found to predominate (61%) and was followed by B. garinii (23%), B. valaisiana (13%), B. burgdorferi sensu stricto (1%), B. lusitaniae (1%), and B. miyamotoi-like (1%). Also, 3% of the ticks were coinfected with multiple strains of B. afzelii. Notably, this is the first report of B. lusitaniae being detected in ticks in Sweden. Our LUX real-time PCR assay proved to be more sensitive than a corresponding TaqMan assay. In conclusion, the novel LUX real-time PCR method is a rapid and sensitive tool for detection and quantification of Borrelia spp. in ticks.

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  • 233.
    Wilsson, Åsa
    et al.
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Majeed, Meytham
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Stendahl, Olle
    Linköping University, Department of Molecular and Clinical Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Calcium requirements for vitronectin-mediated phagocytosis of Staphylococcus aureus by human neutrophilsManuscript (preprint) (Other academic)
    Abstract [en]

    In this paper we have shown that the intracellular free calcium concentration ([Ca2+]i) in human neutrophils regulates phagocytosis of Staphylococcus aureus adherent to vitronectin-coated surfaces. When neutrophils were allowed to phagocytose bacteria bound to vitronectin- or albumin-coated surfaces in tbe presence of Ca2+, there were no obvious differences in the phagocytic activity. Ca2+-depleted neut:ropbils showed a reduced phagocytic activity on vitronectin-coated surfaces. However, the phagocytosis on albumin-coated surfaces was unaffected. Adding extracellularly Ca2+ restored the reduced phagocytic activity in Ca2+-depleted neutrophils on vitronectincoated surfaces. Similarly, using a GRGDSP-peptide to block tbe RGDmediated integrin attachment of tbe neutrophils to vitronectin restored tbe reduced phagocytic activity in Ca2+-depleted cells. Studying phagocytosis in non-migrating neutrophils adherent to vitronectin showed that ingestion of S. aureus occurred independently of Ca2+. This indicate that Ca2+ regulate the phagocytic activity of neutrophils on vitronectin-coated surfaces by regulating integrin-dependent cell directed motility.

  • 234.
    Winberg Tinnerfelt, Martin
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Leukocyte responses to pathogens: integrins, membrane rafts and nitric oxide2008Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    During microbial invasion, leukocytes of the innate immunity are rapidly recruited to the site of infection where they internalize (phagocytose), kill and digest the invaders. To aid this process, leukocytes express surface receptors such as Toll-like receptors, β2-integrins and Fc-receptors. The β2-integrins are also used for attachment to the extracellular matrix and are important for migration. When pro- vs. anti-inflammatory regulation of β2-integrins was investigated, it was found that chemotactic factors modulate neutrophil adhesion through altered affinity and/or avidity of β2-integrins. A bacteria-derived chemoattractant evoked a large increase in affinity as well as in mobility and clustering, while an early, host-derived chemotactic factor induced increased clustering and surface mobility, but only a slight increase in affinity. Anti-inflammatory lipoxin affected β2-integrin avidity, but not affinity.

    The leukocyte membrane is composed of lipids and proteins, which are inhomogeneously distributed. Specific domains in the membrane, membrane rafts, are enriched in signaling proteins and receptors. It was found that lipophosphoglycan (LPG) a virulence factor and membrane component of the parasite Leishmania donovani, accumulated in macrophage rafts during infection, inhibited PKCα translocation to the membrane and halted phagosomal maturation. Membrane rafts were instrumental for LPG to exert its effect. We further showed that nitric oxide (NO) rescued phagosomal maturation halted by Leishmania donovani parasites, possibly through effects on actin dynamics. NO did not affect parasite virulence per se. Moreover, lipoarabinomannan (LAM), a virulence factor on Mycobacterium tuberculosis (Mtb) bacteria, also inserted itself into macrophage membrane rafts. LAM from a less virulent strain (PILAM) was less efficiently inserted. Insertion could to some extent be inhibited by phosphatidylinositol mannoside (PIM), another structural molecule from Mtb. LAM did not activate the p38 MAPK signaling pathway nor did LAM interfere with TLR 2 or 4 signaling. In neutrophil leukocytes we observed a simultaneous, calciumdependent up-regulation of membrane rafts and secretion of azurophilic granules at the site of phagocytosis. Rafts were also found in the phagosome membrane. Wild type Streptococcus pyogenes bacteria, which can survive phagocytosis, modulated raft delivery.

    List of papers
    1. Differential inside-out activation of β2 integrins by leukotriene B4 and fMLP in human neutrophils
    Open this publication in new window or tab >>Differential inside-out activation of β2 integrins by leukotriene B4 and fMLP in human neutrophils
    Show others...
    2004 (English)In: Experimental Cell Research, ISSN 0014-4827, E-ISSN 1090-2422, Vol. 300, no 2, p. 308-319Article in journal (Refereed) Published
    Abstract [en]

    We have investigated how LTB4, an endogenous chemoattractant encountered early in the inflammatory process, and fMLP, a bacteria-derived chemotactic peptide emanating from the site of infection, mediate inside-out regulation of the β2-integrin. The role of the two chemoattractants on β2-integrin avidity was investigated by measuring their effect on β2-integrin clustering and surface mobility, whereas their effect on β2-integrin affinity was measured by the expression of a high affinity epitope, a ligand-binding domain on β2-integrins, and by integrin binding to s-ICAM. We find that the two chemoattractants modulate the β2-integrin differently. LTB4 induces an increase in integrin clustering and surface mobility, but only a modest increase in integrin affinity. fMLP evokes a large increase in β2-integrin affinity as well as in clustering and mobility. Lipoxin, which acts as a stop signal for the functions mediated by pro-inflammatory agents, was used as a tool for further examining the inside-out mechanisms. While LTB4-induced integrin clustering and mobility were inhibited by lipoxin, only a minor inhibition of fMLP-induced β2-integrin avidity and no inhibition of integrin affinity were detected. The different modes of the inside-out regulation of β2-integrins suggest that distinct mechanisms are involved in the β2-integrin modulation induced by various chemoattractants.

    Keywords
    β2-Integrins, Cell adhesion, Chemotactic factors, Eicosanoids, Inflammation, Leukotriene B4, Lipoxins, Human Neutrophils, Signal transduction
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:liu:diva-14629 (URN)10.1016/j.yexcr.2004.07.015 (DOI)
    Available from: 2007-09-13 Created: 2007-09-13 Last updated: 2017-12-13Bibliographically approved
    2. Leishmania donovani lipophosphoglycan inhibits phagosomal maturation via action on membrane rafts
    Open this publication in new window or tab >>Leishmania donovani lipophosphoglycan inhibits phagosomal maturation via action on membrane rafts
    Show others...
    2009 (English)In: Microbes and infection, ISSN 1286-4579, E-ISSN 1769-714X, Vol. 11, no 2, p. 215-222Article in journal (Refereed) Published
    Abstract [en]

    Lipophosphoglycan (LPG), the major surface glycoconjugate on Leishmania donovani promastigotes, is crucial for the establishment of infection inside macrophages. LPG comprises a polymer of repeating Gal beta 1,4Man alpha-PO4 attached to a lysophosphatidylinositol membrane anchor. LPG is transferred from the parasite to the host macrophage membrane during phagocytosis and induces periphagosomal F-actin accumulation correlating with an inhibition of phagosomal maturation. The biophysical properties of LPG suggest that it may be intercalated into membrane rafts of the host-cell membrane. The aim of this study was to investigate if the effects of LPG on phagosomal maturation are mediated via action on membrane rafts. We show that LPG accumulates in rafts during phagocytosis of L. donovani and that disruption of membrane rafts abolished the effects of LPG on periphagosomal F-actin and phagosomal maturation, indicating that LPG requires intact membrane rafts to manipulate host-cell functions. We conclude that LPG associates with membrane rafts in the host cell and exert its actions on host-cell actin and phagosomal maturation through subversion of raft function.

    Keywords
    Leishmania, Lipophosphoglycan, Membrane rafts, Phagosomal maturation, Actin
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:liu:diva-17886 (URN)10.1016/j.micinf.2008.11.007 (DOI)
    Note

    Original Publication: Martin Winberg Tinnerfelt, Åsa Holm, Eva Särndahl, Adrien F Vinet, Albert Descoteaux, Karl-Eric Magnusson, Birgitta Rasmusson and Maria Lerm, Leishmania donovani lipophosphoglycan inhibits phagosomal maturation via action on membrane rafts, 2009, MICROBES AND INFECTION, (11), 2, 215-222. http://dx.doi.org/10.1016/j.micinf.2008.11.007 Copyright: Elsevier Science B.V., Amsterdam. http://www.elsevier.com/

    Available from: 2009-05-20 Created: 2009-04-24 Last updated: 2017-12-13Bibliographically approved
    3. Leishmania donovani: Inhibition of phagosomal maturation is rescued by nitric oxide in macrophages
    Open this publication in new window or tab >>Leishmania donovani: Inhibition of phagosomal maturation is rescued by nitric oxide in macrophages
    2007 (English)In: Experimental parasitology, ISSN 0014-4894, E-ISSN 1090-2449, Vol. 117, no 2, p. 165-170Article in journal (Refereed) Published
    Abstract [en]

    Leishmania donovani promastigotes, the causative agent of visceral leishmaniasis, survive inside macrophages by inhibiting phagosomal maturation. The main surface glycoconjugate on promastigotes, lipophosphoglycan (LPG), is crucial for parasite survival. LPG has several detrimental effects on macrophage function, including inhibition of periphagosomal filamentous actin (F-actin) breakdown during phagosomal maturation. However, in RAW 264.7 macrophages pre-stimulated with lipopolysaccharide (LPS) and interferon ? (IFN?), known to up-regulate inducible nitric oxide synthase (iNOS) and nitric oxide (NO) production, L. donovani promastigotes are unable to inhibit periphagosomal F-actin breakdown and phagosomal maturation proceeds normally. Moreover, the iNOS inhibitor aminoguanidine, blocked the positive effects of LPS/IFN? suggesting that NO is a key player in F-actin remodeling. In conclusion, production of NO by stimulated macrophages seems to allow phagosomal maturation following uptake of L. donovani promastigotes, suggesting a novel mechanism whereby NO facilitates killing of an intracellular pathogen. © 2007 Elsevier Inc. All rights reserved.

    Keywords
    Actin, AG, aminoguanidine, bovine serum albumine, BSA, cGMP, F-actin, filamentous actin, gamma Interferon, GFP, green fluorescent protein, guanosine 3':5'-cyclic monophosphate, IFN?, inducible nitric oxide synthase, iNOS, Leishmania donovani, lipophosphoglycan, lipopolysaccharide, LPG, LPS, Maturation, Nitric oxide, nitric oxide, NO, paraformaldehyde, PFA, Phagosome
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:liu:diva-48433 (URN)10.1016/j.exppara.2007.04.004 (DOI)
    Available from: 2009-10-11 Created: 2009-10-11 Last updated: 2017-12-12Bibliographically approved
    4. Incorporation of Mycobacterium tuberculosis lipoarabinomannan into macrophage membrane rafts is a prerequisite for the phagosomal maturation block.
    Open this publication in new window or tab >>Incorporation of Mycobacterium tuberculosis lipoarabinomannan into macrophage membrane rafts is a prerequisite for the phagosomal maturation block.
    Show others...
    2008 (English)In: Infection and Immunity, ISSN 0019-9567, E-ISSN 1098-5522, Vol. 76, no 7, p. 2882-2887Article in journal (Refereed) Published
    Abstract [en]

    Lipoarabinomannan (LAM) is one of the key virulence factors for Mycobacterium tuberculosis, the etiological agent of tuberculosis. During uptake of mycobacteria, LAM interacts with the cell membrane of the host macrophage and can be detected throughout the cell upon infection. LAM can inhibit phagosomal maturation as well as induce a proinflammatory response in bystander cells. The aim of this study was to investigate how LAM exerts its action on human macrophages. We show that LAM is incorporated into membrane rafts of the macrophage cell membrane via its glycosylphosphatidylinositol anchor and that incorporation of mannose-capped LAM from M. tuberculosis results in reduced phagosomal maturation. This is dependent on successful insertion of the glycosylphosphatidylinositol anchor. LAM does not, however, induce the phagosomal maturation block through activation of p38 mitogen-activated protein kinase, contradicting some previous suggestions.

    National Category
    Microbiology in the medical area
    Identifiers
    urn:nbn:se:liu:diva-20816 (URN)10.1128/IAI.01549-07 (DOI)18426888 (PubMedID)
    Available from: 2009-09-22 Created: 2009-09-22 Last updated: 2018-01-13Bibliographically approved
    5. Phagosomal membrane rafts: azurophilic origin, Ca2+ dependence, and modulation by Streptococcus pyogenes bacteria
    Open this publication in new window or tab >>Phagosomal membrane rafts: azurophilic origin, Ca2+ dependence, and modulation by Streptococcus pyogenes bacteria
    Show others...
    (English)Manuscript (preprint) (Other academic)
    Abstract [en]

    Uptake and killing of microorganisms by neutrophils involve tightly regulated membrane traffic events that are governed by complex signals. Many of these are raft-associated, which implies that raft dynamics may be important during phagosome formation. Locally restricted, calcium-dependent, parallel upregulation of markers for membrane rafts and azurophilic granules was observed at the site of phagocytosis of IgG-opsonized prey in human neutrophils. Subsequent internalization of the prey reduced the levels of these markers in the plasma membrane. Streptococcus pyogenes bacteria, that can survive phagocytosis by neutrophils, modulated phagosomal raft acquisition by means of M proteins. Continued, but not early, delivery of rafts to the membrane of phagosomes in neutrophils and HL-60 cells was independent of calcium, as was fusion between azurophilic granules and phagosomes. Nevertheless, calcium depletion affected bacterial killing kinetics. These findings suggest that early delivery of membrane rafts is important for phagosomal maturation in neutrophils and provide new mechanistic insight into the processes required for generation of bactericidal phagosomes.

    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:liu:diva-52261 (URN)
    Available from: 2009-12-14 Created: 2009-12-14 Last updated: 2009-12-14Bibliographically approved
    Download (pdf)
    Cover
  • 235.
    Winberg Tinnerfelt, Martin
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Holm, Åsa
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Särndahl, Eva
    Linköping University, Department of Clinical and Experimental Medicine, Cell Biology. Linköping University, Faculty of Health Sciences.
    Vinet, Adrien F
    INRS, Canada.
    Descoteaux, Albert
    INRS, Canada.
    Magnusson, Karl-Eric
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Rasmusson, Birgitta
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Lerm, Maria
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Leishmania donovani lipophosphoglycan inhibits phagosomal maturation via action on membrane rafts2009In: Microbes and infection, ISSN 1286-4579, E-ISSN 1769-714X, Vol. 11, no 2, p. 215-222Article in journal (Refereed)
    Abstract [en]

    Lipophosphoglycan (LPG), the major surface glycoconjugate on Leishmania donovani promastigotes, is crucial for the establishment of infection inside macrophages. LPG comprises a polymer of repeating Gal beta 1,4Man alpha-PO4 attached to a lysophosphatidylinositol membrane anchor. LPG is transferred from the parasite to the host macrophage membrane during phagocytosis and induces periphagosomal F-actin accumulation correlating with an inhibition of phagosomal maturation. The biophysical properties of LPG suggest that it may be intercalated into membrane rafts of the host-cell membrane. The aim of this study was to investigate if the effects of LPG on phagosomal maturation are mediated via action on membrane rafts. We show that LPG accumulates in rafts during phagocytosis of L. donovani and that disruption of membrane rafts abolished the effects of LPG on periphagosomal F-actin and phagosomal maturation, indicating that LPG requires intact membrane rafts to manipulate host-cell functions. We conclude that LPG associates with membrane rafts in the host cell and exert its actions on host-cell actin and phagosomal maturation through subversion of raft function.

    Download full text (pdf)
    FULLTEXT01
  • 236.
    Winberg Tinnerfelt, Martin
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Rasmusson, Birgitta
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology .
    Sundqvist, Tommy
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology .
    Leishmania donovani: Inhibition of phagosomal maturation is rescued by nitric oxide in macrophages2007In: Experimental parasitology, ISSN 0014-4894, E-ISSN 1090-2449, Vol. 117, no 2, p. 165-170Article in journal (Refereed)
    Abstract [en]

    Leishmania donovani promastigotes, the causative agent of visceral leishmaniasis, survive inside macrophages by inhibiting phagosomal maturation. The main surface glycoconjugate on promastigotes, lipophosphoglycan (LPG), is crucial for parasite survival. LPG has several detrimental effects on macrophage function, including inhibition of periphagosomal filamentous actin (F-actin) breakdown during phagosomal maturation. However, in RAW 264.7 macrophages pre-stimulated with lipopolysaccharide (LPS) and interferon ? (IFN?), known to up-regulate inducible nitric oxide synthase (iNOS) and nitric oxide (NO) production, L. donovani promastigotes are unable to inhibit periphagosomal F-actin breakdown and phagosomal maturation proceeds normally. Moreover, the iNOS inhibitor aminoguanidine, blocked the positive effects of LPS/IFN? suggesting that NO is a key player in F-actin remodeling. In conclusion, production of NO by stimulated macrophages seems to allow phagosomal maturation following uptake of L. donovani promastigotes, suggesting a novel mechanism whereby NO facilitates killing of an intracellular pathogen. © 2007 Elsevier Inc. All rights reserved.

  • 237.
    Winbladh, Anders
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Surgery. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre for Surgery, Orthopaedics and Cancer Treatment, Department of Surgery in Östergötland.
    Björnsson, Bergthor
    Linköping University, Department of Clinical and Experimental Medicine, Surgery. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre for Surgery, Orthopaedics and Cancer Treatment, Department of Surgery in Östergötland.
    Trulsson, Lena
    Linköping University, Department of Clinical and Experimental Medicine, Surgery. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre for Surgery, Orthopaedics and Cancer Treatment, Department of Surgery in Östergötland.
    Bojmar, Linda
    Linköping University, Department of Clinical and Experimental Medicine. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre for Surgery, Orthopaedics and Cancer Treatment, Department of Surgery in Östergötland.
    Sundqvist, Tommy
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Gullstrand, Per
    Linköping University, Department of Clinical and Experimental Medicine. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre for Surgery, Orthopaedics and Cancer Treatment, Department of Surgery in Östergötland.
    Sandström, Per
    Linköping University, Department of Clinical and Experimental Medicine, Surgery. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre for Surgery, Orthopaedics and Cancer Treatment, Department of Surgery in Östergötland.
    N-acetyl cysteine improves glycogenesis after segmental liver ischemia and reperfusion injury in pigs2012In: Scandinavian Journal of Gastroenterology, ISSN 0036-5521, E-ISSN 1502-7708, Vol. 47, no 2, p. 225-236Article in journal (Refereed)
    Abstract [en]

    Abstract Objective. N-acetylcysteine (NAC) is an antioxidative molecule known to protect liver tissue from oxygen radical species generated during ischemia and reperfusion (IR). Nutritional and toxicology studies have shown that NAC also improves glucose metabolism and glycogen stores. We hypothesized that NAC improves glycogenesis and that impaired glycogenesis is a key element in IR injury. Material and Methods. In an experimental model, 80 min of segmental liver ischemia was induced in 16 pigs and the reperfusion was followed for 360 min. Eight animals received NAC 150 mg/kg as a bolus injection followed by an infusion of NAC 50 mg/kg/h intravenously. Results. AST and leukocyte density were lower in the NAC-treated animals, unrelated to the glutathione levels or apoptosis. Glycogen stores returned to a higher degree in the NAC-treated animals and microdialysis revealed lower levels of lactate during the reperfusion phase. Nitrite/Nitrate levels in the NAC group were lower in both serum and microdialysates, indicating that NAC scavenges radical nitrosative species. Conclusions. NAC treatment improves glycogenesis after liver IR injury and reduces the level of intraparenchymal lactate during reperfusion, possibly due to the scavenging of radical nitrosative species.

  • 238.
    Winbladh, Anders
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Surgery. Linköping University, Faculty of Health Sciences.
    Björnsson, Bergthor
    Linköping University, Department of Clinical and Experimental Medicine, Surgery. Linköping University, Faculty of Health Sciences.
    Trulsson, Lena
    Linköping University, Department of Clinical and Experimental Medicine, Surgery. Linköping University, Faculty of Health Sciences.
    Bojmar, Linda
    Linköping University, Department of Clinical and Experimental Medicine, Surgery. Linköping University, Faculty of Health Sciences.
    Sundqvist, Tommy
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Gullstrand, Per
    Linköping University, Department of Clinical and Experimental Medicine. Linköping University, Faculty of Health Sciences.
    Sandström, Per
    Linköping University, Department of Clinical and Experimental Medicine, Surgery. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre of Surgery and Oncology, Oncology Centre.
    N-Acetylcysteine Improves Glycogenesis after Segmental Liver Ischemia and Reperfusion Injury in PigsManuscript (preprint) (Other academic)
    Abstract [en]

    Objective: N-Acetylcysteine (NAC) is an antioxidative molecule known to protect liver tissue from oxygen radical species generated during ischemia and reperfusion. Nutritional and toxicology studies have shown that NAC also improves glucose metabolism and glycogen stores. We hypothesized that NAC improves glycogenesis and that impaired glycogenesis is a key element in ischemia-reperfusion injury.

    Material and Methods: In an experimental model, 80 minutes of segmental liver ischemia was induced in 16 pigs and the reperfusion was followed for 360 minutes. Eight animals received NAC 150 mg/kg as a bolus injection followed by an infusion of NAC 50 mg/kg/h intravenously.

    Results: AST and leukocyte density were lower in the NAC-treated animals, unrelated to the glutathione levels or apoptosis. Glycogen stores returned to a higher degree in the NAC treated animals and microdialysis revealed lower levels of lactate during the reperfusion phase. Nitrite/Nitrate levels in the NAC group were lower in both serum and microdialysate, indicating that NAC scavenges radical nitrosative species (RNS).

    Conclusions: NAC treatment improves glycogenesis after liver ischemia and reperfusion injury and reduces the level of intraparenchymal lactate during reperfusion, possibly due to the scavenging of RNS.

  • 239.
    Wirén, Michael
    Linköping University, Department of Biomedicine and Surgery, Surgery. Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Growth and integrity of the small intestine in malnutrition and trauma1995Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    The small intestine is an active metabolic organ constituting a functional and immunologic barrier to toxins and microbes in the intestinal lumen. Injuries are repaired by rapid cell replication, which depends on nutritional and humoral growth factors. Glutamine has been suggested to be the most important nutrient for the enterocytes. In the present studies, the effects of glutamine were evaluated using experiments with cultured cells and postoperative supplementation in animals and humans.

    Growth of two enterocyte-like epithelial cell lines, CaCo-2 and HT 29, was studied at different glutamine concentrations, and compared to effects of growth factors and energy substrates. Glutamine effects in starved, operated rats were evaluated by weight, DNA, protein analysis, 3H-thymidine incorporation in intestinal mucosa and urinary recovery of orally administered polyethylene glycols. Three different balanced and complete enteral preparations with no glutamine, 2% (normal) and 4% were used postoperatively . Growth parameters and tissue and plasma concentrations of humoral growth factors were studied 3 and 8 days after intestinal resection in the rat. In a clinical study with total parenteral nutrition for five days after major abdominal surgery, nitrogen balance and humoral growth factors in plasma were evaluated in patients receiving glutamine-containing dipeptide (Gly-Gln) amino acid solution, compared to conventional amino acid solution.

    In the cell cultures, glutamine was shown to be of importance both as a trophic factor and as a metabolic substrate, particularly in cells of intestinal origin. In the animal model of malnutrition and surgery, 3H-thymidine incorporation was higher in the supplemented group compared to glutamine-free and also higher in all operated groups compared to controls. The permeability study showed a higher uptake of small polyethylene glycol molecules in glutamine-supplemented animals, parallel to increases in thymidine incorporation. After major intestinal resection in rats, no major benefit on growth by glutamine supplementation could be found after one week. Rapid PYY increases in plasma and higher IGF-II concentration in ileal mucosa were found. Stimulation of IGF-II concentration suggested an auto- or paracrine action in regulating growth. In the clinical study, no significant differences were seen in the levels of transthyretin, retinol binding protein or nitrogen balance, compared to patients recieving conventional amino acid solution. A positive correlation between insulin and nitrogen balance was found inglutamine treated patients.

    It is concluded that glutamine has important effects as a nutritional substrate for enterocytes and stimulates their proliferation and absorptive capacity in refeeding after malnutrition and surgical trauma in the rat. After intestinal resection, glutamme has no major effects upon growth one week after surgery, but the production of growth factorsincreases earlier in glutamine supplemented animals. In clinical use, a glutamine supplemented amino acid solution appeared no better than conventional amino acd solution. It could, howeyer, represent a more balanced way of supportmg protem metabolism after trauma, by the interaction with insulin and other growth factors.

  • 240.
    Zhang, Jing-Ping
    et al.
    Sun Yat Sen University.
    Zheng, Limin
    Sun Yat Sen University.
    Wang, Jiang-Hai
    Sun Yat Sen University.
    Magnusson, Karl-Eric
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Liu, Xin
    Sun Yat Sen University.
    Lipid Extract from Completely Sporoderm-broken Germinating Ganoderma sinensis Spores Elicits Potent Antitumor Immune Responses in Human Macrophages2009In: PHYTOTHERAPY RESEARCH, ISSN 0951-418X, Vol. 23, no 6, p. 844-850Article in journal (Refereed)
    Abstract [en]

    Ganoderma sinensis has been used widely in Oriental countries for the prevention and treatment of various diseases including cancer. Previous studies have shown that the lipid extract from Ganoderma exhibits direct cytotoxicity against tumor cells. Here, it is reported that the lipid extract from germinating G. sinensis spores, at lower concentrations that have no direct tumoricidal activity, induce potent antitumor immune responses in human monocytes/macrophages. Upon stimulation with the lipid extract, monocytes/macrophages exhibited markedly increased production of proinflammatory cytokines and surface expression of costimulatory molecules. Conditioned medium from stimulated cells effectively suppressed the growth of tumor cells. Apparently, the lipid extract triggered macrophage activation via a mechanism different from that associated with LPS. Moreover, it was observed that the lipid extract could partially re-establish the antitumor activity of the immunosuppressive tumor-associated macrophages. These results indicated that in addition to its direct tumoricidal activity, the lipid extract from G. sinensis spores could exert antitumor activity by stimulating the activation of human monocytes/macrophages.

  • 241.
    Zheng, Limin
    et al.
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology .
    Forsberg, Maria
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Molecular and Clinical Medicine.
    Stendahl, Olle
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology .
    Invasive Salmonella typhimurium induces apoptosis in human U937 cells by activating Rho GTPases2000In: Molecular Biology of the Cell, ISSN 1059-1524, E-ISSN 1939-4586, Vol. 11, p. 1527-Conference paper (Other academic)
  • 242.
    Zheng, Limin
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    He, Min
    Department of Biochemistry, College of Life Sciences, Sun Yatsen (Zhongshan) University, Guangzhou, China.
    Long, Min
    Blomgran, Robert
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Stendahl, Olle
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Pathogen-Induced Apoptotic Neutrophils Express Heat Shock Proteins and Elicit Activation of Human Macrophages2004In: Journal of immunology, ISSN 0022-1767, Vol. 173, no 10, p. 6319-6326Article in journal (Refereed)
    Abstract [en]

    Ingestion of aged or irradiated apoptotic neutrophils actively suppresses stimulation of macrophages (MΦ). Many bacterial pathogens can also provoke apoptosis in neutrophils, but little is known about how such apoptotic cells influence MΦ activation. We found that neutrophils undergoing apoptosis induced by UV irradiation, Escherichia coli, or Staphylococcus aureus could either stimulate or inhibit MΦ activation. In contrast to MΦ that had ingested irradiated apoptotic neutrophils, MΦ that had phagocytosed bacteria-induced apoptotic neutrophils exhibited markedly increased production of the proinflammatory cytokine TNF-α, but not the anti-inflammatory cytokine TGF-β. Moreover, ingestion of bacteria, but not UV-induced apoptotic neutrophils, caused increased expression of FcγRI on MΦ, and this effect was not provoked directly by bacteria associated with the apoptotic neutrophils. Instead, we found that a link between pathogen-induced apoptotic neutrophils and up-regulation of the heat shock proteins HSP60 and HSP70, and we also observed that recombinant HSP60 and HSP70 potentiated LPS-stimulated production of TNF-α in MΦ. The opposing macrophage responses to neutrophils undergoing apoptosis induced in different ways may represent a novel mechanism that regulates the extent of the immune response to invading microbes in two steps: first by aiding the functions of MΦ at an early stage of infection, and subsequently by deactivating those cells through removal of uninfected apoptotic neutrophils. HSP induction in neutrophils may provide the danger signals required to generate a more effective macrophage response.

  • 243.
    Zurek, Birte
    et al.
    University of Cologne, Germany .
    Schoultz, Ida
    Linköping University, Department of Clinical and Experimental Medicine, Surgery. Linköping University, Faculty of Health Sciences.
    Neerincx, Andreas
    University of Cologne, Germany .
    Napolitano, Luisa M
    Telethon Institute Genet and Med, Italy Cluster Biomed CBM, Italy .
    Birkner, Katharina
    University of Cologne, Germany .
    Bennek, Eveline
    University Hospital Aachen, Germany .
    Sellge, Gernot
    University Hospital Aachen, Germany .
    Lerm, Maria
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Meroni, Germana
    Cluster Biomed CBM, Italy .
    Söderholm, Johan D
    Linköping University, Department of Clinical and Experimental Medicine, Surgery. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre for Surgery, Orthopaedics and Cancer Treatment, Department of Surgery in Östergötland.
    Kufer, Thomas A
    University of Cologne, Germany .
    TRIM27 Negatively Regulates NOD2 by Ubiquitination and Proteasomal Degradation2012In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 7, no 7Article in journal (Refereed)
    Abstract [en]

    NOD2, the nucleotide-binding domain and leucine-rich repeat containing gene family (NLR) member 2 is involved in mediating antimicrobial responses. Dysfunctional NOD2 activity can lead to severe inflammatory disorders, but the regulation of NOD2 is still poorly understood. Recently, proteins of the tripartite motif (TRIM) protein family have emerged as regulators of innate immune responses by acting as E3 ubiquitin ligases. We identified TRIM27 as a new specific binding partner for NOD2. We show that NOD2 physically interacts with TRIM27 via the nucleotide-binding domain, and that NOD2 activation enhances this interaction. Dependent on functional TRIM27, ectopically expressed NOD2 is ubiquitinated with K48-linked ubiquitin chains followed by proteasomal degradation. Accordingly, TRIM27 affects NOD2-mediated pro-inflammatory responses. NOD2 mutations are linked to susceptibility to Crohns disease. We found that TRIM27 expression is increased in Crohns disease patients, underscoring a physiological role of TRIM27 in regulating NOD2 signaling. In HeLa cells, TRIM27 is partially localized in the nucleus. We revealed that ectopically expressed NOD2 can shuttle to the nucleus in a Walker A dependent manner, suggesting that NOD2 and TRIM27 might functionally cooperate in the nucleus. We conclude that TRIM27 negatively regulates NOD2-mediated signaling by degradation of NOD2 and suggest that TRIM27 could be a new target for therapeutic intervention in NOD2-associated diseases.

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  • 244.
    Östholm Balkhed, Åse
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Clinical Microbiology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Heart and Medicine Centre, Department of Infectious Diseases in Östergötland.
    Tärnberg, Maria
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Monstein, Hans-Jürg
    Linköping University, Department of Clinical and Experimental Medicine, Clinical Microbiology. Linköping University, Department of Clinical and Experimental Medicine, Infectious Diseases. Linköping University, Faculty of Health Sciences.
    Hällgren, Anita
    Linköping University, Department of Clinical and Experimental Medicine, Infectious Diseases. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Heart and Medicine Centre, Department of Infectious Diseases in Östergötland.
    Hanberger, Håkan
    Linköping University, Department of Clinical and Experimental Medicine, Infectious Diseases. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Heart and Medicine Centre, Department of Infectious Diseases in Östergötland.
    Nilsson, Lennart E.
    Linköping University, Department of Clinical and Experimental Medicine, Clinical Microbiology. Linköping University, Faculty of Health Sciences.
    High frequency of co-resistance in CTX-M-producing Escherichia coli to non-beta-lactam antibiotics, with the exception of amikacin, nitrofurantoin, colistin, tigecycline, and fosfomycin, in a county of Sweden2013In: Scandinavian Journal of Infectious Diseases, ISSN 0036-5548, E-ISSN 1651-1980, Vol. 45, no 4, p. 271-278Article in journal (Refereed)
    Abstract [en]

    Background: The objective of this study was to investigate the in vitro activity of different antibiotics against CTX-M-producing Escherichia coli in a county of Sweden, and to determine the occurrence of multi-resistance and plasmid- mediated quinolone resistance among these isolates. Methods: A total of 198 isolates of E. coli with extended-spectrum beta-lactamase (ESBL) phenotype and mainly CTX-M genotype were studied. The minimum inhibitory concentrations (MICs) for amikacin, chloramphenicol, ciprofloxacin, colistin, fosfomycin, gentamicin, nalidixic acid, nitrofurantoin, tigecycline, tobramycin, trimethoprim, and trimethoprim-sulfamethoxazole were determined with the Etest. Susceptibility was defined according to the breakpoints of the European Committee on Antimicrobial Susceptibility Testing (EUCAST). MIC(50) and MIC(90) values were calculated. Results: Ninety-five percent or more of the isolates were susceptible to amikacin, nitrofurantoin, colistin, tigecycline, and fosfomycin. CTX-M group 9 was more susceptible than CTX-M group 1 to ciprofloxacin, gentamicin, and tobramycin. Sixty-eight percent of the isolates were multi-resistant, and the most common multi-resistance pattern was ESBL phenotype with decreased susceptibility to trimethoprim, trimethoprim-sulfamethoxazole, ciprofloxacin, gentamicin, and tobramycin. Only 1 isolate carried a qnrS1 gene, but 37% carried aac(6')-Ib-cr. Conclusions: A high frequency of co-resistance between ESBL-producing E. coli and non-beta-lactam antibiotics was seen. On the other hand, very high susceptibility was seen for amikacin, nitrofurantoin, colistin, tigecycline, and fosfomycin. These data support the replacement of gentamicin and tobramycin, normally used in Sweden, with amikacin, for severe infections.

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